BioMed Research International: Microbiology The latest articles from Hindawi Publishing Corporation © 2015 , Hindawi Publishing Corporation . All rights reserved. Microbiological Quality of Ready-to-Eat Vegetables Collected in Mexico City: Occurrence of Aerobic-Mesophilic Bacteria, Fecal Coliforms, and Potentially Pathogenic Nontuberculous Mycobacteria Mon, 30 Mar 2015 12:44:05 +0000 The aims of this study were to evaluate the microbiological quality and the occurrence of nontuberculous mycobacteria (NTM) in a variety of salads and sprouts from supermarkets and street vendors in Mexico City. Aerobic-mesophilic bacteria (AMB) were present in 100% of RTE-salads samples; 59% of samples were outside guidelines range (>5.17 log10 CFU per g). Although fecal coliforms (FC) were present in 32% of samples, only 8% of them exceeded the permissible limit (100 MPN/g). Regarding the 100 RTE-sprouts, all samples were also positive for AMB and total coliforms (TC) and 69% for FC. Seven NTM species were recovered from 7 salad samples; they included three M. fortuitum, two M. chelonae, one M. mucogenicum, and one M. sp. Twelve RTE-sprouts samples harbored NTM, which were identified as M. porcinum (five), M. abscessus (two), M. gordonae (two), M. mucogenicum (two), and M. avium complex (one). Most RTE-salads and RTE-sprouts had unsatisfactory microbiological quality and some harbored NTM associated with illness. No correlation between the presence of coliforms and NTM was found. Overall, these results suggest that RTE-salads and RTE-sprouts might function as vehicles for NTM transmission in humans; hence, proper handling and treatment before consumption of such products might be recommendable. Jorge Francisco Cerna-Cortes, Nancy Leon-Montes, Ana Laura Cortes-Cueto, Laura P. Salas-Rangel, Addy Cecilia Helguera-Repetto, Daniel Lopez-Hernandez, Sandra Rivera-Gutierrez, Elizabeth Fernandez-Rendon, and Jorge Alberto Gonzalez-y-Merchand Copyright © 2015 Jorge Francisco Cerna-Cortes et al. All rights reserved. Antibiotic-Resistant Vibrios in Farmed Shrimp Mon, 30 Mar 2015 11:22:44 +0000 Antimicrobial susceptibility pattern was determined in 100 strains of Vibrio isolated from the Litopenaeus vannamei shrimp and identified phenotypically. A high antibiotic-resistance index (75%) was observed, with the following phenotypic profiles: monoresistance (), cross-resistance to β-lactams () and multiple resistance (). Plasmid resistance was characterized for penicillin (), penicillin + ampicillin (), penicillin + aztreonam (), and ampicillin (). Resistance to antimicrobial drugs by the other strains () was possibly mediated by chromosomal genes. The findings of this study support the conclusion that the cultured shrimps can be vehicles of vibrios resistant to β-lactam and tetracycline. Renata Albuquerque Costa, Rayza Lima Araújo, Oscarina Viana Souza, and Regine Helena Silva dos Fernandes Vieira Copyright © 2015 Renata Albuquerque Costa et al. All rights reserved. The Water Cycle, a Potential Source of the Bacterial Pathogen Bacillus cereus Mon, 30 Mar 2015 09:19:27 +0000 The behaviour of the sporulating soil-dwelling Bacillus cereus sensu lato (B. cereus sl) which includes foodborne pathogenic strains has been extensively studied in relation to its various animal hosts. The aim of this environmental study was to investigate the water compartments (rain and soil water, as well as groundwater) closely linked to the primary B. cereus sl reservoir, for which available data are limited. B. cereus sl was present, primarily as spores, in all of the tested compartments of an agricultural site, including water from rain to groundwater through soil. During rain events, leachates collected after transfer through the soil eventually reached the groundwater and were loaded with B. cereus sl. In groundwater samples, newly introduced spores of a B. cereus model strain were able to germinate, and vegetative cells arising from this event were detected for up to 50 days. This first B. cereus sl investigation in the various types of interrelated environments suggests that the consideration of the aquatic compartment linked to soil and to climatic events should provide a better understanding of B. cereus sl ecology and thus be relevant for a more accurate risk assessment of food poisoning caused by B. cereus sl pathogenic strains. Julien Brillard, Christian M. S. Dupont, Odile Berge, Claire Dargaignaratz, Stéphanie Oriol-Gagnier, Claude Doussan, Véronique Broussolle, Marina Gillon, Thierry Clavel, and Annette Bérard Copyright © 2015 Julien Brillard et al. All rights reserved. Listeriosis Outbreaks in British Columbia, Canada, Caused by Soft Ripened Cheese Contaminated from Environmental Sources Mon, 30 Mar 2015 09:17:07 +0000 Soft ripened cheese (SRC) caused over 130 foodborne illnesses in British Columbia (BC), Canada, during two separate listeriosis outbreaks. Multiple agencies investigated the events that lead to cheese contamination with Listeria monocytogenes (L.m.), an environmentally ubiquitous foodborne pathogen. In both outbreaks pasteurized milk and the pasteurization process were ruled out as sources of contamination. In outbreak A, environmental transmission of L.m. likely occurred from farm animals to personnel to culture solutions used during cheese production. In outbreak B, birds were identified as likely contaminating the dairy plant’s water supply and cheese during the curd-washing step. Issues noted during outbreak A included the risks of operating a dairy plant in a farm environment, potential for transfer of L.m. from the farm environment to the plant via shared toilet facilities, failure to clean and sanitize culture spray bottles, and cross-contamination during cheese aging. L.m. contamination in outbreak B was traced to wild swallows defecating in the plant’s open cistern water reservoir and a multibarrier failure in the water disinfection system. These outbreaks led to enhanced inspection and surveillance of cheese plants, test and release programs for all SRC manufactured in BC, improvements in plant design and prevention programs, and reduced listeriosis incidence. Lorraine McIntyre, Lynn Wilcott, and Monika Naus Copyright © 2015 Lorraine McIntyre et al. All rights reserved. Resistance Determinants and Their Association with Different Transposons in the Antibiotic-Resistant Streptococcus pneumoniae Thu, 26 Mar 2015 11:34:44 +0000 Multiple resistance of Streptococcus pneumoniae is generally associated with their unique recombination-mediated genetic plasticity and possessing the mobile genetic elements. The aim of our study was to detect antibiotic resistance determinants and conjugative transposons in 138 antibiotic-resistant pneumococcal strains isolated from nasopharynx of healthy young children from Lublin, Poland. These strains resistant to tetracycline and/or to chloramphenicol/erythromycin/clindamycin were tested by PCR using the specific genes as markers. The presence of Tn916 family transposons, carrying tet(M) and int/xisTn916, was observed in all of the tested strains. Tn916 was detected in 16 strains resistant only to tetracycline. Tn6002 and Tn3872-related element were found among 99 erm(B)-carrying strains (83.8% and 3.0%, resp.). Eight strains harbouring mef(E) and erm(B) genes were detected, suggesting the presence of Tn2010 and Tn2017 transposons. Among 101 chloramphenicol-resistant strains, two variants of Tn5252-related transposon were distinguished depending on the presence of int/xis5252 genes specific for cat gene-containing Tn5252 (75.2% of strains) or gene, specific for cat-containing ICESp23FST81 element (24.8% of strains). In 6 strains Tn916-like and Tn5252-like elements formed a Tn5253-like structure. Besides clonal dissemination of resistant strains of pneumococci in the population, horizontal transfer of conjugative transposons is an important factor of the high prevalence of antibiotic resistance. Izabela Korona-Glowniak, Radoslaw Siwiec, and Anna Malm Copyright © 2015 Izabela Korona-Glowniak et al. All rights reserved. BIODESERT: Exploring and Exploiting the Microbial Resource of Hot and Cold Deserts Thu, 26 Mar 2015 07:25:05 +0000 Ameur Cherif, George Tsiamis, Stéphane Compant, and Sara Borin Copyright © 2015 Ameur Cherif et al. All rights reserved. Vector-Borne Viral Diseases Tue, 24 Mar 2015 12:38:27 +0000 Penghua Wang, Fengwei Bai, Gong Cheng, Jianfeng Dai, and Michael J. Conway Copyright © 2015 Penghua Wang et al. All rights reserved. Evaluation of the Efficacy of a Bacteriophage in the Treatment of Pneumonia Induced by Multidrug Resistance Klebsiella pneumoniae in Mice Tue, 24 Mar 2015 06:18:12 +0000 Multidrug-resistant Klebsiella pneumoniae (MRKP) has steadily grown beyond antibiotic control. However, a bacteriophage is considered to be a potential antibiotic alternative for treating bacterial infections. In this study, a lytic bacteriophage, phage 1513, was isolated using a clinical MRKP isolate KP 1513 as the host and was characterized. It produced a clear plaque with a halo and was classified as Siphoviridae. It had a short latent period of 30 min, a burst size of 264 and could inhibit KP 1513 growth in vitro with a dose-dependent pattern. Intranasal administration of a single dose of 2 × 109 PFU/mouse 2 h after KP 1513 inoculation was able to protect mice against lethal pneumonia. In a sublethal pneumonia model, phage-treated mice exhibited a lower level of K. pneumoniae burden in the lungs as compared to the untreated control. These mice lost less body weight and exhibited lower levels of inflammatory cytokines in their lungs. Lung lesion conditions were obviously improved by phage therapy. Therefore, phage 1513 has a great effect in vitro and in vivo, which has potential to be used as an alternative to an antibiotic treatment of pneumonia that is caused by the multidrug-resistant K. pneumoniae. Fang Cao, Xitao Wang, Linhui Wang, Zhen Li, Jian Che, Lili Wang, Xiaoyu Li, Zhenhui Cao, Jiancheng Zhang, Liji Jin, and Yongping Xu Copyright © 2015 Fang Cao et al. All rights reserved. Vector Borne Infections in Italy: Results of the Integrated Surveillance System for West Nile Disease in 2013 Sun, 22 Mar 2015 09:20:25 +0000 The epidemiology of West Nile disease (WND) is influenced by multiple ecological factors and, therefore, integrated surveillance systems are needed for early detecting the infection and activating consequent control actions. As different animal species have different importance in the maintenance and in the spread of the infection, a multispecies surveillance approach is required. An integrated and comprehensive surveillance system is in place in Italy aiming at early detecting the virus introduction, monitoring the possible infection spread, and implementing preventive measures for human health. This paper describes the integrated surveillance system for WND in Italy, which incorporates data from veterinary and human side in order to evaluate the burden of infection in animals and humans and provide the public health authorities at regional and national levels with the information needed for a fine tune response. Christian Napoli, Simona Iannetti, Caterina Rizzo, Antonino Bella, Daria Di Sabatino, Rossana Bruno, Francesca Sauro, Vanessa Martini, Vincenzo Ugo Santucci, Silvia Declich, and Paolo Calistri Copyright © 2015 Christian Napoli et al. All rights reserved. Sialic Acid Expression in the Mosquito Aedes aegypti and Its Possible Role in Dengue Virus-Vector Interactions Sun, 22 Mar 2015 09:03:44 +0000 Dengue fever (DF) is the most prevalent arthropod-borne viral disease which affects humans. DF is caused by the four dengue virus (DENV) serotypes, which are transmitted to the host by the mosquito Aedes aegypti that has key roles in DENV infection, replication, and viral transmission (vector competence). Mosquito saliva also plays an important role during DENV transmission. In this study, we detected the presence of sialic acid (Sia) in Aedes aegypti tissues, which may have an important role during DENV-vector competence. We also identified genome sequences encoding enzymes involved in Sia pathways. The cDNA for Aedes aegypti CMP-Sia synthase (CSAS) was amplified, cloned, and functionally evaluated via the complementation of LEC29.Lec32 CSAS-deficient CHO cells. AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface. Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome. Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues. In addition, we evaluated the possible role of sialylated molecules in a salivary gland extract during DENV internalization in mammalian cells. The knowledge of early DENV-host interactions could facilitate a better understanding of viral tropism and pathogenesis to allow the development of new strategies for controlling DENV transmission. Jorge Cime-Castillo, Philippe Delannoy, Guillermo Mendoza-Hernández, Verónica Monroy-Martínez, Anne Harduin-Lepers, Humberto Lanz-Mendoza, Fidel de la Cruz Hernández-Hernández, Edgar Zenteno, Carlos Cabello-Gutiérrez, and Blanca H. Ruiz-Ordaz Copyright © 2015 Jorge Cime-Castillo et al. All rights reserved. Occurrence of West Nile Virus Antibodies in Wild Birds, Horses, and Humans in Poland Thu, 19 Mar 2015 14:35:51 +0000 Serum samples of 474 wild birds, 378 horses, and 42 humans with meningitis and lymphocytic meningitis were collected between 2010 and 2014 from different areas of Poland. West Nile virus (WNV) antibodies were detected using competition enzyme linked immunosorbent assays: ELISA-1 ID Screen West Nile Competition, IDvet, ELISA-2 ID Screen West Nile IgM Capture, and ELISA-3 Ingezim West Nile Compac. The antibodies were found in 63 (13.29%) out of 474 wild bird serum samples and in one (0.26%) out of 378 horse serum samples. Fourteen (33.33%) out of 42 sera from patients were positive against WNV antigen and one serum was doubtful. Positive samples obtained in birds were next retested with virus microneutralisation test to confirm positive results and cross-reactions with other antigens of the Japanese encephalitis complex. We suspect that positive serological results in humans, birds, and horses indicate that WNV can be somehow closely related with the ecosystem in Poland. Jowita Samanta Niczyporuk, Elżbieta Samorek-Salamonowicz, Sylvie Lecollinet, Sławomir Andrzej Pancewicz, Wojciech Kozdruń, and Hanna Czekaj Copyright © 2015 Jowita Samanta Niczyporuk et al. All rights reserved. The Date Palm Tree Rhizosphere Is a Niche for Plant Growth Promoting Bacteria in the Oasis Ecosystem Thu, 19 Mar 2015 14:32:03 +0000 In arid ecosystems environmental factors such as geoclimatic conditions and agricultural practices are of major importance in shaping the diversity and functionality of plant-associated bacterial communities. Assessing the influence of such factors is a key to understand (i) the driving forces determining the shape of root-associated bacterial communities and (ii) the plant growth promoting (PGP) services they provide. Desert oasis environment was chosen as model ecosystem where agriculture is possible by the microclimate determined by the date palm cultivation. The bacterial communities in the soil fractions associated with the root system of date palms cultivated in seven oases in Tunisia were assessed by culture-independent and dependent approaches. According to 16S rRNA gene PCR-DGGE fingerprinting, the shapes of the date palm rhizosphere bacterial communities correlate with geoclimatic features along a north-south aridity transect. Despite the fact that the date palm root bacterial community structure was strongly influenced by macroecological factors, the potential rhizosphere services reflected in the PGP traits of isolates screened in vitro were conserved among the different oases. Such services were exerted by the 83% of the screened isolates. The comparable numbers and types of PGP traits indicate their importance in maintaining the plant functional homeostasis despite the different environmental selection pressures. Raoudha Ferjani, Ramona Marasco, Eleonora Rolli, Hanene Cherif, Ameur Cherif, Maher Gtari, Abdellatif Boudabous, Daniele Daffonchio, and Hadda-Imene Ouzari Copyright © 2015 Raoudha Ferjani et al. All rights reserved. Genetic and Technological Characterisation of Vineyard- and Winery-Associated Lactic Acid Bacteria Thu, 19 Mar 2015 13:16:01 +0000 Vineyard- and winery-associated lactic acid bacteria (LAB) from two major PDO regions in Greece, Peza and Nemea, were surveyed. LAB were isolated from grapes, fermenting musts, and winery tanks performing spontaneous malolactic fermentations (MLF). Higher population density and species richness were detected in Nemea than in Peza vineyards and on grapes than in fermenting musts. Pediococcus pentosaceus and Lactobacillus graminis were the most abundant LAB on grapes, while Lactobacillus plantarum dominated in fermenting musts from both regions. No particular structure of Lactobacillus plantarum populations according to the region of origin was observed, and strain distribution seems random. LAB species diversity in winery tanks differed significantly from that in vineyard samples, consisting principally of Oenococcus oeni. Different strains were analysed as per their enological characteristics and the ability to produce biogenic amines (BAs). Winery-associated species showed higher resistance to low pH, ethanol, SO2, and CuSO4 than vineyard-associated isolates. The frequency of BA-producing strains was relatively low but not negligible, considering that certain winery-associated Lactobacillus hilgardii strains were able to produce BAs. Present results show the necessity of controlling the MLF by selected starters in order to avoid BA accumulation in wine. Aspasia A. Nisiotou, Dimitra Dourou, Maria-Evangelia Filippousi, Ellie Diamantea, Petros Fragkoulis, Chryssoula Tassou, and Georgios Banilas Copyright © 2015 Aspasia A. Nisiotou et al. All rights reserved. The Formation of Biofilms by Pseudomonas aeruginosa: A Review of the Natural and Synthetic Compounds Interfering with Control Mechanisms Thu, 19 Mar 2015 12:30:24 +0000 P. aeruginosa is an opportunistic pathogenic bacterium responsible for both acute and chronic infections. Beyond its natural resistance to many drugs, its ability to form biofilm, a complex biological system, renders ineffective the clearance by immune defense systems and antibiotherapy. The objective of this report is to provide an overview (i) on P. aeruginosa biofilm lifestyle cycle, (ii) on the main key actors relevant in the regulation of biofilm formation by P. aeruginosa including QS systems, GacS/GacA and RetS/LadS two-component systems and C-di-GMP-dependent polysaccharides biosynthesis, and (iii) finally on reported natural and synthetic products that interfere with control mechanisms of biofilm formation by P. aeruginosa without affecting directly bacterial viability. Concluding remarks focus on perspectives to consider biofilm lifestyle as a target for eradication of resistant infections caused by P. aeruginosa. Tsiry Rasamiravaka, Quentin Labtani, Pierre Duez, and Mondher El Jaziri Copyright © 2015 Tsiry Rasamiravaka et al. All rights reserved. Identification of the Alternative Splicing of the UL49 Locus of Human Cytomegalovirus Thu, 19 Mar 2015 10:14:57 +0000 The UL49 ORF of human cytomegalovirus (HCMV) is essential for viral replication; conserved among all herpes viruses; however, the function is unclear. Once the UL49 ORF was precisely deleted from the start to stop codon, the mutant did not yield infectious progeny. In this study, we find out many alternatively processed ESTs in UL49 locus in HCMV-infected cells, in which there are two novel transcription termination sites in UL49 locus. Most of these ESTs are rare transcripts that contain directed repeat sequences in the intron splicing regions. There is a typical GU-AG intron splicing site in UL49Y transcripts. The 1847 bp UL49Y cDNA spans an ORF from 335 to 1618 and encodes a putative protein of 427 amino acids with a predicted molecular mass of 47.1 kDa. All the new EST sequences and UL49Y cDNA sequence have been deposited in the GenBank database (GenBank Accession nos. GW314860-GW314900 and GU376796). This study provides us with very important clues for revealing the importance of the UL49 locus alternative splicing. Guang Yang, Wei Li, Wenzhen Liao, Xin Zhang, Yi Zou, Jianfeng Dai, Yueqin Li, Chunxia Jing, and Tianhong Zhou Copyright © 2015 Guang Yang et al. All rights reserved. The Global Ecology and Epidemiology of West Nile Virus Thu, 19 Mar 2015 08:58:25 +0000 Since its initial isolation in Uganda in 1937 through the present, West Nile virus (WNV) has become an important cause of human and animal disease worldwide. WNV, an enveloped virus of the genus Flavivirus, is naturally maintained in an enzootic cycle between birds and mosquitoes, with occasional epizootic spillover causing disease in humans and horses. The mosquito vectors for WNV are widely distributed worldwide, and the known geographic range of WNV transmission and disease has continued to increase over the past 77 years. While most human infections with WNV are asymptomatic, severe neurological disease may develop resulting in long-term sequelae or death. Surveillance and preventive measures are an ongoing need to reduce the public health impact of WNV in areas with the potential for transmission. Caren Chancey, Andriyan Grinev, Evgeniya Volkova, and Maria Rios Copyright © 2015 Caren Chancey et al. All rights reserved. Role of Microorganisms Present in Dairy Fermented Products in Health and Disease Thu, 19 Mar 2015 07:11:00 +0000 Clara G. de los Reyes-Gavilán, María Fernández, John Andrew Hudson, and Riitta Korpela Copyright © 2015 Clara G. de los Reyes-Gavilán et al. All rights reserved. Evaluation of the Inflammatory Response in Macrophages Stimulated with Exosomes Secreted by Mycobacterium avium-Infected Macrophages Mon, 16 Mar 2015 12:49:45 +0000 Exosomes secreted from Mycobacterium avium-infected macrophages contain numerous antigens of both M. avium and the host cell and are involved in the induction and expression of the inflammatory responses in macrophages. The interaction between exosomes secreted from M. avium-infected macrophages and macrophage phagocytosis, cytokine secretion, immunostimulation, and apoptosis was analyzed. Upon stimulation with exosomes secreted from M. avium-infected macrophages, the phagocytosis of dextran by treated macrophages was increased. Furthermore, the expression of CD40, CD80, CD81, CD86, HLA-DR, and most notably CD195 was enhanced. Additionally, the secretion of IL-6, IL-8, IL-10, IFN-γ, and TNF-α was increased by stimulated macrophages. Exosome stimulation did not induce macrophage apoptosis when compared with macrophages infected with M. avium. Caspase expression, including that of caspases 3, 6, and 8, was also not altered in exosome stimulated macrophages. Thus exosomes trigger the inflammatory response in macrophages owing to the presence of bacterial antigens but have no effect on macrophage viability. Jianjun Wang, Yongliang Yao, Jing Xiong, Jianhong Wu, Xin Tang, and Guangxin Li Copyright © 2015 Jianjun Wang et al. All rights reserved. Antimicrobial Activity of Lactic Acid Bacteria in Dairy Products and Gut: Effect on Pathogens Mon, 16 Mar 2015 12:09:17 +0000 The food industry seeks alternatives to satisfy consumer demands of safe foods with a long shelf-life able to maintain the nutritional and organoleptic quality. The application of antimicrobial compounds-producing protective cultures may provide an additional parameter of processing in order to improve the safety and ensure food quality, keeping or enhancing its sensorial characteristics. In addition, strong evidences suggest that certain probiotic strains can confer resistance against infection with enteric pathogens. Several mechanisms have been proposed to support this phenomenon, including antimicrobial compounds secreted by the probiotics, competitive exclusion, or stimulation of the immune system. Recent research has increasingly demonstrated the role of antimicrobial compounds as protective mechanism against intestinal pathogens and therefore certain strains could have an effect on both the food and the gut. In this aspect, the effects of the combination of different strains keep unknown. The development of multistrain probiotic dairy products with good technological properties and with improved characteristics to those shown by the individual strains, able to act not only as protective cultures in foods, but also as probiotics able to exert a protective action against infections, has gained increased interest. Juan L. Arqués, Eva Rodríguez, Susana Langa, José María Landete, and Margarita Medina Copyright © 2015 Juan L. Arqués et al. All rights reserved. Biotechnological and Agronomic Potential of Endophytic Pink-Pigmented Methylotrophic Methylobacterium spp. Tue, 10 Mar 2015 11:08:38 +0000 The genus Methylobacterium is composed of pink-pigmented facultative methylotrophic (PPFM) bacteria, which are able to synthesize carotenoids and grow on reduced organic compounds containing one carbon (C1), such as methanol and methylamine. Due to their high phenotypic plasticity, these bacteria are able to colonize different habitats, such as soil, water, and sediment, and different host plants as both endophytes and epiphytes. In plant colonization, the frequency and distribution may be influenced by plant genotype or by interactions with other associated microorganisms, which may result in increasing plant fitness. In this review, different aspects of interactions with the host plant are discussed, including their capacity to fix nitrogen, nodule the host plant, produce cytokinins, auxin and enzymes involved in the induction of systemic resistance, such as pectinase and cellulase, and therefore plant growth promotion. In addition, bacteria belonging to this group can be used to reduce environmental contamination because they are able to degrade toxic compounds, tolerate high heavy metal concentrations, and increase plant tolerance to these compounds. Moreover, genome sequencing and omics approaches have revealed genes related to plant-bacteria interactions that may be important for developing strains able to promote plant growth and protection against phytopathogens. Manuella Nóbrega Dourado, Aline Aparecida Camargo Neves, Daiene Souza Santos, and Welington Luiz Araújo Copyright © 2015 Manuella Nóbrega Dourado et al. All rights reserved. The High PMNs Phagocytosis Resistance of Enterococcal Isolates from RTx Patients Tue, 10 Mar 2015 09:56:24 +0000 Infections caused by opportunistic pathogens such as enterococci remain difficult to manage, especially in immunocompromised patients. Because of infections’ limited symptoms in such patients the additional problems are to find proper diagnostic criteria and the management of infection. Here we aimed to compare the resistance of commensal enterococcal strains and RTx patients’ isolates, to PMNs phagocytosis. Thirty-six enterococcal urine and faecal isolates from RTx patients and 17 faecal isolates from healthy volunteers were cultured in planktonic and biofilm forms in 37°C or 42°C. Another tested variable was the addition of immunosuppressant to the culture media. Bacterial cells were stained with fluorescent reporter (CFDA, PI) and incubated with PMNs. Results of phagocytosis were estimated as a mean fluorescence intensity (MFI) of PMNs using flow cytometry. Commensal enterococci cultured in all abovementioned (37°C and 42°C/the addition of immunosuppressant) conditions were less resistant to phagocytosis compared to RTx isolates. Observed significant difference in phagocytosis resistance suggests that patients in immunosuppression are colonized with high risk strains which may lead to the development of infection. Tomasz Jarzembowski, Agnieszka Daca, Jacek M. Witkowski, Ewa Bryl, and Bolesław Rutkowski Copyright © 2015 Tomasz Jarzembowski et al. All rights reserved. Delivery of Antibiotics from Cementless Titanium-Alloy Cubes May Be a Novel Way to Control Postoperative Infections Tue, 10 Mar 2015 06:17:12 +0000 Bacterial colonisation and biofilm formation onto orthopaedic devices are difficult to eradicate. In most cases infection is treated by surgical removal of the implant and cleaning of the infected area, followed by extensive treatment with broad-spectrum antibiotics. Such treatment causes great discomfort, is expensive, and is not always successful. In this study we report on the release of vancomycin through polyethersulfone membranes from channels in cementless titanium-alloy cubes. The cubes were constructed with LaserCUSING from Ti6Al4V ELI powder. Vancomycin was released by non-Fickian anomalous (constraint) diffusion. Approximately 50% of the vancomycin was released within the first 17 h. However, sustained delivery of vancomycin for 100 h was possible by reinjecting the channels. Refillable implants may be a novel way to control postoperative infections. Martin B. Bezuidenhout, Anton D. van Staden, Gert A. Oosthuizen, Dimitar M. Dimitrov, and Leon M. T. Dicks Copyright © 2015 Martin B. Bezuidenhout et al. All rights reserved. Impact on Human Health of Microorganisms Present in Fermented Dairy Products: An Overview Mon, 09 Mar 2015 09:56:51 +0000 Fermented dairy products provide nutrients in our diet, some of which are produced by the action of microorganisms during fermentation. These products can be populated by a diverse microbiota that impacts the organoleptic and physicochemical characteristics foods as well as human health. Acidification is carried out by starter lactic acid bacteria (LAB) whereas other LAB, moulds, and yeasts become dominant during ripening and contribute to the development of aroma and texture in dairy products. Probiotics are generally part of the nonstarter microbiota, and their use has been extended in recent years. Fermented dairy products can contain beneficial compounds, which are produced by the metabolic activity of their microbiota (vitamins, conjugated linoleic acid, bioactive peptides, and gamma-aminobutyric acid, among others). Some microorganisms can also release toxic compounds, the most notorious being biogenic amines and aflatoxins. Though generally considered safe, fermented dairy products can be contaminated by pathogens. If proliferation occurs during manufacture or storage, they can cause sporadic cases or outbreaks of disease. This paper provides an overview on the current state of different aspects of the research on microorganisms present in dairy products in the light of their positive or negative impact on human health. María Fernández, John Andrew Hudson, Riitta Korpela, and Clara G. de los Reyes-Gavilán Copyright © 2015 María Fernández et al. All rights reserved. Antimicrobial Peptides: Current and Potential Applications in Biomedical Therapies Thu, 05 Mar 2015 13:59:46 +0000 Joel E. López-Meza, Alejandra Ochoa-Zarzosa, José E. Barboza-Corona, and Dennis K. Bideshi Copyright © 2015 Joel E. López-Meza et al. All rights reserved. Incidence and Diversity of Antimicrobial Multidrug Resistance Profiles of Uropathogenic Bacteria Thu, 05 Mar 2015 13:56:21 +0000 The aim of this study was to assess the most frequent multidrug resistant (MDR) profiles of the main bacteria implicated in community-acquired urinary tract infections (UTI). Only the MDR profiles observed in, at least, 5% of the MDR isolates were considered. A quarter of the bacteria were MDR and the most common MDR profile, including resistance to penicillins, quinolones, and sulfonamides (antibiotics with different mechanisms of action, all mainly recommended by the European Association of Urology for empirical therapy of uncomplicated UTI), was observed, alone or in association with resistance to other antimicrobial classes, in the main bacteria implicated in UTI. The penicillin class was included in all the frequent MDR profiles observed in the ten main bacteria and was the antibiotic with the highest prescription during the study period. The sulfonamides class, included in five of the six more frequent MDR profiles, was avoided between 2000 and 2009. The results suggest that the high MDR percentage and the high diversity of MDR profiles result from a high prescription of antibiotics but also from antibiotic-resistant genes transmitted with other resistance determinants on mobile genetic elements and that the UTI standard treatment guidelines must be adjusted for the community of Aveiro District. Inês Linhares, Teresa Raposo, António Rodrigues, and Adelaide Almeida Copyright © 2015 Inês Linhares et al. All rights reserved. The Relationship of CSF and Plasma Cytokine Levels in HIV Infected Patients with Neurocognitive Impairment Tue, 03 Mar 2015 09:19:15 +0000 Although HAD is now rare due to HAART, the milder forms of HAND persist in HIV-infected patients. HIV-induced systemic and localized inflammation is considered to be one of the mechanisms of HAND. The levels of cytokines in CSF were associated with neurocognitive impairment in HIV infection. However, the changes of cytokines involved in cognition impairment in plasma have not been shown, and their relationships between CSF and plasma require to be addressed. We compared cytokine levels in paired CSF and plasma samples from HIV-infected individuals with or without neurocognitive impairment. Cytokine concentrations were measured by Luminex xMAP. In comparing the expression levels of cytokines in plasma and CSF, IFN-α2, IL-8, IP-10, and MCP-1 were significantly higher in CSF. Eotaxin was significantly higher in plasma, whereas G-CSF showed no difference between plasma and CSF. G-CSF , IL-8 , IP-10 , and MCP-1 in CSF showed significant difference between HIV-CI and HIV-NC group, which may indicate their relationship to HIV associated neurocognitive impairment. In addition, G-CSF and IP-10 in plasma were significantly higher in HIV-CI than HIV-NC. The consistent changes of G-CSF and IP-10 in paired plasma and CSF samples might enhance their potential for predicting HAND. Lin Yuan, An Liu, Luxin Qiao, Bo Sheng, Meng Xu, Wei Li, and Dexi Chen Copyright © 2015 Lin Yuan et al. All rights reserved. cspA Influences Biofilm Formation and Drug Resistance in Pathogenic Fungus Aspergillus fumigatus Tue, 03 Mar 2015 06:09:26 +0000 The microbial cell wall plays a crucial role in biofilm formation and drug resistance. cspA encodes a repeat-rich glycophosphatidylinositol-anchored cell wall protein in the pathogenic fungus Aspergillus fumigatus. To determine whether cspA has a significant impact on biofilm development and sensitivity to antifungal drugs in A. fumigatus, a ΔcspA mutant was constructed by targeted gene disruption, and we then reconstituted the mutant to wild type by homologous recombination of a functional cspA gene. Deletion of cspA resulted in a rougher conidial surface, reduced biofilm formation, decreased resistance to antifungal agents, and increased internalization by A549 human lung epithelial cells, suggesting that cspA not only participates in maintaining the integrity of the cell wall, but also affects biofilm establishment, drug response, and invasiveness of A. fumigatus. Zhongqi Fan, Zhe Li, Zongge Xu, Hongyan Li, Lixiang Li, Cong Ning, Lin Ma, Xiangli Xie, Guangyi Wang, and Huimei Yu Copyright © 2015 Zhongqi Fan et al. All rights reserved. Proteomic Analysis of Human Brain Microvascular Endothelial Cells Reveals Differential Protein Expression in Response to Enterovirus 71 Infection Mon, 02 Mar 2015 12:49:57 +0000 2D DIGE technology was employed on proteins prepared from human brain microvascular endothelial cells (HBMEC), to study the differentially expressed proteins in cells at 0 h, 1 h, 16 h, and 24 h after infection. Proteins found to be differentially expressed were identified with matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDITOF/TOF MS) analysis. We identified 43 spots showing changes of at least 2.5 fold up- or downregulated expressions in EV71-infected cells at different time when comparing to control, and 28 proteins could be successfully identified by MALDI TOF/TOF mass spectrometry analysis. 