Outbreak of vancomycin-resistant Enterococcus faecium ST1133 in paediatric patients with acute lymphoblastic leukaemia from southern Brazil.

OBJECTIVES: We aimed to investigate an outbreak of vancomycin-resistant Enterococcus faecium (VREfm) in paediatric patients from Hospital Pequeno Príncipe. The susceptibility profile was determined, and whole-genome sequencing (WGS) was used to analyse the genetic context of the strains. METHODS: Five VREfm isolates were recovered from sterile sites and surveillance cultures of two paediatric patients with acute lymphoblastic leukaemia. Species identification was performed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), and the minimum inhibitory concentration (MIC) was assessed according to the European Committee for Antimicrobial Susceptibility Testing (EUCAST). WGS was performed to analyse the genetic context of virulence and resistance genes, and in silico multilocus sequence typing was performed to identify the sequence typing of the strains. RESULTS: High-level vancomycin resistance was observed in all isolates (≥256 mg/L). WGS revealed the presence of mobile genetic elements, such as plasmids (rep2, rep11a, repUS15, rep17, and rep18a), insertion sequences, and phages. Multiple resistance genes (aac(6')-aph(2"), dfrG, ermB, and vanA) and virulence genes (acm and efaAfm) were identified. All the isolates were assigned to ST117 (ST1133 - via a novel MLST), an important epidemic lineage associated with nosocomial infections and outbreaks. CONCLUSION: Our results show that the ST117 (ST1133) VREfm isolates are circulating in paediatric patients, which raises a great concern. The development of new drugs as well as the implementation of an antimicrobial stewardship program are necessary for their correct management, limiting the spread of resistance in oncohematological patients.

El desarrollo y los complejos mecanismos de resistencia de S. aureus: una amenaza persistente en la era de los antibióticos

Introducción: S. aureus ha emergido como una amenaza persistente, demostrando una notable habilidad para desarrollar resistencia a lo largo de la evolución de los antibióticos. Desde los primeros enfrentamientos con la penicilina hasta los desafíos actuales con cepas resistentes a la vancomicina y la daptomicina, el estudio de los mecanismos de resistencia de este patógeno ha adquirido una importancia crítica. Objetivos: documentar los cambios en los patrones de resistencia de S. aureus a lo largo del tiempo, además de identificar las etapas críticas en el desarrollo de la resistencia a diferentes antibióticos. Materiales y métodos: el proceso de selección de artículos revisados se llevó a cabo identificando artículos publicados entre 2010 y 2023. Se utilizaron varias bases de datos relevantes, incluyendo PubMed, Scopus, Embase, Cochrane Library y Scielo. Se incluyeron estudios observacionales, artículos de revisión y guías clínicas. Se desarrollaron estrategias de búsqueda específicas para cada base de datos utilizando palabras clave y términos de búsqueda relacionados con S. aureus y su resistencia antimicrobiana, así como los tipos de estudios de interés. Se extrajeron datos relevantes de los estudios seleccionados, incluyendo información sobre los patrones de resistencia, mecanismos de resistencia, impacto clínico y estrategias terapéuticas. Los datos recopilados se analizaron y sintetizaron para documentar los cambios en los patrones de resistencia de S. aureus a lo largo del tiempo y para identificar las etapas críticas en el desarrollo de la resistencia a diferentes antibióticos. Resultados: se incluyeron 100 artículos donde se evidencia una evolución temporal de la resistencia, desde las primeras cepas resistentes a la penicilina hasta las actuales cepas resistentes a la vancomicina y la daptomicina. Estos estudios proporcionaron un análisis detallado de los mecanismos moleculares clave que impulsan la resistencia antimicrobiana, tales como la producción de beta-lactamasas, las alteraciones en las proteínas de unión a penicilina y las modificaciones en la membrana celular. Los hallazgos destacan una evolución significativa en la capacidad de S. aureus para adaptarse a diferentes antibióticos a lo largo del tiempo, subrayando la complejidad y la diversidad de los mecanismos de resistencia desarrollados por esta bacteria. Conclusiones: la evolución de la resistencia de S. aureus ha seguido un patrón marcado por etapas críticas, desde la aparición de cepas productoras de penicilinasa tras la introducción de la penicilina, hasta el surgimiento de MRSA con la meticilina y de VISA y VRSA con la vancomicina. Estos cambios destacan la capacidad de adaptación de S. aureus a nuevas presiones antibióticas. La revisión subraya la necesidad urgente de desarrollar estrategias antimicrobianas innovadoras y sostenibles para controlar esta creciente amenaza. Comprender los mecanismos de resistencia es crucial para desarrollar enfoques más efectivos y personalizados en el tratamiento de las infecciones por este germen.

Introduction: S. aureus has emerged as a persistent threat, demonstrating a remarkable ability to develop resistance throughout the evolution of antibiotics. From the first confrontations with penicillin to the current challenges with strains resistant to vancomycin and daptomycin, the study of the resistance mechanisms of this pathogen has acquired critical importance. Objectives: To document changes in S. aureus resistance patterns over time and identify critical stages in the development of resistance to different antibiotics. Materials and methods: The reviewed articles were selected by identifying articles published between 2010 and 2023. Several relevant databases were used, including PubMed, Scopus, Embase, Cochrane Library, and SciELO. Observational studies, review articles, and clinical guidelines were included. Specific search strategies were developed for each database using keywords and search terms related to S. aureus and its antimicrobial resistance as well as the types of studies of interest. Relevant data were extracted from the selected studies, including information on resistance patterns, resistance mechanisms, clinical impact, and therapeutic strategies. The collected data were analyzed and synthesized to document changes in S. aureus resistance patterns over time and identify critical stages in the development of resistance to different antibiotics. Results: One hundred articles were included where a temporal evolution of resistance is evident, from the first strains resistant to penicillin to the current strains resistant to vancomycin and daptomycin. These studies provided a detailed analysis of the key molecular mechanisms driving antimicrobial resistance, such as beta-lactamase production, alterations in penicillin-binding proteins, and cell membrane modifications. The findings highlight a significant evolution in the ability of S. aureus to adapt to different antibiotics over time, underscoring the complexity and diversity of resistance mechanisms developed by this bacterium. Conclusions: The evolution of S. aureus resistance has followed a pattern marked by critical stages, from the appearance of penicillinase-producing strains after the introduction of penicillin to the emergence of MRSA with methicillin and of VISA and VRSA with vancomycin. These changes highlight the ability of S. aureus to adapt to new antibiotic pressures. The review highlights the urgent need to develop innovative and sustainable antimicrobial strategies to control this growing threat. Understanding resistance mechanisms is crucial to developing more effective and personalized approaches for the treatment of infections caused by this germ.

