RESUMO
Staphylococcus aureus is an important cause of resistant healthcare-associated infections. It has been shown that the wall teichoic acid (WTA) may be an important drug target acting on antibiotic-resistant cells. The UDP-N-acetylglucosamine 2-epimerase, MnaA, is one of the first enzymes on the pathway for the biosynthesis of the WTA. Here, detailed molecular dynamics simulations of S. aureus MnaA were used to characterize the conformational changes that occur in the presence of UDP and UDP-GlcNac and also the energetic landscape associated with these changes. Using different simulation techniques, such as ABMD and GAMD, it was possible to assess the energetic profile for the protein with and without ligands in its active site. We found that there is a dynamic energy landscape that has its minimum changed by the presence of the ligands, with a closed structure of the enzyme being more frequently observed for the bound state while the unbound enzyme favors an opened conformation. Further structural analysis indicated that positively charged amino acids associated with UDP and UDP-GlcNac interactions play a major role in the enzyme opening movement. Finally, the energy landscape profiled in this work provides important conclusions for the design of inhibitor candidates targeting S. aureus MnaA.
Assuntos
Infecções Estafilocócicas/enzimologia , Staphylococcus aureus/enzimologia , Ácidos Teicoicos/química , Sequência de Aminoácidos , Aminoácidos/química , Carboidratos Epimerases/química , Carboidratos Epimerases/metabolismo , Carboidratos Epimerases/ultraestrutura , Domínio Catalítico/efeitos dos fármacos , Parede Celular/enzimologia , Farmacorresistência Bacteriana/genética , Metabolismo Energético/genética , Glucosamina/análogos & derivados , Glucosamina/química , Humanos , Ligantes , Simulação de Dinâmica Molecular , Conformação Proteica/efeitos dos fármacos , Domínios Proteicos/genética , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/patogenicidade , Difosfato de Uridina/químicaRESUMO
Peptidoglycan (PEG) and lipoteichoic acid (LTA) are the main constituents of Gram-positive bacteria cell wall and are described to modulate immune functions. Increased levels of matrix metalloproteinases (MMPs) were described in endotoxemia, suggesting that they participate to tecidual damage, multiple organs failure and vascular disfunction. Staphylococcus aureus PEG is described to increase MMPs 2 and 9 levels in plasma from rat and MMP 9 secretion by human neutrophils, however, the effect of LTA on MMPs is unknown. In this work, was evaluated the modulation of MMPs 2 and 9 expression and secretion in RAW 264.7 macrophages by LTA from S. aureus. The role of A2A and A2B adenosine receptors was also investigated. LTA increased MMP 9 expression and secretion at 12h of treatment. The modulation of MMP 9 secretion was dose dependent, with maximal effect above 1microg/ml. The inhibitor of mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway (U0126, 10microM) prevented LTA stimulation of MMP 9 secretion; however, the inhibitors of p38 (SB203580, 10microM) and Jun N-terminal kinase (JNK; SP600125, 10microM) presented any effect. A2A and A2B adenosine receptors pharmacological blockade or gene knockdown resulted in exacerbated MMP 9 secretion, while an adenosine receptors agonist inhibited LTA-stimulated MMP 9 secretion. These results suggest that LTA increased MMP 9 secretion in macrophages could be involved in complications associated to S. aureus infections. Moreover, LTA modulation of MMP 9 is dependent on MEK/ERK pathway and is regulated by A2A and A2B adenosine receptors.
