Progress 09/15/02 to 09/14/06
Outputs
Sample collection from 56 of 60 proposed farm-slaughter pairs are complete. Four farms dropped out of the study due to various reasons. Completed ones include from all sites in NC, 16 sites from Ohio & 20 from Wisconsin. Questionnaires are being coded & analyzed. Phenotypic & genotypic characterization of three important foodborne pathogens: Salmonella, Campylobacter & Yersinia enterocolitica, was done. Salmonella prevalence & antimicrobial resistance: Pigs from ABF farms had increased odds of Salmonella detection in farm collected fecal samples compared with conventional farms, (OR 2.4, CI 1.5-3.7, p<0.01). No differences were found for carcass swabs. Proportion of Salmonella-positive fecal samples among ABF farms was 27.3% (CI 12.8-41.8%) and 12.3% (CI 5.0-19.5%) for conventional farms. For carcass swabs, proportion of Salmonella-positive samples was as follows across both farm types: A, 9.2% (CI 3.1-15.3%), B, 9.9% (CI 5.5-14.3%), C, 3.9% (CI 0-8.0%), D, 5.0% (CI
1.7-8.3%). No differences in carcass swab positive status detected between the farm categories. Antimicrobial resistance did not significantly differ between isolates from the two farm types for any antimicrobial tested. Proportion of resistance detected was as follows, in declining order: Tetracycline, 65.1% (CI 38.3-91.9%), streptomycin, 23.5% (CI 6.5-40.5%), amoxicillin/clavulanic acid 22.8% (CI 5.6-40.1%), ampicillin 22.1% (CI5.4-38.9%), chloramphenicol 16.8% (CI 0.0- 34.0%), sulfafizoxazole 4.0% (CI 0.0-9.0%), cefoxitin 4.0% (CI 0.0-11.5%) trimethoprim/sulphamethoxazole 1.3% (CI 0.0-3.1%) and kanamycin 1.0% (CI 0.0-3.5%). No resistant isolates were detected for amikacin, ceftriaxone, ciprofloxacin, gentamicin, nalidixic acid or ceftiofur. Overall comparison of Campylobacter from ABF and conventional herds: Campylobacter coli was predominant species, with 1,459 isolates (99%). Found significantly higher prevalence of C. coli on ABF farms (77.3%) than on conventional farms (27.6%)
among pigs at the nursery stage (P < 0.001). At slaughter, we found significantly higher prevalence at the post-evisceration than at the pre-evisceration stage (P < 0.001) in both production systems. The 1,459 C. coli isolates were tested with the agar dilution method for their susceptibility to 6 antimicrobials: chloramphenicol, ciprofloxacin, erythromycin, gentamicin, nalidixic acid, & tetracycline. Resistance was most prevalent against tetracycline (66.2%) followed by erythromycin (53.6%). Resistance to ciprofloxacin at the MIC (> 4 mg/liter) was also found on farms in both systems. Yersinia: For pig samples, 10.9% of fecal samples overall were found to be Yersinia enterocolitica positive. Of these fecal source Y. enterocolitica, 36.9% overall were identified to be harboring the ail gene. NC had the lowest prevalence with 2.5%, while Ohio & Wisconsin had similar Y. enterocolitica prevalence. 21.5 % of Ohio Y. enterocolitica were ail positive. Wisconsin had highest proportion of
isolates as ail positive with 52.6% of Y. enterocolitica isolates harboring the ail gene. Isolation of Y. enterocolitica from carcass samples was rare. All positive carcass samples originated from either Wisconsin or Ohio.
Impacts
This study compared the prevalence of antimicrobial resistant pathogens from conventional and the antimicrobial-free herds at farm and slaughter stages. It is focused on prevalence, antimicrobial resistance and genotyping. This study can be used as a background information to what the role and impact of antimicrobial use in food animals and its association with antimicrobial resistance. The study resulted in several publications that were published in national and international journals and scientific meetings. Among some of the major points that were found and have significant impact on food animal industry and antimicrobial resistance issues include: 1) The high prevalence of Salmonella among ABF herds was unexpected. However, it alerted the swine producers and the public alike to be conscious of the situation and not all antimicrobially-free reared products are full proof safe; 2) Antimicrobial resistant pathogens were commonly isolated from the conventional as well
as ABF systems (including pentaresistant Salmonella R-type ACSSuT). This also warrants concern and points to the possible role of other environmental factors in transmission of these strains in pigs; 3) Multi drug resistant ciprofloxacin resistant Campylobacter coli isolates from swine especially from the ABF reared pigs is alarming since this antimicrobial is used in treatment of severe invasive cases of campylobacteriosis.
