RESUMO
Mixed-genotypes hepatitis C virus (HCV) infections are normally ignored in chronic hemodialysis patients. The aim of this study is to investigate the prevalence of mixed-genotypes infections among hemodialysis patients in Pahang province, Malaysia. Reverse-transcription and polymerase chain reaction methods were performed using two different sets of primers, targeting the 5' untranslated region and nonstructural 5B region. Target region base sequences were obtained by direct sequencing. Discrepancy in outcomes from phylogenetic analysis of both regions suggests double infections. Of 40 subjects in eight hemodialysis centres, evidence of mixed-genotypes infections was found in 5 subjects (12.5%) from three different centres. Four patients were infected with mixed genotypes 3 and 1 and one with genotypes 3 and 4. Cases of mixed HCV genotypes infection were considered high among hemodialysis patients in Pahang. However, further investigation is needed to confirm whether they are true mixed infections or perhaps infection with recombinant virus and also to assess the clinicopathologic characteristics of the infection.
Assuntos
Hepacivirus/genética , Hepatite C Crônica/virologia , Diálise Renal , Sequência de Bases , Feminino , Genótipo , Hepatite C Crônica/epidemiologia , Humanos , Malásia/epidemiologia , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Prevalência , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The current available molecular method to detect infectious bursal disease virus (IBDV) is by reverse transcriptase-polymerase chain reaction (RT-PCR). However, the conventional PCR is time consuming, prone to error and less sensitive. In this study, the performances of Sybr Green I real-time PCR, enzyme-linked immunosorbent assay (ELISA) and conventional agarose detection methods in detecting specific IBDV PCR products were compared. We found the real-time PCR was at least 10 times more sensitive than ELISA detection method with a detection limit of 0.25pg. The latter was also at least 10 times more sensitive than agarose gel electrophoresis detection method. The developed assay detects both very virulent and vaccine strains of IBDV but not other RNA viruses such as Newcastle disease virus and infectious bronchitis virus. Hence, Sybr Green I-based real-time PCR is a highly sensitive assay for the detection of IBDV.
Assuntos
Infecções por Birnaviridae/veterinária , Ensaio de Imunoadsorção Enzimática/métodos , Vírus da Doença Infecciosa da Bursa/isolamento & purificação , Compostos Orgânicos/análise , Reação em Cadeia da Polimerase/métodos , Doenças das Aves Domésticas/virologia , Animais , Benzotiazóis , Infecções por Birnaviridae/virologia , Galinhas , Diaminas , Vírus da Doença Infecciosa da Bursa/genética , Quinolinas , RNA Viral/análise , Sensibilidade e EspecificidadeRESUMO
Mixed-genotypes hepatitis C virus (HCV) infections are normally ignored in chronic hemodialysis patients. The aim of this study is to investigate the prevalence of mixedgenotypes infections among hemodialysis patients in Pahang province, Malaysia. Reverse-transcription and polymerase chain reaction methods were performed using two different sets of primers, targeting the 5’ untranslated region and nonstructural 5B region. Target region base sequences were obtained by direct sequencing. Discrepancy in outcomes from phylogenetic analysis of both regions suggests double infections. Of 40 subjects in eight hemodialysis centres, evidence of mixed-genotypes infections was found in 5 subjects (12.5%) from three different centres. Four patients were infected with mixed genotypes 3 and 1 and one with genotypes 3 and 4. Cases of mixed HCV genotypes infection were considered high among hemodialysis patients in Pahang. However, further investigation is needed to confirm whether they are true mixed infections or perhaps infection with recombinant virus and also to assess the clinicopathologic characteristics of the infection.