Abstract
Objective
To investigate the patterns of environmental contamination by carbapenemase-producing Enterobacterales (CPE) at veterinary hospitals in or around Philadelphia, PA, and to determine genetic relatedness between isolates.
Methods
Between September 2022 and August 2023, participating hospitals (n = 15) completed a survey regarding demographic data, antimicrobial stewardship efforts and prescribing practices, and infection prevention practices. Samples were collected from a variety of environmental surfaces (n = 311) at each hospital and then enriched and cultured on selective agar to screen for CPE. Confirmed CPE isolates were whole genome sequenced.
Results
Of the 15 hospitals that participated in the study, 6 hospitals (40%) had CPE isolated from at least 1 site. All CPE harbored the blaNDM-7 carbapenemase gene. Of the 24 confirmed CPE isolates, 21 isolates (79%) belonged to a clonal cluster of Enterobacter hormaechei subsp xiangfangensis (single nucleotide polymorphism cluster PDS000141048 on National Center for Biotechnology Information Pathogen Detection). The PDS000141048 cluster was represented at 5 of the 6 hospitals from which CPE was isolated. Additionally, the PDS000141048 cluster included previously uploaded isolates from a biosecurity screening program at 1 of the hospitals in the study, as well as isolates from clinical specimens from outside of the Philadelphia area.
Conclusions
These data suggest that CPE can be common in the veterinary hospital environment. We also detected the presence of a network of transmission of carbapenemase-producing E hormaechei among veterinary hospitals in the greater Philadelphia region.
Clinical Relevance
This finding highlights the importance of environmental infection prevention measures in companion animal veterinary medicine and the need for collaboration across veterinary facilities to prevent the spread of CPE.
Antimicrobial-resistant bacterial infections are a growing concern in companion animal veterinary medicine. Among these is carbapenem-resistant Enterobacterales (CRE), which has been designated by the WHO as a point of critical concern in human healthcare.1 Carbapenem-resistant Enterobacterales includes a number of bacterial species in the order Enterobacterales that display a variety of different resistance mechanisms to carbapenems, which are considered a last-resort class of antimicrobial drugs used in the treatment of multidrug-resistant gram-negative bacterial infections.2 Of particular clinical and public health importance is carbapenemase-producing Enterobacterales (CPE), which harbors genes that encode carbapenemases, enzymes that can hydrolyze most β-lactam antibiotics including carbapenems.2 The 5 most prevalent and clinically relevant carbapenemases (commonly referred to as the “big 5”) are Klebsiella pneumoniae carbapenemase, imipenemase, Verona integron-encoded metallo-β-lactamase, New Delhi metallo-β-lactamase (NDM), and oxacillinase (OXA-48).3
Carbapenemase-producing Enterobacterales infections are frequently nosocomial in nature among human patients, and infection rates in hospitalized patients have been shown to be associated with previous exposure to healthcare environments and procedures.4 Specifically, stays in postacute care facilities (such as long-term care facilities) and ICUs are risk factors for CPE infection and colonization.5–7 Other reported risk factors include treatment with fluoroquinolones, exogenous corticosteroid administration, and autoimmune diseases.6,7
The emergence of CPE in animals has been a recent but growing concern. Carbapenemase-producing Enterobacterales has been identified in food animals (pigs, poultry, cattle, and seafood), companion animals (dogs, cats, horses, and pet birds), and wildlife (wild birds and boar).8 In 2018, a cluster of Escherichia coli isolates harboring the gene encoding NDM-5 (blaNDM-5) obtained from the respiratory and urinary tracts of 6 animals (5 dogs and 1 cat) in a tertiary-care veterinary teaching hospital in Philadelphia was described; 5 of the 6 animals had received at least 4 antimicrobial drugs before sample collection.9 Furthermore, K pneumoniae carbapenemase–producing Enterobacterales was obtained from surveillance rectal swabs on 2 subclinical dogs in response to the aforementioned outbreak.10
