An update on the taxonomy and functional properties of the probiotic Enterococcus faecium SF68.

Enterococcus faecium SF68 (SF68) is a well-known probiotic with a long history of safe use. Recent changes in the taxonomy of enterococci have shown that a novel species, Enterococcus lactis, is closely related with E. faecium and occurs together with other enterococci in a phylogenetically well-defined E. faecium species group. The close phylogenetic relationship between the species E. faecium and E. lactis prompted a closer investigation into the taxonomic status of E. faecium SF68. Using phylogenomics and ANI, the taxonomic analysis in this study showed that probiotic E. faecium SF68, when compared to other E. faecium and E. lactis type and reference strains, could be re-classified as belonging to the species E. lactis. Further investigations into the functional properties of SF68 showed that it is potentially capable of bacteriocin production, as a bacteriocin gene cluster encoding the leaderless bacteriocin EntK1 together with putative Lactococcus lactis bacteriocins LsbA, and LsbB-like putative immunity peptide (LmrB) were found located in an operon on plasmid pF9. However, bacteriocin expression was not studied. Competitive exclusion experiments in co-culture over 7 days at 37 °C showed that the probiotic SF68 could inhibit the growth of specific E. faecium and Listeria monocytogenes strains, while showing little or no inhibitory activity towards an entero-invasive Escherichia coli and a Salmonella Typhimurium strain, respectively. In cell culture experiments with colon carcinoma HT29 cells, the probiotic SF68 was also able to strain-specifically inhibit adhesion and/or invasion of enterococcal and L. monocytogenes strains, while such adhesion and invasion inhibition effects were less pronounced for E. coli and Salmonella strains. This study therefore provides novel data on the taxonomy and functional properties of SF68, which can be reclassified as Enterococcus lactis SF68, thereby enhancing the understanding of its probiotic nature.

Optimising genomic approaches for identifying vancomycin-resistant Enterococcus faecium transmission in healthcare settings.

Vancomycin-resistant Enterococcus faecium (VREfm) is a major nosocomial pathogen. Identifying VREfm transmission dynamics permits targeted interventions, and while genomics is increasingly being utilised, methods are not yet standardised or optimised for accuracy. We aimed to develop a standardized genomic method for identifying putative VREfm transmission links. Using comprehensive genomic and epidemiological data from a cohort of 308 VREfm infection or colonization cases, we compared multiple approaches for quantifying genetic relatedness. We showed that clustering by core genome multilocus sequence type (cgMLST) was more informative of population structure than traditional MLST. Pairwise genome comparisons using split k-mer analysis (SKA) provided the high-level resolution needed to infer patient-to-patient transmission. The more common mapping to a reference genome was not sufficiently discriminatory, defining more than three times more genomic transmission events than SKA (3729 compared to 1079 events). Here, we show a standardized genomic framework for inferring VREfm transmission that can be the basis for global deployment of VREfm genomics into routine outbreak detection and investigation.

Genome-based studies indicate that the Enterococcus faecium Clade B strains belong to Enterococcus lactis species and lack of the hospital infection associated markers.

Enterococcus lactis and the heterotypic synonym Enterococcus xinjiangensis from dairy origin have recently been identified as a novel species based on 16S rRNA gene sequence analysis. Enterococcus faecium type strain NCTC 7171T was used as the reference genome for determining E. lactis and E. faecium to be separate species. However, this taxonomic classification did not consider the diverse lineages of E. faecium, and the double nature of hospital-associated (clade A) and community-associated (clade B) isolates. Here, we investigated the taxonomic relationship among isolates of E. faecium of different origins and E. lactis, using a genome-based approach. Additional to 16S rRNA gene sequence analysis, we estimated the relatedness among strains and species using phylogenomics based on the core pangenome, multilocus sequence typing, the average nucleotide identity and digital DNA-DNA hybridization. Moreover, following the available safety assessment schemes, we evaluated the virulence profile and the ampicillin resistance of E. lactis and E. faecium clade B strains. Our results confirmed the genetic and evolutionary differences between clade A and the intertwined clade B and E. lactis group. We also confirmed the absence in these strains of virulence gene markers IS16, hylEfm and esp and the lack of the PBP5 allelic profile associated with ampicillin resistance. Taken together, our findings support the reassignment of the strains of E. faecium clade B as E. lactis.

