A high incidence of the shift of HBV genotypes has been reported in chronic hepatitis B patients dur ing antiviral therapy;however,the underlying molecular mechanisms are largely unknown.In this study,a large sample analysis method based on deep sequencing technology has been established to investigate whether the mixed infection of genotypes is responsible for the shift of the genotypes under drug or immunological pressure.Thirty-eight patients with chronic hepatitis B under adefovir dipivoxil (ADV) treatment were enrolled, and the genotypes of small fragments of serum HBV DNA were compared with deep sequencing technology and Sanger sequencing.With Sanger sequencing, 13 out of 38 patients showed a genotype shift mainly from C to B after ADV treatment for 48 weeks.Based on deep sequencing method,mixed infection with genotype B and C was found in all patients before shift.Further more,the mixed ratios of genotype were analyzed by Solexa in two hundreds chronic hepatitis B patients.The results revealed the existence of mixed infection in 95% (191/200) patients,and mixed ratios of genotype range from >0.02% in 95.5% of cases to >1% in 12% of cases.Our findings revealed that mixed infection of genotype B and C was a common phenomenon in chronic hepatitis B patients evidenced by deep sequencing results.HBV genotypes shift during antiviral therapy may due to pre-existence of mixed in fections of genotypes and different genotypes with different sensitivity to drug selection pressure.The deep se quencing of serum HBV DNA was a high sensitive and reliable method to detect mixed infection.