Enterococcus Strains Research Articles

Green-Synthesis of CuO and Ce-Doped CuO Nanoparticles Using Aqueous Extract of Yam Peel and their Antimicrobial Properties

The rise of drug-resistant microbes presents a critical global health challenge in the 21st century, necessitating the development of potent antimicrobial agents with broad-spectrum activity. Therefore, in this study, we explored the antimicrobial activity of copper oxide (CuO) and cerium-doped CuO (Ce-doped CuO) obtained through a facile green synthesis approach. The nanoparticles were synthesized using aqueous extract of Dioscorea spp (yam peel) and characterized for their physicochemical properties using X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), electron microscopy, and UV-Vis spectroscopy. Their antimicrobial efficacy was tested against three bacterial strains—Enterococcus faecalis, Escherichia coli, and Listeria monocytogenes—and three fungal strains—Aspergillus niger, Mucor mucedo, and Penicillium chrysogenum. The results demonstrated that cerium doping significantly altered the physicochemical properties of CuO, leading to enhanced antimicrobial activity. The antibacterial zone of inhibition ranged from 7.50 to 16.55 mm for CuO and 11.80 to 19.50 mm for Ce-doped CuO. For antifungal activity, the minimum inhibitory concentration (MIC) ranged from 0.25 to 0.30 mg/mL for CuO and 0.025 to 0.03 mg/mL for Ce-doped CuO. Notably, the Ce-doped CuO nanoparticles exhibited antimicrobial effects comparable to standard antibiotics. These findings highlight the potential of green-synthesized Ce-doped CuO nanoparticles as effective antimicrobial agents. By leveraging green synthesis and elemental doping, this study offers a promising solution to combat a wide spectrum of infectious pathogens.

Antimicrobial resistance, virulence gene profiles, and molecular epidemiology of enterococcal isolates from patients with urinary tract infections in Shanghai, China.

This study highlighted the critical need to understand Enterococcal strains causing UTIs in Shanghai, given their high prevalence. By assessing antimicrobial resistance profiles, virulence gene presence, and molecular epidemiology, the research offered valuable insights into the local epidemiology of Enterococcus faecalis and Enterococcus. faecium. Identifying linezolid-resistant strains, all of which carry the optrA gene, including the first report of ST300 in China and recognizing dominant MLST types, such as ST16 and ST179 for E. faecalis and ST78 for E. faecium, are vital for guiding treatment and addressing the challenges these infections present. The data emphasize the need for ongoing surveillance and customized therapeutic approaches to combat emerging resistance and virulence factors in Enterococcal UTIs.

The prevalence of optrA-carrying Enterococci in the vaginal micro-ecology of pregnant women in late pregnancy.

The colonization of Enterococcus in the female vagina leads to neonatal and pediatric enterococcal septicemia. Linezolid (LZD) is a kind of mainstream drug for treating multidrug-resistant Gram-positive infections. OptrA is the main LZD-resistance gene at Enterococci in human isolates. It is essential to explore the prevalence of optrA-carrying Enterococcus in vaginal secretions of late pregnant women and the drug resistance of optrA. From May to June 2023, this study recruited 340 volunteers in late pregnancy (35-40 weeks of pregnancy) to provide non-repetitive vaginal discharge samples. Luria-Bertani broth and florfenicol (10 µg/mL) were used to enrich bacteria. Enterococci was identified through time-of-flight mass spectrometry. Additionally, antimicrobial susceptibility, polymerase chain reaction, and next-generation sequencing assays were applied to this study. Fifty-four optrA-carrying Enterococcus strains were obtained, the proportion of the whole vagina of late pregnant women was 15.88% (54 out of 340), and Enterococcus faecalis account the highest proportion. All optrA-carrying Enterococcus were resistant to at least three drugs. Tetracycline, chloramphenicol, erythromycin, and LZD have higher bacterial resistance rates. Genetic environment analysis revealed that IS1216E, fexA, and erm(A) may synergistically exert multidrug resistance with optrA. It is necessary to strengthen the surveillance of optrA-carrying Enterococcus in pregnant women. This study provides scientific support for controlling hospital infections and managing antibiotic-resistant bacteria, and provides a scientific basis for rational clinical medication.IMPORTANCEThe disruption of cervicovaginal microbiota homeostasis is considered a key factor in causing imbalance in the microenvironment, leading to inflammation, transmission of infections, and illness. Enterococcus is considered a major cause of healthcare-related infections globally. It has resistance to multiple antimicrobial drugs, which pose significant challenges for clinical treatment. Therefore, it is crucial to assess the prevalence of optrA-carrying Enterococcus in vaginal secretions of late pregnant women and the drug resistance of optrA. This study detected 15.88% of optrA-carried Enterococci in 340 pregnant women. Furthermore, we found that optrA-carrying Enterococcus strains are highly resistant to tetracycline, chloramphenicol, erythromycin, and Linezolid. Additionally, genetic environment analysis revealed that IS1216E, fexA, and erm(A) may synergistically exert multidrug resistance with optrA. This study provides scientific support for controlling hospital infections and managing antibiotic-resistant bacteria and provides a scientific basis for rational clinical medication.

