Resistant Enterococcus Species Research Articles

Enterococcus Species and Their Antimicrobial Resistance in an Urban Watershed Affected by Different Anthropogenic Sources

Different anthropogenic sources can have a significant influence on bacterial populations and their antimicrobial activities. In this study, the impact of anthropogenic activities on Enterococcus species was studied in an urban watershed in southern California affected by concentrated animal feeding operations (CAFOs), recreational activities, wastewater treatment plants (WWTPs), urban runoff, and control sites. Water samples were collected quarterly for two years for the enumeration of Enterococcus species based on the Enterolert most probable-number (MPN) assay. Concentrations of enterococci were higher in the sediment compared to surface water (4.5 × 106 CFU/g of sediment vs. 2.3 × 105 MPN/100 mL of water). The species diversity was dominated by E. mundtii (32%), E. faecalis (27%), and E. faecium (25%). E. faecium exhibited the highest antibiotic-resistant phenotype. Resistances were mostly to ciprofloxacin, erythromycin, and tetracycline. Tetracycline and erythromycin resistance genes, encoded by tet (C, K, O, S) and ermB, respectively, were more common in isolates from sediment (42.9%) compared to water (12.7%). E. mundtii was sensitive to ampicillin, chloramphenicol, gentamicin, and high levels of vancomycin. A significant percentage of E. faecalis were also resistant to these antibiotics. E. faecium and E. faecalis exhibited resistance to multiple antibiotics. Our data suggest that resistant Enterococcus species within the watershed might provide some useful data to determine pollutant types and sources in that watershed. Therefore, the widespread occurrence and abundance of E. faecium and E. faecalis, and their resistance genes associated with multiple antibiotics may potentially pose risks to the local populations exposed to these water sources during recreational activities.

A Retrospective Analysis of Culture-Confirmed Enterococci Bloodstream Infections in South Africa, 2016-2020: A Cross-Sectional Study.

(1) Background: The emergence of multidrug resistance enterococci is a major public health concern. This study aimed to determine the prevalence and antimicrobial resistance of enterococci isolated from blood cultures over a five-year period (2016−2020) at public hospitals in South Africa. (2): Methods: A retrospective analysis of clinical enterococci isolated from bloodstream infection samples at the South African public hospitals was conducted. The ESKAPE dataset from January 2016 to December 2020 was obtained from the central data warehouse (CDW) at the National Health Laboratory Service (NHLS). (3): Results: Following de-duplication, a total of 130,352/306,592 organisms isolated from blood cultures were identified as ESKAPE pathogens. In this study, K. pneumoniae (25%; 33,082/130,352), was the most frequently isolated pathogen from blood cultures, followed by S. aureus (23%; 29,922/130,352) and enterococci (16%; 21,339/130,352). Of the enterococci cases, about 43% (9132/21,339) of cases were from the infants aged (<1-year old) and 32% (6745/21,339) from the adult patients. No changes observed in vancomycin, teicoplanin, and linezolid susceptibility; however, E. faecium and E. faecalis blood culture isolates remained highly susceptible (>97%) to these antibiotics. (4): Conclusions: The current study revealed a significant increase of E. faecalis and E. faecium blood culture isolates as compared to the previous national ESKAPE data. Low vancomycin resistance was observed. Continuous monitoring of antimicrobial resistant Enterococcus species is warranted in South Africa.

Effects of age and pasture type on the concentration and prevalence of tetracycline and macrolide resistant Enterococcus species in beef cow-calf production system

