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1.
Braz. j. biol ; 83: e250550, 2023. tab, graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1345536

RESUMO

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.


Assuntos
Benzaldeídos/metabolismo , Aromatizantes/metabolismo , Bacillus subtilis/metabolismo , Microbiologia Industrial , Pseudomonas fluorescens/metabolismo , Enterococcus faecium/metabolismo , Meios de Cultura , Alcaligenes faecalis/metabolismo , Fermentação
2.
Stem Cell Res Ther ; 13(1): 446, 2022 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-36056447

RESUMO

BACKGROUND: Bone has important functions in the body. Several researchers have reported that the polysaccharides and lipopolysaccharide derived from microbes can promote osteogenic differentiation of stem cells. Enterococcus faecium, a lactic acid bacterium (LAB), produces several bioactive metabolites and has been widely applied in the food and nutraceutical industries. The exopolysaccharide (EPS) from LAB has also been extensively examined for its postbiotic effects and for its in vivo and in vitro functionalities. However, studies on promoting bone differentiation using polysaccharides from LAB are lacking. Therefore, the purpose of this study was to investigate the effect of E. faecium L15 extract and EPS on osteogenic differentiation of human dental pulp stem cells (hDPSCs) and to identify the underlying mechanisms. METHODS: hDPSCs were obtained from dental pulp tissue, and L15 extract and EPS were isolated from L15. Gene and protein expression of the osteogenic differentiation markers were analyzed with qPCR and western blotting and the possible signaling pathways were also investigated using western blotting. Osteogenic differentiation potential was examined by alkaline phosphatase (ALP) staining and alizarin red s (ARS) staining. In addition, osteogenic differentiation potential of L15 EPS was explored in ex vivo culture of neonate murine calvaria. RESULTS: The calcium deposition and ALP activity were enhanced by addition of L15 extract or EPS. The expression levels of RUNX2, ALP, and COL1A1 mRNA and the protein expression levels of RUNX2, ALP, and BMP4 were increased in hDPSCs treated with the L15 extract or EPS. The L15 EPS treatment enhanced phosphorylation of the p38 mitogen-activated protein kinase (MAPK). The L15 EPS-induced increases in RUNX2, ALP, and BMP4 expression were suppressed by the p38 MAPK inhibitor SB203580. The promoting effect of L15 EPS on osteogenic differentiation was not only seen in hDPSCs, but also in osteoblast precursors. ALP activity and the expression of RUNX2, ALP, and COL1A1 increased in the L15 EPS-treated osteoblast precursors. In addition, L15 EPS increased bone thickness of neonate murine calvaria in ex vivo culture. CONCLUSIONS: The stimulatory effect of L15 extract and EPS on osteogenic differentiation occurred through the p38 MAPK pathway, and L15 EPS enhanced new bone formation in neonate murine calvaria. These data suggest that L15 EPS has therapeutic potential applicable to bone regeneration.


Assuntos
Enterococcus faecium , Osteogênese , Animais , Diferenciação Celular , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Polpa Dentária/metabolismo , Enterococcus faecium/metabolismo , Humanos , Recém-Nascido , Camundongos , Osteogênese/genética , Células-Tronco/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
3.
Arch Microbiol ; 204(10): 619, 2022 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-36098848

RESUMO

Probiotic attributes of lactic acid bacteria isolated from goat and sheep milk samples were analysed by culturing them on an MRS agar medium. The most potential isolates, GMB24 and SMB16, were identified by biochemical tests which had ability to tolerate different concentrations of acid and bile and phenol resistance. They were further identified as Enterococcus faecium GMB24 and Enterococcus hirae SMB16 by 16S rRNA gene sequencing approach. The probiotic potential of the isolates GMB24 and SMB16 were recorded including antimicrobial activity against pathogenic bacteria viz., Escherichia coli (MTCC118), Staphylococcus aureus (MTCC7443), Pseudomonas aeruginosa (MTCC424), Listeria monocytogens (MTCC657) and Salmonella typhimurium (MTCC733), and antibiotic susceptibility test. The isolates SMB16 and GMB24 exhibited a higher zone of inhibition against P. aeruginosa (19.00 ± 0.57 mm) and S. aureus (25.66 ± 0.88 mm), respectively. The data from these experiments were used for the principal component analysis (PCA) to assess the survivability of the isolates under different factors. The heatmap generated in this study clustered the bacterial isolates based on their phenotype properties. Further, immunomodulating activities of these probiotic bacteria were tested on neutrophil adhesion test, haemagglutinating antibody titer and delayed-type hypersensitivity. Probiotic E. faecium GMB24 and E. hirae SMB16, at 109 cells/mL doses per day, increased the neutrophil adhesion, haemagglutinating antibody titer and DTH in comparison to the untreated control group. The isolates showed negative test for haemolytic and gelatinase activities and hence were considered safe. E. faecium GMB24 and E. hirae SMB16 were shown to have high probiotic potential and immune-stimulant action.


