Prevalence of extended-spectrum cephalosporin-, carbapenem-, and fluoroquinolone-resistant members of the family Enterobacteriaceae isolated from the feces of horses and hospital surfaces at two equine specialty hospitals

Rachael J. Adams Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

Search for other papers by Rachael J. Adams in
Current site
Google Scholar
PubMed
Close
 MS
,
Dixie F. Mollenkopf Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

Search for other papers by Dixie F. Mollenkopf in
Current site
Google Scholar
PubMed
Close
 PhD
,
Dimitria A. Mathys Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

Search for other papers by Dimitria A. Mathys in
Current site
Google Scholar
PubMed
Close
 VMD, PhD
,
Andrea Whittle Rood & Riddle Equine Hospital, Lexington, KY 40511.

Search for other papers by Andrea Whittle in
Current site
Google Scholar
PubMed
Close
,
Greg A. Ballash Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

Search for other papers by Greg A. Ballash in
Current site
Google Scholar
PubMed
Close
 MPH, DVM
,
Margaret Mudge Department of Veterinary Clinical Sciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

Search for other papers by Margaret Mudge in
Current site
Google Scholar
PubMed
Close
 VMD
,
Joshua B. Daniels Department of Veterinary Clinical Sciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.
Veterinary Diagnostic Laboratories, Colorado State University, Fort Collins, CO 80526.

Search for other papers by Joshua B. Daniels in
Current site
Google Scholar
PubMed
Close
 DVM, PhD
,
Bonnie Barr Rood & Riddle Equine Hospital, Lexington, KY 40511.

Search for other papers by Bonnie Barr in
Current site
Google Scholar
PubMed
Close
 VMD
, and
Thomas E. Wittum Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

Search for other papers by Thomas E. Wittum in
Current site
Google Scholar
PubMed
Close
 PhD

Abstract

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.

Abstract

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.

Introduction

The increased use of antimicrobials in veterinary species provides selection pressure for the emergence and dissemination of antimicrobial resistance genes among bacteria and therefore antimicrobial resistance.1 Equine hospitals are infrequently surveyed for antimicrobial resistance patterns of bacteria isolated from horses, but equine hospitals may be an important reservoir for resistant bacteria. Antimicrobial-resistant bacteria in companion animals and livestock have been extensively studied,1,2,3,4,5 but similar studies of horses are few and primarily limited to Europe and Canada.6,7,8,9

Plasmid-mediated antimicrobial resistance genes within the gastrointestinal microbiota can be rapidly conferred through conjugation. Through conjugation, pathogens such as Salmonella spp of the family Enterobacteriaceae can acquire clinically relevant antimicrobial resistance.10,11 Also, commensal bacteria of the family Enterobacteriaceae, such as Escherichia coli and Klebsiella spp, can acquire antimicrobial-resistance genes and produce opportunistic infections. A 20-year study8 at a veterinary university bacteriology laboratory revealed that antimicrobial resistance was more common in gram-negative enteric bacteria, including E coli, isolated from horses. In some situations of life-threatening bacterial infection, cephalosporins and fluoroquinolones may be recommended as treatment, although increasing resistance to these antimicrobials may limit their efficacy.12 Various resistance genes of bacteria encode phenotypic resistance to cephalosporins, such as blaCMY, a plasmid-mediated AmpC-type β-lactamase gene,6 or blaCTX-M, a plasmid-mediated ESBL gene.13 One study14 revealed that cephalosporin administration appeared to have the greatest effect on the selection of antimicrobial-resistant phenotypes for bacteria isolated from horse feces. Several European studies13,15,16,17,18,19,20 have noted ESBL-producing bacteria as part of the gastrointestinal microbiota (through their isolation from horse feces), with a prevalence of 6.5% to 60.5%, suggesting widespread presence of plasmid-mediated antimicrobial resistance genes.16,17 Fluoroquinolones are useful for the treatment of osteomyelitis, septic arthritis, urinary tract infections, and pneumonia caused by gram-negative bacteria,21 but information regarding fluoroquinolone resistance of the gastrointestinal microbiota of horses is limited.22 Carbapenems are also used in horses but as a last resort for the treatment of aggressive infections (eg, neonatal sepsis) caused by bacteria that are resistant to other broad-spectrum antimicrobials.21 Carbapenem-resistant bacteria of the family Enterobacteriaceae can cause fatal infections in people,23 are considered as an urgent public health threat by the CDC, and have also been reported in companion animals24 and livestock25 but not in horses within the United States. A carbapenemase-producing genotype, oxa23, has been identified in 2 Acinetobacter spp isolated from the feces of hospitalized horses in Belgium.20 Although CRE are likely rare in horses, surveillance for the emergence of CRE in horses is critical for limiting the dissemination of resistance genes. Disparities in the prevalence of various CRE genotypes between unrelated horse populations may be present yet currently unidentified; for example, distinct differences in carbapenemase genes among CRE isolated from people have been observed between countries and continents.26

Hospitals may be reservoirs for the dissemination of resistant bacteria among people. A review article27 indicated an association between antimicrobial use and increased presence of nosocomial pathogens that were resistant to various antimicrobials. Hospitalization and antimicrobial use have been associated with the presence of antimicrobial-resistant bacteria in the gastrointestinal microbiota of horses.13,14,18,19 Veterinarians at tertiary-care equine hospitals may be more likely to prescribe antimicrobials that are considered to be of last resort, such as the carbapenems, which may select for resistant bacterial strains such as CRE. Differences in the horse populations and length of hospital stay may also contribute to the types of resistant bacteria that colonize a horse's gastrointestinal tract and contaminate hospital surfaces.

The primary objective of the study presented here was to estimate the prevalence of extended-spectrum cephalosporin-, carbapenem-, and fluoroquinolone-resistant bacteria (eg, E coli, Klebsiella spp, Enterobacter spp) of the family Enterobacteriaceae in the feces of hospitalized horses and on equine hospital surfaces in the United States. An additional objective was to evaluate how antimicrobial-use practices, horse signalment, and hospital layout (floorplan) affected the prevalence of antimicrobial-resistant bacteria. These data were expected to improve our understanding of the effect of hospitalization on the fecal carriage of antimicrobial-resistant Enterobacteriaceae bacteria in horses and on hospital contamination.

Materials and Methods

Animals

Horses hospitalized for ≥ 48 hours at OSUGEC or KYEH with 2 fecal samples collected at different times between May 2015 and June 2016 were eligible for enrollment. A total of 143 horses, 46 from OSUGEC and 97 from KYEH, were enrolled from a convenience sample on the basis of the investigators’ availability to collect fresh feces. Fifty horses (35%) were < 1 year of age, 24 (17%) were ≥ 1 to < 2 years, 36 (25%) were ≥ 2 to < 11 years, and 33 (23%) were ≥ 11 years. Most horses were sexually intact females (n = 103 [72%]); the remainder were males (sexually intact, 24 [17%]; and gelded, 16 [11%]). Of the mares, 59% were without foals, 31% were with foals, and 10% were pregnant. The majority of horses were Thoroughbreds (80%; non-Thoroughbreds, 20%). Horses presented because of gastrointestinal illness (33%), reproductive disease (17%), a wound (9%), and other (eg, corneal ulceration, abscess, diagnostic imaging, respiratory disease, lameness, or neurologic disease; 42%). Beginning on the day of hospital admission, nearly 50% of horses received a combination of potassium penicillin and gentamicin; 43% received formulations of an aminoglycoside, amphenicol, cephalosporin, fluoroquinolone, macrolide, sulfonamide, or tetracycline. Approximately 8% did not receive an antimicrobial. Duration of antimicrobial administration varied, with 0 days for 14 (10%) horses, 1 to 3 days for 33 (23%), 4 to 6 days for 77 (54%), and ≥ 7 days for 19 (13%).

