Vancomycin-resistant Enterococcus Faecium Research Articles

Bad Bugs Move Alike: Regional Transmission of Antibiotic-Resistant Organisms

Background: Upon admission, 56.8% of patients entering 6 Michigan nursing facilities from regional acute-care hospitals (ACHs) were colonized with 1 or more antibiotic-resistant organisms (AROs) (Mody 2018, CID). This observation raises 2 questions critical to regional infection control strategies: (1) Is the high ARO burden entering nursing facilities driven by dominant epidemic lineages or diverse circulating strains? and (2) What are the relative roles of patient characteristics (eg, high-risk patients) and exposure to specific ACHs (eg, high-risk facilities) in determining whether patients are colonized with AROs upon nursing facility admission? Here, we integrated whole-genome sequencing, patient transfer, and clinical data to answer these questions for the 4 most prevalent ARO species in the region: methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus faecalis (VREfc), Enterococcus faecium (VREfm), and ciprofloxacin-resistant Escherichia coli (CipREc). Methods: We studied colonizing isolates collected via active surveillance of 584 patients in 6 Michigan nursing facilities between 2013 and 2016. The whole genome of the first isolate of each ARO species collected from each patient was sequenced and analyzed to identify sequence types (STs) and to infer the transmission network by species. We determined the connectedness between nursing facilities based on the number of patients received from the same ACHs and assigned each ARO to the most recent ACH using curated transfer information. The associations between patient characteristics and recent ACH exposures with colonization by ARO were examined using multivariable models. Results: Most of the sequenced ARO isolates belonged to major healthcare-associated lineages: MRSA (ST5, N = 78 of 117); VREfc (ST6, N = 68 of 75); CipREc (ST131, N = 50 of 64); and closely related VREfm isolates (N = 129). Phylogenetically closely related isolates were found across study facilities, indicating that endemic ARO lineages have permeated local healthcare networks (Fig. 1). Patient characteristics played a dominant role in determining patient risk of ARO colonization on admission to a nursing facility. Only in the case of VREfm was a hospital significantly associated with colonization after adjustment for covariates (Table 1). Conclusions: ARO lineages were widely disseminated and colonization of specific ARO lineages at nursing facility entry could not be attributed to recent exposure to a specific ACH. Thus, for the ARO lineages studied here, a broader transmission system crosses ACHs, nursing facilities and probably the community. Therefore, the best indicators of ARO colonization were patient clinical characteristics, particularly poor functional status and antibiotic exposure. These findings suggest that intervention efforts targeting patients with characteristics associated with ARO colonization may help limit further spread among regional facilities.Funding: NoneDisclosures: None

ONE FOR THE MONEY, TWO FOR THE ECMO: A SUCCESSFUL PHARMACOTHERAPEUTIC STRATEGY FOR VANCOMYCIN-RESISTANT ENTEROCOCCUS FAECIUM AND CANDIDA GLABRATA FUNGEMIA IN VENO-VENOUS EXTRACORPOREAL MEMBRANE OXYGENATION AND CONTINUOUS RENAL REPLACEMENT

