Enterococcus Faecium Isolates Research Articles

Influence of β-lactam pharmacodynamics on the systems microbiology of gram-positive and gram-negative polymicrobial communities.

We sought to evaluate the pharmacodynamics of β-lactam antibacterials against polymicrobial communities of clinically relevant gram-positive and gram-negative pathogens. Two Enterococcus faecalis isolates, two Staphylococcus aureus isolates, and three Escherichia coli isolates with varying β-lactamase production were evaluated in static time-killing experiments. Each gram-positive isolate was exposed to a concentration array of ampicillin (E. faecalis) or cefazolin (S. aureus) alone and during co-culture with an E. coli isolate that was β-lactamase-deficient, produced TEM-1, or produced KPC-3/TEM-1B. The results of the time-killing experiments were summarized using an integrated pharmacokinetic/pharmacodynamics analysis as well as mathematical modelling to fully characterize the antibacterial pharmacodynamics. In the integrated analysis, the maximum killing of ampicillin (Emax) against both E. faecalis isolates was ≥ 4.11 during monoculture experiments or co-culture with β-lactamase-deficient E. coli, whereas the Emax was reduced to ≤ 1.54 during co-culture with β-lactamase-producing E. coli. In comparison to monoculture experiments, culturing S. aureus with KPC-producing E. coli resulted in reductions of the cefazolin Emax from 3.25 and 3.71 down to 2.02 and 2.98, respectively. Two mathematical models were created to describe the interactions between E. coli and either E. faecalis or S. aureus. When in co-culture with E. coli, S. aureus experienced a reduction in its cefazolin Kmax by 24.8% (23.1%RSE). Similarly, β-lactamase-producing E. coli preferentially protected the ampicillin-resistant E. faecalis subpopulation, reducing Kmax,r by 90.1% (14%RSE). β-lactamase-producing E. coli were capable of protecting S. aureus and E. faecalis from exposure to β-lactam antibacterials.

Emergence of Virulent Extensively Drug-Resistant Vancomycin-Resistant Enterococci Among Diarrheic Pet Animals: A Possible Public Health Threat on the Move.

Background: Vancomycin-resistant enterococci (VRE) have become an increasing public health concern in the past few decades, being associated with serious multidrug-resistant (MDR) infections. This study was conducted to investigate the role of diarrheic pet animals as potential reservoirs for virulent extensively drug-resistant (XDR) VRE and their threat on human health. Materials and Methods: Rectal swabs were collected from 153 diarrheic pet animals (80 dogs and 73 cats). The collected swabs were cultured on CHROMagarTMVRE for the isolation of vancomycin-resistant Enterococcus faecalis and Enterococcus faecium, and then suspected colonies were identified as enterococci after Gram staining, conventional biochemical tests, and molecular techniques. VRE were basically identified using the disk diffusion method; however, molecular identification of vanA and vanB genes was carried out among confirmed VRE isolates. Moreover, three virulence genes (cytolysin A, cylA; enterococcal surface protein, esp; and hyaluronidase, hyl) were investigated in VRE isolates. Thereafter, VRE strains that harbored virulence genes were tested for antimicrobial susceptibility. Results: Eighteen out of 153 animals (11.8%) were positive for VRE, which were obtained from 15% and 8.2% of the examined dogs and cats, respectively. None of the obtained isolates carried the vanA gene, whereas the vanB gene was detected in E. faecalis (4/10) with a prevalence rate (40%). Of the obtained VRE isolates, five possessed esp and/or cylA, while all strains were negative for the hyl gene. Furthermore, four virulent VRE isolates exhibited an XDR pattern, and one isolate was MDR. Conclusion: Diarrheic pet animals could represent a potential zoonotic reservoir for virulent XDR vancomycin-resistant E. faecalis, which may have serious public health implications.

