Interlaboratory comparison of Pseudomonas aeruginosa phage susceptibility testing.

Standardized approaches to phage susceptibility testing (PST) are essential to inform selection of phages for study in patients with bacterial infections. There is no reference standard for assessing bacterial susceptibility to phage. We compared agreement between PST performed at three centers: two centers using a liquid assay standardized between the sites with the third, a plaque assay. Four Pseudomonas aeruginosa phages: PaWRA01ø11 (EPa11), PaWRA01ø39 (EPa39), PaWRA02ø83 (EPa83), PaWRA02ø87 (EPa87), and a cocktail of all four phages were tested against 145 P. aeruginosa isolates. Comparisons were made within measurements at the two sites performing the liquid assay and between these two sites. Agreement was assessed based on coverage probability (CP8), total deviation index, concordance correlation coefficient (CCC), measurement accuracy, and precision. For the liquid assay, there was satisfactory agreement among triplicate measurements made on different days at site 1, and high agreement based on accuracy and precision between duplicate measurements made on the same run at site 2. There was fair accuracy between measurements of the two sites performing the liquid assay, with CCCs below 0.6 for all phages tested. When compared to the plaque assay (performed once at site 3), there was less agreement between results of the liquid and plaque assays than between the two sites performing the liquid assay. Similar findings to the larger group were noted in the subset of 46 P. aeruginosa isolates from cystic fibrosis. Results of this study suggest that reproducibility of PST methods needs further development.

Continuing the quality improvement of an electronic personal health record and interactive website for people with diabetes in Scotland (My Diabetes My Way).

AIMS: eHealth applications have the potential to enable patients to take more control over managing their own health, helping to delay and prevent complications. My Diabetes My Way (MDMW) is an electronic personal health record/educational platform available to people with diabetes in Scotland. This study aims to assess user experience with respect to demographic subgroups, examine effectiveness of previous improvements made to the platform and inform its ongoing development. METHODS: All active MDMW users (22,665) were invited to take part in a questionnaire combining Likert scale and free-response items relating to system utility. Likert responses were used to generate a 'utility score'. This was used in regression analyses to determine predictors of system utility scoring. Free-response answers were analysed thematically and themes were generated. RESULTS: A total of 4713 (21%) MDMW users responded to the questionnaire. Most agreed that MDMW helps them to track changes over time, prepare for face-to-face consultations, remember information discussed in consultations and reduced the need to contact their general practitioner. Free-response answers showed that users valued earlier enhancements made to the site (e.g. linking Fitbit data), and highlighted areas needing further improvement. Evidence of the 'digital divide' was seen in respondent demographics, and some users mentioned 'lack of digital skills' as a barrier to engaging with the platform. CONCLUSIONS: User experience of MDMW was positive. Users agreed with statements that MDMW facilitates diabetes self management. Several areas of potential improvement were identified, including linking more wearable device data, and assisting/directing users to gain the digital skills required to engage fully with MDMW.

Student pharmacist practice-based interprofessional education in Scotland: a qualitative study of stakeholders' views and experiences.

Scottish Government funding supports practice-based experiential learning (EL) for student pharmacists. We explored views and experiences of key stakeholders on current practice and future development of interprofessional education (IPE) in EL including barriers and enablers. A pre-piloted schedule was used for online qualitative semi-structured interviews. eMail invitations were sent to 37 stakeholders with an information sheet and consent process. Interviews were analyzed thematically by two researchers independently. Recruitment continued until data saturation and wide representation were achieved. Twenty interviews were conducted with eight EL facilitators, seven faculty and five policy stakeholders. "Nature and experience of current IPE in EL activities" and "Future developments" were the two main themes. Barriers and enablers were also identified at macro, meso, and micro socio-institutional levels. The essence of the analysis highlighted stakeholders' views of the importance of building on current IPE while challenging the ethos and culture of EL practices. All stakeholders should be involved in co-production, training, piloting, and evaluation of curricular developments to overcome logistic barriers and enhanced enablers. Finally, the importance of workload management strategies and continuity of funding for success was also stressed by those interviewed. Future research could include designing frameworks for developing and implementing IPE within EL.