SESSION TITLE: Fellows Critical Care Posters SESSION TYPE: Fellow Case Report Posters PRESENTED ON: October 18-21, 2020 INTRODUCTION: Extracorporeal membrane oxygenation (ECMO) and continuous renal replacement therapy (CRRT) are modalities with a relative paucity in understanding of the disposition of antimicrobials, and even less for when used simultaneously. Preliminary investigations have demonstrated that ECMO leads to pharmacokinetic changes, which include alterations in volume of distribution, drug clearance, and sequestration by the circuit.1 CASE PRESENTATION: Our patient was a 66-year-old-male with a history of alcohol use disorder, transferred to our hospital for acute respiratory distress syndrome due to influenza A pneumonia with superimposed bacterial pneumonia, cannulated for venous-venous ECMO. Encephalopathy, atrial fibrillation, pulmonary embolisms, distributive shock, and anuric acute kidney injury requiring CRRT complicated his course. Following a prolonged course, blood cultures were obtained and Candida glabrata and vancomycin resistant Enterococcus faecium (VRE) were isolated. We initiated high dose micafungin 150 mg IV q 24 h, high dose, high frequency daptomycin 10 mg/kg IV q 24 h. An echocardiogram, Ophthalmology consult, and repeat blood cultures were also requested to document clearance of fungemia and bacteremia. A trans-esophageal echocardiogram was performed, without evidence of masses or vegetations. Fundal exam did not show any signs of fungal endophthalmitis. Repeat cultures were without growth, all existing IV lines were exchanged except for ECMO cannulas. His leukocytosis began to downtrend, and he became afebrile after start of antimicrobial therapy. Given that his most likely source of infection was line associated, a two-week antimicrobial course for uncomplicated fungemia and bacteremia was recommended. DISCUSSION: We hope to add to the limited data on daptomycin and micafungin dosing strategies for patients on ECMO and CRRT. A thorough understanding of the altered pharmacokinetics is essential to apply in these complex patients. Daptomycin has a low volume of distribution, and it is highly protein bound, making it less likely to be affected by ECMO. It is known that for patients on CRRT, daptomycin dosing q 24 h up to 12 mg/kg provides comparable drug exposure to patients with a CrCl = 30 mL/min,2 therefore, a dose of 10 mg/kg q 24 h was deemed appropriate to treat our patient. Creatinine kinase was monitored weekly without any changes from baseline. Micafungin is highly protein bound, minimally renally cleared and does not require CRRT dose adjustment. CONCLUSIONS: Prior studies in ECMO patients have shown decreased plasma concentrations due to drug degradation and increased clearance,3 which warrant a dose increase. As such, micafungin 150 mg q 24 h was used in our patient. This report highlights the need for more research on optimal dosing strategies of antimicrobials in this patient population. Reference #1: Cheng V, Abdul-Aziz MH, Roberts JA, Shekar K. Optimising drug dosing in patients receiving extracorporeal membrane oxygenation. J Thorac Dis. 2018;10(Suppl 5):S629-S641. Reference #2: Xu X, Khadzhynov D, Peters H, et al. Population pharmacokinetics of daptomycin in adult patients undergoing continuous renal replacement therapy. Br J Clin Pharmacol. 2017;83(3):498-509. Reference #3: Watt KM, Cohen-Wolkowiez M, Williams DC, et al. Antifungal extraction by the extracorporeal membrane oxygenation circuit. J Extra Corpor Technol. 2017;49(3):150-159. DISCLOSURES: No relevant relationships by M. Gabriela Cabanilla, source=Web Response No relevant relationships by Nicholas Villalobos, source=Web Response

Evaluation of Sampling Methods for Detection of Pathogens from Steel Surfaces; Contact Plates, Foam Swabs, and Flocked Swabs

Background: Contaminated healthcare surfaces can serve as reservoirs for the transmission of pathogens. Sensitive sampling methods are needed to investigate sources of pathogens for implementing effective disinfection strategies and thereby preventing environmental transmission. Conventional approaches employ swabs to sample environmental surfaces. Contact plates represent an alternative approach for sampling healthcare surfaces that does not require lab processing, though little is known about their performance. A contact plate is an agar plate that is overfilled with selective or nonselective media. It can be gently applied to the surface, then simply incubated at a temperature optimal for target organism (s), thus saving time and resources. Methods: In this study, contact plates containing trypticase soy agar with 5% sheep blood (TSAII), foam swabs, and flocked swabs were evaluated for their ability to recover 4 pathogens that persist on healthcare surfaces. Stainless-steel coupons (4 in2) were inoculated with the following pathogens (102 CFU): Acinetobacter baumannii (AB, strain type 12), carbapenemase-producing KPC+ Klebsiella pneumoniae (KP; ATCC BAA-1705); methicillin-resistant Staphylococcus aureus (MRSA; ATCC 43300); and vancomycin-resistant Enterococcus faecalis (VRE; Van A + 256). The plates were allowed to dry 1 hour. Sampling with CPs was performed in 2 ways; (1) a single contact plate was used to sample 1 stainless-steel surface and (2) a composite was collected by 3 sequential contact-plate samplings of the same stainless-steel surface. The contact plates were then incubated at 351C. Foam and flocked swabs were premoistened with phosphate-buffered saline + 0.02% polysorbate 80 (PBST) and were used to sample the stainless-steel coupons. Swabs were held for 1 hour and processed by sonication and vortexing in 5 mL of PBST, then the eluent was cultured and CFU counted. Mean percentage recoveries (%R) relative to the inoculum were calculated and compared. Results: When the %R for all 4 pathogens were pooled, the composite contact-plate sampling method yielded the highest, (P < .05) (66.0%; SD, 0.22), followed by the single contact plate method (39.7%; SD, 0.12), foam swab (32.9%; SD, 0.18), and flocked swab (20.3%; SD, 0.20). The composite contact plate method yielded the highest %R for VRE (102.1 %; SD, 0.17), and the lowest %R was observed when using flocked swabs to recover KP (6.3%; SD, 0.05). Conclusions: The contact-plate composite method may provide investigators with minimal environmental microbiology capacity an alternative method for environmental sampling and detection of organisms from surface areas (4 in 2) with low bioburden.Funding: NoneDisclosures: None

