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Purpose: China has the world's largest burden of hepatitis B virus (HBV) infection. However, there are few studies reported on the latest prevalence of hepatitis B surface antigen (HBsAg) among pregnant women in China. We aimed to assess the prevalence of HBV infection and its associated risk factors among pregnant women in Yunnan, which is a province with relatively high prevalence of blood-borne viruses.Methods: We conducted a cross-sectional study of 15,641 pregnant women who underwent routine antenatal screening for HBsAg in Yunnan between 2013 and 2016. Logistic regression analyses were used to examine the associations between HBV infection and risk factors.Results: Overall, 7.89% (95% confidence interval (CI), 7.47%-8.31%) of the 15,641 pregnant women tested positive for HBsAg. Multivariable logistic regression analysis showed that advanced age (≥35 years old; adjusted odds ratio [aOR], 2.43-2.59), Hani ethnicity (aOR, 1.97; 95% CI, 1.22-3.17), unemployed women (aOR, 1.88; 95% CI, 1.30-2.72), multigravidas (aOR, 1.21; 95% CI, 1.05-1.40), and multiparas (aOR, 1.28; 95% CI, 1.10-1.49) were more likely to be infected with HBV. HBV infection was associated with cesarean section and preterm birth, with aORs of 1.31 (1.17-1.48) and 1.23 (1.00-1.51), respectively. In the subgroup analysis, the association between Hani ethnicity and HBV infection was significantly higher in women aged 35 years or older (aOR = 5.15; 95% CI, 1.51-17.55) than in women younger than 35 years of age (aOR = 1.69; 95% CI, 1.00-2.86).Conclusions: HBV infection among 15,641 pregnant women was of high-intermediate endemicity in Yunnan, China. Advanced age, Hani ethnicity, unemployment, and multigravid and multiparous statuses were associated with an increased prevalence of HBV infection. High-risk pregnant women should be paid more attention during routine antenatal examination. [ABSTRACT FROM AUTHOR]