Macrolide Resistance in Mycoplasma pneumoniae, Midwestern United States, 2014 to 2021.

The epidemiology of macrolide resistance in Mycoplasma (Mycoplasmoides) pneumoniae in the United States is incompletely described. Using a PCR assay targeting common mutations associated with macrolide resistance in M. pneumoniae (23S rRNA gene, A2063G/A2064G), the frequency of macrolide resistance was estimated to be 10% based on analysis of 114 samples tested from January 2014 to September 2021 at Mayo Clinic Laboratories. Seasonality data showed the highest rates of M. pneumoniae infection in the fall/early winter.

Activity of Omadacycline in Rat Methicillin-Resistant Staphylococcus aureus Osteomyelitis.

Omadacycline, vancomycin, and rifampin, as well as rifampin combination therapies, were evaluated in an experimental rat model of methicillin-resistant Staphylococcus aureus (MRSA) osteomyelitis. All treatment groups had less MRSA recovered than saline-treated animals. The emergence of rifampin resistance was observed in 3 of 16 animals with rifampin monotherapy and none with rifampin combination therapy. After treatment, the median tibial bacterial loads were 6.04, 0.1, 4.81, and 5.24 log10 CFU/g for saline-, rifampin-, vancomycin-, and omadacycline-treated animals, respectively. Omadacycline or vancomycin administered with rifampin yielded no detectable MRSA. Omadacycline administered with rifampin deserves evaluation in humans as a potential treatment for osteomyelitis.

Considerations for the Use of Phage Therapy in Clinical Practice.

Increasing antimicrobial resistance and medical device-related infections have led to a renewed interest in phage therapy as an alternative or adjunct to conventional antimicrobials. Expanded access and compassionate use cases have risen exponentially but have varied widely in approach, methodology, and clinical situations in which phage therapy might be considered. Large gaps in knowledge contribute to heterogeneity in approach and lack of consensus in many important clinical areas. The Antibacterial Resistance Leadership Group (ARLG) has convened a panel of experts in phage therapy, clinical microbiology, infectious diseases, and pharmacology, who worked with regulatory experts and a funding agency to identify questions based on a clinical framework and divided them into three themes: potential clinical situations in which phage therapy might be considered, laboratory testing, and pharmacokinetic considerations. Suggestions are provided as answers to a series of questions intended to inform clinicians considering experimental phage therapy for patients in their clinical practices.

Plasmid Acquisition Alters Vancomycin Susceptibility in Clostridioides difficile.

The increasing incidence of primary and recurring Clostridioides difficile infections (CDI), which evade current treatment strategies, reflects the changing biology of C difficile. Here, we describe a putative plasmid-mediated mechanism potentially driving decreased sensitivity of C difficile to vancomycin treatment. We identified a broad host range transferable plasmid in a C difficile strain associated with lack of adequate response to vancomycin treatment. The transfer of this plasmid to a vancomycin-susceptible C difficile isolate decreased its susceptibility to vancomycin in vitro and resulted in more severe disease in a humanized mouse model. Our findings suggest plasmid acquisition in the gastrointestinal tract to be a possible mechanism underlying vancomycin treatment failure in patients with CDI, but further work is needed to characterize the mechanism by which plasmid genes determine vancomycin susceptibility in C difficile.

Development and Evaluation of a Core Genome Multilocus Sequencing Typing (cgMLST) Scheme for Serratia marcescens Molecular Surveillance and Outbreak Investigations.

