Generation of Infectious HDV Particles

The patient-derived HDV-1 isolate (HDV-1p) was isolated from a male individual with CHD from the university clinic of Hamburg,³³ (link) passaged in human liver chimeric uPA/SCID/beige/IL2RG^-/- (USG) mice, sequenced, cloned as genome-sense tandem dimer in pcDNA3.1(+), and infectious particles were produced in cell culture (Table 1). The HDV-3 isolate (Peru-1) was obtained from a young man from Peru, who developed severe acute hepatitis, which was cloned by Casey et al.³² (link) The origin of the first HDV clone available in the research community is less clear: it is a genotype 1 strain, individual(s) sera were passaged through various chimpanzees at NIH (J. Taylor, pers. commun.), inoculated in a woodchuck and then cloned (Table 1). For the production of the HDV-1a and HDV-3 strains, the HDV recombinant plasmid pSVL(D3) (kindly provided by J. Taylor, Philadelphia, PA, USA)²⁸ (link) and pCMV3-Peru-1.2 (kindly provided by J. Casey, Washington DC, USA)³⁴ (link) were used. Infectious HDV-1a, HDV-1p, and HDV-3 particles were generated in HuH7 cells as previously described.³⁵ (link) In brief, cells were transfected with equimolar amounts of HDV-1a, HDV-1p, or HDV-3 recombinant plasmids and the HBV envelope-expressing vectors pcDNA3.1(+)³⁶ (link) (HDV-1p) or pT7HB2.7³⁷ (link) (HDV-1a, HDV-3) encoding the surface proteins of HBV genotype D using Fugene HD Transfection Reagent (Promega, Madison, WI, USA) (Table 1).