4 proteins were significantly upregulated, and 6 proteins were downregulated, another 18 proteins were different expression at different incubation time. We identified changes in the expression of 12 cellular metabolism-related proteins, 5 molecules involved in cytoskeleton, 3 molecules involved in energy metabolism, 2 molecules involved in signal transduction, 1 molecule involved in the ubiquitin-proteasome pathway, 1 molecule involved in cell cycle, 1 molecule involved in apoptosis-related protein, 1 molecular chaperone, and 2 unknown proteins. These findings build up a comprehensive profile of the HBMEC proteome and provide a useful basis for further analysis of the pathogenic mechanism that underlies EV71 infections to induce severe neural complications. Wenying Luo, Jiayu Zhong, Wei Zhao, Jianjun Liu, Renli Zhang, Liang Peng, Wenxu Hong, Sheng He Huang, and Hong Cao Copyright © 2015 Wenying Luo et al. All rights reserved. Cytotoxicity of Cyclodipeptides from Pseudomonas aeruginosa PAO1 Leads to Apoptosis in Human Cancer Cell Lines Mon, 02 Mar 2015 12:26:15 +0000 Pseudomonas aeruginosa is an opportunistic pathogen of plants and animals, which produces virulence factors in order to infect or colonize its eukaryotic hosts. Cyclodipeptides (CDPs) produced by P. aeruginosa exhibit cytotoxic properties toward human tumor cells. In this study, we evaluated the effect of a CDP mix, comprised of cyclo(L-Pro-L-Tyr), cyclo(L-Pro-L-Val), and cyclo(L-Pro-L-Phe) that were isolated from P. aeruginosa, on two human cancer cell lines. Our results demonstrated that the CDP mix promoted cell death in cultures of the HeLa cervical adenocarcinoma and Caco-2 colorectal adenocarcinoma cell lines in a dose-dependent manner, with a 50% inhibitory concentration (IC50) of 0.53 and 0.66 mg/mL, for HeLa and Caco-2 cells, respectively. Flow cytometric analysis, using annexin V and propidium iodide as apoptosis and necrosis indicators, respectively, clearly showed that HeLa and Caco-2 cells exhibited apoptotic characteristics when treated with the CDP mix at a concentration <0.001 mg/mL. IC50 values for apoptotic cells in HeLa and Caco-2 cells were 6.5 × 10−5 and 1.8 × 10−4 mg/mL, respectively. Our results indicate that an apoptotic pathway is involved in the inhibition of cell proliferation caused by the P. aeruginosa CDP mix. Dolores Vázquez-Rivera, Omar González, Jaquelina Guzmán-Rodríguez, Alma L. Díaz-Pérez, Alejandra Ochoa-Zarzosa, José López-Bucio, Víctor Meza-Carmen, and Jesús Campos-García Copyright © 2015 Dolores Vázquez-Rivera et al. All rights reserved. Cloning and Expression of Synthetic Genes Encoding the Broad Antimicrobial Spectrum Bacteriocins SRCAM 602, OR-7, E-760, and L-1077, by Recombinant Pichia pastoris Mon, 02 Mar 2015 07:12:26 +0000 We have evaluated the cloning and functional expression of previously described broad antimicrobial spectrum bacteriocins SRCAM 602, OR-7, E-760, and L-1077, by recombinant Pichia pastoris. Synthetic genes, matching the codon usage of P. pastoris, were designed from the known mature amino acid sequence of these bacteriocins and cloned into the protein expression vector pPICZαA. The recombinant derived plasmids were linearized and transformed into competent P. pastoris X-33, and the presence of integrated plasmids into the transformed cells was confirmed by PCR and sequencing of the inserts. The antimicrobial activity, expected in supernatants of the recombinant P. pastoris producers, was purified using a multistep chromatographic procedure including ammonium sulfate precipitation, desalting by gel filtration, cation exchange-, hydrophobic interaction-, and reverse phase-chromatography (RP-FPLC). However, a measurable antimicrobial activity was only detected after the hydrophobic interaction and RP-FPLC steps of the purified supernatants. MALDI-TOF MS analysis of the antimicrobial fractions eluted from RP-FPLC revealed the existence of peptide fragments of lower and higher molecular mass than expected. MALDI-TOF/TOF MS analysis of selected peptides from eluted RP-FPLC samples with antimicrobial activity indicated the presence of peptide fragments not related to the amino acid sequence of the cloned bacteriocins. Sara Arbulu, Juan J. Jiménez, Loreto Gútiez, Luis M. Cintas, Carmen Herranz, and Pablo E. Hernández Copyright © 2015 Sara Arbulu et al. All rights reserved. Dengue Patients Exhibit Higher Levels of PrM and E Antibodies Than Their Asymptomatic Counterparts Sun, 01 Mar 2015 14:03:38 +0000 Dengue virus infection is a common tropical disease which often occurs without being detected. These asymptomatic cases provide information in relation to the manifestation of immunological aspects. In this study, we developed an ELISA method to compare neutralizing effects of dengue prM and E antibodies between dengue patients and their asymptomatic household members. Recombinant D2 premembrane (prM) was constructed, cloned, and tested for antigenicity. The recombinant protein was purified and tested with controls by using an indirect ELISA method. Positive dengue serum samples with their asymptomatic pair were then carried out onto the developed ELISA. In addition, commercially available recombinant envelope (E) protein was used to develop an ELISA which was tested with the same set of serum samples in the prM ELISA. Asymptomatic individuals showed preexisting heterotypic neutralizing antibodies. The recombinant prM was antigenically reactive in the developed ELISA. Dengue patients had higher prM and E antibodies compared to their household members. Our study highlights the neutralizing antibodies levels with respect to dengue prM and E between dengue patients and asymptomatic individuals. Adeline Syin Lian Yeo, Anusyah Rathakrishnan, Seok Mui Wang, Sasheela Ponnampalavanar, Rishya Manikam, Jameela Sathar, Santha Kumari Natkunam, and Shamala Devi Sekaran Copyright © 2015 Adeline Syin Lian Yeo et al. All rights reserved. Current and Potential Applications of Host-Defense Peptides and Proteins in Urology Sun, 01 Mar 2015 13:05:52 +0000 The use of antibiotics has become increasingly disfavored as more multidrug resistant pathogens are on the rise. A promising alternative to the use of these conventional drugs includes antimicrobial peptides or host-defense peptides. These peptides typically consist of short amino acid chains with a net cationic charge and a substantial portion of hydrophobic residues. They mainly target the bacterial cell membrane but are also capable of translocating through the membrane and target intracellular components, making it difficult for bacteria to gain resistance as multiple essential cellular processes are being targeted. The use of these peptides in the field of biomedical therapies has been examined, and the different approaches to using them under various settings are constantly being discovered. In this review, we discuss the current and potential applications of these host-defense peptides in the field of urology. Besides the use of these peptides as antimicrobial agents, the value of these biological molecules has recently been expanded to their use as antitumor and anti-kidney-stone agents. Joey Chor Yee Lo and Dirk Lange Copyright © 2015 Joey Chor Yee Lo and Dirk Lange. All rights reserved. Molecular Detection and Sensitivity to Antibiotics and Bacteriocins of Pathogens Isolated from Bovine Mastitis in Family Dairy Herds of Central Mexico Sun, 01 Mar 2015 12:39:47 +0000 Thirty-two farms ( cows) located in the state of Guanajuato, Mexico, were sampled. Pathogens from bovine subclinical mastitis (SCM) and clinical mastitis (CLM) were identified by 16S rDNA and the sensitivity to both antibiotics and bacteriocins of Bacillus thuringiensis was tested. Forty-six milk samples were selected for their positive California Mastitis Test (CMT) (≥3) and any abnormality in the udder or milk. The frequency of SCM and CLM was 39.1% and 9.3%, respectively. Averages for test day milk yield (MY), lactation number (LN), herd size (HS), and number of days in milk (DM) were 20.6 kg, 2.8 lactations, 16.7 animals, and 164.1 days, respectively. MY was dependent on dairy herd (DH), LN, HS, and DM , and correlations between udder quarters from the CMT were around 0.49 . Coagulase-negative staphylococci were mainly identified, as well as Staphylococcus aureus, Streptococcus uberis, Brevibacterium stationis, B. conglomeratum, and Staphylococcus agnetis. Bacterial isolates were resistant to penicillin, clindamycin, ampicillin, and cefotaxime. Bacteriocins synthesized by Bacillus thuringiensis inhibited the growth of multiantibiotic resistance bacteria such as S. agnetis, S. equorum, Streptococcus uberis, Brevibacterium stationis, and Brachybacterium conglomeratum, but they were not active against S. sciuri, a microorganism that showed an 84% resistance to antibiotics tested in this study. Ma. Fabiola León-Galván, José E. Barboza-Corona, A. Arianna Lechuga-Arana, Mauricio Valencia-Posadas, Daniel D. Aguayo, Carlos Cedillo-Pelaez, Erika A. Martínez-Ortega, and Abner J. Gutierrez-Chavez Copyright © 2015 Ma. Fabiola León-Galván et al. All rights reserved. Biologically Active and Antimicrobial Peptides from Plants Sun, 01 Mar 2015 12:31:25 +0000 Bioactive peptides are part of an innate response elicited by most living forms. In plants, they are produced ubiquitously in roots, seeds, flowers, stems, and leaves, highlighting their physiological importance. While most of the bioactive peptides produced in plants possess microbicide properties, there is evidence that they are also involved in cellular signaling. Structurally, there is an overall similarity when comparing them with those derived from animal or insect sources. The biological action of bioactive peptides initiates with the binding to the target membrane followed in most cases by membrane permeabilization and rupture. Here we present an overview of what is currently known about bioactive peptides from plants, focusing on their antimicrobial activity and their role in the plant signaling network and offering perspectives on their potential application. Carlos E. Salas, Jesus A. Badillo-Corona, Guadalupe Ramírez-Sotelo, and Carmen Oliver-Salvador Copyright © 2015 Carlos E. Salas et al. All rights reserved. Antibacterial Activity of Synthetic Peptides Derived from Lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 Sun, 01 Mar 2015 12:13:25 +0000 Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)4K2Ahx2C2) exhibit bigger or similar activity against E. coli (MIC 4–33 μM) and E. faecalis (MIC 10–33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield. María A. León-Calvijo, Aura L. Leal-Castro, Giovanni A. Almanzar-Reina, Jaiver E. Rosas-Pérez, Javier E. García-Castañeda, and Zuly J. Rivera-Monroy Copyright © 2015 María A. León-Calvijo et al. All rights reserved. Ascorbic Acid, Ultraviolet C Rays, and Glucose but not Hyperthermia Are Elicitors of Human β-Defensin 1 mRNA in Normal Keratinocytes Sun, 01 Mar 2015 10:06:07 +0000 Hosts’ innate defense systems are upregulated by antimicrobial peptide elicitors (APEs). Our aim was to investigate the effects of hyperthermia, ultraviolet A rays (UVA), and ultraviolet C rays (UVC) as well as glucose and ascorbic acid (AA) on the regulation of human β-defensin 1 (DEFB1), cathelicidin (CAMP), and interferon-γ (IFNG) genes in normal human keratinocytes (NHK). The indirect in vitro antimicrobial activity against Staphylococcus aureus and Listeria monocytogenes of these potential APEs was tested. We found that AA is a more potent APE for DEFB1 than glucose in NHK. Glucose but not AA is an APE for CAMP. Mild hypo- (35°C) and hyperthermia (39°C) are not APEs in NHK. AA-dependent DEFB1 upregulation below 20 mM predicts in vitro antimicrobial activity as well as glucose- and AA-dependent CAMP and IFNG upregulation. UVC upregulates CAMP and DEFB1 genes but UVA only upregulates the DEFB1 gene. UVC is a previously unrecognized APE in human cells. Our results suggest that glucose upregulates CAMP in an IFN-γ-independent manner. AA is an elicitor of innate immunity that will challenge the current concept of late activation of adaptive immunity of this vitamin. These results could be useful in designing new potential drugs and devices to combat skin infections. Luis Antonio Cruz Díaz, María Guadalupe Flores Miramontes, Paulina Chávez Hurtado, Kirk Allen, Marisela Gonzalez Ávila, and Ernesto Prado Montes de Oca Copyright © 2015 Luis Antonio Cruz Díaz et al. All rights reserved. Plant Antimicrobial Peptides as Potential Anticancer Agents Sun, 01 Mar 2015 09:57:02 +0000 Antimicrobial peptides (AMPs) are part of the innate immune defense mechanism of many organisms and are promising candidates to treat infections caused by pathogenic bacteria to animals and humans. AMPs also display anticancer activities because of their ability to inactivate a wide range of cancer cells. Cancer remains a cause of high morbidity and mortality worldwide. Therefore, the development of methods for its control is desirable. Attractive alternatives include plant AMP thionins, defensins, and cyclotides, which have anticancer activities. Here, we provide an overview of plant AMPs anticancer activities, with an emphasis on their mode of action, their selectivity, and their efficacy. Jaquelina Julia Guzmán-Rodríguez, Alejandra Ochoa-Zarzosa, Rodolfo López-Gómez, and Joel E. López-Meza Copyright © 2015 Jaquelina Julia Guzmán-Rodríguez et al. All rights reserved. Effect of Recombinant Prophenin 2 on the Integrity and Viability of Trichomonas vaginalis Sun, 01 Mar 2015 08:12:34 +0000 Trichomonas vaginalis is the causal agent of trichomoniasis, which is associated with preterm child delivery, low birth weight, and an increased risk of infection by human papilloma virus and human immunodeficiency virus following exposure. Several reports have established increasing numbers of trichomoniasis cases resistant to metronidazole, the agent used for treatment, and it is therefore important to identify new therapeutic alternatives. Previously, our group reported the effect of tritrpticin, a synthetic peptide derived from porcine prophenin, on T. vaginalis; however, the hemolytic activity of this small peptide complicates its possible use as a therapeutic agent. In this study, we report that the propeptide and the processed peptide of prophenin 2 (cleaved with hydroxylamine) affected the integrity and growth of T. vaginalis and that pro-prophenin 2 displays some resistance to proteolysis by T. vaginalis proteinases at 1 h. Its effect on T. vaginalis as well as its low hemolytic activity and short-time stability to parasite proteinases makes prophenin 2 an interesting candidate for synergistic or alternative treatment against T. vaginalis. J. L. Hernandez-Flores, M. C. Rodriguez, A. Gastelum Arellanez, A. Alvarez-Morales, and E. E. Avila Copyright © 2015 J. L. Hernandez-Flores et al. All rights reserved. Bioaccessible Antioxidants in Milk Fermented by Bifidobacterium longum subsp. longum Strains Mon, 23 Feb 2015 13:44:54 +0000 Bifidobacterium longum subsp. longum is among the dominant species of the human gastrointestinal microbiota and could thus have potential as probiotics. New targets such as antioxidant properties have interest for beneficial effects on health. The objective of this study was to evaluate the bioaccessibility of antioxidants in milk fermented by selected B. longum subsp. longum strains during in vitro dynamic digestion. The antioxidant capacity of cell extracts from 38 strains, of which 32 belong to B. longum subsp. longum, was evaluated with the ORAC (oxygen radical absorbance capacity) method. On the basis of screening and gene sequence typing by multilocus locus sequence analysis (MLSA), five strains were chosen for fermenting reconstituted skim milk. Antioxidant capacity varied among the strains tested (). Two strains of B. longum subsp. longum (CUETM 172 and 171) showed significantly higher ORAC values than the other bifidobacteria strains. However, there does not appear to be a relationship between gene sequence types and antioxidant capacity. The milk fermented by each of the five strains selected (CUETM 268, 172, 245, 247, or PRO 16-10) did not have higher initial ORAC values compared to the nonfermented milk samples. However, higher bioaccessibility of antioxidants in fermented milk (175–358%) was observed during digestion. Mérilie Gagnon, Patricia Savard, Audrey Rivière, Gisèle LaPointe, and Denis Roy Copyright © 2015 Mérilie Gagnon et al. All rights reserved. Biodiversity and γ-Aminobutyric Acid Production by Lactic Acid Bacteria Isolated from Traditional Alpine Raw Cow’s Milk Cheeses Mon, 23 Feb 2015 13:34:17 +0000 “Nostrano-cheeses” are traditional alpine cheeses made from raw cow’s milk in Trentino-Alto Adige, Italy. This study identified lactic acid bacteria (LAB) developing during maturation of “Nostrano-cheeses” and evaluated their potential to produce γ-aminobutyric acid (GABA), an immunologically active compound and neurotransmitter. Cheese samples were collected on six cheese-making days, in three dairy factories located in different areas of Trentino and at different stages of cheese ripening (24 h, 15 days, and 1, 2, 3, 6, and 8 months). A total of 1,059 LAB isolates were screened using Random Amplified Polymorphic DNA-PCR (RAPD-PCR) and differentiated into 583 clusters. LAB strains from dominant clusters () were genetically identified to species level by partial 16S rRNA gene sequencing. LAB species most frequently isolated were Lactobacillus paracasei, Streptococcus thermophilus, and Leuconostoc mesenteroides. The 97 dominant clusters were also characterized for their ability in producing GABA by high-performance liquid chromatography (HPLC). About 71% of the dominant bacteria clusters evolving during cheeses ripening were able to produce GABA. Most GABA producers were Lactobacillus paracasei but other GABA producing species included Lactococcus lactis, Lactobacillus plantarum, Lactobacillus rhamnosus, Pediococcus pentosaceus, and Streptococcus thermophilus. No Enterococcus faecalis or Sc. macedonicus isolates produced GABA. The isolate producing the highest amount of GABA ( mg/kg) was a Sc. thermophilus. Elena Franciosi, Ilaria Carafa, Tiziana Nardin, Silvia Schiavon, Elisa Poznanski, Agostino Cavazza, Roberto Larcher, and Kieran M. Tuohy Copyright © 2015 Elena Franciosi et al. All rights reserved. Biocheese: A Food Probiotic Carrier Mon, 23 Feb 2015 09:23:54 +0000 This review describes some aspects related to the technological barriers encountered in the development and stability of probiotic cheeses. Aspects concerning the viability of probiotic cultures in this matrix are discussed and the potential of cheese as a biofunctional food carrier is analyzed, outlying some points related to health and safety. In general, the manufacture of probiotic cheese should have little change when compared with the elaboration of cheese in the traditional way. The physicochemical and technological parameters influencing the quality of these products have also to be measured so as to obtain a process optimization. J. M. Castro, M. E. Tornadijo, J. M. Fresno, and H. Sandoval Copyright © 2015 J. M. Castro et al. All rights reserved. Clostridium butyricum Combined with Bifidobacterium infantis Probiotic Mixture Restores Fecal Microbiota and Attenuates Systemic Inflammation in Mice with Antibiotic-Associated Diarrhea Mon, 23 Feb 2015 08:08:00 +0000 Antibiotic-associated diarrhea (AAD) is one of the most common complications of most types of antibiotics. Our aim was to determine the efficacy of Clostridium butyricum, Bifidobacterium infantis, and their mixture for AAD treatment in mice. AAD models were administered with single probiotic strain and probiotic mixture for short term and long term to evaluate the changes of the composition and diversity of intestinal microbiota, histopathology of the colon, and the systemic inflammation. Our data indicated that long-term probiotic therapy, but not short-term course, exerted beneficial effects on the restoration of the intestinal microbiota, the recovery of the tissue architecture, and attenuation of systemic inflammation. All predominant fecal bacteria reached normal level after the long-term probiotic mixture treatment, while IL-10, IFN-γ, and TNF-α also returned to normal level. However, the efficacy for AAD was time dependent and probiotic strain specific. Short-term administration of probiotic strains or mixture showed no apparent positive effects for AAD. In addition, the beneficial effects of C. butyricum combined with B. infantis probiotic mixture were superior to their single strain. This research showed that supplementation with C. butyricum combined with B. infantis probiotic mixture may be a simple and effective method for AAD treatment. Zongxin Ling, Xia Liu, Yiwen Cheng, Yueqiu Luo, Li Yuan, Lanjuan Li, and Charlie Xiang Copyright © 2015 Zongxin Ling et al. All rights reserved. Bioavailability of Dietary Polyphenols and Gut Microbiota Metabolism: Antimicrobial Properties Mon, 23 Feb 2015 07:01:38 +0000 Polyphenolic compounds are plant nutraceuticals showing a huge structural diversity, including chlorogenic acids, hydrolyzable tannins, and flavonoids (flavonols, flavanones, flavan-3-ols, anthocyanidins, isoflavones, and flavones). Most of them occur as glycosylated derivatives in plants and foods. In order to become bioactive at human body, these polyphenols must undergo diverse intestinal transformations, due to the action of digestive enzymes, but also by the action of microbiota metabolism. After elimination of sugar tailoring (generating the corresponding aglycons) and diverse hydroxyl moieties, as well as further backbone reorganizations, the final absorbed compounds enter the portal vein circulation towards liver (where other enzymatic transformations take place) and from there to other organs, including behind the digestive tract or via blood towards urine excretion. During this transit along diverse tissues and organs, they are able to carry out strong antiviral, antibacterial, and antiparasitic activities. This paper revises and discusses these antimicrobial activities of dietary polyphenols and their relevance for human health, shedding light on the importance of polyphenols structure recognition by specific enzymes produced by intestinal microbial taxa. Laura Marín, Elisa M. Miguélez, Claudio J. Villar, and Felipe Lombó Copyright © 2015 Laura Marín et al. All rights reserved. Isolation and Molecular Characterization of Brucella Isolates in Cattle Milk in Uganda Sun, 22 Feb 2015 11:48:15 +0000 Brucellosis is endemic in livestock and humans in Uganda and its transmission involves a multitude of risk factors like consumption of milk from infected cattle. To shed new light on the epidemiology of brucellosis in Uganda the present study used phenotypic and molecular approaches to delineate the Brucella species, biovars, and genotypes shed in cattle milk. Brucella abortus without a biovar designation was isolated from eleven out of 207 milk samples from cattle in Uganda. These isolates had a genomic monomorphism at 16 variable number tandem repeat (VNTR) loci and showed in turn high levels of genetic variation when compared with other African strains or other B. abortus biovars from other parts of the world. This study further highlights the usefulness of MLVA as an epidemiological tool for investigation of Brucella infections. Denis Rwabiita Mugizi, Shaman Muradrasoli, Sofia Boqvist, Joseph Erume, George William Nasinyama, Charles Waiswa, Gerald Mboowa, Markus Klint, and Ulf Magnusson Copyright © 2015 Denis Rwabiita Mugizi et al. All rights reserved. The Role of Probiotic Lactic Acid Bacteria and Bifidobacteria in the Prevention and Treatment of Inflammatory Bowel Disease and Other Related Diseases: A Systematic Review of Randomized Human Clinical Trials Sun, 22 Feb 2015 08:46:39 +0000 Inflammatory bowel disease (IBD), which includes Crohn’s disease (CD) and ulcerative colitis (UC), is a chronic inflammation of the small intestine and colon caused by a dysregulated immune response to host intestinal microbiota in genetically susceptible subjects. A number of fermented dairy products contain lactic acid bacteria (LAB) and bifidobacteria, some of which have been characterized as probiotics that can modify the gut microbiota and may be beneficial for the treatment and the prevention of IBD. The objective of this review was to carry out a systematic search of LAB and bifidobacteria probiotics and IBD, using the PubMed and Scopus databases, defined by a specific equation using MeSH terms and limited to human clinical trials. The use of probiotics and/or synbiotics has positive effects in the treatment and maintenance of UC, whereas in CD clear effectiveness has only been shown for synbiotics. Furthermore, in other associated IBD pathologies, such as pouchitis and cholangitis, LAB and bifidobacteria probiotics can provide a benefit through the improvement of clinical symptoms. However, more studies are needed to understand their mechanisms of action and in this way to understand the effect of probiotics prior to their use as coadjuvants in the therapy and prevention of IBD conditions. Maria Jose Saez-Lara, Carolina Gomez-Llorente, Julio Plaza-Diaz, and Angel Gil Copyright © 2015 Maria Jose Saez-Lara et al. All rights reserved. A Survey of Modulation of Gut Microbiota by Dietary Polyphenols Sun, 22 Feb 2015 08:12:46 +0000 Dietary polyphenols present in a broad range of plant foods have been related to beneficial health effects. This review aims to update the current information about the modulation of the gut microbiota by dietary phenolic compounds, from a perspective based on the experimental approaches used. After referring to general aspects of gut microbiota and dietary polyphenols, studies related to this topic are presented according to their experimental design: batch culture fermentations, gastrointestinal simulators, animal model studies, and human intervention studies. In general, studies evidence that dietary polyphenols may contribute to the maintenance of intestinal health by preserving the gut microbial balance through the stimulation of the growth of beneficial bacteria (i.e., lactobacilli and bifidobacteria) and the inhibition of pathogenic bacteria, exerting prebiotic-like effects. Combination of in vitro and in vivo models could help to understand the underlying mechanisms in the polyphenols-microbiota-host triangle and elucidate the implications of polyphenols on human health. From a technological point of view, supplementation with rich-polyphenolic stuffs (phenolic extracts, phenolic-enriched fractions, etc.) could be an effective option to improve health benefits of functional foods such as the case of dairy fermented foods. Montserrat Dueñas, Irene Muñoz-González, Carolina Cueva, Ana Jiménez-Girón, Fernando Sánchez-Patán, Celestino Santos-Buelga, M. Victoria Moreno-Arribas, and Begoña Bartolomé Copyright © 2015 Montserrat Dueñas et al. All rights reserved. The Effects of Bifidobacterium breve on Immune Mediators and Proteome of HT29 Cells Monolayers Sun, 22 Feb 2015 06:20:57 +0000 The use of beneficial microorganisms, the so-called probiotics, to improve human health is gaining popularity. However, not all of the probiotic strains trigger the same responses and they differ in their interaction with the host. In spite of the limited knowledge on mechanisms of action some of the probiotic effects seem to be exerted through maintenance of the gastrointestinal barrier function and modulation of the immune system. In the present work, we have addressed in vitro the response of the intestinal epithelial cell line HT29 to the strain Bifidobacterium breve IPLA20004. In the array of 84 genes involved in inflammation tested, the expression of 12 was modified by the bifidobacteria. The genes of chemokine CXCL6, the chemokine receptor CCR7, and, specially, the complement component C3 were upregulated. Indeed, HT29 cells cocultivated with B. breve produced significantly higher levels of protein C3a. The proteome of HT29 cells showed increased levels of cytokeratin-8 in the presence of B. breve. Altogether, it seems that B. breve IPLA20004 could favor the recruitment of innate immune cells to the mucosa reinforcing, as well as the physical barrier of the intestinal epithelium. Borja Sánchez, Irene González-Rodríguez, Silvia Arboleya, Patricia López, Ana Suárez, Patricia Ruas-Madiedo, Abelardo Margolles, and Miguel Gueimonde Copyright © 2015 Borja Sánchez et al. All rights reserved. Determination of the In Vitro and In Vivo Antimicrobial Activity on Salivary Streptococci and Lactobacilli and Chemical Characterisation of the Phenolic Content of a Plantago lanceolata Infusion Thu, 12 Feb 2015 09:30:30 +0000 Introduction. Plant extracts may be suitable alternative treatments for caries. Aims. To investigate the in vitro and in vivo antimicrobial effects of Plantago lanceolata herbal tea (from flowers and leaves) on cariogenic bacteria and to identify the major constituents of P. lanceolata plant. Materials and Methods. The MIC and MBC against cariogenic bacteria were determined for P. lanceolata tea. Subsequently, a controlled random clinical study was conducted. Group A was instructed to rinse with a P. lanceolata mouth rinse, and Group B received a placebo mouth rinse for seven days. The salivary colonisation by streptococci and lactobacilli was investigated prior to treatment and on the fourth and seventh days. Finally, the P. lanceolata tea was analysed for its polyphenolic content, and major phenolics were identified. Results and Discussion. P. lanceolata teas demonstrate good in vitro antimicrobial activity. The in vivo test showed that Group A subjects presented a significant decrease in streptococci compared to Group B. The phytochemical analysis revealed that flavonoids, coumarins, lipids, cinnamic acids, lignans, and phenolic compounds are present in P. lanceolata infusions. Conclusions. P. lanceolata extract could represent a natural anticariogenic agent via an antimicrobial effect and might be useful as an ancillary measure to control the proliferation of cariogenic flora. Gianmaria Fabrizio Ferrazzano, Tiziana Cantile, Lia Roberto, Aniello Ingenito, Maria Rosaria Catania, Emanuela Roscetto, Giuseppe Palumbo, Armando Zarrelli, and Antonino Pollio Copyright © 2015 Gianmaria Fabrizio Ferrazzano et al. All rights reserved. Evaluation of Hemagglutination Activity of Chitosan Nanoparticles Using Human Erythrocytes Wed, 11 Feb 2015 14:10:20 +0000 Chitosan is a polysaccharide composed of randomly distributed chains of β-(1-4) D-glucosamine and N-acetyl-D-glucosamine. This compound is obtained by partial or total deacetylation of chitin in acidic solution. The chitosan-based hemostatic agents have been gaining much attention in the management of bleeding. The aim of this study was to evaluate in vitro hemagglutination activity of chitosan nanoparticles using human erythrocytes. The preparation of nanoparticles was achieved by ionotropic gelification technique followed by neutralization with NaOH 1 mol/L−1. The hemagglutination activity was performed on a solution of 2% erythrocytes (pH 7.4 on PBS) collected from five healthy volunteers. The hemolysis determination was made by spectrophotometric analysis. Chitosan nanoparticle solutions without NaOH addition changed the reddish colour of the wells into brown, suggesting an oxidative reaction of hemoglobin and possible cell lysis. All neutralized solutions of chitosan nanoparticles presented positive haemagglutination, without any change in reaction color. Chitosan nanoparticles presented hemolytic activity ranging from 186.20 to 223.12%, while neutralized solutions ranged from 2.56 to 72.54%, comparing to distilled water. Results highlight the need for development of new routes of synthesis of chitosan nanoparticles within human physiologic pH. Jefferson Muniz de Lima, Ronaldo Rodrigues Sarmento, Joelma Rodrigues de Souza, Fábio André Brayner, Ana Paula Sampaio Feitosa, Rafael Padilha, Luiz Carlos Alves, Isaque Jerônimo Porto, Roberta Ferreti Bonan Dantas Batista, Juliano Elvis de Oliveira, Eliton Souto de Medeiros, Paulo Rogério Ferreti Bonan, and Lúcio Roberto Castellano Copyright © 2015 Jefferson Muniz de Lima et al. All rights reserved. Endophytic Bacillus subtilis Strain E1R-J Is a Promising Biocontrol Agent for Wheat Powdery Mildew Wed, 11 Feb 2015 06:01:49 +0000 In this study, the biocontrol efficacies of 14 endophytic bacterial strains were tested against Blumeria graminis f. sp. tritici (Bgt) in pot experiments under greenhouse conditions. Bacillus subtilis strain E1R-j significantly reduced disease index and exhibited the best control (90.97%). When different formulations of E1R-j were sprayed 24 h before Bgt inoculation, fermentation liquid without bacterial cell and crude protein suspension displayed the similar effects; and they reduced disease index more than bacterial cell suspension (109 cfu mL−1) and fermentation liquid without protein. The control effects were not significantly different between 1011 and 109 cfu mL−1 of bacterial cell suspension but were higher than 107 cfu mL−1. Further observations showed that conidial germination and appressorial formation of Bgt were retarded by spraying E1R-j 24 h before Bgt inoculation. Compared with the water check, conidial germination and appressorial formation were decreased by 43.3% and 42.7%, respectively. In the treatment with E1R-j, the number of houstoria significantly reduced and the speed of mycelial extension was slowed down in the wheat leaves. Scanning electron microscopy observation revealed that E1R-j significantly suppressed the conidial germination and caused rupture and deformation of germ tubes. On the surface of wheat leaves, mycelia and conidiophores became shrinking. Xiaoning Gao, Yufei Gong, Yunxia Huo, Qingmei Han, Zhensheng Kang, and Lili Huang Copyright © 2015 Xiaoning Gao et al. All rights reserved. Comparative Analysis of Quinolone Resistance in Clinical Isolates of Klebsiella pneumoniae and Escherichia coli from Chinese Children and Adults Tue, 10 Feb 2015 12:54:23 +0000 The objective of this study was to compare quinolone resistance and gyrA mutations in clinical isolates of Klebsiella pneumoniae and Escherichia coli from Chinese adults who used quinolone in the preceding month and children without any known history of quinolone administration. The antimicrobial susceptibilities of 61 isolates from children and 79 isolates from adults were determined. The mutations in the quinolone resistance-determining regions in gyrA gene were detected by PCR and DNA sequencing. Fluoroquinolone resistance and types of gyrA mutations in isolates from children and adults were compared and statistically analyzed. No significant differences were detected in the resistance rates of ciprofloxacin and levofloxacin between children and adults among isolates of the two species (all ). The double mutation Ser83Leu + Asp87Asn in the ciprofloxacin-resistant isolates occurred in 73.7% isolates from the children and 67.9% from the adults, respectively (). Children with no known history of quinolone administration were found to carry fluoroquinolone-resistant Enterobacteriaceae isolates. The occurrence of ciprofloxacin resistance and the major types of gyrA mutations in the isolates from the children were similar to those from adults. The results indicate that precautions should be taken on environmental issues resulting from widespread transmission of quinolone resistance. Ying Huang, James O. Ogutu, Jiarui Gu, Fengshu Ding, Yuhong You, Yan Huo, Hong Zhao, Wenjing Li, Zhiwei Zhang, Wenli Zhang, Xiaobei Chen, Yingmei Fu, and Fengmin Zhang Copyright © 2015 Ying Huang et al. All rights reserved. Evaluation of Antibacterial Activity of Some Traditionally Used Medicinal Plants against Human Pathogenic Bacteria Mon, 09 Feb 2015 06:28:10 +0000 The worldwide increase of multidrug resistance in both community- and health-care associated bacterial infections has impaired the current antimicrobial therapy, warranting the search for other alternatives. We aimed to find the in vitro antibacterial activity of ethanolic extracts of 16 different traditionally used medicinal plants of Nepal against 13 clinical and 2 reference bacterial species using microbroth dilution method. The evaluated plants species were found to exert a range of in vitro growth inhibitory action against the tested bacterial species, and Cynodon dactylon was found to exhibit moderate inhibitory action against 13 bacterial species including methicillin-resistant Staphylococcus aureus, imipenem-resistant Pseudomonas aeruginosa, multidrug-resistant Salmonella typhi, and S. typhimurium. The minimum inhibitory concentration (MIC) values of tested ethanolic extracts were found from 31 to >25,000 μg/mL. Notably, ethanolic extracts of Cinnamomum camphora, Curculigo orchioides, and Curcuma longa exhibited the highest antibacterial activity against S. pyogenes with a MIC of 49, 49, and 195 μg/mL, respectively; whereas chloroform fraction of Cynodon dactylon exhibited best antibacterial activity against S. aureus with a MIC of 31 μg/mL. Among all, C. dactylon, C. camphora, C. orchioides, and C. longa plant extracts displayed a potential antibacterial activity of MIC < 100 μg/mL. Bishnu P. Marasini, Pankaj Baral, Pratibha Aryal, Kashi R. Ghimire, Sanjiv Neupane, Nabaraj Dahal, Anjana Singh, Laxman Ghimire, and Kanti Shrestha Copyright © 2015 Bishnu P. Marasini et al. All rights reserved. The Influence of Tea Tree Oil (Melaleuca alternifolia) on Fluconazole Activity against Fluconazole-Resistant Candida albicans Strains Wed, 04 Feb 2015 13:14:16 +0000 The aim of this study was to evaluate the activity of fluconazole against 32 clinical strains of fluconazole-resistant Candida albicans, and C. albicans ATCC 10231 reference strain, after their exposure to sublethal concentrations of tea tree oil (TTO) or its main bioactive component terpinen-4-ol. For all tested fluconazole-resistant C. albicans strains TTO and terpinen-4-ol minimal inhibitory concentrations (MICs) were low, ranging from 0.06% to 0.5%. The 24-hour exposure of fluconazole-resistant C. albicans strains to fluconazole with sublethal dose of TTO enhanced fluconazole activity against these strains. Overall, 62.5% of isolates were classified as susceptible, 25.0% exhibited intermediate susceptibility, and 12.5% were resistant. For all of the tested clinical strains the fluconazole MIC decreased from an average of 244.0 μg/mL to an average of 38.46 μg/mL, and the fluconazole minimal fungicidal concentrations (MFC) decreased from an average of 254.67 μg/mL to an average of 66.62 μg/mL. Terpinen-4-ol was found to be more active than TTO, and strongly enhanced fluconazole activity against fluconazole-resistant C. albicans strains. The results of this study demonstrate that combining natural substances such as TTO and conventional drug such as fluconazole, may help treat difficult yeast infections. Anna Mertas, Aleksandra Garbusińska, Ewelina Szliszka, Andrzej Jureczko, Magdalena Kowalska, and Wojciech Król Copyright © 2015 Anna Mertas et al. All rights reserved. Genetic Variability of Candida albicans Sap8 Propeptide in Isolates from Different Types of Infection Wed, 04 Feb 2015 06:33:53 +0000 The secreted aspartic proteases (Saps) are among the most studied virulence determinants in Candida albicans. These proteins are translated as pre-pro-enzymes consisting of a signal sequence followed by a propeptide and the mature enzyme. The propeptides of secreted proteinases are important for the correct processing, folding/secretion of the mature enzyme. In this study, the DNA sequences of C. albicans Saps were screened and a microsatellite was identified in SAP8 propeptide region. The genetic variability of the repetitive region of Sap8 propeptide was determined in 108 C. albicans independent strains isolated from different types of infection: oral infection (OI), oral commensal (OC), vulvovaginal candidiasis (VVC), and bloodstream