Redefining On-Farm Practices: The Perceived Impact of a Responsible Antimicrobial Use Regulation on Dairy Farmers.

The global food animal industry faces a growing concern regarding antimicrobial resistance (AMR), primarily driven by the use of antimicrobials (AMs) for the treatment, control, and prevention of diseases. Addressing this challenge requires promoting responsible antimicrobial use (AMU) practices. In 2019, the province of Québec, Canada, took a significant step by implementing a regulation that limits the use of AMs of very high importance for human medicine (category I AMs as defined by Health Canada) in the food animal industry. However, the implementation of such regulation can significantly influence behavioral shifts among producers, contributing to the wider effort against AMR. Therefore, the objective of this observational study was to describe the perceived changes in knowledge of dairy producers and on-farm practices following the implementation of this regulation, using a cohort design. Data collection involved administering questionnaires to 87 dairy producers from 3 regions of the province of Québec (Estrie, Montérégie, Centre-Du-Québec) before (2017-2018) and after (2020-2021) the implementation of the regulation. The questionnaires explored the descriptive characteristics of farms, the knowledge of producers about the categorization of AMs, their on-farm treatment practices, and the perceived impacts of the regulation. Statistical analysis included t-tests and McNemar tests to compare the paired data obtained using the 2 questionnaires. The results indicated an increase in the knowledge score (the number of AMs correctly categorized by the producers by their importance for human medicine) after the implementation of the regulation, suggesting an improved understanding of the categorization of AMs based on their importance for human medicine. Trends in AMU practices for treating clinical mastitis and reproductive diseases suggested that category I AMs were less likely to be reported as the primary treatment after the regulation, while category II AMs were more often reported as primary treatment. Adoption of the selective dry cow therapy method significantly increased, while the use of teat sealants remained unchanged. Moreover, producers had divergent perceptions regarding the effect of the regulation on the cure rates and disease frequencies. This disparity emphasizes the need for comprehensive data collection to discern the risks associated with such regulatory shifts. The study acknowledges several limitations, including the potential for recall bias, confirmation bias, and desirability bias. Despite these limitations, this study shows that implementing regulations to encourage responsible AMU drives positive transformations in producers' knowledge and on-farm practices. This underscores the pivotal impact of proactive interventions in combating the escalating threat of AMR within the global food animal industry.

Antimicrobial Resistance and Virulence Gene Profiles Among Escherichia coli Isolates from Retail Chicken Carcasses in Vietnam.

This study evaluated virulence and resistance profiles of Escherichia coli in chicken carcasses from three retail systems in Vietnam. Fresh chicken carcasses from traditional markets and fresh and frozen chicken carcasses from supermarkets were sampled in Vietnam. E. coli isolates from carcass rinses were characterized for extraintestinal pathogenic E. coli (ExPEC) virulence factors (iucD, cnf, papC, tsh, KpsMT II, afa, and sfa) and for phenotypical antimicrobial resistance by Sensititre ARIS® as well as genotypically by polymerase chain reaction. An elevated proportion (30% to 70%) of samples resistant to antimicrobials critically important for human medicine was observed in routine isolates, with no significant differences between the three retail systems. Multidrug-resistant (MDR) ExPEC isolates of phylogroup B1 and, of greater concern, of phylogroup F were detected. Extended-spectrum ß-lactamase (ESBL)- and AmpC ß-lactamase-producing E. coli possessing blaCTX-M or blaCMY-2 resistance genes, respectively, were found. The presence of ExPEC with a high level of antimicrobial resistance (more than 50% of isolates) and MDR (91% of isolates) and detection of ESBL-producing E. coli underline the potential health threat for humans associated with mishandled chicken carcasses or consumption of undercooked chicken meat in Vietnam.

Description of Antimicrobial-Resistant Escherichia coli and Their Dissemination Mechanisms on Dairy Farms.

