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Evaluating the Presence of Pathogenic Bacteria in Fecal Samples of Feral Pigs and Their Transport to Surface Water Dave Bachoon, Jyoti Lama, Brennan Pookwong,

Biology & Environmental Sciences, Georgia College & State University, Georgia

Abstract Methods

Results

Acknowledgements

References

•  The primary goal for this project was to improve understanding of pathogenic bacteria harbored by feral pigs and their impact on water quality. Fecal samples of feral pigs from five counties in Georgia were collected and the DNA was extracted. qPCR screening of fecal samples for the presence of Brucella suis, Campylobacter jejuni, and E. coli O157:H7 indicated that B. suis was present in 25% of the feral pigs and E. coli O157:H7 was present in 5.5% of the pigs but Campylobacter jejuni was not detected in any of the feral pig fecal samples. The majority of majority of pigs carrying zoonotic pathogens came from Morgan County. Phylotyping analysis of E. coli isolates from the feral pigs indicated that these pigs carry a high proportion of virulent strains of E. coli (phylogroups B2 and D) compared to cattle and farm raised pigs. DNA samples are being processed for metagenomic analysis of Brucella and Escherichia groups. Water samples are being collected for analysis of pathogenic bacteria and the presence of fecal bacteria from feral pigs. This project gives Georgia College, a small university steadily building its research portfolio, a valuable opportunity for expanding the institution’s research capabilities and funding potential.

Results

dave.bachoon@gcsu.edu

•  Figure 2. Feral pigs on farmlands. Photos provided by M. Ondovchik USDA-APHIS, Athens Ga.

•  This project is funded through The National Institute of Food and Agriculture (NIFA), USDA. Sustaining water quantity, quality, and availability for agricultural use while maintaining environmental quality through 2050. (2015-68007-23137).

•  We appreciate the collaboration of Dr. Marirosa Molina at USEPA in Athens, GA.

•  We appreciate the assistance of Rob Pinkston of Yager Pro and USDA-APHIS in Athens for help with sample.

Conclusion

Introduction

Specific objectives: Summer sabbatical 1. To evaluate the presence of pathogenic bacteria in fecal samples from feral pigs on cattle farms in relation BMP’s and transport to surface waters. 2. Develop my expertise in metagenomic analysis of bacterial population of fecal samples from feral pigs.

Trapped feral pigs

Figure 3. Water and fecal sampling sites in areas where feral pigs are active. BC, Beef cattle, DC, dairy cattle, NC, no cattle.

Table 1. QPCR and MST primers used

Grant number: 2015-68007-23137

Figure 3. Dichotomous decision tree to determine the phylogenetic group of an E. coli strain (Clermont et al., 2000).

Dick, L. K., and Field, K. G. (2004). Rapid Estimation of Numbers of Fecal Bacteroidetes by Use of a Quantitative PCR Assay for 16S rRNA Genes. Applied and Environmental Microbiology 70(9), 5695-5697. Guion, Chase E., Theresa J. Ochoa, Christopher M. Walker, Francesca Barletta, and Thomas G. Cleary. "Detection of diarrheagenic Escherichia coli by use of melting-curve analysis and real-time multiplex PCR." Journal of clinical microbiology 46, no. 5 (2008): 1752-1757. Haugland, R.A., Varma, M., Sivaganesan, M., Kelty, C., Peed, L., and Shanks, O.C., (2010). Evaluation of genetic markers from the 16S rRNA V2 region for use in quantitative detection of selected Bacteroidales species in human fecal waste by qPCR. Systematic and Applied Microbiology, 22, 348-357 B.J. Kildare, C.M. Leutenegger, B.S. McSwain, D.G. Bambic, V.B. Rajal and S. Wuertz, 16S r RNA-based assays for quantitative detection of universal, human-, cow-, and dog-specific fecal Bacteroidales: a Bayesian approach, Water Res. 41 (16) (2007), pp. 3701–3715 Mieszkin, S., Furet, J. P., Corthier, G., and Gourmelon, M. (2009). Estimation of Pig Fecal Contamination in a River Catchment by Real-Time PCR Using Two Pig-Specific Bacteroidales 16S rRNA Genetic Markers. Applied and Environmental Microbiology 75(10), 3045-3054.

•  qPCR screening of fecal samples for the presence of Brucella suis, Campylobacter jejuni, and E. coli O157:H7 indicated that B. suis was present in 25% of the feral pigs and E. coli O157:H7 was present in 5.5% of the pigs.

•  Campylobacter jejuni was not detected in any of the feral pig fecal samples.

•  Phylotyping analysis of E. coli isolates from the feral pigs indicated that these pigs carry a high proportion of virulent strains of E. coli.

•  With the increasing numbers of feral pigs in the country, more studies are warranted on the diversity of potential pathogenic microbes carried by feral pigs.

Figure 1—In 1998, feral swine could be found in parts of all the counties shown here in green. By 2004, wild pigs had increased their range to counties shown in yellow, while remaining in the green-colored areas as well. Note that wild pigs are now found in two counties in Iowa—America’s #1 swine-producing State. (Map adapted from originals created by the Southeastern Cooperative Wildlife Disease Study, Athens, GA.)