Publications
- No publications reported this period
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Progress 01/01/05 to 12/31/05
Outputs
Sample collection from 56 of 60 proposed farm-slaughter pairs are complete. While collection & prevalence is more or less completed, further phenotypic & genotypic characterization is still underway. Salmonella prevalence & antimicrobial resistance - Pigs from AMF farms had increased odds of Salmonella detection in farm collected fecal samples compared with AMU farms, (OR 2.4, CI 1.5-3.7, p<0.01). No differences were found for carcass swabs. The proportion of Salmonella-positive fecal samples among AMF farms was 27.3% (CI 12.8-41.8%) and 12.3% (CI 5.0-19.5%) for AMU farms. For carcass swabs, the proportion of Salmonella-positive samples was as follows across both farm types: A, 9.2% (CI 3.1-15.3%), B, 9.9% (CI 5.5-14.3%), C, 3.9% (CI 0-8.0%), D, 5.0% (CI 1.7-8.3%). Salmonella in NC: Salmonella prevalence was significantly higher among the antimicrobial-free systems (15.2%) than the conventional systems (4.2%) (odds ratio [OR] 5 4.23; P , 0.05). Frequency of resistance
to most classes of antimicrobials (except tetracycline) was significantly higher among conventional farms than antimicrobial-free farms, with ORs ranging from 2.84 for chloramphenicol to 23.22 for kanamycin at the on-farm level. Campylobacter prevalence and antimicrobial resistance - Overall comparison of ABF & conventional herds: Campylobacter coli was the predominant species, with 1,459 isolates (99%) in the study. Found significantly higher prevalence of C. coli on the ABF farms (77.3%) than on the conventional farms (27.6%) among pigs at the nursery stage (P < 0.001). At slaughter, found significantly higher prevalence at the post-evisceration than at the pre-evisceration stage (P < 0.001) in both production systems. Campylobacter: antimicrobial resistance mechanisms and molecular epidemiology - We compared the use of two genotyping methods for molecular epidemiology purposes, and to determine the genotypic diversity of Campylobacter coli of porcine origin. Evaluation of the
methods was based on their resistance patterns, discriminatory indexes (DI), high test throughputs, costs, and turnaround times. Resistance to erythromycin and tetracycline was the most common. Yersinia prevalence. - Feces: 10.9% of fecal samples overall were found to be Yersinia enterocolitica positive. NC had the lowest prevalence with 2.5% (21.4% of which were ail positive) of samples positive, while Ohio & Wisconsin had similar Y. enterocolitica prevalence (16.8% and 16.1% respectively). Interestingly, 21.5 % of Ohio Y. enterocolitica were ail positive, similar to NC samples. Wisconsin had highest proportion of isolates as ail positive with 52.6% of Y. enterocolitica isolates harboring the ail gene. Carcass samples: Isolation of Y. enterocolitica from carcass samples was rare. Only 4 pre-evisceration carcass samples were positive, & 1 pre-evisceration isolate (25%) was identified to harbor the ail gene. 2 carcasses were identified as Y. enterocolitica positive post-evisceration, &
one of these isolates was ail positive. Y. enterocolitica was isolated from only 6 carcasses post-chill, and none of these isolates were ail positive. All positive carcass samples originated from either Wisconsin or Ohio.