Despite its importance in veterinary medicine, there is little information about antimicrobial resistance (AMR) and its transmission in dairy cattle. The aim of this work is to compare AMR phenotypes and genotypes in resistant Escherichia coli and to determine how the resistance genes spread among the E. coli population on dairy farms in Québec, Canada. From an existing culture collection of E. coli isolated from dairy manure, a convenient selection of the most resistant isolates (a high level of multidrug resistance or resistance to broad-spectrum ß-lactams or fluoroquinolones) was analyzed (n = 118). An AMR phenotype profile was obtained for each isolate. Whole genome sequencing was used to determine the presence of resistance genes, point mutations, and mobile genetic elements. In addition, a subset of isolates from 86 farms was taken to investigate the phylogenetic relationship and geographic distribution of the isolates. The average agreement between AMR phenotypes and genotypes was 95%. A third-generation cephalosporin resistance gene (blaCTX-M-15), a resistance gene conferring reduced susceptibility to fluoroquinolones (qnrS1), and an insertion sequence (ISKpn19) were detected in the vicinity of each other on the genome. These genes were harbored in one triplet of clonal isolates from three farms located >100 km apart. Our study reveals the dissemination of resistant E. coli clones between dairy farms. Furthermore, these clones are resistant to broad-spectrum ß-lactam and fluoroquinolone antimicrobials.

Clonal and plasmidic dissemination of critical antimicrobial resistance genes through clinically relevant ExPEC and APEC-like lineages (ST) in the dairy cattle population of Québec, Canada.

Antimicrobial resistance can be effectively limited by improving the judicious use of antimicrobials in food production. However, its effect on the spread of AMR genes in animal populations is not well described. In the province of Québec, Canada, a new legislation implemented in 2019 has led to an unprecedented reduction in the use of critical antimicrobials in dairy production. We aimed to investigate the potential link between ESBL/AmpC E. coli isolated before and after legislation and to determine the presence of plasmids carrying genes responsible for critical AMR. We collected fecal samples from calves, cows, and manure pit from 87 Québec dairy farms approximately 2 years before and 2 years after the legislation came into effect. The whole genomes of 183 presumptive ESBL/AmpC E. coli isolated after cefotaxime enrichment were sequenced. Their phylogenetic characteristics (MLST, serogroup, cgMLST) and the presence of virulence and resistance genes and replicons were examined. A maximum likelihood phylogenetic tree was constructed based on single nucleotide polymorphism (SNPs). We identified 10 clonal lineages (same cgMLST) and 7 clones (SNPs ≤ 52). Isolates belonging to these clones could be found on different farms before and after the legislation, strongly suggesting a clonal spread of AMR genes in the population during this 4-year period. All isolates were multidrug resistant (MDR), with clone 2 being notable for the presence of macrolide, fluoroquinolone, and third-generation cephalosporin resistance genes. We also identified clinically relevant ExPEC (ST10) and APEC-like lineages (ST117, ST58, ST88) associated with the presence of ExPEC and APEC virulence genes, respectively. Our data also suggests the presence of one epidemic plasmid belonging to the IncY incompatibility group and carrying qnrs1 and blaCTX-M-15. We demonstrated that AMR genes spread through farms and can persist over a 4-year period in the dairy cattle population through both plasmids and E. coli clones, despite the restriction of critical antimicrobial use. MDR ExPEC and APEC-like STs are present in the normal microbiota of cattle (more frequently in calves). These data increase our knowledge on gene dissemination dynamics and highlight the fact that biosecurity measures should be enhanced in this industry to limit such dissemination.

Impact of a Regulation Restricting Critical Antimicrobial Usage on Prevalence of Antimicrobial Resistance in Escherichia coli Isolates From Fecal and Manure Pit Samples on Dairy Farms in Québec, Canada.