Gram Positive Cocci Associated Urinary Tract Infections, their Prevalence and Antibiotic Susceptibility Patterns

Background: Urinary tract infections (UTIs) are the third most common type of infection in humans globally. Gram-positive bacteria are said to be responsible for ten percent of urinary tract (UTI) infections. The study's goal was to profile gram-positive cocci-associated UTIs and their antibiogram, as they were observed at LASUTH. Methods: This was a retrospective assessment of the Medical Microbiology Laboratory records of the LASUTH to review the in vitro antibiotic susceptibility patterns of gram-positive urinary bacterial isolates between April 2020 and March 2021. The bacteria were isolated and identified from routine urine samples using standard bacteriological methods and the API. In vitro antibiotic susceptibility test (AST) was routinely performed by the modified Kirby-Bauer disk diffusion test and susceptibility breakpoints were determined using the Clinical and Laboratory Standards Institute (CLSI) guidelines. Results: 2,253 urine samples were processed in the medical microbiology laboratory over the one year and 662 (29.4%) samples yielded Positive cultures. Of the 662 isolates, 494 (74.6%) were gram-negative bacteria. 164 (24.8%) were gram-positive cocci while 4 (0.6%) were gram positive rod. Among the gram-positive cocci’s isolated Enterococcus faecalis had the highest frequency 58 (35.4%). Aminoglycosides (Gentamycin and Amikacin) and Linezolid antibiotics were found to be the most effective drugs against gram-positive cocci bacteria except Enterococcus spp. For empirical treatment of Enterococcus spp in our facility Fosfomycin and Tigecycline are the best options, while for Streptococcus agalactiae associated UTI, Amikacin, Cefuroxime, Linezolid, and levofloxacin can be used for empirical treatment. Conclusion: The prevalence rate of gram-positive cocci associated UTI in this study was 7.3% (164/2253). The emergence of drug resistance in these pathogens to commonly used antibiotics is a thing of concern. Therefore, efficient antimicrobial stewardship programmes must be in place.

Prevalence and Molecular Characterization of Vancomycin Variable Enterococcus faecium Isolated From Clinical Specimens.

Vancomycin variable Enterococcus (VVE) bacteria are phenotypically susceptible to vancomycin, but they harbor the vanA gene. We aimed to ascertain the prevalence of VVE among clinically isolated vancomycin-susceptible Enterococcus faecium (VSE) isolates, as well as elucidate the molecular characteristics of the vanA gene cluster within these isolates. Notably, we investigated the prevalence and structure of the vanA gene cluster of VVE. Between June 2021 and May 2022, we collected consecutive, non-duplicated vancomycin-susceptible Enterococcus faecium (VSE) samples. Real-time PCR was performed to detect the presence of vanA, vanB, and vanC. Overlapping PCR with sequencing and whole-genome sequencing were performed for structural analysis. Sequence types (STs) were determined by multilocus sequence typing. Exposure testing was performed to assess the ability of the isolates to acquire vancomycin resistance. Among 282 VSE isolates tested, 20 isolates (7.1%) were VVE. Among them, 17 isolates had partial deletions in the IS1216 or IS1542 sequences in vanS (N=10), vanR (N=5), or vanH (N=2). All VVE isolates belonged to the CC17 complex and comprised five STs, namely ST17 (40.0%), ST1421 (25.0%), ST80 (25.0%), ST787 (5.0%), and ST981 (5.0%). Most isolates were related to three hospital-associated clones (ST17, ST1421, and ST80). After vancomycin exposure, 18 of the 20 VVEs acquired vancomycin resistance. Considering the high reversion rate, detecting VVE by screening VSE for vanA is critical for appropriate treatment and infection control.

Enterocins Produced by Enterococci Isolated from Breast-Fed Infants: Antilisterial Potential.

Enterocins are bacteriocins synthesized by Enterococcus strains that show an interesting antimicrobial effectiveness against foodborne pathogens such as Listeria monocytogenes. The objectives of this study were to identify and analyze the expression of enterocin genes of Enterococcus isolated from breast-fed infants and evaluate their ability to inhibit three human isolates of virulent L. monocytogenes, as well as some probiotic bacteria. The susceptibility of the strains of L. monocytogenes to fifteen antibiotics was tested, detecting their resistance to cefoxitin (constitutively resistant), oxacillin, and clindamycin. The production of enterocins A, B, and P was observed in Enterococcus faecium isolates, while enterocin AS-48 was detected in an Enterococcus faecalis isolate. AS-48 showed antilisterial activity by itself, while the joint action of enterocins A and B or B and P was necessary for inhibiting L. monocytogenes, demonstrating the synergistic effect of those combinations. The presence of multiple enterocin genes does not assure the inhibition of L. monocytogenes strains. However, the expression of multiple enterocin genes showed a good correlation with the inhibition capacity of these strains. Furthermore, the potential beneficial strains of lactobacilli and bifidobacteria examined were not inhibited by any of the enterocins produced individually or in combination, with the exception of Bifidobacterium longum BB536, which was inhibited by enterocin AS-48 and the joint production of enterocins A and B or B and P. The enterocins studied here could be candidates for developing alternative treatments against antibiotic-resistant bacterial infections. Moreover, these selected enterocin-producing E. faecium strains isolated from breast-fed infants could be used as probiotic strains due to their antilisterial effect, as well as the absence of virulence factors.

