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Molecular Detection of Verotoxigenic Escherichia coli (VTEC) from Animal Feces for Screening VTEC-shedders
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 Title & Authors
Molecular Detection of Verotoxigenic Escherichia coli (VTEC) from Animal Feces for Screening VTEC-shedders
Kobayashi, Y.; Sato, M.; Taguchi, H.; Koike, S.; Nakatsuji, H.; Tanaka, K.;
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Seventy-six animals including cattle, sheep, horses, 6 species of zoo animals were employed for collection of fresh feces in order to detect verotoxigenic Esherichia coli (VTEC) by safe, quick and sensitive PCR-based molecular methods. Bacterial cell disruption with bead-beating followed by bacterial DNA purification with hydroxyapatide chromatography and gel filtration allowed DNA preparation from animal feces with high recovery and purity. A mountain goat was firstly shown by PCR and sequencing to shed verotoxin 2 gene (vt2) that was used to generate vt2 probe and second primer set for nested PCR to attempt more sensitive detection. Most sensitive nested PCR revealed that 45% of tested cattle and 47% of tested zoo animals were VTEC-positive, while least sensitive normal PCR detected VTEC from none of these animals except a mountain goat. Moderately sensitive detection by PCR in combination with hybridization suggested that the VTEC density varied between the VTEC-positive cattle.
Verotoxin;E. coli;Cattle;Mountain Goat;PCR;Hybridization;
 Cited by
Year-round Monitoring of Verotoxin-producing Escherichia coli from Feces of Dairy Cattle,Kobayashi, Y.;El-Sawy, H.B.I.;

Asian-Australasian Journal of Animal Sciences, 2007. vol.20. 5, pp.789-794 crossref(new window)
Asakura, H., S. Makino, T. Shirahata, T. Tsukamoto, H. Kurazono, T. Ikeda and K. Takeshi. 1998. Detection and genetical characterization of Shiga toxin-producing Escherichia coli from wild deer. Microbiol Immunol. 42:815-22.

Diez-Gonzalez, F., T. R. Callaway, M. G. Kizoulis and J. B. Russell. 1998. Grain feeding and the dissemination of acidresistant Escherichia coli from cattle. Science 281:1666-1668.

Gilgen, M., P. Hubner, C. Hofelein, J. Luthy and U. Candrian. 1998. PCR-based detection of verotoxin-producing Escherichia coli (VTEC) in ground beef. Res. Microbiol. 149:145-154.

Hovde, C. J., P. R. Austin, K. A. Cloud, C. J. Williams and C. W. Hunt. 1999. Effect of cattle diet on Escherichia coli O157:H7 acid resistance. Appl. Environ. Microbiol. 65:3233-3235.

Johnson, R. P., J. B. Wilson, P. Michel, K. Rahn, S. A. Renwick, C. L. Gyles and J. S. Spika. 1999. Human infection with verocytotoxigenic Escherichia coli associated with exposure to farms and rural environments. In: Escherichia coli O157 in farm animals. (Ed. C. S. Stewart and H. J. Flint). CABI Publishing, Willingford, UK. pp. 147-168.

Miyao, Y., Y. Somura, T. Suzuki, A. Kai, T. Itoh, K. Hirayama and K. Itoh. 1996. Isolation of verocytotoxin-producing Escherichia coli from rectum and cecum of healthy cattle. J. Jpn. Vet. Med. Assoc. 49:46-51.

Paton, A. W., E. Voss, P. A. Manning and J. C. Paton. 1997. Shiga toxin-producing Escherichia coli isolates from cases of human disease show enhanced adherence to intestinal epithelial (Henle 407) cells. Infect. Immun. 65:3799-3805.

Purdy, K. J., T. M. Embley, S. Takii and D. B. Nedwell. 1996. Rapid extraction of DNA and rRNA from sediments by a novel hydroxyapatite spin-column method. Appl. Environ. Microbiol. 62:3905-3907.

Rasmussen, M. A., W. C. Cray, Jr., T. A. Casey and S. C. Whipp. 1993. Rumen contents as a reservoir of enterohemorrhagic Escherichia coli. FEMS Mirobiol. Lett. 114:79-84.

Sanz, M. E., M. R. Vinas and A. E. Parma. 1998. Prevalence of bovine verotoxin-producing Escherichia coli in Argentina. Eur. J. Epidemiol. 14:399-403.

Sambrook, J. and D. W. Russell. 2001. Molecular cloning: a laboratory manual, 3rd Ed. Cold Springs Harbor, New York.

Schmidt, H., M. Montag, J. Brockemuhl, J. Heeseman and H. Karch. 1993. Shiga like toxin II related cytotoxins in Citrobacter freundii strains from humans and beef samples. Infect. Immun. 61:534-543.

Stahl, D. A., B. A. Flesher, H. R. Mansfield and L. Montgomery. 1988. Use of phylogenetically based hybridization probes for studies of ruminal microbial ecology. Appl. Environ. Microbiol. 54:1079-1084.

Stewart, D. S., M. L. Tortorello and S. M. Gendel. 1998. Evaluation of DNA preparation techniques for detection of the SLT-1 gene of Escherichia coli O157:H7 in bovine feces using the polymerase chain reaction. Lett. Appl. Microbiol. 26:93-97.

2001. Occurrence of verotoxin-producing Escherichia coli in dairy heifers grazing an irrigated pasture. Toxicol. 159:159-69.

Todd, E. C., R. A. Szabo, J. M. MacKenzie, A. Martin, K. Rahn, C. Gyles, A. Gao, D. Alves and A. J. Yee. 1999. Application of a DNA hybridization-hydrophobic-grid membrane filter method for detection and isolation of verotoxigenic Escherichia coli. Appl. Environ. Microbiol. 65:4775-80.

Tsen, H. Y., C. K. Lin and W. R. Chi. 1998. Development and use of 16S rRNA gene targeted PCR primers for the identification of Escherichia coli cells in water. J. Appl. Microbiol. 85:554-560.