Serratia marcescens can cause a range of severe infections and contributes to nosocomial outbreaks. Although whole-genome sequencing (WGS)-based typing is the standard method for molecular surveillance and outbreak investigation, there is no standardized analytic scheme for S. marcescens core genome multilocus sequence typing (cgMLST). Here, the development and evaluation of a S. marcescens cgMLST scheme is reported with the goal of enabling a standardized methodology and typing nomenclature. Four hundred ninety-one high-quality S. marcescens WGS data sets were extracted from public databases and-using the genomic sequence of NCBI reference strain S. marcescens Db11 (NZ_HG326223.1) as a starting point-all Db11 genes present in ≥97% data sets used to create a cgMLST scheme. The novel scheme was evaluated using WGS data from 24 outbreak investigations (n = 175 isolates) distributed over three continents. Analysis of Db11 genes within the 491 data sets identified 2,692 target genes present in ≥97% of genomes (mean, 99.1%; median, 99.9%). These genes formed the novel cgMLST scheme, covering 47.8% of nucleotides in the Db11 genome. Analyzing 175 isolates from 24 outbreaks using the novel scheme gave comparable results to previous typing efforts for both general groupings and allelic distances within clusters. In summary, a novel cgMLST scheme for S. marcescens was developed and evaluated. The scheme and its associated nomenclature will improve standardization of typing efforts for molecular surveillance and outbreak investigation, allowing better understanding of S. marcescens genomic epidemiology and facilitating interlaboratory comparisons.

Acinetobacter baumannii Genomic Sequence-Based Core Genome Multilocus Sequence Typing Using Ridom SeqSphere+ and Antimicrobial Susceptibility Prediction in ARESdb.

Whole-genome sequencing (WGS) is rapidly replacing traditional typing methods for the investigation of infectious disease outbreaks. Additionally, WGS data are being used to predict phenotypic antimicrobial susceptibility. Acinetobacter baumannii, which is often multidrug-resistant, is a significant culprit in outbreaks in health care settings. A well-characterized collection of A. baumannii was studied using core genome multilocus sequence typing (cgMLST). Seventy-two isolates previously typed by PCR-electrospray ionization mass spectrometry (PCR/ESI-MS) provided by the Antimicrobial Resistance Leadership Group (ARLG) were analyzed using a clinical microbiology laboratory developed workflow for cgMLST with genomic susceptibility prediction performed using the ARESdb platform. Previously performed PCR/ESI-MS correlated with cgMLST using relatedness thresholds of allelic differences of ≤9 and ≤200 allelic differences in 78 and 94% of isolates, respectively. Categorical agreement between genotypic and phenotypic antimicrobial susceptibility across a panel of 11 commonly used drugs was 89%, with minor, major, and very major error rates of 8%, 11%, and 1%, respectively.

An innovative General Practice based Pharmacy Longitudinal Clerkship: using theory to characterise its development, implementation and initial evaluation.

BACKGROUND: Longitudinal Integrated Clerkships exist in undergraduate medicine courses. A pilot Pharmacy Longitudinal Clerkship (pPLC) was funded to investigate delivery of this model of clinical education for student pharmacists. OBJECTIVE(S): To investigate the development, implementation and initial evaluation of a pPLC. METHODS: The 11-week pPLC was delivered to two students in two GP practices in Scotland. Mixed theory-based methods were used to gather information on the pPLC structures and processes required and qualitative semi-structured Theoretical Domains Framework (TDF) based interviews explored outcomes with key stakeholders. Informed written consent was obtained. Interviews were audio-recorded, transcribed verbatim and analysed thematically. University Ethics approval was granted. RESULTS: Data were generated on resources and processes required for a pPLC including funds budgeted for and actually spent on staffing, student travel/subsistence and student clinical 'Kit Bags', learning outcomes, curriculum and training timetable, GP Practice/University contracts. Interviews were completed with the two students, three linked GP clinical supervisors and two Regional Tutors involved. The seven themes were identified and mapped to seven TDF domains including: increased levels of student confidence, and increased student enthusiasm for a career in pharmacy, need for definition of the role of the Regional Tutor for the PLC and GP positivity towards the expected outcomes of clerkship model versus traditional placements. CONCLUSION: Findings are limited by the small number of participants and settings, but evaluation was positive and the work garnered information on requirements for resources and processes. This will inform 'roll out' of the PLC.

