Bayesian evolutionary rate estimate and dated trees
Root-to-tip regression analysis of the temporal signal from datasets
revealed a moderate association between genetic distances and sampling
years.
The best fitting models, Correlation coefficient, the estimated mean
value of the evolutionary rate and the most common recent ancestor
(TMRCA) for each dataset are provided in Table 2. Due to the low
Correlation coefficient, the phylodinamic analysis was not performed on
dataset VII.
In the bayesian tree, built using complete genome (dataset II;
Supplementary Figure 1), two main clades were found: one included HEV-3e
and HEV-3f sequences separated back to 1952 (95% HPD: 1918-1988) and
the other HEV-3a, HEV-3c, HEV-3l, HEV-3n and unclassified strains
originated back to 1936 (95% HPD: 1890-1971). The classification of
Italian strains based on Maximum likelihood tree correlated with the
Bayesian tree.
In dataset III (Supplementary Figure 2), built with HEV-3e and HEV-3f
ORF2 short genomic regions, the WB110VT2017 clustered with wild boar
strains previously reported (WB161VT2017, MK889004; WB125VT2017,
MK889005, WB02VT2016, MG582608) in the same time period and with two
human strains reported in 2018 and 2019 (INMI_1902_2019, MN537876;
ISS_ID_258/2018, MZ274246). The WB03VT2016 clustered with a correlated
wild boar strain WB02VT2016 (MG582608). Both wild boar and human strains
were detected in the same region (Lazio).
Similarly, the 55863/2018 strain clustered with other wild boar strains
reported from the same region (Umbria region, e.g.: 59068.5/UM/2018,
LR699563). However, 55863/2018 clustered also with a human strain from
Lazio (ISS_ID_305/2019, MZ274253) and the cluster also included a
swine HEV-3 strain (14RS333-8_50549-4, MK532935) detected in Northern
Italy in 2014.
In dataset IV (Supplementary Figure 3), built with the few sequences of
HEV-3a ORF2 available for Italian strains, two clusters were apparent.
The WB01VT2016 strain clustered with two strains sequenced from wild
boar hunted in the same area in 2012 (Hepatitis_E_virus_isolate_5,
MH836549; Hepatitis_E_virus_isolate_116, MH836544) and with the
strain INMI_1736_2017 (MN444837) isolated from a patient in 2017; all
strains found in the same region (Lazio). The second HEV-3a cluster
comprised all of the sequences isolated from wild boar in northern Italy
(Supplementary Figure 3).
In dataset V (Supplementary Figure 4), built with HEV-3c ORF2, two main
clusters were apparent: the first grouping Italian swine sequences
reported in northern Italy and the second including Italian human and
wild boar sequences collected in central and northern Italy. The strain
WB119VT2017 clustered with several wild boar and human strains reported
in central Italy (Lazio, Abruzzo and Umbria).
In the bayesian tree built with HEV-3n ORF2, dataset VI (Supplementary
Figure 5), two main clades were observed: the first one only formed by
wild boar strains reported in northern Lazio (Viterbo province) between
2016/2017 and including the wild boar strain WB171VT2017 from this
study; the second clade included strains isolated from wild boar in
Umbria (land border with northern Lazio) in 2016/2017 and wild boar
strains from southern Italy collected in 2017.