Faecium Isolates Research Articles

The presence of Enterococcus faecalis in saliva as a risk factor for endodontic infection.

The aim of the present study was to investigate and correlate the prevalence of Enterococcus faecalis in saliva and in root canals with different pulpal and periapical conditions. Sixty-seven patients were divided into five groups based on pulpaland periapical tissue status: healthy vital teeth (HVT, n=7), healthy treated teeth without lesion (HTT, n=9), irreversible pulpitis (IP, n=13), necrosis (N, n=18), and post-treatment apical periodontitis (PTAP, n=20). Saliva, rubber dam, sterilitycontrol and pre-treatment root canal samples were collected and microbiologically processed by culture method. The phylogenetic relationship of E. faecalis isolates collected from root canals and saliva were investigated by whole genome sequencing. Fisher's exact test was used to correlate the presence of E. faecalis in root canals or saliva with clinical and/or radiographic findings. Linear/logistic regression analyses were performed to establish the relationship between the presence of E. faecalis in root canals, saliva, and the status of periapical tissues. E. faecalis was found in 18 root canal and saliva samples. E. faecalis root canal isolates were recovered with the highest frequency from post-treatment apical periodontitis. The occurrence of E. faecalis in saliva was strongly associated with its detection in the root canals (P < 0.001). The pretreatment presence of E. faecalis in root canals was associated with significantly higher odds of having periapical lesions (OR=11.03; 95% CI, 1.27-95.70; p < 0.05). Saliva and root canal isolates from the same patient were highly correlated at the phylogenetic level (Jaccard index >0.95). This pilot study confirms the role of E. faecalis in developing peri-radicular lesions in secondary endodontic infections and suggests that saliva could be the main source of infection. Further studies are needed to investigate the exact origin of this bacteria and its true role in the pathogenesis of secondary/persistent endodontic infections.

Prevalence and Antibiotic Resistance of ESKAPE Pathogens Isolated from Patients with Bacteremia in Tehran, Iran

Introduction: The ESKAPE acronym refers to a group of deadly hospital-acquired pathogens, including Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. These organisms can evade antibacterial drugs and pose a significant threat to public health. This study investigated the prevalence and antimicrobial resistance of ESKAPE pathogens in patients with bacteremia. Methods: We collected 412 blood samples from patients diagnosed with bacteremia. The ESKAPE isolates were subjected to multidrug-resistant (MDR), extensively drug-resistant (XDR), extended-spectrum beta-lactamase (ESBL), metallo-β-lactamase (MBL), methicillin-resistant S. aureus (MRSA), and vancomycin-resistant Enterococcus (VRE) by the disk diffusion and E-test methods. In the case of VRE, molecular detection was done for vanA and vanB genes. Results: A total of 270 ESKAPE isolates were identified. The frequency of S. aureus was 30%, A. baumannii 22%, P. aeruginosa 17%, K. pneumoniae 13%, E. aerogenes 10.3%, and E. faecium 7.7%. MRSA was 71% and VRE was 19%. ESBL- and MBL-producing strains of A. baumannii were found to account for 39%, P. aeruginosa for 35.7% and 28.2%, and K. pneumoniae for 25.7% and 17.1%. In total, MDR and XDR were present in 52.2% and 15.5% of isolates and were most prevalent in E. aerogenes, A. baumannii, and P. aeruginosa. The vanA gene was detected in all 19% of E. faecium isolates that were VRE. Conclusions: Antibiotic resistance is widespread among ESKAPE pathogens, particularly in patients with bacteremia. Health-care professionals must consider the increasing rates of antibiotic resistance among ESKAPE pathogens and implement new measures to control infections.

Top-Down Genomic Surveillance Approach To Investigate the Genomic Epidemiology and Antibiotic Resistance Patterns of Enterococcus faecium Detected in Cancer Patients in Arkansas.

