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The global food trade provides a means of disseminating antimicrobial resistant (AMR) bacteria and genes. Using selective media, carbapenem-resistant species of Enterobacterales ( Providencia sp. and Citrobacter sp.), were detected in a single package of imported frozen shrimp purchased from a grocery store in Ohio, USA. Polymerase chain reaction confirmed that both isolates harbored bla NDM-1 genes. Following PacBio long read sequencing, the sequences were annotated using the NCBI Prokaryotic Genome Annotation Pipeline. The bla NDM-1 genes were found in IncC plasmids, each with different antimicrobial resistance island configuration. We found that the bla NDM-1 AMR islands had close relationships with previously reported environmental, food, and clinical isolates detected in Asia and the United States, highlighting the importance of the food chain in the global dissemination of antimicrobial resistance.

Since their commercialization, scientists have known that antimicrobial use kills or inhibits susceptible bacteria while allowing resistant bacteria to survive and expand. Today there is widespread antimicrobial resistance (AMR), even to antimicrobials of last resort such as the carbapenems, which are reserved for use in life-threatening infections. It is often convenient to assign responsibility for this global health crisis to the users and prescribers of antimicrobials. However, we know that animals never treated with antimicrobials carry clinically relevant AMR bacteria and genes. The causal pathway from bacterial susceptibility to resistance is not simple, and dissemination is cyclical rather than linear. Amplification of AMR occurs in healthcare environments and on farms where frequent exposure to antimicrobials selects for resistant bacterial populations. The recipients of antimicrobial therapy release antimicrobial residues, resistant bacteria, and resistance genes in waste products. These are reduced but not removed during wastewater and manure treatment and enter surface waters, soils, recreational parks, wildlife, and fields where animals graze and crops are grown for human and animal consumption. The cycle is complete when a patient carrying AMR bacteria is treated with antimicrobials that amplify the resistant bacterial populations. Reducing the development and spread of AMR requires a One Health approach with the combined commitment of governments, medical and veterinary professionals, agricultural industries, food and feed processors, and environmental scientists. In this review and in the companion Currents in One Health by Ballash et al, JAVMA, April 2024, we highlight just a few of the steps of the complex cyclical causal pathway that leads to the amplification, dissemination, and maintenance of AMR.

Once considered to be a simple cause-and-effect relationship with localized impact, the concept of how antimicrobial use drives antimicrobial resistance is now recognized as a complex, transdisciplinary problem on a global scale. While the issue of antimicrobial resistance is often studied and addressed at the antimicrobial-human or antimicrobial-animal treatment interface, the role of the environment in the One Health dynamics of antimicrobial resistance is not as well understood. Antimicrobial-resistant bacteria, including those resistant to carbapenem drugs, are emerging in veterinary clinical environments, on farms, and in natural habitats. These multidrug-resistant bacteria can colonize our livestock and companion animals and are later disseminated into the environment, where they contaminate surface waters and colonize wildlife. From here, the One Health transmission cycle of antimicrobial-resistant bacteria is completed as environmental reservoirs can serve as sources of antimicrobial resistance transmission into human or animal healthcare settings. In this review, we utilize a One Health perspective to evaluate how environments become contaminated and, in turn, become reservoirs that can colonize and infect our veterinary species, and how the veterinary field is combating environmental contamination with antimicrobial stewardship regulations and program implementation. The companion Currents in One Health by Parker et al, AJVR, April 2024, addresses the intensive research that justifies this One Health cycle of antimicrobial resistance transmission and emerging techniques that are dissecting the complex interactions at the One Health interface.

Antibiotic therapy is the standard of care for urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC). However, previous antibiotic therapy may impart a selective pressure that influences the population structure and pathogenic potential of infecting UPEC strains. Here, we conducted a 3-year study using whole-genome-sequencing analysis and retrospective medical record review to characterize how antibiotic exposure influenced the phenotypic antibiotic resistance, acquired resistome, virulome, and population structure of 88 UTI-causing E. coli strains from dogs. A majority of UTI-associated E. coli strains were from phylogroup B2 and clustered within sequence type 372. Previous antibiotic exposure was associated with a population shift toward UPEC from phylogroups other than the typical urovirulent phylogroup B2. The specific virulence profiles within the accessory virulome that were associated with antibiotic use were elicited by the effect of antibiotics on UPEC phylogenetic structure. Among phylogroup B2, antibiotic exposure increased the quantity of genes within the resistome and the odds of developing reduced susceptibility to at least one antibiotic. Non-B2 UPEC strains harbored a more diverse and greater resistome that conferred reduced susceptibility to multiple antibiotic classes following antibiotic exposure. Collectively, these data suggest that previous antibiotic exposure establishes an environment that provides a selective edge to non-B2 UPEC strains through their diverse and abundant antibiotic resistance genes, despite their lack of urovirulence genes. Our findings highlight the necessity for judicious use of antibiotics as we uncover another mechanism by which antibiotic exposure and resistance can influence the dynamics of bacterial infectious disease. IMPORTANCE Urinary tract infections (UTIs) are one of the most common infections of dogs and humans. While antibiotic therapy is the standard of care for UTIs and other infections, antibiotic exposure may influence the pathogenic profile of subsequent infections. We used whole-genome sequencing and retrospective medical record review to characterize the effect of systemic antibiotic therapy on the resistance, virulence, and population structure of 88 UTI-causing UPEC strains isolated from dogs. Our results indicate that antibiotic exposure alters the population structure of infecting UPEC strains, providing a selective edge for non-B2 phylogroups that harbor diverse and abundant resistance gene catalogues but fewer urovirulence genes. These findings highlight how antibiotic resistance can influence pathogen infection dynamics and have clinical implications for the judicious use of antibiotics for bacterial infections., Competing Interests: The authors declare no conflict of interest.

Environmental surfaces can serve as reservoirs for pathogens and antimicrobial-resistant (AMR) bacteria in healthcare settings. Although active surveillance programs are used in veterinary and human healthcare, unconventional settings like zoological facilities are often overlooked, even though antimicrobials are used to maintain the health of their animal collections. Here, we used electrostatic cloths to conduct active environmental surveillance over a 2-year period at two zoological institutions to determine contamination prevalence of human-only and mixed animal-human touch environments with AMR bacteria. We recovered Enterobacterales isolates that expressed quinolone resistance, an AmpC-like phenotype, and an extended-spectrum β-lactamase phenotype from 144 (39%), 141 (38.2%), and 72 (19.5%) of the environmental samples, respectively. The zoological institutions, areas and exhibits within the zoological facility, and sampling surface type affected the odds of recovering AMR bacteria from the environment. Three carbapenemase-producing Enterobacter cloacae complex ST171 isolates recovered from one zoological facility harbored an IncH12 plasmid with a Tn 4401b -KPC-4 transposon conferring multidrug resistance. One isolate maintained three tandem repeats of a Tn 4401b -KPC-4 element on an IncHI2 plasmid, although this isolate was susceptible to the four carbapenem drugs tested. These three isolates and their IncH12 plasmids shared significant genomic similarity with two E. cloacae complex isolates recovered from canine patients at a regional veterinary hospital during year 2 of this study. Our results indicated that surface environments at zoological institutions can serve as reservoirs for AMR bacteria and their genes and have implications for animal and public health. IMPORTANCE Environmental surfaces can be a source of antimicrobial-resistant (AMR) bacteria that pose a risk to human and animal health. Zoological institutions are unique environments where exotic animals, staff, and visitors intermingle and antimicrobials are used to maintain animal health. However, zoological environments are often overlooked as reservoirs of AMR bacteria. Here, we show that zoological environments can serve as reservoirs of fluoroquinolone-resistant and extended-spectrum cephalosporin-resistant bacteria. In addition, we isolated three carbapenemase-producing Enterobacter cloacae complex strains carrying bla KPC-4 , including one with a unique, tandem triplicate of the Tn 4401b -KPC-4 element. Comparative whole genomics of these isolates with two E. cloacae complex isolates from patients at a regional veterinary hospital highlighted the possibility of local KPC-4 spread between animal environments. Our results suggest that environments at zoological institutions serve as reservoirs for AMR bacteria and pose a hypothetical One Health risk to the public, staff, and the wild animal populations in captivity., Competing Interests: The authors declare no conflict of interest.