Exploring the In Vitro Effects of Cassava Diets and Enterococcus Strains on Rumen Fermentation, Gas Production, and Cyanide Concentrations.

This study examined the effects of adding CUB alongside HCN sources from fresh cassava diets on HCN reduction, gas production, and in vitro digestibility. A completely randomized design (CRD) with a 2 × 2 × 3 + 1 factorial approach was used, where Factor A was the HCN source [fresh cassava root (FCR) or leaf (FCL)], Factor B was the HCN concentration (300 and 600 mg/kg dry matter (DM)), and Factor C was the bacterial supplement [no-CUB, E. faecium KKU-BF7 (CUB1), and E. gallinarum KKU-BC15 (CUB2)]. Statistical analysis was performed using the PROC GLM procedure in SAS. No interaction was observed among the main factors on gas kinetics and cumulative gas (p > 0.05). The addition of CUB1 or CUB2 enhanced cumulative gas production compared to the no-CUB group (p = 0.04). Cyanide degradation efficiency was high when FCR was included at a high HCN level. At 12 h post-incubation, HCN degradation efficiency was higher in the CUB2 and CUB1 groups, reaching 98.44-99.07% compared to the no-CUB group. The higher HCN level increased in vitro acid detergent fiber digestibility (IVADFD) (p = 0.01) by 7.20% compared to the low HCN level, and CUB2 further improved IVADFD. Compared to the FCL-fed group, FCR supplementation increased total VFA concentration (p = 0.03) and propionic acid (C3) concentration (p = 0.04). The addition of CUB2 further enhanced propionic acid concentration by 8.97% compared to no-CUB supplementation (p = 0.04). These results indicate that supplementing E. gallinarum KKU-BC15 at the highest HCN levels in FCR boosts HCN degradation efficiency, fiber digestibility, total VFA, and C3 concentration.

Comprehensive Study of Antibiotic Resistance in Enterococcus spp.: Comparison of Influents and Effluents of Wastewater Treatment Plants.

Background/Objectives: The spread of antibiotic resistance, particularly through Enterococcus spp., in wastewater treatment plants (WWTPs) poses significant public health risks. Given that research on antibiotic-resistant enterococci and their antibiotic-resistance genes in aquatic environments is limited, we evaluated the role of Enterococcus spp. in WWTPs by comparing the antibiotic resistance rates, gene prevalence, biofilm formation, and residual antibiotics in the influent and effluent using culture-based methods. Methods: In 2022, influent and effluent samples were collected from 11 WWTPs in South Korea. Overall, 804 Enterococcus strains were isolated, and their resistance to 16 antibiotics was assessed using the microdilution method. Results: High resistance to tetracycline, ciprofloxacin, kanamycin, and erythromycin was observed. However, no significant differences in the overall resistance rates and biofilm formation were observed between the influent and effluent. Rates of resistance to ampicillin, ciprofloxacin, and gentamicin, as well as the prevalence of the tetM and qnrS genes, increased in the effluent, whereas resistance rates to chloramphenicol, florfenicol, erythromycin, and tylosin tartrate, along with the prevalence of the optrA gene, decreased. E. faecium, E. hirae, and E. faecalis were the dominant species, with E. faecalis exhibiting the highest resistance. Conclusions: Our results suggest that WWTPs do not effectively reduce the rates of resistant Enterococcus spp., indicating the need for continuous monitoring and improvement of the treatment process to mitigate the environmental release of antibiotic-resistant bacteria.