Control of hospital-associated Enterococcus faecium infection is a strenuous task due to the difficulty of identifying transmission routes and the persistence of this nosocomial pathogen despite the implementation of infection control measures that have been successful with other important nosocomial pathogens. This study provides a comprehensive analysis of over 100 E. faecium isolates collected from 66 cancer patients at the University of Arkansas for Medical Sciences (UAMS) between June 2018 and May 2019. In the top-down approach used in this study, we employed, in addition to the 106 E. faecium UAMS isolates, a filtered set of 2,167 E. faecium strains from the GenBank database to assess the current population structure of E. faecium species and, consequently, to identify the lineages associated with our clinical isolates. We then evaluated the antibiotic resistance and virulence profiles of hospital-associated strains from the species pool, focusing on antibiotics of last resort, to establish an updated classification of high-risk and multidrug-resistant nosocomial clones. Further investigation of the clinical isolates collected from UAMS patients using whole-genome sequencing analytical methodologies (core genome multilocus sequence typing [cgMLST], core single nucleotide polymorphism [coreSNP] analysis, and phylogenomics), with the addition of patient epidemiological data, revealed a polyclonal outbreak of three sequence types occurring simultaneously in different patient wards. The integration of genomic and epidemiological data collected from the patients increased our understanding of the relationships and transmission dynamics of the E. faecium isolates. Our study provides new insights into genomic surveillance of E. faecium to assist in monitoring and further limiting the spread of multidrug-resistant E. faecium. IMPORTANCE Enterococcus faecium is a member of the gastrointestinal microbiota. Although its virulence is low in healthy, immunocompetent individuals, E. faecium has become the third leading cause of health care-associated infections in the United States. This study provides a comprehensive analysis of over 100 E. faecium isolates collected from cancer patients at the University of Arkansas for Medical Sciences (UAMS). We employed a top-down analytical approach (from population genomics to molecular biology) to classify our clinical isolates into their genetic lineages and thoroughly evaluate their antibiotic resistance and virulence profiles. The addition of patient epidemiological data to the whole-genome sequencing analytical methodologies performed in the study allowed us to increase our understanding of the relationships and transmission dynamics of the E. faecium isolates. This study provides new insights into genomic surveillance of E. faecium to help monitor and further limit the spread of multidrug-resistant E. faecium.

China antimicrobial resistance surveillance network for pets (CARPet), 2018 to 2021

China Antimicrobial Resistance Surveillance Network for Pets (CARPet) was established in 2021 to monitor the resistance profiles of clinical bacterial pathogens from companion animals. From 2018 to 2021, we recovered and tested 4,541 isolates from dogs and cats across 25 Chinese provinces, with Escherichia coli (18.5%) and Staphylococcus pseudintermedius (17.8%) being the most predominant bacterial species. The Enterobacterales were highly susceptible to tigecycline, meropenem, colistin, and amikacin (70.3%–100.0%), but showed moderate resistance to ampicillin, ceftriaxone, doxycycline, florfenicol, levofloxacin, enrofloxacin, and trimethoprim-sulfamethoxazole (29.3%–56.7%). About 66.3% of Acinetobacter spp. were resistant to florfenicol, with relatively low resistance to another 11 antibiotics (1.2%–23.3%). The Pseudomonas spp. showed high susceptibility to colistin (91.7%) and meropenem (88.3%). The coagulase-positive Staphylococcus spp. showed higher resistance rates to most antimicrobial agents than coagulase-negative Staphylococcus isolates. However, over 90.0% of Staphylococcus spp. were susceptible to linezolid, daptomycin and rifampin, and no vancomycin-resistant isolates were detected. E. faecium isolates demonstrated higher resistance rates to most antimicrobial agents than E. faecalis isolates. Streptococcus spp. isolates showed low resistance to most antimicrobial agents except for doxycycline (78.2%) and azithromycin (68.8%). Overall, the tested clinical isolates showed high rates of resistance to commonly used antimicrobial agents in companion animals. Therefore, it is crucial to strengthen the monitoring of bacterial resistance in pets. By timely and effectively collecting, analyzing, and reporting antimicrobial resistance dynamics in pets, the CARPet network will become a powerful platform to provide scientific guidance for both pet medical care and public health.

Determination of the Prevalence and Antimicrobial Resistance of Enterococcus faecalis and Enterococcus faecium Associated with Poultry in Four Districts in Zambia.

The presence of antimicrobial-resistant Enterococci in poultry is a growing public health concern worldwide due to its potential for transmission to humans. The aim of this study was to determine the prevalence and patterns of antimicrobial resistance and to detect drug-resistant genes in Enterococcus faecalis and E. faecium in poultry from four districts in Zambia. Identification of Enterococci was conducted using phenotypic methods. Antimicrobial resistance was determined using the disc diffusion method and antimicrobial resistance genes were detected using polymerase chain reaction and gene-specific primers. The overall prevalence of Enterococci was 31.1% (153/492, 95% CI: 27.1-35.4). Enterococcus faecalis had a significantly higher prevalence at 37.9% (58/153, 95% CI: 30.3-46.1) compared with E. faecium, which had a prevalence of 10.5% (16/153, 95% CI: 6.3-16.7). Most of the E. faecalis and E. faecium isolates were resistant to tetracycline (66/74, 89.2%) and ampicillin and erythromycin (51/74, 68.9%). The majority of isolates were susceptible to vancomycin (72/74, 97.3%). The results show that poultry are a potential source of multidrug-resistant E. faecalis and E. faecium strains, which can be transmitted to humans. Resistance genes in the Enterococcus species can also be transmitted to pathogenic bacteria if they colonize the same poultry, thus threatening the safety of poultry production, leading to significant public health concerns.