Fecal and environmental sampling

A convenience fecal sample was collected from hospitalized horses at 3 time points: within 24 hours of hospital admission (T0), 48 hours after hospital admission (T1), and 3 to 7 days after hospital admission (T2). If a horse was hospitalized for < 3 days, no T2 sample was collected. Two dual-tipped transport swabsa were used to collect fresh feces (estimated 0.5 g of feces collected within 4 hours of defecation) from the center of 1 fecal pile.

Environmental samples were collected with electrostatic cloths from surfaces commonly contacted by hospital personnel or hospitalized horses. These hospitals were selected because of convenience and their focus on equine medicine, their large case load, and the ability of their personnel to collaborate on this study. Environmental surfaces were selected to be representative of each hospital, with a focus on commonly contacted surfaces in high-traffic areas. At OSUGEC, samples were collected from the ICU, orthopedic ward, general-purpose ward, isolation ward, and miscellaneous areas (eg, triage intake area, breezeway, and treadmill room). At KYEH, samples were collected from the general-purpose, gastrointestinal, elective and nonelective surgery, foal, and isolation barns, neonatal ICU, and maternity ward. Sampled environmental surfaces in these locations included but were not limited to stalls, floor drains, water spigots and hoses, preparation counters, cabinet handles, mucking equipment handles, feed room contents, drug carts, office spaces, pharmacies, storage room contents, gurneys, tables, lead ropes, and muzzles. Two electrostatic cloths were used to sample each surface, with a surface's area evenly divided between the 2 cloths. The same surfaces were repeatedly sampled 7 times from May 2015 to April 2016 at KYEH and 5 times from June 2015 to April 2016 at OSUGEC, with sample times distributed throughout the study period on the basis of hospital access and previous environmental culture results. Small surfaces (eg, drains and door and equipment handles) were entirely sampled, whereas representative portions of large surfaces (eg, stalls and surgical suites) were sampled, with a focus on those frequently contacted (eg, stalls and countertops).

One swab of feces and 1 electrostatic cloth were incubated for 18 to 24 hours (overnight) at 37°C (98.6°F) in 9 mL of MacConkey and 90 mL of nutrient broth, respectively, modified with 2 μg of cefotaxime/mL. Then, MacConkey agar plates modified with 8 μg of cefoxitin/mL, 4 μg of cefepime/mL, and 0.5 μg of meropenem/mL with 70 μg of ZnSO4/mL were inoculated to identify AmpC β-lactamase, ESBL, and CRE phenotypes, respectively. A swab saturated with the overnight cultured MacConkey or nutrient broth was used to inoculate the MacConkey agar plates. The second swab of feces and electrostatic cloth were also incubated for 18 to 24 hours in 9 mL of MacConkey and 90 mL of nutrient broth, respectively, at 37°C but instead modified with 16 μg of nalidixic acid/mL. Then, MacConkey agar plates modified with 16 μg of nalidixic acid/mL and 2 μg of ciprofloxacin/mL were inoculated with a swab of the inoculated MacConkey or nutrient broth to identify phenotypic fluoroquinolone-resistant Enterobacteriaceae bacteria. Agar plates were incubated at 37°C for 18 to 24 hours, and then up to 2 morphologically unique lactose-fermenting colonies from each agar plate were selected and stored for genotypic characterization of the resistance phenotype with use of previously described methods.4,25,28

Genotypic analysis

Lactose-fermenting isolates with the AmpC β-lactamse and ESBL phenotypes were further characterized with a standard DNA PCR assay that included previously reported4 primers to confirm resistance genotypes. Of those fecal and environmental bacterial isolates confirmed to have blaCTX-M, 10% of these isolates from fecal swabs and 10% of these isolates from the environment were further characterized to identify specific blaCTX-M alleles. Phenotypic CRE isolates were tested for carbapenemase production with the Carba NP test.29 For Carba NP–positive isolates, MICs of antimicrobials that were part of standardized laboratory panels were generated with the use of a semiautomated broth microdilution systemb that adhered to Clinical and Laboratory Standards Institute guidelines.30 Electrophoresis with a standard plasmid-profiling procedure was performed to determine plasmid size and content for the CRE isolates.31 Species of CRE, antimicrobial-resistance genes, and multilocus sequence types were identified through whole-genome sequencing.c Clonality of E coli isolates with the ESBL phenotype recovered from fecal and environmental samples was determined by following a standardized pulsed-field gel electrophoresis protocol.32 Also by pulsed-field gel electrophoresis, 5 ESBL-producing E coli isolates from the fecal samples collected at both hospitals were compared with corresponding temporal environmental isolates to determine whether the fecal isolates were related to the environmental isolates. Additionally, ESBL-producing E coli isolates recovered from 2 surfaces—the breezeway and a stall in the general-purpose ward—at OSUGEC over consecutive sampling dates were tested for clonality to determine whether the same bacterium persisted in the environment over time. The same analysis was performed on ESBL-producing E coli isolated from 3 surfaces—a stall in the general-purpose barn and the drains and feed room in the isolation barn—at KYEH.

Data analysis

Three marginal multivariable logistic regression models were constructed, fit with generalized estimating equations, to estimate differences in the odds of horses harboring an Enterobacteriaceae isolate resistant to 1 of 3 antimicrobials as follows: cefoxitin, cefepime, and ciprofloxacin. A forward model building approach was used to generate a parsimonious multivariable model.33 In brief, univariable analyses were conducted to identify independent variables to include in the multivariable analysis; variables were included on the basis of a P value ≤ 0.20. These variables were then included in the initial multivariable models and assessed for their inclusion by use of a P value < 0.05. Independent variables not included in the multivariable model on the basis of univariable and multivariable analyses were reintroduced into the revised multivariable model to assess for confounding. Independent variables that resulted in a ≥ 20% change in the multivariable coefficients were considered confounders and included in the analysis for adjustment. Signalment, pregnancy status, presenting complaint, antimicrobial administered, and duration of antimicrobial administration were evaluated as potential confounders. The models were fit to an exchangeable correlation matrix to adjust for repeated measurements for each horse. Robust SEs were calculated by use of the Huber-White sandwich estimator to account for any deviations from the predicted correlation matrix. All models considered that each horse could contribute unequal numbers of repeated time-point results to the analysis over time because of variable length of hospital stays. Statistical testing was performed with a commercially available software program.d

Results

Fecal sampling

From the 143 enrolled hospitalized horses, 358 fecal samples were collected. Ninety-four percent (n = 135) of horses had T1 samples and of those that had T1 samples, 56% (80) also had T2 samples. Enterobacteriaceae bacteria with AmpC β-lactamase and ESBL phenotypes and resistance to fluoroquinolones were isolated from 45% (n = 162), 44% (158), and 41% (148) of fecal samples, respectively. At OSUGEC, 38% (n = 42), 34% (37), and 30% (33) of 110 fecal samples had bacteria with those same resistance phenotypes, respectively. Fecal carriage proportions varied from 13% to 30% within 24 hours of admission (T0), depending on the antimicrobial resistance phenotype (Figure 1). Of the 248 fecal samples from hospitalized horses at KYEH, 48% (n = 120), 49% (121), and 46% (115) of fecal samples had Enterobacteriaceae bacteria with those same resistance phenotypes, respectively, and fecal carriage proportion from 26% to 32% within 24 hours of admission (T0).

Figure 1
Figure 1

Prevalence of Enterobacteriaceae bacteria with AmpC β-lactamase phenotype, ESBL phenotype, and fluoroquinolone resistance isolated from the feces of horses hospitalized at OSUGEC (A) and KYEH (B) within 24 hours of admission (T0; black bars), 48 hours after admission (T1; white bars), and between 3 and 7 days after admission (T2; gray bars).

