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Coastal water quality is facing increasing threats due to human activities. Their contamination by sewage discharges poses significant risks to the environment and public health. We aimed to investigate the presence of antibiotic-resistant Enterococcus in beach waters. Over a 10-month period, samples were collected from four beaches in the State of São Paulo (Brazil). Enterococcus isolates underwent matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) and molecular analysis for accurate genus and species identification. The antimicrobial susceptibility for 14 antibiotics was evaluated using the disc diffusion method followed by a multidrug-resistance (MDR) classification. PCR amplification method was used to detect antimicrobial resistance genes (ARGs). Our findings revealed the prevalence of Enterococcus faecalis, E. faecium and E. hirae. Out of 130 isolates, 118 were resistant to multiple antibiotics. The detection of resistance genes provided evidence of the potential transfer of antibiotic resistance within the environment. Our findings underscore the necessity for continuous research and surveillance to enhance understanding of the pathogenicity and antimicrobial resistance mechanisms of Enterococcus, which is crucial to implement effective measures to preserve the integrity of coastal ecosystems.

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The species included in the ESKAPE group (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and the genus Enterobacter) have a high capacity to develop antimicrobial resistance (AMR), a health problem that is already among the leading causes of death and could kill 10 million people a year by 2050. The generation of new potentially therapeutic molecules has been insufficient to combat the AMR "crisis", and the World Health Organization (WHO) has stated that it will seek to promote the development of rapid diagnostic strategies. The physicochemical properties of metallic nanoparticles (MNPs) have made it possible to design biosensors capable of identifying low concentrations of ESKAPE bacteria in the short term; other systems identify antimicrobial susceptibility, and some have been designed with dual activity in situ (bacterial detection and antimicrobial activity), which suggests that, in the near future, multifunctional biosensors could exist based on MNPs capable of quickly identifying bacterial pathogens in clinical niches might become commercially available. This review focuses on the use of MNP-based systems for the rapid and accurate identification of clinically important bacterial pathogens, exhibiting the necessity for exhaustive research to achieve these objectives. This review focuses on the use of metal nanoparticle-based systems for the rapid and accurate identification of clinically important bacterial pathogens.

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AIMS: This study aimed to prospect and isolate lactic acid bacteria (LAB) from an artisanal cheese production environment, to assess their safety, and to explore their bacteriocinogenic potential against Listeria monocytogenes. METHODS AND RESULTS: Samples were collected from surfaces of an artisanal-cheese production facility and after rep-PCR and 16S rRNA sequencing analysis, selected strains were identified as to be belonging to Lactococcus garvieae (1 strain) and Enterococcus faecium (14 isolates, grouped into three clusters) associated with different environments (worktables, cheese mold, ripening wooden shelves). All of them presented bacteriocinogenic potential against L. monocytogenes ATCC 7644 and were confirmed as safe (γ-hemolytic, not presenting antibiotic resistance, no mucus degradation properties, and no proteolytic or gelatinase enzyme activity). Additionally, cell growth, acidification and bacteriocins production kinetics, bacteriocin stability in relation to different temperatures, pH, and chemicals were evaluated. According to performed PCR analysis all studied strains generated positive evidence for the presence of entA and entP genes (for production of enterocins A and enterocins P, respectively). However, pediocin PA-1 associated gene was recorded only in DNA obtained from E. faecium ST02JL and Lc. garvieae ST04JL. CONCLUSIONS: It is worth considering the application of these safe LAB or their bacteriocins in situ as an alternative means of controlling L. monocytogenes in cheese production environments, either alone or in combination with other antimicrobials.