Enterococci are a normal flora of the gastrointestinal tracts of humans and animals. Enterococci can also cause life-threatening nosocomial infections. Antimicrobial-resistant Enterococcus species have been reported in the feedlot and dairy cattle productions and in meat and milk products, suggesting their foodborne importance. Cow-calf operations represent a significant segment in the beef production system by producing weaned calves. Weaned calves are brought into the feedlot to be finished for meat, and culled cows are also slaughtered for beef, primarily for ground beef products. Infection dynamics in the cow-calf operation can contribute to meat contamination. This study evaluated the effects of age and wheat grazing on the concentration and prevalence of a macrolide antibiotic erythromycin (ERYr) and tetracycline (TETr) resistant enterococci, associated resistance genes and species distribution in a cow-calf production system. In 2017 and 2018, 32 Angus breed cow-calf pairs were randomly assigned to feed on tall fescue or wheat pasture in two independent field experiments. During the grazing experiments of 2-3 weeks, fecal samples were collected weekly and cultured to enumerate, isolate and identify ERYr, TETr, and generic enterococci, using media supplemented with erythromycin, tetracycline or non-supplemented media, respectively. The two main species frequently associated with human illnesses, Enterococcus faecium and E. faecalis, were widely distributed in the cow-calf groups. Generic and TETr- enterococci were prevalent (96-100% prevalence) and abundant (3.2-4.9 log10 CFU/g) in the cow-calf population; however, ERYr enterococci were enumerable by direct plating only from a single cow despite being detected in at least 40% of the fecal samples after enrichment, showing their low abundance. TET- and ERY-resistance were mainly conferred by tet(M) and erm(B), respectively. Wheat grazing reduced the concentration of TETr enterococci and modified enterococcal species and resistance gene distributions. Hence, it is necessary to further investigate wheat grazing in cow-calf production as a potential strategy to mitigate antimicrobial resistance.

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Enterococci, the normal inhabitant of human intestinal tract are now emerging as potential pathogens asssociated with healthcare infections despite their low virulence. The rise in antibiotic resistance has a profound impact on the treatment of infectious diseases and the consequences due to its complications. Stool samples were obtained from children under the age of 14 years, admitted to pediatrics ward of a tertiary care hospital. The identification of enterococcal isolates was followed by suitable biochemical assays, disc diffusion and the E test to assess antibiotic susceptibility and minimum inhibitory concentrations. Vancomycin (van A, van B) and gentamicin resistance genes (aac(6)-Ie-aph(2)-Ia) were detected using PCR. From the total of 300 stool samples, Enterococcus spp were isolated in 132, out of which 39 were E. faecalis and 93 were E. faecium. Highest percentage of resistance was noted against erythromycin (68%) followed by penicillin (48%) and high level gentamicin in (34%) of the isolates. Resistance to high level gentamicin by MIC was found to be 25%. The MIC of vancomycin ranged from 1.5 μg/ml - 4 μg/ml of which 16% of the isolates had an MIC of 4 μg/ml. Resistance gene for aac(6)-Ie-aph(2)-Ia(gentamicin resistance) was found in 25% of isolates and van A (Vancomycin resistance) was detected in 11% of the isolates and none of the isolates harbored van B gene. Feacal carriage of gentamicin resistant enterococcus species was high in pediatric inpatients. No resistance was seen for vancomycin.

Molecular screening the emergence of vancomycin and gentamycin resistant enterococcus species in burn' patients in basrah government, Iraq

Emergence of multidrug resistance bacteria (MDR) in burn infections including Enterococcus yet to be managed and highlighted in most of hospitals in our region. This study included 200 clinical samples collected from wound, urine, stool and blood of 75 burn patients in Al-Faiha Teaching Hospital in Basrah, Iraq, between 2020 -2021. Based on Enterococci chromo agar, biochemical test and PCR for 16S rRNA gene, 50 isolates were identified as Enterococcus spp., involving 20 (40%) E. faecium, 14 (28%) E. faecalis, 7 (14%) E. gallinarum, 6 (12%) E. gilvus, 2 (4%) E. casseliflavus and 1 (2%) E. avium. All detected Enterococci were reported worldly as pathogenic bacteria to human. Six new local strains of Enterococcus were recorded in NCBI and the Gene bank as follow; Enterococcus gallinarum strain IraqYaHa5 and IraqYaHa19, Enterococcus faecium strain IraqYaHa23, IraqYaHa48 and IraqYaHa60, Enterococcus gilvus strain IraqYaHa50. Phylogenetic tree was constructed to all isolated Enterococci. The clinical isolates showed resistance up to nine antibiotics. However, enterococci isolated from healthy people during this study were 100% sensitive to six of those antibiotics. Duplex PCR was applied to detect vancomycin and gentamicin resistance genes. All resistant Enterococcus to gentamicin had harboured aph (3’).

