Medicine

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Author: search.filters.author.Manamperi, A.Subject: search.filters.subject.Hepacivirus

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    Screening of hepatitis C (HCV) antibody reactive donors by RT-PCR in a sample population of blood bank donors in Sri Lanka
    (Malaysian Society of Parasitology and Tropical Medicine, 2007) Manamperi, A.; Gunawardena, N.K.; Nugawela, P.; Bandara, A.; Wellawaththage, C.; Bindusara, R.M.; de Silva, H.J.; Abeyewickreme, W.
    The practice of screening donors for HCV antibodies has substantially lowered the risk of acquiring HCV infection from a transfusion. However, detection of molecular markers in blood is the most reliable means of diagnosing active viral infection. Molecular studies on HCV antibody reactive donors have not been previously performed in Sri Lanka. The present study was carried out to investigate the RNA positive rates in a sample population of anti-HCV antibody reactive blood donors in Sri Lanka, with a view of determining whether RT-PCR testing for HCV RNA should be carried out at the initial donor screening level. Eighty nine (89) HCV antibody reactive donors were tested for the presence of HCV RNA by RT-PCR (sensitivity 200 copies/ml) during the period October 2005 to May 2006. The 89 serology positive donors were initially detected by a third generation ELISA by routine screening of an initial pool of 26,176 blood donors. Of the 89 Anti-HCV antibody positive donors (0.34% of the total donor pool), 6 (0.023% of the total donor pool, and 6.74 % of antibody positive individuals) were positive for HCV RNA. The prevalence of HCVRNA positivity was low in this cohort of Sri Lankan blood donors. This is in keeping with the low prevalence of HCV infection in the community. Routine individual HCV-RNA screening of donors does not seem cost-effective in our setting. The RNA negative, antibody positive profiles reflect either false positive serology results or donors who have been exposed to HCV previously and subsequently resolved their infections.
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    Application of nucleic acid technology (NAT) in the diagnosis of active viral replication in HBV and HCV infections and evidence for HBV surface antigen mutants
    (Sri Lanka Association for the Advancement of Science, 2008) Manamperi, A.; Gunawardene, Y.I.N.S.; Hapuarachchi, C.; Bandara, A.; Wellawaththage, C.; Abeyewickreme, W.; de Silva, J.
    Introduction: The community prevalence of Hepatitis B (HBV) and hepatitis C (HCV) infections, although considered low (< 1%) in Sri Lanka based on serological markers, pose a significant health threat to patients in high risk groups. The early diagnosis of active viral infection is crucial in such situations to prevent further transmission and to enable the clinicians to initiate successful therapeutic interventions. Objective: This study was carried out to investigate the usefulness of polymerase chain reaction (PCR) in the diagnosis of active viral replication in HBV and HCV infections. Methodology: All specimens from patients with serological evidence of hepatitis B (HBV surface antigen and/or antibodies for HBV core protein) or hepatitis C (antibodies for hepatitis C core protein-Anti-HCV) and referred to the Molecular Medicine Unit from May 2005 to May 2008 were analyzed by PCR and reverse-transcription PCR (RT-PCR) for HBV DNA (n=130) and HCV RNA (n=95) respectively. Results: Of the 130 patients tested, 57 (44%) were positive for HBV DNA. The positive group of patients included 10 renal transplant patients, 4 multiply transfused patients, 4 paediatric patients with lymphoma, and 1 patient with cirrhosis. Six HBV DNA positive patients had negative HBsAg serology profiles indicating the possibility of surface antigen mutant strains. The HBV DNA negative patients with positive serology profiles indicate sero-converted/ patients with resolved infections or false positive serology results. Of the 95 patients tested, 14 (15%) were positive for HCV RNA and included 3 paediatric patients with thalassaemia. HCV RNA negative, anti-HCV positive profiles reflect either false positive serology results (due to less specific antibody assays) or donors who have been exposed to HCV previously and subsequently resolved their infections. Conclusions: A major proportion of patients with serological markers for HBV have active viral infection whereas only relatively a minor proportion of patients with serological markers for HCV have active viral replication. We have also found the first possible evidence of hepatitis B surface antigen mutant strains. This underlines the importance of the nucleic acid based technology in the diagnosis and assessment of infection with or suspected to have hepatitis B or C infections. We also emphasize the importance of introducing NAT for screening donors for HBV DNA and HCV RNA to substantially lower the risk of acquiring HBV/HCV infection from a transfusion.
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    Clinical utility of PCR and real time PCR assays for Cytomegalovirus, hepatitis B and hepatitis C infections.
    (Sri Lanka Association for the Advancement of Science, 2008) Dassanayake, R.S.; de Silva, P.; Weerasena, J.; Gunawardene, Y.I.N.S.; Manamperi, A.
    Molecular Medicine Unit, Faculty of Medicine, University of Kelaniya, Reactivation of cytomegalovirus (CMV), Hepatitis B (HBV) and C (HCV) viruses from the status of latency is seen in immunocompromised individuals and such reactivation is often associated with morbidity and mortality in such individuals. The prevalence of these viral infections in a selected population of patients referred to the Molecular Diagnostic Laboratory at the Durdan's Hospital, Colombo, during the period from August 2007 to May 2008 were studied using qualitative PCR assays. All specimens from patients with suspected clinical diagnoses of either CMV or HBV or HCV infections were analyzed. Of 176 samples analyzed for CMV 78 were positive (37 males, 29 females) and majority of them are patients from a nephrology unit. Out of 40 and 10 samples analyzed from males and females, respectively, 22 and 4 were positive for HBV. Twenty six samples were analyzed for HCV and only 6 were fond to be infected with viruses and all of them were from males. Although PCR detection of these viral DNA/RNA is a sensitive method to detect infection, it lacks specificity for the detection of active viral disease and for monitoring the efficacy of antiviral therapy. Therefore, Real-time PCR (RT-PCR) assays for the detection and quantification of CMV-DNA, HBV-DNA and HCV-RNA were developed using SYBRgreen1 chemistry. The assays developed are capable of detecting viral particles in blood samples and quantifying viral DNA accurately over a broad range of input target copies (102 - 108copies/ml) and therefore, can be used to predict the reactivation of viruses by comparing with published kinetic criteria in clinical guidelines. Post PCR analyses of Real-time PCR products by agarose gel electrophoresis revealed bands having the same intensity for a wide range of target copies (103 -108copies/ml). In contrast, RT-PCR elicited higher cycle threshold for the descending order of concentration of target copies. Therefore, based on these results, it is evident that the intensity of conventional PCR bands should not be used for the assessment of viral reactivation or for monitoring therapeutic intervention and for this purpose RT-PCR is the method of choice
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