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Resistance to beta-lactam antimicrobials caused by extended-spectrum beta-lactamase (ESBL)-producing organisms is a global health concern. The objectives of this study were to (1) summarise the prevalence of potential ESBL-producing Escherichia coli (ESBL-EC) and Salmonella spp. (ESBL-SA) isolates from agrifood and human sources in Canada from 2012 to 2017, and (2) describe the distribution of ESBL genotypes among these isolates. All data were obtained from the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS). CIPARS analysed samples for the presence of ESBLs through phenotypic classification and identified beta-lactamase genes (blaTEM, blaSHV, blaCTX, blaOXA, blaCMY−2) using polymerase chain reaction (PCR) and whole genome sequencing (WGS). The prevalence of PCR-confirmed ESBL-EC in agrifood samples ranged from 0.5% to 3% across the surveillance years, and was detected most frequently in samples from broiler chicken farms. The overall prevalence of PCR-confirmed ESBL-SA varied between 1% and 4% between 2012 and 2017, and was most frequently detected in clinical isolates from domestic cattle. The TEM-CMY2 gene combination was the most frequently detected genotype for both ESBL-EC and ESBL-SA. The data suggest that the prevalence of ESBL-EC and ESBL-SA in Canada was low (i.e.

The zoonotic pathogen Campylobacter jejuni is among the leading causes of foodborne diseases worldwide. While C. jejuni colonises many wild animals and livestock, persistence mechanisms enabling the bacterium to adapt to host species' guts are not fully understood. In order to identify putative determinants influencing host preferences of distinct lineages, bootstrapping based on stratified random sampling combined with a k-mer-based genome-wide association was conducted on 490 genomes from diverse origins in Germany and Canada. We show a strong association of both the core and the accessory genome characteristics with distinct host animal species, indicating multiple adaptive trajectories defining the evolution of C. jejuni lifestyle preferences in different ecosystems. Here, we demonstrate that adaptation towards a specific host niche ecology is most likely a long evolutionary and multifactorial process, expressed by gene absence or presence and allele variations of core genes. Several host-specific allelic variants from different phylogenetic backgrounds, including dnaE, rpoB, ftsX or pycB play important roles for genome maintenance and metabolic pathways. Thus, variants of genes important for C. jejuni to cope with specific ecological niches or hosts may be useful markers for both surveillance and future pathogen intervention strategies.

All foods carry microbes, many of which are harmless, but foods can also carry pathogens and/or microbial indicators of contamination. Limited information exists on the co-occurrence of microbes of food safety concern and the factors associated with their presence. Here, a population-based repeated cross-sectional design was used to determine the prevalence and co-occurrence of Escherichia coli, Klebsiella spp., Salmonella spp. and Vibrio spp. in key food commodities - chicken, pork, prawns, salmon and leafy greens. Prevalence in 1369 food samples for these four target bacterial genera/species varied, while 25.6% of all samples had at least two of the target bacteria and eight different combinations of bacteria were observed as co-occurrence profiles in raw prawns. Imported frozen chicken was 6.4 times more likely to contain Salmonella than domestic chicken, and imported salmon was 5.5 times more likely to be contaminated with E. coli. Seasonality was significantly associated with E. coli and Klebsiella spp. contamination in leafy greens, with higher detection in summer and autumn. Moreover, the odds of Klebsiella spp. contamination were higher in summer in chicken and pork samples. These results provide insight on the bacterial species present on foods at retail, and identify factors associated with the presence of individual bacteria, which are highly relevant for food safety risk assessments and the design of surveillance programmes.

Non-typhoidal

In this study, we isolated and molecularly characterized 10 (1.6%) C. difficile isolates from 644 commercially available raw meat samples. Molecular typing by PFGE and ribotyping revealed NAP and ribotypes commonly associated with human clinical cases, suggesting retail meat could be a possible source of transmission warranting further investigation.

Consumption of prawns as a protein source has been on the rise worldwide with seafood identified as the predominant attributable source of human vibriosis. However, surveillance of non-cholera Vibrio is limited both in public health and in food. Using a population- and market share-weighted study design, 211 prawn samples were collected and cultured for Vibrio spp. Contamination was detected in 46 % of samples, and multiple diverse Vibrio isolates were obtained from 34 % of positive samples. Whole genome sequencing (WGS) and phylogenetic analysis illustrated a comprehensive view of Vibrio species diversity in prawns available at retail, with no known pathogenicity markers identified in Vibrio parahaemolyticus and V. cholerae . Antimicrobial resistance genes were found in 77 % of isolates, and 12 % carried genes conferring resistance to three or more drug classes. Resistance genes were found predominantly in V. parahaemolyticus , though multiple resistance genes were also identified in V. cholerae and V. vulnificus . This study highlights the large diversity in Vibrio derived from prawns at retail, even within a single sample. Although there was little evidence in this study that prawns are a major source of vibriosis in the UK, surveillance of non-cholera Vibrio is very limited. This study illustrates the value of expanding WGS surveillance efforts of non-cholera Vibrios in the food chain to identify critical control points for food safety through the production system and to determine the full extent of the public health impact.

Antimicrobial resistance surveillance targeting agricultural animals is practiced in many countries but does not often include media selective for cephalosporin resistance. Here, we compared the frequency of recovery of resistant Escherichia coli using selective and non-selective media from the cecal contents of 116 chickens collected by the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS). Third generation cephalosporin resistance was detected in 24 samples including 12, 10, and 2 on selective, non-selective, and both media, respectively. Isolates producing the CTX-M-1 ESBL were grown from 11 samples, 10 on selective medium only. Our results suggest that current surveillance approaches underestimate the true prevalence of resistance to critically important antimicrobials.