Emergence of Drug Resistant Enterococcus Species from Patients with Urinary Tract Infections in a Tertiary Care CentreA Retrospective Analysis

Emergence of Drug Resistant Enterococcus Species from Patients with Urinary Tract Infections in a Tertiary Care CentreA Retrospective Analysis

Emergence of Drug Resistant Enterococcus Species from Patients with Urinary Tract Infections in a Tertiary Care Centre- A Retrospective Analysis

Introduction: Urinary Tract Infection (UTI) is one of the most common infections, among patients who are hospitalised. Enterococci are one of the frequent isolates among UTI patients, gaining considerable clinical importance, due to their escalating drug resistance, affecting debilitated patients or patients with prolonged hospital stay. Among Multi-Drug Resistant (MDR) enterococci, glycopeptide-resistant enterococci are being increasingly reported. The intrinsic resistance of enterococci and their ability to acquire and disseminate antibiotic resistant genes to other organisms pose a challenge in the treatment of enterococcal infections with MDR. Aim: To investigate the prevalence of MDR, Extensively-Drug Resistant (XDR) and Pan-Drug Resistant (PDR) Enterococci spp. isolated from urine samples in a tertiary care centre. Materials and Methods: The present retrospective study was conducted to find the burden of drug resistant Enterococcus spp. in urine samples, analyse their antibiogram and patients’ socio-demographic information from January to December 2019. Records of microscopic observations to antibiogram of each isolate was noted down from the register for urine samples, and further analysed. The data was coded, verified, entered and analysed using Statistical Package for Social Sciences (SPSS) version 18.0. Results: The majority of patients 127 (20%) belonged to the age group of 21-30 years of age. Among 10,535 urine samples received, 635 culture isolates were identified as Enterococcus spp., of which 17.95%, 3.30% and 0.62% were identified as MDR, XDR and PDR enterococci, respectively. The antimicrobial susceptibility was found to be least for High Level Gentamicin (HLG) (66.14%), and penicillin (68.81%), followed by ampicillin (81.10%) and nitrofurantoin (86.40%). Twenty eight isolates (7.9%) were identified as Vancomycin Resistant Enterococci (VRE), 9 (2.5%) as linezolid resistant and 12 (3.4%) as linezolid intermediate enterococci. Conclusion: Identification and prevention of the alarming increase in MDR, XDR and PDR enterococci spp. is cardinal to prevent morbidity and mortality of affected patients. Analysis of antibiogram periodically, is important in selection of appropriate drugs to prevent the incoherent use of antibiotics.

High-level resistance to aminoglycosides and ampicillin among clinical isolates of Enterococcus species in an Iranian referral hospital.

Background and Objectives:Nowadays, high-level aminoglycosides and ampicillin resistant Enterococcus species are among the most common causes of nosocomial infections. The present study was conducted to determine the prevalence of high-level resistance to aminoglycosides and ampicillin among clinical isolates of Enterococcus species in Ardabil, Iran.Materials and Methods:In this cross–sectional study, a total of 111 Enterococcus species were collected from different clinical specimens between 2013 and 2015. Enterococcus species were identified using standard phenotypic and genotypic methods. BHI agar screen and agar dilution methods were used for detection of high-level gentamicin and streptomycin resistance (HLGR and HLSR) and minimal inhibitory concentration (MIC) of ampicillin, respectively.Results:Of 111 clinical isolates, 59 (53.2%) and 25 (22.5%) isolates were E. faecalis and E. faecium, respectively, based on the PCR results. Totally, 60.3% and 56.7% of isolates were HLGR and HLSR, respectively, as well as 51.35% were HLGR plus HLSR. Among HLGR isolates, 36 (61.01%), 18 (72%) and 13 (48.14%) were E. faecium, E. faecalis and non-faecalis non-faecium species, respectively. Among HLSR isolates, 33 (55.93%), 16 (64%) and 14 (51.85%) were E. faecalis, E. faecium and non-faecalis non-faecium species, respectively. All HLGR isolates contained aac(6′)Ie-aph(2″)Ia gene. Overall, the prevalence of high-level ampicillin resistance among Enterococcus species was 17.1%. For E. faecalis, E. faecium and non-faecalis non-faecium species, ampicillin resistance rates were as follows: 11 (40.74%), 7 (28%) and 1 (1.69%), respectively. For aminoglycoside antibiotics, the resistance rate was significantly higher in E. faecium isolates and for ampicillin it was higher in E. faecalis isolates.Conclusion:The frequency of high-level aminoglycoside resistant enterococcal isolates in our hospital was high and significant ampicillin resistance was noticed. This would require routine testing of enterococcal isolates for HLAR and ampicillin susceptibility.