Evaluation of Enterococcus faecium NRRL B-2354 as a surrogate for Salmonella in ground black pepper at different water activities.

Thermal inactivation kinetics of Salmonella in low moisture foods are necessary for developing proper thermal processing parameters for pasteurization. The effect of water activity on thermal inactivation kinetics of Salmonella and Enterococcus faecium NRRL B-2354 in ground black pepper has not been studied previously. Identification of a suitable surrogate assists in conducting in-plant process validations. Ground black pepper was inoculated with a 5-serotype Salmonella cocktail or E. faecium NRRL B-2354, equilibrated to water activities of 0.25, 0.45 or 0.65 in a humidity-controlled chamber, and isothermally treated at different temperatures. The survivor data were used for fitting the log-linear models to obtain the D and z-values of Salmonella and E. faecium in ground black pepper. Modified Bigelow models were developed to evaluate the effects of temperature and water activity on the thermal inactivation kinetics of Salmonella and E. faecium. Water activity and temperature showed significant negative effects on the thermal resistance of Salmonella and E. faecium in ground black pepper. For example, significantly higher D values of Salmonella were observed at water activity of 0.45 (D70°C = 20.5 min and D75°C = 7.8 min) compared to water activity of 0.65 (D70°C = 3.9 min and D75°C = 2.0 min). D-values of E. faecium were significantly higher than those of Salmonella at all three water activities, indicating that E. faecium is a suitable surrogate for Salmonella in thermal processing validation.

Multidrug-resistant high-risk Enterococcus faecium clones: can we really define them?

Enterococcus faecium is a significant opportunistic human pathogen with a broad host range, including humans, farm animals, pets and wildlife. Specialised subpopulations have globally evolved towards a powerful and convergent adaption to the healthcare environment by acquiring a cocktail of key antimicrobial resistance and virulence genes, enabling them to thrive in the disturbed microbiota of hospitalised patients. These populations can also be found in different community reservoirs, but the relevance of their dispersal in non-human hosts is greatly unknown and is here discussed. This review provides a brief historical overview of what we have been considering E. faecium high-risk clones worldwide alongside the advances in strain typing technologies that have revolutionised our understanding of the genetic evolution of this species over the last three decades.

Genomic rearrangements uncovered by genome-wide co-evolution analysis of a major nosocomial pathogen, Enterococcus faecium.

Enterococcus faecium is a gut commensal of the gastro-digestive tract, but also known as nosocomial pathogen among hospitalized patients. Population genetics based on whole-genome sequencing has revealed that E. faecium strains from hospitalized patients form a distinct clade, designated clade A1, and that plasmids are major contributors to the emergence of nosocomial E. faecium. Here we further explored the adaptive evolution of E. faecium using a genome-wide co-evolution study (GWES) to identify co-evolving single-nucleotide polymorphisms (SNPs). We identified three genomic regions harbouring large numbers of SNPs in tight linkage that are not proximal to each other based on the completely assembled chromosome of the clade A1 reference hospital isolate AUS0004. Close examination of these regions revealed that they are located at the borders of four different types of large-scale genomic rearrangements, insertion sites of two different genomic islands and an IS30-like transposon. In non-clade A1 isolates, these regions are adjacent to each other and they lack the insertions of the genomic islands and IS30-like transposon. Additionally, among the clade A1 isolates there is one group of pet isolates lacking the genomic rearrangement and insertion of the genomic islands, suggesting a distinct evolutionary trajectory. In silico analysis of the biological functions of the genes encoded in three regions revealed a common link to a stress response. This suggests that these rearrangements may reflect adaptation to the stringent conditions in the hospital environment, such as antibiotics and detergents, to which bacteria are exposed. In conclusion, to our knowledge, this is the first study using GWES to identify genomic rearrangements, suggesting that there is considerable untapped potential to unravel hidden evolutionary signals from population genomic data.

Clonal distribution of vancomycin-resistant Enterococcus faecium in Turkey and the new singleton ST733.