Prevention of possible vancomycin resistant enterococcus outbreak caused by infants transferred due to earthquake-trauma

Infection control measures are disrupted due to the large number of patients admitted in a very short time, in the early post-earthquake period. Inter-hospital or intra-hospital transfers can cause the spread of resistant bacteria. We reported the possible outbreak and its prevention caused by vancomycin-resistant enterococcus (VRE) strains carried by infants transferred from another unit to our neonatal intensive care unit after the earthquake. Screening cultures (rectal swab) were obtained from infants on admission and patients were kept in contact isolation. We studied the clonal analysis of VRE strains isolated from patients by Pulse Field Gel Electroforesis.

Antimicrobial Resistance Trends and Epidemiological Characteristics of Isolates from Intra-Abdominal Infections in China: A 6-Year Retrospective Study (2017-2022).

Background: Antimicrobial resistance represents a continuing threat to the health of patients with intra-abdominal infections (IAIs). This study aimed to provide clinicians with guidance to optimize antibiotic therapy. Methods: The clinical data and antibiotic susceptibility results of pathogens from patients with IAIs from 2017 to 2022 were retrospectively collected. The 6-year period was segmented into two stages, namely, the early (2017-2020) and recent stages (2021-2022). The distribution and antibiotic resistance of pathogens were compared between the stages. Results: In total, 5,795 pathogens were isolated from 2,283 patients diagnosed with IAIs. Gram-negative bacteria, Gram-positive bacteria, and fungi accounted for 71.0%, 21.4%, and 7.5% of the isolates, respectively. Klebsiella pneumoniae (1,037, 17.9%) was the primary isolate. The proportion of extended-spectrum β-lactamase-producing Enterobacteriaceae was 89.8% (2,028/2,259), with extended-spectrum β-lactamase-producing Escherichia coli and K. pneumoniae accounting for 27.4% and 43.2%, respectively, of all such isolates. The carbapenem resistance rates of E. coli and K. pneumoniae were 17.1% and 75.9%, respectively. Compared with that in the early stage, the imipenem resistance rate of E. coli was significantly higher in the recent stage (13.8% vs. 25.1%, p < 0.001). Among Gram-positive bacteria, 88 strains of vancomycin-resistant Enterococcus were detected, giving a resistance rate of 10.3%, and the detection rate of methicillin-resistant Staphylococcus aureus was 65.7%. Conclusions: Enterobacteriales and non-fermentative bacteria from IAIs remain highly resistant to carbapenems. The epidemiological characteristics and antibiotic resistance profiles of pathogens in various regions should be closely monitored to mitigate the appearance of drug-resistant bacteria in clinical settings.

What role does antibiotic resistance play in secondary endodontic infections?

The systematic review utilised multiple electronic databases to gather relevant literature, including PubMed, Scopus, EMBASE, Web of Science, and Medline. The search was conducted focusing on clinical studies where Enterococcus strains were isolated to assess their antimicrobial resistance. The search strategy was designed based on the PICO model, targeting patients with secondary/persistent endodontic infections (SPEI) and evaluating the resistance profiles of Enterococci to systemic antibiotics. A total of 584 studies were initially identified through database searches. After removing duplicates, 517 studies were excluded based on title and abstract screening, leading to the final inclusion of 11 clinical trials that met the eligibility criteria. The studies were conducted in various countries, including Brazil, Lithuania, Poland, Germany, Sweden, the United States, and Japan. The focus was on permanent teeth with post-treatment apical periodontitis undergoing endodontic retreatment. Data extraction was performed independently by two reviewers, who compiled details such as authorship, year of publication, country of study, sample groups, bacterial strains tested, antibiotics used, and resistance outcomes. The risk of bias was assessed using the ROB-2 platform, and all studies were considered to have a low risk of bias, with occasional minor deficiencies noted. The analysis revealed fluctuating resistance profiles of Enterococcus faecalis (E. faecalis) and Enterococcus faecium (E. faecium) to various antibiotics over time. E. faecalis isolated from teeth with SPEI showed intermediate resistance to 16 antibiotics. The most effective antibiotics identified were the combination of amoxicillin and clavulanate, followed by amoxicillin and benzylpenicillin. In patients with penicillin allergies, moxifloxacin and azithromycin were suggested as alternatives, albeit with caution due to their varying resistance levels. The antibiotics demonstrating the highest resistance patterns against E. faecalis included clindamycin, gentamicin, metronidazole, and rifampicin, which are contraindicated in SPEI cases. Ongoing monitoring of antibiotic resistance patterns is crucial for effective endodontic treatment protocols. The emergence of resistance among Enterococci, particularly E. faecalis, necessitates regular susceptibility testing to inform clinical decisions and improve treatment outcomes. The findings underscore the need for health professionals to stay informed about the evolving resistance landscape to ensure safe and effective therapeutic regimens in endodontics.