Citation: Journal of the American Veterinary Medical Association 258, 7; 10.2460/javma.258.7.758

Antimicrobial-resistant Enterobacteriaceae bacteria were significantly more likely to be isolated from the feces of horses that were hospitalized for 48 hours (T1) and ≥ 3 to 7 days (T2), compared with the likelihood of being isolated from the feces within 24 hours of admission (T0), for all resistance phenotypes (Table 1). Horses hospitalized at KYEH (vs OSUGEC [reference]) were significantly (P = 0.022) more likely to have fluoroquinolone-resistant bacteria isolated from their feces (OR = 2.2; 95% CI, 1.1 to 4.2; Table 2). However, prevalence of extended-spectrum cephalosporin resistance was not significantly different between hospitals. Potential confounding factors such as signalment, pregnancy status, presenting complaint, antimicrobial administered, and duration of antimicrobial administration did not impact these results.

Table 1

Odds ratios for isolation of Enterobacteriaceae bacteria with AmpC β-lactamase phenotype, ESBL phenotype, and fluoroquinolone resistance from the feces of horses hospitalized at OSUGEC and KYEH within 24 hours of admission (T0), 48 hours after admission (T1), and between 3 and 7 days after admission (T2), with T0 as reference.

Phenotype Time OR (95% CI) P value
AmpC β-lactamase T0 1.0
T1 4.2 (2.7–6.3) < 0.001
T2 2.8 (1.6–4.7) < 0.001
ESBL T0 1.0
T1 3.2 (2.2–4.7) < 0.001
T2 2.9 (1.9–4.7) < 0.001
Fluoroquinolone resistance T0 1.0
T1 4.0 (2.6–6.1) < 0.001
T2 3.8 (2.4–5.9) < 0.001

— = Not applicable.

Table 2

Odds ratios for isolation of Enterobacteriaceae bacteria with AmpC β-lactamase phenotype, ESBL phenotype, and fluoroquinolone resistance from the feces of horses hospitalized at OSUGEC and KYEH, with OSUGEC as reference.

Phenotype Hospital OR (95% CI) P value
AmpC β-lactamase OSUGEC 1.0
KYEH 1.4 (0.8–2.6) 0.227
ESBL OSUGEC 1.0
KYEH 1.8 (0.9–3.5) 0.093
Fluoroquinolone resistance OSUGEC 1.0
KYEH 2.2 (1.1–4.2) 0.022

— = Not applicable.

Environmental sampling

OSUGEC—Bacterial culture of samples collected from surfaces yielded Enterobacteriaceae bacteria that expressed AmpC β-lactamase (82% [135/164]), ESBL (64% [105/164]), and fluoroquinolone-resistant (55% [91/164]) phenotypes (Table 3). Carbapenem-resistant Enterobacteriaceae bacteria were not recovered from OSUGEC. Proportionally more antimicrobial-resistant Enterobacteriaceae bacteria were isolated from the general-purpose ward and ICU. Bacterial cultures of all samples collected from the floor drains (19/19), drug carts (5/5), technician office (5/5), breezeway (5/5), and triage intake area (5/5) at OSUGEC yielded growth of bacteria with the AmpC β-lactamase phenotype. All samples from the ICU-feed room (5/5) yielded growth of ESBL-producing and fluoroquinolone-resistant bacteria. Similarly, ESBL-producing Enterobacteriaceae bacteria were isolated from all breezeway (5/5) samples.

Table 3

Proportion of various locations in OSUGEC and KYEH in which surfaces in these locations were contaminated with Enterobacteriaceae bacteria that expressed AmpC β-lactamase or ESBL phenotypes, produced carbapenemase (CR), or had fluoroquinolone resistance.

Hospital Location AmpC β-lactamase phenotype (%) ESBL phenotype (%) CR (%) Fluoroquinolone resistance (%)
OSUGEC ICU 88 (44/50) 60 (30/50) 0 (0/50) 62 (31/50)
Orthopedic ward 80 (28/35) 57 (20/35) 0 (0/35) 46 (16/35)
General-purpose ward 89 (31/35) 71 (25/35) 0 (0/35) 71 (25/35)
Isolation 68 (13/19) 63 (12/19) 0 (0/19) 26 (5/19)
Miscellaneous* 76 (19/25) 72 (18/25) 0 (0/25) 56 (14/25)
Total 82 (135/164) 64 (105/164) 0 (0/164) 55 (91/164)
KYEH General-purpose barn 62 (29/47) 23 (11/47) 4 (2/47) 40 (19/47)
Gastrointestinal barn 34 (15/44) 7 (3/44) 2 (1/44) 23 (10/44)
Elective surgery barn 38 (15/39) 26 (10/39) 0 (0/39) 31 (12/39)
Neonatal ICU 72 (31/43) 53 (23/43) 0 (0/43) 26 (11/43)
Foal barn 76 (22/29) 45 (13/29) 3 (1/29) 34 (10/29)
Maternity ward 43 (23/54) 20 (11/54) 0 (0/54) 43 (23/54)
Isolation barn 71 (24/34) 85 (29/34) 0 (0/34) 74 (25/34)
Surgical suite 24 (6/25) 4 (1/25) 0 (0/25) 0 (0/25)
Total 52 (165/315) 32 (101/315) 1 (4/315) 35 (110/315)

Included triage intake area, radiology examination room, waiting room, breezeway, and orthopedic treadmill room.

CR bacteria isolated at KYEH were Aeromonas veronii (family Aeromonadaceae).

KYEH—Bacterial culture of samples collected from surfaces yielded Enterobacteriaceae bacteria that expressed AmpC β-lactamase (52% [165/315]) or ESBL (32% [101/315]) phenotypes or resistance to fluoroquinolones (35% [110/315]; Table 2). Unlike at OSUGEC, carbapenem-resistant bacteria (1% [4/315]) were recovered at KYEH. Described34 previously, 4 clonal Aeromonas veronii (family Aeromonadaceae) isolates with the chromosomally mediated carbapenemase gene blaCphA that was confirmed by whole-genome sequencing were isolated. All Aeromonas spp were characterized by low meropenem (≤ 1 μg/mL) and imipenem (≤ 1 μg/mL) MICs. Proportionally more cephalosporin- and fluoroquinolone-resistant Enterobacteriaceae bacteria were isolated from the isolation and foal barns and neonatal ICU. Proportionally fewer resistant Enterobacteriaceae bacteria were isolated from surfaces in the surgical suites. The floor drains had the highest prevalence of Enterobacteriaceae bacteria with antimicrobial resistance phenotypes (AmpC β-lactamase, 84% [31/37]; ESBL, 68% [25/37]; and fluoroquinolone resistance, 73% [27/37].) Feed-room surfaces also had a high prevalence of bacteria with resistance phenotypes (AmpC β-lactamase, 65% [15/23]; and ESBL, 39% [9/23]), whereas stalls had the second-highest prevalence of fluoroquinolone-resistant bacteria (41% [46/112]).

Evaluation of clonality between samples

Evaluation of E coli with the ESBL phenotype isolated from samples collected at OSUGEC revealed no clonal relation. However, 2 distinct clonal pairs (similarity > 95%) were identified at KYEH between 1 fecal sample and 1 environmental sample. Furthermore, 2 horses at KYEH concurrently had a clonal ESBL-producing E coli isolated from their feces at T1. Samples collected contemporaneously from various surfaces yielded multiple E coli clones at both hospitals. These clones did not persist on surfaces over consecutive sampling dates.

Genotypic analysis

Plasmid-mediated resistance genes were commonly identified. Of the 134 Enterobacteriaceae isolates collected from the environment at OSUGEC, 85.1% (114/134) of the isolates that expressed the AmpC β-lactamase phenotype had blaCMY, whereas 76.3% (119/156) of isolates at KYEH had blaCMY (Table 4). Of the OSUGEC and KYEH environmental isolates with the ESBL phenotype, 99.0% (102/103) and 91.9% (91/99), respectively, had blaCTX-M.