The Microbiome Analysis of Hospital Mobile Phones: Hidden Contaminants Revealed

Background: The undisputed versatility of smart devices makes them integral to modern-day society, especially within our National Health System. Despite the benefits, there are increasing concerns regarding their contamination and the associated infection risk. Bacteria under antimicrobial selective pressure can rapidly acquire resistant mechanisms leading to the assumption; mobile phones used within clinical environments may harbor bacteria associated with a higher infection mortality rate. Using next-generation sequencing technology, we characterized the true extent of bacterial contamination on mobile devices of hospital staff to determine the level of antimicrobial resistance within Staphylococcus aureus and Enterococcus faecalis. Smart phones of 250 hospital staff and 191 control group participants were swabbed over a 6-month period. The microbiome of devices were characterized using Illinima MiSeq metabarcoding pipeline. Cultured isolates were quantified and underwent Kirby-Bauer disc diffusion. Primer version 6 and SPSS version 23 software were used to analyze the statistics. Nearly all mobile devices were contaminated with potential pathogens regardless of environment. Metabarcoding revealed far greater bacterial diversity and abundance of gram-negative bacterial contamination than did culture-based methods. In total, 198 bacterial genera of were discovered across both groups, of which 34 were unique to the hospital. Bacillus was significantly higher within the hospital group (P = .036). Differences were also detected within the hospital (high-risk vs low-risk areas, P = .048). Methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcusfaecalis were only isolated from hospital mobile phones (P < .001 and P = .038, respectively). Our results indicate that traditional culture-dependent swabbing methods do not provide an accurate picture of contamination. Metabarcoding reinforces the need for mobile phone infection control practices to mitigate the risks associated with the increase use of smart device technology in clinical environments. These devices are currently exposing immunocompromised patients to unknown levels of pathogenic and multidrug-resistant bacteria. Departmental differences may suggest that the mobile phone is not just an extension to its owners but to their environment and that routine decontamination should be required to prevent the undermining of hand hygiene and the transmission of pathogenic bacteria.Funding: NoneDisclosures: None

Clinical and molecular epidemiology of vancomycin-resistant Enterococcus faecium bacteremia from an Indian tertiary hospital.

Wedetermined the clinical and molecular epidemiology of emerging nosocomial vancomycin-resistant Enterococcus faecium (VREfm)-causing serious bloodstream infections (BSIs) and the correlations between antibiotic resistance and virulence determinantsamong isolates. Allisolates were confirmed by molecular methods (16SrRNA and E. faecium ddl genes) and tested for disk diffusion. PCRwas used to detect aac(6')-aph(2″), vanA and vanB resistance genes, and asa1, cylA, ace, esp, gelE and hyl virulence genes. VREfm and high-level gentamicin-resistant (HLGR) representative isolates were selected to characterize by pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). Of 173 isolates, 73(42.2%), 146(84.4%), and 0(0.0%) were vanA-containing VREfm, aac(6')-aph(2″)-positive HLGR, and vanB-positive. Independent predictors of VREfm infection were hematological malignancies (P = 0.001) and previous hospitalizations (P = 0.007). Observed mortality rate was 34.7%. Independent predictors of BSI-related mortality were endotracheal intubations (P < 0.001), gastrointestinal diseases (P = 0.002), and pulmonary disease (P < 0.001). All VREfm were resistant to vancomycin, teicoplanin, ciprofloxacin, and erythromycin. The esp, hyl, ace, asa1, cylA, and gelE genes were detected at 55.9, 22.5, 2.9, 2.3, 1.7, and 1.2%, respectively. The esp gene was significantly associated with VREfm compared toVSEfm (P = 0.001). PFGE analysis revealed 23 clones, with 7 major clones. The MLST analysis revealed the following five sequence types: ST80, ST17, ST117, ST132, and ST280, all belonging to CC17. The emergence and expansion of VREfm CC17 with limited antibiotic options in our hospital present a serious public health menace and represent challenges to infection control.