Prevalence, serotype, and antimicrobial resistance profiles of children infected with Salmonella in Guangzhou, southern China, 2016-2021.

Salmonella infection is a key global public health concern and has lead to an increased economic burden on society. We investigated the epidemiological characteristics and antimicrobial resistance profiles of clinically isolated Salmonella strains in Guangzhou Women and Children's Medical Center. This was a retrospective study of 1,338 Salmonella strains collected from children in Guangzhou Women and Children's Medical Center during 2016 to 2021. The results revealed that 1,338 cases of Salmonella were mainly isolated from feces and blood samples. The age distribution was dominated by infants under 3 years old. The seasonal distribution was high in summer and autumn. 48 serotypes were detected, and S. typhimurium (78.7%) was the predominant serogroup. The results of antimicrobial susceptibility showed that the highest resistance was observed in ampicillin (84.5%), while lower resistance was observed in piperacillin/tazobactam, cefoperazone/sulbactam and ciprofloxacin. The antimicrobial resistance rate of fecal isolates was higher than that of blood isolates. The five-year average detection rate of multi-drug resistant Salmonella was 8.5% (114/1338) and the MDR rate of S. typhimurium was the lowest (6.9%; 73/1053). We concluded that antibacterial treatment should be carefully selected according to serotype and antimicrobial sensitivity results in children. Antimicrobial resistance monitoring for multi-drug resistant Salmonella is still required.

Nasotracheal enterococcal carriage and resistomes: detection of optrA-, poxtA- and cfrD-carrying strains in migratory birds, livestock, pets, and in-contact humans in Spain

This study determined the carriage rates and antimicrobial resistance (AMR) genes of enterococci from nasotracheal samples of three healthy animal species and in-contact humans. Nasal samples were collected from 27 dog-owning households (34 dogs, 41 humans) and 4 pig-farms (40 pigs, 10 pig-farmers), and they were processed for enterococci recovery (MALDI-TOF–MS identification). Also, a collection of 144 enterococci previously recovered of tracheal/nasal samples from 87 white stork nestlings were characterized. The AMR phenotypes were determined in all enterococci and AMR genes were studied by PCR/sequencing. MultiLocus-Sequence-Typing was performed for selected isolates. About 72.5% and 60% of the pigs and pig-farmers, and 29.4% and 4.9%, of healthy dogs and owners were enterococci nasal carriers, respectively. In storks, 43.5% of tracheal and 69.2% of nasal samples had enterococci carriages. Enterococci carrying multidrug-resistance phenotype was identified in 72.5%/40.0%/50.0%/23.5%/1.1% of pigs/pig-farmers/dogs/dogs’ owners/storks, respectively. Of special relevance was the detection of linezolid-resistant enterococci (LRE) in (a) 33.3% of pigs (E. faecalis-carrying optrA and/or cfrD of ST59, ST330 or ST474 lineages; E. casseliflavus-carrying optrA and cfrD); (b) 10% of pig farmers (E. faecalis-ST330-carrying optrA); (c) 2.9% of dogs (E. faecalis-ST585-carrying optrA); and (d) 1.7% of storks (E. faecium-ST1736-carrying poxtA). The fexA gene was found in all optrA-positive E. faecalis and E. casseliflavus isolates, while fexB was detected in the poxtA-positive E. faecium isolate. The enterococci diversity and AMR rates from the four hosts reflect differences in antimicrobial selection pressure. The detection of LRE carrying acquired and transferable genes in all the hosts emphasizes the need to monitor LRE using a One-Health approach.