Genetic analysis of vancomycin-variable Enterococcus faecium clinical isolates in Italy

PurposeTo investigate the occurrence of vancomycin-variable enterococci (VVE) in a hospital in central Italy.MethodsvanA positive but vancomycin-susceptible Enterococcus faecium isolates (VVE-S) were characterized by antibiotic susceptibility tests, molecular typing (PFGE and MLST), and WGS approach. The reversion of VVE-S to a resistant phenotype was assessed by exposure to increasing vancomycin concentrations, and the revertant isolates were used in filter mating experiments. qPCR was used to analyze the plasmid copy number.ResultsEleven putative VVE-S were selected. WGS revealed two categories of vanA cluster plasmid located: the first type showed the lack of vanR, the deletion of vanS, and an intact vanH/vanA/vanX cluster; the second type was devoid of both vanR and vanS and showed a deletion of 544-bp at the 5′-end of the vanH. Strains (n = 7) carrying the first type of vanA cluster were considered VVE-S and were able to regain a resistance phenotype (VVE-R) in the presence of vancomycin, due to a 44-bp deletion in the promoter region of vanH/vanA/vanX, causing its constitutive expression. VVE-R strains were not able to transfer resistance by conjugation, and the resistance phenotype was unstable: after 11 days of growth without selective pressure, the revertants were still resistant but showed a lower vancomycin MIC. A higher plasmid copy number in the revertant strains was probably related to the resistance phenotype.ConclusionWe highlight the importance of VVE transition to VRE under vancomycin therapy resulting in a potential failure treatment. We also report the first-time identification of VVE-S isolates pstS-null belonging to ST1478.

Possibility of transfer and activation of 'silent' tetracycline resistance genes among Enterococcus faecalis under high-pressure processing

In this study, the tetracycline resistance of Enterococcus faecalis strains isolated from food was determined and molecular analyses of the resistance background were performed by determining the frequency of selected tetracycline resistance genes. In addition, the effect of high-pressure stress (400 and 500 MPa) on the expression of selected genes encoding tetracycline resistance was determined, as well as changes in the frequency of transfer of these genes in isolates showing sensitivity to tetracyclines. In our study, we observed an increase in the expression of genes encoding tetracyclines, especially the tet(L) gene, mainly under 400 MPa pressure. The study confirmed the possibility of transferring genes encoding tetracyclines such as tet(M), tet(L), tet(K), tet(W) and tet(O) by horizontal gene transfer in both control strains and exposed to high-pressure. Exposure of the strains to 400 MPa pressure had a greater effect on the possibility of gene transfer and expression than the application of a higher-pressure. To our knowledge, this study for the first time determined the effect of high-pressure stress on the expression of selected genes encoding tetracycline resistance, as well as the possibility and changes in the frequency of transfer of these genes in Enterococcus faecalis isolates showing sensitivity to tetracyclines and possessing silent genes. Due to the observed possibility of increased expression of some of the genes encoding tetracycline resistance and the possibility of their spread by horizontal gene transfer to other microorganisms in the food environment, under the influence of high-pressure processing in strains phenotypically susceptible to this antibiotic, it becomes necessary to monitor this ability in isolates derived from foods.

Interactions between commensal Enterococcus faecium and Enterococcus lactis and clinical isolates of Enterococcus faecium.