The second N-terminal aromatic residue of the fungal defensin, blapersin, of Blastomyces percursus is essential for its antibacterial activity

Studies have shown that the second N-terminal residue of fungal defensins is closely involved in the binding of defensins to lipid II, a bacterial cell wall precursor, and plays an important role in antibacterial activity. We found that the N-terminal residue is always aromatic in nature. In this study, 29 fungal defensin-like peptides were found via the genomic search strategy. Based on the type of aromatic residue at the second N-terminal site, we mainly divided these peptides into Phe, Trp, and Tyr types. We selected and characterized a defensin, blapersin, derived from Blastomyces percursus as a molecular model to investigate the functional significance of the N-terminal site. The native blapersin killed a wide spectrum of gram-positive bacteria at low molecular concentrations. Its aromatic mutants, W2F and W2Y, displayed enhanced antimicrobial activity, especially against the vancomycin-resistant Enterococcus faecium. The aromatic side chains containing Phe2 and Tyr2 seem to be more favorable for the antibacterial activity of blapersin those containing Trp2. However, the nonaromatic mutant W2A had almost no antibacterial activity. This indicates that the second N-terminal aromatic residue is essential for the antimicrobial action of blapersin. All these defensins have high stability and low toxicity. This is the first report on the enhancement of antibacterial activity by calibration of the N-terminal aromatic residue.

Long-Lasting Decrease of the Acquisition of Enterococcus faecium and Gram-Negative Bacteria Producing Extended Spectrum Beta-Lactamase (ESBL) by Transient Application of Probiotics.

Previously it was shown that application of probiotics stopped the acquisition of vancomycin-resistant Enterococcus faecium (VRE) by patients in an early rehabilitation ward. Once the application of probiotics ended, we examined whether acquisition of VRE reoccurred. Furthermore, we examined whether probiotics altered prevalence of vancomycin-susceptible E. faecium (VSE) and Gram-negative bacteria, which produce extended spectrum beta-lactamase (ESBL). Although probiotic application ceased in April 2018, VRE-colonized patients rarely presented on that ward until 2019. Probiotic treatment also resulted in a decreased number of patients with VSE and ESBL. While decreased incidence of VRE occurred immediately, decreased VSE and ESBL numbers occurred months later. A probiotic-mediated decrease of VSE and ESBL incidence cannot be explained when assuming bacterial transmission exclusively as a linear cause and effect event. The decrease is better understood by considering bacterial transmissions to be stochastic events, which depend on various driving forces similar to an electric current. We hypothesize that VRE, VSE and ESBL uptake by patients and by staff members mutually reinforced each other, leading staff members to form a bacterial reservoir, similar to a condenser that stores electrical energy. Probiotic treatment then inhibited regeneration of that store, resulting in a breakdown of the driving force.

Rhodethrin and Rubrivivaxin as potential source of anti-biofilm agents against vancomycin resistant Enterococcus faecalis (ATCC 19443)

Enterococcus faecalis is frequently present in the hospital environment and readily forms a biofilm that protects from antibiotics and resistance against environmental stress conditions, thereby increasing nosocomial chronic infections. This study aims to assess antimicrobial and antibiofilm activities of two novel terpenoid derivatives Rhodethrin (Rdn) and Rubrivivaxin (Rbn) against vancomycin resistant Enterococcus faecalis strain (ATCC19443). Both terpenoids effectively prevent biofilm formation with >75% attenuation in cell biomass and significantly decrease the production of exopolysaccharides (EPSs) (p = 0.005) and besides their expansion on different surface media. The findings provide new evidence that such terpenoid derivatives could be developed as novel antibacterial drugs.

Microbe-Metabolite Associations Linked to the Rebounding Murine Gut Microbiome Postcolonization with Vancomycin-Resistant Enterococcus faecium.