Table 4

Number and proportion of Enterobacteriaceae isolates with blaCMY coding for the AmpC β-lactamase phenotype or blaCTX-M coding for the ESBL phenotype from commonly contacted surfaces contaminated with Enterobacteriaceae bacteria at OSUGEC and KYEH.

Hospital No. with AmpC β-lactamase phenotype No. (%) with AmpC β-lactamase phenotype that had blaCMY* No. with ESBL phenotype No. (%) with ESBL phenotype that had blaCTX-M
OSUGEC 134 114 (85.1) 103 102 (99.0)
KYEH 156 119 (76.3) 99 91 (91.9)

One OSUGEC- and 9 KYEH-blaCMY isolates did not survive the freezing process to undergo genotypic analysis and therefore could not be included in the table.

Two OSUGEC- and 2 KYEH-blaCTX-M isolates did not survive the freezing process to undergo genotypic analysis and therefore could not be included in the table.

Lower prevalence of plasmid-mediated resistance genes was observed in isolates recovered from fecal samples. The gene blaCMY was identified in 62.5% (n = 25) of isolates from OSUGEC with the AmpC β-lactamase phenotype and 86.6% (103) of isolates from KYEH (Table 5). At OSUGEC, 100% (n = 30) of fecal Enterobacteriaceae bacteria with the ESBL phenotype had blaCTX-M, whereas at KYEH, blaCTX-M was identified in 94.8% (110/116). Further characterization of blaCTX-M for an approximately 10% subset (n = 30) of fecal and environmental samples indicated a mix of genotypes: blaCTX-M-55 (12), blaCTX-M-1 (9), blaCTX-M-15 (7), and blaCTX-M-28 (2). All isolates with blaCTX-M-1 (9) were from OSUGEC.

Table 5

Number and proportion of Enterobacteriaceae with blaCMY coding for the AmpC β-lactamase phenotype or blaCTX-M coding for the ESBL phenotype isolated from the feces of horses hospitalized at OSUGEC and KYEH within 24 hours of admission (T0), 48 hours after admission (T1), and between 3 and 7 days after admission (T2).

Hospital Time No. with AmpC β-lactamase phenotype No. (%) with AmpC β-lactamase phenotype that had blaCMY* No. with ESBL phenotype No. (%) with ESBL phenotype that had blaCTX-M
OSUGEC T0 14 9 (64.3) 7 7 (100)
T1 21 13 (61.9) 16 16 (100)
T2 5 3 (60.0) 7 7 (100)
Total 40 25 (62.5) 30 30 (100)
KYEH T0 25 21 (84.0) 30 29 (96.7)
T1 61 52 (85.2) 52 49 (94.2)
T2 33 30 (90.9) 34 32 (94.1)
Total 119 103 (86.6) 116 110 (94.8)

Two isolates at T2 from OSUGEC and 1 isolate at T2 from KYEH did not survive the freezing process and therefore could not be included in the table.

Two isolates at T0, 3 at T1, and 2 at T2 from OSUGEC and 1 isolate at T0, 3 at T1, and 1 at T2 from KYEH did not survive the freezing process and therefore could not be included in the table.

Discussion

The present study serves as a primary report about the presence of antimicrobial-resistant Enterobacteriaceae bacteria in the feces of hospitalized horses and in equine hospitals. Enterobacteriaceae bacteria expressing clinically relevant resistance phenotypes were isolated from the feces of hospitalized horses and the environment of 2 specialty hospitals in the United States. Hospitalization ≥ 48 hours increased the odds of recovering Enterobacteriaceae isolates with resistant phenotypes from the feces of horses. The number of recovered isolates with resistance to extended-spectrum cephalosporins did not differ between hospitals, but horses hospitalized at KYEH had increased odds of having fluoroquinolone-resistant Enterobacteriaceae bacteria in their feces. The OSUGEC had a consistently higher prevalence of cephalosporin- (57% to 89%) and fluoroquinolone-resistant (26% to 71%) Enterobacteriaceae bacteria across locations within the hospital, versus KYEH (4% to 85% and 0% to 74%, respectively). In both hospitals, the general-purpose wards and barns had numerically higher prevalence of resistant Enterobacteriaceae bacteria, compared with other locations, possibly because of the different, scheduled cleaning procedures of those areas or difference in patient populations (eg, horses with gastrointestinal disease vs horses undergoing outpatient orthopedic procedures). Although Enterobacteriaceae bacteria with extended-spectrum cephalosporin resistance (ie, with AmpC β-lactamase or ESBL phenotypes) were recovered from the surgical suites at KYEH, the surgical suites had the lowest proportion of recovered Enterobacteriaceae bacteria that had resistance to cephalosporins, fluoroquinolones, and carbapenem. Although recovery of resistant bacteria from surgical suites is not ideal, the results of the present study were supportive of the effectiveness of thorough cleaning of commonly contacted surfaces in decreasing nosocomial sources of antimicrobial-resistant bacteria.35 Locations where immunocompromised horses were hospitalized, such as the isolation ward (or barn) and neonatal ICU, yielded higher frequencies of surface contamination with antimicrobial-resistant bacteria, compared with other locations. This was predictable because horses stalled in isolation and the neonatal ICU typically require aggressive antimicrobial treatment and oftentimes administration of multiple antimicrobials.12,36

More variability in the prevalence of environmental Enterobacteriaceae bacteria with AmpC β-lactamase and ESBL phenotypes was noted between locations within KYEH versus OSUGEC, possibly because of the hospital layout or biosecurity protocols. For example, at the time of the present study, KYEH had foot baths throughout the hospital, whereas OSUGEC did not. Also noteworthy was that horses were hospitalized in several barns at KYEH, thereby possibly mitigating the spread of resistant bacteria in the environment; foot traffic, an important source of environmental contamination with bacteria, between barns may have been low.37 The floorplan of OSUGEC had large common areas that were shared by horses and personnel, which may have permitted bacterial dissemination throughout the environment and among horses. Additionally, surgical areas were not sampled at OSUGEC because of a lack of access; therefore, the overall prevalence of resistant bacteria at OSUGEC obtained through this study may have been impacted. Furthermore, horses at the OSUGEC were hospitalized in the same building as farm animal patients, whereas KYEH was an equine-only hospital. Comingling of horses and livestock could have impacted the prevalence of resistant bacteria in the environment because livestock can also play a role in surface contamination through fecal shedding.28 The presence of veterinary students could also have been a factor in the higher overall prevalence of resistant bacteria on surfaces throughout OSUGEC because students, compared with staff and faculty, may have been unfamiliar with appropriate biosecurity protocols during their brief clinical rotations.