Enterococcus faecium (Efm) is a versatile pathogen, responsible for multidrug-resistant infections, especially in hospitalized immunocompromised patients. Its population structure has been characterized by diverse clades (A1, A2, and B (reclassified as E. lactis (Ela)), adapted to different environments, and distinguished by their resistomes and virulomes. These features only partially explain the predominance of clade A1 strains in nosocomial infections. We investigated in vitro interaction of 50 clinical isolates (clade A1 Efm) against 75 commensal faecal isolates from healthy humans (25 clade A2 Efm and 50 Ela). Only 36% of the commensal isolates inhibited clinical isolates, while 76% of the clinical isolates inhibited commensal isolates. The most apparent overall differences in inhibition patterns were presented between clades. The inhibitory activity was mainly mediated by secreted, proteinaceous, heat-stable compounds, likely indicating an involvement of bacteriocins. A custom-made database targeting 76 Bacillota bacteriocins was used to reveal bacteriocins in the genomes. Our systematic screening of the interactions between nosocomial and commensal Efm and Ela on a large scale suggests that, in a clinical setting, nosocomial strains not only have an advantage over commensal strains due to their possession of AMR genes, virulence factors, and resilience but also inhibit the growth of commensal strains.

Evaluation of antibiotic resistance and quorum sensing assay of Enterococcus faecalis isolates from the urine samples

The Gram-positive Enterococci found in the normal flora of the gastrointestinal tract (GIT) and urinary tract infection (UTI) were characterized by high resistance to antibiotics and large virulence. The aim of our study was to evaluate the diversity of antibiotic-resistant and quorum-sensing assays of Enterococcus faecalis isolated from urine samples of children aged between 11 and 15 with UTI. A total of 155 samples were collected from hospitals and diagnostic centres of the Kalaburagi region. The E. faecalis isolates were screened on bile esculin azide agar media. Antibiotic susceptibility test was performed using different antibiotics as per Clinical and Laboratory Standards Institute guidelines. Minimum inhibitory concentrations (MICs) for gentamycin and vancomycin were determined by the method of broth dilution. Quorum sensing assay of E. faecalis was determined by a bioluminescence-based method. The species-level identification of E. faecalis isolates was determined by 16S rRNA polymerase chain reaction amplification. The incidence of E. faecalis in urine samples was found to be 47.74% (74/155). Antimicrobial susceptibility test (AST) indicates that 27% of isolates were multi-drug resistant, showing increased resistance to gentamycin and vancomycin (54.05%). MIC results show that 23 isolates were resistant to gentamycin with a concentration of 128 μg/ml and 17 isolates were resistant to vancomycin with a concentration of 1024 μg/ml. The quorum sensing assay has shown that the normalized relative light units range from 0.4 to 1.2. According to the 16srRNA gene amplification, out of 10 isolates, seven were identified as Enterococcus faecalis and three isolates were Streptococcus gallolyticus. This study emphasizes the development of increased vancomycin resistance of E. faecalis and indicates the presence of virulence factors through the autoinducer assay. It is been suggested that major precautionary measures should be implemented in the eradication of VRE in UTI infection.

The first vanE-type vancomycin resistant Enterococcus faecalis isolates in Norway – phenotypic and molecular characteristics

We aimed to characterize the vanE cluster and its genetic support in the first Norwegian vanE-type isolates and assess genetic relatedness to other vanE isolates. Two vanE-type vancomycin resistant Enterococcus faecalis (vanE-VREfs) isolates (E1 and E2) recovered from the same patient 30 months apart were examined for antimicrobial susceptibility, genome sequence, vancomycin resistance induction, vanE transferability, genome mutation rate, and phylogenetic relationship to E. faecalis closed genomes and two vanE-VREfs from North America. The ST34 E1 and E2 strains expressed low-level vancomycin resistance and susceptibility to teicoplanin. Their vanE gene clusters were part of a non-transferable Tn6202. The histidine kinase part of vanSE was expressed although a premature stop codon (E1) and insertion of a transposase (E2) truncated their vanSE gene. The vancomycin resistance phenotype in E1 was inducible while constitutive in E2. E1 showed a 125-fold higher mutation rate than E2. Variant calling showed 60 variants but nearly identical chromosomal gene content and synteny between the isolates. Their genomes also showed high similarity to another ST34 vanE-VREfs from Canada. In-depth genomic analyses of the first two vanE-VREfs found in Europe identified a single chromosomal insertion site of two variants of vanE-conferring Tn6202. Single nucleotide polymorphism (SNP) and core genome multilocus sequence type (cgMLST) analyses show the genomes are different. This can be explained by the high mutation rate of E1 and acquisition of different mobile genetic elements; thus, we believe the two isolates from the same patient are genetically related. Genome similarities also suggest relatedness between the Canadian and Norwegian vanE-VREfs.

