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  • Title: Non-sequencing molecular approaches to identify preS2-defective hepatitis B virus variants proved to be associated with severe liver diseases.
    Author: Raimondo G, Costantino L, Caccamo G, Pollicino T, Squadrito G, Cacciola I, Brancatelli S.
    Journal: J Hepatol; 2004 Mar; 40(3):515-9. PubMed ID: 15123368.
    Abstract:
    BACKGROUND/AIMS: PreS2-defective hepatitis B virus (HBV) variants may emerge during chronic HBV infection. These variants carry mutation(s) at the ATG-start-codon and/or in-frame deletion into the preS2 genomic region and are commonly detected by sequencing analyses. We evaluated the prevalence of these variants in a large series of chronic HBV infected patients through non-sequencing molecular approaches. METHODS: We examined HBV isolates from 110 HBV carriers: 15 were inactive carriers (IC); 50 had chronic hepatitis (CH); 25 were cirrhotics; 19 had hepatocellular carcinoma (HCC). The entire preS2 genomic region was amplified by PCR technique. The amplicons were processed: (A) through electrophoresis on acrylamide gel to reveal deleted genomes; (B) through electrophoresis on agarose gel after digestion by NlaIII enzyme that cuts the wild ATG-start-codon but not the mutated one. RESULTS: We detected preS2 variants in 56/110 cases (51%). In particular, we found preS2-defective mutants in 2/15 IC, 25/50 CH, 13/26 cirrhotics, and 16/19 HCC. The presence of these variants was thus significantly associated with active infection and liver disease (P<0.002). Moreover, among cases with liver disease preS2-mutants were more prevalent in HCC patients (P<0.02). CONCLUSIONS: Our non-sequencing molecular methods are sensitive and specific, and simplify the identification of all preS2 HBV variant forms. Infection by these variants is significantly associated with active infection and HCC.
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