Assuntos
Enterococcus faecium , Probióticos , Animais , Enterococcus faecium/genética , Streptococcus faecium ATCC 9790/genética , Cabras , Leite/microbiologia , Probióticos/farmacologia , RNA Ribossômico 16S/genética , Ovinos , Staphylococcus aureus/genética
4.
Microb Genom ; 8(9)2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-36129737

RESUMO

Enterococcus faecium is a ubiquitous opportunistic pathogen that is exhibiting increasing levels of antimicrobial resistance (AMR). Many of the genes that confer resistance and pathogenic functions are localized on mobile genetic elements (MGEs), which facilitate their transfer between lineages. Here, features including resistance determinants, virulence factors and MGEs were profiled in a set of 1273 E. faecium genomes from two disparate geographic locations (in the UK and Canada) from a range of agricultural, clinical and associated habitats. Neither lineages of E. faecium, type A and B, nor MGEs are constrained by geographic proximity, but our results show evidence of a strong association of many profiled genes and MGEs with habitat. Many features were associated with a group of clinical and municipal wastewater genomes that are likely forming a new human-associated ecotype within type A. The evolutionary dynamics of E. faecium make it a highly versatile emerging pathogen, and its ability to acquire, transmit and lose features presents a high risk for the emergence of new pathogenic variants and novel resistance combinations. This study provides a workflow for MGE-centric surveillance of AMR in Enterococcus that can be adapted to other pathogens.


Assuntos
Anti-Infecciosos , Enterococcus faecium , Saúde Única , Enterococcus faecium/genética , Humanos , Fatores de Virulência/genética , Águas Residuárias
5.
Microb Pathog ; 171: 105745, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-36057414

RESUMO

This study aimed to investigate the presence of eight virulence genes (ace, asa1, esp, efaA, gelE, cylA, agg, fsr) in Enterococcus from a variety of animals and to explore the drug resistance and pathogenicity. This could provide a theoretical basis for clinical treatment of Enterococcus infections. Anal swabs from pigs, chickens, cattle, and dogs in farms and pet hospitals were collected for Enterococcus isolation and identification. Eight virulence genes were detected (PCR method), and drug resistance was assessed (drug-sensitive paper method). The strains containing different virulence genes were then divided into EV1, EV2, and EV3 groups. The LD50 and pathogenicity was examined by intra-peritoneal injection to infect mice. Differences were found in the detection rates of virulence genes in Enterococcus from the different animals. The highest overall detection rate was for the esp gene (78.0%), and the lowest for the cylA gene (15.5%). Eight genes were detected most frequently in Enterococcus from dogs and least frequently from cattle. Among the Enterococcus strains from four variety of animals, drug resistance was highest against sulfamethoxazole (100%), cefotaxime (>97%), and cefotaxitin (>93%). Drug resistance was lowest against vancomycin (0%), levofloxacin (<12%) and ciprofloxacin (<13%). The LD50 for each of the three groups was EV1LD50=8.71×109CFU, EV2LD50=2.34×1010CFU,and EV3LD50=9.33×1010CFU. The Enterococcus12LD50 dose group caused significant clinical symptoms in mice, with pathological effects on the heart, liver, lungs, and kidneys, and particularly on the urinary system. The abundance of Enterococcus virulence genes, drug resistance, and pathogenicity vary among different animal origins, and the pathology caused by Enterococcus requires effective treatment protocols based on species and regional characteristics.


Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Animais , Animais Domésticos , Antibacterianos/farmacologia , Bovinos , Cefotaxima/farmacologia , Galinhas , Ciprofloxacina/farmacologia , Cães , Resistência a Medicamentos , Farmacorresistência Bacteriana/genética , Enterococcus , Enterococcus faecalis , Infecções por Bactérias Gram-Positivas/veterinária , Levofloxacino/farmacologia , Camundongos , Testes de Sensibilidade Microbiana , Sulfametoxazol/farmacologia , Suínos , Vancomicina/farmacologia , Virulência/genética , Fatores de Virulência/genética
6.
Food Microbiol ; 108: 104101, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36088116

RESUMO

Four batches of Cebreiro-type cheese were made in duplicate from pasteurized milk. A control batch was manufactured with only a commercial O-starter. The other three batches were made with the same starter plus: (i) a commercial culture of Enterococcus faecium; (ii) a selected Kluyveromyces lactis adjunct culture used in a cheese-milk pre-ripening step; and (iii) the combination of both adjunct cultures. The cheeses made with the yeast adjunct were characterized by higher values of overall proteolysis, pH and aw, and showed total and lactic acid bacteria (LAB) counts at least 2 log units than the batches made with only LAB. The volatile profiles of the cheeses made with added K. lactis were distinguished by high contents of esters, branched-chain alcohols, fatty acids, acetoin and 2-pnenylethanol. These batches had a more friable and sticky texture, and exhibited differential piquant, yeasty, alcoholic, acetic and fruity flavors. Furthermore, the addition of enterococci seemed to help achieve more desirable sensory characteristics. The batches manufactured with both adjunct cultures were awarded the highest scores for texture preference, flavor intensity, flavor preference, and overall sensory preference. The sensory profiles of the cheeses made with added yeast closely resembled those of traditional 'good quality' raw-milk Cebreiro cheese.


Assuntos
Queijo , Enterococcus faecium , Kluyveromyces , Queijo/microbiologia , Leveduras
7.
Eur J Clin Microbiol Infect Dis ; 41(10): 1245-1261, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-36057762

RESUMO

The aim of our study was to characterize the epidemiological situation concerning nosocomial vancomycin-resistant Enterococcus faecalis of VanA-phenotype (VREfs-VanA) in Poland by investigating their clonal relationships and the vanA-associated mobilome. One-hundred twenty-five clinical isolates of VREfs-VanA collected between 2004 and 2016 were studied by phenotypic assays, multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), PCR detection of plasmid-specific genes, and Tn1546 structure and localization mapping. Selected isolates were subjected to PFGE-S1, Southern hybridization, genomic sequencing and conjugation experiments. The majority of isolates (97.6%) belonged to clonal complexes CC2 and CC87 of E. faecalis. All isolates were resistant to vancomycin and teicoplanin, and resistance to ciprofloxacin and aminoglycosides (high level) was very prevalent in this group. VanA phenotype was associated with 16 types of Tn1546, carrying insertion sequences IS1216, ISEfa4, IS1251 and IS1542, located on repUS1pVEF1, rep1pIP501, rep2pRE25, rep9pAD1/pTEF2/pCF10 and rep6pS86 replicons. The most common Tn1546 B- and BB-type transposons, harbouring one or two copies of IS1216, were inserted between rep18ap200B and repUS1pVEF1 genes and located on ~ 20 kb and 150-200 kb plasmids. VREfs-VanA in Poland represent a polyclonal group, indicating a number of acquisitions of the vanA determinant. The repUS1pVEF1-vanA plasmids, unique for Poland, were the main factor beyond the acquisition of vancomycin resistance by E. faecalis, circulating in Polish hospitals.


Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Aminoglicosídeos , Proteínas de Bactérias/genética , Carbono-Oxigênio Ligases/genética , Ciprofloxacina , Elementos de DNA Transponíveis , Enterococcus faecalis/genética , Enterococcus faecium/genética , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Hospitais , Humanos , Tipagem de Sequências Multilocus , Polônia/epidemiologia , Teicoplanina , Vancomicina
8.
Anal Bioanal Chem ; 414(24): 7179-7189, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35962141

RESUMO

Early detection of bacterial transmission and outbreaks in hospitals is important because nosocomial infections can result in health complications and longer hospitalization. Current practice to detect outbreaks uses genotyping methods amplified fragment length polymorphism (AFLP) and whole genome sequencing (WGS), which are not suitable methods for real-time transmission screening of both susceptible and resistant bacteria. The aim was to assess the typing technique Fourier transform infrared (FTIR) spectroscopy as real-time screening method to discriminate large amounts of susceptible and resistant bacteria at strain level when there is no evident outbreak in comparison with the WGS reference. Isolates of past hospital outbreak strains of Acinetobacter baumannii/calcoaceticus complex (n = 25), Escherichia coli (n = 31), Enterococcus faecium (n = 22), Staphylococcus aureus (n = 37) and Pseudomonas aeruginosa (n = 30) were used for validation of FTIR. Subsequently, Enterococcus faecalis (n = 106) and Enterococcus faecium (n = 104) isolates from weekly routine screening samples when no potential outbreak was present were analysed. FTIR showed reproducibility and congruence of cluster composition with WGS for A. baumannii/calcoaceticus complex and E. faecium outbreak isolates. The FTIR results of E. faecalis and E. faecium isolates from routine samples showed reproducibility, but the congruence of cluster composition with WGS was low. For A. baumannii/calcoaceticus complex and E. faecium outbreak isolates, FTIR appears to be a discriminatory typing tool. However, our study shows the discriminatory power is too low to screen real-time for transmission of E. faecium and E. faecalis at patient wards based on isolates acquired in routine surveillance cultures when there is no clear suspicion of an ongoing outbreak.