Evaluation of the efficacy of three medical device detergents on bacteria and yeast derived biofilm: a comparative study

Background: This study aimed to evaluate the efficacy of three medical detergents against bacteria and yeast-derived biofilms. Methods: The biofilm removal efficacy of EmpowerTM (Metrex, USA), CidezymeTM (Johnson and Johnson Medical Inc, USA), and Matrix mintTM (Whiteley Medical, Australia) were compared to that of chlorine bleach. Biofilms were produced using Staphylococcus aureus RN9120, Escherichia coli ATCC35218, Pseudomonas aeruginosa ATCC27853, Candida albicans ATCC14053, and clinical isolates of Enterococcus faecalis, E. coli, Klebsiella pneumoniae, Candida auris, and Trichosporon asahii. The organisms were suspended in tryptic soy broth (TSB) in 96-well microplates and cultured for 72 hours. They were treated with the detergents, and the residual biofilm mass was quantified using crystal violet staining followed by optical density measurements at 620 nm (OD620 ). Results: EmpowerTM and CidezymeTM significantly reduced the biofilm mass derived from all species by > 50% of OD620 at 37ºC except those from E. faecalis, T. asahii, and C. auris. Matrix mintTM had no effect on the biofilms under any condition. Conclusion: The culture conditions and the species of the biofilm-producing organism influenced the effectiveness of the detergent. Biofilms produced by E. faecalis, C. auris, and T. asahii were resistant to all detergent treatments under all conditions.

Investigation of some changes and clonal relationship in enterococci isolates due to relocation of a hospital.

To investigate the isolation rates, antimicrobial resistance rates, minimum inhibitory concentration values of antimicrobial agents, and clonal relationships of Enterococcus faecalis and Enterococcus faeciumdue to the relocation of a hospital to a newly constructed building. The comparative, prospective study was conducted at adult general intensive care units of the Mus State Hospital, Mus, Turkey, in two phases; before the relocation from January 25 to December 1, 2014, and after the relocation from February 10 to May 24, 2015. Rectal swab samples were collected 72 hours post-hospitalisation. Identification of Enterococcus faecalis and Enterococcus faeciumisolates was determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and antimicrobial resistance with minimum inhibitory concentration values was detected with Vitek 2 system. The clonal relatedness among the strains was investigated by pulsed-field gel electrophoresis. Data was analysed using SPSS 23. Of the 69 patients, 37(53.62%) were related to pre-relocation phase; 20(54.1%) females and 17(45.9%) males with mean age 62.81±21.71 years. There were 32(46.37%) patients in the post-relocation phase; 13(40.6%) females and 19(59.4%) males with mean age 62.69±21.35 years (p>0.05). Of the 84 enterococci strains isolated, 51(60.7%) were Enterococcus faecium; 28(55%) before relocation and 23(45%) after relocation (p=0.77). The remaining 33(39.3%) isolates were Enterococcus faecalis; 16(48.5%) before relocation and 17(51.5%) after relocation (p=0.73). Multiple strains were located in 7(18.9%) patients before relocation and in 7(21.9%) after relocation. In 1(3.1%) patient after relocation, 2(8.7%) Enterococcus faecium isolates with different resistance and pulsed-field gel electrophoresis patterns were detected. There were no significant differences between the isolation and antibiotic resistance rates before and after relocation (p>0.05), and a clonal relation between the isolates was not detected (p>0.05). Decreased minimum inhibitory concentration values were noted for some antibiotics. Clonal relationship between the isolates and change in the rates of isolation and antimicrobial resistance of Enterococcus faecalis and Enterococcus faecium was not detected due to relocation. Minimum inhibitory concentration values could be used to reveal relocation-related changes in isolates obtained from patients hospitalised in intensive care units.

Antimicrobial Resistance Profiles of Enterococcus faecium and Enterococcus faecalis Isolated from Healthy Dogs and Cats in South Korea.

