Applied Microbiology and Biotechnology

, Volume 63, Issue 2, pp 231–238

A biomarker for the identification of swine fecal pollution in water, using the STII toxin gene from enterotoxigenic Escherichia coli

Short Contribution

Abstract

This research developed a PCR method to identify swine fecal pollution in water, using a portion of the STII toxin gene from enterotoxigenic Escherichia coli as the target sequence. This method showed the gene to have a wide-spread geographical distribution and temporal stability; and the primers demonstrated high specificity, sensitivity, and reliability. A total of 110 DNA extracts from different animal fecal and human sewage samples were screened using the primers and no positives resulted. Centrifugation and filtration methods for concentrating E. coli seeded into stream, ocean, secondary effluent, and dairy lagoon waters resulted in detection limits at the femtogram and attogram levels. E. coli with the biomarker seeded into stream, ocean, and secondary effluent waters remained stable for approximately 2 weeks for all water types. Of the farm lagoon and waste samples tested, 94% were positive for the STII trait, regardless of the number of E. coli screened and 100% were positive when ≥35 E. coli isolates were screened. As the PCR product of the target sequence yielded a single band, the method is applicable to dot blot detection methodology, yielding great accuracy in determining the presence of swine fecal sources.

References

  1. Ackerman EO, Taylor AG (1995) Stream impact due to feedlot runoff. In: Steele K (ed) Animal waste and the land–water interface. Lewis, Baton Rouge, Fla., pp 119–125Google Scholar
  2. Akashi N, Hitotsubashi S, Yamanaka H, Fujii Y, Tsuji T, Miyama A, Joya JE, Okamoto K (1993) Production of heat-stable enterotoxin II by chicken clinical isolates of Escherichia coli. FEMS Microbiol Lett 109:311–315CrossRefPubMedGoogle Scholar
  3. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zheng Z, Miller W, Lipman DJ (1997) Gapped LAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402PubMedGoogle Scholar
  4. Bahirathan ML, Puente L, Seyfried P (1998) Use of yellow-pigmented enterococci as a specific indicator of human and nonhuman sources of faecal pollution. Can J Microbiol 44:1066–1071CrossRefPubMedGoogle Scholar
  5. Bernhard AE, Field KG (2000) Identification of nonpoint sources of fecal pollution in coastal waters by using host-specific 16S ribosomal DNA genetic markers from fecal anaerobes. Appl Environ Microbiol 66:1587–1594PubMedGoogle Scholar
  6. Chern EC (2001) Waste sources in environmental waters: determination by prevalence of cow (LTIIa), swine (STII), and human (STh) biomarkers. MS thesis, University of California, IrvineGoogle Scholar
  7. deGraaf FK, Gaastra W (1994) Fimbriae of enterotoxigenic Escherichia coli. In: Klemm P (ed) Fimbriae: adhesion, genetics, biogenesis, and vaccines. CRC Press, Boca Raton, Fla., pp 57–88Google Scholar
  8. Dufour AP, Strickland ER, Greenberg AE (1981) Membrane filter method for enumerating Escherichia coli. Appl Environ Microbiol 41:1152–1158PubMedGoogle Scholar
  9. Fairbrother JM (1992) Enteric colibacillosis. In: Leman A (ed) Diseases of swine, 7th edn. Iowa State University Press, Ames, Iowa, pp 489–497Google Scholar
  10. Grabow WOK, Neubrech TE, Holtzhausen CS, Jofre J (1995) Bacteriodes fragilis and Escherichia coli bacteriophages: excretion by humans and animals. Water Sci Technol 31:223–230CrossRefGoogle Scholar
  11. Hammermueller J, Kruth S, Prescott J, Gyles C (1995) Detection of toxin genes in Escherichia coli isolated from normal dogs and dogs with diarrhea. Can J Vet Res 59:265–270PubMedGoogle Scholar
  12. Hsu F, Shieh YS, Duin J van, Beekwilder MJ, Sobsey MD (1995) Genotyping male-specific RNA coliphages by hybridization with oligonucleotide probes. Appl Environ Microbiol 61:3960–3966Google Scholar
  13. Khatib L, Tsai YL, Olson BH (2002) A biomarker for the identification of cattle fecal pollution in water using the LTIIa toxin gene from enterotoxigenic E. coli. Appl Microbiol Biotechnol 59:97–104CrossRefPubMedGoogle Scholar
  14. Kreader C (1998) Persistence of PCR-detectable Bacteriodes distasonis from human feces in river water. Appl Environ Microbiol 64:4103–4105PubMedGoogle Scholar
