Rapid recognition of typhoid and non-typhoid Salmonella in poultry samples using multiple polymerase chain reaction method (multiplex-PCR)

Subject Areas : Journal of Comparative Pathobiology

علی Khodadadeh Jigheh ¹ (Graduate of master in microbiology. faculty of basic sciences. Tabriz branch. Islamic Azad university. Tabriz Iran)
یونس Anzabi ² (-Associate Professor. department of pathobiology. faculty of veterinary medicine. Tabriz medical sciences. Islamic Azad university. Tabriz Iran -Department of Microbial Biotechnology, Biotechnology Research Center, Tabriz Branch, Islamic Azad University, Tabriz, Iran)

Received: 2023-04-04 Accepted : 2023-06-20 Published : 2023-08-23

Keywords: Poultry, Typhoid Salmonella, Multiplex polymerase chain rea,

Abstract :

The Salmonella serotypes are detected mainly through traditional microbiologic methods, which are associated with problems. The invention of rapid molecular detection methods has somewhat resolved these problems. This study aimed to assess the possibility of rapid detection of typhoid and non-typhoid Salmonellas in poultry using the multiplex polymerase chain reaction (PCR). A total of 40 isolates of Salmonella from industrial poultry were collected veterinary laboratories in Tabriz, Iran. After microbiological and serological tests, we confirmed that 27 out of 40 isolates belonged to the Salmonella entrica species. Differential tests revealed that 15, 7, 2, and 3 isolates were Salmonella Gallinarum, Salmonella Enteritidis, Salmonella Typhimurium, and Salmonella Pullorum, respectively. We then used specific primers to multiply the genes invA, rfbJ, lygD, I137_08605, and speC. The 27 isolates were then genotypically analyzed through multiplex-PCR. The results showed that all 27 isolates and the standard strains of all 4 bacteria carry the invA gene, while this gene was absent in 13 non-Salmonella isolates. The I137_08605 genes was present in all isolates and the standard strains of S. Gallinarum and S. Pullorum; the rfbJ and lygD genes were present in all isolates of S. Enteritidis and S. Typhimurium and their standard strains; and the speC gene was present in all isolates of S. Gallinarum and some isolates of S. Typhimurium and S. Entritidis and their standard strains. It seems that typhoid Salmonellas of poultry, i.e., S. Gallinarum and S. Pullorum, can be discriminated from non-typhoid Salmonellas through the multiplex-PCR molecular method. This can prevent further damage to the country's poultry industry by preventing their further spread.

References:

1.Jafari R, Fazlara A, Dalirannia A. An investigation into salmonella contamination of native hens'eggs in ahvaz. Scientific-Research Iranian Veterinary Journal. 2006;2(13):58-63.

2. Azizpour A. A survey on prevalence of Salmonella enteritidis and Salmonella typhimurium serotypes in broiler flocks of Ardabil province and determination of their antibiotics resistance to five antibacterial agents widely used in the Iranian medical field. Journal of Health. 2018;9(2):143-51.

3. Azizpour A. Prevalence and antibiotic resistance of Salmonella serotypes in chicken meat of Ardabil, Northwestern Iran. Iranian Journal of Medical Microbiology. 2021;15(2):232-46.

4. Yazdi-Amirkhiz S, Anzabi Y, Mahmazi S. Evaluation of rapid detection and investigation of the presence of spv operon virulence genes in Salmonella isolates using simplex PCR and multiplex PCR molecular methods. Veterinary Clinical Pathology The Quarterly Scientific Journal. 2020;14(55):237-50.

5. Sabeghi M, Anzabi Y. Determination of serogroup and antibiotic resistance pattern of Salmonellas isolated from commercial laying poultry of Tabriz area. Veterinary Clinical Pathology. 2019;13(50).

6. Fardous J, Shamsuzzaman S. Detection of potential pathogenic aerobic bacteria from egg shell and egg contents of hen collected from poultry. Bangladesh Medical Research Council Bulletin. 2015;41(2):67-72.

7. Onuigbo E, Iseghohimhen J, Chah K, Gyang M, Attama A. Chitosan/alginate microparticles for the oral delivery of fowl typhoid vaccine: Innate and acquired immunity. Vaccine. 2018;36(33):4973-8.

8. Xiong D, Song L, Pan Z, Jiao X. Identification and discrimination of Salmonella enterica serovar gallinarum biovars pullorum and gallinarum based on a one-step multiplex PCR assay. Frontiers in microbiology. 2018;9:1718.

9. Mezal EH, Sabol A, Khan MA, Ali N, Stefanova R, Khan AA. Isolation and molecular characterization of Salmonella enterica serovar Enteritidis from poultry house and clinical samples during 2010. Food microbiology. 2014;38:67-74.

10. Azizpour A. A Study of Salmonella Spp. Contamination Rate of Eggs and Assessment of their Antibiotic Resistance Pattern in Ardabil, Iran Qom University of Medical Sciences Journal. 2020;14(1):38-50.

11. Miranzadeh H, T Salehi Z, Karimi V. The count of aerobic mesophill bacteria and isolate salmonella Spp on egg in Isfahan1389. Veterinary Researches & Biological Products. 2012;25(1):31-5.

12. Alzwghaibi AB, Yahyaraeyat R, Fasaei BN, GhalyanchiLangeroudi A, Salehi TZ. Identification and discrimination of Salmonella Enteritidis, S. Pullorum, S. Gallinarum and S. Dublin using Salmonella specific genomic regions amplification assay. Iranian Journal of Veterinary Medicine. 2019;13(2):131-42.

13. Rocha D, Marinho A, Reis M, Borges I, Ramos F, Loureiro E. Perfil epidemiológico e caracterização molecular de Salmonella Typhi isoladas no Estado do Pará, Brasil. Revista Pan-Amazônica de Saúde. 2014;5(4):10.

14. Alzwghaibi A, Yahyaraeyat R, Fasaei BN, Langeroudi AG, Salehi TZ. Rapid molecular identification and differentiation of common Salmonella serovars isolated from poultry, domestic animals and foodstuff using multiplex PCR assay. Archives of microbiology. 2018;200(7):1009-16.

15. Quinn PJ, Markey BK, Leonard FC, Hartigan P, Fanning S, Fitzpatrick E. Veterinary microbiology and microbial disease: John Wiley & Sons; 2011.

16. Xiong D, Song L, Tao J, Zheng H, Zhou Z, Geng S, et al. An efficient multiplex PCR-based assay as a novel tool for accurate inter-serovar discrimination of Salmonella Enteritidis, S. Pullorum/Gallinarum and S. Dublin. Frontiers in microbiology. 2017;8:420.

17. Pal S, Dey S, Batabyal K, Banerjee A, Joardar SN, Isore DP. Evaluation of a Multiplex PCR Assay for Rapid Diagnosis of Fowl Typhoid. Int J Curr Microbiol App Sci. 2019;8(6):2054-8.