infections (BSI). Nine different propeptides for Sap8 processing were identified whose frequencies varied with the type of infection. OC strains presented the highest gene diversity while OI isolated the lowest. The contribution of the Saps to mucosal and systemic infections has been demonstrated and recently Sap8 has been implicated in the cleavage of a signalling glycoprotein that leads to Cek1-MAPK pathway activation. This work is the first to identify a variable microsatellite in the propeptide of a secreted aspartic protease and brings new insights into the variability of Sap8. Joana Carvalho-Pereira, Catarina Vaz, Catarina Carneiro, Célia Pais, and Paula Sampaio Copyright © 2015 Joana Carvalho-Pereira et al. All rights reserved. Guidelines for Optimisation of a Multiplex Oligonucleotide Ligation-PCR for Characterisation of Microbial Pathogens in a Microsphere Suspension Array Tue, 03 Feb 2015 13:54:34 +0000 With multiplex oligonucleotide ligation-PCR (MOL-PCR) different molecular markers can be simultaneously analysed in a single assay and high levels of multiplexing can be achieved in high-throughput format. As such, MOL-PCR is a convenient solution for microbial detection and identification assays where many markers should be analysed, including for routine further characterisation of an identified microbial pathogenic isolate. For an assay aimed at routine use, optimisation in terms of differentiation between positive and negative results and of cost and effort is indispensable. As MOL-PCR includes a multiplex ligation step, followed by a singleplex PCR and analysis with microspheres on a Luminex device, several parameters are accessible for optimisation. Although MOL-PCR performance may be influenced by the markers used in the assay and the targeted bacterial species, evaluation of the method of DNA isolation, the probe concentration, the amount of microspheres, and the concentration of reporter dye is advisable in the development of any MOL-PCR assay. Therefore, we here describe our observations made during the optimisation of a 20-plex MOL-PCR assay for subtyping of Salmonella Typhimurium with the aim to provide a possible workflow as guidance for the development and optimisation of a MOL-PCR assay for the characterisation of other microbial pathogens. Véronique Wuyts, Nancy H. C. Roosens, Sophie Bertrand, Kathleen Marchal, and Sigrid C. J. De Keersmaecker Copyright © 2015 Véronique Wuyts et al. All rights reserved. Locus of Enterocyte Effacement: A Pathogenicity Island Involved in the Virulence of Enteropathogenic and Enterohemorragic Escherichia coli Subjected to a Complex Network of Gene Regulation Mon, 02 Feb 2015 07:15:57 +0000 The locus of enterocyte effacement (LEE) is a 35.6 kb pathogenicity island inserted in the genome of some bacteria such as enteropathogenic Escherichia coli, enterohemorrhagic E.coli, Citrobacter rodentium, and Escherichia albertii. LEE comprises the genes responsible for causing attaching and effacing lesions, a characteristic lesion that involves intimate adherence of bacteria to enterocytes, a signaling cascade leading to brush border and microvilli destruction, and loss of ions, causing severe diarrhea. It is composed of 41 open reading frames and five major operons encoding a type three system apparatus, secreted proteins, an adhesin, called intimin, and its receptor called translocated intimin receptor (Tir). LEE is subjected to various levels of regulation, including transcriptional and posttranscriptional regulators located both inside and outside of the pathogenicity island. Several molecules were described being related to feedback inhibition, transcriptional activation, and transcriptional repression. These molecules are involved in a complex network of regulation, including mechanisms such as quorum sensing and temporal control of LEE genes transcription and translation. In this mini review we have detailed the complex network that regulates transcription and expression of genes involved in this kind of lesion. Fernanda M. Franzin and Marcelo P. Sircili Copyright © 2015 Fernanda M. Franzin and Marcelo P. Sircili. All rights reserved. In Vitro Inhibitory and Cytotoxic Activity of MFM 501, a Novel Codonopsinine Derivative, against Methicillin-Resistant Staphylococcus aureus Clinical Isolates Sun, 01 Feb 2015 12:14:04 +0000 28 new pyrrolidine types of compounds as analogues for natural polyhydroxy alkaloids of codonopsinine were evaluated for their anti-MRSA activity using MIC and MBC value determination assay against a panel of S. aureus isolates. One pyrrolidine compound, MFM 501, exhibited good inhibitory activity with MIC value of 15.6 to 31.3 μg/mL against 55 S. aureus isolates (43 MRSA and 12 MSSA isolates). The active compound also displayed MBC values between 250 and 500 μg/mL against 58 S. aureus isolates (45 MRSA and 13 MSSA isolates) implying that MFM 501 has a bacteriostatic rather than bactericidal effect against both MRSA and MSSA isolates. In addition, MFM 501 showed no apparent cytotoxicity activity towards three normal cell lines (WRL-68, Vero, and 3T3) with IC50 values of >625 µg/mL. Selectivity index (SI) of MFM 501 gave a value of >10 suggesting that MFM 501 is significant and suitable for further in vivo investigations. These results suggested that synthetically derived intermediate compounds based on natural products may play an important role in the discovery of new anti-infective agents against MRSA. Saiful Azmi Johari, Mastura Mohtar, Sharifah Aminah Syed Mohammad, Rohana Sahdan, Zurina Shaameri, Ahmad Sazali Hamzah, and Mohd Fazli Mohammat Copyright © 2015 Saiful Azmi Johari et al. All rights reserved. Antibiotic Therapies in Maxillofacial Surgery in the Context of Prophylaxis Sun, 01 Feb 2015 11:54:04 +0000 Objectives. There is no single pattern for preventive action as to the duration and type of antibiotic therapy in maxillofacial surgery. In these circumstances, it appears reasonable to set relevant standards for prophylactic procedures after such surgeries. Methods. Retrospective analysis of bacteriological tests has been carried out as well as a susceptibility evaluation of cultured bacterial and fungal strains to antibiotics over a five-year period in subjects treated at the Cranio-Maxillo-Facial Clinic in Katowice. A total of 726 bacterial and fungal strains were cultured in 484 patients (200 women and 284 males). The age of the patients was 40.2 on average. Results. The most frequent bacteria isolated from the patients were Gram-positive 541 (74.5%). Gram-negative bacteria were present in 177 (24.4%) cases. Fungi of the Candida genus were isolated in eight cases (1.1%). Conclusions. The most often isolated bacteria were Streptococcus mitis and Streptococcus oralis, whose number has grown over the last two years. Empiric therapies should be based on ciprofloxacin and gentamicin. It has been observed that all the Gram-positive bacteria are becoming more resistant to all antibiotics. Ampicillin and imipenem were antibiotics with the steepest resistance reduction while vancomycin showed the lowest resistance drop. Bogusława Orzechowska-Wylęgała, Adam Wylęgała, Michał Buliński, and Iwona Niedzielska Copyright © 2015 Bogusława Orzechowska-Wylęgała et al. All rights reserved. Studies on Molecular Characterizations of the Outer Membrane Proteins, Lipids Profile, and Exopolysaccharides of Antibiotic Resistant Strain Pseudomonas aeruginosa Sun, 01 Feb 2015 11:24:38 +0000 Susceptibility of the tested Pseudomonas aeruginosa strain to two different antibiotics, tetracycline (TE) and ciprofloxacin (CIP), was carried out using liquid dilution method. Minimum inhibitory concentrations of TE and CIP were 9.0 and 6.0 mg/100 mL, respectively. Some metabolic changes due to both, the mode of action of TE and CIP on P. aeruginosa and its resistance to high concentrations of antibiotics (sub-MIC) were detected. The total cellular protein contents decreased after antibiotic treatment, while outer membrane protein (OMP) contents were approximately constant for both treated and untreated cells. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the OMPs for untreated and TE and CIP treated cells indicated that the molecular changes were achieved as; lost in, induction and stability of some protein bands as a result of antibiotics treatment. Five bands (with mol. wt. 71.75, 54.8, 31.72, 28.63, and 20.33 KDa) were stable in both treated and untreated tested strains, while two bands (with mol. wt. 194.8 and 118.3 KDa) were induced and the lost of only one band (with mol. wt. 142.5 KDa) after antibiotics treatment. On the other hand, total lipids and phospholipids increased in antibiotic treated cells, while neutral lipids decreased. Also, there was observable stability in the number of fatty acids in untreated and treated cells (11 fatty acids). The unsaturation index was decreased to 56% and 17.6% in both TE and CIP treatments, respectively. The produced amount of EPSs in untreated cultures of P. aeruginosa was relatively higher than in treated cultures with sub-MICs of TE and CIP antibiotics. It was also observed that the amounts of exopolysaccharides (EPSs) increased by increasing the incubation period up to five days of incubation in case of untreated and antibiotic treated cultures. Hany M. Yehia, Wesam A. Hassanein, and Shimaa M. Ibraheim Copyright © 2015 Hany M. Yehia et al. All rights reserved. Influence of Culture Media on Biofilm Formation by Candida Species and Response of Sessile Cells to Antifungals and Oxidative Stress Sun, 01 Feb 2015 10:03:38 +0000 The aims of the study were to evaluate the influence of culture media on biofilm formation by C. albicans, C. glabrata, C. krusei, and C. parapsilosis and to investigate the responses of sessile cells to antifungals and reactive oxygen species (ROS) as compared to planktonic cells. For biofilm formation, the Candida species were grown at different periods of time in YP or YNB media supplemented or not with 0.2 or 2% glucose. Sessile and planktonic cells were exposed to increasing concentrations of antifungals, H2O2, menadione or silver nanoparticles (AgNPs). Biofilms were observed by scanning electron microscopy (SEM) and quantified by the XTT assay. C. albicans formed biofilms preferentially in YPD containing 2% glucose (YPD/2%), C. glabrata in glucose-free YNB or supplemented with 0.2% glucose (YNB/0.2%), while C. krusei and C. parapsilosis preferred YP, YPD/0.2%, and YPD/2%. Interestingly, only C. albicans produced an exopolymeric matrix. This is the first report dealing with the in vitro effect of the culture medium and glucose on the formation of biofilms in four Candida species as well as the resistance of sessile cells to antifungals, AgNPs, and ROS. Our results suggest that candidiasis in vivo is a multifactorial and complex process where the nutritional conditions, the human immune system, and the adaptability of the pathogen should be considered altogether to provide an effective treatment of the patient. Isela Serrano-Fujarte, Everardo López-Romero, Georgina Elena Reyna-López, Ma. Alejandrina Martínez-Gámez, Arturo Vega-González, and Mayra Cuéllar-Cruz Copyright © 2015 Isela Serrano-Fujarte et al. All rights reserved. Biofilm-Forming Staphylococcus epidermidis Expressing Vancomycin Resistance Early after Adhesion to a Metal Surface Sat, 31 Jan 2015 07:10:11 +0000 We investigated biofilm formation and time of vancomycin (VCM) resistance expression after adhesion to a metal surface in Staphylococcus epidermidis. Biofilm-forming Staphylococcus epidermidis with a VCM MIC of 1 μg/mL was used. The bacteria were made to adhere to a stainless steel washer and treated with VCM at different times and concentrations. VCM was administered 0, 2, 4, and 8 hours after adhesion. The amount of biofilm formed was evaluated based on the biofilm coverage rates (BCRs) before and after VCM administration, bacterial viability in biofilm was visually observed using the fluorescence staining method, and the viable bacterial count in biofilm was measured. The VCM concentration required to decrease BCR significantly compared with that of VCM-untreated bacteria was 4 μg/mL, even in the 0 hr group. In the 4 and 8 hr groups, VCM could not inhibit biofilm growth even at 1,024 μg/mL. In the 8 hr group, viable bacteria remained in biofilm at a count of 104 CFU even at a high VCM concentration (1,024 μg/mL). It was suggested that biofilm-forming Staphylococcus epidermidis expresses resistance to VCM early after adhesion to a metal surface. Resistance increased over time after adhesion as the biofilm formed, and strong resistance was expressed 4–8 hours after adhesion. Toshiyuki Sakimura, Shiro Kajiyama, Shinji Adachi, Ko Chiba, Akihiko Yonekura, Masato Tomita, Hironobu Koseki, Takashi Miyamoto, Toshiyuki Tsurumoto, and Makoto Osaki Copyright © 2015 Toshiyuki Sakimura et al. All rights reserved. Health Safety of Soft Drinks: Contents, Containers, and Microorganisms Wed, 28 Jan 2015 13:51:37 +0000 Soft drinks consumption is still a controversial issue for public health and public policy. Over the years, numerous studies have been conducted into the possible links between soft drink intake and medical problems, the results of which, however, remain highly contested. Nevertheless, as a result, increasing emphasis is being placed on the health properties of soft drinks, by both the industry and the consumers, for example, in the expanding area of functional drinks. Extensive legislation has been put in place to ensure that soft drinks manufacturers conform to established national and international standards. Consumers trust that the soft drinks they buy are safe and their quality is guaranteed. They also expect to be provided with information that can help them to make informed decisions about the purchase of products and that the information on product labels is not false or misleading. This paper provides a broad overview of available scientific knowledge and cites numerous studies on various aspects of soft drinks and their implications for health safety. Particular attention is given to ingredients, including artificial flavorings, colorings, and preservatives and to the lesser known risks of microbiological and chemical contamination during processing and storage. Dorota Kregiel Copyright © 2015 Dorota Kregiel. All rights reserved. Small Molecule Inhibitor of Type Three Secretion Suppresses Acute and Chronic Chlamydia trachomatis Infection in a Novel Urogenital Chlamydia Model Wed, 28 Jan 2015 06:47:16 +0000 Previously, we reported that a compound from a group of thiohydrazides of oxamic acids, CL-55, possessed antichlamydial activity in vitro that was accompanied by a decreased translocation of the type three secretion effector, IncA, into the host cell. In this study, the antichlamydial activity of CL-55 was tested in vivo in DBA/2 mice infected with C. trachomatis serovar D. We found that intravaginal inoculation of DBA/2 mice with the clinically relevant strain, C. trachomatis serovar D, results in a course of infection and pathology similar to that observed in humans. The early stage of infection in this model was characterized by a shedding of Chlamydia in vaginal secretions followed by an ascending infection and inflammation in the upper genital tract. We found that CL-55 possessed antibacterial activity in vivo and was able to control C. trachomatis vaginal shedding, ascending infection, and inflammation in the upper genital organs in DBA/2 mice. Our data provide a proof of concept for the protective effect of the thiadiazinon, CL-55, against chlamydial infection in vivo and support the feasibility of further studies of its potential therapeutic applications. Ekaterina A. Koroleva, Natalia V. Kobets, Egor S. Zayakin, Sergei I. Luyksaar, Ludmila A. Shabalina, and Naylia A. Zigangirova Copyright © 2015 Ekaterina A. Koroleva et al. All rights reserved. Phylogeny and Taxonomical Investigation of Trichoderma spp. from Indian Region of Indo-Burma Biodiversity Hot Spot Region with Special Reference to Manipur Wed, 28 Jan 2015 06:43:44 +0000 Towards assessing the genetic diversity and occurrence of Trichoderma species from the Indian region of Indo-Burma Biodiversity hotspot, a total of 193 Trichoderma strains were isolated from cultivated soils of nine different districts of Manipur comprising 4 different agroclimatic zones. The isolates were grouped based on the morphological characteristics. ITS-RFLP of the rDNA region using three restriction digestion enzymes: Mob1, Taq1, and Hinf1, showed interspecific variations among 65 isolates of Trichoderma. Based on ITS sequence data, a total of 22 different types of representative Trichoderma species were reported and phylogenetic analysis showed 4 well-separated main clades in which T. harzianum was found to be the most prevalent spp. among all the Trichoderma spp. Combined molecular and phenotypic data leads to the development of a taxonomy of all the 22 different Trichoderma spp., which was reported for the first time from this unique region. All these species were found to produce different extrolites and enzymes responsible for the biocontrol activities against the harmful fungal phytopathogens that hamper in food production. This potential indigenous Trichoderma spp. can be targeted for the development of suitable bioformulation against soil and seedborne pathogens in sustainable agricultural practice. Th. Kamala, S. Indira Devi, K. Chandradev Sharma, and K. Kennedy Copyright © 2015 Th. Kamala et al. All rights reserved. The Type III Secretion System (T3SS) of Chlamydophila psittaci Is Involved in the Host Inflammatory Response by Activating the JNK/ERK Signaling Pathway Thu, 22 Jan 2015 12:53:15 +0000 Chlamydophila psittaci (C. psittaci) is a human zoonotic pathogen, which could result in severe respiratory disease. In the present study, we investigated the role and mechanism of the type III secretion system (T3SS) of C. psittaci in regulating the inflammatory response in host cells. C. psittaci-infected THP-1 cells were incubated with the specific T3SS inhibitor INP0007, inhibitors of ERK, p38, or JNK, and the levels of inflammatory cytokines were analyzed using Q-PCR and ELISA. The levels of ERK, p38, and JNK phosphorylation were analyzed by Western blot. Our results verified that INP0007 inhibited chlamydial growth in vitro, but the coaddition of exogenous iron completely reversed the growth deficit. INP0007 inhibited the growth of C. psittaci and decreased the levels of IL-8, IL-6, TNF-α, and IL-1β. Exogenous iron restored the chlamydial growth but not the production of inflammatory cytokines. These results demonstrated that the expression of inflammatory cytokines during infection was associated with the T3SS which reduced by incubation with ERK and JNK inhibitors, but not with p38 inhibitor. We concluded that the T3SS elicited inflammatory responses by activating the JNK or ERK signaling pathways in the infection of C. psittaci. Qing-zhi He, Huai-cai Zeng, Yan Huang, Yan-qun Hu, and Yi-mou Wu Copyright © 2015 Qing-zhi He et al. All rights reserved. Effects of Mentha suaveolens Essential Oil on Chlamydia trachomatis Thu, 22 Jan 2015 12:10:51 +0000 Chlamydia trachomatis, the most common cause of sexually transmitted bacterial infection worldwide, has a unique biphasic developmental cycle alternating between the infectious elementary body and the replicative reticulate body. C. trachomatis is responsible for severe reproductive complications including pelvic inflammatory disease, ectopic pregnancy, and obstructive infertility. The aim of our study was to evaluate whether Mentha suaveolens essential oil (EOMS) can be considered as a promising candidate for preventing C. trachomatis infection. Specifically, we investigated the in vitro effects of EOMS towards C. trachomatis analysing the different phases of chlamydial developmental cycle. Our results demonstrated that EOMS was effective towards C. trachomatis, whereby it not only inactivated infectious elementary bodies but also inhibited chlamydial replication. Our study also revealed the effectiveness of EOMS, in combination with erythromycin, towards C. trachomatis with a substantial reduction in the minimum effect dose of antibiotic. In conclusion, EOMS treatment may represent a preventative strategy since it may reduce C. trachomatis transmission in the population and, thereby, reduce the number of new chlamydial infections and risk of developing of severe sequelae. Rosa Sessa, Marisa Di Pietro, Fiorenzo De Santis, Simone Filardo, Rino Ragno, and Letizia Angiolella Copyright © 2015 Rosa Sessa et al. All rights reserved. Identification of a New Alcaligenes faecalis Strain MOR02 and Assessment of Its Toxicity and Pathogenicity to Insects Sun, 18 Jan 2015 11:51:46 +0000 We report the isolation of a bacterium from Galleria mellonella larva and its identification using genome sequencing and phylogenomic analysis. This bacterium was named Alcaligenes faecalis strain MOR02. Microscopic analyses revealed that the bacteria are located in the esophagus and intestine of the nematodes Steinernema feltiae, S. carpocapsae, and H. bacteriophora. Using G. mellonella larvae as a model, when the larvae were injected with 24,000 CFU in their hemocoel, more than 96% mortality was achieved after 24 h. Additionally, toxicity assays determined that 1 μg of supernatant extract from A. faecalis MOR02 killed more than 70% G. mellonella larvae 96 h after injection. A correlation of experimental data with sequence genome analyses was also performed. We discovered genes that encode proteins and enzymes that are related to pathogenicity, toxicity, and host/environment interactions that may be responsible for the observed phenotypic characteristics. Our data demonstrates that the bacteria are able to use different strategies to colonize nematodes and kill insects to their own benefit. However, there remains an extensive group of unidentified microorganisms that could be participating in the infection process. Additionally, a nematode-bacterium association could be established probably as a strategy of dispersion and colonization. Rosa Estela Quiroz-Castañeda, Ared Mendoza-Mejía, Verónica Obregón-Barboza, Fernando Martínez-Ocampo, Armando Hernández-Mendoza, Felipe Martínez-Garduño, Gabriel Guillén-Solís, Federico Sánchez-Rodríguez, Guadalupe Peña-Chora, Laura Ortíz-Hernández, Paul Gaytán-Colín, and Edgar Dantán-González Copyright © 2015 Rosa Estela Quiroz-Castañeda et al. All rights reserved. Antibacterial Mechanisms of Polymyxin and Bacterial Resistance Thu, 15 Jan 2015 12:10:09 +0000 Multidrug resistance in pathogens is an increasingly significant threat for human health. Indeed, some strains are resistant to almost all currently available antibiotics, leaving very limited choices for antimicrobial clinical therapy. In many such cases, polymyxins are the last option available, although their use increases the risk of developing resistant strains. This review mainly aims to discuss advances in unraveling the mechanisms of antibacterial activity of polymyxins and bacterial tolerance together with the description of polymyxin structure, synthesis, and structural modification. These are expected to help researchers not only develop a series of new polymyxin derivatives necessary for future medical care, but also optimize the clinical use of polymyxins with minimal resistance development. Zhiliang Yu, Wangrong Qin, Jianxun Lin, Shisong Fang, and Juanping Qiu Copyright © 2015 Zhiliang Yu et al. All rights reserved. Phylogenetic Groups of Escherichia coli Strains from Patients with Urinary Tract Infection in Iran Based on the New Clermont Phylotyping Method Thu, 08 Jan 2015 08:19:42 +0000 Objectives. In 2013, Clermont classified E. coli strains into eight phylogenetic groups using a new quadruplex PCR method. The aims of this study were to identify the phylogenetic groups of E. coli based on this method and to assess their antibiotic resistance patterns in Bushehr, Iran. Methods. In this cross-sectional study, 140 E. coli isolates were subjected to phylogenetic typing by a quadruplex PCR method. Antimicrobial susceptibility testing was performed by disk diffusion method. Results. Phylogenetic group B2 was most predominant (39.3%), followed by unknown (27.1%), E (9.3%), C and clade I (each 6.4%), B1 (5%), F and D (each 2.9%), and A (0.7%). The most common antibiotic resistance was related to amoxicillin (82.1%) and the least to meropenem (0.7%). 82.14% of isolates were multiple drug resistant (MDR). Antibiotic resistance was mainly detected in group B2 (50%). Conclusions. Our findings showed the high prevalence of MDR E. coli isolates with dominance of group B2. About 25% of E. coli isolates belong to the newly described phylogroups C, E, F, and clade I. Such studies need to be done also in other regions to provide greater understanding of the antibiotic resistance pattern and the prevalences of different phylogenetic groups. Darioush Iranpour, Mojtaba Hassanpour, Hossein Ansari, Saeed Tajbakhsh, Gholamreza Khamisipour, and Akram Najafi Copyright © 2015 Darioush Iranpour et al. All rights reserved. Antimicrobial, Antioxidant, and Cytotoxic Properties of Vasicine Acetate Synthesized from Vasicine Isolated from Adhatoda vasica L. Tue, 06 Jan 2015 09:03:54 +0000 Adhatoda vasica (L.) (Acanthaceae) is used in the indigenous system of medicine in India. The alkaloid Vasicine was isolated from ethanolic extract of the leaves of A. vasica using column chromatography. Vasicine acetate was obtained by acetylation of Vasicine. Vasicine acetate exhibited good zone of inhibition against bacteria: 10 mm against E. aerogenes, 10 mm against S. epidermidis, and 10 mm against P. aeruginosa. Vasicine acetate showed minimum inhibitory concentration values against bacteria: M. luteus (125 μg/mL), E. aerogenes (125 μg/mL), S. epidermidis (125 μg/mL), and P. aeruginosa (125 μg/mL). The radical scavenging activity of Vasicine acetate was the maximum at 1000 μg/mL (66.15%). The compound showed prominent cytotoxic activity in vitro against A549 lung adenocarcinoma cancer cell line. Quantification of Vasicine and Vasicine acetate by HPLC-DAD analysis showed their contents to be 0.2293% and 0.0156%, respectively, on dry weight basis of the leaves. Vasicine acetate could be probed further in drug discovery programme. V. Duraipandiyan, N. A. Al-Dhabi, C. Balachandran, S. Ignacimuthu, C. Sankar, and K. Balakrishna Copyright © 2015 V. Duraipandiyan et al. All rights reserved. Molecular Characterization of a New Alkaline-Tolerant Xylanase from Humicola insolens Y1 Mon, 05 Jan 2015 14:06:33 +0000 An endo-1,4-β-xylanase-encoding gene, xyn11B, was cloned from the thermophilic fungus Humicola insolens Y1. The gene encodes a multimodular xylanase that consists of a typical hydrophobic signal sequence, a catalytic domain of glycoside hydrolase (GH) family 11, a glycine-rich linker, and a family 1 carbohydrate binding module (CBM1). Deduced Xyn11B shares the highest identity of 74% with a putative xylanase from Podospora anserina S mat+. Recombinant Xyn11B was successfully expressed in Pichia pastoris and purified to electrophoretic homogeneity. Xyn11B had a high specific activity of 382.0 U mg−1 towards beechwood xylan and showed optimal activity at pH 6.0 and 50°C. Distinct from most reported acidic fungal xylanases, Xyn11B was alkaline-tolerant, retaining 30.7% of the maximal activity at pH 9.0. The and values for beechwood xylan were 2.2 mg mL−1 and 462.8 μmol min−1 mg−1, respectively. The enzyme exhibited a wider substrate specificity and produced a mixture of xylooligosaccharides. All these favorable enzymatic properties make Xyn11B attractive for potential applications in various industries. Pengjun Shi, Yanlong Du, Hong Yang, Huoqing Huang, Xiu Zhang, Yaru Wang, and Bin Yao Copyright © 2015 Pengjun Shi et al. All rights reserved. Pyrosequencing Reveals Fungal Communities in the Rhizosphere of Xinjiang Jujube Mon, 05 Jan 2015 07:14:25 +0000 Fungi are important soil components as both decomposers and plant symbionts and play a major role in ecological and biogeochemical processes. However, little is known about the richness and structure of fungal communities. DNA sequencing technologies allow for the direct estimation of microbial community diversity, avoiding culture-based biases. We therefore used 454 pyrosequencing to investigate the fungal communities in the rhizosphere of Xinjiang jujube. We obtained no less than 40,488 internal transcribed spacer (ITS) rDNA reads, the number of each sample was 6943, 6647, 6584, 6550, 6860, and 6904, and we used bioinformatics and multivariate statistics to analyze the results. The index of diversity showed greater richness in the rhizosphere fungal community of a 3-year-old jujube than in that of an 8-year-old jujube. Most operational taxonomic units belonged to Ascomycota, and taxonomic analyses identified Hypocreales as the dominant fungal order. Our results demonstrated that the fungal orders are present in different proportions in different sampling areas. Redundancy analysis (RDA) revealed a significant correlation between soil properties and the abundance of fungal phyla. Our results indicated lower fungal diversity in the rhizosphere of Xinjiang jujube than that reported in other studies, and we hope our findings provide a reference for future research. Peng Liu, Xiao-Hui Wang, Jian-Gui Li, Wei Qin, Cheng-Ze Xiao, Xu Zhao, Hong-Xia Jiang, Jun-Kang Sui, Rong-Bo Sa, Wei-Yan Wang, and Xun-Li Liu Copyright © 2015 Peng Liu et al. All rights reserved. Understanding Host-Adherent-Invasive Escherichia coli Interaction in Crohn’s Disease: Opening Up New Therapeutic Strategies Mon, 15 Dec 2014 14:43:49 +0000 A trillion of microorganisms colonize the mammalian intestine. Most of them have coevolved with the host in a symbiotic relationship and some of them have developed strategies to promote their replication in the presence of competing microbiota. Recent evidence suggests that perturbation of the microbial community favors the emergence of opportunistic pathogens, in particular adherent-invasive Escherichia coli (AIEC) that can increase incidence and severity of gut inflammation in the context of Crohn’s disease (CD). This review will report the importance of AIEC as triggers of intestinal inflammation, focusing on their impact on epithelial barrier function and stimulation of mucosal inflammation. Beyond manipulation of immune response, restoration of gut microbiota as a new treatment option for CD patients will be discussed. Allison Agus, Sébastien Massier, Arlette Darfeuille-Michaud, Elisabeth Billard, and Nicolas Barnich Copyright © 2014 Allison Agus et al. All rights reserved. Contribution of Crk Adaptor Proteins to Host Cell and Bacteria Interactions Tue, 25 Nov 2014 00:00:00 +0000 The Crk adaptor family of proteins comprises the alternatively spliced CrkI and CrkII isoforms, as well as the paralog Crk-like (CrkL) protein, which is encoded by a different gene. Initially thought to be involved in signaling during apoptosis and cell adhesion, this ubiquitously expressed family of proteins is now known to play essential roles in integrating signals from a wide range of stimuli. In this review, we describe the structure and function of the different Crk proteins. We then focus on the emerging roles of Crk adaptors during Enterobacteriaceae pathogenesis, with special emphasis on the important human pathogens Salmonella, Shigella, Yersinia, and enteropathogenic Escherichia coli. Throughout, we remark on opportunities for future research into this intriguing family of proteins. Narcisa Martinez-Quiles, Leigh Ann Feuerbacher, María Benito-León, and Philip R. Hardwidge Copyright © 2014 Narcisa Martinez-Quiles et al. All rights reserved. Rapid and Sensitive PCR-Dipstick DNA Chromatography for Multiplex Analysis of the Oral Microbiota Mon, 17 Nov 2014 07:11:31 +0000 A complex of species has been associated with dental caries under the ecological hypothesis. This study aimed to develop a rapid, sensitive PCR-dipstick DNA chromatography assay that could be read by eye for multiplex and semiquantitative analysis of plaque bacteria. Parallel oligonucleotides were immobilized on a dipstick strip for multiplex analysis of target DNA sequences of the caries-associated bacteria, Streptococcus mutans, Streptococcus sobrinus, Scardovia wiggsiae, Actinomyces species, and Veillonella parvula. Streptavidin-coated blue-colored latex microspheres were to generate signal. Target DNA amplicons with an oligonucleotide-tagged terminus and a biotinylated terminus were coupled with latex beads through a streptavidin-biotin interaction and then hybridized with complementary oligonucleotides on the strip. The accumulation of captured latex beads on the test and control lines produced blue bands, enabling visual detection with the naked eye. The PCR-dipstick DNA chromatography detected quantities as low as 100 pg of DNA amplicons and demonstrated 10- to 1000-fold higher sensitivity than PCR-agarose gel electrophoresis, depending on the target bacterial species. Semiquantification of bacteria was performed by obtaining a series of chromatograms using serial 10-fold dilution of PCR-amplified DNA extracted from dental plaque samples. The assay time was less than 3 h. The semiquantification procedure revealed the relative amounts of each test species in dental plaque samples, indicating that this disposable device has great potential in analysis of microbial composition in the oral cavity and intestinal tract, as well as in point-of-care diagnosis of microbiota-associated diseases. Lingyang Tian, Takuichi Sato, Kousuke Niwa, Mitsuo Kawase, Anne C. R. Tanner, and Nobuhiro Takahashi Copyright © 2014 Lingyang Tian et al. All rights reserved. Detection of Free-Living Amoebae Using Amoebal Enrichment in a Wastewater Treatment Plant of Gauteng Province, South Africa Tue, 04 Nov 2014 07:29:31 +0000 Free-living amoebae pose a potential health risk in water systems as they may be pathogenic and harbor potential pathogenic bacteria known as amoebae resistant bacteria. Free-living amoebae were observed in 150 (87.2%) of the environmental water samples. In particular, Acanthamoeba sp. was identified in 22 (12.8%) using amoebal enrichment and confirmed by molecular analysis. FLA were isolated in all 8 stages of the wastewater treatment plant using the amoebal enrichment technique. A total of 16 (9.3%) samples were positive for FLA from influent, 20 (11.6%) from bioreactor feed, 16 (9.3%) from anaerobic zone, 16 (9.3%) from anoxic zone, 32 (18.6%) from aerators, 16 (9.3%) from bioreactor effluent, 11 (6.4%) from bioreactor final effluent, and 45 (26.2%) from maturation pond. This study provides baseline information on the occurrence of amoebae in wastewater treatment plant. This has health implications on receiving water bodies as some FLA are pathogenic and are also involved in the transmission and dissemination of pathogenic bacteria. P. Muchesa, O. Mwamba, T. G. Barnard, and C. Bartie Copyright © 2014 P. Muchesa et al. All rights reserved. Effects of Various Antibiotics Alone or in Combination with Doripenem against Klebsiella pneumoniae Strains Isolated in an Intensive Care Unit Tue, 28 Oct 2014 07:54:59 +0000 Colistin, tigecycline, levofloxacin, tobramycin, and rifampin alone and in combination with doripenem were investigated for their in vitro activities and postantibiotic effects (PAEs) on Klebsiella pneumoniae. The in vitro activities of tested antibiotics in combination with doripenem were determined using a microbroth checkerboard technique. To determine the PAEs, K. pneumoniae strains in the logarithmic phase of growth were exposed for 1 h to antibiotics, alone and in combination. Recovery periods of test cultures were evaluated using viable counting after centrifugation. Colistin, tobramycin, and levofloxacin produced strong PAEs ranging from 2.71 to 4.23 h, from 1.31 to 3.82 h, and from 1.35 to 4.72, respectively, in a concentration-dependent manner. Tigecycline and rifampin displayed modest PAEs ranging from 1.18 h to 1.55 h and 0.92 to 1.19, respectively. Because it is a beta-lactam, PAEs were not exactly induced by doripenem (ranging from 0.10 to 0.18 h). In combination, doripenem scarcely changed the duration of PAE of each tested antibiotic alone. The findings of this study may have important implications for the timing of doses during K. pneumoniae therapy with tested antibiotics. Berna Ozbek Celik, Emel Mataraci-Kara, and Mesut Yilmaz Copyright © 2014 Berna Ozbek Celik et al. All rights reserved. High Resolution Melting Analysis for Rapid Mutation Screening in Gyrase and Topoisomerase IV Genes in Quinolone-Resistant Salmonella enterica Sun, 12 Oct 2014 00:00:00 +0000 The increased Salmonella resistance to quinolones and fluoroquinolones is a public health concern in the Southeast Asian region. The objective of this study is to develop a high resolution melt curve (HRM) assay to rapidly screen for mutations in quinolone-resistant determining region (QRDR) of gyrase and topoisomerase IV genes. DNA sequencing was performed on 62 Salmonella strains to identify mutations in the QRDR of gyrA, gyrB, parC, and parE genes. Mutations were detected in QRDR of gyrA (; S83F, S83Y, S83I, D87G, D87Y, and D87N) and parE (; M438I). Salmonella strains with mutations within QRDR of gyrA are generally more resistant to nalidixic acid (MIC  μg/mL). Mutations were uncommon within the QRDR of gyrB, parC, and parE genes. In the HRM assay, mutants can be distinguished from the wild-type strains based on the transition of melt curves, which is more prominent when the profiles are displayed in difference plot. In conclusion, HRM analysis allows for rapid screening for mutations at the QRDRs of gyrase and topoisomerase IV genes in Salmonella. This assay markedly reduced the sequencing effort involved in mutational studies of quinolone-resistance genes. Soo Tein Ngoi and Kwai Lin Thong Copyright © 2014 Soo Tein Ngoi and Kwai Lin Thong. All rights reserved. Pentachlorophenol Degradation by Janibacter sp., a New Actinobacterium