AiM  Vol.5 No.9 , August 2015
The Impact of Enterohemorrhagic Escherichia coli (EHEC) on Ciliate Protozoan Populations in Municipal Sewage
Abstract: Enterohemorrhagic Escherichia coli strains (EHEC) have caused many foodborne outbreaks. Bacterivorous protozoa could remove bacteria from aquatic systems. We analyzed the ciliate protozoan population changes influenced by EHEC co-culture in activated sludge. EHEC and non-EHEC control E. coli cells were added to activated sludge samples in microcosms. The ciliate population changes were monitored by terminal restriction fragment length polymorphism (T-RFLP) analysis. EHEC and non-EHEC fed ciliate protozoan populations were different from each other and the no bacteria added controls based on the additive main effects and multiplicative interaction model (AMMI) analysis. Ciliate species were identified by 18S rDNA clone libraries. The 18S rDNA clones from the original sludge sample were identified as Epistylis wenrichi (70%) and Prorodon teres (30%), while clones from EHEC treated sludge sample were identified as P. teres (52%), Vorticella fusca (41%), Dexitrichides pangi (5%), and Opisthonecta henneguyi (2%). This study could provide helpful information about ciliate protozoan population changes caused by different E. coli strains in wastewater treatment plants, which could be useful for preventing and tracking E. coli outbreaks.
Cite this paper: Li, Z. , Sheridan, P. and Shields, M. (2015) The Impact of Enterohemorrhagic Escherichia coli (EHEC) on Ciliate Protozoan Populations in Municipal Sewage. Advances in Microbiology, 5, 668-676. doi: 10.4236/aim.2015.59069.

[1]   Slayton, R.B., Turabelidze, G., Bennett, S.D., Schwensohn, C.A., Yaffee, A.Q., et al. (2013) Outbreak of Shiga Toxin-Producing Escherichia coli (STEC) O157:H7 Associated with Romaine Lettuce Consumption, 2011. PLoS ONE, 8, e55300.

[2]   Joensen, K.G., Scheutz, F., Lund, O., Hasman, H., Kaas, R.S., Nielsen, E.M. and Aarestrup, F.M. (2014) Real-Time Whole-Genome Sequencing for Routine Typing, Surveillance, and Outbreak Detection of Verotoxigenic Escherichia coli. Journal of Clinical Microbiology, 52, 1501-1510.

[3]   Pennington, T.H. (2014) E. coli O157 Outbreaks in the United Kingdom: Past, Present, and Future. Infection and Drug Resistance, 7, 211-222.

[4]   Riley, L.W., Remis, R.S., Helgerson. S.D., McGee, H.B., Wells, J.G. and Davis, B.R. (1983) Hemorrhagic Colitis Associated with a Rare Escherichia coli Serotype. New England Journal of Medicine, 308, 681-685.

[5]   O’Brien, A.O., Lively, T.A., Chen, M.E., Rothman, R.W. and Formal, S.B. (1983) Escherichia coli O157:H7 Strains Associated with Haemorrhagic Colitis in the United States Produce a Shigella dysenteriae 1 (Shiga) Like Cytotoxin. Lancet, 321, 702.

[6]   Newland, J.W., Strockbine, N.A., Miller, S.F., O’Brien, A.D. and Holmes, R.K. (1985) Cloning of Shiga-Like Toxin Structural Genes from a Toxin Converting Phage of Escherichia coli. Science, 230, 179-181.

[7]   Steinberg, K.M. and Levin, B.R. (2007) Grazing Protozoa and the Evolution of the Escherichia coli O157:H7 Shigatoxin-Encoding Prophage. Proceedings of the Royal Society B, 274, 1921-1929.

[8]   Lainhart, W., Stolfa, G. and Koudelka, G.B. (2009) Shiga Toxin as a Bacterial Defense against a Eukaryotic Predator, Tetrahymena thermophila. Journal of Bacteriology, 191, 5116-5122.

[9]   Lee, S.H., Levy, D.A., Craun, G.F., Beach, M.J. and Calderon, R.L. (2002) Surveillance for Waterborne-Disease Outbreaks—United States, 1999-2000. MMWR Surveillance Summaries, 51, 1-47.

[10]   Ravva, S.V., Sarreal, C.Z., Duffy, B. and Stanker, L.H. (2006) Survival of Escherichia coli O157:H7 in Wastewater from Dairy Lagoons. Journal of Applied Microbiology, 101, 891-902.

[11]   Solomon, E.B., Yaron, S. and Matthews, K.R. (2002) Transmission of Escherichia coli O157:H7 from Contaminated Manure and Irrigation Water to Lettuce Plant Tissue and Its Subsequent Internalization. Applied and Environmental Microbiology, 68, 397-400.

[12]   Bach, S.J., McAllister, T.A., Veira, D.M., Gannon, V.P.J. and Holley, R.A. (2002) Transmission and Control of Escherichia coli 157:H7—A Review. Canadian Journal of Animal Science, 82, 475-490.

[13]   Ravva, S.V., Sarreal, C.Z. and Mandrell, R.E. (2010) Identification of Protozoa in Dairy Lagoon Wastewater That Consume Escherichia coli O157:H7 Preferentially. PLoS ONE, 5, e15671.