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DapE is a Zn2+-metallohydrolase recognized as a drug target for bacterial control. It is a homodimer that requires the exchange of interface strands by an induced fit essential for catalysis. Identifying novel anti-DapE agents requires greater structural details. Most of the characterized DapEs are from the Gram-negative group. Here, two high-resolution DapE crystal structures from Enterococcus faecium are presented for the first time with novel aspects. A loosened enzyme intermediate between the open and closed conformations is observed. Substrates may bind to loose state, subsequently it closes, where hydrolysis occurs, and finally, the change to the open state leads to the release of the products. Mutation of His352 suggests a role, along with His194, in the oxyanion stabilization in the mono-metalated Zn2+ isoform, while in the di-metalated isoform, the metal center 2 complements it function. An aromatic-π box potentially involved in the interaction of DapE with other proteins, and a peptide flip could determine the specificity in the Gram-positive ArgE/DapE group. Finally, details of two extra-catalytic cavities whose geometry changes depending on the conformational state of the enzyme are presented. These cavities could be a target for developing non-competitive agents that trap the enzyme in an inactive state.

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This study aimed to identify and characterize the antimicrobial susceptibility profile of bacteria found in primary endodontic infections in the teeth of patients treated at the Dental Clinic of the University of Ribeirão Preto, São Paulo, Brazil. From September to December 2019, samples were obtained from 21 patients with primary endodontic infections. The collections were carried out in triplicate using paper cones placed close to the total length of the root canal. Bacterial isolation was performed in Brain Heart Infusion agar, Blood agar, and other selective culture media cultured at 37°C for up to 48 h under aerobiosis and microaerophilic conditions. The bacterial species were identified using the Vitek 2 automated system. The disk diffusion method on agar Müeller-Hinton was used to assess antimicrobial susceptibility with the recommended antimicrobials for each identified bacterial species. A total of 49 antibiotics were evaluated. Fifteen of the 21 samples collected showed bacterial growth, and 17 bacterial isolates were found. There were 10 different bacterial species identified: Enterococcus faecalis (four isolates), Streptococcus mitis/oralis (three isolates), Streptococcus anginosus (three isolates) being the most common, followed by Staphylococcus epidermidis, Enterococcus faecium, Streptococcus constellatus, Streptococcus alactolyticus, Enterobacter cloacae, Klebsiella variicola, and Providencia rettgeri (one isolate of each species). The analysis demonstrated significant susceptibility to most of the tested antibiotics. However, some Enterococcus isolates resisted the antibiotic's erythromycin, ciprofloxacin, and tetracycline. A Staphylococcus epidermidis isolate was characterized as multidrug-resistant. Five Streptococcus isolates were non-susceptible to all antibiotics tested.

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Enterococci are ubiquitous microorganisms in almost all environments, from the soil we step on to the food we eat. They are frequently found in naturally fermented foods, contributing to ripening through protein, lipid, and sugar metabolism. On the other hand, these organisms are also leading the current antibiotic resistance crisis. In this study, we performed whole-genome sequencing and comparative genomics of an Enterococcus faecium strain isolated from an artisanal Mexican Cotija cheese, namely QD-2. We found clear genomic differences between commensal and pathogenic strains, particularly in their carbohydrate metabolic pathways, resistance to vancomycin and other antibiotics, bacteriocin production, and bacteriophage and CRISPR content. Furthermore, a bacteriocin transcription analysis performed by RT-qPCR revealed that, at the end of the log phase, besides enterocins A and X, two putative bacteriocins not reported previously are also transcribed as a bicistronic operon in E. faecium QD-2, and are expressed 1.5 times higher than enterocin A when cultured in MRS broth.

