Development of three multiplex-PCR assays for virulence profiling of different iron acquisition systems in Escherichia coli
,
Abstract
Background and Objectives: Escherichia coli is responsible for various enteric and extraintestinal infections in animals and humans. Iron as an essential nutrient, has a proven role in pathogenicity of E. coli. Pathogenic E. coli benefits of having complicated systems for iron acquisition but our current knowledge is limited because of complexity of these systems. In the present study, three multiplex-PCR assays were developed to screen nine different virulence genes related to diverse iron acquisition systems in E. coli.
Materials and Methods: The multiplex-PCR systems were designed and optimized in three panels. Each panel includes a triplex-PCR cocktail. The panels are as follow: panel 1: iroN, iutA and fecA; panel 2: fyuA, sitA and irp2; and panel 3: iucD, chuA and tonB. A total of 39 pathogenic E. coli was screened according to the designed multiplex-PCR.
Results: In total, the top three frequent genes were tonB (100%), fecA (66.6%) and sitA (58.9%). With the exception of fecA and tonB, comparing the prevalence of genes among different origin of isolates (human, cattle, poultry and pigeon) showed significant associations (P < 0.05). Moreover, the iroN, sitA and iucD genes were significantly prevalent (P < 0.05) among members of extraintestinal pathogenic E. coli in comparison with the group of diarrheagenic E. coli.
Conclusion: The current multiplex-PCR assays could be a valuable, rapid and economic tool to investigate diverse iron acquisition systems in E. coli for more precise virulence typing of pathogenic or commensal strains.
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25. Holden KM, Browning GF, Noormohammadi AH, Markham PF, Marenda MS. TonB is essential for virulence in avian pathogenic Escherichia coli. Comp Immunol Microbiol Infect Dis 2012; 35: 129-138.
26. Spurbeck RR, Dinh Jr PC, Walk ST, Stapleton AE, Hooton TM, Nolan LK, et al. Escherichia coli isolates that carry vat, fyuA, chuA, and yfcV efficiently colonize the urinary tract. Infect Immun 2012; 80: 4115-4122.
27. Watts RE, Totsika M, Challinor VL, Mabbett AN, Ulett GC, De Voss JJ, et al. Contribution of siderophore systems to growth and urinary tract colonization of asymptomatic bacteriuria Escherichia coli. Infect Immun 2012; 80: 333-344.
28. Munkhdelger Y, Gunregjav N, Dorjpurev A, Juniichiro N, Sarantuya J. Detection of virulence genes, phylogenetic group and antibiotic resistance of uropathogenic Escherichia coli in Mongolia. J Infect Dev Ctries 2017; 11:51-57.
29. Searle LJ, Méric G, Porcelli I, Sheppard SK, Lucchini S. Variation in siderophore biosynthetic gene distribution and production across environmental and faecal populations of Escherichia coli. PLoS One 2015; 10(3):e0117906.
30. Torres AG, Redford P, Welch RA, Payne SM. TonB-dependent systems of uropathogenic Escherichia coli: aerobactin and heme transport and TonB are required for virulence in the mouse. Infect Immun 2001; 69: 6179-6185.
31. Tu J, Xue T, Qi K, Shao Y, Huang B, Wang X, et al. The irp2 and fyuA genes in high pathogenicity islands are involved in the pathogenesis of infections caused by avian pathogenic Escherichia coli (APEC). Pol J Vet Sci 2016; 19: 21-29.
32. Runyen-Janecky LJ, Reeves SA, Gonzales EG, Payne SM. Contribution of the Shigella flexneri Sit, Iuc, and Feo iron acquisition systems to iron acquisition in vitro and in cultured cells. Infect Immun 2003; 71: 1919-1928.
33. Miajlovic H, Aogáin M Mac, Collins CJ, Rogers TR, Smith SGJ. Characterization of Escherichia coli bloodstream isolates associated with mortality. J Med Microbiol 2016; 65: 71-79.
34. Sarowska J, Futoma-Koloch B, Jama-Kmiecik A, Frej-Madrzak M, Ksiazczyk M, Bugla-Ploskonska G, et al. Virulence factors, prevalence and potential transmission of extraintestinal pathogenic Escherichia coli isolated from different sources: recent reports. Gut Pathog 2019; 11: 10.