Technological and safety properties of bacteriocin-producing &lt;i&gt;Enterococcus&lt;/i&gt; strains isolated from traditional Turkish cheeses

The aim of this study was to identify bacteriocin-producing lactic acid bacteria (LAB) isolated from traditional Turkish cheeses, evaluate their antibacterial properties and technological characteristics, and determine the safety properties of isolates. The isolated strains were identified as Enterococcus faecium DP8.3, DP9.3, and Enterococcus mundtii DP35.1, which are genetically different from each other. The bacteriocins produced by these isolates exhibited stability under high temperatures and maintained their antibacterial activity across a broad pH spectrum. The bacterial strains were fast acid producers, lacked proteolytic or lipolytic activity, were susceptible to antibiotics, and did not show antibiotic resistance or virulence genes. In addition, all the Enterococcus strains decarboxylated tyrosine and showed tdc gene expression. According to the technological properties and safety assessment of the strains, it is believed that they can be used as protective adjunct cultures in the food industry.

Optimization, structural characterization, and biological applications of exopolysaccharide produced by Enterococcus faecium KT990028

The aim of this study was to select the best exopolysaccharide (EPS) producer among the Enterococcus strains to optimize, characterize, and evaluate its biological properties. Among the eleven strains, Enterococcus faecium KT990028 was selected, and the production conditions were optimized: 16.3 % (w/v) sucrose, 0.70 % (w/v) yeast extract, 8.3 % (w/v) reconstituted skimmed milk, at 38 °C in 15 h of incubation, producing 2.880 g/L of EPS. High performance anion exchange chromatography (HPAEC) analysis revealed that the molecular weight was 166.98 kDa. HPAEC, spectroscopy (FTIR), and nuclear magnetic resonance (1H NMR) analyses revealed that the EPS was a heteropolysaccharide composed of galactose (37.74 %), rhamnose (19.79 %), arabinose (17.71 %), glucose (9.50 %), fucose (7.93 %), and mannose (7.33 %). Scanning electron microscopy showed a three-dimensional microstructure in the form of decompressed plates, with wrinkles, and pores. By means of dynamic light scattering (DLS), the EPS showed an average size varying from 135.25 ± 10.56 nm and 410.60 ± 45.20 nm, as the concentration was increased from 0.5 mg/mL to 2.0 mg/mL, respectively. X-ray diffraction revealed that the EPS has an amorphous and crystalline nature, while thermogravimetric analysis indicated stability up to 400 °C. The antioxidant effect (5 mg/mL) against DPPH, ABTS, OH, and O2 was 64.50 ± 0.71 %, 47.50 ± 0.10 %, 68.36 ± 0.59 %, and 44.83 ± 0.86 %, respectively. It was also able to inhibit and biofilm disruption of Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 6057 and had an antimicrobial effect from 50 mg/mL for the strains of against Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, Listeria monocytogenes ATCC 19117, Staphylococcus aureus ATCC 6538, and Enterococcus faecalis ATCC 6057. Cell cytotoxicity carried out using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that the EPS was safe and promoted the proliferation of Vero cells. Thus, the results indicated that the EPS from E. faecium KT990028 is a promising functional biopolymer for possible applications in the food and pharmaceutical fields.

Identification and characterization of Morganella morganii strain YC12-C3 and Enterococcus faecalis strain YC12-C10 and elucidation of its deoxynivalenol-degrading potential.