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Antimicrobial Resistance (AMR) represents one of the main current threats to global public health; where production animals, companion animals, humans, and the environment play a significant role in its dissemination. However, little attention has been given to companion animals as reservoirs and disseminators of relevant antimicrobial resistant bacteria, especially in South American countries such as Chile. For this reason, this research aimed to estimate the prevalence of AMR to different critical antibiotics at a screening level in commensal bacteria such as E. coli and Enterococcus spp., isolated from healthy pet dogs in the Metropolitan Region of Chile, studying their geographical distribution and evaluating associations of phenotypic resistance to different antibiotics. Thus, in E. coli we detected AMR to all critical drugs assessed, including 34.1% to amoxicillin, 20.1% to colistin, 15.7% to enrofloxacin, and 9.2% to cefotaxime. On the other hand, AMR prevalence in E. faecalis was 8.1% for ampicillin and 3.4% for vancomycin; while for E. faecium the AMR prevalence was 19.1% for ampicillin and 10.2% for vancomycin. Additionally, significant differences in prevalence of the different possible AMR were detected according to their geographical distribution, suggesting the existence of various risk factors and stressing the need to establish mitigation measures specific to the differences identified.

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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.

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Mangrove ecosystems are hotspots of biodiversity, but have been threatened by anthropogenic activities. Vancomycin-resistant enterococci (VRE) are nosocomial bacteria classified as high priority by the World Health Organization (WHO). Herein, we describe the identification and genomic characteristics of a vancomycin-resistant Enterococcus faecalis strain isolated from a highly impacted mangrove ecosystem of the northeastern Brazilian, in 2021. Genomic analysis confirmed the existence of the transposon Tn1546-vanA and clinically relevant antimicrobial resistance genes, such as streptogramins, tetracycline, phenicols, and fluoroquinolones. Virulome analysis identified several genes associated to adherence, immune modulation, biofilm, and exoenzymes production. The UFSEfl strain was assigned to sequence type (ST9), whereas phylogenomic analysis with publicly available genomes from a worldwide confirmed clonal relatedness with a hospital-associated Brazilian clone. Our findings highlight the successful expansion of hospital-associated VRE in a mangrove area and shed light on the need for strengthening genomic surveillance of WHO priority pathogens in these vital ecosystems.

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Probióticos são capazes de melhorar o equilíbrio da microbiota intestinal, trazendo benefícios ao hospedeiro. Atualmente no mercado há poucas opções de alimentos, com probióticos em sua composição, destinados a cães e gatos. Portanto, o objetivo deste trabalho foi desenvolver uma matriz alimentar canina (ração úmida) com o probiótico Enterococcus faecium M7AN10. Para tal, avaliou-se a inocuidade, atividade enzimática, atividade antimicrobiana, potencial probiótico e a viabilidade do microrganismo em matriz alimentar canina. O isolado foi considerado inócuo, pois apresentou ausência de atividade hemolítica e de gelatinase, além de ser suscetível a diversos antimicrobianos. E. faecium M7AN10 apresentou atividade proteolítica e capacidade de produção de exoplissacarídeo. Em relação a atividade antimicrobiana pelo método da estria radial, o isolado inibiu Acinetobacter sp. 1, Corynebacterium sp. 4, Micrococcus luteus 33, Micrococcus luteus 43, Micrococcus sp. 3, Micrococcus sp. 20, Micrococcus sp. 36. Além disso, E. faecium M7AN10 apresentou capacidade de autoagregação de 33,50% e resistiu de forma constante quando submetido ao trato gastrointestinal in vitro em conjunto com Lacticaseibacillus rhamnosus LB 1.5 e Lacticaseibacillus paracasei LB 6.4. O cultivo misto manteve-se viável em matriz alimentar canina durante o período de oito dias. Com base nesses resultados, o isolado E. faecium M7AN10 foi considerada uma bactéria candidata a probiótico que pode vir a ser usada como aditivo em alimento para cães.

