Higher baseline viral diversity correlates with lower HBsAg decline following PEGylated interferon-alpha therapy in patients with HBeAg-positive chronic hepatitis B Hu Li, Li Zhang, Hong Ren, Peng Hu Department of Infectious Diseases, Key Laboratory of Molecular Biology for Infectious Diseases, Ministry of Education, Institute for Viral Hepatitis, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, China Background: Viral diversity seems to predict treatment outcomes in certain viral infections. The aim of this study was to evaluate the association between baseline intra-patient viral diversity and hepatitis B surface antigen (HBsAg) decline following PEGylated interferon-alpha (Peg-IFN-α) therapy.Patients and methods: Twenty-six HBeAg-positive patients who were treated with Peg-IFN-α were enrolled. Nested polymerase chain reaction (PCR), cloning, and sequencing of the hepatitis B virus S gene were performed on baseline samples, and normalized Shannon entropy (Sn) was calculated as a measure of small hepatitis B surface protein (SHBs) diversity. Multiple regression analysis was used to estimate the association between baseline Sn and HBsAg decline.Results: Of the 26 patients enrolled in the study, 65.4% were male and 61.5% were infected with hepatitis B virus genotype B. The median HBsAg level at baseline was 4.5 log10 IU/mL (interquartile range: 4.1–4.9) and declined to 3.0 log10 IU/mL (interquartile range: 1.7–3.9) after 48 weeks of Peg-IFN-α treatment. In models adjusted for baseline alanine aminotransferase (ALT) and HBsAg, the adjusted coefficients (95% CI) for ΔHBsAg and relative percentage HBsAg decrease were −1.3 (−2.5, −0.2) log10 IU/mL for higher SHBs diversity (Sn≥0.58) patients and −26.4% (−50.2%, −2.5%) for lower diversity (Sn<0.58) patients. Further analysis showed that the “a” determinant upstream flanking region and the first loop of the “a” determinant (nucleotides 341–359, 371–389, and 381–399) were the main sources of higher SHBs diversity.Conclusion: Baseline intra-patient SHBs diversity was inverse to HBsAg decline in HBeAg-positive chronic hepatitis B (CHB) patients receiving Peg-IFN-α monotherapy. Also, more sequence variations within the “a” determinant upstream flanking region and the first loop of the “a” determinant were the main sources of the higher SHBs diversity. Keywords: HBsAg decline, viral diversity, Shannon entropy, interferon, chronic hepatitis B