Deoxynivalenol ( DON) is one of the most harmful mycotoxins in food or feed or Traditional Chinese Medicine. An efficient and applicable method for the detoxification of DON is urgently developed. 1152 strains were isolated from the intestinal contents of crucian. Morganella morganii YC12-C3 and Enterococcus faecalis YC12-C10 were screened with the highest degradation rate of DON via HPLC methods. The optimal degradation condition of YC12-C3 and YC12-C10 is co-cultured 24 h and 36 h at 28 ℃ in LB medium with pH 7 and 1.0% inoculation dosage, respectively. LC-MS/MS and 1H NMR results show that YC12-C10 and YC12-C3 can transform DON to 3-deoxy-6-demethanol-DON, a new metabolite biotransformed from DON, by deoxidization at C3 hydroxy and de-methanal reaction at methanol moiety of C6. In addition, the DON-degradation in agricultural material assay showed that YC12-C10 and YC12-C3 can degrade 150 μg·kg-1 DON in Coix lacryma-jobi, with a degradation rate of 68.89% and 59.94%, respectively. This result shows that YC12-C10 and YC12-C3 have a sound efficiency in removing DON ability in Coix lacryma-jobi, providing a new strain resource and application technique for biological detoxification of DON in food or feed or TCM industry.

69 Urinary tract infections in preterm infants: Review in two Canadian tertiary centers

Abstract Background Urinary tract infections (UTI) in premature infants are commonly seen in the neonatal intensive care unit (NICU). The reported prevalence of UTI in this population ranges from 1.9-12.2% due to the lack of standardized criteria for UTI among infants less than two months of age. Enterococcus and Enterobacteriaceae are well known to cause UTI and their resistance pattern has been reported to be rising in the neonatal nosocomial infections worldwide. Objectives To describe the epidemiology, microbiology, and antimicrobial treatment for UTI in premature infants based on different diagnostic criteria. Design/Methods This retrospective study (2019-2022) included infants born at &amp;lt;33 weeks of gestational age (GA) with positive urine cultures and ≥5 days of antimicrobials treatment. We collected data from two tertiary level NICUs in Canada, including demographics, microbial susceptibility patterns, urinalysis and imaging results, and antibiotic treatment. Results We identified 43 infants managed as UTI based on physicians’ discretions. The median GA at birth was 26-27 weeks, while the median onset of first UTI ranged 39-80 days, with male predominance [Table 2]. Mixed growth of organisms constituted 21-42% of cases of UTI [Table 1]. Among UTI cases with only single growth organism, we found 53.5% and 18.6% cases fulfilled UTI diagnostic criteria based on the concentration of colony forming units (CFU) of &amp;gt;104/mL or &amp;gt;107/L and &amp;gt;105/mL or &amp;gt;108/L, respectively [Table 1]. Of these cases, 67.4% of infants had urinalysis sent with the culture, and the higher the concentration of CFU, the more likely the samples showed positive nitrites on urinalysis, with 100% correlation among those with CFU &amp;gt;105/mL or &amp;gt;108/L. [Table 1] The most common pathogens were Enterococcus species, E.Coli, Klebsiella species, Citrobacter species and Enterobacter cloacae complex [Table 2]. Among gram negative bacilli (E. Coli and Klebsiella species), greater than 42-80% were resistant to 3rd generation cephalosporins (3GC). We did not identify any vancomycin-resistant Enterococcus strains. Median duration of antimicrobial treatment ranged from 8-13 days, depending on the diagnostic criteria applied. At least 10% of infants with UTI showed abnormalities on kidney ultrasound [Table 2]. Conclusion There exists a large variation in UTI prevalence according to different diagnostic criteria. Abnormal urinalysis with presence of nitrites were frequently associated with UTI of high CFU count. High resistance to 3GC among gram negative bacilli was concerning and carried an impact on the appropriate empirical treatment choice.

Comparative Efficacy of Parenteral and Mucosal Recombinant Probiotic Vaccines Against SARS-CoV-2 and S. pneumoniae Infections in Animal Models.