BACKGROUND: The aim of this study was to provide information about the spread and characteristics of the vancomycin-resistant Enterococcus faecium isolates (VREfm) in Turkey. METHODS: Seventy-one nonduplicate consecutive isolates of VREfm were obtained from various clinical specimens of inpatients treated at university or training hospitals in seven regions of Turkey. Further characteristics included antibiotic susceptibility testing, pulsed-field gel electrophoresis (PFGE) of SmaI-digested genomic DNA, and multilocus sequence typing (MLST) of selected isolates. The presence of vancomycin resistance and virulence genes (esp and hyl) was investigated by polymerase chain reaction (PCR). RESULTS: All VREfm isolates had MICs to vancomycin of ≥32 mg/L and contained the vanA gene. The presence of esp gene was identified in 64 and hyl in eight VREfm isolates. All VREfm showed the multiresistance phenotype, including ampicillin (99%), penicillin (99%), imipenem (99%), ciprofloxacin (87%), moxifloxacin (87%), erythromycin (97%), streptomycin (86%), gentamicin (82%), tetracycline (70%), and teicoplanin (99%). All were susceptible to tigecycline while quinupristin-dalfopristin (97%) and linezolid (93%) were the most active other agents. Analysis of the PFGE profiles showed that 53 (74.6%) VREfm isolates shared a similar electrophoretic profile, designed as type 1, and were closely related (>85%). The sequence type was identified by MLST in 44 VRE isolates with unrelated or closely related PFGE patterns. MLST revealed that nosocomial spread of VREfm resulted from dissemination of lineage C1 E faecium clones. Sequence types ST78, ST203, and ST117 were the most frequently isolated. This is the first report of ST733 around the world. CONCLUSIONS: Lineage C1 clones are disseminated among clinical VREfm isolates in seven different regions in Turkey. Regarding VREfm isolates, the worldwide epidemic strains are in circulation in Turkey.

Trend of clinical vancomycin-resistant enterococci isolated in a regional Italian hospital from 2001 to 2018.

A retrospective study of the epidemiology of vancomycin-resistant enterococci (VRE) in a regional hospital of central Italy in 2001-2018 demonstrated an increased VRE prevalence since 2016. A total of 113 VRE isolates, 89 E. faecium (VREfm) and 24 E. faecalis (VREfs), were collected in the study period. All strains showed high-level resistance to vancomycin; 107 also showed teicoplanin resistance. Altogether, 84 VREfm and 20 VREfs carried vanA, whereas 5 VREfm and 1 VREfs carried vanB. MLST analysis documented that 89 VREfm isolates mainly belonged to ST78, ST80, and ST117. Most strains were isolated from 2001 to 2007, ST78 being the predominant clone. VREfm re-emerged in 2016 with a prevalence of the ST80 lineage. Most VREfs were isolated from 2001 to 2006; although they belonged to 7 different STs, there was a prevalence of ST88 and ST6. Notably, ST88 was sporadically recovered throughout the study period. The increasing rate of VREfm isolation from 2016 to 2018 may be related to the influx of new successful clones and to the renewed and widespread use of vancomycin. Improved infection control measures in hospital wards should be adopted to limit the spread of new epidemic VRE strains.

Comprehensive integrated NGS-based surveillance and contact-network modeling unravels transmission dynamics of vancomycin-resistant enterococci in a high-risk population within a tertiary care hospital.

Vancomycin-resistant E. faecium (VRE) are an important cause of nosocomial infections, which are rapidly transmitted in hospitals. To identify possible transmission routes, we applied combined genomics and contact-network modeling to retrospectively evaluate routine VRE screening data generated by the infection control program of a hemato-oncology unit. Over 1 year, a total of 111 VRE isolates from 111 patients were collected by anal swabs in a tertiary care hospital in Southern Germany. All isolated VRE were whole-genome sequenced, followed by different in-depth bioinformatics analyses including genotyping and determination of phylogenetic relations, aiming to evaluate a standardized workflow. Patient movement data were used to overlay sequencing data to infer transmission events and strain dynamics over time. A predominant clone harboring vanB and exhibiting genotype ST117/CT469 (n = 67) was identified. Our comprehensive combined analyses suggested intra-hospital spread, especially of clone ST117/CT469, despite of extensive screening, single room placement, and contact isolation. A new interactive tool to visualize these complex data was designed. Furthermore, a patient-contact network-modeling approach was developed, which indicates both the periodic import of the clone into the hospital and its spread within the hospital due to patient movements. The analyzed spread of VRE was most likely due to placement of patients in the same room prior to positivity of screening. We successfully demonstrated the added value for this combined strategy to extract well-founded knowledge from interdisciplinary data sources. The combination of patient-contact modeling and high-resolution typing unraveled the transmission dynamics within the hospital department and, additionally, a constant VRE influx over time.