2. Quinn PJ, Markey BK, Leonard FC, Hartigan P, Fanning S, Fitzpatrick ES (2011). Enterobacteriaceae. In: Veterinary Microbiology and Microbial Disease. Wiley-Blackwell Publishing, 2nd ed. Chichester, West Sussex, UK, pp. 263-286.
3. Wilson BA, Salyers AA (2011). Delivery of virulence factors. In: Bacterial pathogenesis: a molecular approach. ASM Press Publishing, 3rd ed. Washington DC.
4. Caza M, Kronstad JW. Shared and distinct mechanisms of iron acquisition by bacterial and fungal pathogens of humans. Front Cell Infect Microbiol 2013; 3: 80.
5. Bullen JJ, Leigh LC, Rogers HJ. The effect of iron compounds on the virulence of Escherichia coli for guinea-pigs. Immunology 1968; 15: 581-588.
6. Braun V. Iron uptake by Escherichia coli. Front Biosci 2003; 8:s1409-1421.
7. Gao Q, Wang X, Xu H, Xu Y, Ling J, Zhang D, et al. Roles of iron acquisition systems in virulence of extraintestinal pathogenic Escherichia coli: salmochelin and aerobactin contribute more to virulence than heme in a chicken infection model. BMC Microbiol 2012; 12: 143.
8. Hajihosein-Tabrizi A, Habibi M, Tabasi M, Asadi Karam MR. Distribution of genes encoding iron uptake systems among the Escherichia coli isolates from diarrheal patients of Iran. J Med Microbiol Infect Dis 2018; 6: 25-30.
9. Rijavec M, Müller-Premru M, Zakotnik B, Žgur-Bertok D. Virulence factors and biofilm production among Escherichia coli strains causing bacteraemia of urinary tract origin. J Med Microbiol 2008; 57: 1329-1334.
10. Derakhshandeh A, Firouzi R, Motamedifar M, Arabshahi S, Novinrooz A, Boroojeni AM, et al. Virulence characteristics and antibiotic resistance patterns among various phylogenetic groups of uropathogenic Escherichia coli isolates. Jpn J Infect Dis 2015; 68: 428-431.
11. Lindstedt B-A, Finton MD, Porcellato D, Brandal LT. High frequency of hybrid Escherichia coli strains with combined Intestinal Pathogenic Escherichia coli (IPEC) and Extraintestinal Pathogenic Escherichia coli (ExPEC) virulence factors isolated from human faecal samples. BMC Infect Dis 2018; 18: 544.
12. Leimbach A, Poehlein A, Vollmers J, Görlich D, Daniel R, Dobrindt U. No evidence for a bovine mastitis Escherichia coli pathotype. BMC Genomics 2017; 18: 359.
13. van der Westhuizen WA, Bragg RR. Multiplex polymerase chain reaction for screening avian pathogenic Escherichia coli for virulence genes. Avian Pathol 2012; 41: 33-40.
14. Ghanbarpour R, Oswald E. Phylogenetic distribution of virulence genes in Escherichia coli isolated from bovine mastitis in Iran. Res Vet Sci 2010; 88: 6-10.
15. Ndlovu T, Le Roux M, Khan W, Khan S. Co-detection of virulent Escherichia coli genes in surface water sources. PLoS One 2015; 10(2): e0116808.
16. Richards VP, Lefébure T, Pavinski Bitar PD, Dogan B, Simpson KW, Schukken YH, et al. Genome based phylogeny and comparative genomic analysis of intra-mammary pathogenic Escherichia coli. PLoS One 2015; 10(7): e0133222.
17. Nataro JP, Bopp CA, Fields PI, Kaper JB, Strockbine NA (2007). Escherichia, Shigella, Salmonella. In: Manual of clinical microbiology. Ed, Murray PR, Baron EJ, Jorgensen JH, Landry ML, Pfaller MA. ASM Press Publishing, 9th ed. Washington D.C.
18. Johnson JR, Brown JJ. A novel multiply primed polymerase chain reaction assay for identification of variant papG genes encoding the Gal(α1–4)Gal-binding PapG adhesins of Escherichia coli. J Infect Dis 1996; 173: 920-926.
19. Johnson TJ, Wannemuehler Y, Doetkott C, Johnson SJ, Rosenberger SC, Nolan LK. Identification of minimal predictors of avian pathogenic Escherichia coli virulence for use as a rapid diagnostic tool. J Clin Microbiol 2008; 46: 3987-3996.