The Emergence of Antimicrobial Resistance and Virulence Characteristics in Enterococcus Species Isolated from Bovine Milk.

Enterococcus spp., including E. faecalis and E. faecium, pose risks to dairy farms as opportunistic pathogens. The study evaluates antimicrobial resistance (AMR) and virulence characteristics of Enterococcus spp. isolated from bovine milk. Bile esculin agar was used to assess 1471 milk samples, followed by colony identification, gram staining, catalase tests, and 45 °C incubation. PCR analysis targeted E. faecalis and E. faecium in characteristic Enterococcus spp. colonies, with MALDI-TOF used for negative samples. Multiple tests, including disk diffusion, chromogenic VRE agar for vancomycin resistance, Vancomycin Etest® for MIC determination, and PCR for virulence factors (cylA, esp, efaA, ace, asa1, gelE, and hyl genes), were performed. Out of 100 identified strains, E. durans (30.66%), E. faecium (26.28%), and E. faecalis (18.25%) were predominant. AMR in Enterococcus spp. varied, with the highest rates against rifampicin (27%), tetracycline (20%), and erythromycin (18%). Linezolid (5%), vancomycin, ciprofloxacin, and teicoplanin (3% each) had lower prevalence. E. faecium and E. faecalis showed high AMR to rifampicin, erythromycin, and tetracycline. Thirty-two strains (18.98%) grew on VRE Chromoselect agar, while 4 (2 E. faecalis and 2 E. faecium) showed vancomycin resistance by MIC values. E. faecalis carried gelE (45.5%) and asa1 (36%), and E. gallinarum had 9.1% with the asa1 gene. Detecting resistant Enterococcus in bovine milk supports control strategies for enterococci on dairy farms, highlighting AMR concerns in the food chain.

In-hospital mortality and one-year survival of critically ill patients with cancer colonized or not with carbapenem-resistant gram-negative bacteria or vancomycin-resistant enterococci: an observational study.

BACKGROUND: Patients with cancer are at risk of multidrug-resistant bacteria colonization, but association of colonization with in-hospital mortality and one-year survival has not been established in critically ill patients with cancer. METHODS: Using logistic and Cox-regression analyses adjusted for confounders, in adult patients admitted at intensive care unit (ICU) with active cancer, we evaluate the association of colonization by carbapenem-resistant Gram-negative bacteria or vancomycin-resistant enterococci with in-hospital mortality and one-year survival. RESULTS: We included 714 patients and among them 140 were colonized (19.6%). Colonized patients more frequently came from ward, had longer hospital length of stay before ICU admission, had unplanned ICU admission, had worse performance status, higher predicted mortality upon ICU admission, and more hematological malignancies than patients without colonization. None of the patients presented conversion of colonization to infection by the same bacteria during hospital stay, but 20.7% presented conversion to infection after hospital discharge. Colonized patients had a higher in-hospital mortality compared to patients without colonization (44.3 vs. 33.4%; p < 0.01), but adjusting for confounders, colonization was not associated with in-hospital mortality [Odds ratio = 1.03 (0.77-1.99)]. Additionally, adjusting for confounders, colonization was not associated with one-year survival [Hazard ratio = 1.10 (0.87-1.40)]. CONCLUSIONS: Adult critically ill patients with active cancer and colonized by carbapenem-resistant Gram-negative bacteria or vancomycin-resistant enterococci active cancer have a worse health status compared to patients without colonization. However, adjusting for confounders, colonization by carbapenem-resistant Gram-negative bacteria or vancomycin-resistant enterococci are not associated with in-hospital mortality and one-year survival.

Susceptibility to Vancomycin of Biofilm Producing Staphylococci Isolated from Tertiary Care Hospital of Nepal / Susceptibilidad a la vancomicina de estafilococos productores de biopelículas aislados del hospital de atención terciaria de Nepal

Background: Methicillin resistance and biofilm-producing Staphylococci are emerging as multidrug-resistant strains narrowing the efficacy of antimicrobial therapy. Although vancomycin is used as the drug of choice to treat such isolates, different studies worldwide have documented the emergence of strains that are intermediately susceptible or resistant to this antibiotic. Objective: The study aimed to determine the minimum inhibitory concentration of vancomycin to methicillin-resistant and biofilm-producing staphylococci isolated from different clinical specimens. Methods: 375 staphylococci isolated from different clinical specimens over one year were included in the study. Biofilm formation was determined by the Tissue culture plate method (TCP), and ica genes were identified by Polymerase Chain Reaction (PCR). Antibiotic susceptibility and methicillin resistance were done following Clinical and Laboratory Standards Institute (CLSI) guidelines. The minimum inhibitory concentration (MIC) of vancomycin in all isolates was determined by the agar dilution method. Results:Among 375 Staphylococci studied, 43% and 57% represented S. aureus and Coagulase-Negative Staphylococci (CNS), respectively. The rate of Methicillin-Resistant S. aureus (MRSA) and Methicillin-Resistant Coagulase Negative Staphylococci (MRCNS) were 81.4% and 66.8% respectively and determined by the disc diffusion method. The most potential antibiotics were tetracycline and chloramphenicol showing sensitivity to more than 90% isolates. The Minimum Inhibitory Concentration (MIC) value of oxacillin for staphylococci ranged from 0.125-32 µg/ml. Oxacillin agar diffusion method showed 51.6% and 79.9% isolates as MRSA and MRCNS, respectively, revealing a very high percentage of S. aureus and CNS isolates as methicillin-resistant. All isolates had susceptible vancomycin MICs that ranged from 0.125-2 µg/ml. Two S. aureus isolated from Central Venous Catheter (CVC) and catheter specimens were detected with intermediate susceptibility to vancomycin. Similarly, three CNS isolated from blood, CVC, and wound/pus (w/p) were intermediately susceptible to vancomycin. Strong biofilm formation was observed in 22.1% of clinical isolates, and the ica gene was detected among 22.9% of isolates. Only one S. aureus detected as a biofilm producer by the TCP method was found to have intermediate susceptibility to vancomycin. Conclusions: The increment in vancomycin MIC among methicillin-resistant and biofilm-producing staphylococci is alarming. Strict control measures to prevent methicillin-resistant isolates spread and routine surveillance for vancomycin-resistant isolates must be incorporated in hospitals to prevent antimicrobial treatment failure