Concern about zoonoses and wildlife has increased. Few studies described the role of wild mammals and environments in the epidemiology of Salmonella . Antimicrobial resistance is a growing problem associated with Salmonella that threatens global health, food security, the economy, and development in the 21st century. The aim of this study is to estimate the prevalence and identify antibiotic susceptibility profiles and serotypes of non-typhoidal Salmonella enterica recovered from non-human primate feces, feed offered, and surfaces in wildlife centers in Costa Rica. A total of 180 fecal samples, 133 environmental, and 43 feed samples from 10 wildlife centers were evaluated. We recovered Salmonella from 13.9% of feces samples, 11.3% of environmental, and 2.3% of feed samples. Non-susceptibility profiles included six isolates from feces (14.6%): four non-susceptible isolates (9.8%) to ciprofloxacin, one (2.4%) to nitrofurantoin, and one to both ciprofloxacin and nitrofurantoin (2.4%). Regarding the environmental samples, one profile was non-susceptible to ciprofloxacin (2.4%) and two to nitrofurantoin (4.8%). The serotypes identified included Typhimurium/I4,[5],12:i:-, S . Braenderup/Ohio, S . Newport, S . Anatum/Saintpaul, and S . Westhampton. The epidemiological surveillance of Salmonella and antimicrobial resistance can serve in the creation of strategies for the prevention of the disease and its dissemination throughout the One Health approach.

As safe agents of last resort, carbapenems are reserved for the treatment of infections caused by multidrug-resistant organisms. The impact of β-lactam antibiotics, cefotaxime, and meropenem on the frequency and diversity of carbapenemase-producing organisms recovered from environmental samples has not been fully established. Therefore, this methodological study aimed at determining β-lactam drugs used in selective enrichment and their impact on the recovery of carbapenemase-producing Enterobacterales (CPE) from untreated wastewater. We used a longitudinal study design where 1L wastewater samples were collected weekly from wastewater treatment plant (WWTP) influent and quarterly from contributing sanitary sewers in Columbus, Ohio USA with 52 total samples collected. Aliquots of 500 mL were passed through membrane filters of decreasing pore sizes to enable all the water to pass through and capture bacteria. For each sample, the resulting filters were placed into two modified MacConkey (MAC) broths, one supplemented with 0.5 μg/mL of meropenem and 70 μg/mL of ZnSO4 and the other supplemented with 2 μg/mL cefotaxime. The inoculated broth was then incubated at 37° C overnight, after which they were streaked onto two types of correspondingly-modified MAC agar plates supplemented with 0.5 μg/mL and 1.0 μg/mL of meropenem and 70 μg/mL of ZnSO4 and incubated at 37°C overnight. The isolates were identified based on morphological and biochemical characteristics. Then, up to four distinct colonies of each isolate's pure culture per sample were tested for carbapenemase production using the Carba-NP test. Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) MALDI-TOF MS was used to identify carbapenemase-producing organisms. In total 391 Carba-NP positive isolates were recovered from the 52 wastewater samples: 305 (78%) isolates had blaKPC, 73 (19%) carried blaNDM, and 14 (4%) harbored both blaKPC and blaNDM resistance genes. CPE genes of both blaKPC and blaNDM were recovered in both types of modified MAC broths, with 84 (21%) having a blaKPC gene, 22 (6%) carrying blaNDM and 9 (2%) harbored both a blaKPC and blaNDM of isolates recovered from MAC medium incorporated with 0.5ug/mL meropenem and 70ug/mL ZnSO4. The most prevalent isolates were Klebsiella pneumoniae, Escherichia coli, and Citrobacter spp., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Surface waters, especially those receiving wastewater flows, can disseminate antimicrobial resistant bacteria (ARB), antimicrobial resistance genes (ARG), and antibiotics. In the Scioto River of central Ohio, United States, we evaluated fishes as potential sentinels of ARB and antimicrobial contamination and investigated the influence of antimicrobial exposure on the fish intestinal resistome. Seventy-seven fish were collected from river reaches receiving inputs from two wastewater treatment plants that serve the greater Columbus Metropolitan Area. Fish were screened for the presence of cephalosporin-resistant (CeRO) and carbapenem-resistant (CRO) organisms, epidemic carbapenemase genes, and antibiotic drugs and metabolites using culture methods, droplet digital PCR, and ultra-high performance liquid chromatography tandem mass spectroscopy (UHPLC-MS/MS). Nearly 21% of fish harbored a CeRO in their resistome, with 19.4% exhibiting bacteria expressing an AmpC genotype encoded by blaCMY, and 7.7% with bacteria expressing an extended-spectrum β-lactamase phenotype encoded by blaCTX-M. blaKPC and blaNDM were present in 87.7% (57/65) and 80.4% (37/46) of the intestinal samples at an average abundance of 104 copies. Three antibiotics-lincomycin (19.5%), azithromycin (31.2%) and sulfamethoxazole (3.9%)-were found in hepatic samples at average concentrations between 25-31 ng/g. Fish harboring blaCTX-M and those exposed to azithromycin were at greater odds of being downstream of a wastewater treatment plant. Fish that bioconcentrated antibiotics in their liver were not at greater odds of harboring CeRO, CRO, or epidemic carbapenemase gene copies in their resistome. Our findings confirm that fishes can be effective bioindicators of surface waters contaminated with ARB, ARG, and antibiotics. Moreover, our findings highlight the varying importance of different mechanisms that facilitate establishment of ARB in aquatic ecosystems., Competing Interests: The authors have declared that no competing interests exist.

Biofilm formation enhances bacteria's ability to colonize unique niches while protecting themselves from environmental stressors. Escherichia coli that colonize the urinary tract can protect themselves from the harsh bladder environment by forming biofilms. These biofilms promote persistence that can lead to chronic and recurrent urinary tract infections (UTI). While biofilm formation is frequently studied among urinary E. coli, its association with other pathogenic mechanisms and adaptations in certain host populations remains poorly understood. Here we utilized whole genome sequencing and retrospective medical record analysis to investigate associations between the population structure, phenotypic resistance, resistome, virulome, and patient demographic and clinical findings of 104 unique urinary E. coli and their capacity to form biofilms. We show that population structure including multilocus sequence typing and Clermont phylogrouping had no association with biofilm capacity. Among clinical factors, exposure to multiple antibiotics within that past 30 days and a clinical history of recurrent UTIs were positively associated with biofilm formation. In contrast, phenotypic antimicrobial reduced susceptibility and corresponding acquired resistance genes were negatively associated with biofilm formation. While biofilm formation was associated with increased virulence genes within the cumulative virulome, individual virulence genes did not influence biofilm capacity. We identified unique virulotypes among different strata of biofilm formation and associated the presence of the tosA/R-ibeA gene combination with moderate to strong biofilm formation. Our findings suggest that E. coli causing UTI in dogs utilize a heterogenous mixture of virulence genes to reach a biofilm phenotype, some of which may promote robust biofilm capacity. Antimicrobial use may select for two populations, non-biofilm formers that maintain an arsenal of antimicrobial resistance genes to nullify treatment and a second that forms durable biofilms to avoid therapeutic insults., Competing Interests: The authors have declared that no competing interests exist.

Wildlife play a role in the acquisition, maintenance, and dissemination of antimicrobial resistance (AMR). This is especially true at the human-domestic animal-wildlife interface, like urbanized areas, where interactions occur that can promote the cross-over of AMR bacteria and genes. We conducted a 2-year fecal surveillance ( n  = 783) of a white-tailed deer (WTD) herd from an urban park system in Ohio to identify and characterize cephalosporin-resistant and carbapenemase-producing bacteria using selective enrichment. Using generalized linear mixed models we found that older (OR = 2.3, P  < 0.001), male (OR = 1.8, P  = 0.001) deer from urbanized habitats (OR = 1.4, P  = 0.001) were more likely to harbor extended-spectrum cephalosporin-resistant Enterobacterales. In addition, we isolated two carbapenemase-producing Enterobacterales (CPE), a Klebsiella quasipneumoniae harboring bla KPC-2 and an Escherichia coli harboring bla NDM-5 , from two deer from urban habitats. The genetic landscape of the plasmid carrying bla KPC-2 was unique, not clustering with other reported plasmids encoding KPC-2, and only sharing 78% of its sequence with its nearest match. While the plasmid carrying bla NDM-5 shared sequence similarity with other reported plasmids encoding NDM-5, the intact IS26 mobile genetic elements surrounding multiple drug resistant regions, including the bla NDM-5 , has been reported infrequently. Both carbapenemase genes were successfully conjugated to a J53 recipient conferring a carbapenem-resistant phenotype. Our findings highlight that urban environments play a significant role on the transmission of AMR bacteria and genes to wildlife and suggest WTD may play a role in the dissemination of clinically and epidemiologically relevant antimicrobial resistant bacteria. IMPORTANCE The role of wildlife in the spread of antimicrobial resistance is not fully characterized. Some wildlife, including white-tailed deer (WTD), can thrive in suburban and urban environments. This may result in the exchange of antimicrobial resistant bacteria and resistance genes between humans and wildlife, and lead to their spread in the environment. We found that WTD living in an urban park system carried antimicrobial resistant bacteria that were important to human health and resistant to antibiotics used to treat serious bacterial infections. This included two deer that carried bacteria resistant to carbapenem antibiotics which are critically important for treatment of life-threatening infections. These two bacteria had the ability to transfer their AMR resistance genes to other bacteria, making them a threat to public health. Our results suggest that WTD may contribute to the spread of antimicrobial resistant bacteria in the environment.