Isolated from Saline Sediment of Arid Land Wed, 17 Sep 2014 11:48:30 +0000 Many pentachlorophenol- (PCP-) contaminated environments are characterized by low or elevated temperatures, acidic or alkaline pH, and high salt concentrations. PCP-degrading microorganisms, adapted to grow and prosper in these environments, play an important role in the biological treatment of polluted extreme habitats. A PCP-degrading bacterium was isolated and characterized from arid and saline soil in southern Tunisia and was enriched in mineral salts medium supplemented with PCP as source of carbon and energy. Based on 16S rRNA coding gene sequence analysis, the strain FAS23 was identified as Janibacter sp. As revealed by high performance liquid chromatography (HPLC) analysis, FAS23 strain was found to be efficient for PCP removal in the presence of 1% of glucose. The conditions of growth and PCP removal by FAS23 strain were found to be optimal in neutral pH and at a temperature of 30°C. Moreover, this strain was found to be halotolerant at a range of 1–10% of NaCl and able to degrade PCP at a concentration up to 300 mg/L, while the addition of nonionic surfactant (Tween 80) enhanced the PCP removal capacity. Amel Khessairi, Imene Fhoula, Atef Jaouani, Yousra Turki, Ameur Cherif, Abdellatif Boudabous, Abdennaceur Hassen, and Hadda Ouzari Copyright © 2014 Amel Khessairi et al. All rights reserved. Emerging Rapid Resistance Testing Methods for Clinical Microbiology Laboratories and Their Potential Impact on Patient Management Wed, 17 Sep 2014 08:57:44 +0000 Atypical and multidrug resistance, especially ESBL and carbapenemase expressing Enterobacteriaceae, is globally spreading. Therefore, it becomes increasingly difficult to achieve therapeutic success by calculated antibiotic therapy. Consequently, rapid antibiotic resistance testing is essential. Various molecular and mass spectrometry-based approaches have been introduced in diagnostic microbiology to speed up the providing of reliable resistance data. PCR- and sequencing-based approaches are the most expensive but the most frequently applied modes of testing, suitable for the detection of resistance genes even from primary material. Next generation sequencing, based either on assessment of allelic single nucleotide polymorphisms or on the detection of nonubiquitous resistance mechanisms might allow for sequence-based bacterial resistance testing comparable to viral resistance testing on the long term. Fluorescence in situ hybridization (FISH), based on specific binding of fluorescence-labeled oligonucleotide probes, provides a less expensive molecular bridging technique. It is particularly useful for detection of resistance mechanisms based on mutations in ribosomal RNA. Approaches based on MALDI-TOF-MS, alone or in combination with molecular techniques, like PCR/electrospray ionization MS or minisequencing provide the fastest resistance results from pure colonies or even primary samples with a growing number of protocols. This review details the various approaches of rapid resistance testing, their pros and cons, and their potential use for the diagnostic laboratory. Hagen Frickmann, Wycliffe Omurwa Masanta, and Andreas E. Zautner Copyright © 2014 Hagen Frickmann et al. All rights reserved. Combating Pathogenic Microorganisms Using Plant-Derived Antimicrobials: A Minireview of the Mechanistic Basis Sun, 14 Sep 2014 12:15:47 +0000 The emergence of antibiotic resistance in pathogenic bacteria has led to renewed interest in exploring the potential of plant-derived antimicrobials (PDAs) as an alternative therapeutic strategy to combat microbial infections. Historically, plant extracts have been used as a safe, effective, and natural remedy for ailments and diseases in traditional medicine. Extensive research in the last two decades has identified a plethora of PDAs with a wide spectrum of activity against a variety of fungal and bacterial pathogens causing infections in humans and animals. Active components of many plant extracts have been characterized and are commercially available; however, research delineating the mechanistic basis of their antimicrobial action is scanty. This review highlights the potential of various plant-derived compounds to control pathogenic bacteria, especially the diverse effects exerted by plant compounds on various virulence factors that are critical for pathogenicity inside the host. In addition, the potential effect of PDAs on gut microbiota is discussed. Abhinav Upadhyay, Indu Upadhyaya, Anup Kollanoor-Johny, and Kumar Venkitanarayanan Copyright © 2014 Abhinav Upadhyay et al. All rights reserved. Molecular Identification and Quantification of Tetracycline and Erythromycin Resistance Genes in Spanish and Italian Retail Cheeses Thu, 11 Sep 2014 09:17:27 +0000 Large antibiotic resistance gene pools in the microbiota of foods may ultimately pose a risk for human health. This study reports the identification and quantification of tetracycline- and erythromycin-resistant populations, resistance genes, and gene diversity in traditional Spanish and Italian cheeses, via culturing, conventional PCR, real-time quantitative PCR (qPCR), and denaturing gradient gel electrophoresis (DGGE). The numbers of resistant bacteria varied widely among the antibiotics and the different cheese varieties; in some cheeses, all the bacterial populations seemed to be resistant. Up to eight antibiotic resistance genes were sought by gene-specific PCR, six with respect to tetracycline, that is, tet(K), tet(L), tet(M), tet(O), tet(S), and tet(W), and two with respect to erythromycin, that is, erm(B) and erm(F). The most common resistance genes in the analysed cheeses were tet(S), tet(W), tet(M), and erm(B). The copy numbers of these genes, as quantified by qPCR, ranged widely between cheeses (from 4.94 to ). DGGE analysis revealed distinct banding profiles and two polymorphic nucleotide positions for tet(W)-carrying cheeses, though the similarity of the sequences suggests this tet(W) to have a monophyletic origin. Traditional cheeses would therefore appear to act as reservoirs for large numbers of many types of antibiotic resistance determinants. Ana Belén Flórez, Ángel Alegría, Franca Rossi, Susana Delgado, Giovanna E. Felis, Sandra Torriani, and Baltasar Mayo Copyright © 2014 Ana Belén Flórez et al. All rights reserved. Bifidobacteria-Host Interactions—An Update on Colonisation Factors Thu, 11 Sep 2014 06:39:36 +0000 Bifidobacteria are one of the predominant bacterial groups of the human intestinal microbiota and have important functional properties making them interesting for the food and dairy industries. Numerous in vitro and preclinical studies have shown beneficial effects of particular bifidobacterial strains or strain combinations on various health parameters of their hosts. This indicates the potential of bifidobacteria in alternative or supplementary therapeutic approaches in a number of diseased states. Based on these observations, bifidobacteria have attracted considerable interest by the food, dairy, and pharmaceutical industries and they are widely used as so-called probiotics. As a consequence of the rapidly increasing number of available bifidobacterial genome sequences and their analysis, there has been substantial progress in the identification of bifidobacterial structures involved in colonisation of and interaction with the host. With the present review, we aim to provide an update on the current knowledge on the mechanisms by which bifidobacteria colonise their hosts and exert health promoting effects. Verena Grimm, Christina Westermann, and Christian U. Riedel Copyright © 2014 Verena Grimm et al. All rights reserved. Effect of High Hydrostatic Pressure Processing on Microbiological Shelf-Life and Quality of Fruits Pretreated with Ascorbic Acid or SnCl2 Thu, 11 Sep 2014 06:30:38 +0000 In the current study, the processing conditions required for the inactivation of Paenibacillus polymyxa and relevant spoilage microorganisms by high hydrostatic pressure (HHP) treatment on apricot, peach, and pear pieces in sucrose (22°Brix) solution were assessed. Accordingly, the shelf-life was determined by evaluating both the microbiological quality and the sensory characteristics (taste, odor, color, and texture) during refrigerated storage after HHP treatment. The microbiological shelf-life of apricots, peaches, and pears was prolonged in the HHP-treated products in comparison with the untreated ones. In all HHP-treated packages for apricots, peaches, and pears, all populations were below the detection limit of the method (1 log CFU/g) and no growth of microorganisms was observed until the end of storage. Overall, no differences of the , , or value among the untreated and the HHP-treated fruit products were observed up to the time at which the unpressurized product was characterized as spoiled. HHP treatment had no remarkable effect on the firmness of the apricots, peaches, and pears. With regard to the sensory assessment, the panelists marked better scores to HHP-treated products compared to their respective controls, according to taste and total evaluation during storage of fruit products. Anthoula A. Argyri, Chrysoula C. Tassou, Fotios Samaras, Constantinos Mallidis, and Nikos Chorianopoulos Copyright © 2014 Anthoula A. Argyri et al. All rights reserved. Epidemiology of West Nile Disease in Europe and in the Mediterranean Basin from 2009 to 2013 Thu, 11 Sep 2014 06:30:22 +0000 West Nile virus (WNV) transmission has been confirmed in the last four years in Europe and in the Mediterranean Basin. An increasing concern towards West Nile disease (WND) has been observed due to the high number of human and animal cases reported in these areas confirming the importance of this zoonosis. A new epidemiological scenario is currently emerging: although new introductions of the virus from abroad are always possible, confirming the epidemiological role played by migratory birds, the infection endemisation in some European territories today is a reality supported by the constant reoccurrence of the same strains across years in the same geographical areas. Despite the WND reoccurrence in the Old World, the overwintering mechanisms are not well known, and the role of local resident birds or mosquitoes in this context is poorly understood. A recent new epidemiological scenario is the spread of lineage 2 strain across European and Mediterranean countries in regions where lineage 1 strain is still circulating creating favourable conditions for genetic reassortments and emergence of new strains. This paper summarizes the main epidemiological findings on WNV occurrence in Europe and in the Mediterranean Basin from 2009 to 2013, considering potential future spread patterns. Daria Di Sabatino, Rossana Bruno, Francesca Sauro, Maria Luisa Danzetta, Francesca Cito, Simona Iannetti, Valeria Narcisi, Fabrizio De Massis, and Paolo Calistri Copyright © 2014 Daria Di Sabatino et al. All rights reserved. Attachment of Asaia bogorensis Originating in Fruit-Flavored Water to Packaging Materials Thu, 11 Sep 2014 00:00:00 +0000 The objective of this study was to investigate the adhesion of isolated spoilage bacteria to packaging materials used in the food industry. Microorganisms were isolated from commercial fruit-flavored mineral water in plastic bottles with flocks as a visual defect. The Gram-negative rods were identified using the molecular method through the amplification of a partial region of the 16S rRNA gene. Based on the sequence identity (99.6%) between the spoilage organism and a reference strain deposited in GenBank, the spoilage isolate was identified as Asaia bgorensis. Experiments on bacterial adhesion were conducted using plates made of glass and polystyrene (packaging materials commonly used in the beverage industry). Cell adhesion ability was determined using luminometry, plate count, and the microscopic method. The strain of A. bogorensis was characterized by strong adhesion properties which were dependent on the surface type, with the highest cell adhesion detected on polystyrene. Dorota Kregiel, Anna Otlewska, and Hubert Antolak Copyright © 2014 Dorota Kregiel et al. All rights reserved. Cytotoxicity of Probiotics from Philippine Commercial Dairy Products on Cancer Cells and the Effect on Expression of cfos and cjun Early Apoptotic-Promoting Genes and Interleukin-1β and Tumor Necrosis Factor-α Proinflammatory Cytokine Genes Sun, 07 Sep 2014 06:19:56 +0000 This study determined cytotoxicity of probiotic Lactobacillus spp. from Philippine dairy products on cancer cells and normal fibroblasts and their effects on expression of early apoptotic-promoting cfos, cjun and proinflammatory cytokine IL-1β, TNF-α genes. Cultures were from Yakult, Bear Brand Probiotic Drink, Nido3+ Powdered Milk. Filter-sterilized supernatants from cultures of Lactobacillus spp. were evaluated for cytotoxicity to colon cancer cells (HT-29 and HCT116), leukemia cells (THP-1), and normal human dermal fibroblasts (HDFn) using PrestoBlue. Bleomycin was the positive control. Absolute quantification of transcript levels was conducted using qRT-PCR. Cytotoxicity index profiles on HDFn, THP-1 of all probiotic supernatants and negative controls suggest nontoxicity to the cells when compared to bleomycin, whereas all probiotic supernatants were found to be cytotoxic to HT-29 and HCT-116 colon cancer cell lines. Expression of cfos, cjun transcripts was significantly upregulated in HT-29 and HCT116 cells treated with probiotic supernatants compared to untreated baseline levels (). Expression of IL-1β and TNF-α by lipopolysaccharide-treated macrophages was significantly downregulated in cells with probiotic supernatants compared to those exposed to MRS medium (). Results provide strong support for the role of Lactobacillus spp. studied in anticancer therapy and in prevention of inflammation that may act as precursor to carcinogenesis. Peter T. Shyu, Glenn G. Oyong, and Esperanza C. Cabrera Copyright © 2014 Peter T. Shyu et al. All rights reserved. Successive Nonstatistical and Statistical Approaches for the Improved Antibiotic Activity of Rare Actinomycete Nonomuraea sp. JAJ18 Wed, 03 Sep 2014 12:59:02 +0000 The selection and optimization of nutritional constituents as well as their levels for the improved production of antibiotic by Nonomuraea sp. JAJ18 were carried out using combination of both nonstatistical one-factor-at-a-time (OFAT) method and statistical response surface methodology (RSM). Using OFAT method, starch and (NH4)2SO4 were identified as suitable carbon and nitrogen sources, respectively. Subsequently, starch, NaCl, and MgSO47H2O were recognized as the most significant media components with confidence level of above 95% using the Plackett-Burman design. The levels of the three media components were further optimized using RSM employed with Box-Behnken design. Accordingly, a second-order polynomial regression model was fitted into the experimental data. By analyzing the response surface plots as well as using numerical optimization method, the optimal levels for starch, NaCl, and MgSO47H2O were determined as 15.6 g/L, 0.8 g/L, and 1.98 g/L, respectively. With the optimized medium, 15.5% increase was observed in antibiotic activity of JAJ18. Results further support the use of RSM for media optimization. To the best of our knowledge, this is the first report of statistical media optimization for antibiotic production in rare actinomycete Nonomuraea species, which will be useful for the development of Nonomuraea cultivation process for efficient antibiotic production on a large scale. Polpass Arul Jose and Solomon Robinson David Jebakumar Copyright © 2014 Polpass Arul Jose and Solomon Robinson David Jebakumar. All rights reserved. Origanum dictamnus Oil Vapour Suppresses the Development of Grey Mould in Eggplant Fruit In Vitro Mon, 01 Sep 2014 13:17:10 +0000 Grey mould rot (Botrytis cinerea) development in vitro or in eggplant (Solanum melongena L.) fruit was evaluated after treatment with dittany (Origanum dictamnus L.) oil (DIT) and storage at 12°C and 95% relative humidity during or following exposure to the volatiles. DIT volatiles used in different concentration (0-50-100-250 μL/L) and times of exposure (up to 120 h) examined the effects on pathogen development as well as fruit quality parameters. In vitro, fungal colony growth was inhibited with the application of DIT oil (during or after exposure) and/or time of application. Continuous exposure to oils reduced conidial germination and production with fungistatic effects observed in 250 μL/L. In vivo, fungal lesion growth and conidial production reduced in DIT-treated fruits. Interesting, in fruits preexposed to volatiles before fungal inoculation, DIT application induced fruit resistance against the pathogen, by reduced lesion growth and conidial production. Conidial viability reduced in >100 μL/L DIT oil. Fruits exposed to essential oil did not affect fruit quality related attributes in general, while skin lightness (L value) increased in 50 and 100 μL/L DIT oil. The results of the current study indicated that dittany volatiles may be considered as an alternative food preservative, eliminating disease spread in the storage/transit atmospheres. Andriana Stavropoulou, Kostas Loulakakis, Naresh Magan, and Nikos Tzortzakis Copyright © 2014 Andriana Stavropoulou et al. All rights reserved. Differences in Extended-Spectrum Beta-Lactamase Producing Escherichia coli Virulence Factor Genes in the Baltic Sea Region Thu, 28 Aug 2014 12:19:42 +0000 The aim of this study was to compare the prevalence of different virulence factor (VF) genes in extended-spectrum beta-lactamase (ESBL) producing Escherichia coli strains isolated from the Baltic Sea region. A total of 432 strains of phenotypically ESBL positive E. coli were collected from 20 institutions located in Estonia, Latvia, Lithuania, and the region of St. Petersburg in Russia from January to May 2012 and analyzed for phylogenetic group and prevalence of 23 VF genes. The strains were collected from clinical material (urine, blood, wound, and respiratory tract). Bacterial isolates were compared according to phylogenetic group, clinical material, and geographical origin. Most of the VF genes were concentrated within phylogenetic group B2 and/or D. When comparing strains isolated from different countries, it was found that strains originating from Estonia and Latvia belonged mainly to group B2 and strains from Lithuania and Russia mainly to groups B2 and D. The P-fimbrial adhesin gene papEF was more prevalent in Russian strains, colicin gene cvaC in Lithuanian strains, and capsular gene kpsMTII in Latvian strains; serum resistant gene traT was less prevalent in Estonian strains. The regional differences of VF genes remained statistically significant after taking into account the phylogenetic distribution in the countries. Jana Lillo, Kristiine Pai, Arta Balode, Mariia Makarova, Kristi Huik, Siiri Kõljalg, Marina Ivanova, Lidia Kaftyreva, Jolanta Miciuleviciene, Paul Naaber, Kristel Parv, Anastasia Pavelkovich, Tiiu Rööp, Karolin Toompere, Ludmila Suzhaeva, and Epp Sepp Copyright © 2014 Jana Lillo et al. All rights reserved. The Probiotic Bifidobacterium breve B632 Inhibited the Growth of Enterobacteriaceae within Colicky Infant Microbiota Cultures Thu, 28 Aug 2014 08:39:14 +0000 Infant colic is a common gastrointestinal disorder of newborns, mostly related to imbalances in the composition of gut microbiota and particularly to the presence of gas-producing coliforms and to lower levels of Bifidobacteria and Lactobacilli. Probiotics could help to contain this disturbance, with formulations consisting of Lactobacillus strains being the most utilized. In this work, the probiotic strain Bifidobacterium breve B632 that was specifically selected for its ability to inhibit gas-producing coliforms, was challenged against the Enterobacteriaceae within continuous cultures of microbiota from a 2-month-old colicky infant. As confirmed by RAPD-PCR fingerprinting, B. breve B632 persisted in probiotic-supplemented microbiota cultures, accounting for the 64% of Bifidobacteria at the steady state. The probiotic succeeded in inhibiting coliforms, since FISH and qPCR revealed that the amount of Enterobacteriaceae after 18 h of cultivation was 0.42 and 0.44 magnitude orders lower in probiotic-supplemented microbiota cultures than in the control ones. These results support the possibility to move to another level of study, that is, the administration of B. breve B632 to a cohort of colicky newborns, in order to observe the behavior of this strain in vivo and to validate its effect in colic treatment. Marta Simone, Caterina Gozzoli, Andrea Quartieri, Giuseppe Mazzola, Diana Di Gioia, Alberto Amaretti, Stefano Raimondi, and Maddalena Rossi Copyright © 2014 Marta Simone et al. All rights reserved. Possible Use of Bacteriophages Active against Bacillus anthracis and Other B. cereus Group Members in the Face of a Bioterrorism Threat Thu, 28 Aug 2014 08:38:43 +0000 Anthrax is an infectious fatal disease with epidemic potential. Nowadays, bioterrorism using Bacillus anthracis is a real possibility, and thus society needs an effective weapon to neutralize this threat. The pathogen may be easily transmitted to human populations. It is easy to store, transport, and disseminate and may survive for many decades. Recent data strongly support the effectiveness of bacteriophage in treating bacterial diseases. Moreover, it is clear that bacteriophages should be considered a potential incapacitative agent against bioterrorism using bacteria belonging to B. cereus group, especially B. anthracis. Therefore, we have reviewed the possibility of using bacteriophages active against Bacillus anthracis and other species of the B. cereus group in the face of a bioterrorism threat. Ewa Jończyk-Matysiak, Marlena Kłak, Beata Weber-Dąbrowska, Jan Borysowski, and Andrzej Górski Copyright © 2014 Ewa Jończyk-Matysiak et al. All rights reserved. Soil Fungal Resources in Annual Cropping Systems and Their Potential for Management Thu, 28 Aug 2014 00:00:00 +0000 Soil fungi are a critical component of agroecosystems and provide ecological services that impact the production of food and bioproducts. Effective management of fungal resources is essential to optimize the productivity and sustainability of agricultural ecosystems. In this review, we (i) highlight the functional groups of fungi that play key roles in agricultural ecosystems, (ii) examine the influence of agronomic practices on these fungi, and (iii) propose ways to improve the management and contribution of soil fungi to annual cropping systems. Many of these key soil fungal organisms (i.e., arbuscular mycorrhizal fungi and fungal root endophytes) interact directly with plants and are determinants of the efficiency of agroecosystems. In turn, plants largely control rhizosphere fungi through the production of carbon and energy rich compounds and of bioactive phytochemicals, making them a powerful tool for the management of soil fungal diversity in agriculture. The use of crop rotations and selection of optimal plant genotypes can be used to improve soil biodiversity and promote beneficial soil fungi. In addition, other agronomic practices (e.g., no-till, microbial inoculants, and biochemical amendments) can be used to enhance the effect of beneficial fungi and increase the health and productivity of cultivated soils. Walid Ellouze, Ahmad Esmaeili Taheri, Luke D. Bainard, Chao Yang, Navid Bazghaleh, Adriana Navarro-Borrell, Keith Hanson, and Chantal Hamel Copyright © 2014 Walid Ellouze et al. All rights reserved. Functional Screening of Antibiotic Resistance Genes from a Representative Metagenomic Library of Food Fermenting Microbiota Thu, 28 Aug 2014 00:00:00 +0000 Lactic acid bacteria (LAB) represent the predominant microbiota in fermented foods. Foodborne LAB have received increasing attention as potential reservoir of antibiotic resistance (AR) determinants, which may be horizontally transferred to opportunistic pathogens. We have previously reported isolation of AR LAB from the raw ingredients of a fermented cheese, while AR genes could be detected in the final, marketed product only by PCR amplification, thus pointing at the need for more sensitive microbial isolation techniques. We turned therefore to construction of a metagenomic library containing microbial DNA extracted directly from the food matrix. To maximize yield and purity and to ensure that genomic complexity of the library was representative of the original bacterial population, we defined a suitable protocol for total DNA extraction from cheese which can also be applied to other lipid-rich foods. Functional library screening on different antibiotics allowed recovery of ampicillin and kanamycin resistant clones originating from Streptococcus salivarius subsp. thermophilus and Lactobacillus helveticus genomes. We report molecular characterization of the cloned inserts, which were fully sequenced and shown to confer AR phenotype to recipient bacteria. We also show that metagenomics can be applied to food microbiota to identify underrepresented species carrying specific genes of interest. Chiara Devirgiliis, Paola Zinno, Mariarita Stirpe, Simona Barile, and Giuditta Perozzi Copyright © 2014 Chiara Devirgiliis et al. All rights reserved. Dyeing Industry Effluent System as Lipid Production Medium of Neochloris sp. for Biodiesel Feedstock Preparation Wed, 27 Aug 2014 05:23:57 +0000 Microalgae lipid feedstock preparation cost was an important factor in increasing biodiesel fuel hikes. This study was conducted with the concept of implementing an effluent wastewater as lipid production medium for microalgae cultivation. In our study textile dyeing industry effluent was taken as a lipid production medium for Neochloris sp. cultivation. The changes in physicochemical analysis of effluent before and after Neochloris sp. treatment were recorded using standard procedures and AAS analysis. There was especially a reduction in heavy metal like lead (Pb) concentration from 0.002 ppm to 0.001 ppm after Neochloris sp. treatment. Neochloris sp. cultivated in Bold Basal Medium (BBM) (specific algal medium) produced 41.93% total lipid and 36.69% lipid was produced in effluent based cultivation. Surprisingly Neochloris sp. cultivated in effluent was found with enhanced neutral lipid content, and it was confirmed by Nile red fluorescence assay. Further the particular enrichment in oleic acid content of the cells was confirmed with thin layer chromatography (TLC) with oleic acid pure (98%) control. The overall results suggested that textile dyeing industry effluent could serve as the best lipid productive medium for Neochloris sp. biodiesel feedstock preparation. This study was found to have a significant impact on reducing the biodiesel feedstock preparation cost with simultaneous lipid induction by heavy metal stress to microalgae. Vidyadharani Gopalakrishnan and Dhandapani Ramamurthy Copyright © 2014 Vidyadharani Gopalakrishnan and Dhandapani Ramamurthy. All rights reserved. Phenotypic Characterization of Mycoplasma synoviae Induced Changes in the Metabolic and Sensitivity Profile of In Vitro Infected Chicken Chondrocytes Tue, 26 Aug 2014 13:06:54 +0000 In infectious synovitis caused by Mycoplasma synoviae chicken chondrocytes (CCH) may come into direct contact with these bacteria that are also capable of invading CCH in vitro. In this study, phenotype microarrays were used to evaluate the influence of Mycoplasma synoviae on the global metabolic activity of CCH. Therefore, CCH were cultured in the presence of 504 individual compounds, spotted in wells of 11 phenotype microarrays for eukaryotic cells, and exposed to Mycoplasma synoviae membranes or viable Mycoplasma synoviae. Metabolic activity and sensitivity of normal cells versus infected cells were evaluated. Metabolic profiles of CCH treated with viable Mycoplasma synoviae or its membranes were significantly different from those of CCH alone. CCH treated with Mycoplasma synoviae membranes were able to use 48 carbon/nitrogen sources not used by CCH alone. Treatment also influenced ion uptake in CCH and intensified the sensitivity to 13 hormones, 5 immune mediators, and 29 cytotoxic chemicals. CCH were even more sensitive to hormones/immune mediators when exposed to viable Mycoplasma synoviae. Our results indicate that exposure to Mycoplasma synoviae or its membranes induces a wide range of metabolic and sensitivity modifications in CCH that can contribute to pathological processes in the development of infectious synovitis. Daliborka Dušanić, Dušan Benčina, Mojca Narat, and Irena Oven Copyright © 2014 Daliborka Dušanić et al. All rights reserved. Nonthermal Pasteurization of Fermented Green Table Olives by means of High Hydrostatic Pressure Processing Mon, 18 Aug 2014 11:18:10 +0000 Green fermented olives cv. Halkidiki were subjected to different treatments of high hydrostatic pressure (HHP) processing (400, 450, and 500 MPa for 15 or 30 min). Total viable counts, lactic acid bacteria and yeasts/moulds, and the physicochemical characteristics of the product (pH, colour, and firmness) were monitored right after the treatment and after 7 days of storage at 20°C to allow for recovery of injured cells. The treatments at 400 MPa for 15 and 30 min, 450 MPa for 15 and 30 min, and 500 MPa for 15 min were found insufficient as a recovery of the microbiota was observed. The treatment at 500 MPa for 30 min was effective in reducing the olive microbiota below the detection limit of the enumeration method after the treatment and after 1 week of storage and was chosen as being more appropriate for storing olives for an extended time period (5 months). After 5 months of storage at 20°C, no microbiota was detected in treated samples, while significant changes for both HHP treated and untreated olives were observed for colour parameters only (minor degradation). In conclusion, HHP treatment may introduce a reliable nonthermal pasteurization method to extend the microbiological shelf-life of fermented table olives. Anthoula A. Argyri, Efstathios Z. Panagou, George-John E. Nychas, and Chrysoula C. Tassou Copyright © 2014 Anthoula A. Argyri et al. All rights reserved. Implementation of a Computerized Decision Support System to Improve the Appropriateness of Antibiotic Therapy Using Local Microbiologic Data Sun, 17 Aug 2014 09:29:17 +0000 A prospective quasi-experimental study was undertaken in 218 patients with suspicion of nosocomial infection hospitalized in a polyvalent ICU where a new electronic device (GERB) has been designed for antibiotic prescriptions. Two GERB-based applications were developed to provide local resistance maps (LRMs) and preliminary microbiological reports with therapeutic recommendation (PMRTRs). Both applications used the data in the Laboratory Information System of the Microbiology Department to report on the optimal empiric therapeutic option, based on the most likely susceptibility profile of the microorganisms potentially responsible for infection in patients and taking into account the local epidemiology of the hospital department/unit. LRMs were used for antibiotic prescription in 20.2% of the patients and PMRTRs in 78.2%, and active antibiotics against the finally identified bacteria were prescribed in 80.0% of the former group and 82.4% of the latter. When neither LMRs nor PMRTRs were considered for empiric treatment prescription, only around 40% of the antibiotics prescribed were active. Hence, the percentage appropriateness of the empiric antibiotic treatments was significantly higher when LRM or PMRTR guidelines were followed rather than other criteria. LRMs and PMRTRs applications are dynamic, highly accessible, and readily interpreted instruments that contribute to the appropriateness of empiric antibiotic treatments. Manuel Rodriguez-Maresca, Antonio Sorlozano, Magnolia Grau, Rocio Rodriguez-Castaño, Andres Ruiz-Valverde, and Jose Gutierrez-Fernandez Copyright © 2014 Manuel Rodriguez-Maresca et al. All rights reserved. Antibacterial Activity of Pseudonocardia sp. JB05, a Rare Salty Soil Actinomycete against Staphylococcus aureus Thu, 14 Aug 2014 06:41:52 +0000 Staphylococcus aureus is a Gram-positive bacterium that causes many harmful and life-threatening diseases. Some strains of this bacterium are resistant to available antibiotics. This study was designed to evaluate the ability of indigenous actinomycetes to produce antibacterial compounds against S. aureus and characterize the structure of the resultant antibacterial compounds. Therefore, a slightly modified agar well diffusion method was used to determine the antibacterial activity of actinomycete isolates against the test microorganisms. The bacterial extracts with antibacterial activity were fractionated by silica gel and G-25 sephadex column chromatography. Also, the active fractions were analyzed by thin layer chromatography. Finally, the partial structure of the resultant antibacterial compound was characterized by Fourier transform infrared spectroscopy. One of the isolates, which had a broad spectrum and high antibacterial activity, was designated as Pseudonocardia sp. JB05, based on the results of biochemical and 16S rDNA gene sequence analysis. Minimum inhibitory concentration for this bacterium was 40 AU mL−1 against S. aureus. The antibacterial activity of this bacterium was stable after autoclaving, 10% SDS, boiling, and proteinase K. Thin layer chromatography, using anthrone reagent, showed the presence of carbohydrates in the purified antibacterial compound. Finally, FT-IR spectrum of the active compound illustrated hydroxyl groups, hydrocarbon skeleton, and double bond of polygenic compounds in its structure. To the best of our knowledge, this is the first report describing the efficient antibacterial activity by a local strain of Pseudonocardia. The results presented in this work, although at the initial stage in bioactive product characterization, will possibly contribute toward the Pseudonocardia scale-up for the production and identification of the antibacterial compounds. Nesa Jafari, Reza Behroozi, Davoud Farajzadeh, Mohammad Farsi, and Kambiz Akbari-Noghabi Copyright © 2014 Nesa Jafari et al. All rights reserved. Helicobacter pylori in Vegetables and Salads: Genotyping and Antimicrobial Resistance Properties Tue, 12 Aug 2014 12:06:40 +0000 From a clinical and epidemiological perspective, it is important to know which genotypes and antibiotic resistance patterns are present in H. pylori strains isolated from salads and vegetables. Therefore, the present investigation was carried out to find this purpose. Three hundred eighty washed and unwashed vegetable samples and fifty commercial and traditional salad samples were collected from Isfahan, Iran. Samples were cultured and those found positive for H. pylori were analyzed using PCR. Antimicrobial susceptibility testing was performed using disk diffusion method. Seven out of 50 (14%) salad and 52 out of 380 (13.68%) vegetable samples harbored H. pylori. In addition, leek, lettuce, and cabbage were the most commonly contaminated samples (30%). The most prevalent virulence genes were oipA (86.44%) and cagA (57.625). VacA s1a (37.28%) and iceA1 (47.45%) were the most prevalent genotypes. Forty different genotypic combinations were recognized. S1a/cagA+/iceA1/oipA+ (33.89%), s1a/cagA+/iceA2/oipA (30.50%), and m1a/cagA+/iceA1/oipA+ (28.81%) were the most prevalent combined genotypes. Bacterial strains had the highest levels of resistance against metronidazole (77.96%), amoxicillin (67.79%), and ampicillin (61.01%). High similarity in the genotyping pattern of H. pylori among vegetable and salad samples and human specimens suggests that vegetable and salads may be the sources of the bacteria. Emad Yahaghi, Faham Khamesipour, Fatemeh Mashayekhi, Farhad Safarpoor Dehkordi, Mohammad Hossein Sakhaei, Mojtaba Masoudimanesh, and Maryam Khayyat Khameneie Copyright © 2014 Emad Yahaghi et al. All rights reserved. Biodegradation Ability and Catabolic Genes of Petroleum-Degrading Sphingomonas koreensis Strain ASU-06 Isolated from Egyptian Oily Soil Sun, 10 Aug 2014 08:25:20 +0000 Polycyclic aromatic hydrocarbons (PAHs) are serious pollutants and health hazards. In this study, 15 PAHs-degrading bacteria were isolated from Egyptian oily soil. Among them, one Gram-negative strain (ASU-06) was selected and biodegradation ability and initial catabolic genes of petroleum compounds were investigated. Comparison of 16S rRNA gene sequence of strain ASU-06 to published sequences in GenBank database as well as phylogenetic analysis identified ASU-06 as Sphingomonas koreensis. Strain ASU-06 degraded 100, 99, 98, and 92.7% of 100 mg/L naphthalene, phenanthrene, anthracene, and pyrene within 15 days, respectively. When these PAHs present in a mixed form, the enhancement phenomenon appeared, particularly in the degradation of pyrene, whereas the degradation rate was 98.6% within the period. This is the first report showing the degradation of different PAHs by this species. PCR experiments with specific primers for catabolic genes alkB, alkB1, nahAc, C12O, and C23O suggested that ASU-06 might possess genes for aliphatic and PAHs degradation, while PAH-RHDαGP gene was not detected. Production of biosurfactants and increasing cell-surface hydrophobicity were investigated. GC/MS analysis of intermediate metabolites of studied PAHs concluded that this strain utilized these compounds via two main pathways, and phthalate was the major constant product that appeared in each day of the degradation period. Abd El-Latif Hesham, Asmaa M. M. Mawad, Yasser M. Mostafa, and Ahmed Shoreit Copyright © 2014 Abd El-Latif Hesham et al. All rights reserved. The Association of Toll-Like Receptor 4 Polymorphism with Hepatitis C Virus Infection in Saudi Arabian Patients Sun, 10 Aug 2014 08:19:43 +0000 Hepatitis C virus (HCV) is a single stranded RNA virus. It affects millions of people worldwide and is considered as a leading cause of liver diseases including