Probiotics are capable of improving the balance of the intestinal microbiota, bringing benefits to the host. Currently, on the market, there are few food options with probiotics in their composition intended for dogs and cats. Therefore, this research aimed to develop a canine food matrix (wet food) with the probiotic Enterococcus faecium M7AN10. To this end, the harmlessness, enzymatic activity, antimicrobial activity, probiotic potential, and viability of the microorganism in the canine food matrix were evaluated. The isolate was considered harmless, as it showed no hemolytic and gelatinase activity and was susceptible to several antimicrobials. E. faecium M7AN10 showed proteolytic activity and exopolysaccharide production capacity. Regarding antimicrobial activity using the radial stria method, the isolate inhibited Acinetobacter sp. 1, Corynebacterium sp. 4, Micrococcus luteus 33, Micrococcus luteus 43, Micrococcus sp. 3, Micrococcus sp. 20, Micrococcus sp. 36. Furthermore, E. faecium M7AN10 showed a self-aggregation capacity of 33.50% and resisted consistently when subjected to the gastrointestinal tract in vitro together with Lacticaseibacillus rhamnosus LB 1.5 and Lacticaseibacillus paracasei LB 6.4. The mixed culture remained viable in a canine food matrix over eight days. Based on these results, the isolate E. faecium M7AN10 was considered a candidate bacterium for a probiotic that could be used as an additive in dog food.

Los probióticos son capaces de mejorar el equilibrio de la microbiota intestinal, aportando beneficios al huésped. Actualmente en el mercado existen pocas opciones de alimentos con probióticos en su composición, destinados a perros y gatos. Por lo tanto, el objetivo de este trabajo fue desarrollar una matriz alimentaria canina (comida húmeda) con el probiótico Enterococcus faecium M7AN10. Para ello se evaluó la inocuidad, actividad enzimática, actividad antimicrobiana, potencial probiótico y viabilidad del microorganismo en matriz alimentaria canina. El aislado fue considerado inofensivo, ya que no mostró actividad hemolítica ni gelatinasa, además de ser susceptible a varios antimicrobianos. E. faecium M7AN10 mostró actividad proteolítica y capacidad de producción de exoplisacáridos. En cuanto a la actividad antimicrobiana mediante el método de las estrías radiales, el aislado inhibió a Acinetobacter sp. 1, Corynebacterium sp. 4, Micrococcus luteus 33, Micrococcus luteus 43, Micrococcus sp. 3, Micrococcus sp. 20, Micrococcus sp. 36. Además, E. faecium M7AN10 mostró una capacidad de autoagregación del 33,50% y resistió consistentemente cuando se sometió al tracto gastrointestinal in vitro junto con Lacticaseibacillus rhamnosus LB 1.5 y Lacticaseibacillus paracasei LB 6.4. El cultivo mixto permaneció viable en una matriz de alimento canino durante un período de ocho días. Con base en estos resultados, el aislado E. faecium M7AN10 se consideró una bacteria candidata para un probiótico que podría usarse como aditivo en la comida para perros.

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Enterococcus faecalis has emerged as an important opportunistic pathogen due to its increasing resistance to antimicrobials, mainly to vancomycin, which leads substantial cases of therapeutic failures. Wastewater treatment plants (WWTP), in turn, are considered hotpots in the spread of antimicrobial resistance according to One Health perspective. In this study, we present the first report of a vancomycin-resistant E. faecalis strain recovered from treated effluent in Brazil. For this purpose, the whole-genome sequencing (WGS) was carried out aiming to elucidate its molecular mechanisms of antimicrobial resistance and its phylogenetic relationships amongst strains from other sources and countries. According to Multilocus Sequence Typing (MLST) analysis this strain belongs to ST21. The WGS pointed the presence of vanA operon, multiple antibiotic resistance and virulence genes, and a significant pathogenic potential for humans. The phylogenomic analysis of E. faecalis 6805 was performed with ST21 representatives from the PubMLST database, including the E. faecalis IE81 strain from clinical sample in Brazil, which had its genome sequenced in this study. Our results demonstrated a strain showing resistance to vancomycin in treated effluent. To the best of our knowledge, this is an unprecedented report of vanA-carrying E. faecalis ST21. Furthermore, it is the first description of a vanA-harboring strain of this species from environmental sample in Brazil. Our data highlight the role of WWTP in the spread of AMR, since these environments are favorable for the selection of multidrug-resistant pathogens and the treated effluents, carrying antibiotic resistance genes, are directed to receiving water bodies.

