Preparing for Mpox Resurgence: Surveillance Lessons From Outbreaks in Toronto, Canada.

BACKGROUND: With many global jurisdictions, Toronto, Canada, experienced an mpox outbreak in spring/summer 2022. Cases declined following implementation of a large vaccination campaign. A surge in early 2023 led to speculation that asymptomatic and/or undetected local transmission was occurring in the city. METHODS: Mpox cases and positive laboratory results are reported to Toronto Public Health. Epidemic curves and descriptive risk factor summaries for the 2022 and 2023 outbreaks were generated. First- and second-dose vaccination was monitored. Mpox virus wastewater surveillance and whole genome sequencing were conducted to generate hypotheses about the source of the 2023 resurgence. RESULTS: An overall 515 cases were reported in spring/summer 2022 and 17 in the 2022-2023 resurgence. Wastewater data correlated with the timing of cases. Whole genome sequencing showed that 2022-2023 cases were distinct from 2022 cases and closer to sequences from another country, suggesting a new importation as a source. At the start of the resurgence, approximately 16% of first-dose vaccine recipients had completed their second dose. CONCLUSIONS: This investigation demonstrates the importance of ongoing surveillance and preparedness for mpox outbreaks. Undetected local transmission was not a likely source of the 2022-2023 resurgence. Ongoing preexposure vaccine promotion remains important to mitigate disease burden.

First Canadian report of transmission of fluconazole-resistant Candida parapsilosis within two hospital networks confirmed by genomic analysis.

Candida parapsilosis is a common cause of non-albicans candidemia. It can be transmitted in healthcare settings resulting in serious healthcare-associated infections and can develop drug resistance to commonly used antifungal agents. Following a significant increase in the percentage of fluconazole (FLU)-nonsusceptible isolates from sterile site specimens of patients in two Ontario acute care hospital networks, we used whole genome sequence (WGS) analysis to retrospectively investigate the genetic relatedness of isolates and to assess potential in-hospital spread. Phylogenomic analysis was conducted on all 19 FLU-resistant and seven susceptible-dose dependent (SDD) isolates from the two hospital networks, as well as 13 FLU susceptible C. parapsilosis isolates from the same facilities and 20 isolates from patients not related to the investigation. Twenty-five of 26 FLU-nonsusceptible isolates (resistant or SDD) and two susceptible isolates from the two hospital networks formed a phylogenomic cluster that was highly similar genetically and distinct from other isolates. The results suggest the presence of a persistent strain of FLU-nonsusceptible C. parapsilosis causing infections over a 5.5-year period. Results from WGS were largely comparable to microsatellite typing. Twenty-seven of 28 cluster isolates had a K143R substitution in lanosterol 14-α-demethylase (ERG11) associated with azole resistance. As the first report of a healthcare-associated outbreak of FLU-nonsusceptible C. parapsilosis in Canada, this study underscores the importance of monitoring local antimicrobial resistance trends and demonstrates the value of WGS analysis to detect and characterize clusters and outbreaks. Timely access to genomic epidemiological information can inform targeted infection control measures.

Monkeypox Virus Detection in Different Clinical Specimen Types.

A global monkeypox outbreak began in May 2022. Limited data exist on specimen type performance in associated molecular diagnostics. Consequently, a diverse range of specimen sources were collected in the initial weeks of the outbreak in Ontario, Canada. Our clinical evaluation identified skin lesions as the optimal diagnostic specimen source.

A Prolonged Outbreak of Human Adenovirus A31 (HAdV-A31) Infection on a Pediatric Hematopoietic Stem Cell Transplantation Ward with Whole Genome Sequencing Evidence of International Linkages.

