Objective To investigate the epidemiology and mutation characteristics of pres/s gene in patients with hepatitis B virus (HBV) infection with double positive HBsAg and anti HBS (anti HBS) in Kunming area. Methods From May 2018 to April 2022, 438 patients with chronic HBV infection who quantitatively detected two and a half pairs of hepatitis B in our hospital and outpatient department were selected as the research objects. 202 HBsAg (+) /anti HBS (+) double positive patients were taken as the experimental group, and 236 HBsAg (+) /anti HBS (-) patients were taken as the control group. The difference of clinical data between the two groups were analyzed. The full-length sequence of pres/s gene was amplified by nested polymerase chain reaction, and the genotype was determined and the mutation characteristics of pres/s gene were analyzed. Results The prevalence rate of patients in the experimental group is higher in the population aged < 10 and above 80, while the prevalence rate of patients in the control group is higher in the population aged 20-49. Compared with the control group, the age of patients in the experimental group, the proportion of HBsAg < 250iu/ml, the infection of type C HBV and the proportion of hepatitis B carriers were significantly increased (P<0.05). A total of 132 HBsAg (+) / anti HBS (+) samples were successfully amplified and sequenced, including 81 B-type genes and 51 C-type genes; In HBsAg (+) /anti HBS (-) samples, there were 110 cases of B-type gene and 65 cases of C-type gene. In patients with type B gene, the mutation rate of N-terminal region in double positive group was significantly higher than that in single positive group (P<0.05). In patients with type C gene, compared with the single positive group, the mutation rates of MHR, a determinant and N-terminal region in the double positive group were significantly increased (P<0.05). Among 81 patients with type B gene, 14 (17.28%) had deletion mutation of pres gene, and 19 (37.25%) had deletion mutation of pres gene in 51 patients with type C gene, but no deletion mutation of pres gene was found in the single positive group.Conclusion HBsAg / anti HBS double positive HBV infection is common in HBsAg positive people aged < 10 and above 80. The rates of HBsAg mutation and PreS gene deletion mutation in double positive patients were higher than those in single positive patients, suggesting that double positive HBsAg/anti HBs may be related to HBsAg mutation and PreS gene mutation