The accumulation of specific IgG antibodies in blood serum is considered a key criterion for the effectiveness of vaccination. For several vaccine-preventable infections, quantitative indicators of the humoral response have been established, which, when reached, provide a high probability of protection against infection. The presence of such a formal correlate of vaccine effectiveness is crucial, for example, in organizing preventive measures and validating newly developed vaccines. However, can effective protection against infection occur when the level of serum antibodies is lower than that provided by parenteral vaccination? Will protection be sufficient if the same vaccine antigen is administered via mucosal membranes without achieving high levels of specific IgG circulating in the blood? In this study, we compared the immunogenicity and protective efficacy of parenteral and mucosal forms of vaccines in experimental animals, targeting infections caused by the SARS-CoV-2 coronavirus and Streptococcus pneumoniae. We investigated the protective properties of a fragment of the coronavirus S1 protein administered intramuscularly with an adjuvant and orally as part of the probiotic strain Enterococcus faecium L3 in a Syrian hamster model. A comparative assessment of the immunogenicity and protective efficacy of a recombinant tandem (PSP) of immunogenic peptides from S. pneumoniae surface proteins, administered either parenterally or orally, was performed in a Balb/c mouse model. Both models demonstrated significant differences in the immunogenicity of parenteral and oral vaccine antigens, but comparable protective efficacy.

Safety assessment of enterocin-producing Enterococcus strains isolated from sheep and goat colostrum

BackgroundThis study investigates the safety evaluation of enterocin-producing 11 E. mundtii and two E. faecium strains previously isolated from small livestock colostrums. Enterococcus species do not possess Generally Recognized as Safe (GRAS) status. Hence, it is critical to scrutinize enterococci’s antibiotic resistance, virulence characteristics, and biogenic amine production capabilities in order to assess their safety before using them as starter or adjunct cultures.ResultsEnterococcus strains showed susceptibility to medically significant antibiotics. Multiple-drug resistance (MDR) was found in only E. faecium HC121.4, and its multiple antibiotic resistance (MAR) index was detected to be 0.22. The tetL and aph(3')-IIIa were the most commonly found antibiotic resistance genes in the strains. However, E. mundtii strains HC56.3, HC73.1, HC147.1, and E. faecium strain HC121.4 were detected to lack any of the antibiotic resistance genes examined in this study. Only E. mundtii HC166.3 showed hemolytic activity, while none of the strains engage in gelatinase activity. The strains were identified to have virulence factor genes with a low rate. None of the virulence factor genes could be detected in E. mundtii HC26.1, HC56.3, HC73.1, HC165.3, HC166.8, and E. faecium HC121.4. The E. mundtii HC73.2 strain displayed the highest presence of virulence factor genes, namely gelE, efaAfs, cpd, and ccf. Similarly, the E. mundtii HC112.1 strain showed a significant presence of genes efaAfm, ccf, and acm. There was no decarboxylation of histidine, ornithine, or lysine seen in any of the strains. Nevertheless, E. faecium HC121.4 and HC161.1 strains could decarboxylate tyrosine, but E. mundtii HC26.1, HC56.3, HC73.1, HC73.2, HC112.1, HC147.1, HC155.2, HC165.3, HC166.3, HC166.5, and HC166.8 strains only showed a limited capacity for tyrosine decarboxylation. None of the strains possessed the hdc, odc, or ldc genes, but all of them had the tdc gene.ConclusionThe E. mundtii HC56.3 and HC73.1 strains were deemed appropriate for utilization in food production. Using the remaining 11 strains as live cultures in food production activities could pose a possible risk to consumer health.

The gut microbiota of wild birds undergoing rehabilitation as a reservoir of multidrug-resistant enterococci in a metropolitan area in Brazil.

Enterococci are ubiquitous usually commensal bacteria that can act as opportunistic pathogens frequently associated with resistance to multiple antimicrobial classes. A variety of animals may carry potentially harmful enterococci. In the present work, the occurrence and characteristics of enterococci recovered from the fecal microbiota of wild birds belonging to four families (Accipitridae, Cathartidae, Falconidae and Strigidae) were investigated. Enterococci were recovered from 104 (92.0%) fecal samples obtained from 113 birds, and 260 strains were selected for additional characterization. Enterococcus faecalis was the predominant species (63.8%), followed by Enterococcus hirae (16.2%), Enterococcus faecium (11.5%), Enterococcus gallinarum (5.4%), Enterococcus avium (1.5%), Enterococcus casseliflavus (0.8%), and Enterococcus raffinosus and Enterococcus cecorum (0.4% each). Major percentages (11.9% 75.0%) of nonsusceptibility were observed to quinolones (particularly to enrofloxacin), erythromycin, rifampin, nitrofurantoin, tetracycline and streptomycin. Gentamicin and ampicillin resistances (13.3% each) were only detected among E. faecium. A total of 133 (51.2%) strains were MDR, showing a large variety of MDR profiles, composed by simultaneous resistance encompassing 3 to 12 antimicrobials. MDR strains were found in 68.2% of the birds. Antimicrobial resistance was associated with the presence of the aac(6')-aph(2″)-Ia, aph(2″)-Id, ant(6)-Ia, ant(9)-Ia, ant(9)-Ib, tet(M), tet(L), tet(S), erm(B), mef(A/E), msrC, and vat(D) genes. The most common virulence genes were efaA, gelE, ace, eeP, and asa1. PFGE analysis revealed a large genetic diversity among most of the strains. MLST performed for 35 E.faecalis strains revealed 23 different STs, whereas 14 STs were found among 18 E.faecium strains. Hospital-associated lineages ST22, ST25, ST56, ST1274 were identified. The results show that the wild birds investigated can carry a diversity of potentially hazardous enterococcal strains displaying multiple antimicrobial resistance and virulence genes, reinforcing the assumption that these animals provide an important target to monitor the circulation of microorganisms that deserve consideration under the One Health perspective.