A surge in hematopoietic stem cell transplantation (HSCT) human adenovirus A31 (HAdV-A31) infections was initially observed in late 2014/2015 at SickKids (SK) Hospital, Toronto, Canada. In response, enhanced laboratory monitoring for all adenovirus infections was conducted. Positive samples underwent genotyping, viral culture, and, in selected cases, whole-genome sequencing (WGS). HAdV-A31 specimens/DNA obtained from four international pediatric HSCT centers also underwent WGS. During the SK outbreak period (27 October 2014 to 31 October 2018), 17/20 HAdV-A31 isolates formed a distinct clade with 0 to 8 mutations between the closest neighbors. Surveillance before and after the outbreak detected six additional HAdV-A31 HSCT cases; three of the four sequenced cases clustered within the outbreak clade. Two SK outbreak isolates were identical to sequences from two patients in an outbreak in England. Three SK non-outbreak sequences also had high sequence similarity to strains from three international centers. Environmental PCR testing of the HSCT ward showed significant adenovirus contamination. Despite intense infection control efforts, we observed re-occurrence of infection with the outbreak strain. Severe but nonfatal infection was observed more commonly with HAdV-A31 compared to other genotypes, except HAdV-C1. Our findings strongly implicate nosocomial spread of HAdV-A31 over 10 years on a HSCT unit and demonstrate the value of WGS in defining and mapping the outbreak. Close linkages among strains in different countries suggest international dissemination, though the mechanism is undetermined. This large, extended outbreak emphasizes the pre-eminent role of HAdV-A31 in causing intractable pediatric HSCT outbreaks of severe illness worldwide.

Phenotypic and Genomic Profiling of Staphylococcus argenteus in Canada and the United States and Recommendations for Clinical Result Reporting.

Staphylococcus argenteus is a newly described species, formerly known as S. aureus clonal complex 75 (CC75). Here, we describe the largest collection of S. argenteus isolates in North America, highlighting identification challenges. We present phenotypic and genomic characteristics and provide recommendations for clinical reporting. Between 2017 and 2019, 22 isolates of S. argenteus were received at 2 large reference laboratories for identification. Identification with routine methods (biochemical, matrix-assisted laser desorption ionization-time of flight mass spectrometry [MALDI-TOF MS], 16S rRNA gene analysis) proved challenging to confidently distinguish these isolates from S. aureus Whole-genome sequencing analysis was employed to confirm identifications. Using several different sequence-based analyses, all clinical isolates under investigation were confirmed to be S. argenteus with clear differentiation from S. aureus Seven of 22 isolates were recovered from sterile sites, 11 from nonsterile sites, and 4 from surveillance screens. While sequence types ST1223/coa type XV, ST2198/coa type XIV, and ST2793/coa type XId were identified among the Canadian isolates, the majority of isolates (73%) belonged to multilocus sequence types (MLST) ST2250/coa type XId and exhibited a high degree of homology at the genomic level. Despite this similarity, 5 spa types were identified among ST2250 isolates, demonstrating some diversity between strains. Several isolates carried mecA, as well as other resistance and virulence determinants (e.g., PVL, TSST-1) commonly associated with S. aureus Based on our findings, the growing body of literature on S. argenteus, the potential severity of infections, and possible confusion associated with reporting, including use of incorrect breakpoints for susceptibility results, we make recommendations for clinical laboratories regarding this organism.

Investigation of a severe SARS-CoV-2 outbreak in a long-term care home early in the pandemic.

BACKGROUND: The implementation of outbreak management measures has decreased the frequency and severity of SARS-CoV-2 outbreaks in Ontario long-term care homes. We describe the epidemiological and laboratory data from one of the first such outbreaks in Ontario to assess factors associated with its severity, and the impact of progressive interventions for infection control over the course of the outbreak. METHODS: We obtained line list and outbreak data from the public health unit to describe resident and staff cases, severity and distribution of cases over time and within the outbreak facility. Where available, we obtained data on laboratory specimens from the Public Health Ontario Laboratory and performed whole genome sequencing and phylogenetic analysis of viral specimens from the outbreak. RESULTS: Among 65 residents of the long-term care home, 61 (94%) contracted SARS-CoV-2, with a case fatality rate of 45% (28/61). Among 67 initial staff, 34 (51%) contracted the virus and none died. When the outbreak was declared, 12 staff, 2 visitors and 9 residents had symptoms. Resident cases were located in 3 of 4 areas of the home. Phylogenetic analysis showed tight clustering of cases, with only 1 additional strain of genetically distinct SARS-CoV-2 identified from a staff case in the third week of the outbreak. No cases were identified among 26 new staff brought into the home after full outbreak measures were implemented. INTERPRETATION: Rapid and undetected viral spread in a long-term care home led to high rates of infection among residents and staff. Progressive implementation of outbreak measures after the peak of cases prevented subsequent staff cases and are now part of long-term care outbreak policy in Ontario.

Diagnosis and Management of First Case of COVID-19 in Canada: Lessons Applied From SARS-CoV-1.

