Simultaneous Detection of invA, STM4497, and fliC183 Genes in Salmonella typhimurium by Multiplex PCR Method in Poultry Meat Samples in Zanjan, Iran

Authors
S. Dousandeh 1
G. Asaadi Tehrani 2
B. Amini 3
P. Maziri 4
1 Department of Microbiology, Faculty of Basic Sciences, Islamic Azad University, Zanjan Branch, Zanjan, IR Iran
2 Department of Genetics, Faculty of Basic sciences, Islamic Azad University, Zanjan Branch, Zanjan, IR Iran
3 Department of Microbiology, Zanjan University of Medical Science, Zanjan, IR Iran
4 Department of Genetics, Faculty of Basic Sciences, Islamic Azad University, Zanjan Branch, Zanjan, IR Iran
Abstract
Background: Salmonella typhimurium is one of the most important species of Salmonella that is intracellular parasite and attacks host mucus membrane. These bacteria can cause gastroenteritis, and their main transmission route is water, poultry, meat, egg, and raw food. The aim of this study was to detect three virulence genes associated with S. typhimurium named invA, STM4497, and fliC183 genes by Multiplex PCR method.
Materials and Methods: 183 samples of poultry were collected from food products in Zanjan (Iran) and cultured in BPW (Buffered Peptone Water) for 18 hr and at 37°C, and in RVS broth (Rappaport Vassiliadis Soya) for 6 hr at 41.5°C. After amplification of genomic DNA by Multiplex PCR method, occurrence of pathogen contamination was checked and compared with standard strain.
Results: From the total of 183 collected samples, 52(28.4%) samples were positive for S. typhimurium. The frequency of STM4497, fliC183, and invA genes were 49 (27%), 3 (2%), and 53 (29%), respectively.
Conclusion: Simultaneous detection of invA, STM4497, and fliC183 genes were recognized as a key for detection of S. typhimurium by Multiplex PCR method.

Keywords

  1. Malorny B, Hoorfar J, Hugas M, Heuvelink A, Fach P, Ellerbyoek L, et al. Interlaboratory diagnostic accuracy of a Salmonella specific PCR-based method. Int J Food Microbiol. 2003; 89(2-3): 241-9.

  2. Suo B, He Y, Tu SI, Shi X. A multiplex real-time polymerase chain reaction for simultaneous detection of Salmonella spp., EscherichiacoliO157, and Listeria monocyto genes in meat products. Foodborne Pathog Dis. 2010; 7(6): 619-28.

  3. 3.Dilmaghani M, Ahmadi M, Zahraei-Salehi T, Talebi A. Detection of Salmonella­ enterica serovar typhimurium from avians using multiplex-PCR. Vet Res Forum. 2011; 2(3): 157-65.

  4. Pui CF, Wong WC, Chai LC, Tunung R, Jeyaletchumi R, Noor Hidayah MS, et al. Salmonella: A foodborne pathogen. Int Food Res J. 2011; 18(2): 465-73.

  5. Tahergorabi R, Matak KE, Jaczynski J. Application of electron beam to inactivate Salmonella in food: Recent developments. Food Res Inter. 2012; 45(2): 685-94.

  6. Bailey JS, Cosby DE. Detection of Salmonellae from chicken rinses and chicken hot dogs with automated Bax PCR system. J Food Prot. 2003; 66(11): 2138-40

  7. Kimura AC, Reddy V, Marcus R, Cieslak PR, Mohle-Boetani JC, Kassenborg HD , et al. Chicken consumption is a newly identified risk factor for sporadic Salmonella enterica serotype enteritidis infections in the United States: a case-control study in FoodNet sites. Clin Infect Dis. 2004; 38(3): 244-52.

  8. Zahraei-Salehi T, Mahzounieh M, Saeedzadeh A. Detection of invA gene in isolated Salmonella from broilers by PCR method. Int J Poult Sci. 2005; 4(8): 557-9.

  9. Shanmugasundaram M, Radhika M, Murali HS, Batra HV. Detection of Salmonella enterica serovar Typhimurium by selective amplification of fliC, fljB, iroB, invA, rfbJ, STM2755, STM4497 genes by polymerase chain reaction in a monoplex and multiplex format. World J Microbiol Biotechnol. 2009; 25(8): 1385–94.

  10. Soumet C, Ermel G, Rose N, Rose V, Drouin P, Salvat G, et al. Evaluation of a multiplex PCR assay for simultaneous identification of Salmonella sp., Salmonella enteritidis and Salmonella typhimurium from environmental swabs of poultry houses. Lett Appl Microbiol. 1999; 28(2): 113-7.