The role of free-ranging wildlife in the epidemiology of enteropathogens causing clinical illness in humans and domestic animals is unclear. Salmonella enterica and anti-microbial resistant bacteria have been detected in the faeces of raccoons (Procyon lotor), but little is known about the carriage of these bacteria in other sympatric meso-mammals. Our objectives were to: (a) report the prevalence of Salmonella and associated anti-microbial resistance, Campylobacter spp, Clostridium difficile, and anti-microbial resistant Escherichia coli in the faeces of striped skunks (Mephitis mephitis) and Virginia opossums (Didelphis virginiana) in southern Ontario; and (b) compare the prevalence of these bacteria in the faeces of these meso-mammal hosts with raccoons from a previously reported study. Faecal swabs were collected from striped skunks and Virginia opossums on five swine farms and five conservation areas from 2011 to 2013. Salmonella was detected in 41% (9/22) and 5% (5/95) of faecal swabs from Virginia opossums and striped skunks, respectively. None of the Salmonella serovars carried resistance to anti-microbials. The prevalence of Campylobacter spp., C. difficile, and anti-microbial resistant E. coli ranged from 6% to 22% in striped skunk and Virginia opossums. Using exact logistic regression, Salmonella was significantly more likely to be detected in faecal swabs of Virginia opossums than skunks and significantly less likely in faecal swabs from skunks than raccoons from a previously reported study. In addition, Campylobacter spp. was significantly more likely to be detected in raccoons than opossums. Salmonella Give was detected in 8/9 (89%) of Salmonella-positive Virginia opossum faecal swabs. Our results suggest that striped skunks and Virginia opossums have the potential to carry pathogenic enteric bacteria in their faeces. The high prevalence of Salmonella Give in Virginia opossum faecal swabs in this study as well as its common occurrence in other Virginia opossum studies throughout North America suggests Virginia opossums may be reservoirs of this serovar.

Antimicrobial resistant bacteria and zoonotic pathogens have previously been isolated from Canada geese. We examined the prevalence of three enteric bacteria (i.e. Campylobacter, Salmonella, Escherichia coli) among Canada geese from three sampling sources in southern Ontario from 2013 through 2015. Samples were obtained by convenience from hunting groups, diagnostic birds submitted for post-mortem, and fresh faeces from live birds in parks. Escherichia coli isolates were isolated and tested for susceptibility to 15 antimicrobials using the Canadian Integrated Program for Antimicrobial Resistance Surveillance test panel. The prevalences of Salmonella, Campylobacter and E. coli were 0%, 11.2% and 72.6%, respectively. Among E. coli isolates, 7.9% were resistant to ≥1 class of antimicrobials and 5.6% were resistant to ≥2 classes of antimicrobials, with some including resistance to antimicrobials of highest importance in human medicine. A significant association between season and E. coli resistance among samples from live birds was noted; summer samples had no resistant E. coli isolates, whereas spring samples demonstrated the highest prevalence of E. coli resistant to ≥1 class of antimicrobials (20.0%) among all sources. In addition, Campylobacter coli were only isolated from the spring faecal samples. Flock-level clustering was an important statistical consideration, as flock was a significant random effect in all but two of our models. Detection of Campylobacter and antimicrobial resistant E. coli from Canada geese suggests that these birds may play a role in disseminating these organisms within the environment.

The spread of antimicrobial resistance from human activity derived sources to natural habitats implicates wildlife as potential vectors of antimicrobial resistance transfer. Wild birds, including corvid species can disseminate mobile genetic resistance determinants through faeces. This study aimed to determine the occurrence of plasmid-mediated quinolone resistance (PMQR) genes in Escherichia coli and Klebsiella spp. isolates obtained from winter roosting sites of American crows (Corvus brachyrhynchos) and common ravens (Corvus corax) in Canada. Faecal swabs were collected at five roosting sites across Canada. Selective media isolation and multiplex PCR screening was utilized to identify PMQR genes followed by gene sequencing, pulse-field gel electrophoresis and multilocus sequence typing to characterize isolates. Despite the low prevalence of E. coli containing PMQR (1·3%, 6/449), qnrS1, qnrB19, qnrC, oqxAB and aac(6')-Ib-cr genes were found in five sequence types (ST), including E. coli ST 131. Conversely, one isolate of Klebsiella pneumoniae contained the plasmid-mediated resistance gene qnrB19. Five different K. pneumoniae STs were identified, including two novel types. The occurrence of PMQR genes and STs of public health significance in E. coli and Klebsiella pneumoniae recovered from corvids gives further evidence of the anthropogenic derived dissemination of antimicrobial resistance determinants at the human activity-wildlife-environment interface. Significance and impact of the study This study examined large corvids as possible vector species for the dissemination of antimicrobial resistance in indicator and pathogenic bacteria as a means to assess the anthropogenic dissemination of plasmid-mediated quinolone resistance (PMQR) genes. Although rare, PMQR genes were found among corvid populations across Canada. The clinically important Escherichia coli strain ST131 containing aac(6')-Ib-cr gene along with a four-class phenotypic antimicrobial resistance (AMR) pattern as well as one Klebsiella pneumoniae strain containing a qnrB19 gene were identified in one geographical location. Corvids are a viable vector for the circulation of PMQR genes and clinically important clones in wide-ranging environments.

