Cattle faecal samples (n = 480) were collected from a cluster of 12 farms, and PCR screened for the presence of the intimin gene (eae). Positive samples were cultured, and colonies were examined for the presence of eae and verocytotoxin (vtx) genes. Colonies which were positive for the intimin gene and negative for the verocytotoxin genes were further screened using PCR for a range of virulence factors including bfpA, espA, espB, tir ehxA, toxB, etpD, katP, saa, iha, lpfAO157/OI-141 and lpfAO157/OI-154. Of the 480 faecal samples, 5.8% (28/480) were PCR positive, and one isolate was obtained from each. All 28 isolates obtained were bfpA negative and therefore atypical EPEC (aEPEC). The serotypes detected included O2:H27, O8:H36, O15:H2, O49:H+, O84:H28, O105:H7 and O132:H34 but half of the isolates could not be serogrouped using currently available antisera. Twenty-two (79%) of the isolates carried the tir gene but only 25% were espB positive, and all other virulence genes tested for were scarce or absent. Several isolates showed intermediate resistance to ciprofloxacin, kanamycin, nalidixic acid, minocycline and tetracycline; full resistance to nalidixic acid or tetracycline with one isolate (O−:H8) displaying resistance to aminoglycosides (kanamycin and streptomycin), quinolones (nalidixic acid) and sulphonamides. This study provides further evidence that cattle are a potential source of aEPEC and add to the very limited data currently available on virulence genes and antibiotic resistance in this pathogenic E. coli group in animals.
Bibliographical noteReference text: Abe, C. M., L. R. Trabulsi, J. Blanco, M. Blanco, G. Dahbi, J. E. Blanco, A. Mora, M. R. Franzolin, C. R. Taddei, M. B. Martinez, R. M. Piazza, and W. P. Elias, 2009: Virulence features of atypical enteropathogenic Escherichia coli identified by the eae(+) EAF-negative stx(−) genetic profile. Diag. Microbiol. Infect. Dis. 64, 357–365.
Aidar-Ugrinovich, L., J. Blanco, M. Blanco, J. E. Blanco, L. Leomil, G. Dahbi, A. Mora, D. L. Onuma, W. D. Silveira, and A. F. P de Castro, 2007: Serotypes, virulence genes, and intimin types of Shiga toxin-producing E. coli (STEC) and enteropathogenic E. coli (EPEC) isolated from calves in Sao Paulo Brazil. Int. J. Food Microbiol. 115, 297–306.
Bieber, D., S. W. Ramer, C. Y. Wu, W. J. Murray, T. Tobe, R. Fernandez, and G. K. Schoolnik, 1998: Type IV pili, transient bacterial aggregates, and virulence of enteropathogenic E. coli. Science 280, 2114–2118.
Blanco, M., J. E. Blanco, G. Dahbi, A. Mora, M. P. Alonso, G. Varela, M. P. Gadea, F. Schelotto, E. A. Gonzalez, and J. Blanco, 2006: Typing of intimin (eae) genes from enteropathogenic E. coli (EPEC) isolated from children with diarrhoea in Montevideo, Uruguay: identification of two novel intimin variants (μB and ξR/β2B). J. Med. Microbiol. 55, 1165–1174.
Bolton, D. J., 2011: Verocytotoxigenic (Shiga Toxin-Producing) E. coli: virulence factors and pathogenicity in the farm to fork paradigm. Food. Path. Dis. 8, 357–365.
Brunder, W., H. Schmidt, and H. Karch, 1996: KatP, a novel catalase peroxidase encoded by the large plasmid of enterohaemorrhagic E. coli O157:H7. Microbiol. 142, 3305–3315.
Carneiro, L. A. M., M. C. Lins, F. R. A. Garcia, A. P. S. Silva, P. M. Mauller, B. Alves, A. C. P. Rosa, J. R. C. Andrade, A. C. Freitas-Almeida, and M. L. P. Queiroz, 2006: Phenotypic and genotypic characterisation of E. coli strains serogrouped as enteropathogenic E. coli (EPEC) isolated from pasteurised milk. Int. J. Food Microbiol. 108, 15–21.