Impacts
Study is among the very few in U.S. comparing the prevalence of antimicrobial resistant C.coli from conventional & the antimicrobial-free herds at farm & slaughter stages. Focused on prevalence, antimicrobial resistance & genotyping. Most of work regarding Campylobacter is complete & majority of work on Salmonella & Yersinia has also been completed. Antimicrobial resistance is a major problem in the food safety & public health arena. Study can be used as a background information to what the role & impact of antimicrobial use in food animals & its association with antimicrobial resistance. Some of the major points that were found and have significant impact on food animal industry and antimicrobial resistance issues include: 1) The high prevalence of Salmonella among ABF herds as compared to conventional ones. This was done at univariate level and when multivariate models (including various factors not yet considered from the questionnaire) are conducted, the results
may be modified. 2) Even though we detected higher number of resistant and multi drug resistant strains in isolates from the conventional system, antimicrobial resistant pathogens commonly isolated from the ABF system (including pentaresistant Salmonella R-type ACSSuT) warrants concern and points to the possible role of other environmental factors in transmission of these strains in pigs. 3)Multi drug resistant ciprofloxacin resistant Campylobacter coli isolates from swine especially from the ABF reared pigs is alarming since this antimicrobial is used in treatment of severe invasive cases of campylobacteriosis.
Publications
- Gebreyes WA, S. Thakur and W.E. Morrow. 2006. Comparison of Prevalence, Antimicrobial Resistance and Occurrence of Multi-drug resistant Salmonella in Antimicrobial-free and Conventional Pig Production Systems. Journal of Food Protection (In press).
- Thakur S. and W.A. Gebreyes. 2005. Prevalence and Antimicrobial Resistance of Campylobacter in Antimicrobial-Free and Conventional Production Systems. Journal of Food Protection 68(11):2402-10.
- S. Thakur and W.A. Gebreyes. 2005. Campylobacter coli in Swine Production: Antimicrobial Resistance Mechanisms and Molecular Epidemiology. J Clin Microbiol. 43(11):5705-14.
- Gebreyes, WA, S. Thakur and W.E. M. Morrow. 2005. Campylobacter coli: Prevalence and antimicrobial resistance in Antimicrobial-free Swine Production Systems. J. Antimicrob Chemother 56(4):765-8.
- Thakur, S. and Gebreyes, W.A. 2005. Prevalence, antimicrobial resistance and genotypic diversity of Campylobacter coli isolated from antimicrobial-free swine production systems. Safe Pork: 6th International Symposium on Epidemiology and Control of Food-borne pathogens in Pork.Sept.6-9, Rohnert Park, California, USA.
- Thakur, S. and Gebreyes, W.A. 2005. Comparative genotyping of Campylobacter coli using Multi Locus Sequence Typing and Pulsed Field Gel Electrophoresis. Safe Pork: 6th International Symposium on Epidemiology and Control of Food-borne pathogens in Pork.Sept.6-9, Rohnert Park, California, USA.
- Gebreyes, W.A., Thakur, S. and Morrow, M.W.E. 2005. Prevalence and antimicrobial resistance of Salmonella isolated from conventional and antimicrobial-free swine herds in North Carolina. Safe Pork: 6th International Symposium on Epidemiology and Control of Food-borne pathogens in Pork.Sept.6-9, Rohnert Park, California, USA.
- Gebreyes, W.A., Morrow, M.W.E., Bahnson, P.B., Funk, J.A. and Thakur, S. (2005). Campylobacter prevalence and antimicrobial resistance in swine reared in antimicrobial-free and conventional production systems. Safe Pork: 6th International Symposium on the Epidemiology and Control of Food-borne pathogens in Pork.Sept.6-9, Rohnert Park, California, USA.