Clostridioides difficile Whole-genome Sequencing Differentiates Relapse With the Same Strain From Reinfection With a New Strain.

BACKGROUND: Current approaches in tracking Clostridioides difficile infection (CDI) and individualizing patient management are incompletely defined. METHODS: We recruited 468 subjects with CDI at Mayo Clinic Rochester between May and December 2016 and performed whole-genome sequencing (WGS) on C. difficile isolates from 397. WGS was also performed on isolates from a subset of the subjects at the time of a recurrence of infection. The sequence data were analyzed by determining core genome multilocus sequence type (cgMLST), with isolates grouped by allelic differences and the predicted ribotype. RESULTS: There were no correlations between C. difficile isolates based either on cgMLST or ribotype groupings and CDI outcome. An epidemiologic assessment of hospitalized subjects harboring C. difficile isolates with ≤2 allelic differences, based on standard infection prevention and control assessment, revealed no evidence of person-to-person transmission. Interestingly, community-acquired CDI subjects in 40% of groups with ≤2 allelic differences resided within the same zip code. Among 18 subjects clinically classified as having recurrent CDI, WGS revealed 14 with initial and subsequent isolates differing by ≤2 allelic differences, suggesting a relapse of infection with the same initial strain, and 4 with isolates differing by >50 allelic differences, suggesting reinfection. Among the 5 subjects classified as having a reinfection based on the timing of recurrence, 3 had isolates with ≤2 allelic differences between them, suggesting a relapse, and 2 had isolates differing by >50 allelic differences, suggesting reinfection. CONCLUSIONS: Our findings point to potential transmission of C. difficile in the community. WGS better differentiates relapse from reinfection than do definitions based on the timing of recurrence.

Core Genome Multilocus Sequence Typing and Prediction of Antimicrobial Susceptibility Using Whole-Genome Sequences of Escherichia coli Bloodstream Infection Isolates.

In total, 50 Escherichia coli bloodstream isolates from the clinical laboratory and 12 E. coli isolates referred for pulsed-field gel electrophoresis (PFGE) were sequenced, assessed for clonality using core genome multilocus sequence typing (cgMLST), and evaluated for genomic susceptibility predictions using ARESdb. Results of sequence typing using whole-genome sequencing (WGS)-based MLST and sequence type (ST)-specific PCR were identical. Overall categorical agreement between genotypic (ARESdb) and phenotypic susceptibility testing for 62 isolates and 11 antimicrobial agents was 91%. Among the referred isolates, high major error rates were found for ceftazidime, cefepime, and piperacillin-tazobactam.

Clinical Evaluation of a Real-Time PCR Assay for Simultaneous Detection of Helicobacter pylori and Genotypic Markers of Clarithromycin Resistance Directly from Stool.

Helicobacter pylori infection is mainly diagnosed noninvasively, with susceptibility testing traditionally requiring endoscopy. Treatment is empirical, with clarithromycin-based triple therapy recommended where resistance rates are below 15%. Rising rates of clarithromycin resistance, resulting in high clarithromycin-based therapy failure rates, are seen worldwide, but U.S. data are limited. We developed a real-time PCR assay for simultaneous detection of H. pylori and genotypic markers of clarithromycin resistance directly from stool specimens. The assay was validated by testing 524 stool samples using an H. pylori stool antigen test as the reference method for detection accuracy and Sanger sequencing to confirm genotypic susceptibility results. A separate set of 223 antigen-positive stool samples was tested and retrospective medical record review conducted to define clinical utility. PCR resulted in 88.6% and 92.8% sensitivity in the validation and clinical study sets, respectively. Sequencing confirmed correct detection of clarithromycin resistance-associated mutations in all positive validation samples. The PCR-predicted clarithromycin resistance rate was 39% in the clinical data set overall and 31% in treatment-naive patients; the clarithromycin-based triple therapy eradication rate in treatment-naive patients was 62%. The clarithromycin-based triple therapy success was lower when resistance was predicted by PCR (41%) than when no resistance was predicted (70%; P = 0.03). PCR results were positive in 98% of antigen-positive stools from patients tested for eradication. The described PCR assay can accurately and noninvasively diagnose H. pylori, provide genotypic susceptibility, and test for eradication. Our findings support the need for susceptibility-guided therapy in our region if a clarithromycin-based regimen is considered.