Assuntos
Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Metaloproteinase 9 da Matriz/imunologia , Receptor A2A de Adenosina/imunologia , Receptor A2B de Adenosina/imunologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/imunologia , Ácidos Teicoicos/farmacologia , Animais , Linhagem Celular , Inibidores Enzimáticos/farmacologia , Regulação Enzimológica da Expressão Gênica/imunologia , Técnicas de Silenciamento de Genes , Humanos , Lipopolissacarídeos/química , Lipopolissacarídeos/imunologia , Lipopolissacarídeos/metabolismo , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/imunologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/imunologia , Macrófagos/enzimologia , Metaloproteinase 9 da Matriz/biossíntese , Camundongos , Ratos , Receptor A2A de Adenosina/metabolismo , Receptor A2B de Adenosina/metabolismo , Infecções Estafilocócicas/enzimologia , Staphylococcus aureus/química , Ácidos Teicoicos/química , Ácidos Teicoicos/imunologia , Ácidos Teicoicos/metabolismo , Fatores de Tempo , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Detailed clinical data, underlying conditions, inflammatory indices and microbiological parameters in 60 patients who had pure growth of coagulase negative staphylococci from their blood culture specimens at the University Hospital of the West Indies, Jamaica, were analyzed and the clinical significance of the isolates ascertained using standard criteria. This study was undertaken between April and September 2003. The isolates were true pathogens of bloodstream infections in only 5 of the 60 patients (8.4%). In the vast majority ie 44 of 60 (73.3%) they were mere blood culture contaminants and in 11 (18.3%), the clinical significance could not be ascertained Fifteen of the 44 patients (34%) with contaminating coagulase negative staphylococci were treated with specific anti-staphylococcal antibiotics; 5 (11.4%) with vancomycin. Although there has been a relative increase of coagulase negative staphylococcal infections including bloodstream infections in recent years, the organisms still remain the most common contaminants in blood cultures. Over 70% of isolates were contaminants in this study which is similar to that in a number of such studies in other parts of the world The findings underline the need for careful evaluation of coagulase negative staphylococci isolated from blood cultures before instituting therapy to avoid unnecessary use of antibiotics, especially vancomycin, and the consequent increase of antibiotic resistance in hospitals.
Assuntos
Antibacterianos/farmacologia , Sepse/microbiologia , Infecções Estafilocócicas/microbiologia , Staphylococcus/isolamento & purificação , Vancomicina/farmacologia , Adulto , Idoso , Coagulase , Infecção Hospitalar/microbiologia , Infecção Hospitalar/prevenção & controle , Resistência a Medicamentos , Feminino , Hospitais Universitários , Humanos , Jamaica , Masculino , Pessoa de Meia-Idade , Sepse/tratamento farmacológico , Sepse/enzimologia , Infecções Estafilocócicas/sangue , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/enzimologia , Staphylococcus/efeitos dos fármacos , Staphylococcus/enzimologiaRESUMO
The prevalence and significance of coagulase negative staphylococci (CoNS) isolated from blood cultures at the University Hospital of the West Indies (UHWI) during a six-month period were investigated. Standard and automated microbiological procedures were used to process 3001 blood culture specimens received from 2363 patients and 658 (21.9%) of the blood cultures yielded 854 bacterial isolates. The highest prevalence of positive blood cultures (60%) and the lowest prevalence of blood isolates of CoNS (12%) were found in the intensive care unit (ICU). The blood isolates of CoNS were most frequent in the surgical wards (13%) and lowest in obstetrics and gynaecology (2%). High rates of resistance to methicillin, other anti-staphylococcal penicillins, and cephalosporins used in the treatment of CoNS were observed All blood isolates of CoNS (100%) were susceptible to vancomycin. In conclusion, the results show that coagulase-negative staphylococci are the most prevalent bacterial isolates in blood cultures at the UHWI occurring mostly as contaminants. The practice of proper venepuncture and hand-washing techniques by medical staff are recommended to facilitate appropriate antibiotic usage.
Assuntos
Infecção Hospitalar/epidemiologia , Hospitais Universitários/estatística & dados numéricos , Infecções Estafilocócicas/epidemiologia , Staphylococcus/isolamento & purificação , Coagulase , Infecção Hospitalar/sangue , Infecção Hospitalar/microbiologia , Humanos , Unidades de Terapia Intensiva , Jamaica/epidemiologia , Testes de Sensibilidade Microbiana , Estudos Soroepidemiológicos , Infecções Estafilocócicas/sangue , Infecções Estafilocócicas/enzimologia , Staphylococcus/enzimologiaRESUMO
Detailed clinical data, underlying conditions, inflammatory indices and microbiological parameters in 60 patients who had pure growth of coagulase negative staphylococci from their blood culture specimens at the University Hospital of the West Indies, Jamaica, were analyzed and the clinical significance of the isolates ascertained using standard criteria. This study was undertaken between April and September 2003. The isolates were true pathogens of bloodstream infections in only 5 of the 60 patients (8.4). In the vast majority ie 44 of 60 (73.3) they were mere blood culture contaminants and in 11 (18.3), the clinical significance could not be ascertained Fifteen of the 44 patients (34) with contaminating coagulase negative staphylococci were treated with specific anti-staphylococcal antibiotics; 5 (11.4) with vancomycin. Although there has been a relative increase of coagulase negative staphylococcal infections including bloodstream infections in recent years, the organisms still remain the most common contaminants in blood cultures. Over 70of isolates were contaminants in this study which is similar to that in a number of such studies in other parts of the world The findings underline the need for careful evaluation of coagulase negative staphylococci isolated from blood cultures before instituting therapy to avoid unnecessary use of antibiotics, especially vancomycin, and the consequent increase of antibiotic resistance in hospitals.