Assuntos
Infecção Hospitalar , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Análise do Polimorfismo de Comprimento de Fragmentos Amplificados , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/genética , Infecção Hospitalar/microbiologia , Enterococcus faecium/genética , Genoma Bacteriano , Genótipo , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/genética , Infecções por Bactérias Gram-Positivas/microbiologia , Hospitais , Humanos , Reprodutibilidade dos Testes , Espectroscopia de Infravermelho com Transformada de Fourier , Sequenciamento Completo do Genoma/métodos
9.
Microb Pathog ; 171: 105715, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35973648

RESUMO

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.


Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Aminoglicosídeos , Antibacterianos/farmacologia , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Desoxirribonucleases/genética , Farmacorresistência Bacteriana/genética , Enterococcus/genética , Enterococcus faecalis , Gelatinases , Infecções por Bactérias Gram-Positivas/epidemiologia , Humanos , Canamicina Quinase/genética , Testes de Sensibilidade Microbiana , Vancomicina , Virulência/genética , Fatores de Virulência/genética
10.
Pak J Pharm Sci ; 35(4): 1125-1134, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-36008911

RESUMO

Lactic acid bacteria are microorganisms that can be present in meat, milk and fermented products, as well as fermented drinks and vegetables, and they inhibit the growth of pathogenic and decaying microorganisms. Our aim was isolating, screening, characterization and estimation of bacteriocin/s of Lactic acid bacteria isolated from Horses milk in Jeddah city, Saudi Arabia. That have antagonistic activity against a wide range of pathogenic bacteria. On MRS agar medium 16 LAB isolates were examined using the agar well diffusion method against the pathogenic bacteria (Gram-positive bacteria: S. aureus ATCCBAA977, ST. Pneumonia ATCC49619) and (Gram-negative bacteria: E. coli ATCC35218, P. aeruginosa ATCC27853). The results showed that 12 isolates out of 16 isolates (75%) worked to inhibit all types of pathogenic bacteria used. The best isolate was identified using molecular methods; the results showed that the isolate (H6) matched 100% with (Enterococcus faecium LR135401.1). This isolate produced the highest bacteriocin at 35°C/24 h/pH (6.5).


Assuntos
Bacteriocinas , Enterococcus faecium , Lactobacillales , Ágar , Animais , Antibacterianos/farmacologia , Bacteriocinas/farmacologia , Escherichia coli , Cavalos , Leite/microbiologia , Staphylococcus aureus
11.
Rev Soc Bras Med Trop ; 55: e0353, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36000617

RESUMO

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.


Assuntos
Enterococcus faecium , Animais , Antibacterianos/farmacologia , Columbidae/microbiologia , Enterococcus faecium/genética , Hospitais , Testes de Sensibilidade Microbiana , Staphylococcus aureus/genética
12.
FEMS Microbiol Lett ; 369(1)2022 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-35922088

RESUMO

Daptomycin is a cyclic lipopeptide used in the treatment of vancomycin-resistant Enterococcus faecium (VREfm). However, the development of daptomycin-resistant VREfm challenges the treatment of nosocomial VREfm infections. Resistance mechanisms of daptomycin are not fully understood. Here, we analyzed the genomic changes leading to a daptomycin-susceptible VREfm isolate becoming resistant after 50 days of daptomycin and linezolid combination therapy. A total of seven isogenic VREfm isolates from the same patient (daptomycin-susceptible and daptomycin-resistant) were analyzed using Illumina whole genome sequencing, and two isolates were further characterized with Nanopore sequencing. One nonsynonymous SNP in the rpoC gene previously shown to harbor mutations in daptomycin-resistant VREfm was identified in the daptomycin-resistant isolates. Whole genome comparative analysis identified the loss of a 46.5 kb fragment, duplication of a 29.7 kb fragment, and integration of two plasmids upon acquisition of daptomycin resistance. Transmission electron microscopy showed similar alterations in cell morphology and cell wall structure as have previously been described in daptomycin-resistant E. faecalis.