Antecedentes: Los estafilococos resistentes a la meticilina y productores de biopelículas están surgiendo como cepas multirresistentes que reducen la eficacia del tratamiento antimicrobiano. Aunque la vancomicina se utiliza como fármaco de elección para tratar dichos aislados, diferentes estudios realizados en todo el mundo han documentado la aparición de cepas intermedias susceptibles o resistentes a este antibiótico. Objetivo: El estudio tenía como objetivo determinar la concentración mínima inhibitoria de la vancomicina para los estafilococos resistentes a la meticilina y productores de biofilm aislados de diferentes muestras clínicas. Métodos: Se incluyeron en el estudio 375 estafilococos aislados de diferentes muestras clínicas durante un año. La formación de biopelículas se determinó mediante el método de la placa de cultivo de tejidos (TCP), y los genes ica se identificaron mediante la reacción en cadena de la polimerasa (PCR). La susceptibilidad a los antibióticos y la resistencia a la meticilina se realizaron siguiendo las directrices del Clinical and Laboratory Standards Institute (CLSI). La concentración inhibitoria mínima (MIC) de vancomicina en todos los aislados se determinó por el método de dilución en agar. Resultados:Entre los 375 estafilococos estudiados, el 43% y el 57% representaban S. aureus y estafilococos coagulasa-negativos (ECN), respectivamente. La tasa de S. aureus resistente a la meticilina (SARM) y de estafilococos coagulasa negativos resistentes a la meticilina (ECNM) fue del 81,4% y el 66,8%, respectivamente, y se determinó por el método de difusión de discos. Los antibióticos más potenciales fueron la tetraciclina y el cloranfenicol, que mostraron una sensibilidad superior al 90% de los aislados. El valor de la concentración inhibitoria mínima (CIM) de la oxacilina para los estafilococos osciló entre 0,125-32 µg/ml. El método de difusión en agar de la oxacilina mostró que el 51,6% y el 79,9% de los aislados eran SARM y MRCNS, respectivamente, lo que revela que un porcentaje muy elevado de los aislados de S. aureus y CNS son resistentes a la meticilina. Todos los aislados tenían MIC de vancomicina susceptibles que oscilaban entre 0,125-2 µg/ml. Se detectaron dos S. aureus aislados de muestras de catéteres venosos centrales (CVC) y catéteres con una susceptibilidad intermedia a la vancomicina. Del mismo modo, tres S. aureus aislados de sangre, CVC y herida/pus (w/p) fueron intermedianamente susceptibles a la vancomicina. Se observó una fuerte formación de biopelículas en el 22,1% de los aislados clínicos, y se detectó el gen ica en el 22,9% de los aislados. Sólo un S. aureus detectado como productor de biopelículas por el método TCP resultó tener una susceptibilidad intermedia a la vancomicina. Conclusiones: El incremento de la MIC de vancomicina entre los estafilococos resistentes a la meticilina y productores de biofilm es alarmante. Para evitar el fracaso del tratamiento antimicrobiano, deben incorporarse en los hospitales medidas de control estrictas para prevenir la propagación de los aislados resistentes a la meticilina y una vigilancia rutinaria de los aislados resistentes a la vancomicina

Risk Factors Associated with Failure of Linezolid Therapy in Vancomycin-Resistant Enterococcus faecium Bacteremia: A Retrospective Cohort Study in a Referral Center in Mexico.

We aimed to assess the factors associated with 30-day mortality in patients with vancomycin-resistant Enterococcus faecium (VREf) bloodstream infection (BSI) who received treatment with linezolid in an 11-year retrospective cohort of patients with VREf BSI. A univariate and stepwise multivariate logistic regression analysis was performed to determine 30-day mortality factors. Moreover, a Cox proportional hazards analysis of predictor covariates of mortality was performed. Eighty patients were included in the final analysis; 42 (53%) died and 38 (47%) survived 30 days after the index bacteremia. Thirteen patients of 42 (31%) died in the first 7 days. The Acute Physiology and Chronic Health Evaluation II (APACHE II) score was significantly associated with 30-day mortality (adjusted odds ratio [aOR], 1.46; 95% confidence interval [CI]: 1.22-1.76; p < 0.001) in the multivariate analysis. Moreover, VREf BSI persisting for more than 48 hours was a strong factor related to 30-day mortality (aOR, 19.6; 95% CI: 1.46-263; p = 0.01). Adequate control of infection source showed a trend to be protective without reaching significance in the multivariate analysis (aOR, 0.19; 95% CI: 0.04-1.0; p = 0.05). The Cox proportional hazards analysis confirmed the same significant mortality predictor besides linezolid treatment within the first 48 hours as a protective factor (hazard ratio 0.46; 95% CI: 0.23-0.92, p = 0.02). Severely ill patients with high APACHE II score and persistent bacteremia have a higher risk of failure with linezolid therapy.

Genomic Analysis of Multidrug-Resistant Enterococcus faecium Harboring vanA Gene from Wastewater Treatment Plants.

The emergence of vancomycin-resistant Enterococcus faecium (Efm) harboring vanA gene and multidrug-resistant determinants is a relevant public health concern. It is an opportunistic pathogen responsible for nosocomial infections widely distributed in the environment, including wastewater treatment plants (WWTPs). Our study addresses a genomic investigation of vanA-carrying Efm from WWTPs in Brazil. Samples from five WWTPs supplied with sewage from different sources were evaluated. Here we present whole-genome sequencing of eight vanA-Efm isolates performed on Illumina MiSeq platform. All these isolates presented multidrug-resistant profile, and five strains were from treated wastewater. Multiple antimicrobial resistance genes (ARGs) were found, such as aph(3')-IIIa, ant(6')-Ia, erm(B), and msrC, some of them being allocated in plasmids. The virulence profile was predominantly constituted by efaAfm and acm genes and all isolates, except for one, were predicted as human pathogens. Multilocus sequence typing analysis revealed a new allele and five different STs, three previously described (ST32, ST168, and ST253) and two novel ones (ST1893 and ST1894). Six strains belonged to CC17, often associated with hospital outbreaks. As far as our knowledge, no genomic studies of vanA-Efm recovered from WWTPs revealed isolates belonging to CC17 in Brazil. Therefore, our findings point to the environmental spread of Efm carrying multiple ARGs.