Los datos clínicos detallados, las condiciones subyacentes, los índices inflamatorios así como los parámetros microbiológicos de 60 pacientes que presentaron crecimiento puro de estafilococos coagu-lasa-negativos de sus cultivos de sangre en el Hospital Universitario de West Indies, Jamaica, fueron sometidos a análisis. De este modo, se pudo determinar la significación clínica de los aislados, usando criterios estándar. En sólo 5 de los 60 pacientes (8.4%), los aislados resultaron ser realmente pató-genos de infecciones del torrente sanguíneo. En la gran mayoría, a saber 44 de 60 (73.3%), se trataba tan sólo de contaminantes en el cultivo de la sangre, y en 11 (18.3%) no pudo determinarse la impor-tancia clínica. Quince de los 44 pacientes (34%) con estafilococos coagulasa-negativos contaminantes, fueron tratados con antibióticos antiestafilocócicos específicos, y 5 (11.4%) con vancomicina. Aunque en años recientes ha habido un aumento relativo de infecciones por estafilococos coagulasa-negativos incluyendo infecciones del torrente sanguíneo los organismos siguen siendo todavía los contamin-antes más comunes en los cultivos de sangre. Más del 70% de los aislados resultaron ser contaminantes en este estudio: un resultado similar al obtenido en una serie de estudios de este tipo realizado en otras partes del mundo. Los hallazgos apuntan a la necesidad de realizar una evaluación cuidadosa de los estreptococos coagulasa-negativos aislados en los cultivos de sangre antes de proceder a instituir la terapia, a fin de evitar el uso innecesario de antibióticos en especial la vancomicina y el consiguiente aumento de la resistencia antibiótica en los hospitales.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Antibacterianos/farmacologia , Infecções Estafilocócicas/microbiologia , Sepse/microbiologia , Staphylococcus/isolamento & purificação , Vancomicina/farmacologia , Coagulase , Hospitais Universitários , Infecção Hospitalar/microbiologia , Infecção Hospitalar/prevenção & controle , Infecções Estafilocócicas/sangue , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/enzimologia , Jamaica , Resistência a Medicamentos , Sepse/tratamento farmacológico , Sepse/enzimologia , Staphylococcus/efeitos dos fármacos , Staphylococcus/enzimologiaRESUMO
The prevalence and significance of coagulase negative staphylococci (CoNS) isolated from blood cultures at the University Hospital of the West Indies (UHWI) during a six-month period were investigated. Standard and automated microbiological procedures were used to process 3001 blood culture specimens received from 2363 patients and 658 (21.9) of the blood cultures yielded 854 bacterial isolates. The highest prevalence of positive blood cultures (60) and the lowest prevalence of blood isolates of CoNS (12) were found in the intensive care unit (ICU). The blood isolates of CoNS were most frequent in the surgical wards (13) and lowest in obstetrics and gynaecology (2). High rates of resistance to methicillin, other anti-staphylococcal penicillins, and cephalosporins used in the treatment of CoNS were observed All blood isolates of CoNS (100) were susceptible to vancomycin. In conclusion, the results show that coagulase-negative staphylococci are the most prevalent bacterial isolates in blood cultures at the UHWI occurring mostly as contaminants. The practice of proper venepuncture and hand-washing techniques by medical staff are recommended to facilitate appropriate antibiotic usage.