The changing epidemiologic role of wildlife as reservoirs of antimicrobial-resistant bacteria (ARB) is poorly understood. In this study, we characterize the phenotypic resistance of commensal Escherichia coli from fecal samples of 879 individual white-tailed (Odocoileus virginianus; WTD) over a ten-year period and analyze resistance patterns. Our results show commensal E. coli from WTD had significant linear increases in reduced susceptibility to 5 of 12 antimicrobials, including broad-spectrum cephalosporins and fluoroquinolones, from 2006 to 2016. In addition, the relative frequency distribution of minimal inhibitory concentrations of two additional antimicrobials shifted towards higher values from across the study period. The prevalence of multidrug-resistant commensal E. coli increased over the study period with a prevalence of 0%, 2.2%, and 3.7% in 2006, 2012, and 2016, respectively. WTD may be persistently and increasingly exposed to antibiotics or their residues, ARB, and/or antimicrobial resistance genes via contaminated environments like surface water receiving treated wastewater effluent., (© 2021. EcoHealth Alliance.)

Retail beef and pork, including processed products, can serve as vehicles for the zoonotic foodborne transmission of pathogens and antimicrobial resistant bacteria. However, processed and seasoned products like sausages, are not often included in research and surveillance programs. The objective of this study was to investigate retail ground beef and pork, including processed products, for the presence of common foodborne pathogens and antimicrobial resistant bacteria. We purchased 763 packages of fresh and fully cooked retail meat products during 29 visits to 17 grocery stores representing seven major grocery chains located in west and central Ohio. Each package of meat was evaluated for contamination with methicillin-resistant Staphylococcus aureus (MRSA), Salmonella spp., Enterobacteriaceae expressing extended-spectrum cephalosporin resistance, and carbapenemase-producing organisms (CPO). Only 3 of the 144 (2.1%) packages of fully cooked meat products contained any of these organisms, 1 with an extended-spectrum β-lactamase-producing (ESBL) Enterobacteriaceae and 2 with CPO. Among the 619 fresh meat products, we found that 85 (13.7%) packages were contaminated with MRSA, 19 (3.1%) with Salmonella, 136 (22.0%) with Enterobacteriaceae expressing an AmpC (bla CMY ) resistance genotype, 25 (4.0%) with Enterobacteriaceae expressing an ESBL (bla CTX-M ) resistance genotype, and 31 (5.0%) with CPO, primarily environmental organisms expressing intrinsic carbapenem resistance. However, one CPO, a Raoultella ornithinolytica, isolated from pork sausage co-harbored both bla KPC-2 and bla NDM-5 on IncN and IncX3 plasmids, respectively. Our findings suggest that fresh retail meat, including processed products can be important vehicles for the transmission of foodborne pathogens and antimicrobial resistant bacteria, including those with epidemic carbapenemase-producing genotypes.

Objective: To estimate the prevalence of extended-spectrum cephalosporin-, carbapenem-, and fluoroquinolone-resistant bacteria of the family Enterobacteriaceae in the feces of hospitalized horses and on hospital surfaces., Sample: Fecal and environmental samples were collected from The Ohio State University Galbreath Equine Center (OSUGEC) and a private referral equine hospital in Kentucky (KYEH). Feces were sampled within 24 hours after hospital admission and after 48 hours and 3 to 7 days of hospitalization., Procedures: Fecal and environmental samples were enriched, and then selective media were inoculated to support growth of Enterobacteriaceae bacteria that expressed resistance phenotypes to extended-spectrum cephalosporins, carbapenems, and fluoroquinolones., Results: 358 fecal samples were obtained from 143 horses. More samples yielded growth of Enterobacteriaceae bacteria that expressed resistance phenotypes (AmpC β-lactamase, OR = 4.2; extended-spectrum beta-lactamase, OR = 3.2; and fluoroquinolone resistance, OR = 4.0) after 48 hours of hospitalization, versus within 24 hours of hospital admission. Horses hospitalized at KYEH were at greater odds of having fluoroquinolone-resistant bacteria (OR = 2.2). At OSUGEC, 82%, 64%, 0%, and 55% of 164 surfaces had Enterobacteriaceae bacteria with AmpC β-lactamase phenotype, extended-spectrum beta-lactamase phenotype, resistance to carbapenem, and resistance to fluoroquinolones, respectively; prevalences at KYEH were similarly distributed (52%, 32%, 1%, and 35% of 315 surfaces)., Conclusions and Clinical Relevance: Results indicated that antimicrobial-resistant Enterobacteriaceae may be isolated from the feces of hospitalized horses and from the hospital environment. Hospitalization may lead to increased fecal carriage of clinically important antimicrobial-resistance genes.

One objective of this study was to determine overall prevalence of Salmonella in ground pork from U.S. retail stores over three seasons including both case-ready and store-ground packages. Package types collected included: overwrap, chub, modified atmosphere packaging, and other (plastic or wax paper wrapped). Because package type represents different production systems and are subject to varied microbiological government regulation and testing methodologies, both USDA-FSIS and FDA Salmonella isolation protocols were performed. Another objective of the study was to determine serotypes and antimicrobial susceptibility profiles of the isolates obtained from the ground pork samples. Ground pork aliquots were subjected to real-time PCR. Recovered isolates were serotyped and minimum inhibitory concentration analysis to 15 antimicrobials was determined using microbroth dilution. Overall prevalence of Salmonella in ground pork from the 865 samples collected was 1.39%. Prevalence was not affected by package type ( p  = 0.29) nor grind location (case-ready vs. store-ground; p  = 0.17). Season affected Salmonella prevalence ( p  = 0.05) with most isolates found during fall, and there was a tendency for geographic region to affect prevalence ( p  = 0.07). The USDA Salmonella isolation method was more effective at recovering isolates ( p  = 0.01) compared with the FDA methodology and yielded a kappa statistic of 0.26 as a measure of agreement. The serotypes isolated included: Infantis, 4,5,12:i:-, Brandenburg, Typhimurium var 5-, Seftenberg, and Johannesburg with only two packages containing multiple serotypes. No isolates were resistant to antibiotics commonly used to treat human Salmonella infections including extended spectrum cephalosporins or fluoroquinolones. Although the recovery of Salmonella from retail ground pork samples was rare, Salmonella Typhimurium (and its monophasic variant 4,5,12:i:-), which are among the most common serovars recovered from human infections, were recovered. Therefore, more effective strategies to further reduce or eliminate these pathogens from retail pork products are warranted.

Widespread use of antimicrobials in human and veterinary medicine drives the emergence and dissemination of resistant bacteria in human, animal, and environmental reservoirs. The AVMA and FDA Center for Veterinary Medicine have both taken public positions emphasizing the importance of incorporating antimicrobial stewardship in veterinary clinical settings; however, a model for implementing a comprehensive antimicrobial stewardship program in veterinary practice is not readily available. In 2015, The Ohio State University College of Veterinary Medicine began developing a veterinary antimicrobial stewardship program modeled on existing programs in human health-care institutions and the 7 core elements of a successful hospital antimicrobial stewardship program, as defined by the CDC. The program includes comprehensive antimicrobial use guidelines, active environmental surveillance, and enhanced infection control procedures in The Ohio State University Veterinary Medical Center, along with routine monitoring and reporting of antimicrobial prescribing practices and antimicrobial susceptibility patterns of common pathogens isolated from patients and the hospital environment. Finally, programs have been developed to educate clinicians, staff, and students on antimicrobial resistance and appropriate antimicrobial prescribing practices. The antimicrobial stewardship program has been designed to help clinicians and students confidently make judicious antimicrobial use decisions and provide them with actionable steps that can help them act as strong stewards while providing the best care for their patients. This report describes our program and the process involved in developing it, with the intent that the program could serve as a potential model for other veterinary medical institutions.

Objectives: To identify the minimum inhibitory concentration (MIC) distribution for commonly used topical antibiotics from isolates of dogs and horses with ulcerative bacterial keratitis, and to investigate changes in MIC values over time and following treatment with topical fluoroquinolones., Animals Studied: One hundred thirty-four client-owned dogs and 20 client-owned horses with bacterial ulcerative keratitis., Procedure: Minimum inhibitory concentration values for 14 topical antibiotics were reported for canine and equine cases of bacterial ulcerative keratitis between 2013 and 2018. Changes in MIC values over time and after treatment with topical fluoroquinolones were reported., Results: The three most common bacterial genera isolated were Staphylococcus, Streptococcus, and Pseudomonas. Together, these represented 79.4% of canine cases and 77.4% of equine cases. Overall, isolates from horses tended to have lower MIC values, as did Pseudomonas isolates from both dogs and horses, compared to other bacterial genera, especially Staphylococcus spp. The MIC values of erythromycin and trimethoprim sulfa for Staphylococcus spp., and the MIC value of moxifloxacin for Pseudomonas significantly increased over time. Previous topical fluoroquinolone use was associated with a significant increase in the MIC value of ofloxacin in canine and equine Staphylococcus isolates and current topical fluoroquinolone use was associated with significant increases in the MIC values of ciprofloxacin, moxifloxacin, and ofloxacin in canine Staphylococcus isolates., Conclusion: Patients previously or currently treated with topical fluoroquinolones, particularly in Staphylococcus infections, may require alternative antibiotics or additional antibiotic classes other than fluoroquinolones. Bacterial culture with MIC susceptibility testing should be highly recommended when a Staphylococcal infection is suspected., (© 2020 American College of Veterinary Ophthalmologists.)

