Molecular Characterization of Hepatitis Delta Virus in the Western Amazon, Acre, Brazil

Background: Hepatitis delta (HDV) is the most severe form of chronic viral hepatitis, with rapid progression to cirrhosis and hepatocellular carcinoma, and remains neglected in the Amazon Basin. The state of Acre, in the Western Brazilian Amazon, is endemic for HDV, with a high prevalence of genotype 3 (HDV-3). This study aimed to analyze epidemiological data, quantify HDV RNA, identify genotypes, and describe the phylogeny and phylogeography of HDV in Acre. Methods: This cross-sectional study included patients positive for HBsAg and anti-HDV under follow-up at the Specialized Assistance Service (SAE) in Rio Branco, Acre, between March and November 2023. Blood samples were collected for HDV RNA quantification using one-step RT-qPCR. Samples with detectable viremia (Ct ≤ 30) underwent Nested-PCR, Sanger sequencing, phylogenetic analysis (Maximum Likelihood), temporal signal evaluation, and Bayesian phylogeographic reconstruction. Results: Among 108 patients (median age 43; 55.6% female), HDV RNA was detected in 48.1%, with viral loads ranging from 140 to 24,000,000 copies/mL. Of these, 55.8% had >100,000 copies/mL. Genotyping (n = 41) identified exclusively HDV-3. Phylogenetic analysis revealed genetic heterogeneity among HDV-3 isolates, with the formation of two major phylogenetic clades. Bayesian analysis estimated tMRCA around 1818 and suggested dispersion from Amazonas to Acre and neighboring regions. Conclusion: HDV-3 predominates in Acre with high genetic diversity, indicating sustained viral circulation in the Western Amazon and reinforcing the need for improved surveillance and diagnosis.