Assuntos
Infecção Hospitalar , Daptomicina , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Enterococos Resistentes à Vancomicina , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Daptomicina/farmacologia , Enterococcus faecium/genética , Genômica , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Humanos , Testes de Sensibilidade Microbiana , Vancomicina/farmacologia , Enterococos Resistentes à Vancomicina/genética
13.
Gut Microbes ; 14(1): 2106105, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35921516

RESUMO

The probiotic bacterial strain Enterococcus faecium SF68 has been shown to alleviate symptoms of intestinal inflammation in human clinical trials and animal feed supplementation studies. To identify factors involved in immunomodulatory effects on host cells, E. faecium SF68 and other commensal and clinical Enterococcus isolates were screened using intestinal epithelial cell lines harboring reporter fusions for NF-κB and JNK(AP-1) activation to determine the responses of host cell innate immune signaling pathways when challenged with bacterial protein and cell components. Cell-free, whole-cell lysates of E. faecium SF68 showed a reversible, inhibitory effect on both NF-κB and JNK(AP-1) signaling pathway activation in intestinal epithelial cells and abrogated the response to bacterial and other Toll-like receptor (TLR) ligands. The inhibitory effect was species-specific, and was not observed for E. avium, E. gallinarum, or E. casseliflavus. Screening of protein fractions of E. faecium SF68 lysates yielded an active fraction containing a prominent protein identified as arginine deiminase (ADI). The E. faecium SF68 arcA gene encoding arginine deiminase was cloned and introduced into E. avium where it conferred the same NF-κB inhibitory effects on intestinal epithelial cells as seen for E. faecium SF68. Our results indicate that the arginine deiminase of E. faecium SF68 is responsible for inhibition of host cell NF-κB and JNK(AP-1) pathway activation, and is likely to be responsible for the anti-inflammatory and immunomodulatory effects observed in prior clinical human and animal trials. The implications for the use of this probiotic strain for preventive and therapeutic purposes are discussed.


Assuntos
Enterococcus faecium , Microbioma Gastrointestinal , Probióticos , Animais , Enterococcus faecium/genética , Humanos , Hidrolases , Imunidade Inata , NF-kappa B/genética , Probióticos/farmacologia , Probióticos/uso terapêutico , Transdução de Sinais , Fator de Transcrição AP-1/genética , Fatores de Virulência/genética
14.
Chem Biol Interact ; 365: 110086, 2022 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-35940279

RESUMO

The emergence and spread of multidrug-resistant (MDR) enterococci and other Gram-positive bacteria represents a severe problem due to the lack of effective therapeutic alternatives. Natural products have long been an important source of new antibacterial scaffolds and can play a key role in the current antibiotic crisis. Enterococci are predominantly non-pathogenic gastrointestinal commensal bacteria, but among them, Enterococcus faecalis and Enterococcus faecium represent the species that account for most clinically relevant infections. The emergence of MDR enterococci has reduced the available antibiotic treatment options and highlights the need to develop new antimicrobial compounds. In the search for new hit compounds against MDR Enterococcus spp., natural-derived compounds represent inspiring scaffolds for drug design studies. In this work, the antimicrobial activity of a fully synthetic chalcone derivative (r4MB) was determined on a clinical panel of 34 MDR Gram-positive bacteria, mostly constituted by E. faecalis and E. faecium, along with Staphylococcus spp., amongst others. Compound r4MB showed activity against 100% of the tested strains, with the minimum inhibitory concentration (MIC) in the range of 5-20 µM. The lethal action of the compound was evaluated using different fluorescent-based assays. The compound showed a time-dependent permeabilisation of the membrane of a vancomycin-resistant E. faecalis, detected by the fluorescent probe SYTOX Green, and digital fluorescent microscopy corroborated the spectrofluorimetric analysis within 6 min of incubation. Flow cytometry analysis of the membrane electric potential demonstrated a significant depolarization, confirming the target of the compound towards the bacterial membrane. No cytotoxic haemolysis was observed with mammalian erythrocytes, and a 99% cytotoxic concentration of 118 µM on NCTC cells demonstrated a promising antimicrobial selectivity. In silico studies using SwissADME and ADMETLabs servers suggest that compound r4MB displayed adequate ADME properties, with no alerts for pan-assay interference compounds (PAINS). Future hit-to-lead optimization of this chalcone derivative can contribute to developing a more potent derivative against infections caused by MDR enterococci.