Antimicrobial Peptides with Antibacterial Activity against Vancomycin-Resistant Staphylococcus aureus Strains: Classification, Structures, and Mechanisms of Action.

The emergence of bacteria resistant to conventional antibiotics is of great concern in modern medicine because it renders ineffectiveness of the current empirical antibiotic therapies. Infections caused by vancomycin-resistant Staphylococcus aureus (VRSA) and vancomycin-intermediate S. aureus (VISA) strains represent a serious threat to global health due to their considerable morbidity and mortality rates. Therefore, there is an urgent need of research and development of new antimicrobial alternatives against these bacteria. In this context, the use of antimicrobial peptides (AMPs) is considered a promising alternative therapeutic strategy to control resistant strains. Therefore, a wide number of natural, artificial, and synthetic AMPs have been evaluated against VRSA and VISA strains, with great potential for clinical application. In this regard, we aimed to present a comprehensive and systematic review of research findings on AMPs that have shown antibacterial activity against vancomycin-resistant and vancomycin-intermediate resistant strains and clinical isolates of S. aureus, discussing their classification and origin, physicochemical and structural characteristics, and possible action mechanisms. This is the first review that includes all peptides that have shown antibacterial activity against VRSA and VISA strains exclusively.

Systematic review and meta-analysis of the epidemiology of vancomycin-resistance Staphylococcus aureus isolates.

BACKGROUND: Vancomycin­resistant Staphylococcus aureus (VRSA) is a serious public health challenging concern worldwide. OBJECTIVES: Therefore, the objective of present study of 62 published studies was to evaluate the prevalence of VRSA based on different years, areas, isolate source, antimicrobial susceptibility testing, and the genetic determinants. METHODS: We searched the relevant articles that focused on the prevalence rates of VRSA in PubMed, Scopus, Embase, and Web of Science from 2000 to 2019. Statistical analyses were conducted using STATA software (version 14.0). RESULTS: The prevalence of VRSA was 2% before 2006, 5% in 2006-2014, and 7% in 2015-2020 that showed a 3.5-fold increase in the frequency of VRSA between before 2006 and 2020 years. The prevalence of VRSA was 5% in Asia, 1% in Europe, 4% in America, 3% in South America, and 16% in Africa. The frequencies of VRSA isolated from clinical, non-clinical, and mixed samples were 6%, 7%, and 14%, respectively. The prevalence of VRSA was 12% using disk diffusion agar method, 7% using MIC-base methods, and 4% using mixed-methods. The prevalence of vanA, vanB, and vanC1 positive were 71%, 26%, and 4% among VRSA strains. The most prevalent genotype was staphylococcal cassette chromosomemec (SCCmec) II, which accounted for 57% of VRSA. The most prevalent staphylococcal protein A (spa) types were t002, t030, and t037. CONCLUSION: The prevalence of VRSA has been increasing in recent years particularly in Africa/Asia than Europe/America. The most prevalent of genetic determinants associated with VRSA were vanA and SCCmec II. This study clarifies that the rigorous monitoring of definite antibiotic policy, regular surveillance/control of nosocomial-associated infections and intensive surveillance of vancomycin-resistance are required for preventing emergence and further spreading of VRSA.

Enterococos resistentes à vancomicina e tetraciclina em carnes cruas e processadas: características fenotípicas e genotípicas / Vancomycin and tetracycline-resistant enterococci from from raw and processed meats: phenotypic and genotypic characteristics of isolates

The ubiquitous nature of enterococci and their ability to colonize different habitats account for their easy spread throughout the food chain. Here, we evaluated the distribution and antimicrobial susceptibility of Enterococcus isolates from meats obtained from different supermarkets. We acquired and cultured 100 products (raw chicken meat, raw pork, and boiled meats) to screen for the presence of Enterococcus spp. In total, 194 isolates were recovered from the samples, with contamination rates of 63.6% in the chicken samples, 31% in the raw pork meat, and 1.4% in the boiled meat samples. PCR amplification with specific primers was performed to screen the DNA of Enterococcus spp. (95/96), E. faecalis (66/96), E. faecium (30/96), and E. casseliflavus/E. flavescens (3/96). The antimicrobial susceptibility tests showed that all the isolates were resistant to at least one of the antibiotics. All E. faecium isolates were resistant to vancomycin, streptomycin, ciprofloxacin, norfloxacin, erythromycin, and tetracycline. The E. casseliflavus/E. flavescens isolates were resistant to gentamicin, streptomycin, ciprofloxacin, norfloxacin, erythromycin, and tetracycline. E. faecalis isolates were resistant to ciprofloxacin, tetracycline, and erythromycin (92%), norfloxacin (83%), vancomycin, and streptomycin (50%). The resistance genes tetL and vanB were detected by genotyping. The presence of these antimicrobial-resistant microorganisms in food might pose problems for public health.(AU)