Enterococcus spp. are typically found in the gastrointestinal tracts of humans and animals. However, they have the potential to produce opportunistic infections that can be transmitted to humans or other animals, along with acquired antibiotic resistance. In this study, we aimed to investigate the antimicrobial resistance profiles of Enterococcus faecium and Enterococcus faecalis isolates obtained from companion animal dogs and cats in Korea during 2020-2022. The resistance rates in E. faecalis towards most of the tested antimicrobials were relatively higher than those in E. faecium isolated from dogs and cats. We found relatively higher resistance rates to tetracycline (65.2% vs. 75.2%) and erythromycin (39.5% vs. 49.6%) in E. faecalis isolated from cats compared to those from dogs. However, in E. faecium, the resistance rates towards tetracycline (35.6% vs. 31.5%) and erythromycin (40.3% vs. 35.2%) were comparatively higher for dog isolates than cats. No or very few E. faecium and E. faecalis isolates were found to be resistant to daptomycin, florfenicol, tigecycline, and quinupristin/dalfopristin. Multidrug resistance (MDR) was higher in E. faecalis recovered from cats (44%) and dogs (33.9%) than in E. faecium isolated from cats (24.1%) and dogs (20.5%). Moreover, MDR patterns in E. faecalis isolates from dogs (27.2%) and cats (35.2%) were shown to encompass five or more antimicrobials. However, E. faecium isolates from dogs (at 13.4%) and cats (at 14.8%) were resistant to five or more antimicrobials. Taken together, the prevalence of antimicrobial-resistant enterococci in companion animals presents a potential public health concern.

Genomic insights into Enterococcus faecium isolates from marine bivalves highlight One Health concerns and healthcare linkages.

Enterococci, especially Enterococcus faecium, are one of today's leading causes of multidrug-resistant infections in hospital settings. The marine environment may harbour enterococci, but its role as an evolutionary niche and as a vector for the spread of enterococci is sparsely investigated. Hence, by applying enterococci in bivalves as a sentinel tool, this study aimed to describe the prevalence of enterocooci along the Norwegian coast and in addition the phylogeny of E. faecium in particular. Enterococci in batch samples of marine bivalves, harvested from 86 different locations, were quantitatively examined by a culture-dependent most probable number (MPN) method. Isolates were identified by MALDI-TOF-MS prior to antimicrobial susceptibility testing by broth microdilution. In-detail analyses of a representative selection of E. faecium isolates (n=148) were done by Illumina whole-genome sequencing, and assembled genomes were compared to closed E. faecium genomes in the public databases and to genomes from commensal and clinical isolates from Norway. Diversity among E. faecium within the same batch sample of bivalves was also explored. Enterococci were detected in 287 of the 471 examined bivalve samples, but in low concentrations with a median value of <18 MPN /100 g. From positive samples, 479 isolates of enterococci were identified belonging to ten different species, where E. faecium (n=247), Enterococcus hirae (n=114) and Enterococcus faecalis (n=66) were most frequently found. Resistance towards one or more antimicrobial agents was observed in 197 isolates (41 %), none of the isolates showed acquired resistance to vancomycin or linezolid. Phylogenetic analyses revealed high diversity among the E. faecium isolates and showed that the marine niche is dominated by strains from the non-clinical setting belonging to clade A2 (n=85) and B (E. lactis) (n=60). Only three isolates belonged to the hospital-associated clade A1 (ST80 and ST117). Two of these clustered with one isolate from a hospitalized patient and one from a non-hospitalized person. This study demonstrated a high prevalence, but low concentrations of enterococci in bivalves, and low levels of antimicrobial resistance. E. faecium genomes showed high population diversity and that very few E. faecium isolates in bivalves may have arisen from the human healthcare system. A systematic surveillance of target micro-organisms applying methods examining multiple isolates from the same bivalve sample provides important data to assess the enterococcal phylogeny, antimicrobial resistance and the level of faecal pollution in the marine environment.

Isolation and Identification of Enterococcus Faecium from Locally (Mukra) and Study some of their Morphological and Biochemical Properties

The study aimed to isolate Enterococcus faecium from pickled turnip and red beet, locally known Mukra and study some of phenotypic characteristics and its ability to antagonism against some pathogenic bacteria. The isolate was selected depending on the shape of the colony growing in MRS-Agar, as it formed smooth, convex, and translucent domed white or creamy colonies. The microscopic characteristics of the bacteria were tested, as they were double spherical or with short chains, non-motile and non-spore-forming. Biochemical tests showed that the isolate is negative for catalase, amylase and gelatinase, does not consume citrate, does not form ammonia from arginine, does not reduce nitrates, and does not produce indole from tryptophan. While the sugars fermentation tests showed the ability of the isolate to ferment Raffinose, Glucose, Sucrose, Xylose, Arabinose, Lactose, Melebose, Trehalose, Rhamnose, Lactose, Cellobiose, Maltose, and the ability to produce gas from the fermentation of Xylose, Raffinose, Glucose. The PCR results showed that the isolate belongs to Enterococcus faecium-K189-3. The isolate showed the ability to inhibit some types of pathogenic bacteria.