Bystander Selection for Antimicrobial Resistance: Implications for Patient Health.

Antimicrobial therapy promotes resistance emergence in target infections and in off-target microbiota. Off-target resistance emergence threatens patient health when off-target populations are a source of future infections, as they are for many important drug-resistant pathogens. However, the health risks of antimicrobial exposure in off-target populations remain largely unquantified, making rational antibiotic stewardship challenging. Here, we discuss the contribution of bystander antimicrobial exposure to the resistance crisis, the implications for antimicrobial stewardship, and some novel opportunities to limit resistance evolution while treating target pathogens.

Identification, antimicrobial susceptibility and virulence factors of Enterococcus species isolated from clinical specimens at an Indian tertiary care hospital

Purpose: Enterococcus species have increasingly emerged as important pathogens in serious nosocomial and community acquired infections. The objective was to determine species identification, evaluate current susceptibility pattern and characterize various virulence determinants among clinical isolates of enterococci at an Indian hospital. Methods & Materials: Consecutive isolates from clinical specimens (April to September 2017) were included. Species identification was performed by conventional biochemical tests. Antimicrobial susceptibility testing was as per CLSI guidelines, including detection of high-level gentamicin resistance (HLGR), high-level streptomycin resistance (HLSR) and glycopeptide resistant enterococci (GRE). All screen-positive GRE isolates were investigated by PCR for species confirmation and presence of vanA/ vanB genes. Virulence genes asa1, esp, hyl, cyl, and gelE were investigated by molecular methods. Haemolysin and biofilm production were studied phenotypically. Results: Of 71 strains isolated, 57 (80.3%), 9 (12.7%) and 5 (7.0%) were from urine, pus and blood respectively consisting of E. faecalis (45, 63.4%), E. faecium (20, 28.2%), E. durans (3, 4.2%), and E. avium, E. hirae, E. mundtii (1, 1.4%). Six (8.4%) were GRE (5 E. faecium, 1 E. faecalis), all vanA phenotypically and genotypically. HLGR and HLSR was observed in 41 (57.7%) and 23 (32.4%) isolates respectively, higher in E. faecium compared to E. faecalis (85.0% and 40.0% vs 46.6% and 28.8%). Resistance to other antibiotics was also higher in E. faecium. All were susceptible to daptomycin. Hemolysin activity was significantly higher in E. faecalis compared to E. faecium (55.5% vs 0%). Biofilm production was observed in 23 (32.4%) isolates. Virulence genes detected were asa1 (9, 45%), gel (9, 45%), esp (7, 35%), cyt (6, 30%) and hyl (1, 5%). Frequency of asa, gel, and cyl was higher in E. faecalis than E. faecium (7, 7, 5 vs 2, 2, 1), whereas esp and hyl was more in E. faecium compared to E. faecalis (5, 1 vs 0, 0). Conclusion: Prevalence of a wide variety of Enterococcus species in clinical samples together with their variable antimicrobial susceptibility pattern and virulence traits emphasizes the need for their routine speciation and susceptibility testing which may help control the spread of multidrug resistant Enterococcus species.