Yersinia occitanica is a later heterotypic synonym of Yersinia kristensenii subsp. rochesterensis and elevation of Yersinia kristensenii subsp. rochesterensis to species status.

The taxonomic position of Yersinia kristensenii subsp. rochesterensis and Yersinia occitanica was re-evaluated by genomic analysis. Average nucleotide identity (ANI), digital DNA-DNA hybridization values, and phylogenetic analyses of the type strains indicate that Y. kristensenii subsp. rochesterensis and Y. occitanica are the same genospecies. Additionally, the overall genomic relatedness index (OGRI) values reveal that Y. kristensenii subsp. rochesterensis should be elevated to species status as Yersinia rochesterensis sp. nov.

Molecular Approach to Diagnosis of Cardiovascular Implantable Electronic Device Infection.

BACKGROUND: Sonicate fluid (SF), a solution derived from vortexing and sonication of explanted cardiovascular implantable electronic devices (CIEDs), is a higher-yield specimen compared with swabs or tissues for culture-based detection of microorganisms associated with CIED infection. Despite this, SF culture fails to identify a causative organism in ~50% of cases. We aimed to evaluate the diagnostic performance of 16S ribosomal RNA gene (rRNA) polymerase chain reaction (PCR)/sequencing of SF and compare it with that of SF culture. METHODS: We identified 322 SF specimens from extracted CIEDs and reviewed clinical data for each patient. Subjects were classified as having or not having CIED infection. Cases were subcategorized as culture negative if no significant growth was reported from SF cultures and as culture positive if an organism was detected above predefined thresholds. 16S rRNA PCR/sequencing was performed, with the organisms identified reported according to Clinical and Laboratory Standards Institute guidelines for sequence data interpretation. RESULTS: A total of 278 SF samples corresponded to infected cases, of which 160 were culture positive and 118 culture negative. The remaining 44 were from noninfected cases, of which 2 were culture positive. Compared with SF culture, the sensitivity of 16S rRNA PCR/sequencing was higher (64% vs 57.5%, P = .003). 16S rRNA PCR/sequencing detected a potential pathogen in 28 of 118 culture-negative cases, identifying staphylococci in the majority (18/28). CONCLUSIONS: 16S rRNA PCR/sequencing has higher sensitivity to detect bacteria in SF from extracted CIEDs than does SF culture.

The use of theory in the development and evaluation of behaviour change interventions to improve antimicrobial prescribing: a systematic review.

