Source: WASHINGTON STATE UNIVERSITY submitted to NRP
CLONAL DISSEMINATION OF ANTIMICROBIAL RESISTANT CAMPYLOBACTER JEJUNI AND ESCHERICHIA COLI
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
NRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
0189511
Grant No.
2001-35212-10844
Cumulative Award Amt.
(N/A)
Proposal No.
2001-02147
Multistate No.
(N/A)
Project Start Date
Oct 1, 2001
Project End Date
Sep 30, 2005
Grant Year
2001
Program Code
[(N/A)]- (N/A)
Recipient Organization
WASHINGTON STATE UNIVERSITY
240 FRENCH ADMINISTRATION BLDG
PULLMAN,WA 99164-0001
Performing Department
ANIMAL HEALTH RESEARCH CENTER
Non Technical Summary
Whenever antibiotics are used, those bacteria very susceptible to the drug die or are eliminated, and become replaced by more antibiotic resistant (AR) strains. Increasingly, there is concern that AR in both pathogenic bacteria and in the normal flora present a risk to the public health, and reduction in the degree of AR in these bacteria is seen as an important public health goal. The AR flora that appear after antibiotic exposure of cattle and other food animals animals may be 'new' AR strains originating on the farm, or may be pre-adapted strains that originated elsewhere and were transferred to the farm by animals, feeds, water, wildlife, humans, or other mechanisms. The origin of the AR flora is important, since different origins call for very different control measures. For one important AR pathogen, multiresistant Salmonella Typhimurium, wide dissemination of AR strains is the predominant process. Whether wide dissemination of AR strains is similarly important in other pathogenic (Campylobacter jejuni) and commensal bacteria (E. coli) in the bovine intestine is the focus of this proposal. In addition, we will determine whether AR E. coli can be competitvely displaced by non-AR strains. These studies will provide basic information about the role of clonal dissemination in the epidemiology of AR on cattle farms, and so aid risk assessment of commensal flora with AR and prediction of effective interventions to reduce AR frequency.
Animal Health Component
60%
Research Effort Categories
Basic
30%
Applied
60%
Developmental
10%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
7123310117010%
7123410117040%
7223310117010%
7223410117040%
Knowledge Area
712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins; 722 - Zoonotic Diseases and Parasites Affecting Humans;

Subject Of Investigation
3310 - Beef cattle, live animal; 3410 - Dairy cattle, live animal;

Field Of Science
1170 - Epidemiology;
Goals / Objectives
Our hypothesis is that AR in the enteric flora of cattle is clonally disseminated on a regional basis and stable over time. This hypothesis predicts that a limited number of genetic strain types will account for most frequently occurring AR profiles, that related or identical AR associated genetic strain types will be detectable on farms over prolonged periods, and that related or identical AR associated genetic strain types will be detectable on different farms within and possibly between regions. This hypothesis also suggests the possibility that clonal types compete for space in a particular intestinal niche in specific cattle types. These predictions are tested by the objectives of this project: Our first objective is to evaluate the association between clonal strain types and AR phenotypes of common commensal (E. coli) and (presumptive human) pathogenic (C. jejuni) enteric bacteria on dairy farms, feedlots, and calf ranches, including an evaluation of their persistence for six months or longer on individual farms and an evaluation of their geographical dissemination among different farm types, in adjacent and distant location. Our second objective is to determine if experimental dosing with frequently occurring non-AR strains of E. coli (biotype 58, as described in Preliminary data section) can competitively displace AR strains in calves after weaning. Objective 1: Determine the association of genetic strain types and AR phenotypes within E. coli and C. jejuni populations of dairy and beef breed calves and feedlot cattle. Objective 2: Determine if experimental dosing with frequently occurring non-AR strains of E. coli (biotype 58, as described in Preliminary data section) can competitively displace AR strains in calves after weaning.
Project Methods
We will identify and characterize potential genetic strain types of C. jejuni and E. coli associated with specific AR profiles, and then determine whether these AR-specific strain types are stable on individual farms for six months periods. Fecal samples will be obtained from calves and cattle on diverse farm types at two sampling visits six months apart. Fecal samples will be cultured to obtain 30 C. jejuni isolates (unselected for AR) and 128 E. coli isolates (selected for at least tetracycline resistance) per farm or ranch. The isolates will be analyzed for AR phenotype (both E. coli and C. jejuni) and strain characterization. Tentatively identified clonal types will be rigorously tested using data from additional characterization of AR genetic determinants encoding selected AR traits, and overall genetic organization (PFGE). Temporal stability of AR clonal types will be assessed within farms by comparison of the results of the two sample sets taken six months or more apart. The regional dissemination of AR clonal types will be assessed by comparisons both within and between states. If our hypothesis is correct, 1) AR clonal types of C. jejuni and E. coli sharing closely related or identical overall genetic organization, and identical AR genotypes will account for a majority of AR paterns within these species, 2) AR clonal types will be stable over six months periods on individual farms, 3) common AR clonal types will be identified on farms on regional or larger areas, 4) common AR clonal types will be identified on farms lacking antibiotic use selection pressures (organic dairies), and 5) AR clonal types will be frequently found on beef cow-calf ranches adjacent to intensive dairy farming areas, but less frequently on beef cow-calf ranches remote from intensive dairy farming areas. In the second objective, we will determine if susceptible clones can competitively displace AR strains, using an experimental 'challenge' model. While the natural replacement of E. coli flora adapted to the enteric environment of milk-fed calves (that has been selected regionally for AR phenotypes due to prolonged and repeated antimicrobial use in the milk and for therapy) by a flora adapted to the enteric environment of ruminants (generally non-AR due to the relative lack of antimicrobial use in these age groups) is gradual over a several months period, it's logical to suspect that the replacement process can be greatly accelerated by increasing the natural level of exposure to the non-AR strains. That orally ingested strain types can colonize the intestinal tract has been confirmed, and competitive displacement is also showing promise for competitive displacement of pathogenic E. coli strains, specifically E. coli O157:H7 (109).