There is some evidence that the veterinary hospital environment, including equipment, may play a role in the spread of CPE in veterinary facilities. A case-control study11 of the aforementioned blaNDM-5 CPE outbreak identified endotracheal intubation, exposure to the anesthesia service, and exposure to the surgery service as statistically significant risk factors for acquisition of blaNDM-5–harboring carbapenem-resistant E coli.11 It was suspected that contamination of endotracheal tubes (or the bin in which they were improperly disinfected) may have played a role in the outbreak. Other outbreaks of CPE in veterinary hospitals have been described and linked specifically to spread within the hospital environment.12,13
Carbapenemase-producing Enterobacterales is a growing concern in veterinary hospitals across the US. The environmental presence of, and genetic relationships between, these resistant organisms in multiple private and academic veterinary hospitals within a metropolitan region with a shared referral base have not been investigated. To better understand the dynamics of CPE spread between veterinary hospitals, the aims of this study were (1) to screen for the environmental presence of CPE in veterinary hospitals in and around Philadelphia, PA, and describe the distribution of CPE within these facilities; and (2) to perform whole genome sequencing on the isolated CPE to assess genetic relatedness.
Methods
Approximately 80 veterinary hospitals in the Philadelphia metropolitan area were sent letters requesting their participation in a study to identify infection prevention practices and potential reservoirs of resistant bacteria. These letters also detailed that their participation would be completely anonymous and would require the completion of a survey along with 1 visit to their practice. To allow for the anonymity of results, separate sampling and laboratory teams were established. The laboratory was blinded to the identity of the practice and the sampling team was blinded to results. Any practice interested in participating contacted the collection team and coordinated survey completion and sample collection at a mutually agreeable time. Practices that opted in to contribute to the study were compensated with certificates for 5 cultures with susceptibility testing from the Penn Vet Ryan Clinical Microbiology Diagnostic Laboratory (valued at $250).
Each participating hospital completed a survey that asked for demographic data including the estimated number of doctors, staff, and patients seen per week, as well as types of care provided (primary, emergency, urgent, low-cost, overnight, exotics, and referral/specialty) and community type (urban, suburban, and rural; survey available in Supplementary Material S1). The survey also asked about hospital participation in antimicrobial stewardship efforts and antimicrobial stocking practices. Finally, the survey asked about specific infection prevention measures including environmental disinfection protocols, hospital-dedicated footwear, glove use policies, specific protocols for patients with multidrug-resistant bacterial infections, hand hygiene protocols, the presence of an infection preventionist on staff, and the use of infectious disease flags in the hospital medical records system.
At the study visit, 29 preestablished surfaces grouped into the following categories were screened for CRE: high-touch points, electronics, water-associated sites, noncritical patient care devices (those not requiring high-level disinfection or sterilization between patients), and patient spaces. The collection team member screened each surface by thoroughly wiping the site with an environmental sampling sponge impregnated with a neutralizing broth (Whirl-pak Hydrated PolySponge Sampling Bag with Sampling Sponge; Thermo Fisher Scientific). Hand hygiene with alcohol-based hand sanitizer and changing gloves was performed between each sample collection to prevent contamination between sites.
The collection team deidentified the participant’s samples by assigning the participating practice an “H number” (H1 to H15). In addition, to further ensure anonymity, the collection team scheduled sampling in clusters (groups of 2 to 3 hospitals within 1 week of each other based on enrollment order and hospital representative availability), so that any hospital that was to redeem its compensation for participation would not be revealed to the laboratory. Each sample collected was labeled with the practice’s H number and a letter (A to Y) that corresponded with the site screened. Practices were given the opportunity to exclude any surfaces that they did not want screened, as well as screen any additional surfaces that were not already included in the study design. At the completion of the visit, the sponges were either delivered directly to the laboratory or shipped overnight at room temperature. The collection team was only informed of the participants’ results if the practice requested their results.