Antibiotic Susceptibility Pattern and Virulence Genes in Enterococcus spp. Isolated From Clinical Samples of Milad Hospital of Tehran, Iran

Background: Enterococcus spp. are part of the normal flora of humans and animals. The nosocomial pathogenicity of Enterococcus spp. has emerged in recent years and has caused great concern due to developing of resistance to many antimicrobial agents. Objectives: The current study aimed to determine the resistance pattern and the type of virulence genes in Enterococcus spp. isolated from Milad hospital of Tehran, Iran. Materials and Methods: The current observational study was conducted from Apr 2014 to Feb 2015 on a total of 149 Enterococcus species isolated from Milad hospital in Tehran, Iran. The antibiotic susceptibility pattern of the bacteria was determined by the disc diffusion method for eight antibiotics. Minimum inhibitory concentration (MIC) of vancomycin was also done using agar-dilution assay by clinical and laboratory standards institute (CLSI) recommendations. The sodA, esp, cyl, ace and gelE genes were detected by polymerase chain reaction (PCR) assay. Results: About 37.5%, 73%, 86.6%, 35.8%, 69%, 60.8%, 45% and 79% of the isolates were resistant to vancomycin, tetracycline, gentamicin, chloramphenicol, ciprofloxacin, penicillin, ampicillin and erythromycin, respectively. MIC on 38% of the isolates was 256 g/mL. Although, the prevalence of vancomycin-resistant Enterococcus (VRE) strains belonged to two species, E. faecium showed high resistance to a broad range of antibiotics. In total, 94 isolates were positive for esp, and 59, 48 and 3 isolates were positive for ace, cylA and gelE, Respectively. Conclusions: The results of the current study designate the important role of medical samples as reservoirs of resistance inducing elements. Early detection of VRE with their virulence trait will help to prevent the spread of vancomycin resistant Enterococcus species. Supervision for antibiotic usage in hospitals, especially for last option antibiotics, can prevent the spread of resistant isolates and losing all treatment options in the future.

Screening municipal wastewater effluent and surface water used for drinking water production for the presence of ampicillin and vancomycin resistant enterococci.

The emergence of clinical enterococcal isolates that are resistant to both ampicillin and vancomycin is a cause of great concern, as therapeutic alternatives for the treatment of infections caused by such organisms are becoming limited. Aquatic environments could play a role in the dissemination of antibiotic resistant enterococci. This study investigated the presence of ampicillin and vancomycin resistant enterococci in the treated effluent of six wastewater treatment plants (WWTPs) and in surface water used as a source for drinking water production in the Netherlands. Membrane filtration in combination with selective media with ampicillin or vancomycin was applied to determine the presence of ampicillin resistant Enterococcus (ARE) and vancomycin resistant Enterococcus (VRE) species. Ampicillin resistant Enterococcus faecium (minimal inhibitory concentration (MIC) >16μg/mL; n=1033) was observed in all studied WWTP effluents. In surface water used for drinking water production (intake locations), no ARE or VRE were observed. At both types of location, intrinsic vancomycin resistant Pediococcus spp., Leuconostoc spp. and Lactobacillus spp. were isolated with the vancomycin medium. The ampicillin resistant E. faecium (AREfm) isolates (n=113) did not contain the vanA or vanB gene, but MIC testing for vancomycin showed intermediate vancomycin resistance (2-8μgmL(-1)) to occur in these AREfm strains. This study documents the discharge of ampicillin resistant E. faecium strains with intermediate vancomycin resistance by the WWTPs into the surface water, but no presence of these strains downstream at intake locations for drinking water production.

Antibiotic resistance patterns of microorganisms isolated from nephrology and kidney transplant wards of a referral academic hospital.