To better understand the contribution of wildlife to the dissemination of Salmonella and antimicrobial resistance in Salmonella and Escherichia coli, we examined whole-genome sequence data from Salmonella and E. coli isolates collected from raccoons (Procyon lotor) and environmental sources on farms in southern Ontario. All Salmonella and phenotypically resistant E. coli collected from raccoons, soil, and manure pits on five swine farms as part of a previous study were included. We assessed for evidence of potential transmission of these organisms between different sources and farms utilizing a combination of population structure assessments (using core-genome multi-locus sequence typing), direct comparisons of multi-drug resistant isolates, and epidemiological modeling of antimicrobial resistance (AMR) genes and plasmid incompatibility (Inc) types. Univariable logistic regression models were fit to assess the impact of source type, farm location, and sampling year on the occurrence of select resistance genes and Inc types. A total of 159 Salmonella and 96 resistant E. coli isolates were included. A diversity of Salmonella serovars and sequence types were identified, and, in some cases, we found similar or identical Salmonella isolates and resistance genes between raccoons, soil, and swine manure pits. Certain Inc types and resistance genes associated with source type were consistently more likely to be identified in isolates from raccoons than swine manure pits, suggesting that manure pits are not likely a primary source of those particular resistance determinants for raccoons. Overall, our data suggest that transmission of Salmonella and AMR determinants between raccoons and swine manure pits is uncommon, but soil-raccoon transmission appears to be occurring frequently. More comprehensive sampling of farms, and assessment of farms with other livestock species, as well as additional environmental sources (e.g., rivers) may help to further elucidate the movement of resistance genes between these various sources.

Wild birds are considered a potential source of zoonotic pathogens. We report on the occurrence of

Until recently, the behaviors of antibiotics and their ecotoxicological impact have been overlooked in the environment. The topic is broad and encompasses a wide range of organisms including microorganisms, algae, invertebrates, and vertebrates inhabiting various aquatic ecosystems. Changing the equilibrium of any 1 component in such systems disrupts the balance of the whole system. The manufacturing and frequent use of fluoroquinolones in human and animal medicine raises great concern over the increase of antibiotic resistance prevalence in microorganisms; however, in addition, the fate of antibiotic parent and metabolite compounds entering environmental ecosystems through various pathways raises environmental impact concerns. Research has focused on the concentration of antibiotics present in environmental samples and the acute toxicity to organisms by way of animal assessment models; however, it remains unclear what role low-level chronic exposure plays in ecotoxicological effects on lifeforms in aquatic environments. The aim of the present review was to assess the levels of fluorquinolone use in animal and human medicine, to determine the pathways of dissemination, and to highlight the ecotoxicological implications in freshwater environments. Environ Toxicol Chem 2016;35:2647-2656. © 2016 SETAC.

One of the core goals of the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS) is to monitor major meat commodities for antimicrobial resistance. Targeted studies with methodologies based on core surveillance protocols are used to examine other foods, e.g., seafood, for antimicrobial resistance to detect resistances of concern to public health. Here we report the discovery of a novel Ambler class A carbapenemase that was identified in a nontoxigenic strain of Vibrio cholerae (N14-02106) isolated from shrimp that was sold for human consumption in Canada. V. cholerae N14-02106 was resistant to penicillins, carbapenems, and monobactam antibiotics; however, PCR did not detect common β-lactamases. Bioinformatic analysis of the whole-genome sequence of V. cholerae N14-02106 revealed on the large chromosome a novel carbapenemase (referred to here as VCC-1, for V ibrio c holerae c arbapenemase 1 ) with sequence similarity to class A enzymes. Two copies of bla VCC-1 separated and flanked by IS Vch9 (i.e., 3 copies of IS Vch9 ) were found in an acquired 8.5-kb region inserted into a VrgG family protein gene. Cloned bla VCC-1 conferred a β-lactam resistance profile similar to that in V. cholerae N14-02106 when it was transformed into a susceptible laboratory strain of Escherichia coli . Purified VCC-1 was found to hydrolyze penicillins, 1st-generation cephalosporins, aztreonam, and carbapenems, whereas 2nd- and 3rd-generation cephalosporins were poor substrates. Using nitrocefin as a reporter substrate, VCC-1 was moderately inhibited by clavulanic acid and tazobactam but not EDTA. In this report, we present the discovery of a novel class A carbapenemase from the food supply.

Toxoplasma gondii is a protozoan parasite which infects a wide variety of mammals and birds worldwide, including humans. Human toxoplasmosis is primarily transmitted through the ingestion of tissue cysts in raw or poorly cooked meat and organs of infected animals, or through the ingestion of oocysts in food, water or soil contaminated with cat faeces. There is a distinct paucity of information on the prevalence and molecular characteristics of T. gondii in retail meats in Canada. This study reports the presence of T. gondii DNA in 4.3% (12 of 281) of packages of fresh ground beef, chicken breasts and ground pork purchased at retail in three Canadian provinces. T. gondii prevalence was very similar among all three meat types tested, and among the provinces sampled. Genotyping of positive samples by means of PCR-RFLP and DNA sequencing demonstrated the presence of both T. gondii type II (66.7%) and type III (33.3%). These findings provide baseline data on the prevalence of T. gondii DNA in fresh meats purchased at retail in Canada and will allow for more accurate and meaningful health risk assessments for the purposes of developing food safety guidelines and policies., Highlights • T. gondii DNA detected in 4.3% (n = 281) of packages of beef, chicken and pork in Canada. • T. gondii prevalence was similar among the meat types tested and provinces sampled. • Genotyping of positive samples demonstrated both T. gondii type II and type III.