We report diagnosis and management of the first laboratory-confirmed case of coronavirus disease 2019 (COVID-19) hospitalized in Toronto, Canada. No healthcare-associated transmission occurred. In the face of a potential pandemic of COVID-19, we suggest sustainable and scalable control measures developed based on lessons learned from severe acute respiratory syndrome.

Importation of Extensively Drug-Resistant Salmonella enterica Serovar Typhi Cases in Ontario, Canada.

A strain of extensively drug-resistant (XDR) Salmonella enterica serovar Typhi has caused a large ongoing outbreak in Pakistan since 2016. In Ontario, Canada, 10 cases of mainly bloodstream infections (n = 9) were identified in patients who traveled to Pakistan. Whole-genome sequencing showed that Canadian cases were genetically related to the Pakistan outbreak strain. The appearance of XDR typhoid cases in Ontario prompted a provincial wide alert to physicians to recommend treatment with carbapenems or azithromycin in suspected typhoid cases with travel history to Pakistan.

Use of Genome Sequencing to Define Institutional Influenza Outbreaks, Toronto, Ontario, Canada, 2014-15.

Adequacy of the current clinical definition of institutional influenza outbreaks is unclear. We performed a retrospective genome sequencing and epidemiologic analysis of institutional influenza outbreaks that occurred during the 2014-15 influenza season in Toronto, Canada. We sequenced the 2 earliest submitted samples positive for influenza A(H3N2) from each of 38 reported institutional outbreaks in long-term care facilities. Genome sequencing showed most outbreak pairs identified by using the current clinical definition were highly related. Inclusion of surveillance samples demonstrated that outbreak sources were likely introductions from broader circulating lineages. Pairwise distance analysis using majority genome and hemagglutinin-specific genes enabled identification of thresholds for discrimination of within and between outbreak pairs; the area under the curve ranged 0.93-0.95. Routine genome sequencing for defining influenza outbreaks in long-term care facilities is unlikely to add significantly to the current clinical definition. Sequencing may prove most useful for investigating sources of outbreak introductions.

Laboratory testing and phylogenetic analysis during a mumps outbreak in Ontario, Canada.

BACKGROUND: In September 2009, a mumps outbreak originated in New York and spread to Northeastern USA and Canada. This study compares the performance of different diagnostic testing methods used in Ontario and describes molecular characteristics of the outbreak strain. METHODS: Between September 2009 and February 2010, specimens from suspect cases were submitted to Public Health Ontario Laboratory for mumps serology, culture and/or real-time reverse-transcriptase PCR (rRT-PCR) testing. rRT-PCR-positive specimens underwent genotyping at Canada's National Microbiology Laboratory. Whole genome sequencing was performed on four outbreak and three sporadic viral culture isolates. RESULTS: Six hundred ninety-eight patients had IgM serology testing, of which 255 (37%) had culture and rRT-PCR. Among those, 35/698 (5%) were IgM positive, 39/255 (15%) culture positive and 47/255 (18%) rRT-PCR-positive. Buccal swabs had the highest rRT-PCR positivity (21%). The outbreak isolates were identical to that in the New York outbreak occurring at the same time. Nucleotide and amino acid identity with the Jeryl Lynn vaccine strain ranged from 85.0-94.5% and 82.4-99.4%, depending on the gene and coding sequences. Homology of the HN protein, the main immunogenic mumps virus protein, was found to be 94.5 and 95.3%, when compared to Jeryl Lynn vaccine major and minor components, respectively. CONCLUSIONS: Despite higher sensitivity than serology, rRT-PCR testing is underutilized. Further work is needed to better understand the suboptimal match of the HN gene between the outbreak strain and the Jeryl Lynn vaccine strain.

Evaluation of Altona Diagnostics RealStar Zika Virus Reverse Transcription-PCR Test Kit for Zika Virus PCR Testing.