Isolation and characterization of antibiotics resistance Enterococcus faecium from mastitic cow's milk

Isolation and characterization of antibiotics resistance Enterococcus faecium from mastitic cow's milk

2110. Enhancing Antibiotic Efficacy Against Enterococcal faecium with Bacteriophage and Antimicrobial Peptides

Abstract Background The urgency to find new alternatives has increased as multidrug-resistant organisms enhance their defense mechanisms. In recent years, there has been a renewed interest in bacteriophages as potential alternatives or adjuncts to antibiotic therapy. However, there is limited information on the impact of bacteriophages with the innate immune system. Cathelicidin LL-37, a cationic peptide with antimicrobial properties, functions in coordination with the immune system to eradicate pathogens. We aim to evaluate how the survival of daptomycin (DAP) and ampicillin (AMP) resistant or non-susceptible Enterococcus faecium isolates are impacted by the interaction between bacteriophage, LL-37, and the addition of DAP/AMP. Methods Two E. faecium clinical isolates were utilized: R497 and HOU503. Bacteriophages provided by the Naval Medical Research Command, NV- 503-01 and NV-497 were quantified and propagated to approximately 108 PFU/mL. LL-37 was prepared at a final concentration of 640μM and diluted in RPMI intentionally to a suboptimal concentration of 0.5μM to adjust for a 70-80% survival rate. This was done to detect a synergistic interaction more easily with the addition of phage. Samples were incubated at 37 °C and plated in triplicate on brain heart infusion agar at 2h. After 24h of incubation, colonies were counted for and analyzed. Data was expressed as a mean percent survival with standard deviation. ANOVA with Tukey's HSD post-hoc test was used to determine the variations between each combination used. Results The targeted bacterial survival rate against E. faecium isolates was attained by LL-37 at a concentration of 0.5μM. However, when R497 and HOU503 was combined with LL-37, bacteriophage, and DAP/AMP the percentage of bacterial survival was significantly (P&amp;lt; 0.05) lower than that of the growth control, LL-37, AMP/DAP, and bacteriophage alone. Conclusion By conducting bacterial survival assays, we observed a notable enhancement in the elimination of multi-drug resistant E. faecium when LL-37, bacteriophage, and DAP/AMP were added. Examining this interaction over longer periods, both with and without varying antimicrobials, will yield more understanding regarding the potential involvement of bacteriophage in the innate immune system's response to fighting infections. Disclosures Michael J. Rybak, PharmD, PhD, MPH, Abbvie, Merck, Paratek, Shionogi, Entasis, La Jolla, T2 Biosystems: Advisor/Consultant

Prevalence and Antimicrobial Resistance Patterns of Hospital Acquired Infections through the COVID-19 Pandemic: Real-Word Data from a Tertiary Urological Centre

Background: Antimicrobial resistance (AMR) remains a significant public health concern, closely linked to antibiotic overuse. During the COVID-19 pandemic, broad-spectrum antibiotics were frequently administered, potentially exacerbating AMR. This study aimed to assess AMR patterns in our urology department before and after the pandemic. Methods: The study encompassed patients admitted to our urology department from January 2016 to December 2022, with confirmed urinary tract infection, bloodstream infection, or wound infection based on positive culture results. Descriptive statistics, including mean, frequency, and percentage, summarized the data. Trends were analyzed using the Joinpoint Regression program. Results: A total of 506 patients were included. Escherichia coli and Klebsiella pneumoniae displayed resistance rates of 65% and 62% to ciprofloxacin, respectively. K. pneumoniae showed resistance rates of 41% to piperacillin tazobactam and 3rd generation cephalosporins (3GC). Carbapenem resistance was observed in 38% of K. pneumoniae isolates. Additionally, 26% of E. coli, 26% of K. pneumoniae, and 59% of Proteus mirabilis isolates were ESBL-positive. Among gram+, 72% of Staphylococcus aureus isolates were MRSA, and 23% of Enterococcus faecium isolates were VRE. Trends in antimicrobial susceptibility patterns over the 7-year study period revealed a statistically significant decrease in E. coli resistance to amoxicillin-clavulanic acid (APC: −5.85; C.I. 95% p &lt; 0.05) and a statistically significant increase in K. pneumoniae resistance to 3GC (APC: 9.93; CI (−19.9–14.4 95% p &lt; 0.05). There were no statistically significant differences in AMR incidence pre- and post-COVID-19. Conclusion: The COVID-19 pandemic did not appear to influence the AMR incidence in our urology department. However, the overall prevalence of AMR and MDROs in our department remains high compared to European AMR.