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Progress 01/01/04 to 12/31/04
Outputs
Sample Collection at Farm & Slaughter in NC, Ohio & Wisconsin: Completed sample collection at farm & slaughter in NC (10 conventional & 10 antimicrobial free farms; ABF). Ohio station sampled 9 farms (7 conventional & 2 ABF) while Wisconsin station completed sampling 5 farms (1 conventional & 4 ABF). Questionnaires collected from individual farm owners are currently being coded & analyzed. Campylobacter prevalence among pigs and carcasses in NC: Did not observe any significant difference in the prevalence of this pathogen at the finishing farms between the conventional (53 percent) & the ABF (55.8%) production systems (P = 0.46). Campylobacter prevalence at the slaughter level was not significantly different between the 2 production systems at the pre-evisceration, post-evisceration or the post-chill (USDA) stages. At post-chill stage, we could isolate Campylobacter only from the carcasses of the pigs reared in the ABF production system. Within both the production
systems, we found significantly higher prevalence of Campylobacter at the post-evisceration stage when compared to pre-evisceration and followed by significantly lower prevalence at the post-chill stage than the post-evisceration stage. Purposely decided to characterize three colonies per positive sample to determine whether phenotypic and genotypic distinct strains of Campylobacter are present in each sample. Total of 1472 (farm 1117 & slaughter 355) out of the 1634 Campylobacter strains that were isolated in this study were speciated with 99% of the isolates were Campylobacter coli. Antimicrobial susceptibility testing of Campylobacter isolates from NC: Completed the susceptibility testing for the Campylobacter isolates generated from North Carolina. Isolates were tested for their susceptibility against six antimicrobials namely chloramphenicol, ciprofloxacin, erythromycin, gentamicin, nalidixic acid and tetracycline. Irrespective of the production system, isolates exhibited maximum
frequency of resistance against tetracycline (66.2%) and erythromycin (59.6%). Resistance against the fluoroquinolone ciprofloxacin were rare and seen only in isolates (n=17) from the farm and not from the slaughter plants. Gentamicin and chloramphenicol resistant isolates were also and seen in 0.4 and 1.8% of the total isolates. Overall, we observed 19 different resistance patterns with significantly higher proportion of isolates from the ABF system at the farm and slaughter were pansusceptible (P < 0.001). Observed 11 different multi drug resistant patterns in 81 of the isolates with Ery Nal Tet (2.7%) being the predominant one. Campylobacter isolates from the conventional system, both on farm and from slaughter, were more often MDR than isolates from the ABF system (P =0.05). Genetic Fingerprinting of Campylobacter isolates from NC: Total of 144 isolates of Campylobacter coli randomly selected from total isolates have been fingerprinted using Pulsed Field Gel Electrophoresis
method. Preliminary results indicate that the Campylobacter coli strains have a diverse genome. Salmonella & Yersinia isolates from Wisconsin & Ohio are currently being tested to determine their antimicrobial resistance profile.
Impacts
This study highlights the prevalence of antimicrobial resistant C.coli from both the conventional and the ABF at farm and slaughter. Even though we detected higher number of resistant and multi drug resistant strains in isolates from the conventional system, antimicrobial resistant Campylobacter coli isolated from the ABF system warrants concern and points to the possible role of other environmental factors in transmission of these strains in pigs. Multi drug resistant ciprofloxacin resistant Campylobacter coli isolates from swine especially from the ABF reared pigs is alarming since this antimicrobial is used in treatment of severe invasive cases of campylobacteriosis.
Publications
- Gebreyes, W.A., Bahnson, P.B., Funk, J.A., Morrow, M.W.E. and Thakur, S. 2003. Campylobacter Prevalence and Diversity in Antimicrobial Free and Conventionally reared market swine. Safe Pork: 5th International Symposium on the Epidemiology and Control of Food-borne pathogens in Pork.Oct.1-4, Crete, Greece.
- Jefferson, A., Thakur, S. and Gebreyes, W.A. 2003. Antibiotic resistance testing of Campylobacter isolates. Second Annual NC State University Undergraduate Summer Research Program Symposium. August 7, Raleigh, NC, USA.