With the emerging Zika virus (ZIKV) epidemic, accessible real-time reverse transcription-PCR (rRT-PCR) assays are needed to streamline testing. The commercial Altona Diagnostics RealStar ZIKV rRT-PCR test kit (Altona PCR) has been approved for emergency use authorization by the U.S. FDA. Our aim was to verify the Altona PCR by comparing it to the CDC-designed dual-target ZIKV rRT-PCR reference assay (reference PCR) and describe the demographics of patients tested for ZIKV by rRT-PCR in Ontario, Canada. A large set of clinical specimens was tested for ZIKV by the Altona PCR and the reference PCR. Positive or equivocal specimens underwent PCR and Sanger sequencing targeting the ZIKV NS5 gene. A total of 671 serum specimens were tested by the reference PCR: 58 (8.6%) were positive, 193 (28.8%) were equivocal, and 420 (62.6%) were negative. Ninety percent of the reference PCR-positive patients were tested in the first 5 days after symptom onset. The Altona PCR was performed on 284/671 specimens tested by the reference PCR. The Altona PCR was positive for 53/58 (91%) reference PCR-positive specimens and 16/193 (8%) reference PCR-equivocal specimens; the ZIKV NS5 PCR was positive for all 68 Altona PCR-positive specimens and negative for all 181 Altona PCR-negative specimens that underwent the NS5 PCR. The Altona PCR has very good sensitivity (91%) and specificity (97%) compared to the reference PCR. The Altona PCR can be used for ZIKV diagnostic testing and has less extensive verification requirements than a laboratory-developed test.

A Perfect Storm: Impact of Genomic Variation and Serial Vaccination on Low Influenza Vaccine Effectiveness During the 2014-2015 Season.

BACKGROUND: The 2014-2015 influenza season was distinguished by an epidemic of antigenically-drifted A(H3N2) viruses and vaccine components identical to 2013-2014. We report 2014-2015 vaccine effectiveness (VE) from Canada and explore contributing agent-host factors. METHODS: VE against laboratory-confirmed influenza was derived using a test-negative design among outpatients with influenza-like illness. Sequencing identified amino acid mutations at key antigenic sites of the viral hemagglutinin protein. RESULTS: Overall, 815/1930 (42%) patients tested influenza-positive: 590 (72%) influenza A and 226 (28%) influenza B. Most influenza A viruses with known subtype were A(H3N2) (570/577; 99%); 409/460 (89%) sequenced viruses belonged to genetic clade 3C.2a and 39/460 (8%) to clade 3C.3b. Dominant clade 3C.2a viruses bore the pivotal mutations F159Y (a cluster-transition position) and K160T (a predicted gain of glycosylation) compared to the mismatched clade 3C.1 vaccine. VE against A(H3N2) was -17% (95% confidence interval [CI], -50% to 9%) overall with clade-specific VE of -13% (95% CI, -51% to 15%) for clade 3C.2a but 52% (95% CI, -17% to 80%) for clade 3C.3b. VE against A(H3N2) was 53% (95% CI, 10% to 75%) for patients vaccinated in 2014-2015 only, significantly lower at -32% (95% CI, -75% to 0%) if also vaccinated in 2013-2014 and -54% (95% CI, -108% to -14%) if vaccinated each year since 2012-2013. VE against clade-mismatched B(Yamagata) viruses was 42% (95% CI, 10% to 62%) with less-pronounced reduction from prior vaccination compared to A(H3N2). CONCLUSIONS: Variation in the viral genome and negative effects of serial vaccination likely contributed to poor influenza vaccine performance in 2014-2015.

Spectrum of Viral Pathogens in Blood of Malaria-Free Ill Travelers Returning to Canada.

Malaria is the most common specific cause of fever in returning travelers, but many other vectorborne infections and viral infections are emerging and increasingly encountered by travelers. We documented common and emerging viral pathogens in malaria-negative specimens from ill travelers returning to Canada. Anonymized, malaria-negative specimens were examined for various viral pathogens by real-time PCR. Samples were positive for herpes simplex viruses 1 or 2 (n = 21, 1.6%), cytomegalovirus (n = 4, 0.3%), Epstein-Barr virus (n = 194, 14.9%), dengue virus types 1-4 (n = 27, 2.1%), chikungunya virus (n = 5, 0.4%), and hepatitis A virus (n = 12, 0.9%). Travel-acquired viral pathogens were documented in >20% of malaria-negative specimens, of which 2.5% were infected with dengue and chikungunya viruses. Our findings support the anecdotal impression that these vectorborne pathogens are emerging among persons who travel from Canada to other countries.

Enquête sur une éclosion importante de SRAS-CoV-2 dans un établissement de soins de longue durée au début de la pandémie.