Rapid transmission and spread of infectious pathogens are enhanced by the agricultural fair environment, where large numbers of livestock and people from numerous backgrounds congregate for several days. The transmission of influenza A virus and zoonotic enteric pathogens to fairgoers is a considerable risk and has occurred several times over the past decade. In an effort to mitigate zoonotic disease transmission in these settings, public health guidelines and recommendations including hand sanitation stations have been implemented. While hand hygiene recommendations to prevent the spread of zoonotic disease are well communicated, it is hypothesized that the adoption of these recommendations by agricultural fairs and fairgoers is low. To test this hypothesis, hand hygiene data collected from 658 agricultural fairs between 2012 and 2019 was analyzed to determine frequency and function of hand sanitation stations at the fairs, as well as utilization of educational signage. In addition, an observational study was performed to calculate the proportion of fairgoers who use hand sanitation stations at the fair. Lastly, samples were taken from working hand sanitation stations present at the exits of livestock barns and tested for the presence of influenza A virus and antimicrobial resistant coliform bacteria. Hand sanitation stations were present at most fairs (77.4 %) as recommended, but only 142 out of 2021 (7.0 %) visitors were observed using the stations. Health risk signage was displayed at more than half of fairs while the proper wash procedure was displayed at less than half. No influenza A virus was detected on any of the hand sanitation stations, however antimicrobial resistant coliform bacteria were recovered from 75.5 % of the sampled hand sanitation stations. Fairs should employ educational material along with functional hand sanitation stations in order to promote hand hygiene at fairs. Stations should be maintained and cleaned often to ensure effectiveness, as hand hygiene continues to be recommended to fairgoers when exiting animal barns to reduce zoonotic disease transmission., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Wastewater flows from metropolitan areas, especially those with healthcare inputs, can serve as transport reservoirs for the dissemination of clinically-relevant antimicrobial resistant bacteria (ARB) such as carbapenem- (CR) and colistin-resistant (CoR) strains. Pulsed electric field (PEF) is an emerging wastewater management tool for reducing bacterial loads without generating environmentally harmful byproducts, but it's ability to reduce ARB and their genetic determinants is not well reported. We collected 86, 10-L raw wastewater influent samples from a large metropolitan wastewater treatment plant in Columbus, Ohio and subjected them to low (34 kV cm -1 for 67 μsec) and high (36 kV cm -1 for 89 μsec) PEF treatment. We quantified the PEF effectiveness by measuring concentrations of total coliform bacteria, CR and CoR bacteria, and the epidemic carbapenemase gene, bla KPC , before and after PEF treatment. Utilizing marginal linear regression models with generalized estimating equations, we observed that low and high PEF treatment resulted in a 1.94 (95% CI 2.06-1.81; P < 0.001) and 2.32 (95% CI 2.46-2.18; P < 0.001) log reduction of total coliform bacteria concentrations, respectively. Low and high PEF treatment produced similar log reductions between CR E. coli (2.01 (95% CI 2.15-1.86; P < 0.001); 2.14 (95% CI: 5.30-4.61; P < 0.001)) and CR Enterobacteriaceae concentrations (1.55 (95% CI 1.70-1.41; P < 0.001); 1.86 (95% CI 2.05-1.68; P < 0.001)), and resulted in a 1.15 log (95% CI 1.38-0.93, P < 0.001) and 1.28 log (95% CI 1.54-1.03, P < 0.001) reduction of absolute bla KPC concentrations. Log CoR E. coli concentrations were reduced by 2.47 (95% CI 2.78-2.15; P < 0.001) and 2.52 (95% CI 2.91-2.15; P < 0.001) and CoR Enterobacteriaceae by 2.24 (95% CI 2.52-1.95; P < 0.001) and 2.50 (95% CI 2.89-2.11; P < 0.001) following low and high PEF application. PEF can be applied for wastewater management as an independent treatment method, particularly at critical control points, such as an on-site management of wastewater from hospitals or other healthcare facilities, or in series with other conventional methods to reduce total bacterial loads and concentrations of clinically-relevant ARB., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The authors declare no conflict of interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Background: Biochemistry analyzers in many high-throughput laboratories use indirect potentiometry to determine serum electrolyte concentrations, which involves a pre-analytical dilution step that may be associated with artifactual increases or decreases in electrolyte concentrations under circumstances of altered serum water fraction (SWF). Severe hypo- and hyperproteinemia, conditions that cause altered SWF, are recognized but under-emphasized causes of falsely measured serum sodium concentrations., Objectives: The goals of this study were to determine the average actual SWF (SWF A ) and establish formulae to correct serum sodium concentration measured by indirect potentiometry in hypo- and hyperproteinemic cats., Methods: Serum samples from 112 feline patients were analyzed for electrolytes (measured by both indirect and direct potentiometry), total protein, albumin, triglycerides, and cholesterol. Each serum sample was also lyophilized to determine the SWF A . A feline-specific formula to estimate SWF (SWF E-FEL ) was developed and evaluated with a multivariable linear model., Results: The mean SWF A in this population of cats was 91.2%, which was significantly different (P < .0001) than the mean (93.9%) calculated using the human estimated formula (SWF E-HUM ). The formula devised for the SWF E-FEL better recapitulated the SWF A than did the SWF E-HUM , and the corrected sodium concentrations calculated using the feline formula were better correlated with serum sodium measured by direct potentiometry than those determined using the human formula., Conclusions: Application of feline-specific formulae is expected to limit the misinterpretation of electrolyte data from indirect potentiometry when altered SWF occurs. To demonstrate this, a case example of a hypoproteinemic cat is provided., (© 2020 American Society for Veterinary Clinical Pathology.)

Transrectal palpation of the reproductive tract is the most common method for pregnancy determination in cattle and is considered a veterinary skill that new veterinary medicine (DVM) graduates should perform proficiently. However, using privately owned cattle to train students can be difficult because producers may believe that transrectal palpation by inexperienced students increases the risk of pregnancy wastage compared with examination by an experienced clinician. We used a randomized field trial of 1,216 healthy Holstein and Jersey cattle in 2 commercial dairy herds to estimate the effect of veterinary student transrectal palpation on early pregnancy loss. All cattle were determined to be pregnant using transrectal ultrasonography at approximately 37 d after artificial insemination. Cattle were then allocated into 2 groups based upon their ear tag number (study group = 598; control group = 618). Cattle in the study group were immediately palpated after ultrasonography by a fourth-year veterinary student, whereas control cattle were not subject to any additional pregnancy assessment. For analysis, the student palpators were divided into 2 groups: students who had previously had formal palpation training via an elective bovine palpation class (n = 30) and students who had not had palpation training (n = 134). All cattle were reevaluated using transrectal ultrasonography approximately 70 d after artificial insemination. A total of 53 (4.36%) animals lost their pregnancy between the first and second pregnancy assessments. Of the animals that lost their pregnancy, 26 (4.35%) were study group cows and 27 (4.37%) were control cows. Of the 26 cows documented to have had pregnancy loss within the study group, 20 out of 378 (5.3%) had been palpated by students who had not taken the palpation elective and 6 out of 220 (2.7%) had been palpated by students who had completed the elective. We found no difference in pregnancy loss between student-palpated and clinician-ultrasounded cattle, supporting the safety of using privately owned animals for student bovine palpation and pregnancy diagnosis training without affecting early pregnancy loss., (Copyright © 2019 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Carbapenemase-producing bacteria (CPB) are rare, multidrug resistant organisms most commonly associated with hospitalized patients. Metropolitan wastewater treatment plants (WWTP) treat wastewater from large geographic areas which include hospitals and may serve as epidemiologic reservoirs for the maintenance or expansion of CPB that originate from hospitals and are ultimately discharged in treated effluent. However, little is known about the potential impact of these WWTP CPB on the local surface water and their risk to the public health. In addition, CPB that are present in surface water may ultimately disseminate to intensively-managed animal agriculture facilities where there is potential for amplification by extended-spectrum cephalosporins. To better understand the role of WWTPs in the dissemination of CPB in surface waters, we obtained samples of treated effluent, and both upstream and downstream nearby surface water from 50 WWTPs throughout the US. A total of 30 CPB with clinically-relevant genotypes were recovered from 15 WWTPs (30%) of which 13 (50%) serviced large metropolitan areas and 2 (8.3%) represented small rural populations (P < 0.05). Recovery of CPB was lowest among WWTPs that utilized ultraviolet radiation for primary disinfection (12%), and higher (P = 0.11) for WWTPs that used chlorination (42%) or that did not utilize disinfection (50%). We did not detect a difference in CPB recovery by sampling site, although fewer CPB were detected in upstream (8%) compared to effluent (20%) and downstream (18%) samples. Our results indicate that WWTP effluent and nearby surface waters in the US are routinely contaminated with CPB with clinically important genotypes including those producing Klebsiella pneumoniae carbapenemase (KPC) and New Delhi metallo-beta-lactamase (NDM). This is a concern for both public health and animal agriculture because introduction of CPB into intensively managed livestock populations could lead to their amplification and foodborne dissemination., Competing Interests: The authors have declared that no competing interests exist.