20. Johnson JR, Stell AL. Extended virulence genotypes of Escherichia coli strains from patience with urosepsis in relation to phylogeny and host compromise. J Infect Dis 2000; 181: 261-272.
21. Sabri M, Caza M, Proulx J, Lymberopoulos MH, Brée A, Moulin-Schouleur M, et al. Contribution of the SitABCD, MntH, and FeoB metal transporters to the virulence of avian pathogenic Escherichia coli O78 strain chi7122. Infect Immun 2008; 76: 601-611.
22. Ewers C, Janßen T, Kießling S, Philipp H-C, Wieler LH. Rapid detection of virulence-associated genes in avian pathogenic Escherichia coli by multiplex polymerase chain reaction. Avian Dis 2005; 49: 269-273.
23. Clermont O, Bonacorsi S, Bingen E. Rapid and simple determination of the Escherichia coli phylogenetic group. Appl Environ Microbiol 2000; 66: 4555-4558.
24. Blum SE, Goldstone RJ, Connolly JPR, Répérant-Ferter M, Germon P, Inglis NF, et al. Postgenomics characterization of an essential genetic determinant of mammary pathogenic Escherichia coli. mBio 2018; 9:e00423-18.
25. Holden KM, Browning GF, Noormohammadi AH, Markham PF, Marenda MS. TonB is essential for virulence in avian pathogenic Escherichia coli. Comp Immunol Microbiol Infect Dis 2012; 35: 129-138.
26. Spurbeck RR, Dinh Jr PC, Walk ST, Stapleton AE, Hooton TM, Nolan LK, et al. Escherichia coli isolates that carry vat, fyuA, chuA, and yfcV efficiently colonize the urinary tract. Infect Immun 2012; 80: 4115-4122.
27. Watts RE, Totsika M, Challinor VL, Mabbett AN, Ulett GC, De Voss JJ, et al. Contribution of siderophore systems to growth and urinary tract colonization of asymptomatic bacteriuria Escherichia coli. Infect Immun 2012; 80: 333-344.
28. Munkhdelger Y, Gunregjav N, Dorjpurev A, Juniichiro N, Sarantuya J. Detection of virulence genes, phylogenetic group and antibiotic resistance of uropathogenic Escherichia coli in Mongolia. J Infect Dev Ctries 2017; 11:51-57.
29. Searle LJ, Méric G, Porcelli I, Sheppard SK, Lucchini S. Variation in siderophore biosynthetic gene distribution and production across environmental and faecal populations of Escherichia coli. PLoS One 2015; 10(3):e0117906.
30. Torres AG, Redford P, Welch RA, Payne SM. TonB-dependent systems of uropathogenic Escherichia coli: aerobactin and heme transport and TonB are required for virulence in the mouse. Infect Immun 2001; 69: 6179-6185.
31. Tu J, Xue T, Qi K, Shao Y, Huang B, Wang X, et al. The irp2 and fyuA genes in high pathogenicity islands are involved in the pathogenesis of infections caused by avian pathogenic Escherichia coli (APEC). Pol J Vet Sci 2016; 19: 21-29.
32. Runyen-Janecky LJ, Reeves SA, Gonzales EG, Payne SM. Contribution of the Shigella flexneri Sit, Iuc, and Feo iron acquisition systems to iron acquisition in vitro and in cultured cells. Infect Immun 2003; 71: 1919-1928.
33. Miajlovic H, Aogáin M Mac, Collins CJ, Rogers TR, Smith SGJ. Characterization of Escherichia coli bloodstream isolates associated with mortality. J Med Microbiol 2016; 65: 71-79.
34. Sarowska J, Futoma-Koloch B, Jama-Kmiecik A, Frej-Madrzak M, Ksiazczyk M, Bugla-Ploskonska G, et al. Virulence factors, prevalence and potential transmission of extraintestinal pathogenic Escherichia coli isolated from different sources: recent reports. Gut Pathog 2019; 11: 10.
| Files | ||
| Issue | Vol 12 No 4 (2020) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v12i4.3930 | |
| Keywords | ||
| Escherichia coli; Iron; Virulence genes; Typing; Multiplex-polymerase chain reaction | ||
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How to Cite
1.
Kalateh Rahmani H, Hashemi Tabar G, Askari Badouei M, Khoramian B. Development of three multiplex-PCR assays for virulence profiling of different iron acquisition systems in Escherichia coli. Iran J Microbiol. 2020;12(4):281-288.