Upon receipt at the laboratory, samples were enriched with 100 mL of buffered peptone water and incubated for 16 to 20 hours at 35 ± 2 °C. Next, 5 mL of the inoculated buffered peptone water was transferred to 5 mL 2X MacConkey broth supplemented with 4 μg/mL cefotaxime (MAC-CTX) for a final concentration of 1X MacConkey broth and 2 μg/mL cefotaxime and then incubated for 16 to 20 hours at 35 ± 2 °C. After incubation, 100 μL of the inoculated MAC-CTX broth was pipetted onto 2 types of chromogenic selective media (CHROMID Carba agar and HardyCHROM Extended Spectrum β-Lactamase [ESBL] agar) and incubated for 16 to 20 hours at 35 ± 2 °C. Growth was interpreted following the manufacturer’s instructions on the chromogenic agar plates for CHROMID Carba. Three colonies of each suspect color and/or morphology from each agar were subcultured to MacConkey agar. If each of the 3 subcultures appeared phenotypically identical on MacConkey agar, only the first subculture received further analysis; however, all subcultures were frozen in 25% glycerol stock for future analysis if necessary. For the HardyCHROM ESBL plates, all colonies were subcultured and then screened with the oxidase test. Oxidase-negative isolates were inferred to be potential Enterobacterales and were subjected to further investigation. Bacteria that met these criteria were initially identified biochemically using a commercial diagnostic system (Vitek2 GNID card; bioMerieux). Confirmed Enterobacterales was screened for carbapenemase production by the modified carbapenem inactivation method, as described in the Clinical and Laboratory Standards Institute M100 standards.14
Confirmed CPE was subsequently sent to a commercial sequencing center for whole genome sequencing (SeqCenter). Briefly, sample libraries were prepared using the Illumina DNA Prep kit and IDT 10-bp UDI indices and sequenced on an Illumina NextSeq 2000, producing 2 X 150-bp reads. Demultiplexing, quality control, and adapter trimming were performed with bcl-convert (version 3.9.3). Raw sequence reads were uploaded to the National Center for Biotechnology Information (NCBI) Pathogen Detection pipeline as part of BioProject PRJNA1029998 for assembly, antimicrobial resistance gene identification, and single nucleotide variant analysis.15–17 Individual biosample identifiers can be found in Supplementary Table S1. Assembled draft genomes were downloaded from the NCBI and used to confirm identification via Kmer Finder and categorized using in silico multilocus sequence typing.18,19
For large clusters of isolates, representative isolates from each unique clade in the NCBI database were selected for additional single nucleotide polymorphism (SNP) analysis. Genomes downloaded from the NCBI were run through the Spriggan 1.1.2 bioinformatics pipeline for genome assembly, multilocus sequence typing determination, and antimicrobial resistance gene prediction.20 Reference-based SNP analysis was performed on the isolate FASTQ files using CFSAN as part of the Dryad 3.0.0 workflow to generate a SNP heatmap.21 A phylogenetic tree based on the SNP matrix was generated using RAxML and visualized using MEGA7.22,23
Results
A total of 15 independent veterinary hospitals (15/80, 18.75%) enrolled in the study between September 2022 and August 2023 (H1 to H15). Of the 15 hospitals, 3 (20%) answered “yes” to participating in antimicrobial stewardship efforts. No hospitals reported regularly stocking imipenem, but 4/15 hospitals (27%) reported routinely stocking meropenem. Most of the hospitals (9/15 [60%]) reported participation in 1 or more infection prevention measures; however, only 1 hospital reported a formal hand hygiene policy, hand hygiene training, hand hygiene auditing, or having an infection preventionist on staff (Table 1).