Experimental Application of Beneficial, Freeze-Dried Strain Enterococcus durans ED 26E/7 with Postbiotic Activity in Different Yogurts, Its Survival and Stability

Yogurt is generally defined as a cultured milk product made using some species of lactic acid bacteria. Moreover, some additive bacteria are frequently involved in yogurts to provide health benefits. The objective of this study was testing the stability and survival of a beneficial strain with postbiotic activity, Enterococcus durans ED 26E/7, in cow, goat, and ewe–goat milk yogurts. The validated methods were used in the study. Postbiotic, concentrated substance (CBs) from the strain ED 26E/7 inhibited growth of indicator bacteria by 60.5%. The strains E. hirae (96%) were susceptible to CBs (inhibitory activity from 200 to 25,600 AU/mL). The growth of staphylococci was inhibited by 79% with activity of 100 up to 25,600 AU/mL. Also, 40 out of 46 fecal E. coli were inhibited (activity 100 AU/mL). CBs was thermo-stable and remained active also after storage for 11 months at −20 °C and −80 °C. Exposing CBs to proteolytic enzymes did not lead to its complete deactivation indicating that it is probably not only a proteinaceous substance. The highest counts of the freeze-dried (encapsulated), safe ED 26E/7 strain and its stability were detected in ewe–goat milk yogurts. They reached up to 5.0 cfu/g. ED 26E/7 represents a further promising additive, although other testing will be performed.

Multicentre evaluation of in vitro activity of contezolid against drug-resistant Staphylococcus and Enterococcus.

To investigate susceptibility to contezolid, a novel oxazolidinone, multicentre surveillance was conducted involving 2449 strains of Staphylococcus and Enterococcus collected from 65 hospitals across China. The MICs of contezolid, linezolid and other clinically significant antibiotics were determined by the broth microdilution method. Consistency with the broth microdilution method for contezolid was assessed using agar dilution method, as well as disc diffusion and ETEST for linezolid, respectively. WGS was conducted on all 20 linezolid-resistant and 30 randomly non-resistant strains to analyse linezolid resistance genes (optrA, poxtA, cfr) and 23S rRNA mutation sites. All strains exhibited WT susceptibility to contezolid, while resistance proportions to daptomycin, vancomycin, teicoplanin, tigecycline and eravacycline ranged from 0% to 5.2% in Staphylococcus, and from 0% to 7.8% in Enterococcus. Linezolid resistance was higher in Enterococcus faecalis (4.4%) compared with Enterococcus faecium (0.2%). Contezolid showed a lower MIC50 (0.5 mg/L) than linezolid (2 mg/L) for methicillin-resistant Staphylococcus. Against Enterococcus, contezolid demonstrated a cumulative MIC percentage of 70% for VRE and 39.1% for E. faecalis (at MIC = 1 mg/L), whereas linezolid showed 0% and 1.1%, respectively. Among the 20 linezolid-resistant Enterococcus strains, all carried the optrA gene without 23S rRNA mutations. For contezolid, MICs were 4 mg/L for 19 strains and 2 mg/L for 1 strain. The ETEST, agar dilution and disc diffusion methods showed essential and categorical agreements of >90% for linezolid, with no major errors or very major errors. Contezolid demonstrated significant in vitro antibacterial activity against methicillin-resistant Staphylococcus, VRE and linezolid-resistant E. faecalis.