  15. Lee CH, Moseley SL, Moon HW, Whipp SC, Gyles CL, So M (1983) Characterization of the gene encoding heat-stable toxin II and preliminary molecular epidemiological studies of enterotoxigenic Escherichia coli heat-stable toxin II producers. Infect Immun 42:264–268PubMedGoogle Scholar
  16. Lortie LA, Dubreuil JD, Hard J (1991) Characterization of Escherichia coli strains producing heat-labile toxin b(STb) isolated from humans with diarrhea. J Clin Microbiol 29:656–659PubMedGoogle Scholar
  17. Moon HW, Schneider RA, Moseley SL (1986) Comparative prevalence of four enterotoxin genes among Escherichia coli isolated from swine. Am J Vet Res 47:210–212PubMedGoogle Scholar
  18. Nagy F, Fekete PZ (1999) Enterotoxigenic Escherichia coli (ETEC) in farm animals. Vet Res 30:259–284PubMedGoogle Scholar
  19. Nair GB, Takeda Y (1997) The heat-labile and heat-stable enterotoxins of Escherichia coli. In: Sussman M (ed) Escherichia coli: mechanisms of virulence. Cambridge University Press, Cambridge, pp 237–256Google Scholar
  20. Okamoto K, Fujii Y, Akashi N, Hitotsubashi S, Kurazono H, Karasawa T, Takeda Y (1993) Identification and characterization of heat-stable enterotoxin II-producing Escherichia coli from patients with diarrhea. Microbiol Immunol 37:411–414PubMedGoogle Scholar
  21. Olson BH, Khatib L, McGee C (2001) Comparison of DNA finger printing methods of E. coli, genotyping male specific phage serotypes and the use of toxin genes as biomarkers to differentiate human and animal waste. Am Water Works Assoc Water Qual Technol Conf 2001:1–21Google Scholar
  22. Parveen S, Murphree RL, Edmiston L, Kaspar CW, Portier KM, Tamplin ML (1997) Association of multiple-antibiotic-resistance profiles with point and nonpoint sources of Escherichia coli in Apalachicola Bay. Appl Environ Microbiol 63:2607–2612PubMedGoogle Scholar
  23. Parveen S, Portier KM, Robinson K, Edmiston L, Tamplin ML (1999) Discriminant analysis of ribotype profiles of Escherichia coli for differentiating human and nonhuman sources of fecal pollution. Appl Environ Microbiol 65:3142–3147PubMedGoogle Scholar
  24. Pell AN (1996) Manure and microbes: public and animal health problem? J Dairy Sci 80:2673–2681Google Scholar
  25. Samadpour M (1997) Letter to regional watershed teams. King County Water Pollution Control Division, Seattle, Wash.Google Scholar
  26. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.Google Scholar
  27. Scott TM, Rose JB, Jenkins TM, Farrah JL (2002) Microbial source tracking: current methodology and future directions. Appl Environ Microbiol 68:5796–5803Google Scholar
  28. Servais P, Billen G, Rego JV (1985) Rate of bacterial mortality in aquatic environments. Appl Environ Microbiol 49:1448–1454Google Scholar
  29. Shin SJ, Chang YF, Timour M, Lauderdale TL, Lein DH (1994) Hybridization of clinical Escherichia coli isolates from calves and piglets in New York State with gene probes for enterotoxins (STaP, STb, LT), Shiga-like toxins (SLT-1, SLT-II) and adhesion factors (K88, K99, F41, 987P). Vet Microbiol 38:217–225CrossRefPubMedGoogle Scholar
  30. Stacy-Phipps S, Mecca JJ, Weiss JB (1995) Multiplex PCR assay and simple preparation method for stool specimen detection of enterotoxigenic Escherichia coli DNA during course of infection. J Clin Microbiol 33:1054–1059PubMedGoogle Scholar
  31. Takagi M, Amorin CRN, Ferreira H, Yano T (1997) Virulence related characteristics of Escherichia coli from sows with mastitis-metritis-agalactia (MMA) syndrome. Rev Microbiol 28:55–60Google Scholar
  32. Tartera C, Joffre J (1987) Bacteriophages active against Bacteriodes fragilis in sewage-polluted waters. Appl Environ Microbiol 53:1632–1637Google Scholar
  33. Whipp S, Moseley SL, Moon HW (1986) Microscopic alterations in jejunal epithelium of 3-week-old pigs induced by pig-specific, mouse negative, heat-stable Escherichia coli enterotoxin. Am J Vet Res 47:615–617PubMedGoogle Scholar
  34. Wiggins BA (1996) Discriminant analysis of antibiotic resistance patterns in fecal streptococci, a method to differentiate human and animal sources of fecal pollution in natural waters. Appl Environ Microbiol 62:3997–4002Google Scholar
  35. Woodward MJ, Kearsley R, Wray C, Roeder PL (1993) DNA probes for the detection of toxin genes in Escherichia coli isolated from diarrheal disease in cattle and pigs. Vet Microbiol 22:277–290CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  1. 1.Department of Environmental Health Science and PolicyUniversity of California at IrvineIrvineUSA

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