The ongoing challenge of vancomycin-resistant Enterococcus faecium and Enterococcus faecalis in Europe: an epidemiological analysis of bloodstream infections.

Vancomycin-resistant enterococci infections are of great public health significance due to limited therapeutic options. We investigated epidemiological trends and risk factors of vancomycin resistance in enterococci isolates from patients with bloodstream infections in the EU/EEA from 2012 to 2018. Routine vancomycin susceptibility data of clinical E. faecium (n = 67,022) and E. faecalis (n = 103,112) blood isolates from the European Antimicrobial Resistance Surveillance Network were analysed using descriptive statistics and multivariable regression analyses. In Europe, proportions of vancomycin-resistant E. faecium (VREFm) increased from 8.1% (95%CI 6.7-9.7%) in 2012 to 19.0% (95%CI 16.8-21.5%) in 2018. Rising VREFm proportions were observed across all European regions, both genders and all age groups except children and adolescents (1-19 years). Adults (20-59 years) and elderly (≥60 years) had an increased likelihood of VREFm compared to children and adolescents (1-19 years) (OR: 1.99 [95%CI 1.42-2.79, p < 0.001] and OR: 1.56 [95%CI 1.09-2.23, p = 0.014], respectively). Inpatients hospital units, including internal medicine and ICUs, were associated with an increased likelihood of VREFm (OR: 2.29 (95%CI 1.58-3.32, p < 0.001) compared to the emergency department which reflects patients with community origin of E. faecium infections. The mean proportion of vancomycin-resistant E. faecalis in Europe was found to be low (1.1% [95%CI 0.9-1.4%]). Local and regional authorities should intensify efforts directed at diagnostic and antimicrobial stewardship for vancomycin and all last resort drugs for the management of VREFm, particularly for hospitalized elderly patients.

Linezolid-resistant Enterococcus faecium strains isolated from one hospital in Poland -commensals or hospital-adapted pathogens?

One of the most pressing problems of enterococci infections is occurring resistance to linezolid, which is an antibiotic used in the treatment of infections caused by vancomycin-resistant strains (VRE). The main objective of our research was to investigate the relationship of 19 linezolid-resistant E. faecium isolates from 18 patients hospitalized at Clinical Hospital in Gdansk (Poland). One of the LZDREF was isolated in 2003 (K2003), and another 18 were collected from 2013 to 2017. Genotyping with PCR MP method indicated 14 main unrelated genetic profiles and no association with K2003 strain. Two isolates with the same genotype and genetically closely related two sub-types (2 isolates for each sub-type) were hospital-derived colonizations of patients. The other unrelated genotypes were discussed in the context of colonization, nosocomial infections, and commensal origin, taking into account prior exposure to linezolid. We determined the presence of a point mutation G2576T in six loci of 23S rDNA. There was also a significant correlation (p<0.0015) between the presence of MIC>32 value and the presence of G2576T point mutation on the sixth rrn. We also detected 5 virulence genes for all isolates: gelE, cylA, asa1, hyl, esp. Correlation (p≤0.0001) was observed between the presence of gelE gene encoding gelatinase and two other genes: cylA and asa1 encoding cytolysin and collagen binding protein responsible for aggregation of bacterial cells, respectively. Significant correlation was also observed between asa1 and cfr genes encoding 23S rRNA rybonuclease responsible for resistance to PhLOPSA antibiotics (p = 0.0004). The multidimensional analysis has also shown the correlation between cfr gene and GI-tract (p = 0, 0491), which suggests horizontal gene transfer inside the gut microbiota and the risk of colonization with linezolid-resistant strains without previously being treated with the antibiotic. The patient could have been colonized with LZDRVREF strains which in the absence of competitive microbiota quickly settle in ecological niches favourable for them and pose a risk for the patient.

Emergence of vancomycin-resistant Enterococcus faecium ST1421 lacking the pstS gene in Korea.