A natureza ubíqua dos enterococos e sua capacidade de colonizar diferentes habitats são responsáveis pela sua fácil disseminação pela cadeia alimentar. No presente estudo, avaliamos a distribuição e a susceptibilidade antimicrobiana de isolados de Enterococcus provenientes de produtos cárneos. Cem produtos (carne de frango cru, carne de porco crua e carne cozida) foram adquiridos e cultivados para a presença de Enterococcus spp. No total, 194 amostras foram avaliadas, com taxas de contaminação de 63,6% nas amostras de frango, 31% na carne de porco crua e 1,4% nas amostras de carne cozida. A amplificação por PCR foi realizada para confirmar a presença de Enterococcus spp. (95/96), E. faecalis (66/96), E. faecium (30/96) E. casseliflavus/E. flavescens (3/96). Resultados de susceptibilidade mostraram que 100% dos isolados foram resistentes a pelo menos um antibiótico, sendo 100% de E. faecium resistentes a vancomicina, estreptomicina, ciprofloxacina, norfloxacina, eritromicina e tetraciclina. E. casseliflavus / E. flavescens resistentes a gentamicina, estreptomicina, ciprofloxacina, norfloxacina, eritromicina e tetraciclina. E. faecalis foram resistentes a ciprofloxacina, tetraciclina e eritromicina (92%), norfloxacina (83%), vancomicina e estreptomicina (50%). Na genotipagem, foram detectados os genes tetL e vanB. A presença desses microrganismos resistentes aos antimicrobianos nos alimentos pode causar problemas para a saúde pública.(AU)

Enterococos resistentes à vancomicina e tetraciclina em carnes cruas e processadas: características fenotípicas e genotípicas / Vancomycin and tetracycline-resistant enterococci from from raw and processed meats: phenotypic and genotypic characteristics of isolates

The ubiquitous nature of enterococci and their ability to colonize different habitats account for their easy spread throughout the food chain. Here, we evaluated the distribution and antimicrobial susceptibility of Enterococcus isolates from meats obtained from different supermarkets. We acquired and cultured 100 products (raw chicken meat, raw pork, and boiled meats) to screen for the presence of Enterococcus spp. In total, 194 isolates were recovered from the samples, with contamination rates of 63.6% in the chicken samples, 31% in the raw pork meat, and 1.4% in the boiled meat samples. PCR amplification with specific primers was performed to screen the DNA of Enterococcus spp. (95/96), E. faecalis (66/96), E. faecium (30/96), and E. casseliflavus/E. flavescens (3/96). The antimicrobial susceptibility tests showed that all the isolates were resistant to at least one of the antibiotics. All E. faecium isolates were resistant to vancomycin, streptomycin, ciprofloxacin, norfloxacin, erythromycin, and tetracycline. The E. casseliflavus/E. flavescens isolates were resistant to gentamicin, streptomycin, ciprofloxacin, norfloxacin, erythromycin, and tetracycline. E. faecalis isolates were resistant to ciprofloxacin, tetracycline, and erythromycin (92%), norfloxacin (83%), vancomycin, and streptomycin (50%). The resistance genes tetL and vanB were detected by genotyping. The presence of these antimicrobial-resistant microorganisms in food might pose problems for public health.

A natureza ubíqua dos enterococos e sua capacidade de colonizar diferentes habitats são responsáveis pela sua fácil disseminação pela cadeia alimentar. No presente estudo, avaliamos a distribuição e a susceptibilidade antimicrobiana de isolados de Enterococcus provenientes de produtos cárneos. Cem produtos (carne de frango cru, carne de porco crua e carne cozida) foram adquiridos e cultivados para a presença de Enterococcus spp. No total, 194 amostras foram avaliadas, com taxas de contaminação de 63,6% nas amostras de frango, 31% na carne de porco crua e 1,4% nas amostras de carne cozida. A amplificação por PCR foi realizada para confirmar a presença de Enterococcus spp. (95/96), E. faecalis (66/96), E. faecium (30/96) E. casseliflavus/E. flavescens (3/96). Resultados de susceptibilidade mostraram que 100% dos isolados foram resistentes a pelo menos um antibiótico, sendo 100% de E. faecium resistentes a vancomicina, estreptomicina, ciprofloxacina, norfloxacina, eritromicina e tetraciclina. E. casseliflavus / E. flavescens resistentes a gentamicina, estreptomicina, ciprofloxacina, norfloxacina, eritromicina e tetraciclina. E. faecalis foram resistentes a ciprofloxacina, tetraciclina e eritromicina (92%), norfloxacina (83%), vancomicina e estreptomicina (50%). Na genotipagem, foram detectados os genes tetL e vanB. A presença desses microrganismos resistentes aos antimicrobianos nos alimentos pode causar problemas para a saúde pública.

Factors associated with vancomycin-resistant enterococci colonization in a pediatric intensive care unit of Paraguay: A cross-sectional study on hospital charts. / Factores asociados a colonización por enterococo sp resistente a vancomicina en una unidad de cuidados intensivos pediátricos del Paraguay: estudio transversal sobre historias clínicas.

INTRODUCTION: Vancomycin-resistant enterococci (VRE) infections have become widespread and a challenge in hospitalized patients. The threat of infection by intractable enterococci and the possibility that vancomycin resistance could involve pneumococci or staphylococci advocate for careful surveillance of resistant strains. OBJECTIVE: To determine the risk factors associated with VRE colonization in pediatric patients admitted to the Pediatric Intensive Care Unit (PICU) in the period between January 2012 and June 2013. METHODS: We conducted a cross-sectional study analyzing the clinical histories of 140 patients admitted to the PICU (children from 1 month to 18 years) who underwent rectal swab cultures within 48 hours of admission. We calculated the odds ratios and confidence intervals of the risk factors for VRE colonization in the PICU, and then we used multiple logistic regression for the statistically significant variables. RESULTS: VRE colonization was present in 18.6% of patients. The following variables were identified as risk factors associated with VRE colonization: prior hospitalization in the past year (odds ratio: 10.8; 95% confidence interval: 2.43 to 47.8; p = 0.001); prior use of one broad-spectrum antibiotic (odds ratio: 5.05; 95% confidence interval: 2.04 to 12.5; p = 0.000); use of two or more broad-spectrum antibiotics in past year (odds ratio: 5.4; 95% confidence interval: 1.5 to 18.4; p = 0.009); prior hospitalization in a high-risk area (odds ratio: 4.91; 95% confidence interval: 1.83 to 13.2; p = 0.000); hospitalization for more than five days in a high-risk area (odds ratio: 5.64; 95% confidence interval: 2.18 to 14.6; p = 0.000); and use of immunosuppressant drugs (odds ratio: 4.84; 95% confidence interval: 1.92 to 11.9; p = 0.001). In a logistic multiple regression the use of two or more broad-spectrum antibiotics (odds ratio: 4.81; 95% confidence interval: 1.01 to 22.8; p = 0.047) and prior hospitalization in past year (odds ratio: 7.84; 95% confidence interval: 1.24 to 49.32; p = 0.028) were identified as independent factors statistically associated with VRE colonization. CONCLUSION: Pediatric patients admitted for intensive care with a history of prior hospitalization in the past year and exposure to two or more broad-spectrum antibiotics have a greater risk of colonization by vancomycin-resistant enterococci.