Estimation of the In-Vitro Synergistic Effect of Vancomycin with Bacteriophage and its Endolysin on Iraqi Local Isolates of Enterococcus faecalis

Enterococcus faecalis is known to cause many serious infections with an upsurge in the development of antibiotic resistant strains, especially vancomycin. Lytic bacteriophage and its endolysin have the potential to affect and lyse specific bacterial cells making them a possible alternative therapy to antibiotic resistant strains. The study aimed to investigate the synergistic effect of phage/ phage cocktail/ endolysin with vancomycin on E. faecalis isolated from urine samples. The antibioresistant profile 17 E. faecalis was done with an emphasis on the sensitivity to vancomycin. The lytic phages were isolated from the local environment. The infectivity and the coverage rate for the phage and phage cocktail were calculated. The enzyme of the lytic phage was isolated by chromatography. The synergistic effect was tested by turbidity reduction assay to evaluate the growth reduction level. It had been found that 6/17 E. faecalis isolates were resistant to vancomycin, 7/17 isolates were intermediate, and 4/17 were susceptible. Three lytic phages were identified with an infectivity rate of 100%, 76.4%, and 88.2%. The phage cocktail yielded 100% rate of infectivity and 91.6 % coverage rate. The reduction in the growth levels of the 17 E.faecalis after treatment with phage / phage cocktail/ vancomycin/ mixture of vancomycin and phage cocktail was effective. The synergistic effect of vancomcyin together with the phage cocktail significantly reduced the growth of 70.6% of the isolates. The antibacterial activity of vancomycin plus endolysin treatment was revealed in a time dependent manner. Thus, the synergistic effect of vancomycin with phage/ endoloysin illustrated an effective method to treat the antibiotic-resistant strains of E. faecalis to overcome this emerging clinical problem.

Molecular characterization and biofilm formation ability of Enterococcus faecium and Enterococcus faecalis bloodstream isolates from a Chinese tertiary hospital in Beijing

To investigate the molecular characteristics and biofilm-forming ability of 116 Enterococcus faecium (Efm) and 72 Enterococcus faecalis (Efs) isolates obtained from patients with bloodstream infections (BSI) at a Chinese hospital between July 2011 and March 2018. The presence of glycopeptide resistance genes and five virulence genes (esp, gelE, asa1, hyl, and cylA) was screened using two multiplex PCR. MLST was used to assess the clonality. Crystal violet staining was used to detect biofilms. Vancomycin resistance was detected in 30.1% of Efm and 2.8% of Efs isolates, respectively. All VRE strains carried the vanA gene. The esp, gelE, asa1, and cylA genes in 72 Efs strains were detected at 62.5%, 84.7%, 84.7%, and 69.4%, respectively. Among the 116 Efm isolates, 74.1% and 25.8% carried esp and hyl, respectively. The esp gene was significantly associated with vancomycin-resistant Efm (VREfm) compared to vancomycin-susceptible Efm (VSEfm). In total, 91.7% of Efs and 20.0% of Efm produced biofilms. Twenty-six STs were identified among the 72 Efs isolates, with ST4 (29.2%) being the predominant. In total, 116 Efm strains were grouped into 26 STs, with ST78 (46.6%) being the predominant. Both VREfm (41.7%) and VSEfm (48.8%) were dominant in ST78. There is no clear evidence suggesting that some STs are associated with vancomycin resistance or biofilm formation. Both Efm and Efs BSI isolates showed a polyclonal pattern with a dominant clone and many unique types, implying the coexistence of clonal dissemination and an influx of new clones. The horizontal transmission of resistance genes may play a more important role in VREfm prevalence than clonal expansion.