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Litopenaeus vannamei, the Pacific whiteleg shrimp, is one of the most marketable species in aquaculture worldwide. However, it is susceptible to different infections causing considerable losses in production each year. Consequently, using prebiotics that promotes the proliferation of beneficial bacteria and strengthen the immune system is a current strategy for disease control. In this study, we isolated two strains of E. faecium from the gut of L. vannamei fed with agavin-supplemented diets. These isolates showed antibacterial activity against Vibrio parahaemolyticus, Vibrio harveyi and Vibrio alginolyticus, most likely due to peptidoglycan hydrolase (PGH) activity. Furthermore, we sequenced the genome of one isolate. As a result, we observed three proteins related to the production of bacteriocins, a relevant trait for selecting probiotic strains since they can inhibit the invasion of potential pathogens. Additionally, the genome annotation showed genes related to the production of essential nutrients for the host. It lacked two of the most common factors associated with virulence in Enterococcus pathogenic strains (esp and hyl). Thus, this host-probiotic-derived strain has potential application not only in shrimp health but also in alternative aquatic environments, as it is adapted to coexist within the gut shrimp microbiota, independently of the diet.

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Abstract Vanillin is the major component which is responsible for flavor and aroma of vanilla extract and is produced by 3 ways: natural extraction from vanilla plant, chemical synthesis and from microbial transformation. Current research was aimed to study bacterial production of vanillin from native natural sources including sewage and soil from industrial areas. The main objective was vanillin bio-production by isolating bacteria from these native sources. Also to adapt methodologies to improve vanillin production by optimized fermentation media and growth conditions. 47 soil and 13 sewage samples were collected from different industrial regions of Lahore, Gujranwala, Faisalabad and Kasur. 67.7% bacterial isolates produced vanillin and 32.3% were non-producers. From these 279 producers, 4 bacterial isolates selected as significant producers were; A3, A4, A7 and A10. These isolates were identified by ribotyping as A3 Pseudomonas fluorescence (KF408302), A4 Enterococcus faecium (KT356807), A7 Alcaligenes faecalis (MW422815) and A10 Bacillus subtilis (KT962919). Vanillin producers were further tested for improved production of vanillin and were grown in different fermentation media under optimized growth conditions for enhanced production of vanillin. The fermentation media (FM) were; clove oil based, rice bran waste (residues oil) based, wheat bran based and modified isoeugenol based. In FM5, FM21, FM22, FM23, FM24, FM30, FM31, FM32, FM34, FM35, FM36, and FM37, the selected 4 bacterial strains produced significant amounts of vanillin. A10 B. subtilis produced maximum amount of vanillin. This strain produced 17.3 g/L vanillin in FM36. Cost of this fermentation medium 36 was 131.5 rupees/L. This fermentation medium was modified isoeugenol based medium with 1% of isoeugenol and 2.5 g/L soybean meal. ech gene was amplified in A3 P. fluorescence using ech specific primers. As vanillin use as flavor has increased tremendously, the bioproduction of vanillin must be focused.