Fluoroquinolone resistance among fecal extended spectrum βeta lactamases positive Enterobacterales isolates from children in Dar es Salaam, Tanzania

BackgroundFluoroquinolones have been, and continue to be, routinely used for treatment of many bacterial infections. In recent years, most parts of the world have reported an increasing trend of fluoroquinolone resistant (FQR) Gram-negative bacteria.MethodsA cross-sectional study was conducted between March 2017 and July 2018 among children admitted due to fever to referral hospitals in Dar es Salaam, Tanzania. Rectal swabs were used to screen for carriage of extended-spectrum β-lactamase-producing Enterobacterales (ESBL-PE). ESBL-PE isolates were tested for quinolone resistance by disk diffusion method. Randomly selected fluroquinolone resistant isolates were characterized by using whole genome sequencing.ResultsA total of 142 ESBL-PE archived isolates were tested for fluoroquinolone resistance. Overall phenotypic resistance to ciprofloxacin, levofloxacin and moxifloxacin was found in 68% (97/142). The highest resistance rate was seen among Citrobacterspp. (100%, 5/5), followed by Klebsiella.pneumoniae (76.1%; 35/46), Escherichiacoli (65.6%; 42/64) and Enterobacter spp. (31.9%; 15/47). Whole genome sequencing (WGS) was performed on 42 fluoroquinolone resistant-ESBL producing isolates and revealed that 38/42; or 90.5%, of the isolates carried one or more plasmid mediated quinolone resistance (PMQR) genes. The most frequent PMQR genes were aac(6’)-lb-cr (74%; 31/42), followed by qnrB1 (40%; 17/42), oqx,qnrB6 and qnS1. Chromosomal mutations in gyrA, parC and parE were detected among 19/42 isolates, and all were in E.coli. Most of the E. coli isolates (17/20) had high MIC values of > 32 µg/ml for fluoroquinolones. In these strains, multiple chromosomal mutations were detected, and all except three strains had additional PMQR genes. Sequence types, ST131 and ST617 predominated among E.coli isolates, while ST607 was more common out of 12 sequence types detected among the K.pneumoniae. Fluoroquinolone resistance genes were mostly associated with the IncF plasmids.ConclusionThe ESBL-PE isolates showed high rates of phenotypic resistance towards fluoroquinolones likely mediated by both chromosomal mutations and PMQR genes. Chromosomal mutations with or without the presence of PMQR were associated with high MIC values in these bacteria strains. We also found a diversity of PMQR genes, sequence types, virulence genes, and plasmid located antimicrobial resistance (AMR) genes towards other antimicrobial agents.

Occult Vancomycin-Resistant Enterococcus faecium ST117 Displaying a Highly Mutated vanB2 Operon

Rare information is available on clinical Enterococcus faecium encountered in Sardinia, Italy. This study investigated the antimicrobial susceptibility profiles and genotypic characteristics of E. faecium isolated at the University Hospital of Sassari, Italy, using the Vitek2 system and PCR, MLST, or WGS. Vitek2 revealed two VanB-type vancomycin-resistant Enterococcus faecium (VREfm) isolates (MICs mg/L = 8 and ≥32) but failed to detect vancomycin resistance in one isolate (MIC mg/L ≤ 1) despite positive genotypic confirmation of vanB gene, which proved to be vancomycin resistant by additional phenotypic methods (MICs mg/L = 8). This vanB isolate was able to increase its vancomycin MIC after exposure to vancomycin, unlike the "classic" occult vanB-carrying E. faecium, becoming detectable by Vitek 2 (MICs mg/L ≥ 32). All three E. faecium had highly mutated vanB2 operons, as part of a chromosomally integrated Tn1549 transposon, with common missense mutations in VanH and VanB2 resistance proteins and specific missense mutations in the VanW accessory protein. There were additional missense mutations in VanS, VanH, and VanB proteins in the vanB2-carrying VREfm isolates compared to Vitek2. The molecular typing revealed a polyclonal hospital-associated E. faecium population from Clade A1, and that vanB2-VREfm, and nearly half of vancomycin-susceptible E. faecium (VSEfm) analyzed, belonged to ST117. Based on core genome-MLST, ST117 strains had different clonal types (CT), excluding nosocomial transmission of specific CT. Detecting vanB2-carrying VREfm isolates by Vitek2 may be problematic, and alternative methods are needed to prevent therapeutic failure and spread.

Draft Genome Sequence of a Preterm Infant-Derived Isolate of Candida parapsilosis.

Candida parapsilosis is a human fungal pathogen of increasing incidence and causes invasive candidiasis, notably in preterm or low-birthweight neonates. Here, we present the genome sequence of C. parapsilosis NCYC 4289, a fecal isolate from a preterm male infant.