Assuntos
Chalcona , Chalconas , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Chalcona/farmacologia , Chalconas/farmacologia , Chalconas/uso terapêutico , Enterococcus , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Mamíferos , Testes de Sensibilidade Microbiana , Permeabilidade
15.
Mikrobiyol Bul ; 56(3): 525-533, 2022 Jul.
Artigo em Turco | MEDLINE | ID: mdl-35960242

RESUMO

Colonized surfaces, equipment, individuals, and infected patients can be sources for the hospital spread of vancomycin-resistant enterococci (VRE), which is one of the important nosocomial pathogens. The basic epidemiological tools for the prevention and control of hospital-acquired infections are the typing methods. Pulsed-field gel electrophoresis (PFGE) is a highly discriminating method used frequently to detect clonal associations in epidemics and hospital-acquired VRE infections. This study aimed to investigate the presence of cross-contamination, which service or services come to the forefront in case of possible cross-contamination and clonal relationship between VRE strains isolated from rectal swab, clinical and environmental swab samples taken in two different periods in various clinics of Istanbul University Istanbul Medical Faculty Hospital by PFGE method. A total of 125 VREs isolated in two different periods were included in the study. Rectal and environmental swab samples were inoculated on Enterococcosel agar and sodium azide broth, urine samples were inoculated on chromogenic agar, and other clinical samples were inoculated on 5% sheep blood agar and incubated for 18-24 hours. VITEK 2 Compact automation system GP panel (bioMerieux, MarcyL'Etoile, France) was used for the species identification of catalase-negative, gram-positive colonies and disc diffusion and minimum inhibitory concentration (MIC) gradient tests were used to determine vancomycin susceptibility. Multiplex polymerase chain reaction was used to search for vanA and vanB resistance genes in isolates identified as VRE, and PFGE was used to determine clonal association. All isolates were identified as Enterococcus faecium with the vanA resistance gene. It was shown that PFGE clones were divided into six types with 65% similarity (A-F), and in this polyclonal spread, the major type was type A, which contained 108 isolates in 37 subtypes existed in the hospital for years. In patients from whom similar isolates were obtained, the high rate of hospitalizations in the same emergency room or in different emergency services in the same building drew attention. Our results showed that the presence of VRE was established in our hospital, new isolates were added from time to time, and there was a cross-contamination. It was observed that emergency services, where infection control measures were neglected due to working conditions, were among the high-risk areas for VRE contamination. In order to better understand the importance of emergency services in cross-contamination, it would be useful to conduct surveillance studies among patients hospitalized in emergency services and monitor the rate of VRE in the services where positive patients were transferred.


Assuntos
Infecção Hospitalar , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Enterococos Resistentes à Vancomicina , Ágar/uso terapêutico , Proteínas de Bactérias/genética , Infecção Hospitalar/tratamento farmacológico , Eletroforese em Gel de Campo Pulsado , Serviço Hospitalar de Emergência , Enterococcus faecium/genética , Infecções por Bactérias Gram-Positivas/diagnóstico , Humanos , Enterococos Resistentes à Vancomicina/genética
16.
Res Vet Sci ; 151: 42-46, 2022 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-35853330

RESUMO

This study investigated the prevalence and antimicrobial susceptibility of enterococci isolated from dogs and cats with urinary tract infections in northwestern Croatia. During this study, the laboratory received 787 urine samples, 651 from dogs and 136 from cats. A total of 260 urine samples (211 from dogs and 49 from cats) were bacteriologically positive. Of these, 29 isolates belonged to Enterococcus spp.; 22 from dogs and seven from cats. Enterococci isolates were identified by PCR method, 12 of which were Enterococcus faecium and 17 were Enterococcus faecalis species. In dogs, 16 E. faecalis and six E. faecium strains were identified, whereas in cats, six E. faecium and only one E. faecalis strain were identified. Antimicrobial susceptibility was determined by the Kirby-Bauer disk diffusion method for nine antimicrobials: penicillin, ampicillin, vancomycin, nitrofurantoin, rifampicin, tetracycline, chloramphenicol, enrofloxacin, ciprofloxacin. The isolates were tested for high-level resistance to streptomycin and gentamicin. The highest resistance of Enterococcus spp. was observed to rifampicin (86%) and enrofloxacin (83%), followed by tetracycline and ciprofloxacin (69%). Resistance to vancomycin was 28%, and the lowest resistance was to chloramphenicol (17%). Multidrug resistance was found in 76% of enterococci isolates. High-level streptomycin resistance was detected in 17% and high-level gentamicin resistance in 10% of the isolated enterococci. When comparing species susceptibility, E. faecium isolates were significantly more resistant to penicillin, ampicillin, nitrofurantoin, and ciprofloxacin (p < 0.05). Eleven E. faecium isolates (92%) and 12 E. faecalis isolates (76%) were multidrug resistant.