[14]   Gantalupo, P.G., Calgua, B., Zhao, G., Hundesa, A., Wier, A.D., Katz, J.P., Grabe, M., Hendrix, R.W., Girones, R., Wang, D. and Pipas, J.M. (2011) Raw Sewage Harbors Diverse Viral Populations. mBio, 2, Article ID: e00180-11.

[15]   Elmund, G.K., Allen, M.J. and Rice, E.W. (1999) Comparison of Escherichia coli, Total Coliform, and Fecal Coliform Populations as Indicators of Wastewater Treatment Efficiency. Water Environment Research, 71, 332-339.

[16]   Grant, S.B., Pendroy, C.P., Mayer, C.L., Bellin, J.K. and Palmer, C.J. (1996) Prevalence of Enterohemorrhagic Escherichia coli in Raw and Treated Municipal Sewage. Applied and Environmental Microbiology, 62, 3466-3469.

[17]   Martinez-Castillo, A., Allue-Guardia, A., Dahbi, G., Blanco, J., Creuzburg, K., Schmidt, H. and Muniesa, M. (2011) Type III Effector Genes and Other Virulence Factors of Shiga toxin-Encoding Escherichia coli Isolated from Wastewater. Environmental Microbiology Reports, 4, 147-155.

[18]   Muniesa, M., Serra-Moreno, R. and Jofre, J. (2004) Free Shiga toxin Bacteriophages Isolated from Sewage Showed Diversity although the stx Genes Appeared Conserved. Environmental Microbiology, 6, 716-725.

[19]   Hahn, M.W. and Hofle, M.G. (2001) Grazing of Protozoa and Its Effect on Populations of Aquatic Bacteria. FEMS Microbiology Ecology, 35, 113-121.

[20]   Curds, C.R. (1982) The Ecology and Role of Protozoa in Aerobic Sewage Treatment Processes. Annual Review of Microbiology, 36, 27-46.

[21]   Snaidr, J., Amann, R., Huber, I., Ludwig, W. and Schleifer, K. (1997) Phylogenetic Analysis and in Situ Identification of Bacteria in Activated Sludge. Applied and Environmental Microbiology, 63, 2884-2896.

[22]   Zhou, K., Xu, M., Liu, B. and Cao, H. (2008) Characteristics of Microfauna and Their Relationships with the Performance of an Activated Sludge Plant in China. Journal of Environmental Sciences, 20, 482-486.

[23]   Madoni, P. (1994) A Sludge Biotic Index (SBI) for the Evaluation of the Biological Performance of Activated Sludge Plants Based on the Microfauna Analysis. Water Research, 28, 67-75.

[24]   Fitzpatrick, A.H. (2010) Detection, Quantification, and Sequencing of stx1 and stx2 in Southeastern Idaho Wastewater Treatment Facilities. Master’s Thesis, Idaho State University, Pocatello.

[25]   King, W.L. and Hurst, A. (1963) A Note on the Survival of Some Bacteria in Different Diluents. Journal of Applied Bacteriology, 26, 504-506.

[26]   Reynolds, J. (2005) Serial Dilution Protocols. ASM Microbe Library.

[27]   Li, Z., Haynes, R., Sato, E., Shields, M.S., Fujita, Y. and Sato, C. (2014) Microbial Community Analysis of a Single Chamber Microbial Fuel Cell Using Potato Wastewater. Water Environment Research, 86, 324-330.

[28]   Dopheide, A., Lear, G., Stott, R. and Lewis, G. (2008) Molecular Characterization of Ciliate Diversity in Stream Biofilms. Applied and Environmental Microbiology, 74, 1740-1747.

[29]   Birnboim, C. and Doly, J. (1979) A Rapid Alkaline Extraction Procedure for Screening Recombinant Plasmid DNA. Nucleic Acids Research, 7, 1513-1523.

[30]   Li, Z., Briggs, B.R., Sheridan, P.P. and Shields, M.S. (2013) An Anion-Exchange Method to Concentrate Dissolved DNA from Aquifer Water. Journal of Microbiological Methods, 93, 1-8.

[31]   Martin, D.P., Lemey, P., Lott, M., Moulton, V., Posada, D. and Lefeuvre, P. (2010) RDP3: A Flexible and Fast Computer Program for Analyzing Recombination. Bioinformatics, 26, 2462-2463.

[32]   Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M. and Kumar, S. (2011) MEGA5: Molecular Evolutionary Genetics Analysis Using Maximum Likelihood, Evolutionary Distance, and Maximum Parsimony Methods. Molecular Biology and Evolution, 28, 2731-2739.

[33]   Liu, W.T., Marsh, T.L., Cheng, H. and Forney, L.J. (1997) Characterization of Microbial Diversity by Determining Terminal Restriction Fragment Length Polymorphisms of Genes Encoding 16S rRNA. Applied and Environmental Microbiology, 63, 4516-4522.

[34]   Culman, S.W., Bukowski, R., Gauch, H.G., Cadillo-Quiroz, H. and Buckley, D.H. (2009) T-REX: Software for the Processing and Analysis of T-RFLP Data. BMC Bioinformatics, 10, 171.

[35]   Culman, S.W., Gauch, H.G., Blackwood, C.B. and Thies, J.E. (2008) Analysis of T-RFLP Data Using Analysis of Variance and Ordination Methods: A Comparative Study. Journal of Microbiological Methods, 75, 55-63.

[36]   Soper, D.S. (2015) Analysis of Variance (ANOVA) Calculator—One-Way ANOVA from Summary Data.