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Progress 01/01/03 to 12/31/03
Outputs
Objectives of this study were to determine the prevalence, antimicrobial resistance and genotypic diversity of three important pathogens: Salmonella enterica, Campylobacter spp. and Yersinia enterocolitica among pigs raised antimicrobial free (ABF) and those raised conventionally. A total of 60 farms were selected for entry into the project, 30 conventionally reared (antibiotics included in the feed) and antibiotic free (no antibiotics included in the feed) during the growing phase. The farms were equivalently distributed in three regions of the United States. Within each region, 10 conventional and 10 antibiotic free herds were recruited. Questionnaire was administered in person to record farm antibiotic use and management practices. Sampling consisted of 10 g of feces obtained from each of 30 pigs on farm within 48 hrs of slaughter. In addition, 10 carcass swabs were obtained at each of three points during the slaughter process; pre-evisceration, post-evisceration
and post-chilling. Salmonella: At time of submission, Salmonella data from a total of 24 farms (16 farms in NC, six farms from Ohio and two farms from WI) has been completed. 16 of the 24 farms were found to be positive for Salmonella. The overall prevalence of Salmonella on-farm was 17% and 6% at pre-evisceration. Comparison of USDA-FSIS carcass swabbing method with that of single swipe along midline revealed that better recovery of Salmonella was found using single swipe (3.7%) than USDA recommended approach (2%). There was no significant difference between prevalence at pre-evisceration (5.7%) and post-evisceration (5.5%). However, there was significant reduction after chilling of carcass (2%). Antimicrobial susceptibility and genotyping is underway. Campylobacter: 26 farms (18 from NC, 7 from Ohio & 2 from Wisonsin) have been processed for Campylobacter. All the 26 farms have been positive for Campylobacter. Campylobacter prevalence was high in both production types (ABF and
conventional). Significant reduction was seen after chilling (p<0.01). However, Campylobacter was still isolated post-chilling. 71% of the Campylobacter positive pigs tested for resistance pattern showed multiple resistance patterns and MIC levels. Resistance against Ciprofloxacin & Chloramphenicol was seen in ABF pigs in one farm. Multidrug resistant strains of Campylobacter were more commonly seen in pigs from ABF farms, including resistant to ciprofloxacin & chloramphenicol. Yersinia: At the time of submission, 5 farms have been completed (3 conventional and 2 antibiotic free). 4 herds (2 conventional & 2 antibiotic free) have had at least one Y. enterocolitica isolate from fecal samples. Among positive herds, the on-farm prevalence ranged from 10-60%. Results of the PCR for presence or absence of the ail gene and antimicrobial resistance are pending. Y. enterocolitica was not isolated from any carcass swabs. These preliminary results support previous research that suggested swine
could serve as a reservoir of Y. enterocolitica. Additionally, although preliminary in nature, there does not appear to be significant contamination of carcasses during slaughter and processing in those pigs sampled to date.
Impacts
The use of antimicrobials in food animal production and its association with antimicrobial resistance has been a debating issue. As this study compares antimicrobial-free farms with that of conventional ones, the results that were generated from this ongoing study has significant impacts on identification of sources of resistant organisms, understanding the diversity of strains and the role of processing practices in slaughter plants. The three food borne pathogens are among the leading cause of gastroenteritis in the United States. In case of pigs, C.coli is reported to be present in higher number than either Salmonella or Yersinia spp. Salmonella was also found commonly in two-third of the farms (16 of 24). There were significant findings during this progress report period. One is the similar level contamination before and after evisceration. This may imply that contamination during evisceration may not be a significant factor.
Publications
- Funk, J.A., Bahnson, P.B., Gebreyes, W.A. W.E.M. Morrow. Yersinia enterocolitica Prevalence in Antibiotic Free and Conventionally Reared Swine. 84th Conference of Research Workers in Animal Diseases (CRWAD). Chicago, IL, November 9-11, 2003. Oral presentation # 73.
- Gebreyes, W.A., Thakur S., Bahnson, P., Funk J., Morrow M. Antimicrobial Resistance and Diversity of Campylobacter in Antimicrobial Free and Conventionally Reared Market Swine. 84th Conference of Research Workers in Animal Diseases (CRWAD). Chicago, IL, November 9-11, 2003. Oral presentation # 44.
- Gebreyes, W.A., Thakur S., Bahnson, P., Funk J., Morrow M. 2003. Prevalence and Antimicrobial Resistance of Campylobacter spp. Among Antimicrobial-free and Conventionally reared market Pigs. 5th International Symposium on Epidemiology and Control of Foodborne Pathogens in pork (SAFEPORK). Crete, Greece, Oct.1-4, 2003.
- Gebreyes, W.A., Thakur S., Bahnson, P., Funk J., Morrow M. 2003. Antimicrobial Resistance of Campylobacter spp. Among Antimicrobial-free and Conventionally reared market Pigs. 12th International Conference of Campylobacter, Helicobacter and Related Organisms (CHRO). Aarhus, Denmark. September 6-10, 2003.
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Progress 01/01/02 to 12/31/02
Outputs
As planned in the original grant proposal, all the necessary preparations to conduct the study including recruiting participating conventional and antibiotic free swine producers, purchasing supplies has been completed in all the three participating states.
Impacts
(N/A)
Publications
- No publications reported this period
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