Although multilocus sequence typing (MLST) has been used to study molecular epidemiology and to explore the population structure of Enterococcus faecium, vancomycin-resistant E. faecium (VREF) strains lacking the pstS gene that were non-typable using conventional MLST methods were reported recently. We found nationwide emergence of VREF isolates lacking pstS in Korea and hereby report the molecular characteristics of these isolates. Forty-six VREF isolates lacking the pstS gene were identified among 300 VREF rectal isolates collected from hospitalized patients between 2014 and 2015. MLST was performed and clonal relatedness was determined by pulsed-field gel electrophoresis (PFGE). Four VREF ST1421 isolates were whole-genome sequenced. Among the VREF rectal isolates lacking pstS, 98% were classified as ST1421, which has identical allelic profiles to ST17 for all housekeeping genes except pstS. PFGE pattern analyses revealed 32 pulsotypes. All isolates harbored Tn1546 components with various transposase and insertion sequences. The whole-genome sequencing of four VREF ST1421 isolates showed that the pstS gene region was deleted at various locations with considerable inversion. The pstS gene was also depleted in 12.1% of 33 VREF clinical isolates in 2006-2007 and in 11.8% of 59 clinical isolates in 2012-2013. VREF ST1421 strains lacking the pstS gene have emerged in Korea. The emergence and spread of pstS-deleted VREF strains pose a serious challenge for epidemiological investigation. Alternative molecular typing methods to MLST will be increasingly necessary.

Assessment of safety aspect and probiotic potential of autochthonous Enterococcus faecium strains isolated from spontaneous fermented sausage.

OBJECTIVE: The objectives of this research project were isolation, identification, and evaluation of the safety aspect and probiotics properties of 21 Enterococcus faecium strains isolated from sausages originated from southeastern Serbia. RESULTS: Analyzed E. faecium isolates showed tolerance to simulated gastrointestinal conditions. All the examined isolates grew well on media with 0.1% and 0.2% of phenol. None of the tested isolates were histamine-producers, while the synthesis of tyramine was observed for E. faecium sk8-1 and sk8-17. Full resistance to antibiotics was not observed for any examined isolate of E. faecium (penicillin, amoxicillin, and ofloxacin showed the effect on all tested isolates). An inhibition zone against examined pathogens was exhibited by all strains, with the largest inhibition zone against Pseudomonas spp., Proteus spp. and E. coli (12-30 mm/MIC values ranged from 0.5 to 12 mg mL-1). CONCLUSION: The results indicated that E. faecium isolates from spontaneously fermented sausage showed a potential for further investigation and possible application as probiotics.

High Phenotypic and Genotypic Diversity of Enterococcus faecium from Clinical and Commensal Isolates in Third Level Hospital.

Background: The use of antimicrobials and myeloablative chemotherapy regimens has promoted multiresistant microorganisms to emerge as nosocomial pathogens, such as vancomycin-resistant Enterococcus faecium (VREfm). We described a polyclonal outbreak of bloodstream infection caused by Efm in a hemato-oncological ward in Mexico. Our aim was to describe the clonal complex (CC) of the Efm strains isolated in the outbreak in comparison with commensal and environmental isolates. Methodology: Sixty Efm clinical, environmental, and commensal strains were included. We constructed a cladogram and a phylogenetic tree using Vitek and Multilocus sequence typing data, respectively. Results: We reported 20 new sequence types (ST), among which 17/43 clinical isolates belonged to CC17. The predominant ST in the clinical strains were ST757, ST1304, ST412, and ST770. Neither environmental nor commensal isolates belonged to CC17. The phylogeny of our collection shows that the majority of the clinical isolates were different from the environmental and commensal isolates, and only a small group of clinical isolates was closely related with environmental and commensal isolates. The cladogram revealed a similar segregation to that of the phylogeny. Conclusions: We found a high diversity among clinical, environmental, and commensal strains in a group of samples in a single hospital. Highest diversity was found between commensal and environmental isolates.

Macrolide-susceptible probiotic Enterococcus faecium ST296 exhibits faecal-environmental-oral microbial community cycling among beef cattle in feedlots.