The role of free-ranging wildlife in the epidemiology of enteropathogens causing clinical illness in humans and domestic animals is unclear. Salmonella enterica and anti-microbial resistant bacteria have been detected in the faeces of raccoons (Procyon lotor), but little is known about the carriage of these bacteria in other sympatric meso-mammals. Our objectives were to: (a) report the prevalence of Salmonella and associated anti-microbial resistance, Campylobacter spp, Clostridium difficile, and anti-microbial resistant Escherichia coli in the faeces of striped skunks (Mephitis mephitis) and Virginia opossums (Didelphis virginiana) in southern Ontario; and (b) compare the prevalence of these bacteria in the faeces of these meso-mammal hosts with raccoons from a previously reported study. Faecal swabs were collected from striped skunks and Virginia opossums on five swine farms and five conservation areas from 2011 to 2013. Salmonella was detected in 41% (9/22) and 5% (5/95) of faecal swabs from Virginia opossums and striped skunks, respectively. None of the Salmonella serovars carried resistance to anti-microbials. The prevalence of Campylobacter spp., C. difficile, and anti-microbial resistant E. coli ranged from 6% to 22% in striped skunk and Virginia opossums. Using exact logistic regression, Salmonella was significantly more likely to be detected in faecal swabs of Virginia opossums than skunks and significantly less likely in faecal swabs from skunks than raccoons from a previously reported study. In addition, Campylobacter spp. was significantly more likely to be detected in raccoons than opossums. Salmonella Give was detected in 8/9 (89%) of Salmonella-positive Virginia opossum faecal swabs. Our results suggest that striped skunks and Virginia opossums have the potential to carry pathogenic enteric bacteria in their faeces. The high prevalence of Salmonella Give in Virginia opossum faecal swabs in this study as well as its common occurrence in other Virginia opossum studies throughout North America suggests Virginia opossums may be reservoirs of this serovar.

This study evaluated the carriage of AmpC and extended-spectrum beta-lactamase (ESBL) genes and associated plasmids in faecal bacteria of Canadian corvids. Faecal samples from 449 birds in five roosting sites across Canada were analyzed using selective media, screening for AmpC and ESBL genes by PCR, and sequencing. Genomic relatedness was determined by PFGE and MLST. Plasmid mobility was studied by conjugation and transformation experiments, followed by plasmid typing. In total, 96 (21%, n = 449) cefotaxime-resistant Escherichia coli and three (0.7%) Klebsiella pneumoniae isolates were identified. ESBL genes blaCTX-M-1 (n = 3), blaCTX-M-14 (n = 2), blaCTX-M-32 (n = 2) and blaCTX-M-124 (n = 1) were detected in eight E. coli isolates, whereas blaSHV-2 (2) was found in two K. pneumoniae. E. coli isolates contained blaCMY-2 (n = 83) and blaCMY-42 (n = 1). The high genetic diversity of the isolates and presence of clinically important E. coli ST69 (n = 1), ST117 (n = 7) and ST131 (n = 1) was revealed. AmpC genes were predominantly carried by plasmids of incompatibility groups I1 (45 plasmids), A/C (10) and K (7). The plasmid IncI1/ST12 was most common and found in diverse E. coli STs in all sites. Highly diverse E. coli isolates containing AmpC and ESBL genes, including clinically important clones and emerging plasmids, are in circulation throughout Canadian wildlife.

From May through October 2016, we conducted a repeated cross-sectional study examining the effects of temporal, spatial, flock and demographic factors (i.e. juvenile vs. adult) on the prevalence of Campylobacter and antimicrobial resistant Enterobacteriaceae among 344 fresh faecal samples collected from Canada geese (Branta canadensis) from four locations where birds nested in Guelph, Ontario, Canada. The overall prevalence of Campylobacter among all fresh faecal samples was 9.3% and was greatest in the fall when these birds became more mobile following the nesting season. Based on 40 gene comparative genomic fingerprinting (CGF40), the increase in prevalence noted in the fall was matched by an increase in the number of unique CGF40 subtypes identified. Resistance to colistin was detected most commonly, in 6% of Escherichia coli isolates, and was highest in the late summer months. All colistin-resistant isolates were negative for the mcr-1 to mcr-5 genes; a chromosomal resistance mechanism (PmrB) was identified in all of these isolates. The prevalence of samples with E. coli exhibiting multi-class resistance or extended spectrum beta-lactamase was low (i.e.

Case Description—A 9-year-old castrated male domestic shorthair cat was evaluated because of hematuria and weight loss after an 8-year history of intermittent signs of feline lower urinary tract disease (FLUTD). A complete diet history revealed that the cat was eating a commercial diet that does not undergo the same processing procedures as most pet foods and so might be at increased risk for bacterial contamination owing to a nonstandard industry cooking procedure. Clinical Findings—The cat had a history consistent with FLUTD, but bacteriologic culture of the urine revealed Salmonella organisms. Additional analysis revealed Salmonella enterica serotype I:ROUGH-O:g,m,s:- in samples of urine and feces as well as Salmonella enterica serotype Johannesburg and Salmonella enterica serotype Senftenberg in the diet. Treatment and Outcome—The cat responded positively to antimicrobial treatment for the Salmonella bacteriuria as well as to dietary and environmental management for the clinical signs associated with FLUTD. Clinical Relevance—Findings in this case highlighted an additional health consequence associated with ingestion of Salmonella-contaminated food. Such contamination is of particular concern with raw meat–based diets or diets that have not undergone standard industry cooking practices. Veterinarians should obtain a diet history for every companion animal during every evaluation to help with diagnosis and optimal treatment. (J Am Vet Med Assoc 2015;247:525–530)

