German bacteriologist H. E. Suchsland announces that the delicate aroma and subtle shades of flavor which affect the palate of the smoker are not due to the tobacco but are attributed to the microbes which aid in the process of tobacco fermentation. A patent based upon this observation was submitted, presumably to improve the poor quality of German tobacco by adding to the harvested tobacco leaves bacteria that he had isolated and grown in his laboratory from high-quality West Indian tobacco.
The microbial degradation of nicotine and nicotinic acid was reported. The morphological and physiological properties of the nicotine-decomposing bacteria were also described.
W. C. Flanders of R. J. Reynolds Tobacco Company issues a 70-page report of a three-year study to determine if the number of microorganisms (bacteria and mold) changed appreciably during aging. Experiments were also conducted to determine if the recorded changes in the microbes follow the changes in the chemical components of tobaccos. These studies were continued and extended for several years.
Pseudomonas aeruginosa and other potentially pathogenic fungi and bacteria were identified in snuff. Similar microbial isolates from a patient was the basis for the physician to theorize that some of the snuff-derived microbes may be responsible in part for chronic bronchitis.
The results of studies were reported that had been undertaken to characterize the deposition of cigarette smoke particles and debris released from the cigarette filter into the human respiratory tract. Popular brand cigarettes were smoked mechanically and in a manner to reflect normal smoking behavior. The studies documented that tobacco flakes and fine tobacco leaf debris were released into mainstream smoke from the cigarette filter of all brands that were tested (Tareyton, Winston, Kent, L&M, Marlboro, and Viceroy). The tobacco flakes and other particulates (filter fibers and carbon from charcoal filters) were studied by light and electron microscopy.
Comparative studies were preformed for microbiological activity in the smoke of popular brand nonfiltered and filtered cigarettes that had been “cold smoked” or lit. Viable bacteria were found in the smoke of all cigarettes tested.
The tobacco from different popular brands of cigarettes was analyzed for bacteria. The number of bacteria was determined on “our own” (Philip Morris) and competitive cigarette fillers. This test was run for several months and each month Viceroy, Brown & Williamson’s product, always showed the lowest degree of “contaminant.” The difference between the brands was statistically significant. Brands tested included Salem, Pall Mall, Chesterfield, Kool, Kent, Viceroy, Winston, and Marlboro. The number of bacteria on Marlboro were “too numerous to count.”
A 189-page report was prepared by investigators at the Brown & Williamson Tobacco Company that presents methods for the microbiological examination of tobacco and tobacco products. The writings include the description of techniques for the quantitative determination of bacteria and fungi and methods for the isolation of potentially human pathogenic microorganisms including Coliform bacteria. Also identified were Staphylococcus aureus, Enterococci, Pseudomonas, Clostridium, and Aspergillus.
A 52-page report that describes a “contact plate method” in which a whole cigarette is rolled over the surface of the nutrient agar dish. Viable microbes that are transferred from the cigarette to the plate are illustrated. Presumably, the intent of the assay was to measure the growth of microbes that would be transferred from the cigarette paper to the hand of the smoker. Other studies showed the growth of microbes from a natural wrapper of a cigar. Also, culture methods were established for testing for coliform bacteria and for counting viable fungi in tobacco.
A 346-page in-house document is produced by the British-American Tobacco Company entitled “Methods for the Microbiological Examination of Tobacco and Tobacco Products.” The authors describe the “Public Health Aspects” of smoking and smokeless tobacco products. They note that “[T]he detection of micro-organisms of health significance in tobacco products must be expected to be regarded as undesirable or even unacceptable by public agencies, regardless of whether there is proof of the significance in initiating or spreading infection in man. Therefore, it is suggested that tobacco products should be substantially free, or contain only minimal numbers, of micro-organisms of potential health significance to man which could conceivably occur on tobacco…” Suggested standards are presented for tobacco products for various bacteria and fungi, and standards that had been established for food products (fish, sausage, meat pies, cream yogurt, soft cheese, and pasteurized milk).
Philip Morris characterizes the microbial population on Marlboro tobaccos throughout the processing line. Five different Marlboro Make-Your-Own tobaccos with various anti-microbial preservatives were evaluated microbiologically for mold and bacteria over time. The microflora of Marlboro raw and tobacco blends were defined for burley, oriental, flue-cured, and other tobacco types.
Bacillus spores were identified in chewing tobacco sold in the USA. Broth of the culture microbes evoked plasma exudation from the oral mucosa when tested using a hamster cheek pouch assay.
Microbiologists in Sweden used a mass-spectrophotometry-based assay to document that tobacco smoking increased dramatically the air concentrations of endotoxin (LPS). The authors note that smoke-derived LPS may be a health risk factor associated with environmental tobacco smoke.
A US Patent was assigned to Philip Morris for an “antibacterial lavage” method to treat tobacco leaves so as to eliminate or reduce bacterial endotoxins (LPS) and tobacco-specific nitrosamines that are formed during the curing process. Bacteria found on tobacco leaves were reported to be primarily Gram-negative bacteria, including pseudomonades and enterobacters. In the awarded patent, Hempling notes that bacterial endotoxins can remain as a residue on the tobacco even after the bacteria have been destroyed.
The microbiological composition of tobacco products was defined using culture and chemical analysis. Tobacco smoke was analyzed chemically, and LPS was measured for tobacco leaves and cigarette tobacco.
US Military publishes a report of an investigation that documents bacterial species diversity of varying brands of cigarettes made in the Middle East that were thought to be associated with illnesses of American soldiers deployed in Operation Iraqi Freedom.
Identification of microflora on tobacco using culture-independent methods based on the amplification of microbial 16S rDNA sequences directly from the leaf surfaces. The investigators discovered also that three of five dominant bacterial species on the tobacco could not be cultivated.
The microbiological composition of tobacco products was defined using culture and chemical analysis (of tobacco leaves) or chemical analysis only (tobacco and tobacco smoke). Mesophilic bacteria dominated among the bacteria in both fresh and cured tobacco leaves; however, a wide range of other bacteria, including Gram-negative bacteria, and fungi were delineated. Microbial flora was compared in studies of tobacco from cigarettes from different countries. LPS was also measured.
Bacteria grown from a single flake of tobacco from all brands of smoking (cigarette, cigar, and pipe) and smokeless (snus, snuff, and long cut) tobacco products. In many instances, the bacteria from the tobacco caused hemolysis of blood in blood agar and liquid broth cultures.
Twenty-seven species of bacteria were identified in an analysis of both unaged tobacco and flue-cured tobacco by 16S rRNA sequence analysis. More species () were identified from the unaged flue-cured tobacco leaves than in the aging leaves ( species).
Fifteen classes of bacteria and a broad range of potentially pathogenic organisms were detected in all cigarette samples studied. In greater than 90% of the tobacco samples, the investigators identified Acinetobacter, Bacillus, Burkholderia, Clostridium, Klebsiella, Pseudomonas aeruginosa, and Serratia. The bacteria were identified using a 16S rRNA-based taxonomic microarray. Cloning and sequencing were used to evaluate total bacterial diversity of four brands of cigarettes. Previous studies have shown that smoking was associated with colonization by pathogenic bacteria and an increased risk of lung infection. This study, however, was the first to show that cigarettes themselves could be the source of exposure to a wide array of potentially pathogenic microbes.