OBJECTIVES: This systematic review (SR) reviews the evidence on use of theory in developing and evaluating behaviour change interventions (BCIs) to improve clinicians' antimicrobial prescribing (AP). METHODS: The SR protocol was registered with PROSPERO. Eleven databases were searched from inception to October 2018 for peer-reviewed, English-language, primary literature in any healthcare setting and for any medical condition. This included research on changing behavioural intentions (e.g. in simulated scenarios) and research measuring actual AP. All study designs/methodologies were included. Excluded were: grey literature and/or those which did not state a theory. Two reviewers independently extracted and quality assessed the data. The Theory Coding Scheme (TCS) evaluated the extent of the use of theory. RESULTS: Searches found 4227 potentially relevant papers after removal of duplicates. Screening of titles/abstracts led to dual assessment of 38 full-text papers. Ten (five quantitative, three qualitative and two mixed-methods) met the inclusion criteria. Studies were conducted in the UK (n = 8), Canada (n = 1) and Sweden (n = 1), most in primary care settings (n = 9), targeting respiratory tract infections (n = 8), and medical doctors (n = 10). The most common theories used were Theory of Planned Behaviour (n = 7), Social Cognitive Theory (n = 5) and Operant Learning Theory (n = 5). The use of theory to inform the design and choice of intervention varied, with no optimal use as recommended in the TCS. CONCLUSIONS: This SR is the first to investigate theoretically based BCIs around AP. Few studies were identified; most were suboptimal in theory use. There is a need to consider how theory is used and reported and the systematic use of the TCS could help.

A cross-sectional survey of the perspectives of older people in the Scottish Highlands on the management of their chronic pain.

BACKGROUND: Although there is evidence of suboptimal outcomes in older people with chronic pain, little emphasis has been placed on those in remote and rural settings. OBJECTIVE: To describe the perspectives of older people in the Scottish Highlands on their chronic pain management. DESIGN: Cross-sectional survey. SETTING: NHS Highland, the most remote and rural geographical health board in Scotland. SUBJECTS: Home-dwelling members of the public aged ≥70 years. METHODS: Anonymised questionnaires were mailed to a random sample of 1800 older people. Questionnaire items were demographics, nature of any chronic pain, management regimens and perceived effectiveness. Validated scales were the Pain Disability Questionnaire and the Tampa Scale for Kinesiophobia. RESULTS: Adjusted response rate was 39.3% (709/1755). One-quarter (25.0%, n = 177) were experiencing chronic pain, being more likely to live in deprived areas (P < 0.05). Median pain intensity was 6 (IQR 4-7, 10 high), causing distress (median 5, IQR 3-7). Respondents largely consulted GPs (66.1%, n = 117) with a minority (16.4%, n = 29) referred to a specialist pain clinic and few consulting other health professionals. Over three quarters (78.0%, n = 138) were receiving prescribed medicines, most commonly paracetamol, alone (35.6%, n = 63) or in combination with opioids (16.4%, n = 29). One-third (31.6%, n = 56) expressed a desire for more effective medicines; few reported using any non-pharmacological therapies. The median scores for the Pain Disability Questionnaire and Tampa Scale for Kinesiophobia were 74 (IQR 34-104.5, 150 high) and 40 (IQR 35-45, 68 high). CONCLUSIONS: Evidence of provision of appropriate integrated and person-centred chronic pain care is lacking.

A cross-sectional survey of the pharmacy workforce in general practice in Scotland.

BACKGROUND: In Scotland, there has been significant investment in pharmacy teams in general medical practices over recent years, aligned to current government policy. OBJECTIVES: To characterize the national pharmacy workforce including activities undertaken, perceived competence and confidence, as well as perception of integration of the intervention. METHODS: A cross-sectional survey of all pharmacists and pharmacy technicians in general practices. Survey items were demographics, activities undertaken and experiences. The NoMAD tool (Improving the Normalization of Complex Interventions) was included as a measure of perspectives of implementation. Post-piloting, a questionnaire link was sent to all pharmacists (n = 471) and pharmacy technicians (n = 112). A total NoMAD score was obtained by assigning 1 (strongly disagree) to 5 (strongly agree) to each item. RESULTS: Responses were received from 393 (83.4%) pharmacists and 101 (91.8%) pharmacy technicians. Three quarters of pharmacists (74.6%) and pharmacy technicians (73.3%) had been qualified for over 10 years. Two-thirds of pharmacists (68.4%) were independent prescribers, with three quarters (72.3%) currently prescribing. Respondents worked in a median of two practices and were providing a range of activities including medication/polypharmacy reviews, medicines reconciliation, prescribing efficiencies and training. Respondents reported high levels of competence and confidence (median 8, scale 0-10 highest). Median NoMAD total score (scale 20-100 highest, Cronbach's alpha 0.89) was 80 for pharmacists and 75 for pharmacy technicians, P ≤ 0.001. CONCLUSIONS: The general practice pharmacy workforce in Scotland is experienced, well-qualified and integrated within general practices, delivering a range of activities. These findings have implications for workforce planning and future education and training.