Companion animals are likely relevant in the transmission of antimicrobial-resistant bacteria. Enterobacter xiangfangensis sequence type 171 (ST171), a clone that has been implicated in clusters of infections in humans, was isolated from two dogs with clinical disease in Ohio. The canine isolates contained IncHI2 plasmids encoding bla KPC-4 Whole-genome sequencing was used to put the canine isolates in phylogenetic context with available human ST171 sequences, as well as to characterize their bla KPC-4 plasmids., (Copyright © 2018 American Society for Microbiology.)

The dissemination of Enterobacteriaceae expressing resistance to clinically important antibiotics such as extended-spectrum cephalosporins (ESC), carbapenems, and fluoroquinolones is of critical concern to both human and veterinary medicine. In healthcare facilities, the movement of patients, personnel, and equipment provides an opportunity for Enterobacteriaceae carrying antibiotic resistance genes to disseminate in the hospital environment where they pose a threat to patients, staff, and hospital visitors. We collected environmental samples using electrostatic cloths to estimate the frequency of resistant Enterobacteriaceae contamination on human and veterinary hospital surfaces. Samples were enriched in nutrient broth modified with antibiotics to provide specific selection pressure to select for bacteria expressing AmpC β-lactamase (bla CMY ), extended-spectrum β-lactamase (ESBL, bla CTX-M ), carbapenemase, quinolone, and fluoroquinolone-resistant phenotypes. A total of 31 contact surfaces were sampled at each of five visits to two human hospitals. In addition, 34 surfaces were sampled at each of three visits to a veterinary hospital serving both farm animal and companion animal patients. Isolates expressing the bla CMY phenotype were recovered from 0.6%, 100%, and 18.2% of human hospitals, veterinary farm animal, and veterinary companion animal hospital surfaces, respectively. Isolates expressing the bla CTX-M phenotype were recovered from 0.6%, 55%, and 16.7% of human hospitals, farm animal, and companion animal veterinary hospital surfaces, respectively. Carbapenemase-producing Enterobacteriaceae (CPE) were detected from 1.3% of human hospital surfaces, but none were recovered from the veterinary hospital. Fluoroquinolone resistance was detected in 0.6%, 5.0%, and 37.9% of human hospitals, farm animal, and companion animal veterinary hospital surfaces, respectively. Our results indicate that ESC and fluoroquinolone-resistant Enterobacteriaceae can contaminate surfaces in both human and veterinary medical settings, with higher prevalence observed in veterinary hospitals, although the recovery of CPE from human hospital environments is concerning. Hospital medical trolleys or carts may serve as fomites for the dissemination of clinically relevant resistant bacteria.

Antimicrobial resistant bacteria in retail meat pose a health hazard to the public, as does contamination of these products with Salmonella. Our aim was to determine the prevalence of Salmonella as well as Escherichia coli expressing AmpC and extended-spectrum beta-lactamase (ESBL) resistance phenotypes contaminating broiler transport cages and fresh, retail ground chicken meat. Sterile gauze sponges were used to collect duplicate cage floor samples from transport trailers that deliver market-ready birds to a single organic poultry-processing facility. With the exception of the first visit (n = 25), 50 duplicate cage floor samples were collected using moistened sterile gauze sponges on each of nine weekly visits during May, June, and July 2013. Additionally, fresh, retail ground chicken meat was sampled at each weekly visit from an on-site retail store located at the same processing facility. A total of 425 cage swabs and 72 ground chicken aliquots from 24 retail packages were collected and screened for the presence of Salmonella as well as E. coli expressing resistance to extended-spectrum cephalosporins using selective culture. We recovered Salmonella from 26.1% of cage swab samples and 2.8% of retail meat samples. E. coli expressing AmpC and ESBL resistance phenotypes were recovered from 84.9% and 22.6% of cage swabs and 77.8% and 11.1% of fresh, retail ground meat samples, respectively. Our results suggest that transport cages could potentially act as a source of broiler exposure to both Salmonella and enteric bacteria resistant to important antimicrobial drugs as they are transported for entry into the food supply as fresh, retail meat products.

OBJECTIVE To determine whether brachycephalic dogs were at greater risk of anesthesia-related complications than nonbrachycephalic dogs and identify other risk factors for such complications. DESIGN Retrospective cohort study. ANIMALS 223 client-owned brachycephalic dogs undergoing general anesthesia for routine surgery or diagnostic imaging during 2012 and 223 nonbrachycephalic client-owned dogs matched by surgical procedure and other characteristics. PROCEDURES Data were obtained from the medical records regarding dog signalment, clinical signs, anesthetic variables, surgery characteristics, and complications noted during or following anesthesia (prior to discharge from the hospital). Risk of complications was compared between brachycephalic and nonbrachycephalic dogs, controlling for other factors. RESULTS Perianesthetic (intra-anesthetic and postanesthetic) complications were recorded for 49.1% (n = 219) of all 446 dogs (49.8% [111/223] of brachycephalic and 48.4% [108/223] of nonbrachycephalic dogs), and postanesthetic complications were recorded for 8.7% (39/446; 13.9% [31/223] of brachycephalic and 3.6% [8/223] of nonbrachycephalic dogs). Factors associated with a higher perianesthetic complication rate included brachycephalic status and longer (vs shorter) duration of anesthesia; the risk of perianesthetic complications decreased with increasing body weight and with orthopedic or radiologic procedures (vs soft tissue procedures). Factors associated with a higher postanesthetic complication rate included brachycephalic status, increasing American Society of Anesthesiologists status, use of ketamine plus a benzodiazepine (vs propofol with or without lidocaine) for anesthetic induction, and invasive (vs noninvasive) procedures. CONCLUSIONS AND CLINICAL RELEVANCE Controlling for other factors, brachycephalic dogs undergoing routine surgery or imaging were at higher risk of peri- and postanesthetic complications than nonbrachycephalic dogs. Careful monitoring is recommended for brachycephalic dogs in the perianesthetic period.

Antimicrobial-resistant bacteria represent an important concern impacting both veterinary medicine and public health. The rising prevalence of extended-spectrum beta-lactamase (ESBL), AmpC beta-lactamase, carbapenemase (CRE) and fluoroquinolone-resistant Enterobacteriaceae continually decreases the efficiency of clinically important antibiotics. Moreover, the potential for zoonotic transmission of antibiotic-resistant enteric bacteria increases the risk to public health. Our objective was to estimate the prevalence of specific antibiotic-resistant bacteria on human contact surfaces in various animal environments. Environmental surface samples were collected from companion animal shelters, private equine facilities, dairy farms, livestock auction markets and livestock areas of county fairs using electrostatic cloths. Samples were screened for Enterobacteriaceae expressing AmpC, ESBL, CRE or fluoroquinolone resistance using selective media. Livestock auction markets and county fairs had higher levels of bacteria expressing both cephalosporin and fluoroquinolone resistance than did equine, dairy, and companion animal environments. Equine facilities harboured more bacteria expressing cephalosporin resistance than companion animal shelters, but less fluoroquinolone resistance. The regular use of extended-spectrum cephalosporins in livestock populations could account for the increased levels of cephalosporin resistance in livestock environments compared to companion animal and equine facilities. Human surfaces, as well as shared human and animal surfaces, were contaminated with resistant bacteria regardless of species environment. Detecting these bacteria on common human contact surfaces suggests that the environment can serve as a reservoir for the zoonotic transmission of antibiotic-resistant bacteria and resistance genes. Identifying interventions to lower the prevalence of antibiotic-resistant bacteria in animal environments will protect both animal and public health., (© 2018 Blackwell Verlag GmbH.)