Of interest was the recovery of carbapenemase-producing A veronii from the environment of KYEH on several sampling dates. Although A veronii did not confer clinical resistance to carbapenems, the recovery of carbapenemase-producing bacteria from the environment of an equine hospital has not been previously reported. At KYEH, carbapenems were primarily used for the treatment of joint infections (intra-articular administration) of hospitalized horses plus some outpatients. Less frequently, hospitalized horses with bacterial ophthalmologic infections or salmonellosis received a carbapenem because the etiologic agents were multidrug resistant. Hospitalized horses that received carbapenems were predominantly hospitalized in the general-purpose barn or ward, where 2 of the 4 isolates of A veronii were recovered. Repeated administration of carbapenems to people has been shown to select for CRE38; thus, judicious antimicrobial usage may decrease the presence of multidrug-resistant bacteria in the environment.34

Enterobacteriaceae bacteria are common to the gastrointestinal microbiota of horses but can become opportunistic pathogens; however, even as opportunistic pathogens, they are less likely to pose an important health risk to horses than a primary pathogen (eg, Salmonella spp). The risk of environmental contamination with antimicrobial-resistant Enterobacteriaceae bacteria to hospitalized horses is not well established, but the probability of a nosocomial opportunistic infection secondary to exposure to contaminated surfaces is likely low. This low risk may be because enteric commensal bacteria do not commonly produce clinical disease and standard hospital cleaning, disinfection, and biosecurity protocols are usually effective. However, the hospital environment has been proposed as a major risk factor for the development of health-care–associated nosocomial infections in people35; thus, considering the veterinary hospital environment as a risk factor of nosocomial infections and reducing environmental contamination are still reasonable. A greater concern with environmental contamination in hospitals is that plasmid-mediated genes that confer antimicrobial resistance can be transferred between nonpathogenic and pathogenic bacteria, thereby increasing the risk of multidrug-resistant pathogenic bacterial infections in hospitalized patients.39 With rare exceptions, the hospital environment is not sterile. Bacterial resistance to disinfectants is another consideration, albeit rare, when evaluating for the presence of antimicrobial-resistant bacteria in the hospital environment.40 Additionally, the increased use of antimicrobials in the hospital may lead to the selection (ie, selection pressure) of resistant bacteria.39 Although expecting no antimicrobial-resistant bacteria in the hospital environment is unrealistic and the risk of nosocomial infection may be low, minimizing the presence or development of antimicrobial-resistant bacteria in hospitalized patients may reduce this risk from low to almost zero.

The prevalence of hospitalized horses colonized with extended-spectrum cephalosporin- and fluoroquinolone-resistant bacteria was high. Overall, Enterobacteriaceae bacteria that expressed an AmpC β-lactamase phenotype were most commonly isolated from horses hospitalized at OSUGEC, whereas an ESBL phenotype was most common at KYEH. The difference in recovery of resistant bacteria from horses at OSUGEC and KYEH could have been because of differences in the 2 populations, such as antimicrobial administration prior to hospitalization, housing style and density, and lifestyle (professional or recreational); for example, more horses at KYEH had a professional lifestyle (ie, horseracing). On admission, 13% to 32% of horses harbored a bacterium with resistance to at least 1 of the 4 evaluated antimicrobials. Because the development of antimicrobial resistance within 24 hours is implausible, previous antimicrobial administration may have impacted the gastrointestinal microbiota of these horses prior to hospitalization or these horses may have had resistant bacteria as part of their microbiota.13

In the present study, hospitalization, independent of antimicrobial administration, for at least 48 hours increased the odds of isolating Enterobacteriaceae bacteria with an AmpC β-lactamase or ESBL phenotype or with fluoroquinolone resistance from the feces. The odds may have increased with hospitalization because of antimicrobial use and subsequent selection pressure for the development of antimicrobial-resistant Enterobacteriaceae bacteria and through contact with an environment contaminated with antimicrobial-resistant Enterobacteriaceae bacteria, including contamination because of increased fecal shedding of these bacteria by hospitalized horses, and subsequent development of a nosocomial infection.15 However, the odds of identifying a resistant bacterium were numerically lower at T2 than at T1 for both extended-spectrum cephalosporin- and fluoroquinolone-resistant Enterobacteriaceae bacteria. This difference may have been because of the smaller sample size at T2 (n = 80), compared with that at T1 (n = 135). Additionally, this difference may have been because of the administration of various antimicrobials of various classes, such as the bacteriocidal antimicrobials trimethoprim-sulfamethoxazole and metronidazole, during hospitalization that slowly killed Enterobacteriaceae bacteria that had resistance to extended-spectrum cephalosporins or fluoroquinolones at T0 or T1. Similarly, the gastrointestinal microbiota may have had time to reestablish itself with prolonged hospitalization, thus decreasing the prevalence of resistant bacteria. The higher prevalence of antimicrobial-resistant Enterobacteriaceae bacteria in the feces of the horses in the present study at T1 and T2 than at T0 suggested a benefit of minimizing the length of hospital stay when possible.

A European study,6 analogous to the present study, indicates 6.5% to 60.5% of horses have ESBL-producing E coli in their feces. For 110 hospitalized horses, the odds of isolating E coli with ciprofloxacin resistance or the ESBL phenotype were 1.6 to 7.0 and 5.6 to 17.2, respectively, after 2, 4, 6, and ≥ 7 days of hospitalization times the odds of isolation on the day of admission (reference).22 Similar to the results of the present study, the prevalence of antimicrobial-resistant E coli increased after hospital admission but decreased after prolonged hospitalization, compared with admission. Combined previous and present study data suggested that similar trends of the impact of hospitalization on the recovery of antimicrobial-resistant Enterobacteriaceae bacteria from horses occurred independent of the country of origin.

The dissemination of mobile genetic elements conferring antimicrobial resistance to bacteria is widespread in any population-dense environment.1,2,3,4,5 At OSUGEC, 85.1% and 99.0% of AmpC β-lactamase and ESBL phenotypes had blaCMY and blaCTX-M, respectively, which are frequently plasmid mediated.4,41 At KYEH, AmpC β-lactamase and ESBL phenotypes had similar prevalences of blaCMY (76.3%) and blaCTX-M (91.9%). The genes blaCMY and blaCTX-M in the fecal isolates of Enterobacteriaceae bacteria were more commonly isolated from horses hospitalized at KYEH. Additionally, at OSUGEC, blaCTX-M was more prevalent in isolates with the ESBL phenotype (100%) than blaCMY in isolates with the AmpC β-lactamase phenotype (62.5%). A similar trend was noted at KYEH (94.8% ESBL and 86.5% AmpC β-lactamase phenotype). Because of some variability, however, other mechanisms of extended-spectrum cephalosporin resistance, such as porin mutations,42 may be more commonly present in the microbiota of horses hospitalized at OSUGEC than at KYEH. Plasmid-mediated resistance genes are highly mobile and can be transferred between bacteria.10,11 Because resistance genes were likely plasmid-mediated and present at high frequencies in the feces of horses and the hospital environment in the present study, the risk of transference between bacteria of the gastrointestinal microbiota and pathogenic bacteria was high. Continued adherence to appropriate disinfectant and biosecurity protocols is recommended to reduce this risk.

Evaluation of the clonal relationship among E coli isolates of the ESBL phenotype from the hospital environment revealed a diverse population. Horses did not appear to frequently exchange clonal E coli strains during hospitalization; however, 2 horses hospitalized simultaneously at KYEH had the same E coli clone in their feces, which suggested possible transference between them. Five sets of fecal and environmental isolates from KYEH were evaluated for clonality. Two genetically identical pairs of fecal and environmental isolates were identified, which suggested that horses can serve as a source of environmental contamination with antimicrobial-resistant bacteria. Genetically identical pairs of fecal and environmental E coli isolates of the ESBL phenotype were not identified at OSUGEC, likely because of the small number of fecal isolates of E coli expressing an ESBL phenotype that were identified concurrent with environmental sampling. Furthermore, the same E coli clones were simultaneously identified on multiple surfaces throughout the hospital at both OSUGEC and KYEH, possibly because patients and hospital staff and visitors served as mechanical vectors for dissemination of these clones throughout the hospitals. The results of the present study indicated the importance of a detailed biosecurity program including diligent cleaning and disinfection of the environment, good hand hygiene, stalling of horses on the basis of health status, and environmental and patient surveillance to determine the effectiveness of the program.

An important limitation of the present study was the lack of quantification of antimicrobial-resistant bacteria in each sample. Antimicrobials were included with the growth media (MacConkey agar plates) in an attempt to select for the growth of specific antimicrobial-resistant phenotypes of Enterobacteriaceae bacteria. Therefore, the quantities of Enterobacteriaceae bacteria with extended-spectrum cephalosporin or fluoroquinolone resistance originally present in the fecal and environmental samples were unknown, such that only qualitative results (presence or absence of specific antimicrobial-resistant bacteria) could be reported.

