RESUMO
Enterococcus faecium is a lactic acid bacterium with applications in food engineering and nutrigenomics, including as starter cultures in fermented foods. To differentiate the E. faecium probiotic from pathogenic bacteria, physiological analyses are often used but they do not guarantee that a bacterial strain is not pathogenic. We report here new findings and an approach based on comparison of the genetic mobility of (1) probiotic, (2) pathogenic, and (3) nonpathogenic and non-probiotic strains, so as to differentiate probiotics, and inform their safe use. The region of the 16S ribosomal DNA (rDNA) genes of different E. faecium strains native to Pernambuco-Brazil was used with the GenBank query sequence. Complete genomes were selected and divided into three groups as noted above to identify the mobile genetic elements (MGEs) (transposase, integrase, conjugative transposon protein and phage) and antibiotic resistance genes (ARGs), and to undertake pan-genome analysis and multiple genome alignment. Differences in the number of MGEs were found in ARGs, in the presence and absence of the genes that differentiate E. faecium probiotics and pathogenic bacteria genetically. Our data suggest that genetic mobility appears to be informative in differentiating between probiotic and pathogenic strains. While the present findings are not necessarily applicable to all probiotics, they offer novel molecular insights to guide future research in nutrigenomics, clinical medicine, and food engineering on new ways to differentiate pathogenic from probiotic bacteria.
Assuntos
Elementos de DNA Transponíveis , Enterococcus faecium/genética , Genômica , Probióticos , Resistência Microbiana a Medicamentos , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/patogenicidade , Microbiologia de Alimentos , Genes Bacterianos , Genoma Bacteriano , Nutrigenômica/métodosRESUMO
The ban on the use of antibiotics as feed additives for animal growth promotion in the European Union and United States and the expectation of this trend to further expand to other countries in the short term have prompted a surge in probiotic research. Multi-species probiotics including safe and compatible strains with the ability to bind different nutritional lectins with detrimental effects on poultry nutrition could replace antibiotics as feed additives. Lactobacillus salivarius LET201, Lactobacillus reuteri LET210, Enterococcus faecium LET301, Propionibacterium acidipropionici LET103 and Bifidobacterium infantis CRL1395 have proved to be compatible as evaluated through three different approaches: the production and excretion of antimicrobial compounds, growth inhibition by competition for essential nutrients and physical contact, and a combination of both. The safety of P. acidipropionici LET103 was confirmed, since no expression of virulence factors or antibiotic resistance was detected. The innocuity of E. faecium LET301 should be further evaluated, since the presence of genes coding for certain virulence factors (gelE, efaAfm and efaAfs) was observed, albeit no expression of gelE was previously detected for this strain and there are no reports of involvement of efaAfm in animal pathogenicity. Finally, a combination of the five strains effectively protected intestinal epithelial cells of broilers from the cytotoxicity of mixtures of soybean agglutinin, wheat germ agglutinin and concanavalin A. To our knowledge, this is the first time that a combination of strains is evaluated for their protection against lectins that might be simultaneously present in poultry feeds.
