Datasets of sequences
To clarify the evolutionary correlation amongst HEV strains, seven sequence datasets (numbered I to VII) were built with sequences from this study. The first dataset (I) used to develop the phylogenetic tree, was built with the eight full genomes of this study and 59 complete HEV-3 genomes downloaded from the NCBI database; these include 18 HEV-3 subtype reference genomes (Smith et al., 2020), 40 HEV-3 complete genomes downloaded from the NCBI database, which share >89.0% BLASTn nucleotide identity (nt. id.) with the Italian sequences of this study. An HEV-4 sequence (HE-JA30, Acc. N° LC02274) serves as outgroup. For Bayesian analysis, six other datasets were built to clarify the evolutionary correlation amongst Italian strains, to determine the most recent common ancestor (TMRCA), and to calculate the HEV evolutionary rate. For each of these six datasets (II-VII), the selection of HEV sequences was based on four criteria: (i) sequences originated from Italy, (ii) year of collection, (iii) classified as genotype HEV-3, and (iv) sourced from swine, wild boar or human. The second dataset (II) comprises the 20 Italian HEV-3 full genome sequences available on the NCBI database and the eight full genome sequences reported upon in this study. Owing to the scarcity of full genome sequences for Italian strains of HEV, datasets III to VII were built using a 290bp short genomic region within ORF2, the most sequenced and represented region on NCBI database; in order to strengthen the analysis of the evolutionary correlation that exist amongst Italian strains. Dataset III comprises 41 HEV-3e and 93 HEV-3f sequences; dataset IV 23 HEV-3a subtype sequences; dataset V 56 HEV-3c subtype sequences; dataset VI 32 HEV-3n subtype sequences, and dataset VII 27 unclassified sequences. For each dataset nucleotide sequences were aligned using Mafft v7.475 (Katoh et al., 2019) and manually edited using Aliview (Larsson, 2014).