Surveillance for common and emerging antimicrobial-resistance mechanisms in horses is warranted to help maintain the efficacy of critically important antimicrobials (eg, carbapenems). Reducing the prevalence of antimicrobial-resistant bacteria in the gastrointestinal microbiota of horses and the hospital environment is crucial for the health and welfare of hospitalized horses and hospital staff and visitors.

Acknowledgments

No external funding was used in this study. The authors declare that there were no conflicts of interest.

Presented in abstract form at the 97th Annual Conference of Research Workers in Animal Diseases, Chicago, December 2016.

The authors thank personnel at the referral equine hospital in Kentucky and The Ohio State University Galbreath Equine Center for their collaboration and assistance with sample collection.

Footnotes

a.

BBL CultureSwab Transport Systems (liquid Stuart, double swab), Becton, Dickinson, and Co, Franklin Lakes, NJ.

b.

CMV3AGNF and ESB1F MIC panels, Thermo Fisher Scientific, Oakwood Village, Ohio.

c.

Illumina MiSeq, San Diego, Calif.

d.

STATA, version 15.1, Stata Corp LLC, College Station, Tex.

Abbreviations

CRE

Carbapenem-resistant Enterobacteriaceae

ESBL

Extended-spectrum β-lactamase

KYEH

Kentucky equine hospital

ICU

Intensive care unit

MIC

Minimum inhibitory concentration

OSUGEC

The Ohio State University Galbreath Equine Center

References

  • 1.

    Schwarz S, Kehrenberg C, Walsh TR. Use of antimicrobial agents in veterinary medicine and food animal production. Int J Antimicrob Agents 2001;17:431437.

    • Search Google Scholar
    • Export Citation
  • 2.

    Dierikx CM, van Duijkeren E, Schoormans AHW, et al. Occurrence and characteristics of extended-spectrum-β-lactamase- and AmpC-producing clinical isolates derived from companion animals and horses. J Antimicrob Chemother 2012;67:13681374.

    • Search Google Scholar
    • Export Citation
  • 3.

    Lutz EA, McCarty MJ, Mollenkopf DF, et al. Ceftiofur use in finishing swine barns and the recovery of fecal Escherichia coli or Salmonella spp resistant to ceftriaxone. Foodborne Pathog Dis 2011;8:12291234.

    • Search Google Scholar
    • Export Citation
  • 4.

    Mollenkopf DF, Weeman MF, Daniels JB, et al. Variable within- and between-herd diversity of CTX-M cephalosporinase-bearing Escherichia coli isolates from dairy cattle. Appl Environ Microbiol 2012;78:45524560.

    • Search Google Scholar
    • Export Citation
  • 5.

    Taylor NM, Davies RH, Ridley A, et al. A survey of fluoroquinolone resistance in Escherichia coli and thermophilic Campylobacter spp on poultry and pig farms in Great Britain. J Appl Microbiol 2008;105:14211431.

    • Search Google Scholar
    • Export Citation
  • 6.

    Boyen F, Smet A, Hermans K, et al. Methicillin resistant staphylococci and broad-spectrum β-lactamase producing Enterobacteriaceae in horses. Vet Microbiol 2013;167:6777.

    • Search Google Scholar
    • Export Citation
  • 7.

    Malo A, Cluzel C, Labrecque O, et al. Evolution of in vitro antimicrobial resistance in an equine hospital over 3 decades. Can Vet J 2016;57:747751.

    • Search Google Scholar
    • Export Citation
  • 8.

    Awosile BB, Heider LC, Saab ME, et al. Antimicrobial resistance in bacteria isolated from horses from the Atlantic Provinces, Canada (1994 to 2013). Can Vet J 2018;59:951957.

    • Search Google Scholar
    • Export Citation
  • 9.

    Maddox TW, Clegg PD, Williams NJ, et al. Antimicrobial resistance in bacteria from horses: epidemiology of antimicrobial resistance. Equine Vet J 2015;47:756765.

    • Search Google Scholar
    • Export Citation
  • 10.

    Huddleston JR. Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes. Infect Drug Resist 2014;7:167176.

    • Search Google Scholar
    • Export Citation
  • 11.

    Leverstein–van Hall MA, Box ATA, Blok HEM, et al. Evidence of extensive interspecies transfer of integron-mediated antimicrobial resistance genes among multidrug-resistant Enterobacteriaceae in a clinical setting. J Infect Dis 2002;186:4956.

    • Search Google Scholar
    • Export Citation
  • 12.

    Dunkel B, Johns IC. Antimicrobial use in critically ill horses. J Vet Emerg Crit Care (San Antonio) 2015;25:89100.

  • 13.

    Johns I, Verheyen K, Good L, et al. Antimicrobial resistance in faecal Escherichia coli isolates from horses treated with antimicrobials: a longitudinal study in hospitalised and non-hospitalised horses. Vet Microbiol 2012;159:381389.

    • Search Google Scholar
    • Export Citation
  • 14.

    Dunowska M, Morley PS, Traub-Dargatz JL, et al. Impact of hospitalization and antimicrobial drug administration on antimicrobial susceptibility patterns of commensal Escherichia coli isolated from the feces of horses. J Am Vet Med Assoc 2006;228:19091917.

    • Search Google Scholar
    • Export Citation
  • 15.

    Ahmed MO, Clegg PD, Williams NJ, et al. Antimicrobial resistance in equine faecal Escherichia coli isolates from North West England. Ann Clin Microbiol Antimicrob 2010;9:1219.

    • Search Google Scholar
    • Export Citation
  • 16.

    Damborg P, Marskar P, Baptiste KE, et al. Faecal shedding of CTX-M-producing Escherichia coli in horses receiving broad-spectrum antimicrobial prophylaxis after hospital admission. Vet Microbiol 2012;154:298304.

    • Search Google Scholar
    • Export Citation
  • 17.

    Dolejska M, Duskova E, Rybarikova J, et al. Plasmids carrying blaCTX-M-1 and qnr genes in Escherichia coli isolates from an equine clinic and a horseback riding centre. J Antimicrob Chemother 2011;66:757764.

    • Search Google Scholar
    • Export Citation
  • 18.

    Maddox TW, Clegg PD, Diggle PJ, et al. Cross-sectional study of antimicrobial-resistant bacteria in horses, part 1: prevalence of antimicrobial-resistant Escherichia coli and methicillin-resistant Staphylococcus aureus. Equine Vet J 2012;44:289296.

    • Search Google Scholar
    • Export Citation
  • 19.

    Maddox TW, Pinchbeck GL, Clegg PD, et al. Cross-sectional study of antimicrobial-resistant bacteria in horses, part 2: risk factors for faecal carriage of antimicrobial-resistant Escherichia coli in horses. Equine Vet J 2012;44:297303.

    • Search Google Scholar
    • Export Citation
  • 20.

    Smet A, Boyen F, Pasmans F, et al. OXA-23-producing Acinetobacter species from horses: a public health hazard? J Antimicrob Chemother 2012;67:30093010.

    • Search Google Scholar
    • Export Citation
  • 21.

    Haggett EF, Wilson WD. Overview of the use of antimicrobials for the treatment of bacterial infections in horses. Equine Vet Educ 2008;20:433448.

    • Search Google Scholar
    • Export Citation
  • 22.

    Maddox TW, Williams NJ, Clegg PD, et al. Longitudinal study of antimicrobial-resistant commensal Escherichia coli in the faeces of horses in an equine hospital. Prev Vet Med 2011;100:134145.

    • Search Google Scholar
    • Export Citation
  • 23.