Cleary, J., L. C. Lai, R. K. Shaw, A. Straatman-Iwanowska, S. Donnenberg, G. Frankel, and S. Knutton, 2004: Enteropathogenic E. coli (EPEC) adhesion to intestinal epithelial cells: role of bundle-forming pili (BFP), EspA filaments and intimin. Microbiology 150, 527–538.
Clinical and Laboratory Standards Institute (CLSI), 2008: Performance Standards for Antimicrobial Disc and Dilution Susceptibility Tests for Bacteria Isolated from Animals: Approved Standard M31-A3, 3rd edn. CLSA, Wayne, PA, USA.
De Verdier, K., A. Nyman, C. Greko, and B. Bengtsson, 2012: Antimicrobial resistance and virulence factors in E. coli from Swedish dairy calves. Acta Vet. Scan. 54.
Donnenberg, M. S., J. A. Giron, J. P. Nataro, and J. B. Kaper, 1992: A plasmid-encoded type IV fimbrial gene of enteropathogenic E. coli associated with localized adherence. Mol. Microbiol. 6, 3427–3437.
Dulguer, M. V., S. H. Fabbricotti, S. Y. Bando, C. A. Moreira-Filho, U. Fagundes-Neto, and I. C. A. Scaletsky, 2003: Atypical enteropathogenic E. coli strains: phenotypic and genetic profiling reveals a strong association between enteroaggregative E. coli heat-stable enterotoxin and diarrhea. J. Infect. Dis. 188, 1685–1794.
Estrada-Garcia, T., C. Lopez-Saucedo, R. Thompson-Bonilla, M. Abonce, D. Lopez-Hernandez, J. I. Santos, J. L. Rosado, H. L. DuPont, and K. Z. Long, 2009: Diarrheagenic E. coli pathotypes association with infection and diarrhea among Mexican children: atypical enteropathogenic E. coli is associated with acute diarrhea. J. Clin. Microbiol. 47, 93–98.
Giron, J. A., A. S. Ho, and G. K. Schoolnik, 1991: An inducible bundle-forming pilus of enteropathogenic Escherichia coli. Science 254, 710–713.
Gunzburg, S. T., N. G. Tornieporth, and L. W. Riley, 1995: Identification of enteropathogenic E. coli by PCR-based detection of the bundle-forming pilus gene. J. Clin. Microbiol. 33, 1375–1377.
Gyles, C. L. (1995): VT toxemia in animal models. In: Karmali, M. A., and A. G. Goglio (eds), Recent Advances in Verocytotoxin-Producing E. coli Infections, pp. 233–240. Elsevier Science, Amsterdam.
Hedberg, C., S. Savarino, J. Besser, C. Paulus, V. Thelen, L. Myers, D. N. Cameron, T. J. Barrett, J. B. Kaper, and M. T. Osterholm, 1997: An outbreak of foodborne illness caused by E. coli O39:NM, an agent not fitting into the existing scheme for classifying diarrheogenic E. coli. J. Infect. Dis. 176, 1625–1628.
Hernandez, R. T., W. P. Elias, M. A. M. Vieira, and T. A. Gomes, 2009: An overview of atypical enteropathogenic E. coli. FEMS Microbiol. Lett. 297, 137–149.
Holland, R. E., R. A. Wilson, M. S. Holland, V. Yuzbasiyan-Gurkan, T. P. Mullaney, and D. G. White, 1999: Characterization of eae+ Escherichia coli isolated from healthy and diarrheic calves. Vet. Microbiol. 66, 251–263.
Horcajo, P., G. Domínguez-Bernal, R. de la Fuente, J. A. Ruiz-Santa-Quiteria, J. E. Blanco, M. Blanco, A. Mora, G. Dahbi, C. López, B. Puentes, M. P. Alonso, and J. A. Orden, 2012: Comparison of ruminant and human attaching and effacing E. coli (AEEC) strains. Vet. Microbiol. 155, 341–348.