INTRODUCCIÓN: Las infecciones por Enterococcus sp resistente a la vancomicina se han diseminado y generan un desafío clínico-terapéutico en los pacientes hospitalizados. La amenaza de que la infección por enterococos intratables y la posibilidad que la resistencia a la vancomicina pueda propagarse a neumococos o estafilococos, abogan por la vigilancia atenta de las cepas resistentes. OBJETIVO: Determinar los factores de riesgos asociados a la portación de Enterococcus sp resistente a la vancomicina en pacientes pediátricos ingresados en una unidad de cuidados intensivos pediátricos del Paraguay en el periodo entre enero de 2012 y junio de 2013. MÉTODOS: Estudio transversal. Se analizaron las historias clínicas previas de 140 pacientes ingresados a terapia intensiva (niños de un mes a 18 años), a quienes se realizaron cultivos de hisopado rectal dentro de las 48 horas del ingreso, para determinar los factores asociados a la portación de Enterococcus sp resistente a la vancomicina en unidad de cuidados intensivos pediátricos. Se calculó el Odd ratio con sus intervalos de confianza y p < 0,05 para las variables de estudio. Posteriormente, se realizó regresión logística múltiple para las variables estadísticamente significativas. RESULTADOS: La portación de Enterococcus sp resistente a la vancomicina se observó en 18,6% de los pacientes. Se identificaron como factores asociados: la hospitalización previa durante el último año (Odds ratio: 10,8; intervalo de confianza 95%: 2,43 a 47,8; p = 0,001), uso previo de antibióticos de amplio espectro (Odds ratio: 5,05; intervalo de confianza 95%: 2,04 a 12,5; p = 0,000), uso de dos o más antibióticos de amplio espectro en el último año (Odds ratio: 5,4; intervalo de confianza 95%: 1,5 a 18,4; p = 0,009), internación previa en área de alto riesgo (Odds ratio: 4,91; intervalo de confianza 95%: 1,83 a 13,2; p = 0,000), internación por igual o mayor a seis días en área de alto riesgo (Odds ratio: 5,64; intervalo de confianza 95%: 2,18 a 14,6; p = 0,000) y uso de inmunosupresores (Odds ratio: 4,84; intervalo de confianza 95%: 1,92 a 11,9; p = 0,001). La regresión múltiple señala a la utilización de dos o más antibióticos de amplio espectro (Odds ratio: 4,81; intervalo de confianza 95%: 1,01 a 22,8; p = 0,047) y a la historia de hospitalización previa dentro del año (Odds ratio: 7,84; intervalo de confianza 95%: 1,24 a 49,32; p = 0,028) como factores independientes asociados estadísticamente con la portación de Enterococcus sp resistente a la vancomicina. CONCLUSIÓN: Los pacientes pediátricos ingresados en la unidad de cuidados intensivos con historia de internación previa dentro del año y la exposición a dos o más antibióticos de amplio espectro, tienen mayor riesgo de colonización por el enterococo resistente a vancomicina.

Draft genome sequence of Enterococcus faecium E86, a strain producing broad-spectrum antimicrobial peptides: Description of a novel bacteriocin immunity protein and a novel sequence type.

OBJECTIVES: The aim of this study was to report the draft genome sequence of the bacteriocinogenic strain Enterococcus faecium E86. Bacteriocins are prokaryotic peptides or proteins with antimicrobial activity. The genome information may contribute to the identification of enterocins produced by this strain that exhibit inhibitory activity against the foodborne pathogen Listeria monocytogenes and vancomycin-resistant enterococci (VRE) involved in human infections, among other bacterial genera and species. METHODS: An Illumina MiSeq platform was used for genome sequencing. De novo assembly of 5 735 838 paired-end reads was done using the A5-miseq pipeline, yielding >300-fold average genome coverage. Genome annotation was performed by the RAST server, and mining of the bacteriocinogenic gene clusters was done using the BAGEL3 and antiSMASH v.4 platforms. RESULTS: The total scaffold size was determined to be 2 689 107 bp, approximately 2.7 Mbp, featuring a G + C content of 38.1%. The genome contains 2858 coding sequences and 74 RNA genes. Genome analyses revealed the presence of: 30 genes involved in drug resistance; 2 bacteriocinogenic gene clusters (for enterocin P and enterocin TW21); EntiTW21, a novel bacteriocin immunity protein and a novel multilocus sequence type (ST1500). CONCLUSION: This work highlights the potential biotechnological application of this strain for the production of enterocin P, a bacteriocin that can be employed in the food industry as a biopreservative against L. monocytogenes and as an alternative to classical antibiotics against VRE.

An epidemiological and molecular study regarding the spread of vancomycin-resistant Enterococcus faecium in a teaching hospital in Bogotá, Colombia 2016.

BACKGROUND: Enterococcus faecium is ranked worldwide as one of the top ten pathogens identified in healthcare-associated infections (HAI) and is classified as one of the high priority pathogens for research and development of new antibiotics worldwide. Due to molecular biology techniques' higher costs, the approach for identifying and controlling infectious diseases in developing countries has been based on clinical and epidemiological perspectives. Nevertheless, after an abrupt vancomycin-resistant Enterococcus faecium dissemination in the Méderi teaching hospital, ending up in an outbreak, further measures needed to be taken into consideration. The present study describes the vancomycin-resistant Enterococcus faecium pattern within Colombian's largest installed-bed capacity hospital in 2016. METHODS: Thirty-three vancomycin-resistant Enterococcus faecium isolates were recovered during a 5-month period in 2016. Multilocus variable-number tandem-repeat analysis was used for molecular typing to determine clonality amongst strains. A modified time-place-sequence algorithm was used to trace VREfm spread patterns during the outbreak period and estimate transmission routes. RESULTS: Four clonal profiles were identified. Chronological clonal profile follow-up suggested a transitional spread from profile "A" to profile "B", returning to a higher prevalence of "A" by the end of the study. Antibiotic susceptibility indicated high-level vancomycin-resistance in most isolates frequently matching vanA gene identification. DISCUSSION: Transmission analysis suggested cross-contamination via healthcare workers. Despite epidemiological control of the outbreak, post-outbreak isolates were still being identified as having outbreak-related clonal profile (A), indicating reduction but not eradication of this clonality. This study supports the use of combined molecular and epidemiological strategies in an approach to controlling infectious diseases. It contributes towards a more accurate evaluation of the effectiveness of the epidemiological measures taken regarding outbreak control and estimates the main cause related to the spread of this microorganism.