    Schwaber MJ, Carmeli Y. Carbapenem-resistant Enterobacteriaceae: a potential threat. JAMA 2008;300:29112913.

  • 24.

    Stolle I, Prenger-Berninghoff E, Stamm I, et al. Emergence of OXA-48 carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in dogs. J Antimicrob Chemother 2013;68:28022808.

    • Search Google Scholar
    • Export Citation
  • 25.

    Mollenkopf DF, Stull JW, Mathys DA, et al. Carbapenemase-producing Enterobacteriaceae recovered from the environment of a swine farrow-to-finish operation in the United States. Antimicrob Agents Chemother 2017;61:e01298e16.

    • Search Google Scholar
    • Export Citation
  • 26.

    Gupta N, Limbago BM, Patel JB, et al. Carbapenem-resistant Enterobacteriaceae: epidemiology and prevention. Clin Infect Dis 2011;53:6067.

  • 27.

    Stiefel U, Donskey CJ. The role of the intestinal tract as a source for transmission of nosocomial pathogens. Curr Infect Dis Rep 2004;6:420425.

    • Search Google Scholar
    • Export Citation
  • 28.

    Adams RJ, Kim SS, Mollenkopf DF, et al. Antimicrobial-resistant Enterobacteriaceae recovered from companion animal and livestock environments. Zoonoses Public Health 2018;65:519527.

    • Search Google Scholar
    • Export Citation
  • 29.

    Nordmann P, Poirel L, Dortet L. Rapid detection of carbapenemase-producing Enterobacteriaceae. Emerg Infect Dis 2012;18:15031507.

  • 30.

    Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. CLSI supplement M100. 27th ed. Wayne, Pa: Clinical and Laboratory Standards Institute, 2017.

    • Search Google Scholar
    • Export Citation
  • 31.

    Kado CI, Liu ST. Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol 1981;145:13651373.

  • 32.

    Wang SH, Khan Y, Hines L, et al. Methicillin-resistant Staphylococcus aureus sequence type 239-iii, Ohio, USA, 2007–2009. Emerg Infect Dis 2012;18:15571565.

    • Search Google Scholar
    • Export Citation
  • 33.

    Hosmer DWJ, Lemeshow S, Sturdivant RX. Model-building strategies and methods for logistic regression. In: Applied logistic regression. 3rd ed. Hoboken, NJ: John Wiley & Sons, Inc, 2013;89152.

    • Search Google Scholar
    • Export Citation
  • 34.

    Adams RJ, Mathys DA, Mollenkopf DF, et al. Carbapenemase-producing Aeromonas veronii disseminated in the environment of an equine specialty hospital. Vector Borne Zoonotic Dis 2017;17:439442.

    • Search Google Scholar
    • Export Citation
  • 35.

    Al-Hamad A, Maxwell S. How clean is clean: proposed methods for hospital cleaning assessment. J Hosp Infect 2008;70:328334.

  • 36.

    Corley KTT, Hollis AR. Antimicrobial therapy in neonatal foals. Equine Vet Educ 2009;21:436448.

  • 37.

    Portner JA, Johnson JA. Guidelines for reducing pathogens in veterinary hospitals: hospital design and special considerations. Compend Contin Educ Pract Vet 2010;32:E1E7.

    • Search Google Scholar
    • Export Citation
  • 38.

    Jeon MH, Choi SH, Kwak YG, et al. Risk factors for the acquisition of carbapenem-resistant Escherichia coli among hospitalized patients. Diagn Microbiol Infect Dis 2008;62:402406.

    • Search Google Scholar
    • Export Citation
  • 39.

    Weinstein RA. Controlling antimicrobial resistance in hospitals: infection control and use of antibiotics. Emerg Infect Dis 2001;7:188192.

    • Search Google Scholar
    • Export Citation
  • 40.

    Weber DJ, Rutala WA, Sickbert-Bennett EE. Outbreaks associated with contaminated antiseptics and disinfectants. Antimicrob Agents Chemother 2007;51:42174224.

    • Search Google Scholar
    • Export Citation
  • 41.

    Pérez-Pérez FJ, Hanson ND. Detection of plasmid-mediated AmpC β-lactamase genes in clinical isolates by using multiplex PCR. J Clin Microbiol 2002;40:21532162.

    • Search Google Scholar
    • Export Citation
  • 42.

    Seiffert SN, Hilty M, Perreten V, et al. Extended-spectrum cephalosporin-resistant gram-negative organisms in livestock: an emerging problem for human health? Drug Resist Updat 2013;16:2245.

    • Search Google Scholar
    • Export Citation
  • Figure 1

    Prevalence of Enterobacteriaceae bacteria with AmpC β-lactamase phenotype, ESBL phenotype, and fluoroquinolone resistance isolated from the feces of horses hospitalized at OSUGEC (A) and KYEH (B) within 24 hours of admission (T0; black bars), 48 hours after admission (T1; white bars), and between 3 and 7 days after admission (T2; gray bars).

  • 1.

    Schwarz S, Kehrenberg C, Walsh TR. Use of antimicrobial agents in veterinary medicine and food animal production. Int J Antimicrob Agents 2001;17:431437.

    • Search Google Scholar
    • Export Citation
  • 2.

    Dierikx CM, van Duijkeren E, Schoormans AHW, et al. Occurrence and characteristics of extended-spectrum-β-lactamase- and AmpC-producing clinical isolates derived from companion animals and horses. J Antimicrob Chemother 2012;67:13681374.

    • Search Google Scholar
    • Export Citation
  • 3.

    Lutz EA, McCarty MJ, Mollenkopf DF, et al. Ceftiofur use in finishing swine barns and the recovery of fecal Escherichia coli or Salmonella spp resistant to ceftriaxone. Foodborne Pathog Dis 2011;8:12291234.

    • Search Google Scholar
    • Export Citation
  • 4.

    Mollenkopf DF, Weeman MF, Daniels JB, et al. Variable within- and between-herd diversity of CTX-M cephalosporinase-bearing Escherichia coli isolates from dairy cattle. Appl Environ Microbiol 2012;78:45524560.

    • Search Google Scholar
    • Export Citation
  • 5.

    Taylor NM, Davies RH, Ridley A, et al. A survey of fluoroquinolone resistance in Escherichia coli and thermophilic Campylobacter spp on poultry and pig farms in Great Britain. J Appl Microbiol 2008;105:14211431.

    • Search Google Scholar
    • Export Citation
  • 6.

    Boyen F, Smet A, Hermans K, et al. Methicillin resistant staphylococci and broad-spectrum β-lactamase producing Enterobacteriaceae in horses. Vet Microbiol 2013;167:6777.

    • Search Google Scholar
    • Export Citation
  • 7.

    Malo A, Cluzel C, Labrecque O, et al. Evolution of in vitro antimicrobial resistance in an equine hospital over 3 decades. Can Vet J 2016;57:747751.

    • Search Google Scholar
    • Export Citation
  • 8.

    Awosile BB, Heider LC, Saab ME, et al. Antimicrobial resistance in bacteria isolated from horses from the Atlantic Provinces, Canada (1994 to 2013). Can Vet J 2018;59:951957.

    • Search Google Scholar
    • Export Citation
  • 9.

    Maddox TW, Clegg PD, Williams NJ, et al. Antimicrobial resistance in bacteria from horses: epidemiology of antimicrobial resistance. Equine Vet J 2015;47:756765.

    • Search Google Scholar
    • Export Citation
  • 10.

    Huddleston JR. Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes. Infect Drug Resist 2014;7:167176.

    • Search Google Scholar
    • Export Citation
  • 11.

    Leverstein–van Hall MA, Box ATA, Blok HEM, et al. Evidence of extensive interspecies transfer of integron-mediated antimicrobial resistance genes among multidrug-resistant Enterobacteriaceae in a clinical setting. J Infect Dis 2002;186:4956.