cirrhosis and hepatocellular carcinoma. A recent study reported that TLR4 gene polymorphisms are good prognostic predictors and are associated with protection from liver fibrosis among Caucasians. This study aims to investigate the implication of genetic polymorphisms of TLR4 gene on the HCV infection in Saudi Arabian patients. Two SNPs in the TLR4 gene, rs4986790 (A/G) and rs4986791 (C/T), were genotyped in 450 HCV patients and 600 uninfected controls. The association analysis confirmed that both SNPs showed a significant difference in their distribution between HCV-infected patients and uninfected control subjects (; , 95% –0.581) and (; , 95% –0.443), respectively. More importantly, haplotype analysis revealed that four haplotypes, AC, GT, GC, and AT (rs4986790, rs4986791), were significantly associated with HCV infection when compared with control subjects. One haplotype AC was more prominently found when chronic HCV-infected patients were compared with cirrhosis/HCC patients (frequency = 94.7% and ). Both TLR4 SNPs under investigation were found to be significantly implicated with HCV-infection among Saudi Arabian population. Ahmed A. Al-Qahtani, Mashael R. Al-Anazi, Fahad Al-Zoghaibi, Ayman A. Abdo, Faisal M. Sanai, Mohammed Q. Khan, Ali Albenmousa, Hamad I. Al-Ashgar, and Mohammed N. Al-Ahdal Copyright © 2014 Ahmed A. Al-Qahtani et al. All rights reserved. Participation of Integrin α5β1 in the Fibronectin-Mediated Adherence of Enteroaggregative Escherichia coli to Intestinal Cells Thu, 07 Aug 2014 09:28:27 +0000 Adherence to the intestinal epithelia is a key feature in enteroaggregative Escherichia coli (EAEC) infection. The aggregative adherence fimbriae (AAFs) are involved in EAEC interaction with receptors at the surface of intestinal cells. We and others have demonstrated that fibronectin is a receptor for AAF/II fimbriae. Considering that the major cellular receptor of fibronectin is integrin , in this study we evaluated the participation of this receptor in the fibronectin-mediated adherence of EAEC strain 042 to intestinal cells. We found that EAEC strain 042 has the ability to bind directly and indirectly to integrin ; direct binding was not mediated by AAF/II fimbriae and indirect binding was mediated by AAF/II and fibronectin. Coimmunoprecipitation assays confirmed the formation of the complex AafA/fibronectin/integrin . To evaluate EAEC adherence to intestinal cells, T84 cells were incubated with fibronectin and an antibody that blocks the interaction region of integrin to fibronectin, the RGD site. Under these conditions, we found the number of adherent bacteria to epithelial cells significantly reduced. Additionally, fibronectin-mediated adherence of EAEC strain 042 was abolished in HEp-2 cells transfected with integrin shRNA. Altogether, our data support the involvement of integrin in the fibronectin-mediated EAEC binding to intestinal cells. Mariana Izquierdo, Alejandra Alvestegui, James P. Nataro, Fernando Ruiz-Perez, and Mauricio J. Farfan Copyright © 2014 Mariana Izquierdo et al. All rights reserved. Eucalyptus Essential Oil as a Natural Food Preservative: In Vivo and In Vitro Antiyeast Potential Thu, 07 Aug 2014 07:21:23 +0000 In this study, the application of eucalyptus essential oil/vapour as beverages preservative is reported. The chemical composition of eucalyptus oil was determined by gas chromatography-mass spectrometry (GC-MS) and solid phase microextraction GC-MS (SPME/GC-MS) analyses. GC-MS revealed that the major constituents were 1,8-cineole (80.5%), limonene (6.5%), -pinene (5%), and -terpinene (2.9%) while SPME/GC-MS showed a relative reduction of 1,8-cineole (63.9%) and an increase of limonene (13.8%), -pinene (8.87%), and -terpinene (3.98%). Antimicrobial potential of essential oil was initially determined in vitro against 8 different food spoilage yeasts by disc diffusion, disc volatilization, and microdilution method. The activity of eucalyptus vapours was significantly higher than the eucalyptus oil. Minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) varied from 0.56 to 4.50 mg/mL and from 1.13 to 9 mg/mL, respectively. Subsequently, the combined efficacy of essential oil and thermal treatment were used to evaluate the preservation of a mixed fruit juice in a time-dependent manner. These results suggest eucalyptus oil as a potent inhibitor of food spoilage yeasts not only in vitro but also in a real food system. Currently, this is the first report that uses eucalyptus essential oil for fruit juice preservation against food spoiling yeast. Amit Kumar Tyagi, Danka Bukvicki, Davide Gottardi, Giulia Tabanelli, Chiara Montanari, Anushree Malik, and Maria Elisabetta Guerzoni Copyright © 2014 Amit Kumar Tyagi et al. All rights reserved. Generation of Dipeptidyl Peptidase-IV-Inhibiting Peptides from β-Lactoglobulin Secreted by Lactococcus lactis Sun, 03 Aug 2014 08:28:25 +0000 Previous studies showed that hydrolysates of β-lactoglobulin (BLG) prepared using gastrointestinal proteases strongly inhibit dipeptidyl peptidase-IV (DPP-IV) activity in vitro. In this study, we developed a BLG-secreting Lactococcus lactis strain as a delivery vehicle and in situ expression system. Interestingly, trypsin-digested recombinant BLG from L. lactis inhibited DPP-IV activity, suggesting that BLG-secreting L. lactis may be useful in the treatment of type 2 diabetes mellitus. Suguru Shigemori, Kazushi Oshiro, Pengfei Wang, Yoshinari Yamamoto, Yeqin Wang, Takashi Sato, Yutaka Uyeno, and Takeshi Shimosato Copyright © 2014 Suguru Shigemori et al. All rights reserved. N-Acyl Homoserine Lactone-Mediated Quorum Sensing with Special Reference to Use of Quorum Quenching Bacteria in Membrane Biofouling Control Thu, 24 Jul 2014 09:59:53 +0000 Membrane biofouling remains a severe problem to be addressed in wastewater treatment systems affecting reactor performance and economy. The finding that many wastewater bacteria rely on N-acyl homoserine lactone-mediated quorum sensing to synchronize their activities essential for biofilm formations; the quenching bacterial quorum sensing suggests a promising approach for control of membrane biofouling. A variety of quorum quenching compounds of both synthetic and natural origin have been identified and found effective in inhibition of membrane biofouling with much less environmental impact than traditional antimicrobials. Work over the past few years has demonstrated that enzymatic quorum quenching mechanisms are widely conserved in several prokaryotic organisms and can be utilized as a potent tool for inhibition of membrane biofouling. Such naturally occurring bacterial quorum quenching mechanisms also play important roles in microbe-microbe interactions and have been used to develop sustainable nonantibiotic antifouling strategies. Advances in membrane fabrication and bacteria entrapment techniques have allowed the implication of such quorum quenching bacteria for better design of membrane bioreactor with improved antibiofouling efficacies. In view of this, the present paper is designed to review and discuss the recent developments in control of membrane biofouling with special emphasis on quorum quenching bacteria that are applied in membrane bioreactors. Harshad Lade, Diby Paul, and Ji Hyang Kweon Copyright © 2014 Harshad Lade et al. All rights reserved. Biotechnological Applications Derived from Microorganisms of the Atacama Desert Wed, 23 Jul 2014 11:07:20 +0000 The Atacama Desert in Chile is well known for being the driest and oldest desert on Earth. For these same reasons, it is also considered a good analog model of the planet Mars. Only a few decades ago, it was thought that this was a sterile place, but in the past years fascinating adaptations have been reported in the members of the three domains of life: low water availability, high UV radiation, high salinity, and other environmental stresses. However, the biotechnological applications derived from the basic understanding and characterization of these species, with the notable exception of copper bioleaching, are still in its infancy, thus offering an immense potential for future development. Armando Azua-Bustos and Carlos González-Silva Copyright © 2014 Armando Azua-Bustos and Carlos González-Silva. All rights reserved. Alpha-Melanocyte Stimulating Hormone: An Emerging Anti-Inflammatory Antimicrobial Peptide Wed, 23 Jul 2014 00:00:00 +0000 The alpha-melanocyte stimulating hormone (-MSH) is a neuropeptide belonging to the melanocortin family. It is well known for its anti-inflammatory and antipyretic effects and shares several characteristics with antimicrobial peptides (AMPs). There have been some recent reports about the direct antimicrobial activity of -MSH against various microbes belonging to both fungal and bacterial pathogens. Similar to -MSH’s anti-inflammatory properties, its C-terminal residues also exhibit antimicrobial activity parallel to that of the entire peptide. This review is focused on the current findings regarding the direct antimicrobial potential and immunomodulatory mechanism of -MSH and its C-terminal fragments, with particular emphasis on the prospects of -MSH based peptides as a strong anti-infective agent. Madhuri Singh and Kasturi Mukhopadhyay Copyright © 2014 Madhuri Singh and Kasturi Mukhopadhyay. All rights reserved. Recombinant Lysostaphin Protects Mice from Methicillin-Resistant Staphylococcus aureus Pneumonia Wed, 16 Jul 2014 12:08:55 +0000 The advent of methicillin-resistant Staphylococcus aureus (MRSA) and the frequent and excessive abuse of ventilators have made MRSA pneumonia an inordinate threat to human health. Appropriate antibacterial therapies are crucial, including the use of lysostaphin as an alternative to antibiotics. To explore the potential use of lysostaphin as a therapeutic agent for MRSA pneumonia, mice were intranasally infected with MRSA and then treated with recombinant lysostaphin (rLys; 45 mg/kg in the high-dose group and 1 mg/kg in the low-dose group) (0.33 mg/mL, 15 mg/mL), vancomycin (120 mg/kg) (40 mg/mL), or phosphate-buffered saline (PBS, negative control) 4 h after infection. Therapeutic efficacy was assessed by mouse survival, lung histopathology, bacterial density in the lungs, bodyweight, lung weight, temperature, white blood cells counts, lymphocytes counts, granulocytes counts, and monocytes counts. The mice treated with rLys showed lower mortality, less lung parenchymal damage, and lower bacterial density at metastatic tissue sites than mice treated with PBS or vancomycin. The overall mortality was 100%, 60%, 40%, and 60% for the control, vancomycin, high-dose rLys, and low-dose rLys groups, respectively. These findings indicate that, as a therapeutic agent for MRSA pneumonia, lysostaphin exerts profound protective effects in mice against the morbidity and mortality associated with S. aureus pneumonia. Chen Chen, Huahao Fan, Yong Huang, Fan Peng, Hang Fan, Shoujun Yuan, and Yigang Tong Copyright © 2014 Chen Chen et al. All rights reserved. Diversity and Enzymatic Profiling of Halotolerant Micromycetes from Sebkha El Melah, a Saharan Salt Flat in Southern Tunisia Wed, 16 Jul 2014 12:04:04 +0000 Twenty-one moderately halotolerant fungi have been isolated from sample ashes collected from Sebkha El Melah, a Saharan salt flat located in southern Tunisia. Based on morphology and sequence inference from the internal transcribed spacer regions, 28S rRNA gene and other specific genes such as β-tubulin, actin, calmodulin, and glyceraldehyde-3-phosphate dehydrogenase, the isolates were found to be distributed over 15 taxa belonging to 6 genera of Ascomycetes: Cladosporium (), Alternaria (), Aspergillus (), Penicillium (), Ulocladium (), and Engyodontium (). Their tolerance to different concentrations of salt in solid and liquid media was examined. Excepting Cladosporium cladosporioides JA18, all isolates were considered as alkali-halotolerant since they were able to grow in media containing 10% of salt with an initial pH 10. All isolates were resistant to oxidative stresses and low temperature whereas 5 strains belonging to Alternaria, Ulocladium, and Aspergillus genera were able to grow at 45°C. The screening of fungal strains for sets of enzyme production, namely, cellulase (CMCase), amylase, protease, lipase, and laccase, in presence of 10% NaCl, showed a variety of extracellular hydrolytic and oxidative profiles. Protease was the most abundant enzyme produced whereas laccase producers were members of the genus Cladosporium. Atef Jaouani, Mohamed Neifar, Valeria Prigione, Amani Ayari, Imed Sbissi, Sonia Ben Amor, Seifeddine Ben Tekaya, Giovanna Cristina Varese, Ameur Cherif, and Maher Gtari Copyright © 2014 Atef Jaouani et al. All rights reserved. A Novel Promising Strain of Trichoderma evansii (WF-3) for Extracellular α-Galactosidase Production by Utilizing Different Carbon Sources under Optimized Culture Conditions Sun, 13 Jul 2014 08:51:16 +0000 A potential fungal strain of Trichoderma sp. (WF-3) was isolated and selected for the production of α-galactosidase. Optimum conditions for mycelial growth and enzyme induction were determined. Basal media selected for the growth of fungal isolate containing different carbon sources like guar gum (GG), soya bean meal (SM), and wheat straw (WS) and combinations of these carbon substrates with basic sugars like galactose and sucrose were used to monitor their effects on α-galactosidase production. The results of this study indicated that galactose and sucrose enhanced the enzyme activity in guar gum (GG) and wheat straw (WS). Maximum α-galactosidase production (213.63 UmL−1) was obtained when the basic medium containing GG is supplemented with galactose (5 mg/mL). However, the presence of galactose and sucrose alone in the growth media shows no effect. Soya meal alone was able to support T. evansii to produce maximum enzyme activity (170.36 UmL−1). The incubation time, temperature, and pH for the maximum enzyme synthesis were found to be 120 h (5 days), 28°C, and 4.5–5.5, respectively. All the carbon sources tested exhibited maximum enzyme production at 10 mg/mL concentration. Among the metal ions tested, Hg was found to be the strongest inhibitor of the enzyme. Among the chelators, EDTA acted as stronger inhibitor than succinic acid. Aishwarya Chauhan, Nikhat Jamal Siddiqi, and Bechan Sharma Copyright © 2014 Aishwarya Chauhan et al. All rights reserved. Geodermatophilus poikilotrophi sp. nov.: A Multitolerant Actinomycete Isolated from Dolomitic Marble Wed, 09 Jul 2014 14:05:46 +0000 A novel Gram-reaction-positive, aerobic actinobacterium, tolerant to mitomycin C, heavy metals, metalloids, hydrogen peroxide, desiccation, and ionizing- and UV-radiation, designated G18T, was isolated from dolomitic marble collected from outcrops in Samara (Namibia). The growth range was 15–35°C, at pH 5.5–9.5 and in presence of 1% NaCl, forming greenish-black coloured colonies on GYM Streptomyces agar. Chemotaxonomic and molecular characteristics of the isolate matched those described for other representatives of the genus Geodermatophilus. The peptidoglycan contained meso-diaminopimelic acid as diagnostic diaminoacid. The main phospholipids were phosphatidylethanolamine, phosphatidylcholine, phosphatidylinositol, and small amount of diphosphatidylglycerol. MK-9(H4) was the dominant menaquinone and galactose was detected as diagnostic sugar. The major cellular fatty acids were branched-chain saturated acids iso-C16:0 and iso-C15:0 and the unsaturated C17:1ω8c and C16:1ω7c. The 16S rRNA gene showed 97.4–99.1% sequence identity with the other representatives of genus Geodermatophilus. Based on phenotypic results and 16S rRNA gene sequence analysis, strain G18T is proposed to represent a novel species, Geodermatophilus poikilotrophi. Type strain is G18T (= DSM 44209T = CCUG 63018T). The INSDC accession number is HF970583. The novel R software package lethal was used to compute the lethal doses with confidence intervals resulting from tolerance experiments. Maria del Carmen Montero-Calasanz, Benjamin Hofner, Markus Göker, Manfred Rohde, Cathrin Spröer, Karima Hezbri, Maher Gtari, Peter Schumann, and Hans-Peter Klenk Copyright © 2014 Maria del Carmen Montero-Calasanz et al. All rights reserved. Fungi Treated with Small Chemicals Exhibit Increased Antimicrobial Activity against Facultative Bacterial and Yeast Pathogens Wed, 09 Jul 2014 11:04:36 +0000 For decades, fungi have been the main source for the discovery of novel antimicrobial drugs. Recent sequencing efforts revealed a still high number of so far unknown “cryptic” secondary metabolites. The production of these metabolites is presumably epigenetically silenced under standard laboratory conditions. In this study, we investigated the effect of six small mass chemicals, of which some are known to act as epigenetic modulators, on the production of antimicrobial compounds in 54 spore forming fungi. The antimicrobial effect of fungal samples was tested against clinically facultative pathogens and multiresistant clinical isolates. In total, 30 samples of treated fungi belonging to six different genera reduced significantly growth of different test organisms compared to the untreated fungal sample (growth log reduction 0.3–4.3). For instance, the pellet of Penicillium restrictum grown in the presence of butyrate revealed significant higher antimicrobial activity against Staphylococcus (S.) aureus and multiresistant S. aureus strains and displayed no cytotoxicity against human cells, thus making it an ideal candidate for antimicrobial compound discovery. Our study shows that every presumable fungus, even well described fungi, has the potential to produce novel antimicrobial compounds and that our approach is capable of rapidly filling the pipeline for yet undiscovered antimicrobial substances. Christoph Zutz, Dragana Bandian, Bernhard Neumayer, Franz Speringer, Markus Gorfer, Martin Wagner, Joseph Strauss, and Kathrin Rychli Copyright © 2014 Christoph Zutz et al. All rights reserved. The Bacterial Contamination of Allogeneic Bone and Emergence of Multidrug-Resistant Bacteria in Tissue Bank Tue, 08 Jul 2014 12:49:31 +0000 Present study was carried out for the microbiological evaluation of allogeneic bone processed from femoral heads. A total 60 bacterial isolates comprising five different species including Streptococcus spp., Staphylococcus spp., Klebsiella spp., Bacillus spp., and Pseudomonas spp. were characterized based on their cultural and biochemical characteristics. Average bioburden was ranged from to  cfu/gm. The majority (81.7%) of the microbial contaminants were detected as Gram positive with the predominant organism being skin commensal coagulase negative Staphylococci (43.3%). Antimicrobial resistance was evaluated by the activities of 14 broad and narrow spectrum antibiotic discs. Comparing the overall pattern, marked resistance was noted against Penicillin and Amoxicillin 100% (60/60). The most effective single antibiotics were Gentamicin, Tobramycin, and Ofloxacin which were bactericidal against 100% (60/60) isolates. Multidrug resistance (MDR) was confirmed in 70% (42/60) of the samples. Among them, the most prevalent antibiotypes were Penicillin, Amoxicillin, Oxacillin, Polymyxin, and Cefpodoxime (80% of total MDR). The study results revealed higher contamination rate on bone allografts and recommend the implementation of good tissue banking practices during tissue procurement, processing, and storage in order to minimize the chances of contamination. Fahmida Binte Atique and Md. Masudur Rahman Khalil Copyright © 2014 Fahmida Binte Atique and Md. Masudur Rahman Khalil. All rights reserved. Specific Growth Rate Determines the Sensitivity of Escherichia coli to Lactic Acid Stress: Implications for Predictive Microbiology Tue, 08 Jul 2014 00:00:00 +0000 This study tested the hypothesis that sensitivity of Escherichia coli to lactic acid at concentrations relevant for fermented sausages (pH 4.6, 150 mM lactic acid, , temperature = 20 or 27°C) increases with increasing growth rate. For E. coli strain 683 cultured in TSB in chemostat or batch, subsequent inactivation rates when exposed to lactic acid stress increased with increasing growth rate at harvest. A linear relationship between growth rate at harvest and inactivation rate was found to describe both batch and chemostat cultures. The maximum difference in T90, the estimated times for a one-log reduction, was 10 hours between bacteria harvested during the first 3 hours of batch culture, that is, at different growth rates. A 10-hour difference in T90 would correspond to measuring inactivation at 33°C or 45°C instead of 37°C based on relationships between temperature and inactivation. At similar harvest growth rates, inactivation rates were lower for bacteria cultured at 37°C than at 15–20°C. As demonstrated for E. coli 683, culture conditions leading to variable growth rates may contribute to variable lactic acid inactivation rates. Findings emphasize the use and reporting of standardised culture conditions and can have implications for the interpretation of data when developing inactivation models. Roland Lindqvist and Gunilla Barmark Copyright © 2014 Roland Lindqvist and Gunilla Barmark. All rights reserved. Production of Conjugated Linoleic and Conjugated α-Linolenic Acid in a Reconstituted Skim Milk-Based Medium by Bifidobacterial Strains Isolated from Human Breast Milk Sun, 06 Jul 2014 12:09:59 +0000 Eight bifidobacterial strains isolated from human breast milk have been tested for their abilities to convert linoleic acid (LA) and α-linolenic acid (LNA) to conjugated linoleic acid (CLA) and conjugated α-linolenic acid (CLNA), respectively. These bioactive lipids display important properties that may contribute to the maintenance and improvement human health. Three selected Bifidobacterium breve strains produced CLA from LA and CLNA from LNA in MRS (160–170 and 210–230 μg mL−1, resp.) and, also, in reconstituted skim milk (75–95 and 210–244 μg mL−1, resp.). These bifidobacterial strains were also able to simultaneously produce both CLA (90–105 μg mL−1) and CLNA (290–320 μg mL−1) in reconstituted skim milk. Globally, our findings suggest that these bifidobacterial strains are potential candidates for the design of new fermented dairy products naturally containing very high concentrations of these bioactive lipids. To our knowledge, this is the first study describing CLNA production and coproduction of CLA and CLNA by Bifidobacterium breve strains isolated from human milk in reconstituted skim milk. María Antonia Villar-Tajadura, Luis Miguel Rodríguez-Alcalá, Virginia Martín, Aránzazu Gómez de Segura, Juan Miguel Rodríguez, Teresa Requena, and Javier Fontecha Copyright © 2014 María Antonia Villar-Tajadura et al. All rights reserved. Emerging Microbial Concerns in Food Safety and New Control Measures Sun, 06 Jul 2014 06:18:06 +0000 Moreno Bondi, Patrizia Messi, Prakash M. Halami, Chrissanthy Papadopoulou, and Simona de Niederhausern Copyright © 2014 Moreno Bondi et al. All rights reserved. Clonality and Resistome Analysis of KPC-Producing Klebsiella pneumoniae Strain Isolated in Korea Using Whole Genome Sequencing Thu, 03 Jul 2014 13:40:18 +0000 We analyzed the whole genome sequence and resistome of the outbreak Klebsiella pneumoniae strain MP14 and compared it with those of K. pneumoniae carbapenemase- (KPC-) producing isolates that showed high similarity in the NCBI genome database. A KPC-2-producing multidrug-resistant (MDR) K. pneumoniae clinical isolate was obtained from a patient admitted to a Korean hospital in 2011. The strain MP14 was resistant to all tested β-lactams including monobactam, amikacin, levofloxacin, and cotrimoxazole, but susceptible to tigecycline and colistin. Resistome analysis showed the presence of -lactamase genes including , , , and . MP14 also possessed aac(6′-)Ib, aadA2, and aph(3′-)Ia as aminoglycoside resistance-encoding genes, mph(A) for macrolides, oqxA and oqxB for quinolone, catA1 for phenicol, sul1 for sulfonamide, and dfrA12 for trimethoprim. Both SNP tree and cgMLST analysis showed the close relatedness with the KPC producers (KPNIH strains) isolated from an outbreak in the USA and colistin-resistant strains isolated in Italy. The plasmid-scaffold genes in plasmids pKpQil, pKpQil-IT, pKPN3, or pKPN-IT were identified in MP14, KPNIH, and Italian strains. The KPC-2-producing MDR K. pneumoniae ST258 stain isolated in Korea was highly clonally related with MDR K. pneumoniae strains from the USA and Italy. Global spread of KPC-producing K. pneumoniae is a worrying phenomenon. Yangsoon Lee, Bong-Soo Kim, Jongsik Chun, Ji Hyun Yong, Yeong Seon Lee, Jung Sik Yoo, Dongeun Yong, Seong Geun Hong, Roshan D’Souza, Kenneth S. Thomson, Kyungwon Lee, and Yunsop Chong Copyright © 2014 Yangsoon Lee et al. All rights reserved. Prophages in Enterococcal Isolates from Renal Transplant Recipients: Renal Failure Etiologies Promote Selection of Strains Thu, 03 Jul 2014 09:11:19 +0000 Infections caused by commensal bacteria may be fatal for the patients under immunosuppressive therapy. This results also from difficulty in identification of high risk strains. Enterococcal infections are increasingly frequent but despite many studies on virulence traits, the difference between commensal and pathogenic strains remains unclear. Prophages are newly described as important elements in competition between strains during colonization, as well as pathogenicity of the strains. Here we evaluate a difference in presence of pp4, pp1, and pp7 prophages and ASA (aggregation substance) gene expression in enterococcal isolates from renal transplant recipients (RTx) with different etiology of the end-stage renal failure. Prophages sequence was screened by PCR in strains of Enterococcus faecalis isolated from urine and feces of 19 RTx hospitalized at Medical University of Gdansk and 18 healthy volunteers. FLOW-FISH method with use of linear locked nucleic acid (LNA) probe was used to assess the ASA gene expression. Additionally, ability of biofilm formation was screened by crystal violet staining method. Presence of prophages was more frequent in fecal isolates from immunocompromised patients than in isolates from healthy volunteers. Additionally, both composition of prophages and ASA gene expression were related to the etiology of renal disease. Agnieszka Daca, Tomasz Jarzembowski, Jacek M. Witkowski, Ewa Bryl, Bolesław Rutkowski, and Alicja Dębska-Ślizień Copyright © 2014 Agnieszka Daca et al. All rights reserved. Probiotic Potential of Lactobacillus Strains with Antimicrobial Activity against Some Human Pathogenic Strains Thu, 03 Jul 2014 08:23:53 +0000 The objective of this study was to isolate, identify, and characterize some lactic acid bacterial strains from human milk, infant feces, and fermented grapes and dates, as potential probiotics with antimicrobial activity against some human pathogenic strains. One hundred and forty bacterial strains were isolated and, after initial identification and a preliminary screening for acid and bile tolerance, nine of the best isolates were selected and further identified using 16 S rRNA gene sequences. The nine selected isolates were then characterized in vitro for their probiotic characteristics and their antimicrobial activities against some human pathogens. Results showed that all nine isolates belonged to the genus Lactobacillus. They were able to tolerate pH 3 for 3 h, 0.3% bile salts for 4 h, and 1.9 mg/mL pancreatic enzymes for 3 h. They exhibited good ability to attach to intestinal epithelial cells and were not resistant to the tested antibiotics. They also showed good antimicrobial activities against the tested pathogenic strains of humans, and most of them exhibited stronger antimicrobial activity than the reference strain L. casei Shirota. Thus, the nine Lactobacillus strains could be considered as potential antimicrobial probiotic strains against human pathogens and should be further studied for their human health benefits. Parisa Shokryazdan, Chin Chin Sieo, Ramasamy Kalavathy, Juan Boo Liang, Noorjahan Banu Alitheen, Mohammad Faseleh Jahromi, and Yin Wan Ho Copyright © 2014 Parisa Shokryazdan et al. All rights reserved. Predictive Symptoms and Signs of Severe Dengue Disease for Patients with Dengue Fever: A Meta-Analysis Tue, 01 Jul 2014 11:44:09 +0000 The aim of the meta-analysis was to provide more solid evidence for the reliability of the new classification. A systematic literature search was performed using PubMed, Armed Forces Pest Management Board Literature Retrieval System, and Google Scholar up to August 2012. A pooled odds ratio (OR) was calculated using either a random-effect or a fixed-effect model. A total of 16 papers were identified. Among the 11 factors studied, five symptoms demonstrated an increased risk for SDD, including bleeding [OR: 13.617; 95% confidence interval (CI): 3.281, 56.508], vomiting/nausea (OR: 1.692; 95% CI: 1.256, 2.280), abdominal pain (OR: 2.278; 95% CI: 1.631, 3.182), skin rashes (OR: 2.031; 95% CI: 1.269, 3.250), and hepatomegaly (OR: 4.751; 95% CI: 1.769, 12.570). Among the four bleeding-related symptoms including hematemesis, melena, gum bleeding, and epistaxis, only hematemesis (OR: 6.174; 95% CI: 2.66, 14.334; ) and melena (OR: 10.351; 95% CI: 3.065, 34.956; ) were significantly associated with SDD. No significant associations with SDD were found for gender, lethargy, retroorbital pain, diarrhea, or tourniquet test, whereas headache appeared protective (OR: 0.555; 95% CI: 0.455, 0.676). The meta-analysis suggests that bleeding (hematemesis/melena), vomiting/nausea, abdominal pain, skin rashes, and hepatomegaly may predict the development of SDD in patients with DF, while headache may predict otherwise. H. Zhang, Y. P. Zhou, H. J. Peng, X. H. Zhang, F. Y. Zhou, Z. H. Liu, and X. G. Chen Copyright © 2014 H. Zhang et al. All rights reserved. Blood Stream Infections Tue, 01 Jul 2014 08:48:56 +0000 Renu Bharadwaj, Abhijit Bal, Ketoki Kapila, Vidya Mave, and Amita Gupta Copyright © 2014 Renu Bharadwaj et al. All rights reserved. Amelioration of Colitis in Mouse Model by Exploring Antioxidative Potentials of an Indigenous Probiotic Strain of Lactobacillus fermentum Lf1 Tue, 01 Jul 2014 07:38:49 +0000 Based on the preliminary screening of eight indigenous putative probiotic Lactobacilli, Lactobacillus fermentum Lf1 was selected for assessing its antioxidative efficacy in DSS colitis mouse model based on its ability to enhance the expression of “Nrf2” by 6.43-fold and malondialdehyde (MDA) inhibition by 78.1  ±  0.24% in HT-29 cells under H2O2 stress. The Disease Activity Index and histological scores of Lf1-treated mice were lower than the control group. However, expression of “Nrf2” was not observed in Lf1-treated mice. A significant increase in the expression of antioxidative enzymes such as SOD2 and TrxR-1 was recorded in both of the groups. The expression of SOD2 was significantly downregulated in colitis-induced mice by −100.00-fold relative to control group, and the downregulation was considerably reduced to −37.04-fold in colitis Lf1 treatment group. Almost, a similar trend was recorded in case of “thioredoxin” expression, though “CAT” was refractile to expression. The Lf1-treated group had decreased malondialdehyde level as compared to colitis control (37.92  ±  6.31 versus 91.13  ±  5.76 μM/g). These results point towards Lf1-induced activation of the antioxidant enzyme system in the mouse model and its prospects to be explored as a new strategy for IBD management. Ritu Chauhan, Aparna Sudhakaran Vasanthakumari, Harsh Panwar, Rashmi H. Mallapa, Raj Kumar Duary, Virender Kumar Batish, and Sunita Grover Copyright © 2014 Ritu Chauhan et al. All rights reserved. Reversal of Ampicillin Resistance in MRSA via Inhibition of Penicillin-Binding Protein 2a by Acalypha wilkesiana Mon, 30 Jun 2014 12:28:32 +0000 The inhibitory activity of a semipure fraction from the plant, Acalypha wilkesiana assigned as 9EA-FC-B, alone and in combination with ampicillin, was studied against methicillin-resistant Staphylococcus aureus (MRSA). In addition, effects of the combination treatment on PBP2a expression were investigated. Microdilution assay was used to determine the minimal inhibitory concentrations (MIC). Synergistic effects of 9EA-FC-B with ampicillin were determined using the fractional inhibitory concentration (FIC) index and kinetic growth curve assay. Western blot experiments were carried out to study the PBP2a expression in treated MRSA cultures. The results showed a synergistic effect between ampicillin and 9EA-FC-B treatment with the lowest FIC index of 0.19 (synergism 0.5). The presence of 9EA-FC-B reduced the MIC of ampicillin from 50 to 1.56 μg mL−1. When ampicillin and 9EA-FC-B were combined at subinhibitory level, the kinetic growth curves were suppressed. The antibacterial effect of 9EA-FC-B and ampicillin was shown to be synergistic. The synergism is due the ability of 9EA-FC-B to suppress the activity of PBP2a, thus restoring the susceptibility of MRSA to ampicillin. Corilagin was postulated to be the constituent responsible for the synergistic activity showed by 9EA-FC-B. Carolina Santiago, Ee Leen Pang, Kuan-Hon Lim, Hwei-San Loh, and Kang Nee Ting Copyright © 2014 Carolina Santiago et al. All rights reserved. Fresh-Cut Pineapple as a New Carrier of Probiotic Lactic Acid Bacteria Sun, 29 Jun 2014 08:30:31 +0000 Due to the increasing interest for healthy foods, the feasibility of using fresh-cut fruits to vehicle probiotic microorganisms is arising scientific interest. With this aim, the survival of probiotic lactic acid bacteria, belonging to Lactobacillus plantarum and Lactobacillus fermentum species, was monitored on artificially inoculated pineapple pieces throughout storage. The main nutritional, physicochemical, and sensorial parameters of minimally processed pineapples were monitored. Finally, probiotic Lactobacillus were further investigated for their antagonistic effect against Listeria monocytogenes and Escherichia coli O157:H7 on pineapple plugs. Our results show that at eight days of storage, the concentration of L. plantarum and L. fermentum on pineapples pieces ranged between 7.3 and 6.3 log cfu g−1, respectively, without affecting the final quality of the fresh-cut pineapple. The antagonistic assays indicated that L. plantarum was able to inhibit the growth of both pathogens, while L. fermentum was effective only against L. monocytogenes. This study suggests that both L. plantarum and L. fermentum could be successfully applied during processing of fresh-cut pineapples, contributing at the same time to inducing a protective effect against relevant foodborne pathogens. Pasquale Russo, Maria Lucia Valeria de Chiara, Anna Vernile, Maria Luisa Amodio, Mattia Pia Arena, Vittorio Capozzi, Salvatore Massa, and Giuseppe Spano Copyright © 2014 Pasquale Russo et al. All rights reserved. Differential Response of Oyster Shell Powder on Enzyme Profile and Nutritional Value of Oyster Mushroom Pleurotus florida PF05 Thu, 26 Jun 2014 13:27:56 +0000 Oyster mushroom Pleurotus florida was cultivated on different combinations of wheat straw (WS) as basal substrate and oyster shell powder (OSP) supplement. The OSP supplementation considerably responded to different cultivation phases. The mycelium grew fast and showed rapid growth rate (8.91 mmd−1) in WS + OSP (97 + 3) combination while WS + OSP (92 + 8) showed maximum laccase (3.133 U/g) and Mn peroxidase (MnP) activities (0.091 U/g). The climax level of laccase (5.433 U/g) and MnP (0.097 U/g) was recorded during fruit body initiation in WS + OSP (97 + 3) and WS + OSP (98 + 2) combinations, respectively. The WS + OSP (97 + 3) combination represented the best condition for mushroom cultivation and produced the highest biological efficiency (147%). In addition, protein and lipid contents in fruit bodies were slightly improved in response to OSP. The carbohydrate was significantly increased by raising concentration of OSP. The highest values of protein, carbohydrate, and lipid noted were 31.3 μg/g, 0.0639 (g/g), and 0.373 (g/g) correspondingly. Conclusively it was evident that lower concentrations of OSP acted positively and relatively to higher concentrations and improved nutritional content which may suitably be used to enhance both yield and nutritional values of mushroom. Ram Naraian, Om Prakash Narayan, and Jatin Srivastava Copyright © 2014 Ram Naraian et al. All rights reserved. In Vitro Antimicrobial and Antiprotozoal Activities, Phytochemical Screening and Heavy Metals Toxicity of Different Parts of Ballota nigra Thu, 26 Jun 2014 11:07:19 +0000 The study was done to assess the phytochemicals (flavonoids, terpenoids, saponins, tannin, alkaloids, and phenol) in different parts (root, stem, and leaves) of Ballota nigra and correlated it to inhibition of microbes (bacteria and fungi), protozoan (Leishmania), and heavy metals toxicity evaluation. In root and stem flavonoids, terpenes and phenols were present in ethanol, chloroform, and ethyl acetate soluble fraction; these were found to be the most active inhibiting fractions against all the tested strains of bacteria, fungi, and leishmania. While in leaves flavonoids, terpenes, and phenols were present in ethanol, chloroform, and n-butanol fractions which were the most active fractions against both types of microbes and protozoan (leishmania) in in vitro study. Ethanol and chloroform fractions show maximum inhibition against Escherichia coli (17 mm). The phytochemical and biological screenings were correlated with the presence of heavy metals in selected plant Ballota nigra. Cr was found above permissible value (above 1.5 mg/kg) in all parts of the plant. Ni was above WHO limit in B. nigra root and leaves (3.35 ± 1.20 mg/kg and 5.09 ± 0.47 mg/kg, respectively). Fe was above permissible value in all parts of B. nigra (above 20 mg/kg). Cd was above permissible value in all parts of the plant (above 0.3 mg/kg). Pb was above WHO limit (above 2 mg/kg) in all parts of Ballota nigra. Najeeb Ullah, Ijaz Ahmad, and Sultan Ayaz Copyright © 2014 Najeeb Ullah et al. All rights reserved. High-Level Antimicrobial Efficacy of Representative Mediterranean Natural Plant Extracts against Oral Microorganisms Thu, 26 Jun 2014 06:59:34 +0000 Nature is an unexplored reservoir of novel phytopharmaceuticals. Since biofilm-related oral diseases often correlate with antibiotic resistance, plant-derived antimicrobial agents could enhance existing treatment options. Therefore, the rationale of the present report was to examine the antimicrobial impact of Mediterranean natural extracts on oral microorganisms. Five different extracts from Olea europaea, mastic gum, and Inula viscosa were tested against ten bacteria and one Candida albicans strain. The extraction protocols were conducted according to established experimental procedures. Two antimicrobial assays—the minimum inhibitory concentration (MIC) assay and the minimum bactericidal concentration (MBC) assay—were applied. The screened extracts were found to be active against each of the tested microorganisms. O. europaea presented MIC and MBC ranges of 0.07–10.00 mg mL−1 and 0.60–10.00 mg mL−1, respectively. The mean MBC values for mastic gum and I. viscosa were 0.07–10.00 mg mL−1 and 0.15–10.00 mg mL−1, respectively. Extracts were less effective against C. albicans and exerted bactericidal effects at a concentration range of 0.07–5.00 mg mL−1 on strict anaerobic bacteria (Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, and Parvimonas micra). Ethyl acetate I. viscosa extract and total mastic extract showed considerable antimicrobial activity against oral microorganisms and could therefore be considered as alternative natural anti-infectious agents. Lamprini Karygianni, Manuel Cecere, Alexios Leandros Skaltsounis, Aikaterini Argyropoulou, Elmar Hellwig, Nektarios Aligiannis, Annette Wittmer, and Ali Al-Ahmad Copyright © 2014 Lamprini Karygianni et al. All rights reserved. Preliminary Evaluation of Probiotic Properties of Lactobacillus Strains Isolated from Sardinian Dairy Products Thu, 26 Jun 2014 05:28:19 +0000 Twenty-three Lactobacillus strains of dairy origin were evaluated for some functional properties relevant to their use as probiotics. A preliminary subtractive screening based on the abilities to inhibit the growth of microbial pathogens and hydrolyze conjugated bile salts was applied, and six strains were selected for further characterization including survival under gastrointestinal environmental conditions, adhesion to gut epithelial tissue, enzymatic activity, and some safety properties. All selected strains maintained elevated cell numbers under conditions simulating passage through the human gastrointestinal tract, well comparable to the values obtained for the probiotic strain Lactobacillus rhamnosus GG, and were able to adhere to Caco-2 cells to various extents (from 3 to 20%). All strains exhibited high aminopeptidase, and absent or very low proteolytic and strong β-galactosidase activities; none was found to be haemolytic or to produce biogenic amines and all were susceptible to tetracycline, chloramphenicol, erythromycin, ampicillin, and amoxicillin/clavulanic acid. Our results indicate that the Lactobacillus strains analyzed could be considered appropriate probiotic candidates, due to resistance to GIT simulated conditions, antimicrobial activity, adhesion to Caco-2 cell-line, and absence of undesirable properties. They could be used as adjunct cultures for contributing to the quality and health related functional properties of dairy products. Maria Barbara Pisano, Silvia Viale, Stefania Conti, Maria Elisabetta Fadda, Maura Deplano, Maria Paola Melis, Monica Deiana, and Sofia Cosentino Copyright © 2014 Maria Barbara Pisano et al. All rights reserved. Use of Antimicrobial Films and Edible Coatings Incorporating Chemical and Biological Preservatives to Control Growth of Listeria monocytogenes on Cold Smoked Salmon Wed, 25 Jun 2014 08:10:23 +0000 The relatively high incidence of Listeria monocytogenes in cold smoked salmon (CSS) is of concern as it is a refrigerated processed food of extended durability (REPFED). The objectives of this study were to compare and optimize the antimicrobial effectiveness of films and coatings incorporating nisin (Nis) and sodium lactate (SL), sodium diacetate (SD), potassium sorbate (PS), and/or sodium benzoate (SB) in binary or ternary combinations on CSS. Surface treatments incorporating Nis (25000 IU/mL) in combination with PS (0.3%) and SB (0.1%) had the highest inhibitory activity, reducing the population of L. monocytogenes by a maximum of 3.3 log CFU/cm2 (films) and 2.9 log CFU/cm2 (coatings) relative to control samples after 10 days of storage at 21°C. During refrigerated storage, coatings were more effective in inhibiting growth of L. monocytogenes than their film counterparts. Cellulose-based coatings incorporating Nis, PS, and SB reduced the population of L. monocytogenes, and anaerobic and aerobic spoilage flora by a maximum of 4.2, 4.8, and 4.9 log CFU/cm2, respectively, after 4 weeks of refrigerated storage. This study highlights the effectiveness of cellulose-based edible coatings incorporating generally regarded as safe (GRAS) natural and chemical antimicrobials to inhibit the development of L. monocytogenes and spoilage microflora thus enhancing the safety and quality of CSS. Hudaa Neetoo and Fawzi Mahomoodally Copyright © 2014 Hudaa Neetoo and Fawzi Mahomoodally. All rights reserved. Sinefungin, a Natural Nucleoside Analogue of S-Adenosylmethionine, Inhibits Streptococcus pneumoniae Biofilm Growth Mon, 23 Jun 2014 10:56:57 +0000 Pneumococcal colonization and disease is often associated with biofilm formation, in which the bacteria exhibit elevated resistance both to antibiotics and to host defense systems, often resulting in infections that are persistent and difficult to treat. We evaluated the effect of sinefungin, a nucleoside analogue of S-adenosylmethionine, on pneumococcal in vitro biofilm formation and in vivo colonization. Sinefungin is bacteriostatic to pneumococci and significantly decreased biofilm growth and inhibited proliferation and structure of actively growing biofilms but did not alter growth or the matrix structure of established biofilms. Sinefungin significantly reduced pneumococcal colonization in rat middle ear. The quorum sensing molecule (autoinducer-2) production was significantly reduced by 92% in sinefungin treated samples. The luxS, pfs, and speE genes were downregulated in biofilms grown in the presence of sinefungin. This study shows that sinefungin inhibits pneumococcal biofilm growth in vitro and colonization in vivo, decreases AI-2 production, and downregulates luxS, pfs, and speE gene expressions. Therefore, the S-adenosylmethionine (SAM) inhibitors could be used as lead compounds for the development of novel antibiofilm agents against pneumococci. Mukesh Kumar Yadav, Seok-Won Park, Sung-Won Chae, and Jae-Jun Song Copyright © 2014 Mukesh Kumar Yadav et al. All rights reserved. Potential Probiotic Escherichia coli 16 Harboring the Vitreoscilla Hemoglobin Gene Improves Gastrointestinal Tract Colonization and Ameliorates Carbon Tetrachloride Induced Hepatotoxicity in Rats Thu, 19 Jun 2014 07:05:36 +0000 The present study describes the beneficial effects of potential probiotic E. coli 16 (pUC8:16gfp) expressing Vitreoscilla hemoglobin (vgb) gene, associated with bacterial respiration under microaerobic condition, on gastrointestinal (GI) colonization and its antioxidant activity on carbon tetrachloride (CCl4) induced toxicity in Charles Foster rats. In vitro, catalase activity in E. coli 16 (pUC8:16gfp) was 1.8 times higher compared to E. coli 16 (pUC-gfp) control. In vivo, E. coli 16 (pUC8:16gfp) not only was recovered in the fecal matter after 70 days of oral administration but also retained antibacterial activities, whereas E. coli 16 (pUC-gfp) was not detected. Oral administration of 200 and 500 μL/kg body weight of CCl4 to rats at weekly interval resulted in elevated serum glutamyl pyruvate transaminase (SGPT) and serum glutamyl oxalacetate transaminase (SGOT) levels compared to controls. Rats prefed with E. coli 16 (pUC8:16gfp) demonstrated near to normal levels for SGPT and SGOT, whereas the liver homogenate catalase activity was significantly increased compared to CCl4 treated rats. Thus, pUC8:16gfp plasmid encoding vgb improved the growth and GI tract colonization of E. coli 16. In addition, it also enhanced catalase activity in rats harboring E. coli 16 (pUC8:16gfp), thereby preventing the absorption of CCl4 to GI tract. Prasant Kumar, Ayush V. Ranawade, and Naresh G. Kumar Copyright © 2014 Prasant Kumar et al. All rights reserved. Carbapenemases in Gram-Negative Bacteria: Laboratory Detection and Clinical Significance Sun, 15 Jun 2014 11:05:30 +0000 Branka Bedenić, Vanda Plečko, Sanda Sardelić, Selma Uzunović, and Karmen Godič Torkar Copyright © 2014 Branka Bedenić et al. All rights reserved. Inhibitory and Toxic Effects of Volatiles Emitted by Strains of Pseudomonas and Serratia on Growth and Survival of Selected Microorganisms, Caenorhabditis elegans, and Drosophila melanogaster Wed, 11 Jun 2014 12:02:08 +0000 In previous research, volatile organic compounds (VOCs) emitted by various bacteria into the chemosphere were suggested to play a significant role in the antagonistic interactions between microorganisms occupying the same ecological niche and between bacteria and target eukaryotes. Moreover, a number of volatiles released by bacteria were reported to suppress quorum-sensing cell-to-cell communication in bacteria, and to stimulate plant growth. Here, volatiles produced by Pseudomonas and Serratia strains isolated mainly from the soil or rhizosphere exhibited bacteriostatic action on phytopathogenic Agrobacterium tumefaciens and fungi and demonstrated a killing effect on cyanobacteria, flies (Drosophila melanogaster), and nematodes (Caenorhabditis elegans). VOCs emitted by the rhizospheric Pseudomonas chlororaphis strain 449 and by Serratia proteamaculans strain 94 isolated from spoiled meat were identified using gas chromatography-mass spectrometry analysis, and the effects of the main headspace compounds—ketones (2-nonanone, 2-heptanone, 2-undecanone) and dimethyl disulfide—were inhibitory toward the tested microorganisms, nematodes, and flies. The data confirmed the role of bacterial volatiles as important compounds involved in interactions between organisms under natural ecological conditions. Alexandra A. Popova, Olga A. Koksharova, Valentina A. Lipasova, Julia V. Zaitseva, Olga A. Katkova-Zhukotskaya, Svetlana Iu. Eremina, Alexander S. Mironov, Leonid S. Chernin, and Inessa A. Khmel Copyright © 2014 Alexandra A. Popova et al. All rights reserved. Association between Giardia duodenalis and Coinfection with Other Diarrhea-Causing Pathogens in India Mon, 09 Jun 2014 15:53:56 +0000 Giardia duodenalis, is often seen as an opportunistic pathogen and one of the major food and waterborne parasites. Some insights of Giardia infestation in a diarrhoea-prone population were investigated in the present study. Our primary goal was to understand the interaction of this parasite with other pathogens during infection and to determine some important factors regulating the diarrhoeal disease spectrum of a population. Giardia showed a steady rate of occurrence throughout the entire study period with a nonsignificant association with rainfall . Interestingly coinfecting pathogens like Vibrio cholerae and rotavirus played a significant role in the occurrence of this parasite. Moreover, the age distribution of the diarrhoeal cases was very much dependent on the coinfection rate of Giardia infection. As per our findings, Giardia infection rate seems to play a vital role in regulation of the whole diarrhoeal disease spectrum in this endemic region. Avik K. Mukherjee, Punam Chowdhury, Krishnan Rajendran, Tomoyoshi Nozaki, and Sandipan Ganguly Copyright © 2014 Avik K. Mukherjee et al. All rights reserved. Statistical Optimization of Fibrinolytic Enzyme Production Using Agroresidues by Bacillus cereus IND1 and Its Thrombolytic Activity In Vitro Mon, 09 Jun 2014 12:50:30 +0000 A potent fibrinolytic enzyme-producing Bacillus cereus IND1 was isolated from the Indian food, rice. Solid-state fermentation was carried out using agroresidues for the production of fibrinolytic enzyme. Among the substrates, wheat bran supported more enzyme production and has been used for the optimized enzyme production by statistical approach. Two-level full-factorial design demonstrated that moisture, supplementation of beef extract, and sodium dihydrogen phosphate have significantly influenced enzyme production (). A central composite design resulted in the production of 3699 U/mL of enzyme in the presence of 0.3% (w/w) beef extract and 0.05% (w/w) sodium dihydrogen phosphate, at 100% (v/w) moisture after 72 h of fermentation. The enzyme production increased fourfold compared to the original medium. This enzyme was purified to homogeneity by ammonium sulfate precipitation, diethylaminoethyl-cellulose ion-exchange chromatography, Sephadex G-75 gel filtration chromatography, and casein-agarose affinity chromatography and had an apparent molecular mass of 29.5 kDa. The optimum pH and temperature for the activity of fibrinolytic enzyme were found to be 8.0 and 60°C, respectively. This enzyme was highly stable at wide pH range (7.0–9.0) and showed 27% ± 6% enzyme activity after initial denaturation at 60°C for 1 h. In vitro assays revealed that the enzyme could activate plasminogen and significantly degraded the fibrin net of blood clot, which suggests its potential as an effective thrombolytic agent. Ponnuswamy Vijayaraghavan and Samuel Gnana Prakash Vincent Copyright © 2014 Ponnuswamy Vijayaraghavan and Samuel Gnana Prakash Vincent. All rights reserved. Effect of Experimentally Induced Hepatic and Renal Failure on the Pharmacokinetics of Topiramate in Rats Mon, 09 Jun 2014 11:46:12 +0000 We aimed to investigate the effect of induced hepatic and renal failure on the pharmacokinetics of topiramate (TPM) in rats. Twenty-four Sprague-Dawley rats were used in this study. Renal or hepatic failure was induced by a single i.p. dose of 7.5 mg/kg cisplatin () or 0.5 mL/kg carbon tetrachloride (CCl4) (), respectively. Three days after cisplatin dose or 24 h after CCl4 dose, the rats were administered a single oral dose of 20 mg/kg TPM. The plasma samples were quantified by LC-MS/MS method. Compared to control, plasma concentration-time profile in CCl4-treated and, to a lesser extent, in cisplatin-treated rats decreased more slowly particularly in the elimination phase. TPM oral clearance (CL/F) in CCl4-treated group was significantly lower than that in control (), whereas , T1/2, and Vd/F were significantly higher in CCl4-treated rats compared to the control (). The CL/F was not significantly different between cisplatin-treated rats and control (). However, in cisplatin-treated rats, the T1/2 and Vd/F were significantly higher than that in the control group (). Both conditions failed to cause a significant effect on or . The present findings suggest that induced hepatic or renal failure could modify the pharmacokinetic profile of TPM in the rat. Kamal M. Matar and Yasin I. Tayem Copyright © 2014 Kamal M. Matar and Yasin I. Tayem. All rights reserved. Phenotypic and Molecular Characterization of Plasmid Mediated AmpC β-Lactamases among Escherichia coli, Klebsiella spp., and Proteus mirabilis Isolated from Urinary Tract Infections in Egyptian Hospitals Mon, 09 Jun 2014 11:44:52 +0000 The incidence of resistance by Enterobacteriaceae to β-lactam/β-lactamase inhibitors combination is increasing in Egypt. Three phenotypic techniques, comprising AmpC disk diffusion and inhibition dependent methods using phenylboronic acid (PBA) and cloxacillin, were compared to PCR based method for detection of plasmid mediated AmpC β-lactamase in common urinary tract isolates. A total of 143 isolates, including E. coli, Klebsiella pneumonia, and Proteus mirabilis, were collected from urinary tract infections cases in Egyptian hospitals. Plasmid encoded AmpC genes were detected by PCR in 88.46% of cefoxitin resistant isolates. The most prevalent AmpC gene family was CIT including CMY-2, CMY-4, and two CMY-2 variants. The second prevalent gene was DHA-1 which was detected in E. coli and Klebsiella pneumonia. The genes EBC, FOX, and MOX were also detected but in small percentage. Some isolates were identified as having more than one pAmpC gene. The overall sensitivity and specificity of phenotypic tests for detection of AmpC β-lactamase showed that AmpC disk diffusion and inhibition dependent method by cloxacillin were the most sensitive and the most specific disk tests. PCR remains the gold standard for detection of AmpC β-lactamases. This study represents the first report of CMY-2 variants of CMY-42 and CMY-102 β-lactamase-producing E. coli, Klebsiella pneumonia, and Proteus mirabilis isolates in Egypt. Mai M. Helmy and Reham Wasfi Copyright © 2014 Mai M. Helmy and Reham Wasfi. All rights reserved. Molecular Analysis of VanA Outbreak of Enterococcus faecium in Two Warsaw Hospitals: The Importance of Mobile Genetic Elements Mon, 09 Jun 2014 11:42:38 +0000 Vancomycin-resistant Enterococcus faecium represents a growing threat in hospital-acquired infections. Two outbreaks of this pathogen from neighboring Warsaw hospitals have been analyzed in this study. Pulsed-field gel electrophoresis (PFGE) of SmaI-digested DNA, multilocus VNTR analysis (MLVA), and multilocus sequence typing (MLST) revealed a clonal variability of isolates which belonged to three main lineages (17, 18, and 78) of nosocomial E. faecium. All isolates were multidrug resistant and carried several resistance, virulence, and plasmid-specific genes. Almost all isolates shared the same variant of Tn1546 transposon, characterized by the presence of insertion sequence ISEf1 and a point mutation in the vanA gene. In the majority of cases, this transposon was located on 50 kb or 100 kb pRUM-related plasmids, which lacked, however, the axe-txe toxin-antitoxin genes. 100 kb plasmid was easily transferred by conjugation and was found in various clonal backgrounds in both institutions, while 50 kb plasmid was not transferable and occurred solely in MT159/ST78 strains that disseminated clonally in one institution. Although molecular data indicated the spread of VRE between two institutions or a potential common source of this alert pathogen, epidemiological investigations did not reveal the possible route by which outbreak strains disseminated. Ewa Wardal, Katarzyna Markowska, Dorota Żabicka, Marta Wróblewska, Małgorzata Giemza, Ewa Mik, Hanna Połowniak-Pracka, Agnieszka Woźniak, Waleria Hryniewicz, and Ewa Sadowy Copyright © 2014 Ewa Wardal et al. All rights reserved. In Vitro Antifungal Evaluation of Seven Different Disinfectants on Acrylic Resins Thu, 05 Jun 2014 10:53:19 +0000 Objective. The aim of this study was to evaluate alternative methods for the disinfection of denture-based materials. Material and Methods. Two different denture-based materials were included in the study. Before microbial test, the surface roughness of the acrylic resins was evaluated. Then, the specimens were divided into 8 experimental groups , according to microorganism considered and disinfection methods used. The specimens were contaminated in vitro by standardized suspensions of Candida albicans ATCC#90028 and Candida albicans oral isolate. The following test agents were tested: sodium hypochlorite (NaOCl 1%), microwave (MW) energy, ultraviolet (UV) light, mouthwash containing propolis (MCP), Corega Tabs, 50% and 100% white vinegar. After the disinfection procedure, the number of remaining microbial cells was evaluated in CFU/mL. Kruskal-Wallis, ANOVA, and Dunn’s test were used for multiple comparisons. Mann Whitney U test was used to compare the surface roughness. Results. Statistically significant difference was found between autopolymerised and heat-cured acrylic resins. The autopolymerised acrylic resin surfaces were rougher than surfaces of heat-cured acrylic resin. The most effective disinfection method was 100% white vinegar for tested microorganisms and both acrylic resins. Conclusion. This study showed that white vinegar 100% was the most effective method for tested microorganisms. This agent is cost-effective and easy to access and thus may be appropriate for household use. A. Z. Yildirim-Bicer, I. Peker, G. Akca, and I. Celik Copyright © 2014 A. Z. Yildirim-Bicer et al. All rights reserved. Experimental Protection of Diabetic Mice against Lethal P. aeruginosa Infection by Bacteriophage Thu, 05 Jun 2014 08:26:44 +0000 The emergence of antibiotic-resistant bacterial strains has become a global crisis and is vulnerable for the exploration of alternative antibacterial therapies. The present study emphasizes the use of bacteriophage for the treatment of multidrug resistant P. aeruginosa. P. aeruginosa was used to induce septicemia in streptozotocin (STZ) induced diabetic and nondiabetic mice by intraperitoneal (i.p.) injection of 3 × 108 CFU, resulting in a fatal bacteremia within 48 hrs. A single i.p. injection of 3 × 109 PFU phage GNCP showed efficient protection in both diabetic (90%) and nondiabetic (100%) bacteremic mice. It was further noted that the protection rate was reduced in diabetic mice when phage GNCP was administered after 4 h and 6 h of lethal bacterial challenge. In contrast, nondiabetic bacteremic mice were rescued even when treatment was delayed up to 20 h after lethal bacterial challenge. Evaluation of results confirmed that a single intraperitoneal injection of the phage dose (3 × 109 PFU/mL) was more effective than the multiple doses of imipenem. These results uphold the efficacy of phage therapy against pernicious P. aeruginosa infections, especially in cases of immunocompromised host. Nagaveni Shivshetty, Rajeshwari Hosamani, Liyakat Ahmed, Ajay Kumar Oli, Syed Sannauallah, Shivshetty Sharanbassappa, S. A. Patil, and Chandrakanth R. Kelmani Copyright © 2014 Nagaveni Shivshetty et al. All rights reserved. Association of Levels of Antibodies from Patients with Inflammatory Bowel Disease with Extracellular Proteins of Food and Probiotic Bacteria Wed, 04 Jun 2014 11:10:55 +0000 Inflammatory bowel disease (IBD) is an autoimmune disease characterized by a chronic inflammation of the gastrointestinal tract mucosa and is related to an abnormal immune response to commensal bacteria. Our aim of the present work has been to explore the levels of antibodies (IgG and IgA) raised against extracellular proteins produced by LAB and its association with IBD. We analyzed, by Western-blot and ELISA, the presence of serum antibodies (IgA and IgG) developed against extracellular protein fractions produced by different food bacteria from the genera Bifidobacterium and Lactobacillus. We used a sera collection consisting of healthy individuals (HC, ), Crohn's disease patients (CD, ), and ulcerative colitis patients (UC, ). Levels of IgA antibodies developed against a cell-wall hydrolase from Lactobacillus casei subsp. rhamnosus GG (CWH) were significantly higher in the IBD group (; ). The specificity of our measurements was confirmed by measuring IgA antibodies developed against the CWH peptide 365-VNTSNQTAAVSAS-377. IBD patients appeared to have different immune response to food bacteria. This paper sets the basis for developing systems for early detection of IBD, based on the association of high levels of antibodies developed against extracellular proteins from food and probiotic bacteria. Arancha Hevia, Patricia López, Ana Suárez, Claudine Jacquot, María C. Urdaci, Abelardo Margolles, and Borja Sánchez Copyright © 2014 Arancha Hevia et al. All rights reserved. Safe-Site Effects on Rhizosphere Bacterial Communities in a High-Altitude Alpine Environment Wed, 04 Jun 2014 07:37:33 +0000 The rhizosphere effect on bacterial communities associated with three floristic communities (RW, FI, and M sites) which differed for the developmental stages was studied in a high-altitude alpine ecosystem. RW site was an early developmental stage, FI was an intermediate stage, M was a later more matured stage. The N and C contents in the soils confirmed a different developmental stage with a kind of gradient from the unvegetated bare soil (BS) site through RW, FI up to M site. The floristic communities were composed of 21 pioneer plants belonging to 14 species. Automated ribosomal intergenic spacer analysis showed different bacterial genetic structures per each floristic consortium which differed also from the BS site. When plants of the same species occurred within the same site, almost all their bacterial communities clustered together exhibiting a plant species effect. Unifrac significance value () on 16S rRNA gene diversity revealed significant differences () between BS site and the vegetated sites with a weak similarity to the RW site. The intermediate plant colonization stage FI did not differ significantly from the RW and the M vegetated sites. These results pointed out the effect of different floristic communities rhizospheres on their soil bacterial communities. Sonia Ciccazzo, Alfonso Esposito, Eleonora Rolli, Stefan Zerbe, Daniele Daffonchio, and Lorenzo Brusetti Copyright © 2014 Sonia Ciccazzo et al. All rights reserved. MiR-15b Targets Cyclin D1 to Regulate Proliferation and Apoptosis in Glioma Cells Wed, 04 Jun 2014 00:00:00 +0000 Aim. To investigate the role and mechanism of miR-15b in the proliferation and apoptosis of glioma. Methods. The miR-15b mimics were transfected into human glioma cells to upregulate the miR-15b expression. Cyclin D1 was determined by both western blotting analysis and luciferase reporter assay. Methylthiazol tetrazolium (MTT) and flow cytometry were employed to detect the cell proliferation, cell cycle, and apoptosis. Results. Overexpression of miR-15b inhibits proliferation by arrested cell cycle progression and induces apoptosis, possibly by directly targeting Cyclin D1. Both luciferase assay and bioinformatics search revealed a putative target site of miR-15b binding to the 3′-UTR of Cyclin D1. Moreover, expression of miR-15b in glioma tissues was found to be inversely correlated with Cyclin D1 expression. Enforced Cyclin D1 could abrogate the miR-15b-mediated cell cycle arrest and apoptosis. Conclusions. Our findings identified that miR-15b may function as a glioma suppressor by targeting the Cyclin D1, which may provide a novel therapeutic strategy for treatment of glioma. Guan Sun, Lei Shi, Shushan Yan, Zhengqiang Wan, Nan Jiang, Linshan Fu, Min Li, and Jun Guo Copyright © 2014 Guan Sun et al. All rights reserved. Complex Links between Natural Tuberculosis and Porcine Circovirus Type 2 Infection in Wild Boar Tue, 03 Jun 2014 09:26:02 +0000 Individuals in natural populations are exposed to a diversity of pathogens which results in coinfections. The aim of this study was to investigate the relation between natural infection with tuberculosis (TB) due to infection by bacteria of the Mycobacterium tuberculosis complex and porcine circovirus type 2 (PCV2) in free-ranging Eurasian wild boar (Sus scrofa). Apparent prevalence for TB lesions and PCV2 infection was extremely high in all age classes, including piglets (51% for TB; 85.7% for PCV2). Modeling results revealed that the relative risk of young (less than 2 years old) wild boar to test positive to PCV2 PCR was negatively associated with TB lesion presence. Also, an interaction between TB, PCV2, and body condition was evidenced: in wild boar with TB lesions probability of being PCV2 PCR positive increased with body condition, whereas this relation was negative for wild boar without TB lesions. This study provides insight into the coinfections occurring in free-ranging host populations that are naturally exposed to several pathogens at an early age. Using TB and PCV2 as a case study, we showed that coinfection is a frequent event among natural populations that takes place early in life with complex effects on the infections and the hosts. Iratxe Díez-Delgado, Mariana Boadella, MariPaz Martín-Hernando, José Angel Barasona, Beatriz Beltrán-Beck, David González-Barrio, Marina Sibila, Joaquín Vicente, Joseba M. Garrido, Joaquim Segalés, and Christian Gortazar Copyright © 2014 Iratxe Díez-Delgado et al. All rights reserved. Pretreatment of Cottage Cheese to Enhance Biogas Production Tue, 03 Jun 2014 08:08:12 +0000 This study evaluated the possibility of pretreating selected solid fraction of an anaerobic digester treating food waste to lower the hydraulic retention time and increase the methane production. The study investigated the effect of different pretreatments (thermal, chemical, thermochemical and enzymatic) for enhanced methane production from cottage cheese. The most effective pretreatments were thermal and enzymatic. Highest solubilisation of COD was observed in thermal pretreatment, followed by thermochemical. In single enzyme systems, lipase at low concentration gave significantly higher methane yield than for the experiments without enzyme additions. The highest lipase dosages decreased methane yield from cottage cheese. However, in case of protease enzyme an increase in concentration of the enzyme showed higher methane yield. In the case of mixed enzyme systems, pretreatment at 1 : 2 ratio of lipase : protease showed higher methane production in comparison with 1 : 1 and 2 : 1 ratios. Methane production potentials for different pretreatments were as follows: thermal 357 mL/g VS, chemical 293 mL/g VS, and thermochemical 441 mL/g VS. The average methane yield from single enzyme systems was 335 mL/g VS for lipase and 328 mL/g VS for protease. Methane potentials for mixed enzyme ratios were 330, 360, and 339 mL/g VS for 1 : 1, 1 : 2, and 2 : 1 lipase : protease, respectively. Vidhya Prabhudessai, Bhakti Salgaonkar, Judith Braganca, and Srikanth Mutnuri Copyright © 2014 Vidhya Prabhudessai et al. All rights reserved. Comparative Examination of the Olive Mill Wastewater Biodegradation Process by Various Wood-Rot Macrofungi Mon, 02 Jun 2014 09:17:41 +0000 Olive mill wastewater (OMW) constitutes a major cause of environmental pollution in olive-oil producing regions. Sixty wood-rot macrofungi assigned in 43 species were evaluated for their efficacy to colonize solidified OMW media at initially established optimal growth temperatures. Subsequently eight strains of the following species were qualified: Abortiporus biennis, Ganoderma carnosum, Hapalopilus croceus, Hericium erinaceus, Irpex lacteus, Phanerochaete chrysosporium, Pleurotus djamor, and P. pulmonarius. Fungal growth in OMW (25%v/v in water) resulted in marked reduction of total phenolic content, which was significantly correlated with the effluent’s decolorization. A. biennis was the best performing strain (it decreased phenolics by 92% and color by 64%) followed by P. djamor and I. lacteus. Increase of plant seeds germination was less pronounced evidencing that phenolics are only partly responsible for OMW’s phytotoxicity. Laccase production was highly correlated with all three biodegradation parameters for H. croceus, Ph. chrysosporium, and Pleurotus spp., and so were manganese-independent and manganese dependent peroxidases for A. biennis and I. lacteus. Monitoring of enzymes with respect to biomass production indicated that Pleurotus spp., H. croceus, and Ph. chrysosporium shared common patterns for all three activities. Moreover, generation of enzymes at the early biodegradation stages enhanced the efficiency of OMW treatment. Georgios Koutrotsios and Georgios I. Zervakis Copyright © 2014 Georgios Koutrotsios and Georgios I. Zervakis. All rights reserved. Biofilm Formation by Mycobacterium bovis: Influence of Surface Kind and Temperatures of Sanitizer Treatments on Biofilm Control Sun, 01 Jun 2014 12:53:34 +0000 Mycobacterium bovis causes classic bovine tuberculosis, a zoonosis which is still a concern in Africa. Biofilm forming ability of two Mycobacterium bovis strains was assessed on coupons of cement, ceramic, or stainless steel in three different microbiological media at 37°C with agitation for 2, 3, or 4 weeks to determine the medium that promotes biofilm. Biofilm mass accumulated on coupons was treated with 2 sanitizers (sanitizer A (5.5 mg L−1 active iodine) and sanitizer B (170.6 g1 alkyl dimethylbenzyl ammonium chloride, 78 g−1 didecyldimethyl ammonium chloride, 107.25 g L−1 glutaraldehyde, 146.25 g L−1 isopropanol, and 20 g L−1 pine oil) at 28 and 45°C and in hot water at 85°C for 5 min. Residual biofilms on treated coupons were quantified using crystal violet binding assay. The two strains had a similar ability to form biofilms on the three surfaces. More biofilms were developed in media containing 5% liver extract. Biofilm mass increased as incubation time increased till the 3rd week. More biofilms were formed on cement than on ceramic and stainless steel surfaces. Treatment with hot water at 85°C reduced biofilm mass, however, sanitizing treatments at 45°C removed more biofilms than at 28°C. However, neither treatment completely eliminated the biofilms. The choice of processing surface and temperatures used for sanitizing treatments had an impact on biofilm formation and its removal from solid surfaces. Victoria O. Adetunji, Aderemi O. Kehinde, Olayemi K. Bolatito, and Jinru Chen Copyright © 2014 Victoria O. Adetunji et al. All rights reserved. Identification, Typing, Antifungal Resistance Profile, and Biofilm Formation of Candida albicans Isolates from Lebanese Hospital Patients Sun, 01 Jun 2014 08:37:36 +0000 As leading opportunistic fungal pathogens identification and subtyping of Candida species are crucial in recognizing outbreaks of infection, recognizing particularly virulent strains, and detecting the emergence of drug resistant strains. In this study our objective was to compare identification of Candida albicans by the hospitals through the use of conventional versus identification based on the ITS (Internal Transcribed Spacer) and to assess biofilm forming capabilities, drug resistance patterns and correlate these with MLST typing. ITS typing revealed a 21.2% hospital misidentification rate. Multidrug resistance to three drugs out of four tested was detected within 25% of the isolates raising concerns about the followed treatment regimens. Drug resistant strains as well as biofilm formers were phylogenetically related, with some isolates with significant biofilm forming capabilities being correlated to those that were multidrug resistant. Such isolates were grouped closely together in a neighbor-joining tree generated by MLST typing indicating phylogenetic relatedness, microevolution, or recurrent infection. In conclusion, this pilot study gives much needed insight concerning C. albicans isolates circulating in Lebanese hospitals and is the first study of its kind correlating biofilm formation, antifungal resistance, and evolutionary relatedness. Ibrahim Bitar, Roy A. Khalaf, Houda Harastani, and Sima Tokajian Copyright © 2014 Ibrahim Bitar et al. All rights reserved. Mass Spectrometry and Multiplex Antigen Assays to Assess Microbial Quality and Toxin Production of Staphylococcus aureus Strains Isolated from Clinical and Food Samples Thu, 29 May 2014 11:51:57 +0000 The aim of our study was to investigate the microbial quality of meat products and on some clinical samples in Abidjan focused on Staphylococcus genus and the toxin production profile of Staphylococcus aureus (S. aureus) isolated. Bacteria were collected from 240 samples of three meat products sold in Abidjan and 180 samples issued from clinical infections. The strains were identified by both microbiological and MALDI-TOF-MS methods. The susceptibility to antibiotics was determined by the disc diffusion method. The production of Panton-Valentine Leukocidin, LukE/D, and epidermolysins was screened using radial gel immunodiffusion. The production of staphylococcal enterotoxins and TSST-1 was screened by a Bio-Plex Assay. We observed that 96/240 of meat samples and 32/180 of clinical samples were contaminated by Staphylococcus. Eleven species were isolated from meats and 4 from clinical samples. Forty-two S. aureus strains were isolated from ours samples. Variability of resistance was observed for most of the tested antibiotics but none of the strains displays a resistance to imipenem and quinolones. We observed that 89% of clinical S. aureus were resistant to methicillin against 58% for those issued from meat products. All S. aureus isolates issued from meat products produce epidermolysins whereas none of the clinical strains produced these toxins. The enterotoxins were variably produced by both clinical and meat product samples. Paul Attien, Haziz Sina, Wardi Moussaoui, Gaëlle Zimmermann-Meisse, Thomas Dadié, Daniel Keller, Philippe Riegel, Vincent Edoh, Simeon O. Kotchoni, Marcellin Djè, Gilles Prévost, and Lamine Baba-Moussa Copyright © 2014 Paul Attien et al. All rights reserved. Development of a Potential Probiotic Fresh Cheese Using Two Lactobacillus salivarius Strains Isolated from Human Milk Thu, 29 May 2014 09:41:20 +0000 Cheeses have been proposed as a good alternative to other fermented milk products for the delivery of probiotic bacteria to the consumer. The objective of this study was to assess the survival of two Lactobacillus salivarius strains (CECT5713 and PS2) isolated from human milk during production and storage of fresh cheese for 28 days at 4°C. The effect of such strains on the volatile compounds profile, texture, and other sensorial properties, including an overall consumer acceptance, was also investigated. Both L. salivarius strains remained viable in the cheeses throughout the storage period and a significant reduction in their viable counts was only observed after 21 days. Globally, the addition of the L. salivarius strains did not change significantly neither the chemical composition of the cheese nor texture parameters after the storage period, although cheeses manufactured with L. salivarius CECT5713 