Hornitzky, M., K. Mercieca, K. A. Bettelheim, and S. P. Djordjevic, 2005: Bovine feces from animals with gastrointestinal infections are a source of serologically diverse atypical enteropathogenic E. coli and Shiga toxin-producing E. coli strains that commonly possess intimin. App. Environ. Microbiol. 71, 3405–3412.
Knutton, S., R. K. Shaw, R. P. Anantha, M. S. Donnenberg, and A. A. Zorgani, 1999: The type IV bundle-forming pilus of enteropathogenic E. coli undergoes dramatic alterations in structure associated with bacterial adherence, aggregation and dispersal. Mol. Microbiol. 33, 499–509.
Kobayashi, H., J. Shimada, M. Nakazawa, T. Morozumi, T. Pohjanvirta, S. Pelkonen, and K. Yamamoto, 2001: Prevalence and characteristics of Shiga toxin-producing E. coli from healthy cattle in Japan. App. Environ. Microbiol. 67, 484–489.
Machado, J., F. Grimont, and P. A. D. Grimont, 2000: Identification of E. coli flagellar types by restriction of the amplified fliC gene. Res. Microbiol. 151, 535–546.
McNally, A., A. J. Roe, S. Simpson, F. M. Thomson-Carter, D. E. E. Hoey, C. Currie, T. Chakraborty, D. G. E. Smith, and D. L. Gally, 2001: Differences in levels of secreted locus of enterocyte effacement proteins between human disease-associated and bovine E. coli O157. Infect. Immun. 69, 5107–5114.
Medina, A., P. Horcajo, S. Jurado, R. De La Fuente, J. A. Ruiz-Santa-Quiteria, G. Dominguez-Bernal, and J. A. Orden, 2011: Phenotypic and genotypic characterisation of antimicrobial resistance in enterohemorrhagic E. coli and atypical enteropathogenic E. coli strains from ruminants. J. Vet. Diag. Inv. 23, 91–95.
Moller-Stray, J., H. M. Eriksen, T. Bruheim, B. A. Kapperud, A. Skeie, M. Sunde, A. M. Urdahl, B. Oygard, and L. Vold, 2012: Two outbreaks of diarrhoea in nurseries in Norway after farm visits, April to May 2009. Euro. Surveill. 17, 947.
Monaghan, A., B. Byrne, D. A. McDowell, A. Carroll, E. McNamara, and D. J. Bolton, 2013: Serotypes and virulence profiles of atypical enteropathogenic E. coli (EPEC) isolated from bovine farms and abattoirs. J. Appl. Microbiol. 114, 595–603.
Nataro, J. P., 2006: Atypical enteropathogenis E. coli: typical pathogens? Emer. Infect. Dis. 12, 696.
Nataro, J. P., and J. B. Kaper, 1998: Diarrheagenic E. coli. Clin. Microbiol. Rev. 11, 142–201.
Orden, J. A., D. Cid, J. A. Ruiz-Santa-Quiteria, S. Garcia, S. Martinez, and R. de la Feunte, 2002: Verotoxin-producing E. coli (VTEC) enteropathogenic E. coli (EPEC) and necrotoxigenic E. coli (NTEC) isolated from healthy cattle in Spain. J. Appl. Microbiol. 93, 29–35.
Orskov, I., F. Orskov, B. Jann, and K. Jann, 1977: Serology, chemistry, and genetics of O and K antigens of E. coli. Microbiol. Mol. Biol. Rev. 41, 667–710.
Paton, A. W., and J. C. Paton, 1998: Detection and Characterization of Shiga Toxigenic E. coli by Using Multiplex PCR Assays for stx1, stx2, eaeA, Enterohemorrhagic E. coli hlyA, rfbO111, and rfbO157. J. Clin. Microbiol. 36, 598–602.
Paton, P. W., P. Srimanote, M. C. Woodrow, and J. C. Paton, 2002: Characterization of Saa, a novel autoagglutinating adhesin produced by locus of enterocyte effacement-negative Shiga-toxigenic E. coli strains that are virulent for humans. Inf. Immun. 69, 6999–7009.
Schmidt, H., B. Henkel, and H. Karch, 1997: A gene cluster closely related to type II secretion pathway operons of Gram-negative bacteria is located on the large plasmid of enterohemorrhagic in E. coli O157 strains. FEMS Microbiol. Lett. 148, 265–272.