Vancomycin-resistant Enterococcus faecium (VREfm) is an emerging antibiotic-resistant pathogen. Strain-level investigations are beginning to reveal the molecular mechanisms used by VREfm to colonize regions of the human bowel. However, the role of commensal bacteria during VREfm colonization, in particular following antibiotic treatment, remains largely unknown. We employed amplicon 16S rRNA gene sequencing and metabolomics in a murine model system to try and investigate functional roles of the gut microbiome during VREfm colonization. First-order taxonomic shifts between Bacteroidetes and Tenericutes within the gut microbial community composition were detected both in response to pretreatment using ceftriaxone and to subsequent VREfm challenge. Using neural networking approaches to find cooccurrence profiles of bacteria and metabolites, we detected key metabolome features associated with butyric acid during and after VREfm colonization. These metabolite features were associated with Bacteroides, indicative of a transition toward a preantibiotic naive microbiome. This study shows the impacts of antibiotics on the gut ecosystem and the progression of the microbiome in response to colonization with VREfm. Our results offer insights toward identifying potential nonantibiotic alternatives to eliminate VREfm through metabolic reengineering to preferentially select for Bacteroides IMPORTANCE This study demonstrates the importance and power of linking bacterial composition profiling with metabolomics to find the interactions between commensal gut bacteria and a specific pathogen. Knowledge from this research will inform gut microbiome engineering strategies, with the aim of translating observations from animal models to human-relevant therapeutic applications.

Unexpected Cell Wall Alteration-Mediated Bactericidal Activity of the Antifungal Caspofungin against Vancomycin-Resistant Enterococcus faecium.

Enterococcus faecium has become a major opportunistic pathogen with the emergence of vancomycin-resistant enterococci (VRE). As part of the gut microbiota, they have to cope with numerous stresses, including effects of antibiotics and other xenobiotics, especially in patients hospitalized in intensive care units (ICUs) who receive many medications. The aim of this study was to investigate the impact of the most frequently prescribed xenobiotics for ICU patients on fitness, pathogenicity, and antimicrobial resistance of the vanB-positive E. faecium Aus0004 reference strain. Several phenotypic analyses were carried out, and we observed that caspofungin, an antifungal agent belonging to the family of echinocandins, had an important effect on E. faecium growth in vitro We confirmed this effect by electron microscopy and peptidoglycan analysis and showed that, even at a subinhibitory concentration (1/4× MIC, 8 mg/liter), caspofungin had an impact on cell wall organization, especially with respect to the abundance of some muropeptide precursors. By transcriptome sequencing (RNA-seq), it was also shown that around 20% of the transcriptome was altered in the presence of caspofungin, with 321 and 259 significantly upregulated and downregulated genes, respectively. Since the fungal target of caspofungin (i.e., β-1,3-glucan synthase) was absent in bacteria, the mechanistic pathway of caspofungin activity was investigated. The repression of genes involved in the metabolism of pyruvate seemed to have a drastic impact on bacterial cell viability, while a decrease of glycerol metabolism could explain the conformational modifications of peptidoglycan. This is the first report of caspofungin antibacterial activity against E. faecium, highlighting the potential impact of nonantibiotic xenobiotics against bacterial pathogens.

Antimicrobial O-Alkyl Derivatives of Naringenin and Their Oximes Against Multidrug-Resistant Bacteria.

New antimicrobial agents are needed to address infections caused by multidrug-resistant bacteria. Here, we are reporting novel O-alkyl derivatives of naringenin and their oximes, including novel compounds with a naringenin core and O-hexyl chains, showing activity against clinical strains of clarithromycin-resistant Helicobacter pylori, vancomycin-resistant Enterococcus faecalis, methicillin-resistant Staphylococcus aureus, and beta-lactam-resistant Acinetobacter baumannii and Klebsiella pneumoniae. The minimum inhibitory concentrations (MICs), which provide a quantitative measure of antimicrobial activity, were in the low microgram range for the selected compounds. Checkerboard assays for the most active compounds in combination with antibiotics revealed interactions that varied from synergistic to neutral.

In Vitro Activity and Potency of the Novel Oxazolidinone Contezolid (MRX-I) Tested against Gram-Positive Clinical Isolates from the United States and Europe.

Contezolid, a new oxazolidinone antibacterial agent currently in development for the treatment of skin and skin structure infections, was susceptibility tested against Gram-positive clinical isolates (n = 1,211). Contezolid demonstrated potent activity against Staphylococcus aureus (MIC50/90, 0.5/1 mg/liter), coagulase-negative Staphylococcus (MIC50/90, 0.25/0.5 mg/liter), Enterococcus spp. (MIC50/90, 0.5/1 mg/liter), and streptococci (MIC50/90, 1/1 mg/liter). Moreover, methicillin-resistant S. aureus and vancomycin-resistant Enterococcus faecium isolates were all inhibited by contezolid at ≤1 mg/liter. These results support the clinical development of contezolid.

