Genetic variability of hepatitis A virus genotype I strains in Bulgaria, 2011-2023

The HAV transmission generally occurs through faecal-oral transmission, contaminated water or food, poor sanitary conditions or poor hygiene. Bulgaria is considered a country in transition between an intermediate and a low endemicity profile with significant number of cases and cyclic outbreaks. A phylogenetic, comparative analysis was applied to genotype I Bulgarian sequences collected from 2011 to 2023 to investigate the HAV circulation, the molecular epidemiology, the phylogenetic relationships respect to foreign sequences and gene flows between different Bulgarian towns.

Five different HAV IA and IB VP1-2A dataset were investigated through phylogenetic analysis. The sequence alignments were conducted through MAFFT, under the Galaxy platform, followed by manual editing with Bioedit. Bayesian phylogeny was obtained with BEAST, the gene flow migration analysis in Bulgaria was explored through Mac Clade.

Genotyping of the HAV sequences from Bulgaria revealed two main supported clades corresponding to the IA (n = 115) and IB (n = 120) subtypes. Phylogenetic analysis in IA highlighted a major cluster with strains strictly related to the frozen berries European outbreak. Other minor clusters indicated the simultaneous circulation of different variants. In HAV IB a major cluster of very similar sequences was reported, confirming the low variability. The Bayesian tree indicated intermixing between Bulgarian and foreign strains both in IA and IB. Time scaled phylogeny revealed that HAV IA began circulating in Bulgaria before the genotype IB, confirming previous data. For HAV genotype IA the gene flow analysis indicated bidirectional exchanges involving Sofia and Samokov (each 3.3% of exchange) and Samokov with Dolna Banja (each 3.3% of exchange). Other supported flows (3.3% of exchange) were identified. For IB gene flow analysis showed a 30% from Sofia to Samokov, a 10% from Sofia to Pernik, from Sofia to Novachene and from Novachene to Botevgrad.

The Bayesian approach allowed to estimate the origin, circulation of the HAV epidemic in Bulgaria, to follow the trend of the infections during time. Gene flows could be explained by infection transmitted through trade, importation/exchange of contaminated food, population movements and geographic proximity. The findings can be useful in public health to plan more effective preventive measures.