Surveillance is an important component of an overall strategy to address antimicrobial resistant bacteria in food animals and the food chain. The poultry market has many points of entry into the Canadian food chain, and some production practices are underrepresented in terms of surveillance. For example, pathogen carriage and antimicrobial resistance surveillance data are limited in smallholder chicken flocks raised for slaughter at provincially inspected abattoirs. In Canada, antimicrobial resistance in Escherichia coli and Salmonella isolated from commercial broiler chicken flocks, slaughtered at federally inspected abattoirs, is monitored by the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS). The objective of this study was to establish baseline information of antimicrobial resistance presence in E. coli and Salmonella isolated from smallholder flocks in Ontario, utilizing CIPARS collection and isolation methodologies, and to compare findings with CIPARS federally inspected abattoir data from Ontario, Canada. Five chickens per flock were sampled from 205 smallholder flocks. Of 1,025 samples, the E. coli prevalence was 99% (1,022/1,025), and 47% (483/1,022) of positive E. coli isolates were resistant to one or more of the 14 antimicrobials. Furthermore, as compared to results reported for the CIPARS commercial flocks, E. coli isolates from smallholder flocks had significantly lower resistance prevalence to six of 14 individual antimicrobials. Recovery of E. coli did not differ between federally inspected and provincially inspected flocks. Salmonella prevalence at the bird level in smallholder flocks was 0.3% (3/1,025), significantly lower (p ≪ 0.0001, 95% CI 0.080%-0.86%) than federally inspected commercial flocks. The overall differences found between the commercial and smallholder flocks may be explained by differences in poultry husbandry practices and hatchery sources.

Over the past 15 years, hepatitis E virus (HEV), norovirus (NoV), and rotavirus (RV) have been hypothesized to be potentially zoonotic; swine and pork have been suggested as possible human infection sources for all 3 viruses. Our objective was to estimate HEV, NoV, and RV prevalence and load on Canadian retail pork chops and livers. Using the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS) sampling platform, pork livers (n = 283) and chops (n = 599) were collected, processed, and assayed for the 3 viruses by four collaborating federal laboratories using validated real time reverse transcriptase polymerase chain reactions (qRT-PCR). Follow-up qRT-PCR estimating viral load in genomic copies/g was followed by nested classical RT-PCR and isolate sequencing of a partial segment of the ORF2 gene. Local alignments were performed using MUSCLE (Multiple Sequence Comparison by Log-Expectation); a phylogenetic tree was created. Twenty-five livers and 6 chops were classified ‘positive’ (thresholds for viral RNA detected in both replicates of the assay) or ‘suspect’ (thresholds detected in one of two replicates) for HEV. Follow-up qRT-PCR detected HEV on 16 livers, 0 chops, and nested classical RT-PCR, on 14 livers and 0 chops. Initial qRT-PCR classified 12 chops ‘suspect’ for NoV. Follow-up qRT-PCR detected viral RNA on only one sample with thresholds greater than 40 in both replicates. No amplicon was yielded, and therefore no isolate was sequenced from this sample. Partial ORF2 genes from 14 HEV isolates were sequenced, and compared via sequence identity and phylogenetic analysis with selected human case isolates listed in NCBI-GenBank. Overall, HEV prevalence on retail pork was comparable with other published reports.

Wild birds are considered a potential source of zoonotic pathogens. We report on the occurrence of Campylobacter, Salmonella, and antimicrobial-resistant, nonspecific Escherichia coli in ducks, grebes, and swans obtained by convenience while conducting related research with Canada Geese (Branta canadensis). Samples were obtained in southern Ontario, Canada, between 2013 and 2015 from hunter-caught birds, birds submitted for postmortem diagnosis, and fresh feces from live birds in parks. A secondary objective was to characterize Campylobacter genotypes using comparative genomic fingerprinting. Salmonella and E. coli isolates were tested for susceptibility to 15 antimicrobials using the Canadian Integrated Program for Antimicrobial Resistance Surveillance test panel. A total of 71 samples were collected from 15 different waterfowl species. We detected Campylobacter, Salmonella, and E. coli in 17, 3, and 84% of samples, respectively. Ten unique Campylobacter subtypes were identified, some of which had been identified previously in water, poultry, waterfowl, and human clinical cases. Both Salmonella isolates were pansusceptible and 15% of E. coli isolates were resistant to at least one antimicrobial, including resistance to antimicrobials of highest importance to human health. Source attribution studies should examine the role of waterfowl in the dissemination of these pathogens.

Impacts  Eleven percent of generic Escherichia coli isolates from dogs that visited dog parks in southern Ontario during the summer of 2009 were resistant to at least one anti-microbial.  Based on multilevel multivariable logistic regression models, factors positively associated with shedding of anti-microbial-resistant generic E. coli included attending a dog day care, being a large mixed breed dog and exposure to compost; consumption of commercial dry and home cooked diets was sparing factor.  Dogs that visit dog parks may be a reservoir for anti-microbial-resistant bacteria that may pose a risk to humans. Summary Anti-microbial resistance can threaten health by limiting treatment options and increasing the risk of hospitalization and severity of infection. Companion animals can shed anti-microbial-resistant bacteria that may result in the exposure of other dogs and humans to anti-microbial-resistant genes. The prevalence of anti-micro- bial-resistant generic Escherichia coli in the faeces of dogs that visited dog parks in south-western Ontario was examined and risk factors for shedding anti-micro- bial-resistant generic E. coli identified. From May to August 2009, canine faecal samples were collected at ten dog parks in three cities in south-western Ontario, Canada. Owners completed a questionnaire related to pet characteristics and man- agement factors including recent treatment with antibiotics. Faecal samples were collected from 251 dogs, and 189 surveys were completed. Generic E. coli was iso- lated from 237 of the faecal samples, and up to three isolates per sample were tested for anti-microbial susceptibility. Eighty-nine percent of isolates were pan-susceptible; 82.3% of dogs shed isolates that were pan-susceptible. Multiclass resistance was detected in 7.2% of the isolates from 10.1% of the dogs. Based on multilevel multivariable logistic regression, a risk factor for the shedding of generic E. coli resistant to ampicillin was attending dog day care. Risk factors for the shed- ding of E. coli resistant to at least one anti-microbial included attending dog day care and being a large mixed breed dog, whereas consumption of commercial dry and home cooked diets was protective factor. In a multilevel multivariable model for the shedding of multiclass-resistant E. coli, exposure to compost and being a large mixed breed dog were risk factors, while consumption of a commercial dry