Objective:Antibiotic use pattern and emergence of resistant bacteria are major concerns in clinical settings. This study aimed to detect common bacteria and their antibiotic sensitivity patterns in nephrology and kidney transplant wards.Methods:This 1-year, observational study was performed in the nephrology and kidney transplant wards of Imam Khomeini Hospital Complex, Tehran, Iran. All patients treated with antimicrobial agents for confirmed or suspected infections were included. Their demographic, clinical, and laboratory data (including biological media used for microbial culture, growth organisms, and antibiograms) were collected. Adherence of antimicrobial regimen to standard guidelines was also assessed.Findings:About half of the patients received antibiotic. The most common infecting bacteria were Escherichia coli followed by Enterococcus sp. and Staphylococcus aureus. E. coli showed high rate of sensitivity to carbapenems and nitrofurantoin and high rate of resistance to co-trimoxazole and ciprofloxacin. Enterococcus sp. in both wards had high rate of resistance to ampicillin and were all sensitive to linezolid. Unlike to the nephrology ward, more than 50% of Enterococcus sp. from kidney transplant ward was resistant to vancomycin. The most common type of S. aureus in this nephrology ward was methicillin-resistant S. aureus (MRSA). Most commonly-prescribed antibiotics were carbapenems followed by vancomycin, ciprofloxacin, and ceftriaxone. Antibiotic regimens were 75% and 83%, 85% and 91%, and 80% and 87% compatible with international guidelines in antibiotic types, dosages, and treatment durations, respectively, in nephrology and kidney transplant wards, respectively.Conclusion:MRSA, fluoroquinolone-resistant E. coli, and vancomycin resistant Enterococcus species are major threats in nephrology and kidney transplant wards. Most commonly-prescribed antibiotics were carbapenems that necessitate providing internal guidelines by the teamwork of clinical pharmacist, infectious disease specialists, and nephrologists to avoid the widespread use of broad-spectrum antibiotics.

Characterization of aminoglycoside resistance and virulence genes among Enterococcus spp. isolated from a hospital in China.

This study investigated the aminoglycoside resistance phenotypes and genotypes, as well as the prevalence of virulence genes, in Enterococcus species isolated from clinical patients in China. A total of 160 enterococcal isolates from various clinical samples collected from September 2013 to July 2014 were identified to the species level using the VITEK-2 COMPACT system. The antimicrobial susceptibilities of the identified Enterococcus strains were determined by the Kirby-Bauer (K-B) disc diffusion method. PCR-based assays were used to detect the aminoglycoside resistance and virulence genes in all enterococcal isolates. Of 160 Enterococcus isolates, 105 were identified as E. faecium, 35 as E. faecalis, and 20 isolates were classified as “other” Enterococcus species. High-level aminoglycoside resistance (HLAR) for gentamicin, streptomycin, and both antibiotics was identified in 58.8, 50, and 34.4% of strains, respectively. The most common virulence gene (50.6% of isolates) was efaA, followed by asa1 (28.8%). The most prevalent aminoglycoside resistance genes were aac(6')-Ie-aph(2''), aph(2')-Id, aph(3')-IIIa, and ant(6')-Ia, present in 49.4%, 1.3%, 48.8% and 31.3% of strains, respectively. Overall, E. faecium and E. faecalis were most frequently associated with hospital-acquired enterococcal infections in Zhejiang Province. All aminoglycoside resistance genes, except aph(2'')-Id, were significantly more prevalent in HLAR strains than amongst high level aminoglycoside susceptible (HLAS) strains, while there was no significant difference between HLAR and HLAS strains in regard to the prevalence of virulence genes, apart from esp, therefore, measures should be taken to manage infections caused by multi-drug resistant Enterococcus species.

Influence Factors for Duration of Vancomycin Resistant Enterococci's Spontaneous Decolonization

Purpose: The aim of this study was to identify the factors influencing the spontaneous decolonization period of vancomycin resistant enterococcus (VRE) species in pediatric patients. Methods: The medical records of patients presenting positive VRE cultures between January 2005 and November 2010 at a tertiary hospital in Seoul, Korea, were reviewed retrospectively. The subjects were divided into two groups according to the average number of days for decolonization (325 days). Clinical characteristics were compared between shorter VRE colonization patients (

Antibacterial Efficacy Testing of a Bioelectric Wound Dressing Against Clinical Wound Pathogens