presented significantly higher values of hardness. A total of 59 volatile compounds were identified in the headspace of experimental cheeses, and some L. salivarius-associated differences could be identified. All cheeses presented good results of acceptance after the sensory evaluation. Consequently, our results indicated that fresh cheese can be a good vehicle for the two L. salivarius strains analyzed in this study. Nivia Cárdenas, Javier Calzada, Ángela Peirotén, Esther Jiménez, Rosa Escudero, Juan M. Rodríguez, Margarita Medina, and Leónides Fernández Copyright © 2014 Nivia Cárdenas et al. All rights reserved. Immune Modulating Capability of Two Exopolysaccharide-Producing Bifidobacterium Strains in a Wistar Rat Model Thu, 29 May 2014 06:30:22 +0000 Fermented dairy products are the usual carriers for the delivery of probiotics to humans, Bifidobacterium and Lactobacillus being the most frequently used bacteria. In this work, the strains Bifidobacterium animalis subsp. lactis IPLA R1 and Bifidobacterium longum IPLA E44 were tested for their capability to modulate immune response and the insulin-dependent glucose homeostasis using male Wistar rats fed with a standard diet. Three intervention groups were fed daily for 24 days with 10% skimmed milk, or with 109 cfu of the corresponding strain suspended in the same vehicle. A significant increase of the suppressor-regulatory TGF-β cytokine occurred with both strains in comparison with a control (no intervention) group of rats; the highest levels were reached in rats fed IPLA R1. This strain presented an immune protective profile, as it was able to reduce the production of the proinflammatory IL-6. Moreover, phosphorylated Akt kinase decreased in gastroctemius muscle of rats fed the strain IPLA R1, without affecting the glucose, insulin, and HOMA index in blood, or levels of Glut-4 located in the membrane of muscle and adipose tissue cells. Therefore, the strain B. animalis subsp. lactis IPLA R1 is a probiotic candidate to be tested in mild grade inflammation animal models. Nuria Salazar, Patricia López, Pablo Garrido, Javier Moran, Estefanía Cabello, Miguel Gueimonde, Ana Suárez, Celestino González, Clara G. de los Reyes-Gavilán, and Patricia Ruas-Madiedo Copyright © 2014 Nuria Salazar et al. All rights reserved. Phenotypic and Genotypic Characterization of Atypical Listeria monocytogenes and Listeria innocua Isolated from Swine Slaughterhouses and Meat Markets Wed, 28 May 2014 12:22:59 +0000 In the last decade, atypical Listeria monocytogenes and L. innocua strains have been detected in food and the environment. Because of mutations in the major virulence genes, these strains have different virulence intensities in eukaryotic cells. In this study, we performed phenotypic and genotypic characterization of atypical L. monocytogenes and L. innocua isolates obtained from swine slaughterhouses and meat markets. Forty strains were studied, including isolates of L. monocytogenes and L. innocua with low-hemolytic activity. The isolates were characterized using conventional phenotypic Listeria identification tests and by the detection and analysis of L. monocytogenes-specific genes. Analysis of 16S rRNA was used for the molecular identification of the Listeria species. The L. monocytogenes isolates were positive for all of the virulence genes studied. The atypical L. innocua strains were positive for hly, plcA, and inlC. Mutations in the InlC, InlB, InlA, PI-PLC, PC-PLC, and PrfA proteins were detected in the atypical isolates. Further in vitro and transcriptomic studies are being developed to confirm the role of these mutations in Listeria virulence. Luisa Zanolli Moreno, Renata Paixão, Debora Dirani Sena de Gobbi, Daniele Cristine Raimundo, Thais Sebastiana Porfida Ferreira, Andrea Micke Moreno, Ernesto Hofer, Cristhiane Moura Falavina dos Reis, Glavur Rogério Matté, and Maria Helena Matté Copyright © 2014 Luisa Zanolli Moreno et al. All rights reserved. Contrasted Reactivity to Oxygen Tensions in Frankia sp. Strain CcI3 throughout Nitrogen Fixation and Assimilation Wed, 28 May 2014 10:47:49 +0000 Reconciling the irreconcilable is a primary struggle in aerobic nitrogen-fixing bacteria. Although nitrogenase is oxygen and reactive oxygen species-labile, oxygen tension is required to sustain respiration. In the nitrogen-fixing Frankia, various strategies have been developed through evolution to control the respiration and nitrogen-fixation balance. Here, we assessed the effect of different oxygen tensions on Frankia sp. strain CcI3 growth, vesicle production, and gene expression under different oxygen tensions. Both biomass and vesicle production were correlated with elevated oxygen levels under both nitrogen-replete and nitrogen-deficient conditions. The mRNA levels for the nitrogenase structural genes (nifHDK) were high under hypoxic and hyperoxic conditions compared to oxic conditions. The mRNA level for the hopanoid biosynthesis genes (sqhC and hpnC) was also elevated under hyperoxic conditions suggesting an increase in the vesicle envelope. Under nitrogen-deficient conditions, the hup2 mRNA levels increased with hyperoxic environment, while hup1 mRNA levels remained relatively constant. Taken together, these results indicate that Frankia protects nitrogenase by the use of multiple mechanisms including the vesicle-hopanoid barrier and increased respiratory protection. Faten Ghodhbane-Gtari, Karima Hezbri, Amir Ktari, Imed Sbissi, Nicholas Beauchemin, Maher Gtari, and Louis S. Tisa Copyright © 2014 Faten Ghodhbane-Gtari et al. All rights reserved. Synergistic Antimicrobial Action of Chlorhexidine and Ozone in Endodontic Treatment Wed, 28 May 2014 00:00:00 +0000 Objectives. The aim of this study was to determine whether irrigation with sodium hypochlorite, chlorhexidine, and ozone gas, alone or in combination, were effective against Enterococcus faecalis and Candida albicans; these are microorganisms frequently isolated from teeth with periapical lesions resistant to endodontic treatment. Material and Methods. 220 single root teeth, recently extracted, were inoculated with Candida albicans and Enterococcus faecalis. The formulations tested were sodium hypochlorite at 1, 3, and 5% chlorhexidine at 0.2% and 2% and ozone gas applied for different periods of time. The combination of sodium hypochlorite at 5% and chlorhexidine at 2%, with gaseous ozone, were also assessed. For the most active treatments the mechanism of action was assessed through flow cytometry. Results. Sodium hypochlorite, chlorhexidine, and gaseous ozone alone were ineffective in completely eliminating the microorganisms. The association of chlorhexidine at 2% followed by ozone gas for 24 seconds promoted the complete elimination of Candida albicans and Enterococcus faecalis. Flow cytometry shows that ozone and chlorhexidine act differently, which could explain its synergic activity. Conclusions. This new disinfection protocol, combining irrigation with chlorhexidine at 2% and ozone gas for 24 seconds, may be advantageous when treating infected root canals. Rita Noites, Cidália Pina-Vaz, Rita Rocha, Manuel Fontes Carvalho, Acácio Gonçalves, and Irene Pina-vaz Copyright © 2014 Rita Noites et al. All rights reserved. Screening for Genes Coding for Putative Antitumor Compounds, Antimicrobial and Enzymatic Activities from Haloalkalitolerant and Haloalkaliphilic Bacteria Strains of Algerian Sahara Soils Tue, 27 May 2014 10:23:57 +0000 Extreme environments may often contain unusual bacterial groups whose physiology is distinct from those of normal environments. To satisfy the need for new bioactive pharmaceuticals compounds and enzymes, we report here the isolation of novel bacteria from an extreme environment. Thirteen selected haloalkalitolerant and haloalkaliphilic bacteria were isolated from Algerian Sahara Desert soils. These isolates were screened for the presence of genes coding for putative antitumor compounds using PCR based methods. Enzymatic, antibacterial, and antifungal activities were determined by using cultural dependant methods. Several of these isolates are typical of desert and alkaline saline soils, but, in addition, we report for the first time the presence of a potential new member of the genus Nocardia with particular activity against the yeast Saccharomyces cerevisiae. In addition to their haloalkali character, the presence of genes coding for putative antitumor compounds, combined with the antimicrobial activity against a broad range of indicator strains and their enzymatic potential, makes them suitable for biotechnology applications. Okba Selama, Gregory C. A. Amos, Zahia Djenane, Chiara Borsetto, Rabah Forar Laidi, David Porter, Farida Nateche, Elizabeth M. H. Wellington, and Hocine Hacène Copyright © 2014 Okba Selama et al. All rights reserved. Statistical Approach for Production of PUFA from Kocuria sp. BRI 35 Isolated from Marine Water Sample Sun, 25 May 2014 11:02:48 +0000 In this study, Plackett-Burman design was used to identify the most influential parameters affecting PUFA production by Kocuria sp. BRI 35 isolated from Antarctic water sample. Amongst 10 variables evaluated, magnesium chloride, protease peptone, glucose, and temperature were significant. Response surface methodology consisting of a central composite design was developed to study the interactions between the variables and to determine optimal values of significant variables. A quadratic model (R = 0.9652, F = 14.64, P < 0.0001) was built. The contour plots indicated that the isolate produced maximum PUFA at lower concentrations of magnesium sulfate (0.9 g/L) and higher concentrations of protease peptone (5 g/L) and glucose (10 g/L) at 15°C. MgSO4 and glucose exhibited quadratic as well as interactive effect on PUFA production whereas protease peptone and temperature showed interactive effects only. After optimization, PUFA production per unit biomass increased from 0.94 mg/g to 11.12 mg/g. This represented an increase from 3% to 58.62% of the total fatty acids. Among PUFAs, the yield of ω-6 fatty acids increased from 9.66 mg/L to 107.71 mg/L with significant increase in linoleic acid (20.36 mg/L) whereas ω-3 fatty acids increased up to 12.37 mg/L with DHA being the major ω-3 fatty acid produced. Swanandi Pote and Rama Bhadekar Copyright © 2014 Swanandi Pote and Rama Bhadekar. All rights reserved. Evaluation of Microbial Enzymes in Normal and Abnormal Cervicovaginal Fluids of Cervical Dysplasia: A Case Control Study Thu, 22 May 2014 11:46:52 +0000 The aim of the present study was to evaluate the role of microbial enzymes in normal and abnormal cervicovaginal fluids of cervical dysplasia. The cervicovaginal infections were evaluated through the estimation of microbial enzymes in patients with and without abnormal cervical cytology like bacterial and fungal infections. The patients were categorized based on infection caused by organism and stages of dysplasia. The pH, Whiff test, and Pap smear tests were conducted for normal and abnormal cervical swabs based on standard protocols. Microbial enzymes include mucinase, sialidases, and proteases of the cervical swabs and are estimated according to standard methods. The results of abnormal cervical cytological smears showed increased pH and the presence of amines with different levels of Pap smear test. Increased levels of microbial enzymes were observed in patients with abnormal cytology than normal cytology. Three microbial enzymes mucinase, sialidase, and protease were significantly () more elevated in patients with bacterial infections (, , ) than without dysplasia (, , ). The results reinforce that the microbial infection seems to be more prone to cervical dysplasia and may act as risk-factor for the development of cervical cancer along with HPV infection. Subramanyam Dasari, Wudayagiri Rajendra, and Lokanatha Valluru Copyright © 2014 Subramanyam Dasari et al. All rights reserved. Three-Dimensional In Vitro Models of Granuloma to Study Bacteria-Host Interactions, Drug-Susceptibility, and Resuscitation of Dormant Mycobacteria Wed, 21 May 2014 07:15:24 +0000 Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium bovis, and Mycobacterium avium subsp. paratuberculosis can survive within host macrophages in a dormant state, encased within an organized aggregate of immune host cells called granuloma. Granulomas consist of uninfected macrophages, foamy macrophages, epithelioid cells, and T lymphocytes accumulated around infected macrophages. Within granulomas, activated macrophages can fuse to form multinucleated giant cells, also called giant Langhans cells. A rim of T lymphocytes surrounds the core, and a tight coat of fibroblast closes the structure. Several in vivo models have been used to study granuloma’s structure and function, but recently developed in vitro models of granuloma show potential for closer observation of the early stages of host’s responses to live mycobacteria. This paper reviews culture conditions that resulted in three-dimensional granulomas, formed by the adhesion of cell populations in peripheral blood mononuclear cells infected with mycobacteria. The similarities of these models to granulomas encountered in clinical specimens include cellular composition, granulomas’ cytokine production, and cell surface antigens. A reliable in vitro dormancy model may serve as a useful platform to test whether drug candidates can kill dormant mycobacteria. Novel drugs that target dormancy-specific pathways may shorten the current long, difficult treatments necessary to cure mycobacterial diseases. Liam E. Fitzgerald, Naiara Abendaño, Ramon A. Juste, and Marta Alonso-Hearn Copyright © 2014 Liam E. Fitzgerald et al. All rights reserved. Antibacterial Activity of pH-Dependent Biosynthesized Silver Nanoparticles against Clinical Pathogen Wed, 21 May 2014 00:00:00 +0000 Simple, nontoxic, environmental friendly method is employed for the production of silver nanoparticles. In this study the synthesized nanoparticles UV absorption band occurred at 400 nm because of the surface Plasmon resonance of silver nanoparticles. The pH of the medium plays important role in the synthesis of control shaped and sized nanoparticles. The colour intensity of the aqueous solution varied with pH. In this study, at pH 9, the colour of the aqueous solution was dark brown, whereas in pH 5 the colour was yellowish brown; the colour difference in the aqueous solution occurred due to the higher production of silver nanoparticles. The antibacterial activity of biosynthesized silver nanoparticles was carried out against E. coli. The silver nanoparticles synthesized at pH 9 showed maximum antibacterial activity at 50 μL. Kethirabalan Chitra and Gurusamy Annadurai Copyright © 2014 Kethirabalan Chitra and Gurusamy Annadurai. All rights reserved. In Vitro Synergistic Effect of Curcumin in Combination with Third Generation Cephalosporins against Bacteria Associated with Infectious Diarrhea Sun, 18 May 2014 12:47:08 +0000 Diarrhea is one of the leading causes of morbidity and mortality in humans in developed and developing countries. Furthermore, increased resistance to antibiotics has resulted in serious challenges in the treatment of this infectious disease worldwide. Therefore, there exists a need to develop alternative natural or combination drug therapies. The aim of the present study was to investigate the synergistic effect of curcumin-1 in combination with three antibiotics against five diarrhea causing bacteria. The antibacterial activity of curcumin-1 and antibiotics was assessed by the broth microdilution method, checkerboard dilution test, and time-kill assay. Antimicrobial activity of curcumin-1 was observed against all tested strains. The MICs of curcumin-1 against test bacteria ranged from 125 to 1000 μg/mL. In the checkerboard test, curcumin-1 markedly reduced the MICs of the antibiotics cefaclor, cefodizime, and cefotaxime. Significant synergistic effect was recorded by curcumin-1 in combination with cefotaxime. The toxicity of curcumin-1 with and without antibiotics was tested against foreskin (FS) normal fibroblast and no significant cytotoxicity was observed. From our result it is evident that curcumin-1 enhances the antibiotic potentials against diarrhea causing bacteria in in vitro condition. This study suggested that curcumin-1 in combination with antibiotics could lead to the development of new combination of antibiotics against diarrhea causing bacteria. Nishanth Kumar Sasidharan, Sreerag Ravikumar Sreekala, Jubi Jacob, and Bala Nambisan Copyright © 2014 Nishanth Kumar Sasidharan et al. All rights reserved. Effect of Er,Cr:YSGG Laser Irradiation with Radial Firing Tips on Candida albicans in Experimentally Infected Root Canals Thu, 15 May 2014 13:35:04 +0000 Aim. To compare the disinfection effect of Er,Cr:YSGG laser using radial firing tips with NaOCI in root canals infected with C. albicans and to evaluate the irradiation effect on the dentinal surfaces. Material and Methods. In total seventy-six mandibular premolar teeth were used. In order to standardize the incubation and sterilization procedure, eight teeth were used. Sixty-eight of the root canals were incubated with C. albicans suspension for 72 hours. The specimens were divided into 5 experimental groups. Two groups were constituted as Group 1 was irradiated with 1.5 W laser () and group 2, which was irradiated with 2 W laser (). Two more groups were formed as Group 3 (2 W laser () and Group 4 NaOCI (5%) (). Group 5 () did not receive any treatment. Mann-Whitney U and Kruskal-Wallis H tests were used to compare the different laser output powers. Wilcoxon Signed Ranks Test was used in order to compare the Candida cfu/ml levels according to treatment protocols (). Results. Both 1.5 W and 2 W laser resulted in a major reduction of C. albicans without a significant difference. The comparison of the dentin surfaces irradiated with Er,Cr:YSGG laser at two power settings resulted in similar morphological changes. However, NaOCI was found to be more effective in reduction of C. albicans than 2 W laser application. Conclusion. According to the results of the present study, the Er,Cr:YSGG laser with radial firing tips presented less antifungal effects on C. albicans in root canals of infected teeth than NaOCl solution. Leman Ozkan, Serap Cetiner, and Tamer Sanlidag Copyright © 2014 Leman Ozkan et al. All rights reserved. A Fast, Reliable, and Sensitive Method for Detection and Quantification of Listeria monocytogenes and Escherichia coli O157:H7 in Ready-to-Eat Fresh-Cut Products by MPN-qPCR Thu, 15 May 2014 06:38:04 +0000 In the present work we developed a MPN quantitative real-time PCR (MPN-qPCR) method for a fast and reliable detection and quantification of Listeria monocytogenes and Escherichia coli O157:H7 in minimally processed vegetables. In order to validate the proposed technique, the results were compared with conventional MPN followed by phenotypic and biochemical assays methods. When L. monocytogenes and E. coli O157:H7 were artificially inoculated in fresh-cut vegetables, a concentration as low as 1 CFU g−1 could be detected in 48 hours for both pathogens. qPCR alone allowed a limit of detection of 101 CFU g−1 after 2 hours of enrichment for L. monocytogenes and E. coli O157:H7. Since minimally processed ready-to-eat vegetables are characterized by very short shelf life, our method can potentially address the consistent reduction of time for microbial analysis, allowing a better management of quality control. Moreover, the occurrences of both pathogenic bacteria in mixed salad samples and fresh-cut melons were monitored in two production plants from the receipt of the raw materials to the early stages of shelf life. No sample was found to be contaminated by L. monocytogenes. One sample of raw mixed salad was found positive to an H7 enterohemorrhagic serotype. Pasquale Russo, Giuseppe Botticella, Vittorio Capozzi, Salvatore Massa, Giuseppe Spano, and Luciano Beneduce Copyright © 2014 Pasquale Russo et al. All rights reserved. Antimicrobial Activity of Coronarin D and Its Synergistic Potential with Antibiotics Thu, 15 May 2014 00:00:00 +0000 Coronarin D is a labdane-type diterpene from the rhizomes of Hedychium coronarium. In the view of our ongoing effort to explore its novel biological activity, antimicrobial activity study of coronarin D was performed. The results showed that coronarin D was active against tested Gram-positive bacteria, inactive for tested Gram-negative bacteria, and weakly active against tested fungi. The antibacterial effect of the combination of coronarin D with nine classical antibiotics against four Gram-positive bacteria was also evaluated. The fractional inhibitory concentration indices (FICI) of coronarin D-antibiotics combinations, calculated from the checkerboard assay, were used as synergism indicator. Out of 36 combinations, 47% showed total synergism, 33% had partial synergistic interaction, 17% showed no effect, and 3% showed antagonism. By combination with coronarin D at concentration of 0.25 minimal inhibitory concentration (MIC), the activities of antibiotics were boosted to 4- to 128-fold. These finding suggested an attractive approach to combat the infectious diseases by using coronarin D-antibiotic drug combination. Nanthawan Reuk-ngam, Nitirat Chimnoi, Nisachon Khunnawutmanotham, and Supanna Techasakul Copyright © 2014 Nanthawan Reuk-ngam et al. All rights reserved. Antifungal and Antiproliferative Protein from Cicer arietinum: A Bioactive Compound against Emerging Pathogens Wed, 14 May 2014 08:36:46 +0000 The emergence of epidemic fungal pathogenic resistance to current antifungal drugs has increased the interest in developing alternative antibiotics from natural sources. Cicer arietinum is well known for its medicinal properties. The aim of this work was to isolate antimicrobial proteins from Cicer arietinum. An antifungal protein, C-25, was isolated from Cicer arietinum and purified by gel filtration. C-25 protein was tested using agar diffusion method against human pathogenic fungi of ATCC strains and against clinical isolates of Candida krusei, Candida tropicalis, and Candida parapsilosis, and MIC values determined were varied from 1.56 to 12.5 μg/mL. The SEM study demonstrated that C-25 induces the bleb-like surface changes, irregular cell surface, and cell wall disruption of the fungi at different time intervals. Cytotoxic activity was studied on oral cancer cells and normal cells. It also inhibits the growth of fungal strains which are resistant to fluconazole. It reduced the cell proliferation of human oral carcinoma cells at the concentration of 37.5 μg/mL (IC50) and no toxic effect was found on normal human peripheral blood mononuclear cells even at higher concentration of 600 μg/mL. It can be concluded that C-25 can be considered as an effective antimycotic as well as antiproliferative agent against human oral cancer cells. Suresh Kumar, Vaishali Kapoor, Kamaldeep Gill, Kusum Singh, Immaculata Xess, Satya N. Das, and Sharmistha Dey Copyright © 2014 Suresh Kumar et al. All rights reserved. Epidemiology of Carbapenemase-Producing Enterobacteriaceae and Acinetobacter baumannii in Mediterranean Countries Tue, 13 May 2014 13:04:43 +0000 The emergence and global spread of carbapenemase-producing Enterobacteriaceae and Acinetobacter baumannii are of great concern to health services worldwide. These β-lactamases hydrolyse almost all β-lactams, are plasmid-encoded, and are easily transferable among bacterial species. They are mostly of the KPC, VIM, IMP, NDM, and OXA-48 types. Their current extensive spread worldwide in Enterobacteriaceae is an important source of concern. Infections caused by these bacteria have limited treatment options and have been associated with high mortality rates. Carbapenemase producers are mainly identified among Klebsiella pneumoniae, Escherichia coli, and A. baumannii and still mostly in hospital settings and rarely in the community. The Mediterranean region is of interest due to a great diversity and population mixing. The prevalence of carbapenemases is particularly high, with this area constituting one of the most important reservoirs. The types of carbapenemase vary among countries, partially depending on the population exchange relationship between the regions and the possible reservoirs of each carbapenemase. This review described the epidemiology of carbapenemases produced by enterobacteria and A. baumannii in this part of the world highlighting the worrisome situation and the need to screen and detect these enzymes to prevent and control their dissemination. Nassima Djahmi, Catherine Dunyach-Remy, Alix Pantel, Mazouz Dekhil, Albert Sotto, and Jean-Philippe Lavigne Copyright © 2014 Nassima Djahmi et al. All rights reserved. Prevalence and Correlation with Clinical Diseases of Helicobacter pylori cagA and vacA Genotype among Gastric Patients from Northeast China Tue, 13 May 2014 08:59:39 +0000 Helicobacter pylori vacA and cagA genes have significant genetic heterogenicity, resulting in different clinical outcomes. Northeast part of China has reported high prevalence of H. pylori infections and gastric cancer. Hence, we investigated the H. pylori cagA and vacA genotypes with clinical outcomes in Northeast China. Gastric tissue samples (), chronic gastritis (GIs), gastric ulcer (GU), and gastric cancer (GC) were analysed for 16S rRNA ureA, cagA, and cagA genotypes by PCR. A total of 141 (84%) cases were found positive for H. pylori by 16S rRNA and ureA. GC showed high H. pylori infection (93%) compared with GIs (72%) and GU (84%). The vacAs1am1 was highly found in GC (40%) and GU (36%), vacAs1am2 in GIs (33%), vacAs1bm1 (14%) and vacAs1bm2 (8%) in GU cases, and s2m1 in normal cases (33%), while vacAs1cm1 showed low frequency in GIs (2%) and GU (3%) and GC showed negative result. The East-Asian cagA strain was highly observed in GC (43%), as compared to GIs (41%) and GU (20%). The East-Asian cagA/vacAs1am1 was significantly higher in GC (23%) than in GU (22%) and GIs (145) patients. The East-Asian type cagA with vacAs1a and vacAm1 is the most predominant genotype in H. pylori strains of Northeast China. Faisal Aziz, Xin Chen, Xuesong Yang, and Qiu Yan Copyright © 2014 Faisal Aziz et al. All rights reserved. Safety Characterization and Antimicrobial Properties of Kefir-Isolated Lactobacillus kefiri Tue, 13 May 2014 08:00:47 +0000 Lactobacilli are generally regarded as safe; however, certain strains have been associated with cases of infection. Our workgroup has already assessed many functional properties of Lactobacillus kefiri, but parameters regarding safety must be studied before calling them probiotics. In this work, safety aspects and antimicrobial activity of L. kefiri strains were studied. None of the L. kefiri strains tested caused - or -hemolysis. All the strains were susceptible to tetracycline, clindamycin, streptomycin, ampicillin, erythromycin, kanamycin, and gentamicin; meanwhile, two strains were resistant to chloramphenicol. On the other hand, all L. kefiri strains were able to inhibit both Gram(+) and Gram(−) pathogens. Regarding the in vitro results, L. kefiri CIDCA 8348 was selected to perform in vivo studies. Mice treated daily with an oral dose of 108 CFU during 21 days showed no signs of pain, lethargy, dehydration, or diarrhea, and the histological studies were consistent with those findings. Moreover, no differences in proinflammatory cytokines secretion were observed between treated and control mice. No translocation of microorganisms to blood, spleen, or liver was observed. Regarding these findings, L. kefiri CIDCA 8348 is a microorganism isolated from a dairy product with a great potential as probiotic for human or animal use. Paula Carasi, Mariángeles Díaz, Silvia M. Racedo, Graciela De Antoni, María C. Urdaci, and María de los Angeles Serradell Copyright © 2014 Paula Carasi et al. All rights reserved. Emetic Bacillus cereus Are More Volatile Than Thought: Recent Foodborne Outbreaks and Prevalence Studies in Bavaria (2007–2013) Thu, 08 May 2014 00:00:00 +0000 Several Bacillus cereus strains possess the genetic fittings to produce two different types of toxins, the heat-stable cereulide or different heat-labile proteins with enterotoxigenic potential. Unlike the diarrheal toxins, cereulide is (pre-)formed in food and can cause foodborne intoxications shortly after ingestion of contaminated food. Based on the widely self-limiting character of cereulide intoxications and rarely performed differential diagnostic in routine laboratories, the real incidence is largely unknown. Therefore, during a 7-year period about 4.300 food samples linked to foodborne illness with a preliminary report of vomiting as well as food analysed in the context of monitoring programs were investigated to determine the prevalence of emetic B. cereus in food environments. In addition, a lux-based real-time monitoring system was employed to assess the significance of the detection of emetic strains in different food matrices and to determine the actual risk of cereulide toxin production in different types of food. This comprehensive study showed that emetic strains are much more volatile than previously thought. Our survey highlights the importance and need of novel strategies to move from the currently taxonomic-driven diagnostic to more risk orientated diagnostics to improve food and consumer safety. Ute Messelhäusser, Elrike Frenzel, Claudia Blöchinger, Renate Zucker, Peter Kämpf, and Monika Ehling-Schulz Copyright © 2014 Ute Messelhäusser et al. All rights reserved. In Vitro Antibacterial, Antioxidant, and Cytotoxic Activities of Parthenium hysterophorus and Characterization of Extracts by LC-MS Analysis Wed, 07 May 2014 11:45:30 +0000 Present work reports the biological activities of P. hysterophorus leaf, stem, flower, and root. Dried samples were sequentially extracted with many solvents. Hexane (HX), benzene (BZ), and chloroform (CH) extracts of leaf showed considerable antibacterial activity against Streptococcus mutans (MTCC 497), Proteus vulgaris (MTCC 7299), and Salmonella typhi (MTCC 3917). Flower extracts exhibited presence of higher amount of flavonoids (13.9–59.6 μgQE/mg) followed by leaf, stem, and root. Stem (HX, BZ, and CH), leaf ethanol (ET), and root (HX, BZ, and CH) fractions showed noticeable antioxidant capacity in phosphomolybdate assay. Most of the extracts demonstrated beta carotene bleaching inhibition capability. BZ, ethyl acetate (EA), and ET fractions of leaves, stem aqueous (AQ), and flower EA extracts showed membrane protective activities (40–55%). Middle fractions of the plant parts displayed moderate antihemolytic potential. Most of the flower extracts exhibited cytotoxic activity (80–100%) against lung and colon cancer cell lines. Root (HX and ET) and leaf ET extracts showed considerable inhibition (90–99%) of colon and ovary cancer cell lines. The LC-MS scan demonstrated presence of different compounds showing 3–20 min retention time. The study revealed considerable antibacterial, antioxidant, lipo-protective, antihemolytic, and anticancer potential in all parts of P. hysterophorus. Shashank Kumar, Sanjay Pandey, and Abhay K. Pandey Copyright © 2014 Shashank Kumar et al. All rights reserved. Synthesis and Antimicrobial Activities of 5-Arylidene-thiazolidine-2,4-dione Derivatives Wed, 07 May 2014 07:48:51 +0000 Antibiotic resistance is considered one of the world's major public health concerns. The main cause of bacterial resistance is the improper and repeated use of antibiotics. To alleviate this problem, new chemical substances against microorganisms are being synthesized and tested. Thiazolidines are compounds having many pharmacological activities including antimicrobial activities. For this purpose some thiazolidine derivatives substituted at position 5 in the thiazolidine nucleus were synthesized and tested against several microorganisms. Using a disc diffusion method, antimicrobial activity was verified against Gram-positive, Gram-negative, and alcohol acid resistant bacteria and yeast. The minimum inhibition concentrations (MIC) and minimum bactericidal concentrations (MBC) were determined. All derivatives showed antimicrobial activity mainly against Gram-positive bacteria, with MIC values ranging from 2 to 16 µg/mL. Ivanildo Mangueira da Silva, João da Silva Filho, Priscila Brandão Gomes da Silva Santiago, Micalyne Soares do Egito, Carlos André de Souza, Frederico Leite Gouveia, Rafael Matos Ximenes, Kêsia Xisto da Fonseca Ribeiro de Sena, Antonio Rodolfo de Faria, Dalci José Brondani, and Julianna Ferreira Cavalcanti de Albuquerque Copyright © 2014 Ivanildo Mangueira da Silva et al. All rights reserved. Bioactive Compounds Derived from the Yeast Metabolism of Aromatic Amino Acids during Alcoholic Fermentation Mon, 05 May 2014 14:08:43 +0000 Metabolites resulting from nitrogen metabolism in yeast are currently found in some fermented beverages such as wine and beer. Their study has recently attracted the attention of researchers. Some metabolites derived from aromatic amino acids are bioactive compounds that can behave as hormones or even mimic their role in humans and may also act as regulators in yeast. Although the metabolic pathways for their formation are well known, the physiological significance is still far from being understood. The understanding of this relevance will be a key element in managing the production of these compounds under controlled conditions, to offer fermented food with specific enrichment in these compounds or even to use the yeast as nutritional complements. Albert Mas, Jose Manuel Guillamon, Maria Jesus Torija, Gemma Beltran, Ana B. Cerezo, Ana M. Troncoso, and M. Carmen Garcia-Parrilla Copyright © 2014 Albert Mas et al. All rights reserved. Antagonistic Interactions of “Ya-Sa-Marn-Phlae” Ethanol Extract in Combination with Topical Antiseptics against Clinical Isolates of Staphylococcus aureus Mon, 05 May 2014 08:26:25 +0000 This investigation was aimed at assessing a possible interaction of a traditional Thai herbal recipe, “Ya-Sa-Marn-Phlae (YSMP),” used for wound treatments with topical antiseptics, povidone-iodine (PI) solution and hydrogen peroxide (H2O2), and effects of THR-SK010 alone and the combinations on Staphylococcus aureus. Antibacterial activities of ethanol extracts from the herbal recipe were determined against both methicillin resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA). YSMP exhibited remarkable antistaphylococcal activity with MIC values of 3.9–7.8 μg/mL. This recipe possessed bacteriostatic activity and did not reduce the tolerance of both MRSA and MSSA isolates to the high ionic strength. Interaction between THR-SK010 and the antiseptics was carried out by checkerboard testing and time-kill assay. Both indifferent and slightly antagonistic effects were observed with THR-SK010/PI and THR-SK010/H2O2 combinations against the tested isolates. In addition to commercially available antiseptics, THR-SK010 offered additional therapeutic options for the decolonization of MRSA and MSSA. Topical application of plant extracts with antioxidant activity, such as THR-SK010, should not be used immediately with PI or H2O2 and further investigation on this interaction is needed. Sasitorn Chusri, Sirirat Tongrod, Julalak Chokpaisarn, Surasak Limsuwan, and Supayang Piyawan Voravuthikunchai Copyright © 2014 Sasitorn Chusri et al. All rights reserved. Recent Developments in Mycobacteriology: A Clinical and Diagnostic Perspective Wed, 30 Apr 2014 09:46:50 +0000 Tomasz Jagielski, Jakko van Ingen, Nalin Rastogi, and Jarosław Dziadek Copyright © 2014 Tomasz Jagielski et al. All rights reserved. The Kampo Medicine Rokumigan Possesses Antibiofilm, Anti-Inflammatory, and Wound Healing Properties Wed, 30 Apr 2014 09:09:36 +0000 Periodontal diseases, which are inflammatory diseases of bacterial origin affecting the tooth-supporting tissues, are characterized by inflammation and destruction of gingival connective tissue and alveolar bone, and may lead to tooth loss. The aim of the study was to investigate Rokumigan, a Kampo Japanese traditional medicine made of six different plants, for its capacity to prevent biofilm formation by Fusobacterium nucleatum, to inhibit interleukin-6 (IL-6) and interleukin-8 (IL-8) secretion by mucosal cells, and to promote wound healing in a fibroblast model. Using a microplate colorimetric assay, Rokumigan prevented biofilm formation by F. nucleatum, while it had no effect on bacterial growth. Rokumigan inhibited IL-6 secretion in both epithelial cells and fibroblasts stimulated with lipopolysaccharide. However, it caused no significant inhibition of IL-8 secretion by both cell types. Rokumigan significantly increased proliferation and migration of gingival fibroblasts in a wound healing assay. In conclusion, the Kampo formulation Rokumigan, through suppression of biofilm formation by F. nucleatum, inhibition of IL-6 secretion by gingival epithelial cells and fibroblasts, and promotion of wound healing in a fibroblast model, may have potential application for periodontal diseases. James Liao, Jabrane Azelmat, Lei Zhao, Masami Yoshioka, Daisuke Hinode, and Daniel Grenier Copyright © 2014 James Liao et al. All rights reserved. Emergent Bacteria in Cystic Fibrosis: In Vitro Biofilm Formation and Resilience under Variable Oxygen Conditions Tue, 29 Apr 2014 13:17:52 +0000 Concurrent to conventional bacterial pathogens, unusual microbes are emerging from cystic fibrosis (CF) airways. Nonetheless, little is known about the contribution of these newly microbes to the resilience of CF-associated biofilms, particularly under variable-oxygen concentrations that are known to occur in vivo in the mucus of CF patients. Two CF-emergent bacterial species, Inquilinus limosus and Dolosigranulum pigrum, and the major pathogen Pseudomonas aeruginosa were studied in terms of biofilm development and antibiotic susceptibilities under in vitro atmospheres with different oxygen availabilities. All species were able to develop in vitro biofilms under different oxygen-available