Biovolume and spatial distribution of foodborne Gram-negative and Gram-positive pathogenic bacteria in mono- and dual-species biofilms

The objective of this study was to characterize the biofilms formed by Salmonella enterica serotype Agona, Listeria monocytogenes, methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE) after 12, 48, 72, 120 and 240 h of incubation at 10 °C. Biofilms containing a single species, together with dual-species biofilms in which S. enterica and a Gram-positive bacterium existed in combination, were formed on polystyrene and evaluated by using confocal laser scanning microscopy (CLSM). All strains were able to form biofilm. The greatest biovolume in the observation field of 14,161 μm2 was observed for mono-species biofilms after 72 h, where biovolumes of 94,409.0 μm3 ± 2131.0 μm3 (S. enterica), 58,418.3 μm3 ± 5944.9 μm3 (L. monocytogenes), 68,020.8 μm3 ± 5812.3 μm3 (MRSA) and 59,280.0 μm3 ± 4032.9 μm3 (VRE) were obtained. In comparison with single-species biofilms, the biovolume of S. enterica was higher in the presence of MRSA or VRE after 48, 72 and 120 h. In dual-species biofilms, the bacteria showed a double-layer distribution pattern, with S. enterica in the top layer and Gram-positive bacteria in the bottom layer. This spatial disposition should be taken into account when effective strategies to eliminate biofilms are being developed.

A single-center study on the distribution and antibiotic resistance of pathogens causing bloodstream infection in adult patients with hematological disease during the period 2014-2018

目的探讨2014–2018年成人血液病患者发生血流感染的病原菌分布和耐药情况，为该类患者合理使用抗菌药物提供数据参考。方法回顾性分析2014年1月至2018年12月中国医学科学院血液病医院收治的血液病合并血流感染患者的临床资料、病原菌种类分布和药物敏感性数据。结果5年内共1 478例血液病患者发生1 935次血流感染，1 700例次（87.9%）发生在中性粒细胞缺乏期，14 d与30 d全因死亡率分别为5.5%和8.2%。52.7%的血流感染存在伴随症状，主要为呼吸道症状、肛周黏膜损伤及消化道症状，比例分别为12.4%、12.3%和9.1%。共分离出2 025株病原菌，革兰阴性菌1 551株（76.6%），主要为大肠埃希菌、肺炎克雷伯菌和铜绿假单胞菌；革兰阳性球菌423株（20.9%），主要为葡萄球菌属与草绿色链球菌；真菌51株（2.5%），以热带念珠菌为主。肠杆菌科细菌对哌拉西林/他唑巴坦、碳青霉烯类、阿米卡星耐药率<10%。其中肺炎克雷伯菌对头孢吡肟、哌拉西林/他唑巴坦、美罗培南耐药率呈逐年上升趋势。铜绿假单胞菌对哌拉西林/他唑巴坦、喹诺酮类、氨基糖苷类抗菌药物耐药率<5%，低于碳青霉烯类。共检出耐甲氧西林金黄色葡萄球菌11株，耐万古霉素肠球菌1株。结论成人血液病患者血流感染病原菌分布广泛，不同菌种耐药率差异较大，且部分菌种耐药呈上升趋势，应根据不同人群血流感染病原菌分布及耐药情况合理使用抗菌药物。

Vancomycin-resistant Enterococcus faecium colonizing patients on hospital admission in Germany: prevalence and molecular epidemiology.

To analyse the rectal carriage rate and the molecular epidemiology of vancomycin-resistant Enterococcus faecium (VREfm) recovered from patients upon hospital admission. Adult patients were screened at six German university hospitals from five different federal states upon hospital admission for rectal colonization with VREfm between 2014 and 2018. Molecular characterization of VREfm was performed by WGS followed by MLST and core-genome MLST analysis. Of 16350 patients recruited, 263 were colonized with VREfm, with increasing prevalence rates during the 5 year study period (from 0.8% to 2.6%). In total, 78.5% of the VREfm were vanB positive and 20.2% vanA positive, while 1.2% harboured both vanA and vanB. The predominant ST was ST117 (56.7%) followed by ST80 (15%), ST203 (10.9%), ST78 (5.7%) and ST17 (3.2%). ST117/vanB VREfm isolates formed a large cluster of 96 closely related isolates extending across all six study centres and four smaller clusters comprising 13, 5, 4 and 3 isolates each. In contrast, among the other STs inter-regional clonal relatedness was rarely observed. To our knowledge, this is the largest admission prevalence and molecular epidemiology study of VREfm. These data provide insight into the epidemiology of VREfm at six German university hospitals and demonstrate the remarkable inter-regional clonal expansion of the ST117/vanB VREfm clone.