CONTEXTE: Le déploiement de mesures de gestion des éclosions de SRAS-CoV-2 dans les établissements de soins de longue durée en Ontario a permis d'en réduire la fréquence et la gravité. Nous décrivons ici les données épidémiologiques et de laboratoire d'une de ces premières éclosions en Ontario afin de déterminer les facteurs associés à son importance et les impacts des interventions progressives de lutte contre les infections appliquées pendant la durée de l'éclosion. MÉTHODES: Nous avons obtenu du bureau de santé la liste des cas et les données de l'éclosion afin de décrire les cas chez les résidents et le personnel, leur gravité et leur distribution dans le temps et à l'intérieur de l'établissement touché. Quand elles étaient disponibles, nous avons obtenu des données concernant les échantillons soumis au laboratoire de Santé publique Ontario et effectué un séquençage complet et une analyse phylogénétique des échantillons viraux de l'éclosion. RÉSULTATS: Sur les 65 résidents de l'établissement de soins de longue durée, 61 (94 %) ont contracté le SRAS-CoV-2, le taux de létalité étant de 45 % (28/61). Parmi les 67 employés initiaux, 34 (51 %) ont contracté le virus, et aucun n'est décédé. Lorsque l'éclosion a été déclarée, 12 employés, 2 visiteurs et 9 résidents présentaient des symptômes. Parmi les résidents, les cas se trouvaient dans 3 des 4 secteurs de l'établissement. L'analyse phylogénétique a montré une forte similitude des séquences; une seule autre souche de SRAS-CoV-2 génétiquement distincte a été identifiée chez un employé à la troisième semaine de l'éclosion. Après le déploiement de toutes les mesures de gestion de l'éclosion, aucun cas n'a été identifié parmi les 26 nouveaux employés appelés en renfort. INTERPRÉTATION: La propagation rapide et non détectée du virus dans un établissement de soins de longue durée a donné lieu à des taux élevés d'infection chez les résidents et le personnel. L'application progressive de mesures de gestion après le pic de l'éclosion a permis d'éviter la contamination du personnel appelé en renfort et fait désormais partie des politiques à long terme de prévention des éclosions en Ontario.

Mutations acquired during cell culture isolation may affect antigenic characterisation of influenza A(H3N2) clade 3C.2a viruses.

As elsewhere, few (< 15%) sentinel influenza A(H3N2) clade 3C.2a viruses that dominated in Canada during the 2014/15 season could be antigenically characterised by haemagglutination inhibition (HI) assay. Clade 3C.2a viruses that could be HI-characterised had acquired genetic mutations during in vitro cell culture isolation that modified the potential glycosylation motif found in original patient specimens and the consensus sequence of circulating viruses at amino acid positions 158-160 of the haemagglutinin protein. Caution is warranted in extrapolating antigenic relatedness based on limited HI findings for clade 3C.2a viruses that continue to circulate globally.

Interim estimates of 2015/16 vaccine effectiveness against influenza A(H1N1)pdm09, Canada, February 2016.

Using a test-negative design, the Canadian Sentinel Practitioner Surveillance Network (SPSN) assessed interim 2015/16 vaccine effectiveness (VE) against influenza A(H1N1)pdm09 viruses. Adjusted VE showed significant protection of 64% (95% confidence interval (CI): 44-77%) overall and 56% (95%CI: 26-73%) for adults between 20 and 64 years-old against medically attended, laboratory-confirmed A(H1N1)pdm09 illness. Among the 67 A(H1N1)pdm09-positive specimens that were successfully sequenced, 62 (> 90%) belonged to the emerging genetic 6B.1 subclade, defined by S162N (potential gain of glycosylation) and I216T mutations in the haemagglutinin protein. Findings from the Canadian SPSN indicate that the 2015/16 northern hemisphere vaccine provided significant protection against A(H1N1)pdm09 illness despite genetic evolution in circulating viruses.

Integrated Sentinel Surveillance Linking Genetic, Antigenic, and Epidemiologic Monitoring of Influenza Vaccine-Virus Relatedness and Effectiveness During the 2013-2014 Influenza Season.