  11. Malorny B, Hoorfar J, Bunge C, Helmuth R. Multicenter validation of the analytical accuracy of Salmonella PCR: towards an international standard. Appl Environ Microbiol. 2003; 69(1): 290-6.

  12. Rahn K, De Grandis SA, Clarke RC, McEwen SA, Galán JE, Ginocchio C, et al. Amplification of an invA gene sequence of Salmonella typhimurium by polymerase chain reaction as a specific method of detection of Salmonella. See comment in PubMed Commons belowMol Cell Probes. 1992; 6(4): 271-9.

  13. Mousavi SL, Rasooli I, Nazarian S, Amani J. Simultaneous detection of Escherichia coli O157:H7, toxigenic Vibrio cholerae, and Salmonella typhimurium by multiplex PCR. Iran J Clin Infect Dis. 2009; 4(2): 97-103.

  14. Chen Z, Zhang K, Yin H, Li Q, Wang L, Liu Z. Detection of Salmonella and several common Salmonella serotypes in food by loop-mediated isothermal amplification method. Food Science and Human Wellness. 2015; 4(2): 75-9. See comment in PubMed Commons below

  15. Aldridge P, Gnerer J, Karlinsey JE, Hughes KT. Transcriptional and translational control of the Salmonella fliC gene. J Bacteriol. 2006; 188(12): 4487-96.

  16. Millar BC, Xu J, Moore JE. Molecular diagnostics of medically important bacterial infections.  Curr Issues Mol Biol. 2007; 9(1): 21-39.

  17. O'Regan E, McCabe E, Burgess C, McGuinness S, Barry T, Duffy G, et al. Development of a real-time multiplex PCR assay for the detection of multiple Salmonella serotypes in chicken samples. BMC Microbiol. 2008; 8: 156.

  18. Malkawi HI, Gharaibeh R. Rapid and simultaneous identification of two Salmonella enterica serotypes, enteritidis and typhimurium, from chicken and meat products by multiplex PCR. Biotech. 2004; 3(1): 44-8.

  19. Rathnayaka R, Nguyen TMT. Simultaneous detection of four food borne bacterial pathogens by metal hydroxide immobilised multiplex PCR method. Trop Agric Res Ext. 2010; 12(1): 42-6.

  20. Jamshidi A, Bassami MR, Afshari-Nic S. Identification of Salmonella spp. and Salmonella typhimurium by a multiplex PCR-based assay from poultry carcasses in Mashhad, Iran. Int J Vet Res. 2009; 3(1): 43-8.

  21. Karami A, Ranjbar R, Ahmadi Z, Safiri Z. Rapid detection of different serovars of Salmonella enterica by multiplex PCR. Iran J Public Health. 2007; 36(2): 38-42.

  22. Aseel AS, Mayada FH, Majed HM. Isolation and molecular identification of Salmonella typhimurium from chicken meat in Iraq. J World's Poult Res. 2011; 3(2): 63-6.

  23. Jeong ES, Lee KS, Heo SH, Seo JH, Choi YK. Triplex PCR for the simultaneous detection of Pseudomonas aeruginosa, Helicobacter hepaticus and Salmonella typhimurium. Exp Anim. 2011; 60(1): 65-70.

  24. De-Freitas CG, Santana AP, Da-Silva PH, Gonçalves VS, Barros Mde A, Torres FA , et al . PCR multiplex for detection of Salmonella enteritidis, typhi and typhimurium and occurrence in poultry meat. Int J Food Microbiol. 2010; 139(1-2): 15-22.

  25. 25. Guo L, Killefer J, Kenney PB, Amick-Morris JD. Use of arbitrarily primed polymerase chain reaction to study Salmonella ecology in a turkey production environment. Poult Sci. 1999; 78(1): 24-31.

  26. 26. Ferretti R, Mannazzu I, Cocolin L, Comi G, Clementi F. Twelve-hour PCR-based method for detection of Salmonella spp. in food. Appl Environ Microbiol. 2001; 67(2): 977–8.

  27. 27. Itoh Y, Hirose K, Miyake M, Khan AQ, Hashimoto Y, Ezaki T. Amplification of rfbE and fliC genes by polymerase chain reaction for identification and detection of Salmonella serovar Enteritidis, Dublin and Gallinarum-Pullorum. Microbiol Immunol. 1997; 41(10): 791-4.

  28. 28. Saeki EK, Alves J, Bonfante RC, Hirooka EY. De Oliveira TCRM. Multiplex PCR (mPCR) for the detection of Salmonella spp. and the differentiation of the Typhimurium and Enteritidis serovars in Chicken Meat. J Food Safety. 2013; 33(1): 25-9.

Infection Epidemiology and Microbiology
Volume 2, Issue 4 - Serial Number 5
October 2016
Pages 20-23