Results of the veterinary hospital survey administered to a hospital representative by study site.
| H1 | H2 | H3 | H4 | H5 | H6 | H7 | H8 | H9 | H10 | H11 | H12 | H13 | H14 | H15 | |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| No. of doctors | 7 | 8 | 3 | 1.5 | 1.5 | 44 | 10 | 15.5 | 18 | 36 | 4 | 2 | 21.5 | 5 | 176 |
| No. of staff | 27 | approx 30 | 25 | 8 | approx 10 | 220 | 55 | 88 | approx 50 | > 200 | approx 10 | 12 | 115 | >25 | approx 200 |
| Average No. of patients/week | > 200 | 100–200 | 100–200 | 100–200 | 100–200 | > 200 | > 200 | > 200 | > 200 | > 200 | > 200 | 100–200 | > 200 | 100–200 | > 200 |
| Primary care | Y | N | Y | Y | Y | N | Y | N | Y | Y | Y | Y | Y | Y | Y |
| Emergency care | N | Y | N | Y | Y | Y | N | Y | Y | Y | N | N | Y | N | Y |
| Urgent care | N | Y | N | Y | N | N | Y | N | N | N | N | Y | N | N | N |
| Low-cost care | N | N | N | N | N | N | N | N | N | N | N | N | N | N | N |
| Overnight care | N | Y | N | N | N | Y | N | Y | N | Y | N | N | Y | N | Y |
| Exotics | N | Y | N | N | N | N | N | N | N | N | N | Y | N | N | N |
| Referral/specialty | N | Y | N | N | N | Y | N | Y | Y | Y | N | N | Y | N | Y |
| Antimicrobial stewardship efforts | N | Y | N | N | X | N | X | N | X | X | Y | N | N | X | Y |
| Disinfection training | N | Y | N | N | X | Y | N | Y | X | Y | Y | N | N | X | Y |
| Disinfection auditing | N | N | N | N | X | N | Y | N | X | N | N | N | N | X | Y |
| Dedicated footwear | N | N | N | N | X | N | N | N | X | N | N | N | N | X | N |
| Multidrug-resistant protocols | Y | Y | N | N | X | Y | Y | N | X | Y | N | N | Y | X | Y |
| Infectious disease flags | Y | Y | N | N | X | Y | Y | Y | X | Y | N | N | Y | X | Y |
| Hand hygiene policy | N | N | X | N | X | N | N | X | N | N | N | N | N | N | Y |
| Hand hygiene training | N | N | X | N | X | N | N | X | N | N | N | N | N | N | Y |
| Hand hygiene auditing | N | N | X | N | X | N | N | X | N | N | N | N | N | N | Y |
| Infection preventionist | N | N | X | N | X | N | N | X | N | N | N | N | N | N | Y |
| Community type | S | U | S | S | S | S | U | S | S | S | S | U | S | S | U |
| Glove policy | I | A | NP | NP | X | I | A | I | X | A | NP | NP | I | X | A |
Hospitals are designated H1 to H15. The answers for yes (Y)/no (N) questions of types of care offered, participation in antimicrobial stewardship efforts, disinfection training in place, disinfection auditing in place, dedicated footwear required, multidrugresistant protocols in place, infectious disease flags used in medical records, hand hygiene policy in place, hand hygiene training in place, hand hygiene auditing in place, and presence of an infection preventionist on staff are provided as well as hospitals that did not respond to a certain question (X). For community type, suburban (S) and urban (U) are indicated. For glove policy, required for all patients (A), required for infectious disease patients only (I), no formal policy in place (N), and hospital did not respond to that question (X) are indicated.