BACKGROUND: Canada's Sentinel Physician Surveillance Network links genetic, antigenic, and vaccine effectiveness (VE) measures in an integrated platform of influenza monitoring, described here for the 2013-2014 influenza season of resurgent A(H1N1)pdm09 and late-season type B activity. METHODS: VE was estimated as [1 - odds ratio] × 100% and compared vaccination status between individuals who tested positive (cases) and those who tested negative (controls) for influenza virus. Vaccine-virus relatedness was assessed by genomic sequence analysis and hemagglutination inhibition assays. RESULTS: Analyses included 1037 controls (of whom 33% were vaccinated) and 663 cases (of whom 14% were vaccinated). A total of 415 cases tested positive for A(H1N1)pdm09 virus, 15 tested positive for A(H3N2) virus, 191 tested positive for B/Yamagata-lineage virus, 6 tested positive for B/Victoria-lineage virus, and 36 tested positive for viruses of unknown subtype or lineage. A(H1N1)pdm09 viruses belonged to clade 6B, distinguished by a K163Q substitution, but remained antigenically similar to the A/California/07/2009-like vaccine strain, with an adjusted VE of 71% (95% confidence interval [CI], 58%-80%). Most B/Yamagata-lineage viruses (83%) clustered phylogenetically with the prior (ie, 2012-2013) season's B/Wisconsin/01/2010-like clade 3 vaccine strain, while only 17% clustered with the current (ie, 2013-2014) season's B/Massachusetts/02/2012-like clade 2 vaccine strain. The adjusted VE for B/Yamagata-lineage virus was 73% (95% CI, 57%-84%), with a lower VE obtained after partial calendar-time adjustment for clade-mismatched B/Wisconsin/01/2010-like virus (VE, 63%; 95% CI, 41%-77%), compared with that for clade-matched B/Massachusetts/02/2012-like virus (VE, 88%; 95% CI, 48%-97%). No A(H3N2) viruses clustered with the A/Texas/50/2012-like clade 3C.1 vaccine strain, and more than half were antigenically mismatched, but sparse data did not support VE estimation. CONCLUSIONS: VE corresponded with antigenically conserved A(H1N1)pdm09 and lineage-matched B/Yamagata viruses with clade-level variation. Surveillance linking genotypic, phenotypic, and epidemiologic measures of vaccine-virus relatedness and effectiveness could better inform predictions of vaccine performance and reformulation.

Vancomycin-variable enterococci can give rise to constitutive resistance during antibiotic therapy.

Vancomycin-resistant enterococci (VRE) are notorious clinical pathogens restricting the use of glycopeptide antibiotics in the clinic setting. Routine surveillance to detect VRE isolated from patients relies on PCR bioassays and chromogenic agar-based test methods. In recent years, we and others have reported the emergence of enterococcal strains harboring a "silent" copy of vancomycin resistance genes that confer a vancomycin-susceptible phenotype (vancomycin-susceptible enterococci [VSE]) and thus escape detection using drug sensitivity screening tests. Alarmingly, these strains are able to convert to a resistance phenotype (VSE→VRE) during antibiotic treatment, severely compromising the success of therapy. Such strains have been termed vancomycin-variable enterococci (VVE). We have investigated the molecular mechanisms leading to the restoration of resistance in VVE isolates through the whole-genome sequencing of resistant isolates, measurement of resistance gene expression, and quantification of the accumulation of drug-resistant peptidoglycan precursors. The results demonstrate that VVE strains can revert to a VRE phenotype through the constitutive expression of the vancomycin resistance cassette. This is accomplished through a variety of changes in the DNA region upstream of the resistance genes that includes both a deletion of a likely transcription inhibitory secondary structure and the introduction of a new unregulated promoter. The VSE→VRE transition of VVE can occur in patients during the course of antibiotic therapy, resulting in treatment failure. These VVE strains therefore pose a new challenge to the current regimen of diagnostic tests used for VRE detection in the clinic setting.

Characterization of an Enterococcus gallinarum Isolate Carrying a Dual vanA and vanB Cassette.

The ability of vancomycin resistance determinants to be horizontally transferred within enterococci species is a concern. Identification and characterization of vancomycin-resistant enterococci (VRE) in a clinical isolate have a significant impact on infection control practices. In this study, we describe a clinical isolate of Enterococcus gallinarum exhibiting high-level resistance to vancomycin and teicoplanin. The genetic characterization of this isolate showed the presence of vanA and vanB genes in addition to the naturally carried vanC gene. vanA was identified on pA6981, a 35,608-bp circular plasmid with significant homology to plasmid pS177. The vanB operon was integrated into the bacterial chromosome and showed a high level of homology to previously reported Tn1549 and Tn5382. To the best of our knowledge, this is the first report of E. gallinarum carrying both vanA and vanB operons, indicating the importance of identifying the vancomycin resistance mechanism in non-E. faecium and non-E. faecalis enterococcal species.