environments, with D. pigrum accumulating high amounts of biomass and respiratory activities. When established, biofilms were of difficult eradication, with antibiotics losing their effectiveness in comparison with the corresponding planktonic populations. Surprisingly, biofilms of each emergent organism displayed multidrug resistance under aerobic environments, enduring even in low-oxygen atmospheres. This study suggests a potential prospect on the impact of nonconventional organisms I. limosus and D. pigrum on CF lung infections, demonstrating capacity to adapt to biofilm mode of life under restricted-oxygen atmospheres resembling CF airways, which may ultimately endanger the efficacy of currently used antibiotic regimens. Susana P. Lopes, Nuno F. Azevedo, and Maria O. Pereira Copyright © 2014 Susana P. Lopes et al. All rights reserved. The Structural Modeling of the Interaction between Levofloxacin and the Mycobacterium tuberculosis Gyrase Catalytic Site Sheds Light on the Mechanisms of Fluoroquinolones Resistant Tuberculosis in Colombian Clinical Isolates Mon, 28 Apr 2014 14:07:17 +0000 We compared the prevalence of levofloxacin (LVX) resistance with that of ofloxacin (OFX) and moxifloxacin (MFX) among multidrug resistant (MDR) MTB clinical isolates collected in Medellin, Colombia, between 2004 and 2009 and aimed at unraveling the underlying molecular mechanisms that explain the correlation between QRDR-A mutations and LVX resistance phenotype. We tested 104 MDR isolates for their susceptibility to OFX, MFX, and LVX. Resistance to OFX was encountered in 10 (9.6%) of the isolates among which 8 (7.7%) were also resistant to LVX and 6 (5.7%) to MFX. Four isolates resistant to the 3 FQ were harboring the Asp94Gly substitution, whilst 2 other isolates resistant to OFX and LVX presented the Ala90Val mutation. No mutations were found in the QRDR-B region. The molecular modeling of the interaction between LVX and the DNA-DNA gyrase complex indicates that the loss of an acetyl group in the Asp94Gly mutation removes the acid base interaction with LVX necessary for the quinolone activity. The Ala90Val mutation that substitutes a methyl for an isopropyl group induces a steric modification that blocks the LVX access to the gyrase catalytic site. N. Alvarez, E. Zapata, G. I. Mejía, T. Realpe, P. Araque, C. Peláez, F. Rouzaud, and J. Robledo Copyright © 2014 N. Alvarez et al. All rights reserved. Predicting Mycobacterium tuberculosis Complex Clades Using Knowledge-Based Bayesian Networks Wed, 23 Apr 2014 00:00:00 +0000 We develop a novel approach for incorporating expert rules into Bayesian networks for classification of Mycobacterium tuberculosis complex (MTBC) clades. The proposed knowledge-based Bayesian network (KBBN) treats sets of expert rules as prior distributions on the classes. Unlike prior knowledge-based support vector machine approaches which require rules expressed as polyhedral sets, KBBN directly incorporates the rules without any modification. KBBN uses data to refine rule-based classifiers when the rule set is incomplete or ambiguous. We develop a predictive KBBN model for 69 MTBC clades found in the SITVIT international collection. We validate the approach using two testbeds that model knowledge of the MTBC obtained from two different experts and large DNA fingerprint databases to predict MTBC genetic clades and sublineages. These models represent strains of MTBC using high-throughput biomarkers called spacer oligonucleotide types (spoligotypes), since these are routinely gathered from MTBC isolates of tuberculosis (TB) patients. Results show that incorporating rules into problems can drastically increase classification accuracy if data alone are insufficient. The SITVIT KBBN is publicly available for use on the World Wide Web. Minoo Aminian, David Couvin, Amina Shabbeer, Kane Hadley, Scott Vandenberg, Nalin Rastogi, and Kristin P. Bennett Copyright © 2014 Minoo Aminian et al. All rights reserved. Absence of Cospeciation between the Uncultured Frankia Microsymbionts and the Disjunct Actinorhizal Coriaria Species Tue, 22 Apr 2014 12:25:40 +0000 Coriaria is an actinorhizal plant that forms root nodules in symbiosis with nitrogen-fixing actinobacteria of the genus Frankia. This symbiotic association has drawn interest because of the disjunct geographical distribution of Coriaria in four separate areas of the world and in the context of evolutionary relationships between host plants and their uncultured microsymbionts. The evolution of Frankia-Coriaria symbioses was examined from a phylogenetic viewpoint using multiple genetic markers in both bacteria and host-plant partners. Total DNA extracted from root nodules collected from five species: C. myrtifolia, C. arborea, C. nepalensis, C. japonica, and C. microphylla, growing in the Mediterranean area (Morocco and France), New Zealand, Pakistan, Japan, and Mexico, respectively, was used to amplify glnA gene (glutamine synthetase), dnaA gene (chromosome replication initiator), and the nif DK IGS (intergenic spacer between nifD and nifK genes) in Frankia and the matK gene (chloroplast-encoded maturase K) and the intergenic transcribed spacers (18S rRNA-ITS1-5.8S rRNA-ITS2-28S rRNA) in Coriaria species. Phylogenetic reconstruction indicated that the radiations of Frankia strains and Coriaria species are not congruent. The lack of cospeciation between the two symbiotic partners may be explained by host shift at high taxonomic rank together with wind dispersal and/or survival in nonhost rhizosphere. Imen Nouioui, Faten Ghodhbane-Gtari, Maria P. Fernandez, Abdellatif Boudabous, Philippe Normand, and Maher Gtari Copyright © 2014 Imen Nouioui et al. All rights reserved. Two-Dimensional PCA Highlights the Differentiated Antitumor and Antimicrobial Activity of Methanolic and Aqueous Extracts of Laurus nobilis L. from Different Origins Wed, 16 Apr 2014 16:06:46 +0000 Natural matrices are important sources of new antitumor and antimicrobial compounds. Species such as Laurus nobilis L. (laurel) might be used for this purpose, considering its medicinal properties. Herein, in vitro activity against human tumor cell lines, bacteria, and fungi was evaluated in enriched phenolic extracts. Specifically, methanol and aqueous extracts of wild and cultivated samples of L. nobilis were compared considering different phenolic groups. Principal component analysis (PCA) was applied to understand how each extract acts differentially against specific bacteria, fungi, and selected human tumor cell lines. In general, the extract type induced the highest differences in bioactivity of laurel samples. However, from the PCA biplot, it became clear that wild laurel samples were higher inhibitors of tumor cell lines (HeLa, MCF7, NCI-H460, and HCT15). HepG2 had the same response to laurel from wild and cultivated origin. It was also observed that methanolic extracts tended to have higher antimicrobial activity, except against A. niger, A. fumigatus, and P. verrucosum. The differences in bioactivity might be related to the higher phenolic contents in methanolic extracts. These results allow selecting the extract type and/or origin with highest antibacterial, antifungal, and antitumor activity. Maria Inês Dias, João C. M. Barreira, Ricardo C. Calhelha, Maria-João R. P. Queiroz, M. Beatriz P. P. Oliveira, Marina Soković, and Isabel C. F. R. Ferreira Copyright © 2014 Maria Inês Dias et al. All rights reserved. Molecular Screening of Virulence Genes in Extraintestinal Pathogenic Escherichia coli Isolated from Human Blood Culture in Brazil Tue, 15 Apr 2014 09:55:01 +0000 Extraintestinal pathogenic Escherichia coli (ExPEC) is one of the main etiological agents of bloodstream infections caused by Gram-negative bacilli. In the present study, 20 E. coli isolates from human hemocultures were characterized to identify genetic features associated with virulence (pathogenicity islands markers, phylogenetic group, virulence genes, plasmid profiles, and conjugative plasmids) and these results were compared with commensal isolates. The most prevalent pathogenicity island, in strains from hemoculture, were PAI IV536, described by many researchers as a stable island in enterobacteria. Among virulence genes, iutA gene was found more frequently and this gene enconding the aerobactin siderophore receptor. According to the phylogenetic classification, group B2 was the most commonly found. Additionally, through plasmid analysis, 14 isolates showed plasmids and 3 of these were shown to be conjugative. Although in stool samples of healthy people the presence of commensal strains is common, human intestinal tract may serve as a reservoir for ExPEC. Vanessa L. Koga, Geizecler Tomazetto, Paula S. Cyoia, Meiriele S. Neves, Marilda C. Vidotto, Gerson Nakazato, and Renata K. T. Kobayashi Copyright © 2014 Vanessa L. Koga et al. All rights reserved. Detection of Food Spoilage and Pathogenic Bacteria Based on Ligation Detection Reaction Coupled to Flow-Through Hybridization on Membranes Thu, 10 Apr 2014 11:02:14 +0000 Traditional culturing methods are still commonly applied for bacterial identification in the food control sector, despite being time and labor intensive. Microarray technologies represent an interesting alternative. However, they require higher costs and technical expertise, making them still inappropriate for microbial routine analysis. The present study describes the development of an efficient method for bacterial identification based on flow-through reverse dot-blot (FT-RDB) hybridization on membranes, coupled to the high specific ligation detection reaction (LDR). First, the methodology was optimized by testing different types of ligase enzymes, labeling, and membranes. Furthermore, specific oligonucleotide probes were designed based on the 16S rRNA gene, using the bioinformatic tool Oligonucleotide Retrieving for Molecular Applications (ORMA). Four probes were selected and synthesized, being specific for Aeromonas spp., Pseudomonas spp., Shewanella spp., and Morganella morganii, respectively. For the validation of the probes, 16 reference strains from type culture collections were tested by LDR and FT-RDB hybridization using universal arrays spotted onto membranes. In conclusion, the described methodology could be applied for the rapid, accurate, and cost-effective identification of bacterial species, exhibiting special relevance in food safety and quality. K. Böhme, P. Cremonesi, M. Severgnini, Tomás G. Villa, I. C. Fernández-No, J. Barros-Velázquez, B. Castiglioni, and P. Calo-Mata Copyright © 2014 K. Böhme et al. All rights reserved. Evaluation of Three Formulations of Culture Media for Isolation of Brucella spp. regarding Their Ability to Inhibit the Growth of Contaminating Organisms Thu, 10 Apr 2014 08:16:26 +0000 Three culture media (Brucella agar, Farrell medium, and CITA) were compared for their effectiveness in inhibiting contamination and for isolating Brucella spp. One hundred lymph nodes from pigs (n = 50) and wild boars (n = 50) with lymphadenitis were collected in slaughterhouses in the State of São Paulo and were assessed on these three selective media for Brucella spp. All of the samples were negative for Brucella spp. on the three culture media. On the agar medium, fungal (70 plates) and Gram-positive bacterial (59 plates) contaminants were observed; in the CITA medium, the absence of fungal and Gram-positive bacteria on 15 plates was observed; no bacterial or fungal growth was observed on the Farrell media. The results demonstrated that the CITA and Farrell media inhibited the growth of contaminants better than the Brucella agar. Acácia F. Vicente, João M. A. P. Antunes, Gustavo H. B. Lara, Mateus S. R. Mioni, Susan D. Allendorf, Marina G. Peres, Camila M. Appolinário, Fernando J. P. Listoni, Marcio G. Ribeiro, and Jane Megid Copyright © 2014 Acácia F. Vicente et al. All rights reserved. Molecular Analysis of Ciprofloxacin Resistance Mechanisms in Malaysian ESBL-Producing Klebsiella pneumoniae Isolates and Development of Mismatch Amplification Mutation Assays (MAMA) for Rapid Detection of gyrA and parC Mutations Thu, 10 Apr 2014 08:08:42 +0000 Ninety-three Malaysian extended-spectrum -lactamase (ESBL)-producing Klebsiella pneumoniae isolates were investigated for ciprofloxacin resistance. Two mismatch amplification mutation (MAMA) assays were developed and used to facilitate rapid detection of gyrA and parC mutations. The isolates were also screened for plasmid-mediated quinolone resistance (PMQR) genes including aac(6′)-Ib-cr, qepA, and qnr. Ciprofloxacin resistance (MICs  μg/mL) was noted in 34 (37%) isolates, of which 33 isolates had multiple mutations either in gyrA alone or in both gyrA and parC regions . aac(6′)-Ib-cr was the most common PMQR gene detected in this study , followed by qnrB and qnrS ( and 1, resp.). Low-level ciprofloxacin resistance (MICs 1-2 μg/mL) was noted in 40 (43%) isolates carrying qnrB accompanied by either aac(6′)-Ib-cr or a single gyrA 83 mutation . Ciprofloxacin resistance was significantly associated with the presence of multiple mutations in gyrA and parC regions. While the isolates harbouring gyrA and/or parC alteration were distributed into 11 PFGE clusters, no specific clusters were associated with isolates carrying PMQR genes. The high prevalence of ciprofloxacin resistance amongst the Malaysian ESBL-producing K. pneumoniae isolates suggests the need for more effective infection control measures to limit the spread of these resistant organisms in the hospital. Farah Al-Marzooq, Mohd Yasim Mohd Yusof, and Sun Tee Tay Copyright © 2014 Farah Al-Marzooq et al. All rights reserved. Improving the Diagnosis of Bloodstream Infections: PCR Coupled with Mass Spectrometry Wed, 09 Apr 2014 09:11:26 +0000 The reference method for the diagnosis of bloodstream infections is blood culture followed by biochemical identification and antibiotic susceptibility testing of the isolated pathogen. This process requires 48 to 72 hours. The rapid administration of the most appropriate antimicrobial treatment is crucial for the survival of septic patients; therefore, a rapid method that enables diagnosis directly from analysis of a blood sample without culture is needed. A recently developed platform that couples broad-range PCR amplification of pathogen DNA with electrospray ionization mass spectrometry (PCR/ESI-MS) has the ability to identify virtually any microorganism from direct clinical specimens. To date, two clinical evaluations of the PCR/ESI-MS technology for the diagnosis of bloodstream infections from whole blood have been published. Here we discuss them and describe recent improvements that result in an enhanced sensitivity. Other commercially available assays for the molecular diagnosis of bloodstream infections from whole blood are also reviewed. The use of highly sensitive molecular diagnostic methods in combination with conventional procedures could substantially improve the management of septic patients. Elena Jordana-Lluch, Montserrat Giménez, M. Dolores Quesada, Vicente Ausina, and Elisa Martró Copyright © 2014 Elena Jordana-Lluch et al. All rights reserved. Antibacterial Potential of Northeastern Portugal Wild Plant Extracts and Respective Phenolic Compounds Tue, 08 Apr 2014 08:20:24 +0000 The present work aims to assess the antibacterial potential of phenolic extracts, recovered from plants obtained on the North East of Portugal, and of their phenolic compounds (ellagic, caffeic, and gallic acids, quercetin, kaempferol, and rutin), against bacteria commonly found on skin infections. The disk diffusion and the susceptibility assays were used to identify the most active extracts and phenolic compounds. The effect of selected phenolic compounds on animal cells was assessed by determination of cellular metabolic activity. Gallic acid had a higher activity, against gram-positive (S. epidermidis and S. aureus) and gram-negative bacteria (K. pneumoniae) at lower concentrations, than the other compounds. The caffeic acid, also, showed good antibacterial activity against the 3 bacteria used. The gallic acid was effective against the 3 bacteria without causing harm to the animal cells. Gallic and caffeic acid showed a promising applicability as antibacterial agents for the treatment of infected wounds. Eva Pinho, Isabel C. F. R. Ferreira, Lillian Barros, Ana Maria Carvalho, Graça Soares, and Mariana Henriques Copyright © 2014 Eva Pinho et al. All rights reserved. Genomic and Proteomic Characterization of Bacteriocin-Producing Leuconostoc mesenteroides Strains Isolated from Raw Camel Milk in Two Southwest Algerian Arid Zones Mon, 07 Apr 2014 16:33:57 +0000 Information on the microbiology of camel milk is very limited. In this work, the genetic characterization and proteomic identification of 13 putative producing bacteriocin Leuconostoc strains exhibiting antilisterial activity and isolated from camel milk were performed. DNA sequencing of the 13 selected strains revealed high homology among the 16S rRNA genes for all strains. In addition, 99% homology with Leuconostoc mesenteroides was observed when these sequences were analysed by the BLAST tool against other sequences from reference strains deposited in the Genbank. Furthermore, the isolates were characterized by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDITOF MS) which allowed for the identification of 2 mass peaks 6242 m/z and 5118 m/z that resulted to be specific to the species L. mesenteroides. Remarkably, the phyloproteomic tree provided more intraspecific information of L. mesenteroides than phylogenetic analysis. Accordingly, phyloproteomic analysis grouped L. mesenteroides strains into different subbranches, while all L. mesenteroides isolates were grouped in the same branch according to phylogenetic analysis. This study represents, to our knowledge, the first report on the use of MALDI-TOF MS on the identification of LAB isolated from camel milk. Zineb Benmechernene, Inmaculada Fernández-No, Marcos Quintela-Baluja, Karola Böhme, Mebrouk Kihal, Pilar Calo-Mata, and Jorge Barros-Velázquez Copyright © 2014 Zineb Benmechernene et al. All rights reserved. Bacteriological Profile and Drug Resistance Patterns of Blood Culture Isolates in a Tertiary Care Nephrourology Teaching Institute Mon, 07 Apr 2014 00:00:00 +0000 Blood stream infections can lead to life threatening sepsis and require rapid antimicrobial treatment. The organisms implicated in these infections vary with the geographical alteration. Infections caused by MDR organisms are more likely to increase the risk of death in these patients. The present study was aimed to study the profile of organisms causing bacteremia and understand antibiotic resistance patterns in our hospital. 1440 blood samples collected over a year from clinically suspected cases of bacteremia were studied. The isolates were identified by standard biochemical tests and antimicrobial resistance patterns were determined by CLSI guidelines. Positive blood cultures were obtained in 9.2% of cases of which Gram-positive bacteria accounted for 58.3% of cases with staph aureus predominance; gram negative bacteria accounted for 40.2% with enterobactereciea predominence; and 1.5% were fungal isolates. The most sensitive drugs for Gram-positive isolates were vancomycin, teicoplanin, daptomycin, linezolid, and tigecycline and for Gram-negative were carbapenems, colistin, aminoglycosides, and tigecycline. The prevalence of MRSA and vancomycin resistance was 70.6% and 21.6%, respectively. ESBL prevalence was 39.6%. Overall low positive rates of blood culture were observed. Kalpesh Gohel, Amit Jojera, Shailesh Soni, Sishir Gang, Ravindra Sabnis, and Mahesh Desai Copyright © 2014 Kalpesh Gohel et al. All rights reserved. Sequence and Apoptotic Activity of VacA Cytotoxin Cloned from a Helicobacter pylori Thai Clinical Isolate Wed, 02 Apr 2014 12:46:12 +0000 The vacuolating cytotoxin VacA produced by Helicobacter pylori induces the formation of large cytoplasmic vacuoles in host gastric epithelial cells as well as a release of cytochrome C from mitochondria resulting in cell apoptosis. Considerable sequence diversity in VacA relating to different degrees of disease severity is observed with clinical samples from a multitude of geographic places. In this study we describe expression in Escherichia coli, purification to homogeneity and in vitro assay of its apoptotic activity of a VacA toxin from a H. pylori isolate of a Thai patient with gastrointestinal lymphoma. Sequencing revealed that the deduced amino acid sequence of the cloned Thai isolate VacA is similar to H. pylori s1/m2 type strains. The percent sequence similarity to the model strain 60190 was lower due to the presence of extra amino acids in the mid (m) region. The purified VacA toxin exhibited significant apoptotic activity on both T84 and MDCK epithelial cell lines, as revealed by DAPI staining, whereby the observed activity was significantly higher on MDCK cells. These findings could relate to a modulation of VacA activity on host cells in the Thai isolate-VacA toxin that may differ from those of the model strain. Muhammad Junaid, Sarbast Al-Gubare, Muhammad Yousef, Mathukorn Na Ubol, Somphob Leetachewa, Chatchai Muanprasat, Chanan Angsuthanasombat, Wanpen Chaicumpa, Niaz Ali, and Gerd Katzenmeier Copyright © 2014 Muhammad Junaid et al. All rights reserved. Staphylococcus aureus and Staphylococcal Food-Borne Disease: An Ongoing Challenge in Public Health Tue, 01 Apr 2014 16:24:57 +0000 Staphylococcal food-borne disease (SFD) is one of the most common food-borne diseases worldwide resulting from the contamination of food by preformed S. aureus enterotoxins. It is one of the most common causes of reported food-borne diseases in the United States. Although several Staphylococcal enterotoxins (SEs) have been identified, SEA, a highly heat-stable SE, is the most common cause of SFD worldwide. Outbreak investigations have found that improper food handling practices in the retail industry account for the majority of SFD outbreaks. However, several studies have documented prevalence of S. aureus in many food products including raw retail meat indicating that consumers are at potential risk of S. aureus colonization and subsequent infection. Presence of pathogens in food products imposes potential hazard for consumers and causes grave economic loss and loss in human productivity via food-borne disease. Symptoms of SFD include nausea, vomiting, and abdominal cramps with or without diarrhea. Preventive measures include safe food handling and processing practice, maintaining cold chain, adequate cleaning and disinfection of equipment, prevention of cross-contamination in home and kitchen, and prevention of contamination from farm to fork. This paper provides a brief overview of SFD, contributing factors, risk that it imposes to the consumers, current research gaps, and preventive measures. Jhalka Kadariya, Tara C. Smith, and Dipendra Thapaliya Copyright © 2014 Jhalka Kadariya et al. All rights reserved. Biomarkers for Sepsis Sun, 30 Mar 2014 12:25:56 +0000 Bloodstream infections are a major concern because of high levels of antibiotic consumption and of the increasing prevalence of antimicrobial resistance. Bacteraemia is identified in a small percentage of patients with signs and symptoms of sepsis. Biomarkers are widely used in clinical practice and they are useful for monitoring the infectious process. Procalcitonin (PCT) and C-reactive protein (CRP) have been most widely used, but even these have limited abilities to distinguish sepsis from other inflammatory conditions or to predict outcome. PCT has been used to guide empirical antibacterial therapy in patients with respiratory infections and help to determine if antibacterial therapy can be stopped. New biomarkers such as those in this review will discuss the major types of biomarkers of bloodstream infections/sepsis, including soluble triggering receptor expressed on myeloid cells-1 (sTREM-1), soluble urokinase-type plasminogen receptor (suPAR), proadrenomedullin (ProADM), and presepsin. Cesar Henriquez-Camacho and Juan Losa Copyright © 2014 Cesar Henriquez-Camacho and Juan Losa. All rights reserved. Worldwide Dissemination of the NDM-Type Carbapenemases in Gram-Negative Bacteria Wed, 26 Mar 2014 13:36:23 +0000 The emergence of one of the most recently described carbapenemases, namely, the New Delhi metallo-lactamase (NDM-1), constitutes a critical and growingly important medical issue. This resistance trait compromises the efficacy of almost all lactams (except aztreonam), including the last resort carbapenems. Therapeutical options may remain limited mostly to colistin, tigecycline, and fosfomycin. The main known reservoir of NDM producers is the Indian subcontinent whereas a secondary reservoir seems to have established the Balkans regions and the Middle East. Although the spread of -like genes (several variants) is derived mostly by conjugative plasmids in Enterobacteriaceae, this carbapenemase has also been identified in P. aeruginosa and Acinetobacter spp. Acinetobacter sp. may play a pivotal role for spreading genes for its natural reservoir to Enterobacteriaceae. Rapid diagnostic techniques (Carba NP test) and screening of carriers are the cornerstone to try to contain this outbreak which threatens the efficacy of the modern medicine. Laurent Dortet, Laurent Poirel, and Patrice Nordmann Copyright © 2014 Laurent Dortet et al. All rights reserved. Performance of Quantification of Modified Hodge Test: An Evaluation with Klebsiella pneumoniae Carbapenemase-Producing Enterobacteriaceae Isolates Wed, 26 Mar 2014 08:58:43 +0000 Modified Hodge Test (MHT) has been suggested as screening tests for carbapenemases, but concerns regarding its difficult interpretation and common false-positive results obtained in the presence of other β-lactamases have been noted. This study aimed to quantify the enhanced growth formed by the indicator strain and thus evaluate the performance of a quantitative interpretation of MHT for KPC screening. MHT was performed in 50 KPC-producing isolates and 334 non-carbapenemase-producing isolates, using ertapenem (ETP) and meropenem (MEM) as substrates. The size of enhanced growth of indicator strain was measured for each isolate tested and for the positive control used, and a ratio was calculated. Our results revealed 17 different ETP and MEM ratios, with distinct sensitivity (SN) and specificity (SP). Higher SN combined to higher SP was achieved when ETP and MEM ratios were 0.45, with a SN value of 96% for both substrates and SP values of 99.4% and 100% for ETP and MEM, respectively. The quantification with both substrates increased SP of the test for KPC detection. Considering that MHT is the unique phenotypic test that is referred to by CLSI, a more accurate approach for its interpretation could be applied to make it a more useful tool. Vanessa Bley Ribeiro, Adriano Rostirolla Linhares, Alexandre P. Zavascki, and Afonso Luis Barth Copyright © 2014 Vanessa Bley Ribeiro et al. All rights reserved. Potential Synergy Activity of the Novel Ceragenin, CSA-13, against Carbapenem-Resistant Acinetobacter baumannii Strains Isolated from Bacteremia Patients Mon, 24 Mar 2014 16:36:24 +0000 Carbapenem-resistant Acinetobacter baumannii is an important cause of nosocomial infections, particularly in patients in the intensive care units. As chronic infections are difficult to treat, attempts have been made to discover new antimicrobials. Ceragenins, designed to mimic the activities of antimicrobial peptides, are a new class of antimicrobial agents. In this study, the in vitro activities of CSA-13 either alone or in combination with colistin (sulphate), tobramycin, and ciprofloxacin were investigated using 60 carbapenem-resistant A. baumannii strains isolated from bacteremia patients blood specimens. MICs and MBCs were determined by microbroth dilution technique. Combinations were assessed by using checkerboard technique. The MIC50 values (mg/L) of CSA-13, colistin, tobramycin, and ciprofloxacin were 2, 1, 1.25, and 80, respectively. The MIC90 (mg/L) of CSA-13 and colistin were 8 and 4. The MBCs were equal to or twice greater than those of the MICs. Synergistic interactions were mostly seen with CSA-13-colistin (55%), whereas the least synergistic interactions were observed in the CSA-13-tobramycin (35%) combination. No antagonism was observed. CSA-13 appears to be a good candidate for further investigations in the treatment of A. baumannii infections. However, future studies should be performed to correlate the safety, efficacy, and pharmacokinetic parameters of this molecule. Cagla Bozkurt-Guzel, Paul B. Savage, Alper Akcali, and Berna Ozbek-Celik Copyright © 2014 Cagla Bozkurt-Guzel et al. All rights reserved. A Novel Electronic Nose as Adaptable Device to Judge Microbiological Quality and Safety in Foodstuff Mon, 24 Mar 2014 07:37:40 +0000 This paper presents different applications, in various foodstuffs, by a novel electronic nose (EN) based on a mixed metal oxide sensors array composed of thin films as well as nanowires. The electronic nose used for this work has been done, starting from the commercial model EOS835 produced by SACMI Scarl. The SENSOR Lab (CNR-INO, Brescia) has produced both typologies of sensors, classical MOX and the new technologies with nanowire. The aim of this work was to test and to illustrate the broad spectrum of potential uses of the EN technique in food quality control and microbial contamination diagnosis. The EN technique was coupled with classical microbiological and chemical techniques, like gas chromatography with mass spectroscopy (GC-MS) with SPME technique. Three different scenarios are presented: (a) detection of indigenous mould in green coffee beans, (b) selection of microbiological spoilage of Lactic Acid Bacteria (LAB), and (c) monitoring of potable water. In each case, the novel EN was able to identify the spoiled product by means of the alterations in the pattern of volatile organic compounds (VOCs), reconstructed by principal component analysis (PCA) of the sensor responses. The achieved results strongly encourage the use of EN in industrial laboratories. Finally, recent trends and future directions are illustrated. V. Sberveglieri, E. Nunez Carmona, Elisabetta Comini, Andrea Ponzoni, Dario Zappa, Onofrio Pirrotta, and A. Pulvirenti Copyright © 2014 V. Sberveglieri et al. All rights reserved. Aetiology of Bacteraemia as a Risk Factor for Septic Shock at the Onset of Febrile Neutropaenia in Adult Cancer Patients Thu, 20 Mar 2014 00:00:00 +0000 Septic shock (SS) at the onset of febrile neutropaenia (FN) is an emergency situation that is associated with high morbidity and mortality. The impact of the specific aetiology of bloodstream infections (BSIs) in the development of SS at the time of FN is not well established. The aim of this study was to evaluate the association between the aetiology of BSIs and SS at the time of FN in hospitalised adult cancer patients. This prospective cohort study was performed at a single tertiary hospital from October 2009 to August 2011. All adult cancer patients admitted consecutively to the haematology ward with FN were evaluated. A stepwise logistic regression was conducted to verify the association between the microbiological characteristics of BSIs and SS at the onset of FN. In total, 307 cases of FN in adult cancer patients were evaluated. There were 115 cases with documented BSI. A multivariate analysis showed that polymicrobial bacteraemia () was associated with SS. The specific blood isolates independently associated with SS were viridans streptococci () and Escherichia coli (). Neutropaenic cancer patients with polymicrobial bacteraemia or BSI by viridans streptococci or Escherichia coli are at increased risk for SS at the time of FN. Regis Goulart Rosa and Luciano Zubaran Goldani Copyright © 2014 Regis Goulart Rosa and Luciano Zubaran Goldani. All rights reserved. Biomarkers for Sepsis: A Review with Special Attention to India Wed, 19 Mar 2014 08:01:55 +0000 Sepsis is a serious infection and still a common cause of morbidity and mortality in resource-limited settings such as India. Even when microbiologic diagnostics are available, bacteremia is only identified in a proportion of patients who present with sepsis and bloodstream infections. Biomarkers have been used in a variety of disease processes and can help aid in diagnosing bacterial infections. There have been numerous biomarkers investigated to aid with diagnosis and prognostication in sepsis with the majority suffering from lack of sensitivity or specificity. Procalcitonin has been heralded as the biomarker that holds the most promise for bloodstream infections. Data are emerging in India, and in this review, we focus on the current data of biomarkers in sepsis with particular attention to how biomarkers could be used to augment diagnosis and treatment in India. George E. Nelson, Vidya Mave, and Amita Gupta Copyright © 2014 George E. Nelson et al. All rights reserved. Experimental Inoculation of BFDV-Positive Budgerigars (Melopsittacus undulatus) with Two Mycobacterium avium subsp. avium Isolates Thu, 13 Mar 2014 16:07:17 +0000 Beak and feather disease virus- (BFDV-) positive (naturally infected) but clinically healthy budgerigars (Melopsittacus undulatus) were inoculated with two isolates of Mycobacterium avium subsp. avium isolated from naturally infected golden pheasant (Chrysolophus pictus) and peafowl (Pavo cristatus). During a period of more than two months after inoculation, samples of cloacal and crop swabs, faeces, and blood were obtained for BFDV and Mycobacterium avium testing with PCR. Birds were euthanized nine weeks after inoculation. All infected budgerigars developed signs typical of mycobacteriosis, but more advanced clinical and pathological changes were visible in the group infected with the pheasant isolate. Only a few cloacal and crop swab samples were positive for Mycobacterium avium subsp. avium despite advanced pathological changes in the internal organs. In the groups infected with mycobacterium isolates the frequency of BFDV-positive samples was higher than in the control group. In the infected groups the frequency of BFDV was substantially higher in the cloacal swabs of birds inoculated with the pheasant isolate than in the peafowl-isolate-infected group. Aleksandra Ledwoń, Rafał Sapierzyński, Ewa Augustynowicz-Kopeć, Piotr Szeleszczuk, and Marcin Kozak Copyright © 2014 Aleksandra Ledwoń et al. All rights reserved. A Systematic Follow-Up of Mycobacterium tuberculosis Drug-Resistance and Associated Genotypic Lineages in the French Departments of the Americas over a Seventeen-Year Period Thu, 13 Mar 2014 13:52:08 +0000 The population of the French Departments of the Americas (FDA) is highly influenced by the intense migratory flows with mainland France and surrounding countries of the Caribbean and Latin America, some of which have high incidence rates of tuberculosis (Haiti: 230/100,000; Guyana: 111/100,000; and Suriname: 145/100,000) and drug resistance. Since the development of drug resistance to conventional antituberculous drugs has a major impact on the treatment success of tuberculosis, we therefore decided to review carefully Mycobacterium tuberculosis drug resistance and associated genotypic lineages in the FDA over a seventeen-year period (January 1995–December 2011). A total of 1239 cases were studied, including 153 drug-resistant and 26 multidrug-resistant- (MDR-) TB cases, representing 12.3% and 2.1% of the TB cases in our study setting. A significantly higher proportion of M. tuberculosis isolates among relapse cases showed drug resistance to isoniazid (22.5%, ), rifampicin (20.0%, ), or both (MDR-TB, 17.5%; ). Determination of spoligotyping based phylogenetic clades showed that among the five major lineages observed—T family (30.1%); Latin-American and Mediterranean (LAM, 23.7%); Haarlem (H, 22.2%); East-African Indian (EAI, 7.2%); and X family (6.5%)—two lineages, X and LAM, were overrepresented in drug-resistant and MDR-TB cases, respectively. Finally, 19 predominant spoligotypes were identified for the 1239 isolates of M. tuberculosis in our study among which 4 were significantly associated with drug resistance corresponding to SIT20/LAM1, SIT64/LAM6, SIT45/H1, and SIT46/undefined lineage. Julie Millet, Elisabeth Streit, Mylène Berchel, Anne-Gaël Bomer, Franziska Schuster, Delaina Paasch, Jessica Vanhomwegen, Gilbert Cadelis, and Nalin Rastogi Copyright © 2014 Julie Millet et al. All rights reserved. A First Assessment of Mycobacterium tuberculosis Genetic Diversity and Drug-Resistance Patterns in Twelve Caribbean Territories Mon, 10 Mar 2014 14:19:26 +0000 With the exception of some French-speaking islands, data on tuberculosis (TB) in the Caribbean are scarce. In this study, we report a first assessment of genetic diversity of a convenience sample of Mycobacterium tuberculosis strains received from twelve Caribbean territories by spoligotyping and describe their drug-resistance patterns. Of the 480 isolates, 40 (8.3%) isolates showed resistance to at least one anti-TB drug. The proportion of drug-resistant strains was significantly higher in The Bahamas (21.4%; ), and Guyana (27.5%; ), while it was significantly lower in Jamaica (2.4%; ) than in other countries of the present study. Regarding genetic diversity, 104 distinct spoligotype patterns were observed: 49 corresponded to clustered strains (2 to 93 strains per cluster), while 55 remained unclustered among which 16 patterns were not reported previously. Combining the study results with regional data retrieved from the international SITVIT2 database underlined a connection between frequency of certain M. tuberculosis phylogenetic lineages and the language spoken, suggesting historical (colonial) and ongoing links (trade, tourism, and migratory flows) with European countries with which they shared a common past. Julie Millet, Shirematee Baboolal, Elisabeth Streit, Patrick E. Akpaka, and Nalin Rastogi Copyright © 2014 Julie Millet et al. All rights reserved.