Each of the 15 independent hospitals that opted to participate was environmentally screened for CPE (Table 2). Up to 29 environmental sites were sampled in each hospital, although not all sites were sampled at each facility due to practice-elected exclusion or absence/lack of access to the site. In total, 311 samples across H1 to H15 were sampled. Of the 15 hospitals, 6 hospitals (40%) had CPE isolated from at least 1 environmental site resulting in a total of 24 CPE organisms isolated. The maximum number of CPE-positive sites at a single hospital was at H6, which had positive isolates from 11/20 (55%) sampled sites. Three hospitals (H5, H7, and H9) had 3/20 (15%) CPE-positive sites each, 1 hospital (H8) had 2/22 (9%) CPE-positive sites, and 1 hospital (H15) had 2/21 (9.5%) CPE-positive sites. The most common sampling sites from which CPE was isolated across the hospitals were sinks in the treatment area, floors, and cages with 3/15 (20%) of hospitals testing CPE positive from these types of sites. The following sites were CPE positive at 2/15 (13.3%) of the hospitals: door to treatment area, ultrasound/radiography station, and defecation area. The following sites were CPE positive at 1/15 (6.6%) of the hospitals: cabinet/drawer handles, anesthesia machine, computer keyboards, bloodwork machine, floor drain, stethoscopes, pulse oximeter, runs, and exam table.
Carbapenemase-producing Enterobacterales (CPE) screening results by veterinary hospital and sampling site.
| H1 | H2 | H3 | H4 | H5 | H6 | H7 | H8 | H9 | H10 | H11 | H12 | H13 | H14 | H15 | |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Cabinet/drawer handles | − | − | − | − | − | − | + | − | − | − | − | − | − | − | − |
| Cage handles/latches | − | − | − | − | − | − | − | − | − | − | − | − | − | − | − |
| Door to treatment area | − | − | − | − | − | + | + | − | − | − | − | − | − | − | − |
| Anesthesia machine | − | − | − | − | − | + | − | − | − | − | − | − | − | − | − |
| Light switch | − | − | − | − | − | − | − | − | − | − | − | − | − | − | − |
| Computer keyboards | − | − | − | − | − | + | − | − | − | − | − | − | − | − | − |
| Staff cell phones | − | − | ns | − | − | − | − | − | − | − | − | − | − | − | − |
| Clinic phone | − | − | − | − | − | − | − | − | − | − | − | − | − | − | − |
| Bloodwork machine | − | − | − | − | − | + | − | − | − | − | − | − | − | − | − |
| Ultrasound/radiography station | − | − | ns | − | + | + | − | − | − | ns | − | − | − | − | − |
| Sink (treatment area) | − | − | − | − | − | + | + | − | + | − | − | − | − | − | − |
| Sink (nontreatment area) | − | − | − | − | − | − | − | − | − | − | − | − | − | − | − |
| Floor drain | ns | − | ns | ns | ns | ns | ns | ns | ns | ns | ns | ns | ns | ns | + |
| Run drain | ns | − | ns | − | ns | ns | ns | ns | ns | − | − | ns | ns | ns | − |
| Dental water source | − | ns | − | − | − | ns | − | ns | ns | ns | − | − | ns | ns | ns |
| Stethoscopes | − | − | ns | − | − | + | − | − | − | − | ns | − | − | − | − |
| Thermometers | − | − | − | − | − | − | − | − | − | − | − | − | − | − | − |
| Pulse oximeter | − | − | − | − | + | − | − | ns | − | − | − | − | − | − | − |
| Blood pressure cuffs | − | − | − | ns | − | − | − | − | ns | − | − | ns | − | − | − |
| Pill guns | − | − | − | ns | ns | ns | − | ns | − | ns | ns | − | ns | ns | ns |
| Runs | ns | ns | ns | ns | ns | ns | ns | − | + | − | − | − | − | ns | − |
| Cages | − | − | − | − | + | + | − | − | − | − | − | ns | − | − | + |
| Defecation area | ns | − | − | − | ns | + | ns | + | ns | − | ns | ns | ns | ns | − |
| Exam table | − | − | − | − | − | + | ns | − | − | − | − | − | − | − | ns |
| Floor | − | − | − | − | − | + | − | + | + | − | ns | − | − | − | ns |
| Sponge | ns | − | ns | ns | ns | ns | ns | − | ns | ns | ns | ns | ns | ns | ns |
| Isolation cage | ns | ns | ns | ns | ns | ns | ns | − | ns | ns | ns | ns | ns | − | ns |
| Laundry bin | ns | ns | ns | ns | ns | ns | ns | ns | ns | − | ns | ns | ns | ns | ns |
| Scrub container | ns | ns | ns | ns | ns | ns | ns | ns | ns | ns | ns | ns | − | ns | ns |
Results for CPE not isolated (−), CPE isolated (+), and site not sampled (ns) are shown.
