RESUMO
Vellore, a region in southern India, has a high incidence of severe human infections with Beta-hemolytic group C and G streptococci (GCGS). To determine the causative species in these infections, we conducted 16S rRNA gene sequencing: Streptococcus dysgalactiae subsp. equisimilis (81%) and S. anginosus (19%) were the causative organisms in the 2-year study period (2006-2007). We used PCR to detect the virulence-related emm gene; results showed that it was restricted to S. dysgalactieae subsp. equisimilis isolates of 99.2% tested positive. Due to a novel marker, S. anginosus and S. constellatus can be quickly and accurately distinguished from other members of the genus. The notable contribution of the anginosus group to human infections suggests that this group of obligate pathogens deserves more attention in healthcare and research.
Assuntos
Infecções Estreptocócicas/microbiologia , Streptococcus anginosus , DNA Bacteriano/genética , Genes Bacterianos/genética , Marcadores Genéticos/genética , Humanos , Índia/epidemiologia , Fases de Leitura Aberta/genética , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Infecções Estreptocócicas/epidemiologia , Streptococcus/genética , Streptococcus anginosus/genética , Streptococcus constellatus/genética , Fatores de Virulência/genéticaRESUMO
A real-time quantitative PCR was developed to assess antiviral activity of molecules against human herpesvirus 8 (HHV-8) and the Epstein-Barr virus (EBV). The antiviral activity of the reference molecules acyclovir, ganciclovir, cidofovir, adefovir and brivudin, as assessed by this methodology, proved very similar to the activity as determined by a DNA-DNA hybridisation method.
Assuntos
Antivirais/farmacologia , Citosina/análogos & derivados , Herpesvirus Humano 4/efeitos dos fármacos , Herpesvirus Humano 8/efeitos dos fármacos , Organofosfonatos , Aciclovir/farmacologia , Antivirais/sangue , Linhagem Celular , Cidofovir , Citosina/farmacologia , Ganciclovir/farmacologia , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/imunologia , Herpesvirus Humano 8/genética , Herpesvirus Humano 8/imunologia , Humanos , Testes de Sensibilidade Microbiana , Compostos Organofosforados/farmacologia , Reação em Cadeia da PolimeraseRESUMO
Oral streptococci are a heterogeneous group of human commensals, with a potential to cause serious infections. Activation of plasminogen has been shown to increase the virulence of typical human pathogenic streptococci such as S. pneumoniae. One important factor for plasminogen activation is the streptococcal α-enolase. Here we report that plasminogen activation is also common in oral streptococci species involved in clinical infection and that it depends on the action of human plasminogen activators. The ability to activate plasminogen did not require full conservation of the internal plasminogen binding sequence motif FYDKERKVY of α-enolase that was previously described as crucial for increased plasminogen binding, activation and virulence. Instead, experiments with recombinant α-enolase variants indicate that the naturally occurring variations do not impair plasminogen binding. In spite of these variations in the internal plasminogen binding motif oral streptococci showed similar activation of plasminogen. We conclude that the pathomechanism of plasminogen activation is conserved in oral streptococci that cause infections in human. This may contribute to their opportunistic pathogenic character that is unfurled in certain niches.
Assuntos
Proteínas de Bactérias/metabolismo , Fosfopiruvato Hidratase/metabolismo , Plasminogênio/metabolismo , Streptococcus/patogenicidade , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Sítios de Ligação/genética , Eletroforese em Gel de Poliacrilamida , Fibrinolisina/metabolismo , Humanos , Immunoblotting , Dados de Sequência Molecular , Boca/microbiologia , Mutação , Fosfopiruvato Hidratase/genética , Ativadores de Plasminogênio/metabolismo , Ligação Proteica , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Infecções Estreptocócicas/metabolismo , Infecções Estreptocócicas/microbiologia , Streptococcus/enzimologia , Streptococcus/genética , Streptococcus oralis/enzimologia , Streptococcus oralis/genética , Streptococcus oralis/patogenicidade , Virulência/genéticaRESUMO
The molecular mode of cell killing by the antiviral drug (E)-5-(2-bromovinyl-2'-deoxyuridine (BVDU) was studied in Chinese hamster ovary (CHO) cells stably transfected with the thymidine kinase gene (tk) of varicella zoster virus (CHO-VZVtk). The colony-forming ability of the cells was reduced to <1% at a concentration of approximately 1 microM BVDU, whereas for nontransfected cells or cells transfected with tk gene of herpes simplex virus type 1 (CHO-HSVtk), a 1000-fold higher dose was required to achieve the same response. BVDU inhibited thymidylate synthase in CHO-VZVtk but not in CHO-HSVtk and control cells. On the other hand, the drug was incorporated into DNA of VZVtk- and HSVtk-expressing cells to nearly equal amounts. Because coexposure of CHO-VZVtk cells to exogenous thymidine protected them from BVDU-induced cell killing, the cells obviously die because of thymidine depletion. At highly cytotoxic BVDU doses (50 microM) and longer exposure times (24-48 h), VZVtk cells were blocked to some extent in S and G2/M phase and underwent apoptosis (48-72 h). Not only apoptosis but also necrosis was induced. The findings also show that the drug causes the induction of c-Jun and the activation of activator protein-1 resulting in increased level of Fas ligand (FasL) and caspase-8/-3 activation. Bid and poly(ADP-ribose) polymerase were cleaved by caspases. Expression of Bax increased, whereas Bcl-2/Bcl-x(L) remained unchanged. Transfection of dominant-negative Fas-associated death domain and inhibition of caspase-8 by N-benzyloxycarbonyl-IETD-fluoromethyl ketone strongly abrogated BVDU-induced apoptosis, indicating Fas/FasL to be crucially involved. Thus, BVDU-triggered apoptosis differs significantly from that induced by ganciclovir, which induces in the same cellular background the mitochondrial damage pathway.