Epidemiology and source of infection in cancer patients with febrile neutropenia: an experience from a developing country

BackgroundFebrile neutropenia (FN) is a life-threatening complication that predisposes cancer patients to serious infections. This study aims to describe the epidemiology and source of infection in cancer patients with FN in a tertiary care hospital.MethodsA hospital-based retrospective study was conducted in a large tertiary care hospital from January 2020 to December 2021. Data on cancer patients with FN were collected from the hospital information system.Results150 cancer patients with FN were identified during the study period. Most patients were males (98; 65.3%), and the mean age of participants was 42.2 ± 16.0 years. Most patients (127; 84.7%) had hematologic malignancies, and acute myeloid leukemia was the most common diagnosis (42; 28%), followed by acute lymphocytic leukemia (28; 18.7%) and Hodgkin’s lymphoma (20; 13.3%). Fifty-four (36%) patients had a median Multinational Association for Supportive Care in Cancer (MASCC) scores greater than 21. Regarding the outcome, nine (6%) died, and 141(94%) were discharged. The focus of fever was unknown in most patients (108; 72%). Among the known origins of fever were colitis (12; 8%), pneumonia (8; 5.3%), cellulitis (6; 4%), bloodstream infections (7; 4.6%), perianal abscess (2; 1.3%) and others. The median duration of fever was two days, and the median duration of neutropenia was seven days. Sixty-three (42%) patients had infections: 56 (73.3%) were bacterial, four (2.6%) were viral, two (1%) were fungal and 1 (0.7%) was parasitic. Among the bacterial causes, 50 cases (89.2%) were culture-positive. Among the culture-positive cases, 34 (68%) were gram-positive and 22 (44%) were gram-negative. The most frequent gram-positive bacteria were E. faecalis (9; 18% of culture-positive cases), and the most frequent gram-negative organisms were Klebsiella pneumoniae (5; 10%). Levofloxacin was the most commonly used prophylactic antibiotic (23; 15.33%), followed by acyclovir (1610.7%) and fluconazole in 15 patients (10%). Amikacin was the most popular empiric therapy, followed by piperacillin/tazobactam (74; 49.3%), ceftazidime (70; 46.7%), and vancomycin (63; 42%). One-third of E. faecalis isolates were resistant to ampicillin. Approximately two-thirds of Klebsiella pneumoniae isolates were resistant to piperacillin/tazobactam and ceftazidime. Amikacin resistance was proven in 20% of isolates.ConclusionsThe majority of patients suffered from hematologic malignancies. Less than half of the patients had infections, and the majority were bacterial. Gram-positive bacteria comprised two-thirds of cases. Therefore, empiric therapy was appropriate and in accordance with the antibiogram of the isolated bacteria.

Whole Genome Sequencing of Escherichia coli and Enterococcus spp. in wildlife-livestock interface: a pilot study

ObjectivesThis pilot study provides a multidisciplinary investigation to monitor livestock-wildlife interface. Ecological data, microbiological investigations, and whole genome sequencing were used to characterize eight bacterial isolates obtained from sympatric domestic and wild ruminants in Maiella National Park (Italy) in terms of genetic patterns of antimicrobial resistance. MethodsUsing selective culturing of fresh fecal samples of monitored and georeferenced populations of Apennine chamois, goats, red deer, and sheep, Escherichia coli, Enterococcus faecium, and Enterococcus faecalis isolates were isolated and subjected to minimum inhibitory concentration determination and whole genome sequencing. ResultsThe analyzed isolates showed phenotypic and genotypic resistance to tetracycline and critically important antibiotics such as linezolid and carbapenems. Virulence genes related to biofilm regulation and Shiga toxins were also detected. Furthermore, serotypes related to nosocomial infections, harbouring plasmids recognized as important mobile resistance gene transmitters, were identified. ConclusionsThis multidisciplinary pilot study represents a promising initial step to identify the environmental drivers and the transmission routes of antimicrobial resistance and virulence factors, providing new data on bacteria from rare and endangered species such as Apennine chamois.

Techno-functional characterization of fecal lactobacilli isolates of Bos indicus calves for probiotic properties.

