METHODOLOGY: Sixty-seven laboratory Salmonella enterica strains were tested. Six sets of primers targeting defined regions of the O antigen synthesis genes (rfb gene cluster) and Vi antigen gene (viaB) were selected and combined into a multiplex PCR for O-grouping. Four primers (H-for, Ha-rev, Hb-rev and Hd-rev) were used in the second step multiplex PCR for H-typing. The optimized mPCR assays were further evaluated with 58 blind-coded Salmonella strains.
RESULTS: The multiplex PCR results obtained showed 100% concordance to the conventionally typed serogroups. Validation with 58 blind coded Salmonella strains yield 100% accuracy and specificity.
CONCLUSION: Based on this study, PCR serogrouping proved to be a rapid, alternative method for further differentiation of Salmonella enterica.
DESIGN: A molecular epidemiology study was conducted among HIV-1 seropositive patients attending the University Malaya Medical Center (UMMC) from July 2003 to June 2004.
METHODS: Protease (PR) and reverse transcriptase (RT) gene sequences were derived from drug resistance genotyping assay of 100 newly diagnosed or antiretroviral-naive patients. These were phylogenetically analysed to determine the subtypes and recombination breakpoint analyses were performed on intersubtype recombinants to estimate the recombination breakpoint(s).
RESULTS: CRF01_AE predominated in Kuala Lumpur with 65% in both PR and RT genes. B subtype was detected at 14% and 12% in PR and RT genes, respectively. C subtype was present at 1% in both genes. Overall, the concordance of PR and RT genes in discriminating subtypes/circulating recombinant forms (CRF) was high at 96%. In this study, novel CRF01_AE/B intersubtype recombinants were detected at high prevalence (22%), including those isolates with subtype discordance. Thai variants of CRF01_AE and B subtype were involved in the genesis of these unique recombinant forms (URF). Interestingly, 19 CRF01_AE/B intersubtype recombinant isolates shared similar recombination breakpoints in both PR and RT genes. Several distinct URF were also identified.
CONCLUSION: PR and RT genes can be utilized for subtype/CRF assessment with high degree of agreement, allowing concurrent surveillance of circulating HIV-1 subtypes with antiretroviral drug resistance genotyping tests. The emergence of highly identical CRF01_AE/B intersubtype recombinants suggests the possibility of the appearance of a new circulating recombinant form in Kuala Lumpur.