Schmidt, H., W. L. Zhang, U. Hemmrich, S. Jelacic, W. Brunder, P. I. Tarr, U. Dobrindt, J. Hacker, and H. Karch, 2001: identification and characterization of a novel genomic island integrated at selC in locus of enterocyte effacement-negative, Shiga toxin-producing E. coli. Infect. Immun. 69, 6863–6873.
Shaw, R. K., S. Daniell, F. Ebel, G. Frankel, and S. Knutton, 2001: EspA-filament-mediated protein translocation into red blood cells. Cell. Microbiol. 3, 213–222.
Staples, M., C. J. Doyle, R. M. A. Graham, and A. V. Jennison, 2013: Molecular epidemiological typing of enteropathogenic E. coli strains from Australian patients. Diag. Microbiol. Infect. Dis. 75, 320–324.
Szalo, I. M., F. Goffaux, V. Pirson, D. Pierard, H. Ball, and J. Mainil, 2002: Presence in bovine enteropathogenic (EPEC) and enterohaemorrhagic (EHEC) E. coli of genes encoding for putative adhesins of human EHEC strains. Res. Microbiol. 153, 653–658.
Tarr, C. L., T. M. Large, C. L. Moeller, D. W. Lacher, P. I. Tarr, D. W. Acheson, and T. S. Whittam, 2002: Molecular characterization of a serotype O121:H19 clone, a distinct Shiga toxin-producing clone of pathogenic E. coli. Infect. Immun. 70, 6853–6859.
Tobe, T., and C. Sasakawa, 2002: Species-specific cell adhesion of enteropathogenic E. coli is mediated by type IV bundle forming pili. Cell. Microbiol. 4, 29–42.
Toma, C., E. Martínez Espinosa, T. Song, E. Miliwebsky, I. Chinen, S. Iyoda, M. Iwanaga, and M. Rivas, 2004: Distribution of putative adhesins in different seropathotypes of Shiga toxin-producing E. coli. J. Clinical Microbiol. 42, 4937–4946.
Trabulsi, L., R. Keller, and T. Tardelli-Gomes, 2002: Typical and atypical enteropathogenic E. coli. Emerg. Infect. Dis. 8, 508–513.
Vignoli, R., G. Varela, M. I. Mota, N. F. Cordeiro, P. Power, E. Ingold, P. Gadea, A. Sirok, F. Schelotto, J. A. Ayala, and G. Gutkind, 2005: Enteropathogenic E. coli strains carrying genes encoding the PER-2 and TEM-116 extended spectrum β-lactamases isolated from children with diarrhoea in Uruguay. J. Clin. Microbiol. 43, 2940–2943.
Viljanen, M., T. Peltola, S. Junnila, L. Olkkonen, H. Järvinen, M. Kuistila, and P. Huovinen, 1990: Outbreak of diarrhoea due E. coli O111:B4 in school-children and adults: association of Vi antigen-like reactivity. Lancet 336, 831–834.
Walk, S. T., J. M. Mladonicky, J. A. Middleton, A. J. Heidt, J. R. Cunningham, P. Bartlett, K. Sato, and T. S. Whittam, 2007: Influence of antibiotic selection on genetic composition of E. coli populations from conventional and organic dairy farms. Appl. Environ. Microbiol. 73, 5982–5989.
Walsh, C., G. Duffy, R. O'Mahony, S. Fanning, I. Blair, and D.A. McDowell, 2006: Antimicrobial resistance in Irish isolates of verocytotoxigenic E. coli (E. coli)-VTEC. Int. J. Food Microbiol. 109, 173–178.
Wells, J. G., B. R. Davis, I. K. Wachsmuth, L. W. Riley, R. S. Remis, R. Sokolow, and G. K. Morris, 1983: Laboratory investigation of hemorrhagic colitis outbreaks associated with a rare E. coli serotype. J. Clin. Microbiol. 18, 512–520.
- Enteropathogenic E. coli
- virulence genes
- antibiotic resistance