Assuntos
Anti-Infecciosos/metabolismo , Bifidobacterium longum subspecies infantis/metabolismo , Enterococcus faecium/metabolismo , Lactobacillus/metabolismo , Doenças das Aves Domésticas/prevenção & controle , Probióticos/farmacologia , Propionibacterium/metabolismo , Animais , Antibiose , Bifidobacterium longum subspecies infantis/genética , Bifidobacterium longum subspecies infantis/crescimento & desenvolvimento , Bifidobacterium longum subspecies infantis/patogenicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Concanavalina A/toxicidade , Farmacorresistência Bacteriana , Enterococcus faecium/genética , Enterococcus faecium/crescimento & desenvolvimento , Enterococcus faecium/patogenicidade , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/fisiologia , Lactobacillus/genética , Lactobacillus/crescimento & desenvolvimento , Lactobacillus/patogenicidade , Lectinas/metabolismo , Modelos Teóricos , Lectinas de Plantas/toxicidade , Probióticos/efeitos adversos , Propionibacterium/genética , Propionibacterium/crescimento & desenvolvimento , Propionibacterium/patogenicidade , Ligação Proteica , Proteínas de Soja/toxicidade , Virulência , Fatores de Virulência/genética , Aglutininas do Germe de Trigo/toxicidadeRESUMO
Enterococcus faecium MXVK29 has the ability to produce an antimicrobial compound that belongs to Class IIa of the Klaenhammer classification, and could be used as part of a biopreservation technology through direct inoculation of the strain as a starter or protective culture. However, Enterococcus is considered as an opportunistic pathogen, hence, the purpose of this work was to study the food safety determinants of E. faecium MXVK29. The strain was sensitive to all of the antibiotics tested (penicillin, tetracycline, vancomycin, erythromycin, chloramphenicol, gentamicin, neomycin, kanamycin and netilmicin) and did not demonstrate histamine, cadaverine or putrescine formation. Furthermore, tyrosine-decarboxylase activity was detected by qualitative assays and PCR. Among the virulence factors analysed for the strain, only the genes encoding the sexual pheromone cCF10 precursor lipoprotein (ccf) and cell-wall adhesion (efaAfm ) were amplified. The presence of these genes has low impact on pathogenesis, as there are no other genes encoding for virulence factors, such as aggregation proteins. Therefore, Enterococcus faecium could be employed as part of a bioconservation method, because it does not produce risk factors for consumer's health; in addition, it could be used as part of the hurdle technology in foods. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of molecular techniques has allowed, in recent years, to detect pathogenicity genes present in the genome of starter cultures used in food processing and preservation. The presence of these genes is undesirable, because horizontal transfer may occur with the natural biota of consumers. For this reason, it is important to analyse the presence of pathogenicity genes in such cultures. In this work, virulence factors and antibiotic resistance of Enterococcus faecium strain MXVK29, producing an antimicrobial compound with high antilisterial activity, were analysed. The results indicate that the strain is safe to be used in food processing as starter culture.
Assuntos
Enterococcus faecium , Conservação de Alimentos , Inocuidade dos Alimentos , Produtos da Carne/microbiologia , Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/enzimologia , Enterococcus faecium/isolamento & purificação , Enterococcus faecium/patogenicidade , Humanos , México , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase , Tiramina/biossíntese , Tirosina Descarboxilase/metabolismo , Virulência/genética , Fatores de Virulência/genéticaRESUMO
Abstract Objectives To evaluate the prevalence of some virulence genes among 510 clinical Enterococcus spp. isolates and to assess the association of those genes with the species, infection site, and patient group (inpatients/outpatients). Methods Adhesins genes (aggregation substances agg and asa1 of Enterococcus faecalis and Enterococcus faecium, respectively), enterococcal surface protein (esp), endocarditis-specific antigen A (efaA), collagen-binding proteins (ace/acm)); invasins (hyaluronidase (hyl) and gelatinase (gelE)); cytotoxines (activation of cytolysin (cylA) in E. faecalis); and modulators of the host immunity and inflammation (enhanced expression pheromone (eep) in E. faecalis) were detected by polymerase chain reaction. Results The overall prevalence was: esp – 44.3%, agg/asa1 – 38.4%, ace/acm – 64.3%, efaA – 85.9%, eep – 69.4%, gelE – 64.3%, hyl – 25.1%, and cylA – 47.1%. E. faecalis isolates had significantly higher frequency of adhesin genes (esp and agg/asa1) and gelatinase in comparison to E. faecium. Multiple virulence genes in E. faecalis were significantly more prevalent than in E. faecium isolates. Domination of E. faecium with or without only one gene compared to the isolates of E. faecalis were found. Enterococcus spp. isolates obtained from outpatients compared to inpatients isolates had significantly higher frequency of agg/asa1, eep, gelE and cylA. Some adhesins genes (esp, agg/asa1 and efaA) had higher prevalence among the non-invasive Enterococcus spp. isolates compared to those causing invasive bacteremia, while ace/acm revealed higher dissemination in isolates causing invasive infections compared to non-invasive isolates. Conclusion Most E. faecalis attaches to abiotic surfaces in hospital environment, which correlates with higher prevalence of gene encoding for virulence factors involved in biofilm formation, such as enterococcal surface protein, aggregation substance, and gelatinase. The intestinal tract is an important reservoir for opportunistic enterococcal pathogens and allows them to access infectious sites through different virulence factors, demonstrated in outpatient isolates in this study.
