Simultaneous Screening of 24 Target Genes of Foodborne Pathogens in 35 Foodborne Outbreaks Using Multiplex Real-Time SYBR Green PCR Analysis

Table 1

Eight sets of real-time multiplex PCR with 4 primer pairs for 3 target genes and an IAC gene prior to comprehensive and rapid analysis of foodborne outbreak.

Primer set		Species	Target gene	Primer name	Sequence (5^′-3^′)	Gene bank accession no.	Location	Product size (bp)	T 𝑚^a	Tm distance	Refer-ences
	∗^b	Clostridium perfringens	cpe	GAP-11	GGTTCATTAATTGAAACTGGTG	X81849	583–604	154	75.8 ± 0.37		[10]
				GAP-12	AACGCCAATCATATAAATTACAGC		712–736			3.7
A		Providencia alcalifaciens	gyrB	PAG38-F	TCTGCACGGTGTGGGTGTT	AJ300547	38–56	73	79.5 ± 0.79		[2]
				PAG110-R	ACCGTCACGGCGGATTACT		110–92			1
		EHEC (Stx 2)	Stx2	JMS2-F	CGACCCCTCTTGAACATA	EF441616	140–157	108	80.5 ± 0.76		[11]
				JMS2-R	GATAGACATCAAGCCCTCGT		228–247
		Campylobacter coli	ceuE	ceuE-For	CAAGTACTGCAATAAAAACTAGCACTACG	X88849	2777–2805	72	73.7 ± 0.43		[12]
				ceuE-Rev	AGCTATCACCCTCATCACTCATACTAATAG		2848–2819			4
B	*	Campylobacter jejuni	specific	AB-F	CTGAATTTGATACCTTAAGTGCAGC	AL111168	381121–	86	77.7 ± 0.96		[13]
				AB-R	AGGCACGCCTAAACCTATAGCT		381185–			1.9
		TRH-positive Vibrio parahaemolyticus	trh	trh250-F	GGCTCAAAATGGTTAAGCG	AY742213	705–687	250	79.6 ± 0.21		[14]
				trh250-R	CATTTCCGCTCTCATATGC		456–474
		Listeria monocytogenes	hly	Lm-hly-F	GGGAAATCTGTCTCAGGTGATGT	AF253320	973–995	106	77.4 ± 0.78		[15]
				Lm-hly-R	CGATGATTTGAACTTCATCTTTTGC		1078–1054			1.5
C	*	Emetic Bacillus cereus	ces	ces-TM-F	GATGTTTGCGACGATGCAA	DQ360825	8689–8707	65	78.9 ± 0.87		[4]
				ces-TM-R	CTTTCGGCGTGATACCCATT		8734–8793			1.6
		ETEC (LT)	lt	LT-1	TTACGGCGTTACTATCCTCTCTA	X83966	233–255	275	80.5 ± 0.21		[16]
				LT-2	GGTCTCGGTCAGATATGTGATTC		507–485
		ETEC (ST)	st	STa-F	GCTAATGTTGGCAATTTTTATTTCTGTA	M25607	294–321	190	77.1 ± 0.55		[17]
				STa-R	AGGATTACAACAAAGTTCACAGCAGTAA		456–483			1.7
D	*	EHEC and EPEC	eaeA	eae-F2	CATTGATCAGGATTTTTCTGGTGATA	Z11541	899–924	106	78.8 ± 0.54		[18]
				eae-R	CTCATGCGGAAATAGCCGTTA		979–1000			2.6
		V. cholerae	ompW	ompW-F	AACATCCGTGGATTTGGCATCTG	X51948	675–692	89	81.4 ± 0.69		[19]
				ompW-R	GCTGGTTCCTCAACGCTTCTG		741–763
		EAEC	aggR	aggRks1	GTATACACAAAAGAAGGAAGC	Z18751	18–38	254	79.2 ± 0.25		[20]
				aggRKas2	ACAGAATCGTCAGCATCAGC		170–151			0.8
E		EHEC (Stx 1)	Stx1	JMS1-F	GTCACAGTAACAAACCGTAACA	EF441598	509–488	95	80.0 ± 0.72		[11]
				JMS1-R	TCGTTGACTACTTCTTATCTGGA		415–437			1.6
	*	Staphylococcus aureus	femB	FemB-fw	AATTAACGAAATGGGCAGAAACA	AF106850	277–299	93	81.6 ± 0.62		[21]
				FemB-rv	TGCGCAACACCCTGAACTT		351–370
	*	TDH-positive Vibrio parahaemolyticus	tdh	tdh-F176	TCCATCTGTCCCTTTTCCTG	X54341	176–195	247	80.1 ± 0.22		This study
				tdh-R422	AGACACCGCTGCCATTGTAT		422–403			1.9
F		Y. enterocolitica and Y. pseudotuberculosis	yadA	yadA-F1757	ACGAGTTGACAAAGGTTTAGCC	X13882	1757–1778	129	82.0 ± 0.38		This study
				yadA-R1885	GAACCAACCGCTAATGCCTGA		1885–1865			4.3
		Plesiomonas shigelloides	gyrB	PSG-F64	TTAACGCCCTGTCGGATAAG	AJ300545	64–83	250	86.3 ± 0.26		This study
				PSG-R313	TCGAGCAGATGAATCGACAC		313–294
	*	EAEC	astA	EAST-1-S	GCCATCAACACAGTATATCC	L11241	63–82	106	83.7 ± 0.88		[22]
				EAST-AS	GAGTGACGGCTTTGTAGTCC		148–168			1.5
G		EIEC and Shigella spp.	ipaH	ipaH1672-F	CTCTCAGAGGGTGGCTGACC	M32063	1672–1691	90	85.2 ± 0.31		This study
				ipaH1761-R	TCACGCATCACCTGTGCA		1761–1743			3.1
		Aeromonas hydrophila	ahh1	AHH1-F	GCCGAGCGCCCAGAAGGTGAGTT	CP000462	1653360–82	130	88.3 ± 0.48		[23]
				AHH1-R	GAGCGGCTGGATGCGGTTGT		1653473–92
		Enterotoxigenic B. cereus	nheB	SG-F3	GCACTTATGGCAGTATTTGCAGC	DQ153257	2101–2123	152	80.5 ± 0.84		[24]
				SG-R3	GCATCTTTTAAGCCTTCTGGTC		2231–2252			1.9
H	*	Salmonella spp.	invA	invA2-F	GAT TCT GGT ACT AAT GGT GAT GAT C	M90846	132–156	288	82.4 ± 0.28		[25]
				invA2-R	GCCAGGCT AT CGCCAAT AAC		419–400			6.2
		DAEC	daaD	daaD-F31	GTCACCTGCGGGATGTTACT	AF233530	31–50	233	88.6 ± 0.32		This study
				daaD-R263	AGCTCATGACGACCATCCTT		263–244
IAC for sets A-E; IAC for; sets; F,G,H		Yersinia ruckeri	16S rRNA	yersH2-F	GGCTCACCTAGGCGACGA	X75275	245–262	211	86.1 ± 0.53		This study
				yersH2-R	TCAGTGCTATTAACACTTAACCCTTCC		455–429
		Yersinia ruckeri	16S rRNA	yers-F	GGAGGAAGGGTTAAGTGTTA	X75275	426–443	68	77.2 ± 0.53		[9]
				yers-R	GAGTTAGCCGGTGCTTCTT		475–493

^aValues represent means ± standard deviations of 15 to 60 tests. ^bEight main foodborne bacteria.