Clostridium perfringens is an important pathogen of animals and humans and is the causative agent of necrotic enteritis (NE) in poultry. This study focuses on the typing of intestinal C. perfringens isolates (n ‫؍‬ 61) from outbreaks of NE collected from several areas of Southern Ontario, using a recently developed multilocus sequence typing (MLST) technique. For comparison, C. perfringens isolates from healthy birds were also obtained and typed. An additional locus, the pfoS locus, was included in our analysis, in an attempt to increase the discriminatory ability of the method previously published. Birds were collected from two major poultry processors in Canada, and isolates from processor 2 formed a distinct MLST cluster. Isolates from healthy birds also collected from the outbreak flocks clustered together with isolates from the birds with NE. Although isolates from eight outbreaks clustered together, MLST types were also occasionally different between outbreaks. Strong linkage disequilibrium was observed between loci, suggesting a clonal C. perfringens population structure. Detection assays for toxin genes cpb2 (beta-2 toxin), tpeL, and the newly described netB (NetB toxin) were also performed. netB was almost always found in outbreak isolates, whereas cpb2 was found exclusively in healthy bird isolates. The toxin gene tpeL, which has not been previously identified in C. perfringens type A strains, was also found, but only in the presence of netB. Resistance to bacitracin was found in 34% of isolates from antimicrobial agent-free birds and in 100% of isolates from conventionally raised birds.

The study objective was to determine the effects of two treatment regimens on quantities of ceftiofur and tetracycline resistance genes in feedlot cattle. The two regimens were ceftiofur crystalline-free acid (CCFA) administered to either one or all steers within a pen and subsequent feeding/not feeding of therapeutic doses of chlortetracycline. A 26-day randomized controlled field trial was conducted on 176 steers. Real-time PCR was used to quantify blaCMY-2, blaCTX-M, tet(A), tet(B), and 16S rRNA gene copies/gram of feces from community DNA. A significant increase in ceftiofur resistance and a decrease in tetracycline resistance elements were observed among the treatment groups in which all steers received CCFA treatment, expressed as gene copies/gram of feces. Subsequent chlortetracycline administration led to rapid expansion of both ceftiofur and tetracycline resistance gene copies/gram of feces. Our data suggest that chlortetracycline is contraindicated when attempting to avoid expansion of resistance to critically important third-generation cephalosporins.

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This study analyzed antimicrobial resistance (AMR) and resistance genes in generic Escherichia coli isolated from retail meat samples purchased (2007-2008) in Alberta, Canada, and determined potential associations between resistance phenotypes and resistance genes with relation to the meat types. A total of 422 E. coli isolates from retail chicken, turkey, beef, and pork meats were tested for antimicrobial susceptibility. Multiplex PCRs were used to detect major resistance genes for tetracyclines [tet(A), tet(B), tet(C)], sulfonamides (sul1, sul2, sul3), aminoglycosides (strA/B, aadA, aadB, aac(3)IV, aphA1, aphA2), and β-lactamase (bla(CMY-2), bla(TEM), bla(SHV), bla(PSE-1)). Resistance to ciprofloxacin was not found in any isolate. Overall resistances to clinically important antimicrobials amoxicillin-clavulanic acid (16.8% of isolates) and ceftriaxone (12.6% isolates) were observed. These resistances were observed more frequently (p<0.0001) in chicken-derived E. coli than those from the other meat types. Resistance to multiple antimicrobials (≥ 5) was found in more chicken derived E. coli (32%) than E. coli from other meat types. The β-lactamase genes of clinical importance, including bla(CMY-2) and bla(TEM), were found in about 18% of poultry-derived E. coli and in only 5% of ground beef. The bla(CMY-2) gene was more likely to be found in E. coli from chicken than turkey, beef, or pork meats. The tet(A) gene was associated with bla(CMY-2), whereas bla(CMY-2) and bla(TEM) genes were associated with strA/B genes. Resistance genes for tetracycline, sulfonamides, and aminoglycosides were associated with the phenotypic expression of resistance to unrelated classes of antimicrobials. These data suggest the prevalence of AMR and select resistance genes were higher in poultry-derived E. coli. The multiple associations found between AMR phenotypes and resistance genes suggest a complex nature of resistance in E. coli from retail meat, and hence the use of a single antimicrobial could result in the selection of resistant E. coli not only to the drug being used but to other unrelated classes of antimicrobials as well.

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There is little information on the genetic basis of resistance to the critically important extended-spectrum cephalosporins (ESCs) in Enterobacteriaceae from dogs in Canada. This study assessed the frequency of ESC resistance in Enterobacteriaceae isolated from dogs in Ontario and the distribution of major ESC resistance genes in these bacteria. A total of 542 Enterobacteriaceae were isolated from 506 clinical samples from two diagnostic laboratories in Ontario. Eighty-eight ESC-resistant Enterobacteriaceae and 217 Escherichia coli were isolated from 234 fecal samples from dogs collected at leash-free dog parks. These fecal isolates were tested for ESC resistance along with the clinical isolates. Isolates with reduced ESC susceptibility were screened for bla, bla, and bla, and all CTX-M-positive isolates underwent whole-genome sequencing. The prevalence of ESC resistance in clinical Enterobacteriaceae was 10.4%. The average frequency of fecal carriage of ESC-resistant Enterobacteriaceae in healthy dogs was 26.5%. The majority of ESC-resistant isolates were E. coli and the other major Enterobacteriaceae carrying ESC resistance genes were Klebsiella pneumoniae and Proteus mirabilis. The results show that the same ESC resistance genes can be found in clinical and fecal Enterobacteriaceae in dogs. The identified E. coli sequence types (including ST131 and ST648) and CTX-M variants (including CTX-M-14, -15, and -27) support the hypothesis of transfer of resistant bacteria between humans and dogs. CTX-M-1 was frequently found in canine fecal Enterobacteriaceae, while it is still rare in human Enterobacteriaceae in Canada, thus suggesting transfer of resistant bacteria to dogs from food animals or other sources.

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