To tackle antimicrobial resistance (AMR), one of the major health threats of this century, the World Health Organization (WHO) endorsed a global action plan in 2015. This plan calls countries to develop national actions to address AMR. The province of Québec, Canada, adopted a new regulation on the 25th of February 2019, to limit the use in food animals of antimicrobials of very high importance in human medicine. We aimed to establish the impact of this regulation by comparing the AMR situation in dairy cattle in Québec ~2 years before and 2 years after its introduction. We sampled calves, cows, and the manure pit in 87 farms. Generic and putative ESBL/AmpC E. coli were tested for susceptibility to 20 antimicrobials. Logistic regression was used to investigate whether the probability of antimicrobial resistance differed between isolates obtained from the pre and post regulation periods by sample type (calves, cows, manure pit) and in general. To identify AMR genes dissemination mechanisms, we sequenced the whole genome of 15 generic isolates. In the generic collection, at the herd level, the proportion of multidrug resistant (MDR) isolates, decreased significantly from 83 to 71% (p = 0.05). Folate inhibitor and aminoglycoside resistances demonstrated a significant decrease. However, when analyzed by sample type (calves, cows, manure pit), we did not observe a significant AMR decrease in any of these categories. In the ESBL/AmpC collection, we did not detect any significant difference between the two periods. Also, the general resistance gene profile was similar pre and post regulation. We identified both clonal and plasmidic dissemination of resistance genes. In conclusion, as early as 2 years post regulation implementation, we observed a significant decrease in MDR in the dairy industry in Quebec in the generic E. coli collection with folate inhibitor and aminoglycoside resistances showing the most significant decrease. No other significant decreases were yet observed.

High Risk Clone: A Proposal of Criteria Adapted to the One Health Context with Application to Enterotoxigenic Escherichia coli in the Pig Population.

The definition of a high risk clone for antibiotic resistance dissemination was initially established for human medicine. We propose a revised definition of a high risk clone adapted to the One Health context. Then, we applied our criteria to a cluster of enrofloxacin non susceptible ETEC:F4 isolates which emerged in 2013 in diseased pigs in Quebec. The whole genomes of 183 ETEC:F4 strains isolated in Quebec from 1990 to 2018 were sequenced. The presence of virulence and resistance genes and replicons was examined in 173 isolates. Maximum likelihood phylogenetic trees were constructed based on SNP data and clones were identified using a set of predefined criteria. The strains belonging to the clonal lineage ST100/O149:H10 isolated in Quebec in 2013 or later were compared to ETEC:F4 whole genome sequences available in GenBank. Prior to 2000, ETEC:F4 isolates from pigs in Quebec were mostly ST90 and belonged to several serotypes. After 2000, the isolates were mostly ST100/O149:H10. In this article, we demonstrated the presence of a ETEC:F4 high risk clone. This clone (1) emerged in 2013, (2) is multidrug resistant, (3) has a widespread distribution over North America and was able to persist several months on farms, and (4) possesses specific virulence genes. It is crucial to detect and characterize high risk clones in animal populations to increase our understanding of their emergence and their dissemination.

Prevalence, Risk Factors, and Characterization of Multidrug Resistant and ESBL/AmpC Producing Escherichia coli in Healthy Horses in Quebec, Canada, in 2015-2016.

Although antimicrobial resistance is an increasing threat in equine medicine, molecular and epidemiological data remain limited in North America. We assessed the prevalence of, and risk factors for, shedding multidrug-resistant (MDR) and extended-spectrum ß-lactamase (ESBL) and/or AmpC ß-lactamase-producing E. coli in healthy horses in Quebec, Canada. We collected fecal samples in 225 healthy adult horses from 32 premises. A questionnaire on facility management and horse medical history was completed for each horse. Indicator (without enrichment) and specific (following enrichment with ceftriaxone) E. coli were isolated and tested for antimicrobial susceptibility. The presence of ESBL/AmpC genes was determined by PCR. The prevalence of isolates that were non-susceptible to antimicrobials and to antimicrobial classes were estimated at the horse and the premises level. Multivariable logistic regression was used to assess potential risk factors for MDR and ESBL/AmpC isolates. The shedding of MDR E. coli was detected in 46.3% of horses. Non-susceptibility was most commonly observed to ampicillin, amoxicillin/clavulanic acid or streptomycin. ESBL/AmpC producing isolates were detected in 7.3% of horses. The most commonly identified ESBL/AmpC gene was blaCTX-M-1, although we also identified blaCMY-2. The number of staff and equestrian event participation were identified as risk factors for shedding MDR isolates. The prevalence of healthy horses harboring MDR or ESBL/AmpC genes isolates in their intestinal microbiota is noteworthy. We identified risk factors which could help to develop guidelines to preclude their spread.