Progress 10/01/01 to 09/30/05

Outputs
OUTPUTS: This project was original scientific research and as a result the outputs were primarily scientific presentations at meetings, full length proceedings from meetings, and general information presentations to affected industry stakeholders. These included: 1) Steering committee for a colloquium on Pre-harvest Food Safety organized by the American Academy of Microbiologists. The colloquium was held in Perthshire, Scotland on December 4-5, 2003; 2) The veterinarian's role in controlling the emergence and dissemination of drug resistant bacteria. Annual Meeting of the Korean Society for Veterinary Science, Seoul National University, Seoul, Republic of Korea. April 20, 2006; 3) Besser TE, Hancock D, Davis MA. 2003. The veterinarian's role in controlling the emergence and dissemination of drug-resistant bacteria. J Vet Med Educ. 30:136-40. Full length proceedings. 4) Morley PS, Apley MD, Besser TE, Burney DP, Fedorka-Cray PJ, Papich MG, Traub-Dargatz JL, Weese JS. 2005. ACVIM Consensus: Antimicrobial Drug Use in Veterinary Medicine. J Vet Intern Med 19:617-629. Full length proceedings. 5) The veterinarians role in emergence and control of antimicrobial drug resistant bacteria. AAVMC Agenda for Action: Veterinary Medicine's Role in Biodefense and Public Health Conference, Washington DC, November, 2002. 6) Campylobacter in the Food Supply. WSU Annual Food Safety Farm to Table Conference. Best Western Hotel, Moscow ID, May 25, 2005. 7) Antimicrobial Resistance and Agriculture Invited address at the Academy of Dairy Veterinary Consultants Fall Meeting, LaJolla CA, Oct 7 - 8, 2005; 8) Clonal dissemination of antimicrobial resistant E. coli and thermophilic Campylobacter sp. in cattle. Invited address at the meeting of the USDA CSREES Epidemiologic Approaches for Food Safety meeting, Washington D.C., October 6, 2005; 9) Besser TE. Prevalence and Antimicrobial Resistance of Campylobacter spp. in the bovine reservoir. NW Chapter AOAC Annual Conference. Tacoma, WA. June 22, 2006. 10) Besser, Sischo, Moore. Series of producer talks in three Washington counties (Yakima, Whatcom and Adams) regarding antimicrobial use in calves and antimicrobial resistance. March 4-6, 2008. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
1) MDR E. coli were highly prevalent in all cattle production systems. 2) Calf ranches had higher prevalence of MDR E. coli compared to feedlots and dairies. Beef cow-calf ranges had lower prevalence of MDR E. coli compared to feedlots and dairies. 3) Cattle production systems located in California had higher prevalence of MDR E. coli compared to those located in Washington. 4) Conventional dairies and beef cow-calf ranches had higher prevalence of MDR E. coli than did organic dairies and ranches. 5) While some evidence of clonality of MDR E. coli across different production systems was identified, in general there was high diversity among MDR E. coli suggesting that there are many such disseminated MDR strains. 6) Resistant Campylobacter sp. were highly prevalent in all cattle production systems. 7) C. coli were frequently MDR, whereas only doxycycline resistance occurred at high frequency within C. jejuni cattle isolates. 8) No evidence of clonality of doxycycline resistant C. jejuni was observed. 9) Disseminated clonal MDR strains of C. coli were observed.