Increase of vancomycin-resistant Enterococcus faecium strain type ST117 CT71 at Charit\xe9 - Universit\xe4tsmedizin Berlin, 2008 to 2018

BackgroundIn addition to an overall rise in vancomycin-resistant Enterococcus faecium (VREfm), an increase in certain strain types marked by sequence type (ST) and cluster type (CT) has been reported in Germany over the past few years. Outbreak analyses at Charité - Universitätsmedizin Berlin revealed the frequent occurrence of VREfm ST117 CT71 isolates in 2017 and 2018. To investigate whether ST117 CT71 have emerged in recent years or whether these strains have been circulating for a longer time, we retrospectively analyzed non-outbreak strains that occurred between 2008 and 2018 to identify frequent STs and CTs.MethodsIn total, 120 VREfm isolates obtained from clinical and surveillance cultures from the years 2008, 2013, 2015, and 2018 were analyzed. Thirty isolates per year comprising the first 7–8 non-outbreak isolates of each quarter of the respective year were sequenced using whole genome sequencing. MLST and cgMLST were determined as well as resistance genes and virulence factors. Risk factors for VREfm ST117 were analyzed in a multivariable analysis with patient characteristics as possible confounders.ResultsThe percentage of VREfm of type ST117 increased from 17% in 2008 to 57% in 2018 (p = 0.012). In 2008, vanA genotype accounted for 80% of all ST117 isolates compared to 6% in 2018. VanB CT71 first appeared in 2018 and predominated over all other ST117 at 43% (p < 0.0001).The set of resistance genes (msrC, efmA, erm(B), dfrG, aac(6′)-Ii, gyrA, parC and pbp5) and virulence factors (acm, esp, hylEfm, ecbA and sgrA) in CT71 was also found in other ST117 non-CT71 strains, mainly in CT36. The study population did not differ among the different calendar years analyzed in terms of age, gender, length of stay, or ward type (each p > 0.2).ConclusionThis study revealed an increase in ST117 strains from 2008 to 2018, accompanied by a shift toward CT71 strains with the vanB genotype in 2018. We did not detect resistance or virulence traits in CT71 that could confer survival advantage compared to other CTs among ST117 strains. To date, it is not clear why ST117 and in particular strain type ST117 CT71 predominates over other strains.

Protective Functions of Group 3 Late Embryogenesis Abundant (G3LEA) Proteins in Enterococcus faecium During Vancomycin Treatment.

Late embryogenesis abundant (LEA) proteins protect organisms from various environmental stresses; however, the underlying mechanism of LEA mediated therapeutic evasion is still unclear in both eukaryotes and prokaryotes. In this study, group 3 LEA protein (G3LEA) of vancomycin-resistant Enterococcus faecium under sublethal concentration of vancomycin stress was evaluated and shown to have two functions: the first is the reduction of reactive oxygen species (ROS) content, preventing apoptosis by suppressing apoptotic proteins Cas3 and MAOB, and the second is activating specific drug efflux pumps. Sublethal vancomycin model was established with using Propidium Iodide (PI) stain. Real-time PCR was conducted to evaluate the expression of G3lea. Flow cytometry and confocal microscope using Anti- G3LEA, anti- MAOB, and anti- Cas3 were performed to assess the expression of G3LEA. Under sublethal vancomycin stress, G3LEA is upregulated, suppressing the expression of apoptotic markers and increasing specific efflux markers. These results suggest that G3LEA protein suppresses antibiotic mediated apoptosis in prokaryotic cells and plays a key role in understanding and preventing antibiotic resistance.

Clinical Utility of Lefamulin: If Not Now, When?