Spread of multidrug-resistant IncHI1 plasmids carrying ESBL gene blaCTX-M-1 and metabolism operon of prebiotic oligosaccharides in commensal Escherichia coli from healthy horses, France.

The objective of the study was to identify the genetic determinants and characteristics of expanded-spectrum cephalosporin (ESC) resistance in commensal Escherichia coli from healthy horses in France in 2015. Faecal samples from 744 adult horses were screened for ESC-resistant E. coli isolates. The extended-spectrum beta-lactamase (ESBL)/AmpC resistance genes were identified using polymerase chain reaction (PCR) and sequencing. ESC phenotypes were horizontally transferred by conjugation or transformation. Plasmids carrying ESBL/AmpC genes were typed by PCR-based replicon typing, restriction fragment length polymorphism (RFLP), and plasmid multilocus sequence typing (pMLST). The ESC-resistant E. coli isolates were typed by XbaI macrorestriction analysis. Sixteen of 41 stables harboured at least one horse carrying ESC-resistant E. coli. The proportion of individually tested horses carrying ESC-resistant E. coli was 8.5% (28/328). Fifty non-redundant ESC-resistant E. coli isolates showing a great diversity of XbaI macrorestriction profiles belonged mainly to phylogroup B1, and were negative for major E. coli virulence genes, indicating they are commensal isolates. ESBL blaCTX-M genes were dominant (blaCTX-M-1, n=34; blaCTX-M-2, n=8; blaCTX-M-14, n=2) and located on conjugative plasmids belonging to various incompatibility groups (IncHI1, IncI1, IncN, IncY, or non-typeable). Among these, the multidrug-resistant IncHI1-pST9 plasmids were dominant and simultaneously harboured the blaCTX-M-1/2 genes and an operon enabling the metabolism of short-chain fructo-oligosaccharides (scFOS). In conclusion, commensal E. coli of French horses displayed a significant distribution of IncHI1-pST9 plasmids carrying both the blaCTX-M-1/2 gene and the fos metabolism operon. This finding highlights the risk of co-selection of multidrug-resistant IncHI1 plasmids carrying ESBL genes possibly mediated by the use of scFOS as prebiotic in horses.

Prevalence, risk factors, and characterization of multidrug resistant and extended spectrum ß-lactamase/AmpC ß-lactamase producing Escherichia coli in healthy horses in France in 2015.

BACKGROUND: Although antimicrobial resistance is increasingly common in equine medicine, molecular and epidemiological data remains scarce. OBJECTIVES: We estimated the prevalence of, and risk factors for, shedding of multidrug resistant (MDR), extended spectrum ß-lactamase (ESBL)-producing, and AmpC ß-lactamase-producing, or some combination of these in Escherichia coli in horses in France. We characterized ESBL/AmpC isolates for antimicrobial susceptibility and the presence of virulence and ESBL/AmpC-associated resistance genes. ANIMALS: Fecal samples from healthy adult horses at 41 premises were collected. A questionnaire was completed by each premises manager. A subset of these samples was tested to build 2 bacterial collections. METHODS: Indicator (without enrichment) and specific (enrichment with ceftriaxone) E. coli tested for antimicrobial susceptibility. Prevalence of isolates nonsusceptible to antimicrobials was estimated at the horse and the premises level. The ESBL/AmpC and virulence genes were identified by PCR. Multivariable logistic regression was used to investigate risk factors for MDR and ESBL/AmpC isolates at premises. RESULTS: Approximately 44% of horses shed MDR E. coli. Resistance most commonly was observed to ampicillin, streptomycin, and amoxicillin/clavulanic acid. Twenty-nine percent of premises housed horses shedding ESBL/AmpC-producing isolates. The ESBL/AmpC gene most commonly identified was blaCTX-M-1 . Virulence gene iutA was identified in 1 ESBL/AmpC-producing isolate. Medical treatment, staff numbers, and activity were identified as risk factors for housing horses shedding ESBL/AmpC-producing E. coli isolates. CONCLUSIONS AND CLINICAL IMPORTANCE: Prevalence of healthy horses harboring ESBL/AmpC genes and MDR isolates in their intestinal microbiota is substantial. Risk factors could be used to elaborate guidelines to prevent their dissemination.