Molecular basis for the emergence of a new hospital endemic tigecycline-resistant Enterococcus faecalis ST103 lineage.

Enterococcus faecalis are a major cause of nosocomial infection worldwide, and the spread of vancomycin resistant strains (VRE) limits treatment options. Tigecycline-resistant VRE began to be isolated from inpatients at a Brazilian hospital within months following the addition of tigecycline to the hospital formulary. This was found to be the result of a spread of an ST103 E. faecalis clone. Our objective was to identify the basis for tigecycline resistance in this lineage. The genomes of two closely related tigecycline-susceptible (MIC = 0.06 mg/L), and three representative tigecycline-resistant (MIC = 1 mg/L) ST103 isolates were sequenced and compared. Further, efforts were undertaken to recapitulate the emergence of resistant strains in vitro. The specific mutations identified in clinical isolates in several cases were within the same genes identified in laboratory-evolved strains. The contribution of various polymorphisms to the resistance phenotype was assessed by trans-complementation of the wild type or mutant alleles, by testing for differences in mRNA abundance, and/or by examining the phenotype of transposon insertion mutants. Among tigecycline-resistant clinical isolates, five genes contained non-synonymous mutations, including two genes known to be related to enterococcal tigecycline resistance (tetM and rpsJ). Finally, within the in vitro-selected resistant variants, mutation in the gene for a MarR-family response regulator was associated with tigecycline resistance. This study shows that E. faecalis mutates to attain tigecycline resistance through the complex interplay of multiple mechanisms, along multiple evolutionary trajectories.

Factores asociados a colonización por enterococo sp resistente a vancomicina en una unidad de cuidados intensivos pediátricos del Paraguay: estudio transversal sobre historias clínicas / Factors associated with vancomycin-resistant enterococcus colonization in a pediatric intensive care unit of Paraguay: a cross-sectional study on hospital charts

INTRODUCCIÓN Las infecciones por Enterococcus sp resistente a la vancomicina se han diseminado y generan un desafío clínico-terapéutico en los pacientes hospitalizados. La amenaza de que la infección por enterococos intratables y la posibilidad que la resistencia a la vancomicina pueda propagarse a neumococos o estafilococos, abogan por la vigilancia atenta de las cepas resistentes. OBJETIVO Determinar los factores de riesgos asociados a la portación de Enterococcus sp resistente a la vancomicina en pacientes pediátricos ingresados en una unidad de cuidados intensivos pediátricos del Paraguay en el periodo entre enero de 2012 y junio de 2013. MÉTODOS Estudio transversal. Se analizaron las historias clínicas previas de 140 pacientes ingresados a terapia intensiva (niños de un mes a 18 años), a quienes se realizaron cultivos de hisopado rectal dentro de las 48 horas del ingreso, para determinar los factores asociados a la portación de Enterococcus sp resistente a la vancomicina en unidad de cuidados intensivos pediátricos. Se calculó el Odd ratio con sus intervalos de confianza y p < 0,05 para las variables de estudio. Posteriormente, se realizó regresión logística múltiple para las variables estadísticamente significativas. RESULTADOS La portación de Enterococcus sp resistente a la vancomicina se observó en 18,6% de los pacientes. Se identificaron como factores asociados: la hospitalización previa durante el último año (Odds ratio: 10,8; intervalo de confianza 95%: 2,43 a 47,8; p = 0,001), uso previo de antibióticos de amplio espectro (Odds ratio: 5,05; intervalo de confianza 95%: 2,04 a 12,5; p = 0,000), uso de dos o más antibióticos de amplio espectro en el último año (Odds ratio: 5,4; intervalo de confianza 95%: 1,5 a 18,4; p = 0,009), internación previa en área de alto riesgo (Odds ratio: 4,91; intervalo de confianza 95%: 1,83 a 13,2; p = 0,000), internación por igual o mayor a seis días en área de alto riesgo (Odds ratio: 5,64; intervalo de confianza 95%: 2,18 a 14,6; p = 0,000) y uso de inmunosupresores (Odds ratio: 4,84; intervalo de confianza 95%: 1,92 a 11,9; p = 0,001). La regresión múltiple señala a la utilización de dos o más antibióticos de amplio espectro (Odds ratio: 4,81; intervalo de confianza 95%: 1,01 a 22,8; p = 0,047) y a la historia de hospitalización previa dentro del año (Odds ratio: 7,84; intervalo de confianza 95%: 1,24 a 49,32; p = 0,028) como factores independientes asociados estadísticamente con la portación de Enterococcus sp resistente a la vancomicina. CONCLUSIÓN Los pacientes pediátricos ingresados en la unidad de cuidados intensivos con historia de internación previa dentro del año y la exposición a dos o más antibióticos de amplio espectro, tienen mayor riesgo de colonización por el enterococo resistente a vancomicina.

