RESUMO
A common epitope region of enteroviruses was identified by sequence-independent single-primer amplification (SISPA), followed by immunoscreening of 11 cDNA libraries from two Korean enterovirus isolates (echoviruses 7 and 30) and a coxsackievirus B3 (ATCC-VR 30). The putative common epitope region was localized in the N terminus of VP1 when the displayed recombinant proteins from the phages were chased by the convalescent-phase sera. The genomic region encoding the common epitope region was amplified and then expressed by using the vector pGEX-5X-1. The antigenicity of the expressed recombinant protein was identified by Western blotting with guinea pig antisera for six different serotypes of enteroviruses. After successive immunization of mice with the recombinant common epitope protein, splenocytes were extracted and hybridized with P3X63-Ag8-653 cells. A total of 24 hybridomas that produced monoclonal antibodies (MAbs) against the putative common epitope of enteroviruses were selected. Four of these were immunoglobulin G1 isotypes with a kappa light chain. These MAbs recognized 15 Korean endemic serotypes and prototypes of enteroviruses in an indirect immunofluorescence assay. These results suggest that the expressed protein might be a useful antigen for producing group common antibodies and that the use of the MAbs against the putative common epitope of enteroviruses might be a valuable diagnostic tool for rapidly identifying a broad range of enteroviruses.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Enterovirus/classificação , Epitopos/imunologia , Proteínas Virais/imunologia , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Clonagem Molecular , Primers do DNA , DNA Complementar/genética , Enterovirus/imunologia , Feminino , Humanos , Hibridomas , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Técnicas de Amplificação de Ácido Nucleico , Proteínas Virais/genética , Proteínas Virais/metabolismoRESUMO
In order to investigate the prevalence of sexually transmitted viruses (STVs) in Korean populations, the prevalence of specific antibodies to herpes simplex type 2 (HSV-2) and hepatitis C virus (HCV) and surface antigen to hepatitis B virus (HBsAg) was evaluated in blood donors (n = 200), voluntary visitors for STI testing in public health centre (n = 204), HIV seropositive individuals (n = 200) and commercial sex workers (CSWs) (n = 200). All blood samples were tested using commercial enzyme-linked immunosorbent assay (ELISA) kits that detect type-specific IgG to HSV-2 and HBsAg and anti-HCV. The prevalence of type-specific IgG to HSV-2 among the above four groups was 5.0%, 21.9%, 65.0% and 71.0%, respectively. The seroprevalence of HBsAg was 0% in blood donors, 7.0% of visitors for STI, 7.5% of HIV infected individuals and 1.2% in CSWs. That of anti-HCV was 0%, 2.5%, 5.0% and 10.3%, respectively. This study underlines the very high seroprevalence of STVs, especially HSV-2, in the group at high risk of STI. The prevalences of HCV in CSWs and HIV infected individuals were two to three times higher than STI patients. It means sexual transmission might be a possible route of transmission of HCV, because HIV infection is closely related with sexual behaviour in Korea. The spread of HSV-2 is dependent on sexual transmission and that the presence of antibody to HSV-2 may be suitable for use as a serological marker of the degree at risk of STI.