Publications

  • DeFrancesco KA, Cobbold RN, Rice DH, Besser TE, Hancock DD. 2004. Antimicrobial resistance of commensal Escherichia coli from dairy cattle associated with recent salmonellosis outbreaks. Veterinary Microbiology 98:55-61.
  • Bae W, Kaya KN, Hancock DD, Call DR, Park YH, Besser TE. 2005. Prevalence and antimicrobial resistance of thermophilic Campylobacter spp. from cattle farms in Washington State. Appl. Environ. Microbiol. 71:169-174.
  • Besser TE, LeJeune JT, Rice DH, Berg J, Stilborn RP, Kaya KN, Bae W, Hancock DD. 2005. Increasing prevalence of Campylobacter jejuni in feedlot cattle through the feeding period. Appl. Environ Microbiol. 71:5752-5758, 2005.
  • Bae W, Hancock DD, Call DR, ParkYH, Berge ACB, Finger RM, Sischo WM, Besser TE. 2007. Dissemination of Antimicrobial Resistant Strains of Campylobacter coli and Campylobacter jejuni among Cattle in Washington State and California. Veterinary Microbiology. 122:306-15.
  • Daniels JB, Call DR, Besser TE. 2007. Molecular Epidemiology of blaCMY-2 Plasmids Carried by Salmonella enterica and Escherichia coli Isolated from Cattle in the Pacific Northwest. Appl Environ Microbiol. 73(24):8005-11.
  • Berge AC, Hancock DD, Sischo WM, Besser TE. 2010. Geographic, farm, and animal factors associated with multiple antimicrobial resistance in fecal Escherichia coli isolates from cattle in the western United States. J Am Vet Med Assoc. 236:1338-44.


Progress 01/01/02 to 12/31/02

Outputs
The first years' efforts were largely directed towards 1) development of the needed methods to perform the study, 2) fieldwork: sampling the E. coli and C. jejuni flora of a variety of farms in the two regions to collect the bacterial isolates needed to identify strain-associated resistance, and 3) lab work: performing preliminary screening of the isolates obtained for antibiotic resistance and for genetic relatedness. The methods that have been developed to date include 96-well replica plating method to assess the metabolic profile and antibiotic resistance profiles of E. coli isolates, and antibiotic resistance profiles of C. jejuni. These methods have been applied to several thousand E. coli isolates obtained in each region's dairy farms, calf ranches, feedlots, and beef cow-calf operations. The profiling of the C. jejuni isolates is also underway. Specifically, 9 dairies, 6 cow-calf ranches, 4 beef feedlots, and 4 dairy calf ranches have been sampled, and over 6,000 E. coli and 2,000 thermophilic Campylobacter isolates have been obtained and banked. Analysis of these strains' antimicrobial resistance and genetic fingerprints is well underway. The first evaluations of candidate resistant clones, those E. coli carrying resistance to ceftiofur or other third generation cephalosporins, is in progress.

Impacts
Pending.

Publications

  • No publications reported this period


Progress 10/01/01 to 12/31/01

Outputs
Herd sampling has commenced. A dairy herd and a beef-cow-calf herd have been sampled, E. coli and Campylobacter isolates have been obtained, biotyped and tested for antimicrobial resistance, and banked. Work is beginning on comparing farms to identify isolates with shared biotypes/antibiograms in order to identify potential clonal strain types.

Impacts
To the extent that antimicrobial resistant clonal types of commensal E. coli and Campylobacters are identified among the antimicrobial resistant flora, dissemination of these strain types rather than de novo emergence of new antimicrobial resistant strain types will be supported as the principal mechanism accounting for changes in the frequency of antimicrobial resistance on cattle farms.

Publications

  • No publications reported this period