ABSTRACT
Background & objectives: Multiple transfusions in ?-thalassaemia patients undergoing regular transfusion regimen are at a risk of developing transfusion transmitted infections, including hepatitis C virus (HCV). The present study was conducted to investigate the association of HCV viraemia and genotype with clinical parameters in HCV seroreactive ?-thalassaemic individuals. Methods: A total of 172 HCV seroreactive ?-thalassaemic individuals aged between 2-35 yr with at least 25 units of blood transfusion were catagorized into four groups (2-12 yr, group 1; 13-19 yr, group 2; 20-29 yr, group 3; 30-35 yr, group 4). Aged matched control samples (n=87; ?-thalassaemics without HCV infection) were also included. HCV RNA was detected by nested reverse transcriptase polymerase chain reaction (RT-PCR) based on 5� UTR of HCV genome, viral load was determined by real-time RT-PCR. Nested RT-PCR amplified partial core region was used for DNA sequencing. Liver function parameters [serum total bilirubin, alanine aminotransferase (ALT) and aspartate aminotransferase (AST)] were also determined. Results: Of the 172 HCV seroreactive individuals, 59.30 per cent (n=102) were HCV RNA positive. HCV viral load ranged from 173 to 32.04�[5] IU/ml; 87.65 per cent were infected with HCV genotype 3. Liver enzymes, such as ALT, AST and serum total bilirubin were significantly elevated in all age groups compared to control groups. Serum ferritin levels were found to be high in all individuals, but 16.27 per cent of HCV-infected individuals with >10,000 IU/ml viral load also showed high ferritin levels (>1500 ?g/l) where the majority of them were infected with HCV genotype 3. Interpretation & conclusions: HCV genotype 3 was the major circulating genotype among ?-thalassaemia patients in this region. Our findings indicated an association between HCV replication and hepatic iron load and also highlighted the need for sensitive quantitative RT-PCR-based detection of HCV RNA in the high risk population
ABSTRACT
Purpose: Rapid and specific detection of viral nucleic acid is increasingly important in the diagnosis of infectious diseases. The objective was to develop a rapid, efficient process of nucleic acid based detection of hepatitis C virus (HCV) infection for its diagnosis and treatment follow‑up. Materials and Methods: A two‑step nested reverse‑transcriptase polymerase chain reaction (RT‑PCR) has been standardised on a sample set of 125 individuals from different liver clinics in Kolkata. The method utilises a novel fast nested RT‑PCR for HCV detection and genotyping from HCV infected patient plasma with high processivity. Results: The overall time required from ribonucleic acid (RNA) isolation to nested PCR amplified product detection is reduced to 42% when compared with conventional nested RT‑PCR amplification. The method is sensitive as conventional PCR and detected all HCV RNA positive samples. Sequencing, phylogenetic analysis of the PCR amplified product by this method showed concordant genotypes with conventional PCR. Conclusion: Though being a two‑step process, this method is fast, cost‑efficient, reliable and feasible for regular HCV RNA screening and apt even in resource limited settings. This method could be translated to regular nucleic acid screening for other infectious diseases as regular PCR regimen.