Enterococci are included in the United States National Antimicrobial Resistance Monitoring System to track antibiotic resistance among commensal Gram-positive enteric bacteria, largely due to their high abundance in food animals and in retail meat. In the U.S. cattle industry, macrolides are used to prevent and control liver abscesses, which cause significant economic losses. Previous studies have suggested that feeding tylosin and the intensity of the pen environment, both expand and sustain respectively the prevalence of multidrug resistance among enterococci in feedlot cattle. This has led to research into alternative feed supplements and improved stewardship practices. In a randomized controlled trial, we measured the impact of a probiotic and an altered pen environment on antimicrobial resistance among faecal Enterococcus spp. in cattle fed tylosin. Supplementing cattle with an Enterococcus faecium and Saccharomyces cerevisiae-based probiotic yielded the isolation of E. faecium of the probiotic sequence type (ST296) from faecal and environmental samples in treatment groups, as well as from cattle and the manure pack in nearby pens. Of importance, the probiotic strain also was found in a desiccated and milled manure pack sample taken 120 days after the initial trial ended. Phylogenetic and SNP analyses revealed clonal survival and spread compatible with faecal-environmental-oral recycling of the probiotic strain within and among cattle and pens. The increase in prevalence of the ST296 strain occurred concomitant with a decrease in ST240, the dominant sequence type associated with ermB and tet(M) resistance genes in this trial. SIGNIFICANCE AND IMPACT OF THE STUDY: We demonstrate that a macrolide-susceptible probiotic Enterococcus faecium ST296 strain fed to beef cattle becomes fully embedded in the microbial community cycling of bacteria via faecal-environmental-oral transmission within and among feedlot pens. An initial investment in feeding the probiotic is thereby leveraged into expanding numbers of susceptible bacteria in cattle and their environment, even among those cattle fed tylosin.

Amplified fragment length polymorphism and whole genome sequencing: a comparison of methods in the investigation of a nosocomial outbreak with vancomycin resistant enterococci.

Background: Recognition of nosocomial outbreaks with antimicrobial resistant (AMR) pathogens and appropriate infection prevention measures are essential to limit the consequences of AMR pathogens to patients in hospitals. Because unrelated, but genetically similar AMR pathogens may circulate simultaneously, rapid high-resolution molecular typing methods are needed for outbreak management. We compared amplified fragment length polymorphism (AFLP) and whole genome sequencing (WGS) during a nosocomial outbreak of vancomycin-resistant Enterococcus faecium (VRE) that spanned 5 months. Methods: Hierarchical clustering of AFLP profiles was performed using unweighted pair-grouping and similarity coefficients were calculated with Pearson correlation. For WGS-analysis, core single nucleotide polymorphisms (SNPs) were used to calculate the pairwise distance between isolates, construct a maximum likelihood phylogeny and establish a cut-off for relatedness of epidemiologically linked VRE isolates. SNP-variations in the vanB gene cluster were compared to increase the comparative resolution. Technical replicates of 2 isolates were sequenced to determine the number of core-SNPs derived from random sequencing errors. Results: Of the 721 patients screened for VRE carriage, AFLP assigned isolates of 22 patients to the outbreak cluster. According to WGS, all 22 isolates belonged to ST117 but only 21 grouped in a tight phylogenetic cluster and carried vanB resistance gene clusters. Sequencing of technical replicates showed that 4-5 core-SNPs were derived by random sequencing errors. The cut-off for relatedness of epidemiologically linked VRE isolates was established at ≤7 core-SNPs. The discrepant isolate was separated from the index isolate by 61 core-SNPs and the vanB gene cluster was absent. In AFLP analysis this discrepant isolate was indistinguishable from the other outbreak isolates, forming a cluster with 92% similarity (cut-off for identical isolates ≥90%). The inclusion of the discrepant isolate in the outbreak resulted in the screening of 250 patients and quarantining of an entire ward. Conclusion: AFLP was a rapid and affordable screening tool for characterising hospital VRE outbreaks. For in-depth understanding of the outbreak WGS was needed. Compared to AFLP, WGS provided higher resolution typing of VRE isolates with implications for outbreak management.

Effect of probiotic Enterococcus faecium SF68 on liver function in healthy dogs.