Assuntos
Anti-Infecciosos , Doenças do Gato , Doenças do Cão , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Ampicilina , Animais , Antibacterianos/farmacologia , Doenças do Gato/tratamento farmacológico , Doenças do Gato/epidemiologia , Gatos , Cloranfenicol , Ciprofloxacina , Croácia/epidemiologia , Doenças do Cão/epidemiologia , Cães , Farmacorresistência Bacteriana , Enrofloxacina , Enterococcus , Gentamicinas , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/veterinária , Testes de Sensibilidade Microbiana/veterinária , Nitrofurantoína , Penicilinas , Prevalência , Rifampina , Estreptomicina , Tetraciclina , Vancomicina
17.
J Antibiot (Tokyo) ; 75(9): 498-508, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35896611

RESUMO

Eravacycline (Erava) is a synthetic fluorocycline with potent antimicrobial activity against a wide range of Gram-positive bacteria. This study aimed to investigate the in vitro antimicrobial activity and resistance mechanism of Erava in clinical E. faecium isolates from China. Erava minimum inhibitory concentrations (MICs) against clinical E. faecium isolates-including those resistant to linezolid (LZD) or harboring the tetracycline (Tet) resistance genes was ≤0.25 mg l-1. Moreover, our data indicated that clinical isolates of E. faecium with Erava MIC 0.25 mg l-1 were predominantly shown to belong to Sequence-type 78 (ST78) and ST80. The prevalence of Erava heteroresistance in clinical E. faecium strain was 2.46% (3/122). The increased Erava MIC values of heteroresistance-derived E. faecium clones could be significantly reduced by efflux pump inhibitors (EPIs). Furthermore, comparative proteomics results showed that efflux pumps lmrA, mdlA, and mdlB contributed significantly to the acquisition of Erava resistance in E. faecium. In addition, a genetic mutation in 16 S rRNA (G190A) were detected in resistant E. faecium isolates induced by Erava. In summary, Erava exhibits potent in vitro antimicrobial activity against E. faecium, but mutation of Tet target sites and elevated expression of efflux pumps under Erava selection results in Erava resistance.


Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Antibacterianos/farmacologia , Farmacorresistência Bacteriana/genética , Enterococcus faecalis , Humanos , Testes de Sensibilidade Microbiana , Tetraciclinas/farmacologia
18.
Bratisl Lek Listy ; 123(8): 543-549, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35852503

RESUMO

BACKGROUND: Enterococcus species account for most of the human enterococcal HAI and multidrug-resistant infections and have become a major threat to modern public health. We examine the rise in the number of vancomycin resistant E. faecium blood stream and urinary tract infections in a COVID-19 department during an epidemiologic outbreak investigation to detect and eliminate nosocomial clusters of the bacteria. METHODS:  Strain identification was performed by classical isolation and biochemical and cultivation methods. Antibiotic testing results were interpreted according to European committee on antimicrobial susceptibility testing (EUCAST) guidelines. Six isolated samples underwent the whole genome sequencing (WGS) during the outbreak investigation. Isolate relatedness was determined using the core genome multi-locus sequence typing. RESULTS:  WGS revealed two genotypically distinct VRE clusters, one of which had genetically closely related patients and environmental isolates. The cluster was terminated by enhanced infection control strategies. CONCLUSIONS:  This study provides the first description of an outbreak caused by vanB-ST117 and vanA-ST17 E. faecium strains among COVID-19 patients in Slovakia. This study can help to raise the awareness about the need for strict adherence to infection control measures and the implementation of rational antimicrobial stewardship as a routine part of COVID-19 management (Tab. 3, Fig. 3, Ref. 27). Text in PDF www.elis.sk Keywords: vancomycin-resistant Enterococcus faecium, antibiotic resistant, COVID-19, SARS-CoV-2, bacterial outbreak, healthcare-associated infection.


