A combination of genetic polymorphisms increases the risk of progressive disease in chronic hepatitis C.

BACKGROUND: There is increasing interest in the influence of host genetic factors on hepatic fibrosis, and whether genetic markers can reliably identify subjects at risk of developing severe disease. We hypothesised that hepatitis C virus (HCV) infected subjects with progressive fibrosis, classified using strict criteria based on histology at biopsy in addition to disease duration would be more likely to inherit several genetic polymorphisms associated with disease progression compared with subjects with a low rate of disease progression. METHODS: We examined polymorphisms in eight genes that have been reported to have an association with hepatic fibrosis. RESULTS: Associations between polymorphisms in six genes and more rapidly progressing fibrosis were observed, with individual adjusted odds ratios ranging from 2.1 to 4.5. The relationship between rapidly progressing fibrosis and possession of > or =3, > or =4, or > or =5 progression associated alleles was determined and the adjusted odds ratios increased with increasing number of progression associated alleles (9.1, 15.5, and 24.1, respectively). Using logistic regression analysis, a predictive equation was developed and tested using a second cohort of patients with rapidly progressing fibrosis. The predictive equation correctly classified 80% of patients in this second cohort. CONCLUSIONS: This approach may allow determination of a genetic profile predictive of rapid disease progression in HCV and identify patients warranting more aggressive therapeutic management.

Use of denaturing gradient gel electrophoresis to detect mutation in VS2 of the 16S-23S rDNA spacer amplified from Staphylococcus aureus isolates.

To develop a double gradient denaturing gradient gel electrophoresis (DG-DGGE) based typing method that rapidly and accurately types clinical isolates of Staphylococcus aureus, the VS2 region of the 16S-23S rRNA spacer region (ISR) was chosen because of its potential high variation. The VS2 region was amplified with a 40-mer GC-clamp attached to the 5'-end of the reverse primer. The 145 bp PCR product was then separated by DG-DGGE using denaturant concentrations of 25-40% and polyacrylamide concentrations of 6-12%. Of the five mutations identified in 336 S. aureus isolates, one mutation was found to be highly specific for 161/171 (94%) of methicillin-resistant S. aureus (MRSA) isolates from different geographic locations and isolation times. This same mutation was found in 15/160 (9%) of penicillin- or methicillin-sensitive S. aureus isolates. In some isolates two mutations occured together in the one genome suggesting some S. aureus isolates have two copies of VS2. In these 336 isolates nine genotypes with different combinations of the five mutations were identified. In 18 coagulase-negative staphylococci (CNS), the MRSA-specific mutation was found along with two other mutations in all isolates demonstrating consistent differences in the presence of these mutations between CNS and S. aureus. The marked differences in VS2 sequences found between MRSA, methicillin- or penicillin-sensitive S. aureus (SSA), and CNS by DGGE in the present study may be useful in evolutionary studies and in the development of a specific assay for MRSA from clinical specimens.