Optimization of exopolysaccharide production from the novel Enterococcus species, using statistical design of experiment

The Exopolysaccharide (EPS) producing novel strains of Enterococcus previously isolated from the vaginal source of pregnant women were selected based on ropy structure formation. The two selected strains, E.villorum SB-2 and E.rivorum S22-3, were found to be producing 2.87 g/l and 3.14 g/l EPS, respectively, in the minimal media (M17 media) after 24-hour fermentation under anaerobic condition. Both the strains have probiotic properties and have the potential to be used for industrial applications. The production media and fermentation conditions were optimized to enhance the EPS production using the one-factor method, Placket-Burman factorial designing and Central composite design (CCD) of Response surface methodology (RSM). The most relevant factors affecting the EPS yield were sucrose, yeast extract and pH for E.villorum SB2 and sucrose, yeast extract and magnesium sulfate for the E.rivorum S22-3 as determined by Placket-Burman design, whose concentrations were further optimized using CCD. The optimized fermentation conditions gave the total EPS of 9.76 g/l (4 times the initial production) from E.villorum SB-2 and 7.74 g/l (2.5 times the initial production) from E.rivorum S22-3, respectively, after 36-hour incubation at 37 °C. These optimization studies might be helpful during scale-up process for the industrial scale production of these exopolysaccharide.

Year-round monitoring of antibiotic-resistant bacteria in pristine uppermost stream and estimation of pollution sources

Studies on the conditions and pollution routes of antibiotic-resistant bacteria (ARB) in rivers can help provide countermeasures against the spread of ARB. This study focused on the pristine uppermost stream of a river, where Escherichia coli (E. coli) and enterococci were detected, although the stream flows through a pristine forest catchment. Antibiotic resistance of E. coli and enterococci isolated from the river water, riverbed sediment, and feces of waterside animals, such as birds and Mustelidae, were investigated throughout the year in the pristine uppermost sites. Antibiotic resistance was present in 1.4% (7/494) of the E. coli strains and 3.0% (24/812) of the enterococcal strains, and was low throughout the year. Although antibiotic resistance of bacteria isolated from feces was not detected in this watershed, the prevalence of multidrug-resistant E. coli was 0.4% (1/246) and 0.6% (1/172) in river water and riverbed sediment samples, respectively were observed. The presence of extended-spectrum β-lactamase (ESBL)-producing E. coli was confirmed in river water samples, and genomic analysis revealed that the samples possessed the CTX-M-15 group. Multidrug-resistant strains and ESBL-producing strains were classified as phylogroups B1 and A, respectively, which are E. coli phenotypes isolated from wild animals. Pulsed-field gel electrophoresis revealed analysis targeting enterococci that strains isolated from river water and bird feces were in the same cluster with 100% similarity. Therefore, bird feces are a source of enterococci in the uppermost stream of the river. Because multidrug-resistant bacteria and ESBL-producing bacteria were present in the pristine uppermost stream of the pristine river, urgent elucidation of the spreading routes of ARB is important.

An enterococcal phage protein inhibits type IV restriction enzymes involved in antiphage defense

The prevalence of multidrug resistant (MDR) bacterial infections continues to rise as the development of antibiotics needed to combat these infections remains stagnant. MDR enterococci are a major contributor to this crisis. A potential therapeutic approach for combating MDR enterococci is bacteriophage (phage) therapy, which uses lytic viruses to infect and kill pathogenic bacteria. While phages that lyse some strains of MDR enterococci have been identified, other strains display high levels of resistance and the mechanisms underlying this resistance are poorly defined. Here, we use a CRISPR interference (CRISPRi) screen to identify a genetic locus found on a mobilizable plasmid from Enterococcus faecalis involved in phage resistance. This locus encodes a putative serine recombinase followed by a Type IV restriction enzyme (TIV-RE) that we show restricts the replication of phage phi47 in vancomycin-resistant E. faecalis. We further find that phi47 evolves to overcome restriction by acquiring a missense mutation in a TIV-RE inhibitor protein. We show that this inhibitor, termed type IV restriction inhibiting factor A (tifA), binds and inactivates diverse TIV-REs. Overall, our findings advance our understanding of phage defense in drug-resistant E. faecalis and provide mechanistic insight into how phages evolve to overcome antiphage defense systems.