Purpose of ReviewThe looming threat of antimicrobial resistance requires robust stewardship and new developments in infectious diseases pharmacotherapy. This review discusses the pertinent spectrum and clinical data of lefamulin (Xenleta®), with a focus on potential real-world use.Recent FindingsLefamulin is a novel pleuromutilin antibiotic that obtained Food and Drug Administration labeling for community-acquired bacterial pneumonia (CABP) in 2019. Lefamulin is available in both intravenous and oral formulations, and it inhibits bacterial protein synthesis inhibition through interactive binding to unique sites of the peptidyl transferase center of the 50s bacterial ribosome subunit. Resistance, including cross-resistance with other antibiotics, is infrequent. Lefamulin demonstrates activity against most Gram-positive pathogens and other organisms commonly associated with CABP, i.e., Streptococcus pneumoniae, Haemophilus influenzae, Mycoplasma pneumoniae, Legionella pneumophila, and Chlamydophila pneumoniae. Lefamulin may also be an option for serious public health threats like methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus faecium, and multi-drug-resistant organisms associated with sexually transmitted infections, e.g., Neisseria gonorrhoeae, Mycoplasma genitalium. Lefamulin lacks activity against Pseudomonas aeruginosa, Acinetobacter baumannii, Enterobacterales, most anaerobes, and E. faecalis. In Phase III trials, lefamulin monotherapy was non-inferior to moxifloxacin with or without linezolid for CABP.SummaryLefamulin is a well-tolerated agent with a unique mechanism, availability in both IV and PO formulations, and it has been rigorously studied for safety and efficacy for CABP.

Moving away from traditional antibiotic treatment: can macrocyclic lactones from marine macroalga-associated heterotroph be the alternatives?

Intertidal red algae Hypnea valentiae associated Bacillus amyloliquefaciens MTCC 12716 revealed potential inhibitory effects on the growth of drug-resistant pathogens. In the genome of B. amyloliquefaciens MTCC 12716, biosynthetic gene clusters encoding antibacterial metabolites were predicted, which might be expressed and contributed to the broad-spectrum anti-infective activity. Three homologue members of the 24-membered macrocyclic lactone family, named as bacvalactones 1-3 bearing 13-O-ethyl (1); 15-O-furanyl-13-O-isobutyl-7-O-propyl-propanoate (2); and 15-O-furanyl-13-O-isobutyl-7-O-propyl-propanoate-7,24-dimethyl (3) functionalities, were acquired through bioactivity-guided purification. The macrocyclic lactones displayed bactericidal activity against opportunistic pathogens causing nosocomial infections, for instance, methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus faecalis (VREfs), and multidrug-resistant strains of Pseudomonas aeruginosa and Klebsiella pneumonia with MIC ≤ 3.0μg/mL, whereas standard antibiotics ampicillin and chloramphenicol were active only at concentrations of ≥ 6.25mg/mL. The biosynthetic pathway of macrocyclic lactones that are generated by trans-AT polyketide synthases through stepwise extension of an acetyl starter unit by eleven sequential Claisen condensations with malonyl-CoA was established, and the structures were correlated with the gene organization of the mln operon, which encompasses nine genes mln A-I (approximately 47kb in size). The best binding poses for each compounds (1-3) with Staphylococcus aureus peptide deformylase (SaPDF) unveiled docking scores (≥ 9.70kcal/mol) greater than that of natural peptide deformylase inhibitors, macrolactin N and actinonin (9.14 and 6.96kcal/mol, respectively), which supported their potential in vitro bioactivities. Thus, the present work demonstrated the potential of macrocyclic lactone for biotechnological and pharmaceutical applications against emerging multidrug-resistant pathogens. Key Points •Three antibacterial bacvalactones were identified from the symbiotic bacterium. •The symbiotic bacterial genome was explored to identify the biosynthetic gene clusters. •Trans-AT pks-assisted mln biosynthetic pathway of the macrocyclic lactone was proposed. •In silicomolecular interactions of the bacvalactones with S. aureus PDF were analyzed.

Detection of vancomycin-resistant enterococci in samples from broiler flocks and houses in Turkey.

Vancomycin-resistant enterococcus (VRE) is a global threat to public health. Knowledge about the occurrence of vanA-carrying enterococci in broiler and environmental samples is important as antibiotic resistance can be transferred to human bacteria. The aim of this study was to investigate the presence of VRE in broiler cloacal and environmental (house) samples and to genotype the isolates. In this study, 350 swabs were collected from broiler farms. All samples were plated onto enterococcus selective agar containing 6 mg/L vancomycin and 64 mg/L ceftazidime. Minimum inhibitory concentration (MIC) values were determined for vancomycin and teicoplanin. Vancomycin-resistant Enterococcus faecium (VREfm) was isolated from 6 out of 300 (2%) broiler cloacal samples and 13 out of 50 (26%) house samples. All E. faecium isolates had vanA genes. All VREfm isolates (19 isolates) were confirmed to be 95% similar to each other. In conclusion, although 20 years have passed since the ban on avoparcin in Turkey, the present study shows that VREfm isolates are still present in broiler production and especially in broiler houses, and most importantly, a major VREfm clone was isolated from broiler cloacal and house samples.