Positive CPE isolates were further characterized by whole genome sequencing (Supplementary Table S1). All CPE isolates in this study were found to produce the carbapenemase gene blaNDM-7. The most common species identified was Enterobacter hormaechei subsp xiangfangensis, representing 21/24 (87.5%) of isolates. Single nucleotide polymorphisms were used to group isolates into clusters, with clusters being defined as isolates differing by < 50 SNPs. One E hormaechei isolate from H6 did not cluster with any isolates of CPE in the NCBI Pathogen Detection database. All 21 of the E hormaechei subsp xiangfangensis isolates were ST171 and grouped in the PDS000141048 SNP cluster using the NCBI Pathogen Detection pipeline, with SNP distances ranging from 0 to 28. Isolates belonging to cluster PDS000141048 were identified in 5 of the 6 hospitals from which CPE was isolated. Isolates within the PDS000141048 cluster were most closely related to those isolated from within the same facility (H5 isolates within 4 SNPs, H6 isolates within 5 SNPs, H7 isolates within 1 SNP, H8 isolates within 1 SNP, and H9 isolates within 8 SNPs). Isolates from H6 and H7 had a high degree of relatedness with all isolates clustering within 4 SNPs. Between other hospitals, there was more genetic diversity found (range, 9 to 27 SNPs).
The same cluster (PDS000141048 on NCBI Pathogen Detection) also included previously uploaded isolates from animals including 22 fecal surveillance isolates (18 dogs and 4 cats) from a clinical biosecurity program at H15 and a 2020 environmental specimen (a sponge) from H15. In addition, 5 animal isolates from clinical specimens (2 urine, 1 endotracheal wash, 1 wound, and 1 esophageal tube site) from outside of the Philadelphia area (3 from Washington, DC; 1 from Connecticut; and 1 from Texas) were also found to be within the PDS000141048 cluster.
For the large cluster of E hormaechei subsp xiangfangensis, representative isolates from the NCBI cluster PDS000141048 (including both previously uploaded isolates and isolates from this study) were selected for additional SNP characterization by an in-house pipeline used for assessment of clinical outbreaks of CPE in human healthcare settings.21 These analyses identified close genetic relationships between isolates from the present study, those from the biosecurity program at H15, and those from the clinical specimens from outside of the Philadelphia area. The SNP distances between representative isolates from the PDS000141048 cluster ranged from 4 to 41 SNPs (Figure 1).
Single nucleotide polymorphism (SNP) characterization of representative isolates from a National Center for Biotechnology Information cluster of Enterobacter hormaechei subsp xiangfangensis. A—A SNP distance matrix using heatmap color coding. B—A SNP-based phylogenetic tree. Purple-highlighted isolates are those from this study (hospital [H]6P, H8Y, H9Y, H7C, and H5V), and blue-highlighted isolates are from distinct investigations. The SNP heatmap utilizes a color gradient to represent the degree of genetic similarity among isolates. Red indicates the smallest SNP differences (n = 4), corresponding to the highest genetic similarity. Yellow represents intermediate SNP differences (28 to 30), while green denotes the largest SNP differences (n = 41), reflecting the lowest genetic similarity. This color scheme provides a visual framework for interpreting the extent of genetic relatedness, with red areas highlighting closely related isolates and green areas denoting more distantly related ones. ETW = Endotracheal wash.