In this study, 105 bacterial colonies were isolated from the feces of newborn healthy Bos indicus calves and 37 isolates were confirmed using morphological, biochemical tests, and genus-specific PCR as lactobacilli. 11 isolates were then short-listed for in vitro probiotic testing based on their ability to dwell under acid and bile stress. Species-level identification using 16S rRNA gene sequencing revealed that they were Ligilactobacillus salivarius. These isolates flourished in 0.4% phenol, depicting resistance in adverse conditions encountered in the gastrointestinal tract. The results of cell surface hydrophobicity were found to be 74.50% for RBL12 and 62.62% for RBL09 in hexadecane and xylene, respectively, and that of auto-aggregation was highest in RBL26 (58.92%). These isolates also produced digestive enzymes like amylase, protease, and β-galactosidase. Further assays reiterated their antimicrobial and coaggregation potential against diarrhea-causing pathogens like Escherichia coli ATCC-25922 and Salmonella arizonae ATCC-13314. Biosafety assessment revealed that none of the tested isolates were hemolytic and mucinolytic in nature. Furthermore, the antioxidant potential of the isolates was also confirmed using 1,1‑diphenyl‑2‑picrylhydrazyl (DPPH) and ferric ion-reducing antioxidant power (FRAP) assay. Along with efficient utilization of inulin, isolates showed promising adhesion ability to the HT-29 cell line. The current findings hence conclude that these Lactobacillus isolates can be exploited as animal probiotics for potentialapplication in young calves to foster gut health and immunity.

Virulence gene detection and antimicrobial resistance analysis of Enterococcus faecium in captive giant pandas (Ailuropoda melanoleuca) in China

BackgroundThe emergence of multidrug resistance among enterococci makes effective treatment of enterococcal infections more challenging. Giant pandas (Ailuropoda melanoleuca) are vulnerable to oral trauma and lesions as they feast on bamboo. Enterococci may contaminate such oral lesions and cause infection necessitating treatment with antibiotics. However, few studies have focused on the virulence and drug resistance of oral-derived enterococci, including Enterococcus faecium, in giant pandas. In this study, we analyzed the prevalence of 8 virulence genes and 14 drug resistance genes in E. faecium isolates isolated from saliva samples of giant pandas held in captivity in China and examined the antimicrobial drug susceptibility patterns of the E. faecium isolates.ResultsTwenty-eight isolates of E. faecium were successfully isolated from the saliva samples. Four virulence genes were detected, with the acm gene showing the highest prevalence (89%). The cylA, cpd, esp, and hyl genes were not detected. The isolated E. faecium isolates possessed strong resistance to a variety of drugs; however, they were sensitive to high concentrations of aminoglycosides. The resistance rates to vancomycin, linezolid, and nitrofurantoin were higher than those previously revealed by similar studies in China and other countries.ConclusionsThe findings of the present study indicate the drugs of choice for treatment of oral E. faecium infection in the giant panda.

Prevalence of Class 1 Integron and In Vitro Effect of Antibiotic Combinations of Multidrug-Resistant Enterococcus Species Recovered from the Aquatic Environment in the Eastern Cape Province, South Africa.

Enterococci are regarded as a better indication of faecal pollution in freshwater and marine waters. Their levels in seawater are positively connected with swimming-related gastrointestinal disorders. This study used an Enterococcus-specific polymerase chain reaction (PCR) to characterize the isolates. Classes 1 and 2 integrons were examined for environmental Enterococcus isolates using a standard biological procedure. All strains were assessed against a panel of 12 antibiotics from various classes using disc diffusion methods. The microdilution method was used to work out the minimum inhibitory concentration (MIC) according to the CLSI guiding principles. The combination therapy of the resistant drugs was evaluated using a checkerboard assay and a time-dependent test for assessing their bactericidal or bacteriostatic activity. The gene diversity of the tested organisms was analyzed with the aid of Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR. In total, 57 Enterococcus spp. environmental samples were recovered, in which Enterococcus faecalis (33.33%) and Enterococcus faecium (59.65%) were the dominant species. Resistance to linezolid, ciprofloxacin, erythromycin, gentamicin, vancomycin, rifampicin, and tetracycline was prevalent. Fifty (50) strains tested positive for class 1 integron, more frequent in Enterococcus faecium and Enterococcus faecalis isolates, with no gene cassette array discovered. A combination of gentamicin (MIC 4 µg/mL) with vancomycin (MIC 256 µg/mL) antibiotics against Enterococcus faecalis showed antibacterial activity. In contrast, the combination of ciprofloxacin (1 µg/mL) with Ampicillin (16 µg/mL) antibiotics against Enterococcus faecalis showed a bacteriostatic effect. The ERIC-PCR analysis pointed out that most of the assessed isolates have close genetic similarities.

Detection of Antibiotic Resistance Genes (CTX-M, Van A and Van B) of Enterococcus faecalis Isolated from Children with Bacteremia by RT-PCR.