Silver-based wound dressings have been developed for the control of bioburden in wounds. However, the popularity and extensive use of silver-based dressings has been associated with emerging microbial resistances to silver. In this study we examined in vitro antibacterial efficacy of a bioelectric dressing containing silver and zinc against various wound pathogens. Antibiotic-sensitive clinical wound isolates showed a 100% reduction in bacterial growth, except that Enterococcus faecalis isolate was shown to survive with a bacterial log10 reduction rate of less than 102 CFU. We also investigated antibacterial efficacy against the extended spectrum β-lactamase (ESBL) bacteria, multidrug-resistant (MDR) bacteria, and methicillin-resistant Staphylococcus aureus (MRSA). The bioelectric dressing was effective in killing wound pathogens including ESBL, MDR, and MRSA in vitro. Furthermore, based on the primary results against E. faecalis, we carried out extensive studies against several nosocomial Enterococcus species including vancomycin-resistant species. Overall, the vancomycin-sensitive or -resistant Enterococcus species were resistant to this dressing at up to 48 h, except for the vancomycin-resistant Enterococcus raffinosus isolate only showing a 100% bacterial reduction at 48 h, but not at 24 h. The results demonstrated the effective bactericidal activity of a bioelectric dressing against antibiotic-sensitive and MDR strains, but Enterococcus species are bacteriostatic.

Antimicrobial resistance of Enterococcus species from meat and fermented meat products isolated by a PCR-based rapid screening method

Enterococci are predominantly found in the gastrointestinal tract of humans and animals, but species commonly resident on vegetation are known. Their presence in large numbers in foods may indicate a lapse in sanitation and their ability to serve as a genetic reservoir of transferable antibiotic resistance is of concern. Conventional culture methods for identification of enterococci are slow and sometimes give false results because of the biochemical diversity of the organisms in this genus. This work reports the development of a PCR-based assay to detect enterococci at the genus level by targeting a 16S rRNA sequence. Published 16S rRNA sequences were aligned and used to design genus specific primers (EntF and EntR). The primers were able to amplify a 678bp target region from Enterococcus faecalis ATCC 7080 and 20 other strains of enterococci from 11 different species, but there was no amplification by 32 species from closely related genera (Pediococcus, Lactobacillus, Streptococcus and Listeria) or species of Escherichia coli and Salmonella. The PCR positive samples were plated, screened by a colony patch technique and their identities were confirmed by API 20 Strep panels and sequencing. When dry fermented sausage and ham as well as fresh meat batter for dry cured sausage manufacture were tested for enterococci by the method, 29 Enterococcus strains (15 E. faecalis, 13 E. faecium, and one E. gallinarum) were identified. When susceptibility of these enterococci to 12 antibiotics was tested, the highest incidence of resistance was to clindamycin (89.6%), followed by tetracycline hydrochloride (65.5%), tylosin (62%), erythromycin (45%), streptomycin and neomycin (17%), chloramphenicol (10.3%), penicillin (10.3%), ciprofloxacin (10.3%) and gentamicin (3.4%). None was resistant to the clinically important drugs vancomycin or ampicillin. Most strains (27/29) were resistant to more than one antibiotic while 17 of 29 strains were resistant to three to 8 antibiotics. The molecular method developed was validated for speciation of enterococci and was useful in assessing uncooked processed meat products as a reservoir for multi-drug resistant Enterococcus species.

Antibacterial activity and mutagenicity of leaves of &amp;lt;i&amp;gt;Indigofera tinctoria&amp;lt;/i&amp;gt; Linn