Citation: American Journal of Veterinary Research 86, 6; 10.2460/ajvr.25.01.0024
Discussion
We identified CPE on environmental surfaces in 6 of the 15 Philadelphia veterinary hospitals from which samples were taken. In our study, the most common sites from which CPE was isolated were sinks, floors, and cages. By comparison, a previous study13 on environmental contamination with CPE and other multidrug-resistant organisms in a group of companion animal hospitals found consultation rooms, ICUs, and utensils (eg, dental cleaning devices, animal transport boxes, mobile phones, stethoscopes, and ultrasonography devices) to be the most frequently contaminated sites. In that study,13 specific sampling sites within consultation rooms and ICUs included both human contact surfaces (eg, desktops) and animal contact surfaces (eg, examination tables). In human healthcare facilities, numerous studies24–26 have identified sinks as reservoirs of CRE, and it has been implied that the splashing of water from the faucet into contaminated drains may result in aerosolization of organisms and contamination of surrounding surfaces as well as human hands. Notably, in 1 such study,25 an isolate of carbapenem-resistant Serratia marcescens was persistently isolated from ICU sink drains/grates after 6 attempts at decontamination, highlighting the role of biofilms as environmental reservoirs. In addition, the isolation of carbapenem-resistant K pneumoniae from floors of patient rooms in human tertiary care hospitals has been documented.27 This current study, and those referenced, indicate the importance of focusing infection prevention efforts on both human and animal contact surfaces, as well as water-associated environments such as sinks, in both veterinary and human medical settings. Particularly, the documentation of floor contamination suggests that dedicated footwear should be emphasized to limit spread outside of a facility.
While these results underscore the importance of infection prevention and control in mitigating the spread of antimicrobial-resistant bacteria in veterinary hospitals, they also suggest a role for active environmental surveillance in directing and improving infection prevention measures. Environmental sampling often occurs in the aftermath of a known outbreak; for example, methicillin-resistant Staphylococcus aureus was identified on stalls, personal items, and animal restraint devices following a cluster of equine and human methicillin-resistant S aureus infections in a veterinary teaching hospital.28 More recent efforts to implement active surveillance for antimicrobial-resistant bacteria have been undertaken in veterinary teaching hospitals with the goals of addressing environmental reservoirs before an outbreak and improving infection prevention strategies. Notably, 1 such active surveillance program isolated target organisms (including CPE) at similar rates from both human-only contact surfaces and human-animal contact surfaces, which led to a campaign for improved hand hygiene in that hospital.29 Likewise, an investigation of the environmental presence of multidrug-resistant bacteria across 4 small animal hospitals in Hungary identified human-only contact surfaces (computer keyboards) and human-animal contact surfaces (muzzles, surgical gowns, and cage floors) as important predilection sites.30
Our study also demonstrated that CPE may spread within veterinary facilities where carbapenems are not routinely used, since CPE was found within the environment of a veterinary hospital (H9) that did not stock a carbapenem drug. This finding correlates with a retrospective study31 of carbapenem-prescribing practices before an outbreak of CPE at a veterinary teaching hospital where carbapenem use was very rare (0.2% of all antimicrobial prescriptions in the study period). The same study31 also revealed that carbapenems were prescribed empirically (before culture and susceptibility results) in over half of the cases in the period preceding the CPE outbreak and suggested a need for specific antimicrobial stewardship actions surrounding the use of carbapenems. However, the spread of CPE even with no or low carbapenem use suggests that antimicrobial stewardship of carbapenems alone will not control its spread. Many CPE isolates are extensively or pan-drug resistant; therefore, the use of any antimicrobial drugs may lead to selection pressure. Stewardship must be bundled with strong infection prevention programs and actions including hand hygiene and environmental control as demonstrated in human medicine.32 It is well established in human medicine that hand hygiene is vitally important in mitigating the transmission of CPE and other multidrug-resistant organisms in hospital settings.33,34 The rarity of formal hand hygiene programs among the hospitals surveyed in the present study indicates an opportunity for improvement in the veterinary medical field with respect to core principles of infection prevention. This premise is supported by a study35 that found the widespread presence of multidrug-resistant organisms in the hands of students working in the intensive care unit of a veterinary teaching hospital. Hand hygiene is thus likely of particular importance in mitigating the transmission of CPE and other multidrug-resistant organisms from veterinary patients to staff and clients.