Impacts  Based on this study, dogs that visited dog parks may be a potential source of zoonotic pathogen exposure to humans.  Exposure to compost, increasing age and consumption of a commercial dry diet were associated with a decreased risk of shedding Campylobacter spp. in dogs.  Dogs less than 1 year of age and those exposed to outdoor water sources had an increased risk of shedding C. upsaliensis. Summary An estimated 6 million pet dogs live in Canadian households with the potential to transmit zoonotic pathogens to humans. Dogs have been identified as carriers of Salmonella, Giardia and Campylobacter spp., particularly Campylobacter upsali- ensis, but little is known about the prevalence and risk factors for these pathogens in pet dogs that visit dog parks. This study examined the prevalence of these organisms in the faeces of dogs visiting dog parks in three cities in south-western Ontario, as well as risk factors for shedding Campylobacter spp. and C. upsalien- sis. From May to August 2009, canine faecal samples were collected at ten dog parks in the cities of Guelph and Kitchener-Waterloo, Ontario, Canada. Owners were asked to complete a questionnaire related to pet characteristics and manage- ment factors including age, diet and activities in which the dog participates. Fae- cal samples were collected from 251 dogs, and 189 questionnaires were completed. Salmonella, Giardia and Campylobacter spp. were present in 1.2%, 6.4% and 43.0% of faecal samples, respectively. Of the Campylobacter spp. detected, 86.1% were C. upsaliensis, 13% were C. jejuni and 0.9% were C. coli. Statistically significant sparing factors associated with the shedding of Campylob- acter spp. included the feeding of a commercial dry diet and the dog's exposure to compost. Age of dog had a quadratic effect, with young dogs and senior dogs having an increased probability of shedding Campylobacter spp. compared with adult dogs. The only statistically significant risk factor for shedding C. upsaliensis was outdoor water access including lakes and ditches, while dogs >1 year old were at a lower risk than young dogs. Understanding the pet-related risk factors for Campylobacter spp. and C. upsaliensis shedding in dogs may help in the devel- opment of awareness and management strategies to potentially reduce the risk of transmitting this pathogen from dogs to humans.

Escherichia coli isolated from colon, cecal, and fecal samples are commonly used as indicator organisms to monitor antimicrobial resistance (AMR) in wild animals. The effect of sample type on E. coli recovery and AMR detection is unknown. We compared the prevalence of E. coli, the frequency of AMR, and the level of agreement between cecal, colon, and fecal samples collected from individual wild house mice (Mus musculus). Samples were collected from 49 mice trapped at swine farms, May-October 2008. We found no difference in the proportion of E. coli-positive samples (82%) among sample types and the agreement among sample types for E. coli recovery ranged from moderate to almost perfect. The percentage of E. coli positive samples resistant to one or more antimicrobial varied from 53% for colon samples to 71% for cecum samples; however, there was no significant difference in the proportion of resistant samples among sample types. The agreement among sample types for resistance to one or more antimicrobial ranged from fair to substantial. These findings indicate that there is no definitive sample type for studies of AMR in house mice. However, we suggest that fecal samples, which have direct contact with the environment, are likely the best sample to use in studies assessing the potential impact of AMR in wildlife on environmental and public health.

To the Editor: Carbapenems, antimicrobial drugs of last resort, are recommended only for severe community- and healthcare-associated multidrug-resistant bacterial infections. In Canada, carbapenem-resistant infection rates in hospitals remained low (

One of the core goals of the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS) is to monitor major meat commodities for antimicrobial resistance. Targeted studies with methodologies based on core surveillance protocols are used to examine other foods, e.g., seafood, for antimicrobial resistance to detect resistances of concern to public health. Here we report the discovery of a novel Ambler class A carbapenemase that was identified in a nontoxigenic strain of Vibrio cholerae (N14-02106) isolated from shrimp that was sold for human consumption in Canada. V. cholerae N14-02106 was resistant to penicillins, carbapenems, and monobactam antibiotics; however, PCR did not detect common β-lactamases. Bioinformatic analysis of the whole-genome sequence of V. cholerae N14-02106 revealed on the large chromosome a novel carbapenemase (referred to here as VCC-1, for Vibrio cholerae carbapenemase 1) with sequence similarity to class A enzymes. Two copies of blaVCC-1 separated and flanked by ISVch9 (i.e., 3 copies of ISVch9) were found in an acquired 8.5-kb region inserted into a VrgG family protein gene. Cloned blaVCC-1 conferred a β-lactam resistance profile similar to that in V. cholerae N14-02106 when it was transformed into a susceptible laboratory strain of Escherichia coli. Purified VCC-1 was found to hydrolyze penicillins, 1st-generation cephalosporins, aztreonam, and carbapenems, whereas 2nd- and 3rd-generation cephalosporins were poor substrates. Using nitrocefin as a reporter substrate, VCC-1 was moderately inhibited by clavulanic acid and tazobactam but not EDTA. In this report, we present the discovery of a novel class A carbapenemase from the food supply.

Numerous serotypes of Salmonella have been detected in a variety of wild animals, including raccoons (Procyon lotor). Raccoons are common, mid-size omnivores that live in close association with people in urban and rural areas in Ontario. Although raccoons are known to shed Salmonella, little is known about their potential long-term role in maintaining Salmonella infections. We sampled feces from raccoons in three areas of Ontario: one primarily urban site around Niagara, one primarily rural site north of Guelph, and the grounds of the Toronto Zoo, in 2007 to identify which serotypes of Salmonella were commonly shed by raccoons in southern Ontario. In addition, we conducted a longitudinal study at the Toronto Zoo site to determine if raccoons remain persistently infected with Salmonella. Salmonella was found in 45% of samples. The prevalence of Salmonella in raccoon feces ranged from 27% at the rural site to 65% at the urban site. We detected 16 serotypes of Salmonella in 83 positive samples. The most common serotype detected in raccoons from the rural and zoo sites was Salmonella enterica serotype Typhimurium, whereas Salmonella Newport was detected most commonly in the urban site. Only one raccoon of 11 that were captured in four or more consecutive trapping sessions shed the same Salmonella serotype for two consecutive months, suggesting that raccoons regularly acquire new Salmonella serotypes from the environment.