Lack of correlation of virulence gene profiles of Staphylococcus aureus bacteremia isolates with mortality.

PURPOSE: Whole genome sequencing (WGS) analysis of Staphylococcus aureus is increasingly used in clinical practice. Although bioinformatics tools used in WGS analysis readily define the S. aureus virulome, the clinical value of this type of analysis is unclear. Here, virulence genes in S. aureus bacteremia (SAB) isolates were evaluated by WGS, with superantigens (SAgs) further evaluated by conventional PCR and functional assays, and results correlated with mortality. METHODS: 152 SAB isolates collected throughout 2015 at a large Minnesota medical center were studied and associated clinical data analyzed. Virulence genes were identified from previously-reported WGS data (https://doi.org/10.1371/journal.pone.0179003). SAg genes sea, seb, sec, sed, see, seg, seh, sei, sej, and tst were also assessed by individual PCR assays. Mitogenicity of SAgs was assessed using an in vitro proliferation assay with splenocytes from HLA-DR3 transgenic mice. RESULTS: Of the 152 SAB isolates studied, 106 (69%) were methicillin-susceptible S. aureus (MSSA). The number of deaths attributed and not attributed to SAB, and 30-day survivors were 24 (16%), 2 (1%), and 128 (83%), respectively. From WGS data, both MSSA and MRSA had high proportions of adhesion (>80%) and immune-evasion (>70%) genes. There was no difference in virulomes between survivor- and non-survivor-associated isolates. Although over 60% of SAB isolates produced functional SAgs, there were no differences in the distribution or prevalence of SAg genes between survivor- and non-survivor-associated isolates. CONCLUSION: In this study of one year of SAB isolates from a large medical center, the S. aureus virulome, as assessed by WGS, and also for SAgs using individual PCRs and phenotypic characterization, did not impact mortality.

Yersinia kristensenii subsp. rochesterensis subsp. nov., isolated from human feces.

A single bacterial isolate, EPLC-04T, was isolated from human feces and identified as representing a member of the genus Yersinia on the basis of phenotypic characteristics, matrix assisted laser desorption ionization time-of-flight mass spectrometry and partial 16S rRNA gene sequencing. The isolate's phenotypic profile differed from that described for the most closely related species, Yersinia kristensenii, by exhibiting lipase production and lacking pyrazinamidase activity. Multiple genetic targets, including the complete (1465 bp) 16S rRNA gene sequence and partial sequences of groEL (539 bp), gyrB (935 bp), glnA (525 bp) and recA (535 bp) indicated that the isolate exhibited 98.91, 92.16, 90.81, 92.78 and 89.01 % identity with Yersinia aldovae, 98.98, 91.99, 90.17, 89.77 and 89.55 % identity with Yersinia intermedia, and 99.66, 98.11, 98.50, 98.49 and 98.51 % identity with Y. kristensenii, respectively. Phylogenetic reconstructions based on the combination of the four housekeeping genes indicated that the isolate formed a unique branch, supported by a bootstrap value of 100 %. Digital DNA-DNA homology and 16S rRNA gene sequencing identified EPLC-04T as representing Y. kristensenii. However, the unique phenotypic traits and results of phylogenetic analysis indicate that it represents a novel subspecies of Y. kristensenii. The name Yersinia kristenseniisubsp. rochesterensis subsp. nov. is proposed for this novel taxon (type strain EPLC-04T=ATCC BAA-2637T, DSMZ 28595T).