Carbapenemase-producing Enterobacteriaceae (CPE) threaten both agriculture and public health. While carbapenems are restricted in food-producing animals, other β-lactams, such as ceftiofur, are frequently applied in livestock. While the relationship is not fully elucidated, ceftiofur use may provide selective pressure that promotes carbapenem resistance. Recently reported in U.S. livestock, plasmid-mediated CPE are also present in livestock in Europe and Asia. We previously reported the rare carbapenemase gene, bla IMP-64 , in the environment of a large farrow-to-finish swine operation. To better understand CPE in this swine production system, in 2016 we followed a cohort of 350+ pigs over 5 months from late sow gestation to the final finishing phase. We screened both environmental and fecal samples for CPE using our selective enrichment protocol, with resulting phenotypic CPE isolates further characterized. Of 55 environmental and 109 sow fecal samples collected from a farrowing barn on our initial visit, 35 (64%) environmental and 15 (14%) sow fecal samples yielded isolates of multiple Enterobacteriaceae species carrying the metallo-β-lactamase gene bla IMP-64 on an IncQ plasmid. The frequency of IMP-64-positive environmental (n = 32), sow fecal (n = 30), and piglet fecal swab (n = 120) samples was highest for all groups when the market pig cohort was between 1 and 10 days, with observed prevalence of 97%, 28%, and 18%, respectively. After weaning, bla IMP-64 was detected in a single environmental sample from a nursery pen, with no CPE recovered in the finishing phase. Used in U.S. swine production to treat and control disease, ceftiofur is administered to piglets on this farm at birth, with males receiving a second dose at castration (≈day 7). Once introduced into animal agriculture, the common use of ceftiofur may provide the selection pressure required for CPE dissemination throughout large, intensively managed food animal populations housed in animal-dense environments.

Objective: To identify abnormalities of the celiac artery (CA) and major branches in dogs and cats by computed tomography angiography (CTA)., Study Design: Multi-institutional retrospective case series., Animals: Two hundred fifty-four dogs and 13 cats., Methods: Abdominal CTA images from 2009 to 2017 were reviewed. Logistic regression models were used to evaluate the relationship between CA abnormalities and sex, age, size of dog, concurrent venous anomaly, or presence of gastrointestinal signs., Results: Abnormalities in the CA were observed in 32 animals (11.9%) including 9 with abnormal branching (3.4%) and 23 with CA compression (8.6%). A celiacomesenteric trunk was observed in 8 (2.9%; 6 dogs, 2 cats). The splenic artery originated from the cranial mesenteric artery in 1 dog; the hepatic arterial branches originated from the left gastric artery in another. Four out of 32 animals (12.5%) with an arterial anomaly had another vascular abnormality. Large breed dogs were more likely to have an arterial anomaly (OR 4.3, 95% CI: 1.18-15.5, P = .02) and 12 times more likely to have CA compression (OR 12.0, 95% CI: 1.4-97.7, P = .02) compared to small breed dogs. Dogs with CA compression were more likely to present for gastrointestinal signs (OR 3.6, 95% CI: 1.2-10.3, P = .01)., Conclusion: Anomalies of the celiac trunk are apparent on CTA and may impact surgical or image-guided intervention. Compression at the origin of the CA was apparent on imaging, similar to the median arcuate ligament syndrome in people, although the significance of this finding in dogs is unknown., (© 2017 The American College of Veterinary Surgeons.)

Every year, multiple outbreaks of salmonellosis in humans are linked to contact with mail-order chicks and ducks. The objective of this study was to describe the temporal changes in the prevalence of serovars, genotypes and antimicrobial resistance (AMR) phenotypes of non-typhoidal Salmonella (NTS) recovered from shipped boxes of mail-order hatchling poultry in the United States during 2013 to 2015. In each year, a sample of feed stores belonging to a single national chain participated in the study. The store employees submitted swabs or hatchling pads from hatchling boxes and shipment tracking information of the arriving boxes to the investigators. NTS was cultured from the samples and isolates were sent to the National Veterinary Services Laboratories (Ames, IA) for serotyping, pulsed-field gel electrophoresis (PFGE) and AMR phenotyping. The PFGE patterns of Salmonella serovars isolated from hatchling boxes were compared with those from human outbreaks of salmonellosis linked to live poultry contact. The box-level prevalence of NTS was significantly higher in 2015 compared to 2014. Also, the population of Salmonella serovars recovered in 2015 was more diverse and substantially different from those recovered in the previous two years. Of PFGE patterns recovered from hatchling boxes, seven distinct patterns in 2015, three in 2014 and four in 2013 were indistinguishable from the PFGE patterns of human outbreaks-associated strains in the respective years. Importantly, a significant positive correlation was found between the box-level prevalence of PFGE patterns and the number of human illnesses associated with the same patterns. Also, the proportion of multidrug-resistant isolates was higher in 2014 and 2015 compared to that in 2013. The results demonstrate that shipments of mail-order hatchling poultry are frequently contaminated with Salmonella genotypes indistinguishable from human outbreaks-associated strains each year, and control efforts at hatchery level are likely to have an important public health impact., (© 2017 Blackwell Verlag GmbH.)

In order to estimate the prevalence of AmpC- and ESBL β-lactamase-producing Enterobacteriaceae in the faecal flora of a healthy domestic canine population, faecal samples were obtained from healthy dogs receiving routine parasitology screening at the Ohio State University Veterinary Medical Center, between January 2013 and April 2013. Samples were screened for the presence of AmpC and ESBL β-lactamase phenotypes, and the clinically important genotypes, bla CMY and bla CTX -M , were confirmed via conventional PCR. Minimum inhibitory concentrations were determined for isolates and plasmids were characterized. Two hundred and twelve canine faecal samples were screened, of which 30 harboured isolates carrying the AmpC bla CMY , representing 14.2% of the population (95% CI: 9.4-18.9%). Nine samples harboured isolates that carried the ESBL bla CTX -M , representing 4.2% of the population (95% CI: 1.5-7.0%). Isolates containing bla CMY harboured multiple plasmid replicon types, while isolates containing bla CTX -M harboured few plasmid replicon types. Our results suggest that domestic dogs may serve as a reservoir for extended-spectrum cephalosporin resistance genes for other domestic animal populations as well as for their human companions. This represents a potential veterinary and public health risk that warrants further investigation and continued surveillance to ascertain the nature and extent of the risk. The high level of diversity of plasmid content among isolates harbouring bla CMY suggests broader dissemination relative to bla CTX -M isolates., (© 2017 Blackwell Verlag GmbH.)

In the US, nontyphoidal Salmonellae are a common foodborne zoonotic pathogen causing gastroenteritis. Invasive Salmonella infections caused by extended-spectrum cephalosporin resistant (ESCR) phenotypes are more likely to result in treatment failure and adverse health outcomes, especially in severe pediatric Salmonella infections where the extended-spectrum β-lactams are the therapy of choice. To examine the genetic and epidemiologic characteristics of ESCR Salmonellae which may enter the food chain, we characterized 44 ceftiofur-resistant Salmonella isolates from the National Animal Health Monitoring System (NAHMS) 2011 beef cattle feedlot health and management study. As part of the NAHMS Feedlot 2011 study, 5050 individual fecal samples from 68 large (1000+ head capacity) feedlots were cultured for Salmonella spp. The resulting 460 positive samples yielded 571 Salmonella isolates with 44 (8%) expressing an AmpC β-lactamase phenotype. These phenotypic bla CMY-2 Salmonella isolates represented 8 serotypes, most commonly S. Newport (n=14, 32%), S. Typhimurium (n=13, 30%), and S. Reading (n=5, 11%), followed by S. Dublin, S. Infantis, S. Montevideo, S. Rough O:i;v:1;7, and S. Uganda. Carriage of the bla CMY-2 gene was confirmed for all isolates expressing an AmpC β-lactamase phenotype by PCR. Additionally, all 44 isolates were shown to carry the bla CMY-2 gene on a large IncA/C plasmid, a gene/plasmid combination which has been previously reported in multiple species. Other plasmids, including IncN, FIC, and FIIA, were also detected in some isolates. Cattle fed chlortetracycline were less likely to be positive for a bla CMY-2 Salmonella isolate in their enteric flora compared to those not receiving chlortetracycline during the feeding period. Carriage of bla CMY-2 was more prevalent in Salmonella isolates originating from lighter weight cattle, cattle fed tylosin and dairy breeds. Our characterization of the NAHMS Feedlot 2011 study Salmonella isolates with ESCR phenotype shows that while other cephalosporin resistance mechanisms have been reported in US cattle, specific serotypes harboring bla CMY-2 on IncA/C plasmids may be the dominant resistance genotype., (Copyright © 2017 Elsevier B.V. All rights reserved.)

We tested 772 cloacal swabs from wild ducks to estimate the prevalence of enteric bacteria resistant to extended-spectrum cephalosporins (ESC). We found a low prevalence of the important ESC resistance genotypes, bla CMY (5.7%) and bla CTX-M (0.3%). This suggests a minor role for wild ducks in the movement of resistant bacteria in the environment.