Fever is one of the most common diseases affecting humans, as it results from any disease or development and worsening of the disease for most people with widespread infections in the body. Therefore, this study aimed to evaluate antibiotic resistance genes (CTX-M, Van A and Van B) of Enterococcus faecalis isolated from children with bacteremia by RT-PCR. A total of 200 children was enrolled in the study, 100 children with fever and 100 healthy children (not suffering from any problem); that is, they are a control group for the detection of antibiotic resistance genes (CTX-M, Van A and Van B) of Enterococcus faecalis by RT-PCR. The age of the two groups ranged from one to five years. Four ml of venous blood sample was collected from each child; the venipuncture area was sterilized first with alcohol at a rate of 70%, followed by medical iodine and then sterilized with alcohol again to avoid contamination with skin flora. The blood samples were cultured on media for isolating bacteria. Then, the resistant isolates of E. faecalis to Vancomycin and cefotaxime antibiotics were taken and kept in special nutrient agar media where the DNA of the bacteria was extracted using (Zymogene Extraction kit, Japan). The detection of the exact genes (CTX-M, Van A and Van B) was done using Real-Time PCR technology according to the protocol mentioned by the company (Sacace biotechnology, Italy). The study presented that 40% of children with fever have positive blood cultures compared with 5% in the control group, with a significant difference between the two groups (P<0.001). The study found that 32.5% of bacteremic children were due to S. aureus, 30%, 5%, and 4% were due to E. faecalis, E. coli, P. aeruginosa and Klebsiella spp, respectively, with significant difference (P<0.01). The study showed that 91.67% of E. faecalis isolates were sensitive to Levofloxacin, 83.33% to Amoxiclav, 66.67% to Erythromycin, 58.33% to Amikacin, 50% to Ampicillin, 33.33% to cefotaxime and Ceftriaxone and 25% toward Vancomycin. From 9 isolates resistant to Vancomycin, the study presented that 88.89% of them were observed with Van A gene production as detected by real-time PCR (P<0.001). The study also showed that 77.78% were observed with Van B gene production as detected by real-time PCR (P<0.001). The study revealed that all E. faecalis isolates resistant to cefotaxime and Ceftriaxone were characterized by CTX gene production as detected by real-time PCR (P<0.001).

Genomic Characterization of a Vancomycin-Resistant Strain of Enterococcus faecium Harboring a rep2 Plasmid.

In China, vancomycin-resistant enterococci (VRE) was not a common occurrence, and research on the genetic context and transmission mechanism of vanA-plasmid was scarce. The aim of this study was to molecularly characterise a vancomycin-resistant Enterococcus faecium isolate from a bloodstream infection and determine the genetic environment and delivery pattern of the plasmid carrying vancomycin-resistant gene. In May 2022, a vancomycin-resistant strain of Enterococci was identified during routine screening for VRE bacteria at the First Affiliated Hospital, Zhejiang University School of Medicine. Utilizing matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), the isolate was accurately identified. Antimicrobial susceptibility and whole-genome sequencing (WGS) were employed to perform phenotypic and genomic analysis, respectively. Further bioinformatics analyses was carried out to characterize the vanA-bearing plasmid. The antimicrobial susceptibility test showed that SJ2 strain was resistant to multiple antimicrobials, including ampicillin, benzylpenicillin, ciprofloxacin, erythromycin, levofloxacin, streptomycin, and vancomycin. Whole-genome analysis revealed that SJ2 strain carries several antimicrobial resistance genes and virulence determinants. MLST analysis found that SJ2 strain belongs to an unknown ST type. Plasmid analysis confirmed that the vanA gene was located on a variant of ~50 kb rep2 plasmid. Our study found that vanA-bearing rep2 plasmid is a potential source of dissemination and outbreak, and continuous surveillance is necessary to control its spread in Hangzhou, China.

ANTIMICROBIAL ACTION OF PUNICA GRANATUM (POMEGRANATE) AND PASSIFLORA EDULIS (PASSION FRUIT) AGAINST ORGANISMS ISOLATED FROM INTESTINAL INFECTIONS –AN IN VITRO STUDY