Of the 6 CPE-positive hospitals, 5 harbored a clonal population of carbapenemase-producing E hormaechei subsp xiangfangensis in the environment. The presence of a closely genetically related strain of organisms across multiple facilities suggests the presence of a referral network in which colonized or infected animals transmit CPE between veterinary hospitals. Alternatively, the environment serves as a potential source within the community that has led to contamination within the multiple facilities. Except for isolates from 2 hospitals (H6 and H7), evidence suggests periodic selection was occurring within each hospital as less genetic variability was found between isolates from the same facility (0 to 8 SNPs) than was found between those facilities (0 to 28 SNPs). Even though isolates in the PDS000141048 SNP cluster were not found in the environment at H15 on sampling, isolates from a significant number of animals (n = 22) found previously as part of a biosecurity screening program at H15 were also found to be part of the cluster suggesting spread to and/or from H15 as well.
In addition to the closely related isolates of blaNDM-7–harboring E hormaechei subsp xiangfangensis from animals and environmental surfaces within the Philadelphia region, we also identified genomic evidence of several related isolates that caused infections in animals from outside of the Philadelphia region (Texas; Washington, DC; and Connecticut). This may suggest a more extensive spread across the US of the primary strain (NDM-7–producing E hormaechei subsp xiangfangensis). One previous national study36 of CPE demonstrated the potentially important role of animal travel when a CPE-positive animal in Virginia was linked to a case of CPE in a dog in Florida and at both veterinary hospitals. More research and surveillance are needed to characterize the role that travel may play in the spread of CPE among animals and geographical regions.
One limitation of this study is its relatively small sample size of 15 hospitals all within the same metropolitan region, as well as the potential for selection bias as hospitals with greater awareness of, and concern for, CPE may have been more likely to participate. Furthermore, each hospital had the option to opt out of any given sampling site, which may similarly result in undetected bias. CHROMID Carba agar has been shown to be more effective at isolating organisms of CPE with high-carbapenem MICs and less effective at isolating bacteria with reduced susceptibility due to carbapenemase production.37 To circumvent this issue, we also used a carbapenem-free enrichment medium followed by ESBL agar devoid of carbapenems, but it is possible that another culture strategy may have yielded slightly different results.
Future directions for investigation may include (1) multicenter surveillance studies of CPE in veterinary patients, (2) evaluation of genetic relatedness among isolates from colonized animals at different hospitals, and (3) epidemiologic studies to identify risk factors for high environmental CPE burdens in veterinary hospitals.
In summary, there appears to exist a transmission network of CPE between veterinary hospitals in the Philadelphia region, and veterinarians and hospital administrators in all sectors of the profession should prioritize best practices in environmental decontamination and hygiene to mitigate the risk of CPE infections in their patients and potentially owners and staff.
Supplementary Materials
Supplementary materials are posted online at the journal website: avmajournals.avma.org.
Acknowledgments
None reported.
Disclosures
The authors have nothing to disclose. No AI-assisted technologies were used in the composition of this manuscript.
Funding
Sampling and sequencing were financially supported by an internal grant funded by the Institute for Infectious and Zoonotic Disease at Penn Vet. This publication (bioinformatic analysis) was supported by the Office of Advanced Molecular Detection, CDC, through Cooperative Agreement No. CK22-2204. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the CDC.
ORCID
E. Hsieh https://orcid.org/0000-0001-9687-9723
J. Dietrich https://orcid.org/0000-0003-2975-495X
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