The emergence of bacterial resistance to carbapenem antibiotics is an urgent public health threat. Carbapenem drugs are a last resort treatment option for life-threatening infections. The frequent use of broad-spectrum antibiotics to treat hospitalized patients provides significant selection pressure favoring the emergence and dissemination of resistant organisms, including carbapenem-resistant Enterobacteriaceae (CRE). CREs have been reported in animal populations, but only rarely in horses. Our objective was to determine the prevalence of CRE in the environment of a referral equine specialty hospital. Environmental samples were collected on seven different sampling dates. Four clonal carbapenemase-producing Aeromonas veronii were recovered from 315 sampled surfaces (1.3%). All four isolates harbored the carbapenemase-producing, metallo-β-lactamase gene bla cphA , although corresponding minimum inhibitory concentrations were within the susceptible range for imipenem and meropenem. All had an identical multilocus sequence type with a previously unreported allelic profile and contained multiple plasmids. To our knowledge, this recovery of bla cphA -harboring A. veronii is the first report of carbapenemase-producing bacteria in the environment of an equine veterinary hospital. However, the low recovery rate suggests that environmental contamination is uncommon. Appropriate hospital cleaning and disinfection protocols are necessary to maintain a low risk of contamination for patients and personnel.

Extended-spectrum β-lactamases (ESBLs) confer bacterial resistance to critically important antimicrobials, including extended-spectrum cephalosporins (ESCs). Livestock are important reservoirs for the zoonotic food-borne transmission of ESC-resistant enteric bacteria. Our aim is to describe the potential role of migratory and resident wild birds in the epidemiology of ESBL-mediated bacterial resistance on dairy farms. Using mist nets, we sampled wild migratory and resident birds either immediately adjacent to or 600 ft away from free-stall barns on three Ohio dairy farms during the 2014 and 2015 spring migrations. Individual swabs were used to obtain both a cloacal and external surface swab from each bird. Samples were inoculated into MacConkey broth containing cefotaxime then inoculated onto MacConkey agar with cefoxitin, cefepime, or meropenem to identify the bla CMY, bla CTX-M, and carbapenemase phenotypes, respectively. Six hundred twenty-three birds were sampled, 19 (3.0%) of which harbored bacteria with bla CMY and 32 (5.1%) harbored bacteria with bla CTX-M from either their cloacal sample or from their external swab. There was no difference in the prevalence of either gene between migratory and resident birds. Prevalence of bla CMY and bla CTX-M was higher among birds sampled immediately outside the barns compared with those sampled 600 ft away. Our results suggest that wild birds can serve as mechanical and/or biological vectors for Enterobacteriaceae with resistance to ESCs. Birds live in close contact with dairy cows and their feed, therefore, transmission locally between farms is possible. Finding a similar prevalence in migratory and nonmigratory birds suggests the potential for regional and intercontinental movement of these resistance genes via birds.

The dissemination of Enterobacteriaceae expressing resistance to extended-spectrum cephalosporins, which are therapeutically used in both human and veterinary medicine, is of critical concern. The normal commensal flora of food animals may serve as an important reservoir for the zoonotic food-borne transmission of Enterobacteriaceae harbouring β-lactam resistance. We hypothesized that the predominant AmpC and ESBL genes reported in US livestock and fresh retail meat products, bla CMY -2 and bla CTX -M , would also be predominant in human enteric flora. We recovered enteric flora from a convenience sample of patients included in a large tertiary medical centre's Clostridium difficile surveillance programme to screen for and estimate the frequency of carriage of AmpC and ESBL resistance genes. In- and outpatient diarrhoeic submissions (n = 692) received for C. difficile testing at the medical centre's clinical diagnostic laboratory from July to December, 2013, were included. Aliquoted to a transport swab, each submission was inoculated to MacConkey broth with cefotaxime, incubated at 37°C and then inoculated to MacConkey agars supplemented with cefoxitin and cefepime to select for the AmpC and ESBL phenotypes, with bla CMY and bla CTX -M genotypes confirmed by PCR and sequencing. From the 692 diarrhoeic submissions, our selective culture yielded 184 isolates (26.6%) with reduced susceptibility to cefotaxime. Of these, 46 (6.7%) samples harboured commensal isolates carrying the AmpC bla CMY . Another 21 (3.0%) samples produced isolates harbouring the ESBL bla CTX -M : 19 carrying CTX-M-15 and 2 with CTX-M-27. Our results indicate that β-lactam resistance genes likely acquired through zoonotic food-borne transmission are present in the enteric flora of this hospital-associated population at lower levels than reported in livestock and fresh food products., (© 2016 Blackwell Verlag GmbH.)

Carbapenem-resistant Enterobacteriaceae (CRE) present an urgent threat to public health. While use of carbapenem antimicrobials is restricted for food-producing animals, other β-lactams, such as ceftiofur, are used in livestock. This use may provide selection pressure favoring the amplification of carbapenem resistance, but this relationship has not been established. Previously unreported among U.S. livestock, plasmid-mediated CRE have been reported from livestock in Europe and Asia. In this study, environmental and fecal samples were collected from a 1,500-sow, U.S. farrow-to-finish operation during 4 visits over a 5-month period in 2015. Samples were screened using selective media for the presence of CRE, and the resulting carbapenemase-producing isolates were further characterized. Of 30 environmental samples collected from a nursery room on our initial visit, 2 (7%) samples yielded 3 isolates, 2 sequence type 218 (ST 218) Escherichia coli and 1 Proteus mirabilis, carrying the metallo-β-lactamase gene bla IMP-27 on IncQ1 plasmids. We recovered on our third visit 15 IMP-27-bearing isolates of multiple Enterobacteriaceae species from 11 of 24 (46%) environmental samples from 2 farrowing rooms. These isolates each also carried bla IMP-27 on IncQ1 plasmids. No CRE isolates were recovered from fecal swabs or samples in this study. As is common in U.S. swine production, piglets on this farm receive ceftiofur at birth, with males receiving a second dose at castration (≈day 6). This selection pressure may favor the dissemination of bla IMP-27 -bearing Enterobacteriaceae in this farrowing barn. The absence of this selection pressure in the nursery and finisher barns likely resulted in the loss of the ecological niche needed for maintenance of this carbapenem resistance gene., (Copyright © 2017 American Society for Microbiology.)

Multistate outbreaks of salmonellosis associated with live poultry contact have been occurring with increasing frequency. In 2013, multistate outbreaks of salmonellosis were traced back to exposure to live poultry, some of which were purchased at a national chain of farm stores (Farm store chain Y). This study was conducted at 36 stores of Farm store chain Y and was concurrent with the timing of exposure for the human outbreaks of salmonellosis in 2013. We used environmental swabs of arriving shipment boxes of hatchling poultry and shipment tracking information to examine the distribution, diversity and anti-microbial resistance of non-typhoidal Salmonella (NTS) across farm stores and hatcheries. Isolates recovered from shipment boxes underwent serotyping, anti-microbial resistance (AMR) testing and pulsed-field gel electrophoresis (PFGE). Postal service tracking codes from the shipment boxes were used to determine the hatchery of origin. The PFGE patterns were compared with the PFGE patterns of NTS causing outbreaks of salmonellosis in 2013. A total of 219 hatchling boxes from 36 stores in 13 states were swabbed between 15 March 2013 and 18 April 2013. NTS were recovered from 59 (27%) of 219 hatchling boxes. Recovery was not significantly associated with species of hatchlings, number of birds in the shipment box, or the presence of dead, injured or sick birds. Four of the 23 PFGE patterns and 23 of 50 isolates were indistinguishable from strains causing human outbreaks in 2013. For serotypes associated with human illnesses, PFGE patterns most frequently recovered from shipment boxes were also more frequent causes of human illness. Boxes positive for the same PFGE pattern most frequently originated from the same mail-order hatchery. Only one of 59 isolates was resistant to anti-microbials used to treat Salmonella infections in people. This study provides critical information to address recurrent human outbreaks of salmonellosis associated with mail-order hatchling poultry., (© 2014 Blackwell Verlag GmbH.)