Objective: Indigofera tinctoria Linn was traditionally been used in the ancient herbal medicine in India/China for epilepsy, nervous disorders, bronchitis, and liver ailments; and also used as an anticardiovascular, but there was no literature evidence substantiating its antibacterial activity. The purpose of this study is to determine the antibacterial and mutagenic properties of an extract prepared from the leaves of I.tinctoria, commonly known as indigo plant. Methods: Minimum inhibitory concentrations (MICs) of the extract are determined with respect to bacterial species by agar dilution as per Clincial and Laboratory Standard Institute and mutagenicity of the extract is determined by Ames test. Results: The respective MICs of the methanol extract of I.tinctoria were methicillin sensitive Staphylococcus aureus 0.125 µg/ml, methicillin resistant S.aureus 0.25 µg/ml, coagulase negative S.aureus 0.25-0.5 µg/ml, vancomycin sensitive Enterococcus faecalis 0.5 µg/ml, vancomycin resistant Enterococcus species 0.125-0.5 µg/ml, Moraxella 2 µg/ml, Streptococcus species 0.125-0.25 µg/ml, Haemophilus influenzae 2-4 µg/ml and against anaerobes is 1-2 µg/ml. Time kill assay were analyzed for methanol extract of I.tinctoria and it showed bacteriostatic and nonmutagenic activity up to 5 mg per plate in Ames test both in the presence and absence of S9 fraction. Conclusion: Methanol extract of I.tinctoria exhibited antibacterial activity, which was more active than that of standard linezolid, against a variety of clinically important bacteria isolated from near by selected hospitals in India and was found to be nonmutagenic up to 5 mg/plate in Ames test.

The anti-sigma factor TcdC modulates hypervirulence in an epidemic BI/NAP1/027 clinical isolate of Clostridium difficile.

Nosocomial infections are increasingly being recognised as a major patient safety issue. The modern hospital environment and associated health care practices have provided a niche for the rapid evolution of microbial pathogens that are well adapted to surviving and proliferating in this setting, after which they can infect susceptible patients. This is clearly the case for bacterial pathogens such as Methicillin Resistant Staphylococcus aureus (MRSA) and Vancomycin Resistant Enterococcus (VRE) species, both of which have acquired resistance to antimicrobial agents as well as enhanced survival and virulence properties that present serious therapeutic dilemmas for treating physicians. It has recently become apparent that the spore-forming bacterium Clostridium difficile also falls within this category. Since 2000, there has been a striking increase in C. difficile nosocomial infections worldwide, predominantly due to the emergence of epidemic or hypervirulent isolates that appear to possess extended antibiotic resistance and virulence properties. Various hypotheses have been proposed for the emergence of these strains, and for their persistence and increased virulence, but supportive experimental data are lacking. Here we describe a genetic approach using isogenic strains to identify a factor linked to the development of hypervirulence in C. difficile. This study provides evidence that a naturally occurring mutation in a negative regulator of toxin production, the anti-sigma factor TcdC, is an important factor in the development of hypervirulence in epidemic C. difficile isolates, presumably because the mutation leads to significantly increased toxin production, a contentious hypothesis until now. These results have important implications for C. difficile pathogenesis and virulence since they suggest that strains carrying a similar mutation have the inherent potential to develop a hypervirulent phenotype.

Evaluation of the automated system Vitek2 for identification and antimicrobial susceptibility testing of Brazilian Gram-positive cocci strains

Automated instruments offer many advantages for clinical laboratories. Nevertheless, they can have problems identifying and determining susceptibilities of some pathogens. Vitek2 (bioMérieux) is an automated system that was recently introduced to Brazil. We evaluated the performance of this equipment for Brazilian isolates that had been characterized using reference identification and antimicrobial susceptibility testing methods. Ninety-nine strains of Gram-positive cocci from a local reference center collection were analyzed, consisting of 50 coagulase-negative Staphylococcus (CoNS) and 49 Enterococcus and related species. Vitek2 correctly identified 79.8% (79/99) of the isolates. Oxacillin resistance was detected in 76% (19/25) of resistant S. epidermidis strains and in 88% (22/25) of other resistant CoNS species strains. Vancomycin resistance was detected in 100% (20/20) of resistant Enterococcus and related species strains. Vitek2 performed very well for the identification of S. epidermidis and non-epidermidis staphylococci, and for the detection of vancomycin resistance in Enterococcus and related species. However, the system needs improvement in order to provide reliable results for the characterization of some CoNS species, identification of Enterococcus and related species and for detecting oxacillin resistance in CoNS.