To investigate whether transcription of hepatitis B virus (HBV) gene occurs in human sperm,total RNA was extracted from sperm of patients with chronic HBV infection (test-1),from donor sperm transfected with a plasmid containing the full-length HBV genome (test-2),and from nontransfected donor sperm (control),used as the template for reverse transcription-polymerase chain reaction (RT-PCR).Positive bands for HBV DNA were observed in the test groups but not in the control.Next,to identify the role of host genes in regulating viral gene transcription in sperm,total RNA was extracted from 2-cell embryos derived from hamster oocytes fertilized in vitro by HBV-transfected (test) or nontransfected (control) human sperm and successively subjected to SMART-PCR,suppression subtractive hybridization,T/A cloning,bacterial amplification,microarray hybridization,sequencing and the Basic Local Alignment Search Tool (BLAST) search to isolate differentially expressed genes.Twenty-nine sequences showing significant identity to five human gene families were identified,with chorionic somatomammotropin hormone 2 (CSH2),eukaryotic translation initiation factor 4 gamma 2 (EIF4G2),pterin-4 alpha-carbinolamine dehydratase 2 (PCBD2),pregnancy-specific beta-1-giycoprotein 4 (PSG4) and titin (TTN) selected to represent target genes.Using real-time quantitative RT-PCR (qRT-PCR),when CSH2 and PCBD2 (or EIF4G2,PSG4 and TTN) were silenced by RNA interference,transcriptional levels of HBV s and x genes significantly decreased (or increased) (P ＜ 0.05).Silencing of a control gene in sperm did not significantly change transcription of HBV s and x genes (P ＞ 0.05).This study provides the first experimental evidence that transcription of HBV genes occurs in human sperm and is regulated by host genes.