Resumo A vanilina é o principal componente responsável pelo sabor e aroma do extrato de baunilha e é produzida de três formas: extração natural da planta da baunilha, síntese química e transformação microbiana. A pesquisa atual teve como objetivo estudar a produção bacteriana de vanilina a partir de fontes naturais nativas, incluindo esgoto e solo de áreas industriais. O objetivo principal era a bioprodução de vanilina por meio do isolamento de bactérias dessas fontes nativas. Também para adaptar metodologias para melhorar a produção de vanilina por meio de fermentação otimizada e condições de crescimento. Foram coletadas 47 amostras de solo e 13 de esgoto de diferentes regiões industriais de Lahore, Gujranwala, Faisalabad e Kasur; 67,7% dos isolados bacterianos produziram vanilina e 32,3% eram não produtores. Desses 279 produtores, 4 isolados bacterianos selecionados como produtores significativos foram: A3, A4, A7 e A10. Esses isolados foram identificados por ribotipagem como fluorescência A3 Pseudomonas (KF408302), A4 Enterococcus faecium (KT356807), A7 Alcaligenes faecalis (MW422815) e A10 Bacillus subtilis (KT962919). Os produtores de vanilina foram posteriormente testados para produção aprimorada de vanilina e foram cultivados em diferentes meios de fermentação sob condições de crescimento otimizadas para produção aprimorada de vanilina. Os meios de fermentação (FM) foram: à base de óleo de cravo, à base de resíduos de farelo de arroz (resíduos de óleo), à base de farelo de trigo e à base de isoeugenol modificado. Em FM5, FM21, FM22, FM23, FM24, FM30, FM31, FM32, FM34, FM35, FM36 e FM37, as 4 cepas bacterianas selecionadas produziram quantidades significativas de vanilina. A10 B. subtilis produziu quantidade máxima de vanilina. Essa cepa produziu 17,3 g / L de vanilina em FM36. O custo desse meio de fermentação 36 foi de 131,5 rúpias / L. Esse meio de fermentação foi um meio à base de isoeugenol modificado com 1% de isoeugenol e 2,5 g / L de farelo de soja. O gene ech foi amplificado em A3 P. fluorescence usando primers específicos para ech. Como o uso da vanilina como sabor aumentou tremendamente, a bioprodução da vanilina deve ser focada.

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La aparición de cepas de enterococos resistentes a daptomicina es un tema de preocupación clínica y epidemiológica en años recientes. Se presenta el caso de un paciente de 50 años con antecedente de artritis reumatoide e inmunosupresión crónica hospitalizado en contexto de neumonía viral por COVID-19, con sobreinfección bacteriana y choque séptico, en quien se documentó en tres oportunidades diferentes aislamientos de Enterococcus faecium vancomicino-resistente VAN A y B con falla terapéutica a daptomicina, por deterioro clínico y persistencia de hemocultivos positivos. Se inició manejo con linezolid con control de la infección, negativización de hemocultivos y evolución clínica satisfactoria. Se realiza reporte del presente caso para dar a conocer la aparición de enterococos resistentes a daptomicina, la cual es una creciente preocupación epidemiológica, con el fin de realizar identificación temprana, prevenir falla terapéutica y poder conocer la epidemiología local

n recent years, the emergence of daptomycin-resistant enterococcus strains is a growing clinical and epidemio-logical topic of concern. We report the case of a 50 year old patient with an antecedent of rheumatoid arthritis and chronic immunosuppression hospitalized in the con-text of COVID-19 pneumonia with bacterial co-infection and septic shock in which a three different moments an isolate of a "vancomycin-resistant enterococcus faecium"(VRE) Van A and B that presented therapeutic failure with daptomycin was documented after clinical deterioration and persistence of positive blood cultures. Linezolid was initiated, with clinical recovery and negativization of blood cultures following the change in antibiotic treatment. This case is reported in order to expose the ever growing con-cern of daptomycin-resistant enterococcus strains in or-der to prevent therapeutic failure, make early identifica-tion and get to know the local epidemiological status.

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BACKGROUND: Domestic pigeons carry pathogens in their droppings, posing a potential public health problem. METHODS: The phenotypic and genotypic antimicrobial resistances of Staphylococcus aureus and Enterococcus faecium in the feces of urban pigeons near hospitals with intensive care units were measured. RESULTS: Twenty-nine samples showed Enterococcus growth, whereas one was positive for S. aureus. The S. aureus isolate was sensitive to the antibiotics tested via antibiogram, however resistance genes were identified. E. faecium isolates showed phenotypic resistance to gentamicin, erythromycin, and ciprofloxacin. CONCLUSIONS: Antimicrobial profiles harmful to health were demonstrated in bacterial pathogens isolated from the external environment of hospitals.