We investigated the implied health benefits of retail chicken breast labeled as "organic" or "antibiotic-free" when compared to conventional products based on frequency of contamination by Salmonella spp., Campylobacter spp., and coliform bacteria resistant to fluoroquinolones, extended-spectrum cephalosporins, or carbapenems. We purchased 231 prepackaged chicken breasts from 99 grocery stores representing 17 retail chains in Ohio, Michigan, and Pennsylvania from June to September 2012. Ninety-six (41.5%) packages were labeled "antibiotic free" and 40 (17.3%) were labeled "organic," with the remaining 95 (41.1%) making neither label claim. Salmonella were recovered from 56 (24.2%) packages, and the recovery rate was not different between product types. Five percent of packages contained Salmonella carrying the extended-spectrum cephalosporin resistance gene bla(CMY-2), representing 21.4% of Salmonella isolates. Campylobacter spp. were recovered from 10.8% of packages, with observed recovery rates similar for the three product types. Using selective media, we recovered Escherichia coli harboring bla(CMY-2) from over half (53.7%) of packages, with similar recovery rates for all product types. In addition, we recovered E. coli carrying bla(CTX-M) from 6.9% of packages, and E. coli with QRDR mutations from 8.2% of packages. Fluoroquinolone-resistant E. coli recovered using selective media were more common (p<0.05) in conventional (18.9%) compared to organic (0) and antibiotic-free (2.1%) packages. Our results indicate that, regardless of product type, fresh retail chicken breast is commonly contaminated with enteric pathogens associated with foodborne illness and commensal bacteria harboring genes conferring resistance to critically important antimicrobial drugs.

We report the recovery of Escherichia coli or Klebsiella pneumoniae containing the extended-spectrum β-lactamase gene bla(CTX-M) from 24 of 1,495 (1.6%) swine fecal samples in 8 of 50 (16%) finishing barns located in 5 U.S. states. We did not detect an association between antimicrobial use and recovery of bla(CTX-M).

We report the detection of Salmonella carrying bla(CTX-M) in U.S. livestock populations. We identified 12 of 2,034 (0.6%) Salmonella isolates originating from turkeys, horses, and pigs from at least 6 U.S. states, all carrying bla(CTX-M-1), many on a pandemic sequence type 1 IncN plasmid.

bla(CTX-M) beta-lactamases confer resistance to critically important cephalosporin drugs. Recovered from both hospital- and community-acquired infections, bla(CTX-M) was first reported in U.S. livestock in 2010. It has been hypothesized that veterinary use of cephalosporins in livestock populations may lead to the dissemination of beta-lactamase-encoding genes. Therefore, our objectives were to estimate the frequency and distribution of coliform bacteria harboring bla(CTX-M) in the fecal flora of Ohio dairy cattle populations. In addition, we characterized the CTX-M alleles carried by the isolates, their plasmidic contexts, and the genetic diversity of the bacterial isolates themselves. We also evaluated the association between ceftiofur use and the likelihood of recovering cephalosporinase-producing bacteria. Thirty fresh fecal samples and owner-reported ceftiofur use data were collected from each of 25 Ohio dairy farms. Fecal samples (n = 747) yielded 70 bla(CTX-M)-positive Escherichia coli isolates from 5/25 herds, 715 bla(CMY-2) E. coli isolates from 25/25 herds, and 274 Salmonella spp. from 20/25 herds. The within-herd prevalence among bla(CTX-M)-positive herds ranged from 3.3 to 100% of samples. Multiple pulsed-field gel electrophoresis (PFGE) patterns, plasmid replicon types, and CTX-M genes were detected. Plasmids with CTX-M-1, -15, and -14 alleles were clonal by restriction fragment length polymorphism (RFLP) within herds, and specific plasmid incompatibility group markers were consistently associated with each bla(CTX-M) allele. PFGE of total bacterial DNA showed similar within-herd clustering, with the exception of one herd, which revealed at least 6 different PFGE signatures. We were unable to detect an association between owner-reported ceftiofur use and the probability of recovering E. coli carrying bla(CTX-M) or bla(CMY-2).

The objective of this study was to investigate the association between ceftiofur use policy in finishing swine barns and recovery of fecal Escherichia coli or Salmonella spp. resistant to ceftriaxone. The study population included 54 finishing swine barns from three companies located in North Carolina. The barns were each classified according to their reported therapeutic ceftiofur use rates of "Rare," "Moderate," and "Common." Fecal samples from the barns were cultured for the presence of E. coli and Salmonella spp. resistant to ceftriaxone using selective media designed to recover rare organisms expressing the AmpC β-lactamase phenotype. A total of 1899 swine fecal samples yielded 1193 E. coli (63%) resistant to ceftriaxone. Recovery rates by ceftiofur use classification were 45% for Rare, 73% for Moderate, and 68% Common ceftiofur use groups. Barns reporting Rare ceftiofur use had a lower odds of recovery of E. coli (OR=0.32; p<0.001) resistant to ceftriaxone compared to Common use barns. The overall Salmonella spp. prevalence was 63.8% (n=714). Of these, 65 Salmonella were resistant to ceftriaxone with the highest rate (6%) found in the Common ceftiofur use group, followed by Rare (4.1%) and Moderate (0.15%). The odds of recovery of Salmonella resistant to ceftriaxone were similar for barns with ceftiofur use classified as Rare and Common. Samples from barns with ceftiofur use classified as Moderate had a lower odds (OR=0.02; p<0.01) of recovery of Salmonella resistant to ceftriaxone than barns classified as Common. Our result is consistent with the hypothesis that the use of ceftiofur in finishing swine barns, beyond its rare application, may influence the recovery of enteric E. coli with resistance to cephalosporin drugs, although other unmeasured factors appear to be important in the recovery of cephalosporin-resistant Salmonella. The dissemination of enteric bacteria with resistance to cephalosporins has the potential to impact both veterinary and human therapeutic treatment options.

We estimated the proportion of retail beef and pork products containing bla(CMY)-mediated third-generation cephalosporin resistance in commensal Escherichia coli and Salmonella enterica. Samples were obtained from 50 grocery stores located in two U.S. states. From each store, 20 fresh meat products were purchased, including 7 packages of ground beef, 3 packages of beef steak, 6 packages of pork chops, and 4 packages of pork ribs. The resulting 1000 packages of fresh meat product were individually screened for the presence of E. coli or Salmonella harboring bla(CMY). Over 8% of all retail meat packages contained E. coli with bla(CMY), whereas 4% contained Salmonella and only 0.5% contained Salmonella with bla(CMY). Retail pork products more frequently yielded E. coli with bla(CMY) than did beef products (12.2% vs. 4.0%; p = 0.06). Salmonella were also recovered more frequently from pork than beef (5.8% vs. 2.4%; p < 0.01). In addition, all five Salmonella isolates with bla(CMY) were recovered from pork products. Our data suggest that enteric bacteria carrying bla(CMY) are frequently present in fresh retail meat products. However, we did not quantify the number of resistant bacteria present in these products, which makes the public health implications of this result unclear.

CTX-M extended-spectrum β-lactamases are enzymes produced by bacteria that are capable of inhibiting the antimicrobial effects of cephalosporin drugs. Recently, the first domestically acquired Salmonella in the United States expressing bla(CTX-M) was reported. This is a concern because expanded-spectrum cephalosporins are the treatment of choice for invasive Gram-negative infections, including salmonellosis in children. Because Salmonella transmission is primarily foodborne, there is also concern that resistant enteric bacteria from livestock can be transferred through the food supply chain to consumers. bla(CTX-M) has not been previously identified in bacterial isolates from food animal populations in the United States. We report the recovery of CTX-M-type extended-spectrum β-lactamases from fecal Escherichia coli of sick and healthy dairy cattle in Ohio. Four individual fecal samples yielded E. coli isolates representing three clonal strains that carried bla(CTX-M) on transferable plasmids. Two distinguishable plasmids were identified, each encoding bla(CTX-M-1) or bla(CTX-M-79). Transferrable bla(CTX-M) genes in bovine E. coli have the potential to serve as a reservoir of resistance for pathogens and may represent a public health concern.

The prophylactic use of intramammary antimicrobial drugs at the end of lactation in dairy cows, known as dry cow therapy (DCT), is widely practiced in US dairy herds. This extremely common use of high-dose, slow-release antimicrobials may influence the ecology of bacterial flora on dairy farms. We investigated the association between the antimicrobial used for intramammary DCT and the relative number of fecal coliform bacteria with reduced susceptibility to three antimicrobial drugs in dairy cattle. Most probable number (MPN) data were estimated from 463 individual fecal samples collected from lactating cows in 15 dairy herds in Ohio, USA. These data were used to calculate the relative number of fecal coliform bacteria with reduced susceptibility to cephalothin, streptomycin, and tetracycline for individual cow samples. The farms included in this project were classified based on DCT, with 8 farms using a cephalosporin-based product and the remaining 7 using a penicillin/streptomycin therapy. Results of a linear mixed model indicate that herds using a cephalosporin DCT had higher (P<0.01) relative numbers of fecal coliform bacteria with reduced susceptibility to cephalothin and streptomycin compared to those using a penicillin/streptomycin intramammary therapy. Relative numbers of fecal coliform bacteria with reduced susceptibility to tetracycline was not associated with DCT. These results suggest that high-dose slow-release antimicrobials applied locally in the udder to populations of dairy cows might influence the antimicrobial susceptibility of the enteric flora. However, the potential animal and public health implications of this result are not clear., (Copyright 2010 Elsevier B.V. All rights reserved.)