Assuntos
Humanos , Virulência/genética , Enterococcus faecium/patogenicidade , Enterococcus faecalis/patogenicidade , Fatores de Virulência/genética , Bulgária , Reação em Cadeia da Polimerase , Incidência , Eletroforese em Gel de Campo PulsadoRESUMO
OBJECTIVES: To evaluate the prevalence of some virulence genes among 510 clinical Enterococcus spp. isolates and to assess the association of those genes with the species, infection site, and patient group (inpatients/outpatients). METHODS: Adhesins genes (aggregation substances agg and asa1 of Enterococcus faecalis and Enterococcus faecium, respectively), enterococcal surface protein (esp), endocarditis-specific antigen A (efaA), collagen-binding proteins (ace/acm)); invasins (hyaluronidase (hyl) and gelatinase (gelE)); cytotoxines (activation of cytolysin (cylA) in E. faecalis); and modulators of the host immunity and inflammation (enhanced expression pheromone (eep) in E. faecalis) were detected by polymerase chain reaction. RESULTS: The overall prevalence was: esp - 44.3%, agg/asa1 - 38.4%, ace/acm - 64.3%, efaA - 85.9%, eep - 69.4%, gelE - 64.3%, hyl - 25.1%, and cylA - 47.1%. E. faecalis isolates had significantly higher frequency of adhesin genes (esp and agg/asa1) and gelatinase in comparison to E. faecium. Multiple virulence genes in E. faecalis were significantly more prevalent than in E. faecium isolates. Domination of E. faecium with or without only one gene compared to the isolates of E. faecalis were found. Enterococcus spp. isolates obtained from outpatients compared to inpatients isolates had significantly higher frequency of agg/asa1, eep, gelE and cylA. Some adhesins genes (esp, agg/asa1 and efaA) had higher prevalence among the non-invasive Enterococcus spp. isolates compared to those causing invasive bacteremia, while ace/acm revealed higher dissemination in isolates causing invasive infections compared to non-invasive isolates. CONCLUSION: Most E. faecalis attaches to abiotic surfaces in hospital environment, which correlates with higher prevalence of gene encoding for virulence factors involved in biofilm formation, such as enterococcal surface protein, aggregation substance, and gelatinase. The intestinal tract is an important reservoir for opportunistic enterococcal pathogens and allows them to access infectious sites through different virulence factors, demonstrated in outpatient isolates in this study.