    • Search Google Scholar
    • Export Citation
  • 12.

    Dunkel B, Johns IC. Antimicrobial use in critically ill horses. J Vet Emerg Crit Care (San Antonio) 2015;25:89100.

  • 13.

    Johns I, Verheyen K, Good L, et al. Antimicrobial resistance in faecal Escherichia coli isolates from horses treated with antimicrobials: a longitudinal study in hospitalised and non-hospitalised horses. Vet Microbiol 2012;159:381389.

    • Search Google Scholar
    • Export Citation
  • 14.

    Dunowska M, Morley PS, Traub-Dargatz JL, et al. Impact of hospitalization and antimicrobial drug administration on antimicrobial susceptibility patterns of commensal Escherichia coli isolated from the feces of horses. J Am Vet Med Assoc 2006;228:19091917.

    • Search Google Scholar
    • Export Citation
  • 15.

    Ahmed MO, Clegg PD, Williams NJ, et al. Antimicrobial resistance in equine faecal Escherichia coli isolates from North West England. Ann Clin Microbiol Antimicrob 2010;9:1219.

    • Search Google Scholar
    • Export Citation
  • 16.

    Damborg P, Marskar P, Baptiste KE, et al. Faecal shedding of CTX-M-producing Escherichia coli in horses receiving broad-spectrum antimicrobial prophylaxis after hospital admission. Vet Microbiol 2012;154:298304.

    • Search Google Scholar
    • Export Citation
  • 17.

    Dolejska M, Duskova E, Rybarikova J, et al. Plasmids carrying blaCTX-M-1 and qnr genes in Escherichia coli isolates from an equine clinic and a horseback riding centre. J Antimicrob Chemother 2011;66:757764.

    • Search Google Scholar
    • Export Citation
  • 18.

    Maddox TW, Clegg PD, Diggle PJ, et al. Cross-sectional study of antimicrobial-resistant bacteria in horses, part 1: prevalence of antimicrobial-resistant Escherichia coli and methicillin-resistant Staphylococcus aureus. Equine Vet J 2012;44:289296.

    • Search Google Scholar
    • Export Citation
  • 19.

    Maddox TW, Pinchbeck GL, Clegg PD, et al. Cross-sectional study of antimicrobial-resistant bacteria in horses, part 2: risk factors for faecal carriage of antimicrobial-resistant Escherichia coli in horses. Equine Vet J 2012;44:297303.

    • Search Google Scholar
    • Export Citation
  • 20.

    Smet A, Boyen F, Pasmans F, et al. OXA-23-producing Acinetobacter species from horses: a public health hazard? J Antimicrob Chemother 2012;67:30093010.

    • Search Google Scholar
    • Export Citation
  • 21.

    Haggett EF, Wilson WD. Overview of the use of antimicrobials for the treatment of bacterial infections in horses. Equine Vet Educ 2008;20:433448.

    • Search Google Scholar
    • Export Citation
  • 22.

    Maddox TW, Williams NJ, Clegg PD, et al. Longitudinal study of antimicrobial-resistant commensal Escherichia coli in the faeces of horses in an equine hospital. Prev Vet Med 2011;100:134145.

    • Search Google Scholar
    • Export Citation
  • 23.

    Schwaber MJ, Carmeli Y. Carbapenem-resistant Enterobacteriaceae: a potential threat. JAMA 2008;300:29112913.

  • 24.

    Stolle I, Prenger-Berninghoff E, Stamm I, et al. Emergence of OXA-48 carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in dogs. J Antimicrob Chemother 2013;68:28022808.

    • Search Google Scholar
    • Export Citation
  • 25.

    Mollenkopf DF, Stull JW, Mathys DA, et al. Carbapenemase-producing Enterobacteriaceae recovered from the environment of a swine farrow-to-finish operation in the United States. Antimicrob Agents Chemother 2017;61:e01298e16.

    • Search Google Scholar
    • Export Citation
  • 26.

    Gupta N, Limbago BM, Patel JB, et al. Carbapenem-resistant Enterobacteriaceae: epidemiology and prevention. Clin Infect Dis 2011;53:6067.

  • 27.

    Stiefel U, Donskey CJ. The role of the intestinal tract as a source for transmission of nosocomial pathogens. Curr Infect Dis Rep 2004;6:420425.

    • Search Google Scholar
    • Export Citation
  • 28.

    Adams RJ, Kim SS, Mollenkopf DF, et al. Antimicrobial-resistant Enterobacteriaceae recovered from companion animal and livestock environments. Zoonoses Public Health 2018;65:519527.

    • Search Google Scholar
    • Export Citation
  • 29.

    Nordmann P, Poirel L, Dortet L. Rapid detection of carbapenemase-producing Enterobacteriaceae. Emerg Infect Dis 2012;18:15031507.

  • 30.

    Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. CLSI supplement M100. 27th ed. Wayne, Pa: Clinical and Laboratory Standards Institute, 2017.

    • Search Google Scholar
    • Export Citation
  • 31.

    Kado CI, Liu ST. Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol 1981;145:13651373.

  • 32.

    Wang SH, Khan Y, Hines L, et al. Methicillin-resistant Staphylococcus aureus sequence type 239-iii, Ohio, USA, 2007–2009. Emerg Infect Dis 2012;18:15571565.

    • Search Google Scholar
    • Export Citation
  • 33.

    Hosmer DWJ, Lemeshow S, Sturdivant RX. Model-building strategies and methods for logistic regression. In: Applied logistic regression. 3rd ed. Hoboken, NJ: John Wiley & Sons, Inc, 2013;89152.

    • Search Google Scholar
    • Export Citation
  • 34.

    Adams RJ, Mathys DA, Mollenkopf DF, et al. Carbapenemase-producing Aeromonas veronii disseminated in the environment of an equine specialty hospital. Vector Borne Zoonotic Dis 2017;17:439442.

    • Search Google Scholar
    • Export Citation
  • 35.

    Al-Hamad A, Maxwell S. How clean is clean: proposed methods for hospital cleaning assessment. J Hosp Infect 2008;70:328334.

  • 36.

    Corley KTT, Hollis AR. Antimicrobial therapy in neonatal foals. Equine Vet Educ 2009;21:436448.

  • 37.

    Portner JA, Johnson JA. Guidelines for reducing pathogens in veterinary hospitals: hospital design and special considerations. Compend Contin Educ Pract Vet 2010;32:E1E7.

    • Search Google Scholar
    • Export Citation
  • 38.

    Jeon MH, Choi SH, Kwak YG, et al. Risk factors for the acquisition of carbapenem-resistant Escherichia coli among hospitalized patients. Diagn Microbiol Infect Dis 2008;62:402406.

    • Search Google Scholar
    • Export Citation
  • 39.

    Weinstein RA. Controlling antimicrobial resistance in hospitals: infection control and use of antibiotics. Emerg Infect Dis 2001;7:188192.

    • Search Google Scholar
    • Export Citation
  • 40.

    Weber DJ, Rutala WA, Sickbert-Bennett EE. Outbreaks associated with contaminated antiseptics and disinfectants. Antimicrob Agents Chemother 2007;51:42174224.

    • Search Google Scholar
    • Export Citation
  • 41.

    Pérez-Pérez FJ, Hanson ND. Detection of plasmid-mediated AmpC β-lactamase genes in clinical isolates by using multiplex PCR. J Clin Microbiol 2002;40:21532162.

    • Search Google Scholar
    • Export Citation
  • 42.

    Seiffert SN, Hilty M, Perreten V, et al. Extended-spectrum cephalosporin-resistant gram-negative organisms in livestock: an emerging problem for human health? Drug Resist Updat 2013;16:2245.

    • Search Google Scholar
    • Export Citation

Advertisement