Summary The purpose of this study was to determine pet-related management factors that may be associated with the presence of Salmonella spp. in feces of pet dogs from volunteer households. From October 2005 until May 2006, 138 dogs from 84 households in Ontario were recruited to participate in a cross-sectional study. Five consecutive daily fecal samples were collected from each dog and enrichment culture for Salmonella spp. was performed. A higher than expected number of the dogs (23.2%; 32/138) had at least one fecal sample positive for Salmonella, and 25% (21/84) of the households had at least one dog shedding Salmonella. Twelve serotypes of Salmonella enterica subsp. enterica were identified, with the predominant serotypes being Typhimurium (33.3%; 13/39), Kentucky (15.4%; 6/39), Brandenburg (15.4%; 6/39) and Heidelberg (12.8%; 5/39). Univariable logistic regression models were created with a random effect for household to account for clustering. Statistically significant risk factors for a dog testing positive included having contact with livestock, receiving a probiotic in the previous 30 days, feeding a commercial or homemade raw food diet, feeding raw meat and eggs, feeding a homemade cooked diet, and having more than one dog in the household. In two-variable models that controlled for feeding raw food, the non-dietary variables were no longer statistically significant. These results highlight the potential public health risk of including raw animal products in canine diets.

SUMMARYFrom July 2008 until May 2009, 240 client-owned pet dogs from seven veterinary clinics in the Region of Waterloo, Ontario, Canada participated in a study to determine pet-related management factors that may be associated with the presence ofCampylobacterspp. in dogs. The prevalence ofCampylobacterspp. carriage in our study population of pet dogs was 22%, with 19% of the dogs positive forC. upsaliensis, and 3% positive forC. jejuni. A significant risk factor from multivariable logistic regression models for bothCampylobacterspp. andC. upsaliensiscarriage was having homemade cooked food as the dog's diet or added to its diet, and a significant sparing factor for both models was treatment with antibiotics in the previous month. Increasing age of the dog decreased the odds ofCampylobacterspp. andC. upsaliensiscarriage. Based on the high prevalence ofCampylobacter, and specificallyC. upsaliensis, further research concerning pet dogs as a risk factor for campylobacteriosis in humans is warranted.

Clostridium difficile infection (CDI) has been associated with increased illness and death in Canada since 2000 (1,2). Although multiple genotypes with higher levels of virulence and antimicrobial resistance have been recognized (1,3), little is known about risk factors for CDI acquisition outside healthcare facilities. In a 2005 study, we found C. difficile in 20% of retail meats sampled in Canada (4). Limitations to that study included limited geographic representation, nonsystematic sampling, and the use of a nonvalidated culture method. These sampling limitations prevent valid extrapolations. Broader sampling and a better understanding of the culture methods were thus required to reassess the prevalence of retail meat contamination with C. difficile. Here, we determined the prevalence of C. difficile in retail meat by using a broad-based government sampling infrastructure, compared 3 culture methods, characterized recovered isolates, and evaluated month-to-month variability in C. difficile recovery.

Wild animals not normally exposed to antimicrobial agents can acquire antimicrobial agent-resistant bacteria through contact with humans and domestic animals and through the environment. In this study we assessed the frequency of antimicrobial resistance in generic Escherichia coli isolates from wild small mammals (mice, voles, and shrews) and the effect of their habitat (farm or natural area) on antimicrobial resistance. Additionally, we compared the types and frequency of antimicrobial resistance in E. coli isolates from swine on the same farms from which wild small mammals were collected. Animals residing in the vicinity of farms were five times more likely to carry E. coli isolates with tetracycline resistance determinants than animals living in natural areas; resistance to tetracycline was also the most frequently observed resistance in isolates recovered from swine (83%). Our results suggest that E. coli isolates from wild small mammals living on farms have higher rates of resistance and are more frequently multiresistant than E. coli isolates from environments, such as natural areas, that are less impacted by human and agricultural activities. No Salmonella isolates were recovered from any of the wild small mammal feces. This study suggests that close proximity to food animal agriculture increases the likelihood that E. coli isolates from wild animals are resistant to some antimicrobials, possibly due to exposure to resistant E. coli isolates from livestock, to the resistance genes of these isolates, or to antimicrobials through contact with animal feed.

OBJECTIVE To determine pet-related management factors associated with the carriage of antimicrobial-resistant Salmonella spp and Escherichia coli in a population of pet dogs. SAMPLE 138 dogs from 84 households in Ontario, Canada. PROCEDURES From October 2005 through May 2006, dogs and households in Ontario, Canada, were recruited to participate in a cross-sectional study. Fecal samples were submitted for culture of Salmonella spp and E coli, which provided 515 bacterial isolates for antimicrobial susceptibility testing. Multilevel logistic regression models with random effects for household and dog were created to identify pet-related management factors associated with antimicrobial resistance. RESULTS Bacterial species, feeding a homemade diet or adding homemade food to the diet, feeding a raw diet or adding anything raw to the diet, feeding a homemade raw food diet, and feeding raw chicken in the past week were significant risk factors for antimicrobial resistance in this population of dogs. CONCLUSIONS AND CLINICAL RELEVANCE In this study, several potentially important pet-related risk factors for the carriage of antimicrobial-resistant Salmonella spp and E coli in pet dogs were identified. Further evaluation of risk factors for antimicrobial resistance in dogs may lead to development of evidence-based guidelines for safe and responsible dog ownership and management to protect the public, especially pet owners who are immunocompromised.