Assuntos
Enterococcus faecalis/patogenicidade , Enterococcus faecium/patogenicidade , Fatores de Virulência/genética , Virulência/genética , Bulgária , Eletroforese em Gel de Campo Pulsado , Humanos , Incidência , Reação em Cadeia da PolimeraseRESUMO
In this work, the sources of contamination by Enterococcus spp. in a ricotta processing line were evaluated. The isolated strains were tested for virulence genes (gelE, cylA,B, M, esp, agg, ace, efaA, vanB), expression of virulence factors (hemolysin and gelatinase), and the resistance to 10 different antibiotics. Enterococcus faecium and Enterococcus faecalis were subjected to discriminatory identification by intergenic spacer region (ITS)-polymerase chain reaction and sequencing of the ITS region. The results showed that Enterococcus spp. was detected in the raw materials, environment samples and the final product. None of the 107 Enterococcus isolates were completely free from all virulence genes considered. A fraction of 21.5% of the isolates containing all of the genes of the cylA, B, M operon also expressed ß-hemolysis. Most of the isolates showed the gelE gene, but only 9.3% were able to hydrolyze gelatin. In addition, 23.5% of the observed Enterococcus isolates had the vanB gene but were susceptible to vancomycin in vitro. The dissemination of antibiotic-resistant enterococci was revealed in this study: 19.3% of the E. faecium samples and 78.0% of the E. faecalis samples were resistant to at least one of the antibiotics tested. Sequencing of region discriminated 5 and 7 distinct groups among E. faecalis and E. faecium, respectively. Although some similarity was observed among some of the isolates, all E. faecalis and E. faecium isolates had genetic differences both in the ITS region and in the virulence profile, which makes them different from each other.
Assuntos
Laticínios/microbiologia , Resistência Microbiana a Medicamentos , Enterococcus faecalis/crescimento & desenvolvimento , Enterococcus faecium/crescimento & desenvolvimento , Microbiologia de Alimentos , Fatores de Virulência , Antibacterianos/farmacologia , DNA Intergênico , Resistência Microbiana a Medicamentos/genética , Enterococcus , Enterococcus faecalis/isolamento & purificação , Enterococcus faecalis/patogenicidade , Enterococcus faecium/isolamento & purificação , Enterococcus faecium/patogenicidade , Genes Bacterianos , Humanos , Reação em Cadeia da Polimerase , Virulência , Fatores de Virulência/genéticaRESUMO
Despite the increasing importance of Enterococcus as opportunistic pathogens, their virulence factors are still poorly understood. This study determines the frequency of virulence factors in clinical and commensal Enterococcus isolates from inpatients in Porto Alegre, Brazil. Fifty Enterococcus isolates were analysed and the presence of the gelE, asa1 and esp genes was determined. Gelatinase activity and biofilm formation were also tested. The clonal relationships among the isolates were evaluated using pulsed-field gel electrophoresis. The asa1, gelE and esp genes were identified in 38%, 60% and 76% of all isolates, respectively. The first two genes were more prevalent in Enterococcus faecalis than in Enterococcus faecium, as was biofilm formation, which was associated with gelE and asa1 genes, but not with the esp gene. The presence of gelE and the activity of gelatinase were not fully concordant. No relationship was observed among any virulence factors and specific subclones of E. faecalis or E. faecium resistant to vancomycin. In conclusion, E. faecalis and E. faecium isolates showed significantly different patterns of virulence determinants. Neither the source of isolation nor the clonal relationship or vancomycin resistance influenced their distribution.
Assuntos
Antibacterianos/farmacologia , Enterococcus faecalis , Enterococcus faecium , Vancomicina/farmacologia , Fatores de Virulência/genética , Biofilmes/crescimento & desenvolvimento , Eletroforese em Gel de Campo Pulsado , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/enzimologia , Enterococcus faecalis/patogenicidade , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/enzimologia , Enterococcus faecium/patogenicidade , Gelatinases/metabolismo , Testes de Sensibilidade Microbiana , Resistência a Vancomicina/genéticaRESUMO
Despite the increasing importance of Enterococcus as opportunistic pathogens, their virulence factors are still poorly understood. This study determines the frequency of virulence factors in clinical and commensal Enterococcus isolates from inpatients in Porto Alegre, Brazil. Fifty Enterococcus isolates were analysed and the presence of the gelE, asa1 and esp genes was determined. Gelatinase activity and biofilm formation were also tested. The clonal relationships among the isolates were evaluated using pulsed-field gel electrophoresis. The asa1, gelE and esp genes were identified in 38%, 60% and 76% of all isolates, respectively. The first two genes were more prevalent in Enterococcus faecalis than in Enterococcus faecium, as was biofilm formation, which was associated with gelE and asa1 genes, but not with the esp gene. The presence of gelE and the activity of gelatinase were not fully concordant. No relationship was observed among any virulence factors and specific subclones of E. faecalis or E. faecium resistant to vancomycin. In conclusion, E. faecalis and E. faecium isolates showed significantly different patterns of virulence determinants. Neither the source of isolation nor the clonal relationship or vancomycin resistance influenced their distribution.