INTRODUCTION Vancomycin-resistant enterococcus (VRE) infections have become widespread and a challenge in hospitalized patients. The threat of infection by intractable enterococci and the possibility that vancomycin resistance could involve pneumococci or staphylococci advocate for careful surveillance of resistant strains. OBJECTIVE To determine the risk factors associated with VRE colonization in pediatric patients admitted to the Pediatric Intensive Care Unit (PICU) in the period between January 2012 and June 2013. METHODS We conducted a cross-sectional study analyzing the clinical histories of 140 patients admitted to the PICU (children from 1 month to 18 years), who underwent rectal swab cultures within 48 hours of admission. We calculated the odds ratios and confidence intervals of the risk factors for VRE colonization in the PICU, and then we used multiple logistic regression for the statistically significant variables. RESULTS VRE colonization was present in 18.6% of patients. The following were identified as risk factors associated to VRE colonization: hospitalization during the previous year (odds ratio: 10.8, 95% confidence interval: 2.43 to 47.8; p = 0.001), prior use of one broad-spectrum antibiotic (odds ratio: 5.05; 95% confidence interval: 2.04 to 12.5; p = 0.000), use of two or more broad-spectrum antibiotics in the last year (odds ratio: 5.4, 95% confidence interval: 1.5 to 18.4; p = 0.009), prior hospitalization in the risk area (odds ratio: 4.91, 95% confidence interval: 1.83 to 13.2; p = 0.000), hospitalization for more than five days in high-risk area (odds ratio: 5.64, 95% confidence interval: 2.18 to 14.6; p = 0.000), and use of immunosuppressant drugs (odds ratio: 4.84, 95% confidence interval: 1.92 to 11.9; p = 0.001). In a logistic multiple regression the use of two or more broad-spectrum antibiotics (odds ratio: 4.81, 95% confidence interval: 1.01 to 22.8; p = 0.047) and the history of prior hospitalization in the last year (odds ratio: 7.84, 95% confidence interval: 1.24 to 49.32, p = 0.028) were identified as independent factors statistically associated with VRE colonization. CONCLUSION Pediatric patients admitted to the Intensive Care Unit with a history of prior hospitalization in the previous year, and exposure to two or more broad-spectrum antibiotics have a greater risk of colonization by vancomycin-resistant enterococcus.

A New Kind of Quinonic-Antibiotic Useful Against Multidrug-Resistant S. aureus and E. faecium Infections.

A rapid emergence of resistant bacteria is occurring worldwide, endangering the efficacy of antibiotics and reducing the therapeutic arsenal available for treatment of infectious diseases. In the present study, we developed a new class of compounds with antibacterial activity obtained by a simple, two step synthesis and screened the products for in vitro antibacterial activity against ATCC® strains using the broth microdilution method. The compounds exhibited minimum inhibitory concentrations (MIC) of 1⁻32 µg/mL against Gram-positive ATCC® strains. The structure⁻activity relationship indicated that the thiophenol ring is essential for antibacterial activity and the substituents on the thiophenol ring module, for antibacterial activity. The most promising compounds detected by screening were tested against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VREF) clinical isolates. We found remarkable activity against VREF for compounds 7 and 16, were the MIC50/90 were 2/4 µg/mL and 4/4 µg/mL, respectively, while for vancomycin the MIC50/90 was 256/512 µg/mL. Neither compound affected cell viability in any of the mammalian cell lines at any of the concentrations tested. These in vitro data show that compounds 7 and 16 have an interesting potential to be developed as new antibacterial drugs against infections caused by VREF.

Vancomycin-resistant enterococci isolates colonizing and infecting haematology patients: clonality, and virulence and resistance profile.

BACKGROUND: Vancomycin-resistant enterococci (VRE) are an important agent of colonization and infection in haematology patients. However, the role of virulence on VRE colonization and infection is controversial. AIM: To characterize the lineage, virulence and resistance profile of VRE infection and colonization isolates; as well as their impact on outcome of haematology patients using a regression logistic model. METHODS: Eighty-six isolates (80 Enterococcus faecium and six E. faecalis) from 76 patients were evaluated. Polymerase chain reaction for resistance and virulence genes, and pulsed-field gel electrophoresis and whole genome sequencing of the major clusters, were performed. Bivariate and multivariate analyses were carried out to evaluate the role of virulence genes on outcome. FINDINGS: All isolates harboured the vanA gene. Regarding the virulence genes, 96.5% of isolates were positive for esp, 69.8% for gelE and asa1 genes. VRE infection isolates were more virulent than colonization isolates and harboured more often the gelE gene (P = 0.008). Infections caused by VRE carrying asa1 gene resulted more frequently in death (P = 0.004), but only the predominant clone remained as protector in the multivariate model. The E. faecium strains were assigned to seven STs (ST78, ST412, ST478, ST792, ST896, ST987, ST963) that belonged to CC17. The E. faecalis sequenced belonged to ST9 (CC9). CONCLUSION: E. faecium was predominant, and infection isolates were more virulent than colonization isolates and harboured more often the gene gelE. Infections caused by VRE carrying the asa1 gene appeared to be associated with a fatal outcome.

Changed epidemiology during intra and interhospital spread of high-risk clones of vanA-containing Enterococcus in Brazilian hospitals.

We report changes in the molecular epidemiology of vanA-containing Enterococcus during the intra and interhospital spread of high-risk clones, in Southeastern Brazil. While VRE faecalis predominated during 1998 to 2006, a reversal has been observed in the last years, where VRE faecium belonging to ST114, ST203, ST412, ST478 and ST858 have become endemic.

A polyclonal outbreak of bloodstream infections by Enterococcus faecium in patients with hematologic malignancies.

BACKGROUND: Enterococcus faecium causes bloodstream infection (BSI) in patients with hematologic malignancies (HMs). We studied the clinical features and outcomes of patients with HM with vancomycin-sensitive E faecium (VSE) and vancomycin-resistant E faecium (VRE) BSI and determined the genetic relatedness of isolates and circumstances associated with the upsurge of E faecium BSI. METHODS: Case-control study of patients with HM and E faecium-positive blood culture from January 2008-December 2012; cases were patients with VRE and controls were VSE isolates. The strains were tested for Van genes by polymerase chain reaction amplification and we performed pulsed-field gel electrophoresis to determine genetic relatedness. RESULTS: Fifty-eight episodes of E faecium BSI occurred: 35 sensitive and 23 resistant to vancomycin. Mortality was 46% and 57%, attributable 17% and 40%, respectively. Early stage HM was associated with VSE (P = .044), whereas an episode of BSI within the 3 months before the event (P = .039), prophylactic antibiotics (P = .013), and vancomycin therapy during the previous 3 months (P = .001) was associated with VRE. The VanA gene was identified in 97% of isolates studied. E faecium isolates were not clonal. CONCLUSIONS: E faecium BSI was associated with high mortality. This outbreak of VRE was not clonal; it was associated with antibiotic-use pressure and highly myelosuppressive chemotherapy.