BACKGROUND: Probiotics are widely used in dogs but can be associated with alterations in some serum biochemistry test results. OBJECTIVE: To assess the effect of Enterococcus faecium SF68 administration for 14 days on serum alanine transferase (ALT) and alkaline phosphatase (ALP) activity and total cholesterol and triglyceride concentrations in healthy dogs. ANIMALS: Thirty-six healthy privately owned neutered dogs were randomly allocated, stratified by sex, to control or probiotic groups. Dogs were clinically healthy, with normal physical examination findings, blood, urine, and fecal analyses and ultrasonographic examinations. METHOD: In this blinded, controlled study E. faecium SF68 was administered to the probiotic group for 14 days. Blood samples were taken from all dogs at days 0, 14, and 28. Serum ALT and ALP activity and total cholesterol and triglyceride concentrations were determined on these 3 days. RESULTS: The probiotic induced no significant changes in mean ALT and ALP activity. Mean cholesterol concentration did not change during probiotic administration but a significant decrease was seen on day 28 (P < .01). Mean triglyceride concentration increased progressively, becoming significant at day 28 (P < .05), with 1 dog developing hypertriglyceridemia. CONCLUSIONS AND CLINICAL IMPORTANCE: E. faecium SF68 would not create confusion when monitoring dogs with hepatobiliary disease because ALT and ALP activity did not change significantly. A significant decrease in cholesterol and significant increase in triglyceride concentrations were seen at day 28 but were not clinically relevant, with 1 dog showing hypertriglyceridemia. A longer trial is warranted to assess if the probiotic effects could be clinically relevant and to assess its potential use in hypertriglyceridemic dogs.

Hospital outbreak caused by linezolid resistant Enterococcus faecium in Upper Austria.

Background: Enterococcus faecium is part of the human gastrointestinal flora but may act as opportunistic pathogen. Environmental persistence, high colonization capability and diverse intrinsic and acquired resistance mechanisms make it especially successful in nosocomial high-risk settings. In March 2014, an outbreak of Linezolid resistant Enterococcus faecium (LREfm) was observed at the hematooncology department of a tertiary care center in Upper Austria. Methods: We report on the outbreak investigation together with the whole genome sequencing (WGS)-based typing results including also non-outbreak LREfm and susceptible isolates. Results: The 54 investigated isolates could be divided in six clusters based on cgMLST. Cluster one comprised LREfm isolates of genotype ST117 and CT24, which was identified as the causative clone of the outbreak. In addition, the detection of four other clusters comprising isolates originating from hematooncology patients but also at other hospitals, pointed to LREfm transmission between local healthcare facilities. LREfm patients (n = 36) were typically at risk for acquisition of nosocomial pathogens because of immunosuppression, frequent hospitalization and antibiotic therapies. Seven of these 36 patients developed LREfm infection but were successfully treated. After termination of the initial outbreak, sporadic cases occurred despite a bundle of applied outbreak control interventions. Conclusions: WGS proved to be an effective tool to differentiate several LREfm clusters in an outbreak. Active screening for LREfm is important in a high-risk setting such as hematooncology, where multiple introductions are possible and occur despite intensified infection control measures.

Australian Group on Antimicrobial Resistance (AGAR) Australian Enterococcal Sepsis Outcome Programme (AESOP) Annual Report 2017.

From 1 January to 31 December 2017, 36 institutions around Australia participated in the Australian Enterococcal Sepsis Outcome Programme (AESOP). The aim of AESOP 2017 was to determine the proportion of enterococcal bacteraemia isolates in Australia that were antimicrobial resistant, and to characterise the molecular epidemiology of the E. faecium isolates. Of the 1,137 unique episodes of bacteraemia investigated, 95.2% were caused by either E. faecalis (52.9%) or E. faecium (42.3%). Ampicillin resistance was not detected in E. faecalis but in 89.6% of E. faecium. Vancomycin non-susceptibility was reported in 0.3% and 47.0% of E. faecalis and E. faecium respectively. Overall 50.9% of E. faecium harboured vanA or vanB genes. For the vanA/B positive E. faecium isolates, 49.6% harboured vanB genes and 49.2% vanA genes; 1.2% harboured vanA and vanB genes. The percentage of E. faecium bacteraemia isolates resistant to vancomycin in Australia is significantly higher than that seen in most European countries. E. faecium consisted of 76 multilocus sequence types (STs) of which 77% of isolates were classified into nine major STs containing ten or more isolates. All major STs belong to clonal cluster (CC) 17, a major hospital-adapted polyclonal E. faecium cluster. Seven of the nine predominant STs (ST80, ST1421, ST17, ST296, ST555, ST203 and ST18) were found across most regions of Australia. The most predominant clone was ST17 which was identified in all regions except the Australian Capital Territory, the Northern Territory and Tasmania. Overall 60.7% of isolates belonging to the nine predominant STs harboured vanA or vanB genes. The AESOP 2017 has shown enterococcal bacteraemias in Australia are frequently caused by polyclonal ampicillin-resistant high-level gentamicin resistant vanA or vanB E. faecium which have limited treatment options.