Assuntos
Antibacterianos/farmacologia , Enterococcus faecalis , Enterococcus faecium , Vancomicina/farmacologia , Fatores de Virulência/genética , Biofilmes/crescimento & desenvolvimento , Eletroforese em Gel de Campo Pulsado , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/enzimologia , Enterococcus faecalis/patogenicidade , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/enzimologia , Enterococcus faecium/patogenicidade , Gelatinases/metabolismo , Testes de Sensibilidade Microbiana , Resistência a Vancomicina/genéticaAssuntos
Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecium/efeitos dos fármacos , Infecções por Bactérias Gram-Positivas/epidemiologia , Resistência a Vancomicina , Classificação , Eletroforese em Gel de Campo Pulsado , Enterococcus faecalis/genética , Enterococcus faecalis/patogenicidade , Enterococcus faecium/genética , Enterococcus faecium/patogenicidade , Hospitais/estatística & dados numéricos , Humanos , México/epidemiologia , Estudos RetrospectivosRESUMO
INTRODUCTION: Vancomycin-resistant enterococci isolates (VRE) have caused numerous outbreaks in intensive care units (ICUs). A contaminated hospital environment, the hands of health care workers (HCW), and carrier patients may play important roles in perpetuating the chain of transmission in these outbreaks. The aims of this study were to report the first VRE outbreak in our center and assess the role of environmental contamination and HCW hands in the spread of new cases of enterococcal infection. MATERIAL AND METHOD: Between August and December 2003, surveillance cultures were performed with samples from all patients (n = 113) admitted to the ICU, as well as cultures of samples from the environment (n = 69) and HCW hands (n = 23). RESULTS: Eighteen clinical samples from 8 patients and 7 environmental samples yielded Enterococcus faecium (24 strains) and E. avium (1 strain). VRE was not detected on HCW hands. All the VRE isolates belonged to a single clone and carried the vanA gene. CONCLUSION: Environmental contamination provides an important reservoir for future outbreaks of VRE, perpetuating transmission of the microorganism in the hospital setting.
Assuntos
Surtos de Doenças , Reservatórios de Doenças , Enterococcus faecium/patogenicidade , Contaminação de Equipamentos , Infecções por Bactérias Gram-Positivas/epidemiologia , Resistência a Vancomicina , Idoso , Idoso de 80 Anos ou mais , Argentina/epidemiologia , Proteínas de Bactérias/análise , Carbono-Oxigênio Ligases/análise , Células Clonais , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , Infecção Hospitalar/prevenção & controle , Infecção Hospitalar/transmissão , Surtos de Doenças/prevenção & controle , Transmissão de Doença Infecciosa/prevenção & controle , Enterococcus faecium/classificação , Enterococcus faecium/enzimologia , Enterococcus faecium/genética , Feminino , Infecções por Bactérias Gram-Positivas/microbiologia , Infecções por Bactérias Gram-Positivas/transmissão , Mãos/microbiologia , Humanos , Higiene , Controle de Infecções/normas , Unidades de Terapia Intensiva , Masculino , Pessoa de Meia-Idade , Recursos Humanos em Hospital , Resistência a Vancomicina/genéticaRESUMO
Os Enterococos são cocos gram-positivos com baixo potencial de causar doenças. No entanto, tornaram-se temidos nas últimas décadas devido ao desenvolvimento de cepas bastante resistentes aos aminoglicosídeos, à ampicilina, penicilina e vancomicina. Os locais de infecções enterocócicas mais comuns são as vias urinárias e a corrente sangüínea. O controle e a prevenção dessas infecções é difícil devido ao elevado grau de resistência, o que torna importante a restrição da antibioticoterapia e o controle de pacientes infectados para reduzir a disseminação dessas bactérias