Until recently, the behaviors of antibiotics and their ecotoxicological impact have been overlooked in the environment. The topic is broad and encompasses a wide range of organisms including microorganisms, algae, invertebrates, and vertebrates inhabiting various aquatic ecosystems. Changing the equilibrium of any 1 component in such systems disrupts the balance of the whole system. The manufacturing and frequent use of fluoroquinolones in human and animal medicine raises great concern over the increase of antibiotic resistance prevalence in microorganisms; however, in addition, the fate of antibiotic parent and metabolite compounds entering environmental ecosystems through various pathways raises environmental impact concerns. Research has focused on the concentration of antibiotics present in environmental samples and the acute toxicity to organisms by way of animal assessment models; however, it remains unclear what role low-level chronic exposure plays in ecotoxicological effects on lifeforms in aquatic environments. The aim of the present review was to assess the levels of fluorquinolone use in animal and human medicine, to determine the pathways of dissemination, and to highlight the ecotoxicological implications in freshwater environments. Environ Toxicol Chem 2016;35:2647-2656. © 2016 SETAC.

Raccoons are common in urban and rural environments and can carry a wide range of bacteria, including Salmonella, that can negatively affect human and livestock health. Although previous studies have reported that raccoons shed a variety of Salmonella serovars in their faeces, it is unknown whether Salmonella is carried on raccoon paws. Our objective was to compare the prevalence of Salmonella on the paws and in the faeces of raccoons in south-western Ontario. Raccoons were sampled in a repeat cross-sectional study on five swine farms and five conservation areas from May to October 2012. A total of 416 paired faecal and paw samples were collected from 285 individual raccoons. Salmonella was detected in 18% (75/416; 95% CI, 14-22%) and 27% (111/416; 95% CI, 22-31%) of paw and faecal samples, respectively. Salmonella was detected only on paws in 8% (35/416; 95% CI, 5.9-11.5%), only in faeces in 17% (71/416; 95% CI, 13.6-21.0%) and on both paws and in faeces in 10% (40/416; 95% CI, 7.0-12.9%) of raccoon captures. Multilevel logistic regression models were used to examine associations between the presence of Salmonella and age (adult, juvenile), sex (male, female), location type (swine farm, conservation area), sample type (faeces, paw) and season (May-July and August-October). Random intercepts were included to account for clustering by individual animal and location. Significant differences, that varied by sample type and season, were noted in the prevalence of Salmonella carriage between sexes. Raccoons can carry Salmonella serovars known to infect humans and livestock on their paws and/or in their faeces and therefore have the potential to mechanically and biologically disseminate Salmonella among livestock facilities and human recreational areas.

Objectives The purpose of this study was to compare the distribution of chloramphenicol and kanamycin resistance genes across three populations of porcine Escherichia coli. Methods PCR was used to assess the distribution of the major chloramphenicol and kanamycin resistance genes catA1, cmlA and floR, and aphA1, aphA2 and aadB in enterotoxigenic E. coli (ETEC), non-ETEC isolates from cases of diarrhoea and commensal E. coli from healthy pigs. Associations between these genes and resistance genes for other antimicrobials or virulence genes were assessed. Results The chloramphenicol and kanamycin resistance genes were distributed differently among the three E. coli populations. While aphA1, aphA2 and aadB were evenly distributed among resistant ETEC, non-ETEC and commensals, the catA1 gene was significantly more frequent in ETEC than in non-ETEC and commensals. Transformation experiments confirmed statistical associations by demonstrating that elt, estB, astA, aadA and sul1 were located with catA1 on a large ETEC plasmid. Plasmids carrying cmlA also carried sul3 and aadA. Other plasmids carrying floR and aadB also carried tet(A), sul2, strA/strB, bla(CMY-2) and occasionally aac(3)IV. Conclusions The clustering of genes observed is a likely cause for chloramphenicol resistance persistence. Similar to tetracycline, chloramphenicol resistance genes are physically linked to virulence genes. This is not the case for kanamycin resistance determinants, which were linked to other resistance genes only.

A total of 318 Escherichia coli isolates obtained from diarrheic and healthy pigs in Ontario from 2001 to 2003 were examined for their susceptibility to 19 antimicrobial agents. They were tested by PCR for the presence of resistance genes for tetracycline, streptomycin, sulfonamides, and apramycin and of 12 common virulence genes of porcine E. coli . Antimicrobial resistance frequency among E. coli isolates from swine in Ontario was moderate in comparison with other countries and was higher in isolates from pigs with diarrhea than in isolates from healthy finisher pigs. Resistance profiles suggest that cephamycinases may be produced by ≥8% of enterotoxigenic E. coli (ETEC). Resistance to quinolones was detected only in enterotoxigenic E. coli (≤3%). The presence of sul3 was demonstrated for the first time in Canada in porcine E. coli isolates. Associations were observed among tetA , sul1 , aadA , and aac(3)IV and among tetB , sul2 , and strA / strB , with a strong negative association between tetA and tetB . The paa and sepA genes were detected in 92% of porcine ETEC, and strong statistical associations due to colocation on a large plasmid were observed between tetA , estA , paa , and sepA . Due at least in part to gene linkages, the distribution of resistance genes was very different between ETEC isolates and other porcine E. coli isolates. This demonstrates that antimicrobial resistance epidemiology differs significantly between pathogenic and commensal E. coli isolates. These results may have important implications with regards to the spread and persistence of resistance and virulence genes in bacterial populations and to the prudent use of antimicrobial agents.