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In this study, we evaluated the antimicrobial susceptibility, the presence of gene-encoding virulence factors and CRISPR systems, as well as the ability to produce lytic enzymes among clinical E. faecalis and E. faecium isolates (n = 44). All enterococci isolates showed phenotypes of multidrug resistance. E. faecalis and E. faecium isolates exhibited high-level aminoglycoside resistance phenotype, several of them harboring the aac(6')Ie-aph(2″)Ia and aph(3')-IIIa genes. The gene vanA was the most frequent among vancomycin-resistant E. faecium. High prevalence of the virulence genes esp and efaA were observed; hyl gene was more associated with E. faecium, while ace and efaA genes were more frequently detected in E. faecalis. Caseinase activity was frequently detected among the isolates. Gelatinase and DNAse activities predominated among E. faecalis, while hemolytic capability was frequent among E. faecium isolates. Twenty-nine isolates showed at least one CRISPR system investigated. Several enterococci isolates harbored the aac(6')-Ie-aph(2″)-Ia or aph(3')-IIIa genes and a CRISPR loci. CRISPR loci were positively correlated to efaA and gelE genes, and gelatinase and DNAse activities, while CRISPR loci absence was related to hyl gene presence. These results show that clinical isolates of E. faecalis and E. faecium harboring virulence genes show the concomitant presence of CRISPR loci and antibiotic resistance determinants.

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The conditions of aquatic environments have a great influence on the microbiota of several animals, many of which are a potential source of microorganisms of biotechnological interest. In this study, bacterial strains isolated from aquatic environments were bioprospected to determine their probiotic profile and antimicrobial effect against fish and food pathogens. Two isolates, identified via 16S rRNA sequencing as Lactococcus lactis (L1 and L2) and one as Enterococcus faecium 135 (EF), produced a bacteriocin-like antimicrobial substance (BLIS), active against Listeria monocytogenes, Salmonella Choleraesuis and Salmonella Typhimurium. Antimicrobial activity of BLIS was reduced when exposed to high temperatures and proteolytic enzymes (trypsin, pepsin, papain and pancreatin). All strains were sensitive to 7 types of antibiotics (vancomycin, clindamycin, streptomycin, gentamicin, chloramphenicol, rifampicin and ampicillin), exhibited a high rate of adherence to Caco-2 cells and expressed no hemolysin and gelatinase virulence factors. EF showed some resistance at pH 2.5 and 3.0, and L2/EF showed higher resistance to the action of bile salts. Finally, the presence of bacteriocin genes encoding for proteins, including Nisin (L1 and L2), Enterocin A, B, P, and Mundticin KS (EF) was detected. The molecular and physiological evidence suggests that the bacterial isolates in this study could be used as natural antimicrobial agents and may be considered safe for probiotic application.

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Aloe vera has been medicinally used for centuries. Its bioactive compounds have been shown to be very effective in the treatment of numerous diseases. In this work, a novel functional beverage was developed and characterized to combine the health benefits of probiotic bacteria with the Aloe vera plant itself. Two Aloe vera juices were obtained by fermentation either by a novel isolated Enterococcus faecium or a commercial Lactococcus lactis. The extraction of Aloe vera biocompounds for further fermentation was optimized. Extraction with water plus cellulase enhanced the carbohydrates and phenolic compounds in the obtained extracts. The biotransformation of the bioactive compounds from the extracts during fermentation was assessed. Both probiotic bacteria were able to grow on the Aloe vera extract. Lactic acid and short-chain fatty acids (SCFA) together with fourteen individual phenolic compounds were quantified in the produced Aloe vera juice, mainly epicatechin, aloin, ellagic acid, and hesperidin. The amount of total phenolic compounds was maintained through fermentation. The antioxidant activity was significantly increased in the produced juice by the ABTS method. The novel produced Aloe vera juice showed great potential as a functional beverage containing probiotics, prebiotics, SCFA, and phenolic compounds in its final composition.

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