Background and objectives: The overuse and misuse of antibiotics coupled with the intrinsic and acquired resistance of bacteria resulted in the emergence of multi-drug resistant strains. Pomegranate and Passion fruit are two fruits which have been documented to have signicant antimicrobial activity against harmful organisms which colonize the gut. The present study evaluates the antimicrobial activity of Punica granatum and Passiora edulis against organisms isolated from intestinal infections and compares the antimicrobial efcacy of the ethanolic and aqueous extracts of fruit rind of Punica granatum and leaves of Passiora edulis. Fruit ri Methods: nd of Punica granatum and the fresh leaves of Passiora edulis plants were dried, ground into powder and made into extracts. A total of 100 gastrointestinal pathogens were isolated and included in the study. The antibacterial efcacy of the aqueous and alcoholic extracts against 100 human faecal isolates was tested by employing Kirby Bauer disc diffusion technique. Sensitivity to the ethanolic and aqueous extracts of Punica Results: granatum were found to be maximum in Staphylococcus aureus. MRSA was sensitive to these extracts. Aqueous extract of Punica granatum were effective against a large number of organisms, however, better antimicrobial action was seen with the ethanolic extract of Punica granatum. No encouraging results were obtained with the extracts of Passiora edulis. Our study shows that puried forms of Pomegranate (Punica granat Interpretation and conclusions: um) can be used as an adjunct in the treatment, in conjunction with antibiotics against intestinal pathogens and can open new areas of research.

Penicillin-Binding Proteins and Alternative Dual-Beta-Lactam Combinations for Serious Enterococcus faecalis Infections with Elevated Penicillin MICs.

Ampicillin-ceftriaxone has become a first-line therapy for Enterococcus faecalis endocarditis. We characterized the penicillin-binding protein (PBP) profiles of various E. faecalis strains and tested for synergy to better inform beta-lactam options for the treatment of E. faecalis infections. We assessed the affinity of PBP2B from elevated-MIC strain E. faecalis LS4828 compared to type strain JH2-2 using the fluorescent beta-lactam Bocillin FL. We also characterized pbp4 and pbpA structures and PBP4 and PBP2B expression and used deletion and complementation studies to assess the impact of PBP2B on the levels of resistance. We tested penicillin-susceptible and -resistant E. faecalis isolates against ceftriaxone or ceftaroline combinations with other beta-lactams in 24-h time-kill studies. Two penicillin-susceptible strains (JH2-2 and L2052) had identical pbp sequences and similar PBP expression levels. One reduced-penicillin-susceptibility strain (L2068) had pbp sequences identical to those of the susceptible strains but expressed more PBP4. The second decreased-penicillin-susceptibility strain (LS4828) had amino acid substitutions in both PBP4 and PBP2B and expressed increased quantities of both proteins. PBP2B did not appear to contribute significantly to the elevated beta-lactam MICs. No synergy was demonstrable against the strains with both mutated PBPs and increased expression (L2068 and LS4828). Meropenem plus ceftriaxone or ertapenem plus ceftriaxone demonstrated the most consistent synergistic activity. PBP2B of strain LS4828 does not contribute significantly to reduced penicillin susceptibility. Neither the MIC nor the level of PBP expression correlated directly with the identified synergistic combinations when tested at static subinhibitory concentrations.

Enterococcus Species Isolated from Urine Cultures and Antibiotic Resistance Rates

While enterococci are found as flora elements in many parts of the body, they have recently emerged as different infectious agents such as urinary tract infections, bacteremia, and meningitis. Increasing antibiotic resistance also poses additional challenges in treatment. In this study, it was aimed to determine the species identification and antibiotic resistance rates of enterococci isolated from the urinary tract. Enterococcus strains isolated from urine culture samples sent to the routine microbiology laboratory from various clinics of our hospital between 01.01.2020 and 31.12.2021 with the preliminary diagnosis of urinary tract infection were included in this study. Samples were cultivated on sheep blood agar and eosin methylene blue agar, and species determination and antibiogram evaluation were carried out in accordance with the recommendations of EUCAST (European Committee on Antimicrobial Susceptibility Testing-2020-2021) using conventional methods and automated identification device (VITEK 2 Compact-BioMerieux-France). All enterococci isolates (n=349) were susceptible to vancomycin, teicoplanin, tigecycline and linezolid. Of Enterococcus faecalis isolates (n=238), 32.8% were resistant to ciprofloxacin, 23.7% to high-level gentamicin, 33.2% to imipenem, 2.5% to nitrofurantoin, and 26.9% to streptomycin. Of the Enterococcus faecium isolates (n=111), 84.7% were resistant to ciprofloxacin, 58.6% to high-level gentamicin, 91% to imipenem, 31.5% to nitrofurantoin, and 83.8% to streptomycin. In conclusion, it is important to determine the resistance patterns of enterococcus strains, which are most frequently isolated as the causative agent of urinary tract infection, and also cause other clinical presentations such as wound infection or bacteremia, and to establish appropriate treatment protocols and to determine new antibiotic resistances.