Assuntos
Infecções Bacterianas , Enterococcus/patogenicidade , Resistência Microbiana a Medicamentos , Enterococcus faecalis/patogenicidade , Enterococcus faecium/patogenicidade , Vancomicina/farmacologiaRESUMO
Vancomycin-resistant enterococci (VRE) have caused hospital outbreaks worldwide, and the vancomycin-resistance gene (vanA) has crossed genus boundaries to methicillin-resistant Staphylococcus aureus. Spread of VRE, therefore, represents an immediate threat for patient care and creates a reservoir of mobile resistance genes for other, more virulent pathogens. Evolutionary genetics, population structure, and geographic distribution of 411 VRE and vancomycin-susceptible Enterococcus faecium isolates, recovered from human and nonhuman sources and community and hospital reservoirs in 5 continents, identified a genetic lineage of E. faecium (complex-17) that has spread globally. This lineage is characterized by 1) ampicillin resistance, 2) a pathogenicity island, and 3) an association with hospital outbreaks. Complex-17 is an example of cumulative evolutionary processes that improved the relative fitness of bacteria in hospital environments. Preventing further spread of this epidemic E. faecium subpopulation is critical, and efforts should focus on the early disclosure of ampicillin-resistant complex-17 strains.
Assuntos
Infecção Hospitalar/transmissão , Enterococcus faecium/efeitos dos fármacos , Infecções por Bactérias Gram-Positivas/transmissão , Resistência a Vancomicina , África/epidemiologia , Animais , Austrália/epidemiologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Gatos , Bovinos , Infecção Hospitalar/epidemiologia , Infecção Hospitalar/microbiologia , Cães , Enterococcus faecium/classificação , Enterococcus faecium/genética , Enterococcus faecium/patogenicidade , Europa (Continente)/epidemiologia , Evolução Molecular , Infecções por Bactérias Gram-Positivas/epidemiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , América do Norte/epidemiologia , Resistência às Penicilinas , Recombinação Genética , América do Sul/epidemiologiaRESUMO
Objetivo: caracterizar clínica y epidemiológicamente, y establecer los factores de riesgo de un brote de infección nosocomial por Enterococcus faecium resistente a vancomicina en el HUSVP de Medellín. Metodología: se realizó un estudio con una fase descriptiva (27 casos) y un diseño tipo casos (27) y controles (54). Resultados: los 27 casos de E. faecium tuvieron un alto nivel de resistencia a vancomicina (128 ug/ml), y además fueron resitentes a teicoplanina, ampicilina, penicilina, ciprofloxacina, y gentamicina y estreptomicina de alto nivel. En el 67 por ciento de los casos se interpretó cono infección; el 33 por ciento de las cepas se aislaron de orina y el 22 por ciento de cavidad abdominal. Los factores de riesgo asociados fueron la insuficiencia renal (RD 12.6), nutrición parenteral total (RD 6.2), la cirugía abdominal (RD 4.1), uso previo de metronidazol (RD 19.5) ceftazidima (RD 15.9), cefotaxima (RD 12.6), imipenem (RD 11.6), ciproflaxina (RD 6.7), vancomicina (RD 5.4), ampicilina/sulbactam (RD 4.4), días estancia (p=0.004), días entre el ingreso y la infección (p=0.0007), número de cirugias (p=0.0004) y número de antibióticos previos (p=0.00005). Conclusiones: esta es la primera epidemia de ERV que se publica en Colombia, con un análisis clínico, epidemiológico y de factores de riesgo, y nos muestra una realidad ya vivida por otros países y de muy difícil manejo, y nos alerta sobre la posibilidad de aparición de Staphylococcus aureus con sensibilidad disminuida a la vancomicina