Se investigó la prevalencia e importancia de los estafilococos coagulasa negativos (ECoN) aislados de cultivos de sangre en el Hospital Universitario de West Indies (HUWI) por un período de seis meses. Se utilizaron procedimientos microbiológicos estándar y automatizados para procesas 3001 cultivos de sangre recibidos de 2363 pacientes y 658 (21.9%) de los cultivos dieron 854 aislados bacterianos. La más alta prevalencia de cultivos de sangre positivos (60%) y la más baja prevalencia de aislados de ECoN (12%) se encontraron en la Unidad de Cuidados Intensivos (UCI). Los aislados de sangre de ECoN fueron más frecuentes en las salas de cirugía (13%) y más bajos en las de obstetricia y ginecología (2%). Se observaron altas tasas de resistencia a la meticilina, así como a otras penicilinas anti-estafilocócicas y cefalosporinas usadas en el tratamiento de ECoN. Todos los aislados de sangre de ECoN (100%) fueron susceptibles a la vancomicina. En conclusión, los resultados muestran que los estafilococos coagulasa negativos son los aislados bacterianos más prevalentes en cultivos de sangre en el HUWI, presentándose en la mayor parte de los casos como contaminantes. Se recomienda la práctica de técnicas adecuadas de venepuntura y lavado de manos por parte del personal médico a fin de facilitar un uso antibiótico correcto.
Assuntos
Humanos , Hospitais Universitários/estatística & dados numéricos , Infecção Hospitalar/epidemiologia , Infecções Estafilocócicas/epidemiologia , Staphylococcus/isolamento & purificação , Coagulase , Estudos Soroepidemiológicos , Infecção Hospitalar/sangue , Infecção Hospitalar/microbiologia , Infecções Estafilocócicas/sangue , Infecções Estafilocócicas/enzimologia , Jamaica/epidemiologia , Staphylococcus/enzimologia , Testes de Sensibilidade Microbiana , Unidades de Terapia IntensivaRESUMO
Se aislaron cepas de Staphylococcus aureus resistentes a penicilina y productoras de ß-lactamasas. Mediante el método de dilución en tubos, se calculó la concentración inhibitoria CIM para amoxicilina-sulbactam, con diferente tensión de oxígeno y frente a heparina. Los valores de CIM se redujeron significativamente al sumar sulbactam al antibiótico. La disminución del oxígeno no afectó la capacidad inhibidora de ß-lactamasas del sulbactam, del mismo modo que la presencia de heparina no interfirió en su efecto protector sobre la amoxicilina. Estos resultados reafirmaron el concepto de efectividad de la combinación amoxicilina-sulbactam sobre los procesos infecciosos a S. aureus productores de ß-lactamasas, aun en afecciones donde se genera anaerobiosis relativa y también en enfermos con tratamiento simultáneo con heparina
Assuntos
Amoxicilina/antagonistas & inibidores , Anaerobiose/efeitos dos fármacos , beta-Lactamases/antagonistas & inibidores , Resistência Microbiana a Medicamentos/fisiologia , Infecções Estafilocócicas/tratamento farmacológico , Sulbactam/uso terapêutico , Amoxicilina/uso terapêutico , beta-Lactamases/efeitos adversos , Adjuvantes Farmacêuticos/uso terapêutico , Heparina/efeitos adversos , Infecções Estafilocócicas/enzimologia , Infecções Estafilocócicas/fisiopatologia , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/enzimologia , Sulbactam/antagonistas & inibidores , Sulbactam/farmacocinéticaRESUMO
Se aislaron cepas de Staphylococcus aureus resistentes a penicilina y productoras de ß-lactamasas. Mediante el método de dilución en tubos, se calculó la concentración inhibitoria CIM para amoxicilina-sulbactam, con diferente tensión de oxígeno y frente a heparina. Los valores de CIM se redujeron significativamente al sumar sulbactam al antibiótico. La disminución del oxígeno no afectó la capacidad inhibidora de ß-lactamasas del sulbactam, del mismo modo que la presencia de heparina no interfirió en su efecto protector sobre la amoxicilina. Estos resultados reafirmaron el concepto de efectividad de la combinación amoxicilina-sulbactam sobre los procesos infecciosos a S. aureus productores de ß-lactamasas, aun en afecciones donde se genera anaerobiosis relativa y también en enfermos con tratamiento simultáneo con heparina (AU)