Assuntos
COVID-19 , Infecção Hospitalar , Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Enterococos Resistentes à Vancomicina , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , COVID-19/epidemiologia , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , Surtos de Doenças , Enterococcus faecium/genética , Infecções por Bactérias Gram-Positivas/diagnóstico , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Tipagem de Sequências Multilocus , SARS-CoV-2 , Eslováquia/epidemiologia , Vancomicina/farmacologia , Vancomicina/uso terapêutico , Enterococos Resistentes à Vancomicina/genética
19.
Antimicrob Resist Infect Control ; 11(1): 99, 2022 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-35871001

RESUMO

BACKGROUND: Spread of vancomycin-resistant Enterococcus (VRE) is a global concern as a significant cause of healthcare-associated infections. A series of VRE faecium (VREf) outbreaks caused by clonal propagation due to interhospital transmission occurred in six general hospitals in Aomori prefecture, Japan. METHODS: The number of patients with VREf was obtained from thirty seven hospitals participating in the local network of Aomori prefecture. Thirteen hospitals performed active screening tests for VRE. Whole genome sequencing analysis was performed. RESULTS: The total number of cases with VREf amounted to 500 in fourteen hospitals in Aomori from Jan 2018 to April 2021. It took more than three years for the frequency of detection of VRE to return to pre-outbreak levels. The duration and size of outbreaks differed between hospitals according to the countermeasures available at each hospital. Whole genome sequencing analysis indicated vanA-type VREf ST1421 for most samples from six hospitals. CONCLUSIONS: This was the first multi-jurisdictional outbreak of VREf sequence type 1421 in Japan. In addition to strict infection control measures, continuous monitoring of VRE detection in local medical regions and smooth and immediate communication among hospitals are required to prevent VREf outbreaks.


Assuntos
Enterococcus faecium , Infecções por Bactérias Gram-Positivas , Enterococos Resistentes à Vancomicina , Enterococcus faecium/genética , Infecções por Bactérias Gram-Positivas/prevenção & controle , Humanos , Japão/epidemiologia , Vancomicina/farmacologia , Enterococos Resistentes à Vancomicina/genética
20.
J Appl Microbiol ; 133(3): 1989-2001, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35808847

RESUMO

AIMS: The objective of this study was to evaluate the inhibitory activity of compounds secreted by bacteria isolated from a hydrogen-producing bioreactor to understand how these microorganisms interact in this community. METHODS AND RESULTS: In vitro inhibitory assays were performed using samples secreted by bacteria subject to different treatments to determine if their inhibitory effect was due to organic acids, non-proteinaceous compounds or bacteriocin-like inhibitory substances (BLIS). Bacterial isolated were suppressed 43%, 30% and 27% by neutralized, precipitated and non-neutralized cell-free supernatants, respectively. Non-hydrogen producers (non-H2 P) lactic acid bacteria (LAB) (Lactobacillus plantarum LB1, Lactobacillus pentosus LB7, Pediococcus acidilactici LB4) and hydrogen producers (H2 P) LAB (Enterococcus faecium F) were inhibited by the production of organic acids, non-proteinaceous compounds and BLIS. Meanwhile, the obligate anaerobe H2 P (Clostridium beijerinckii B) inhibited by the production of non-proteinaceous compounds and BLIS. The presence of BLIS was confirmed when proteolytic enzymes affected the inhibitory activity of secreted proteins in values ranging from 20% to 42%. The BLIS produced by L. plantarum LB1, P. acidilactici LB4, L. pentosus LB7 and E. faecium F showed molecular masses of ~11, 25, 20 and 11 kDa, respectively. CONCLUSIONS: It was demonstrated antagonistic interactions between Lactobacillus-Enterococcus and Pediococcus-Enterococcus species, generated by the secretion of organic acids, non-proteinaceous compounds and BLIS. SIGNIFICANCE AND IMPACT OF THE STUDY: We report the interactions between LAB isolated from hydrogen-producing bioreactors. These interactions might impact the dynamics of the microbial population during hydrogen generation. Our work lays a foundation for strategies that allow controlling bacteria that can affect hydrogen production.


Assuntos
Bacteriocinas , Enterococcus faecium , Lactobacillales , Bacteriocinas/metabolismo , Bacteriocinas/farmacologia , Reatores Biológicos , Enterococcus/metabolismo , Fermentação , Hidrogênio/metabolismo , Lactobacillales/metabolismo , Pediococcus/metabolismo , Triticum/metabolismo
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