Objective: To assess the prevalence and related factors of HIV infection among male clients of the female sex workers in Hekou Yao autonomous county of Honghe Hani Yi autonomous prefecture (Hekou county) in Yunnan province in China, 2014-2015. Methods: Serial cross-sectional survey was conducted during June 2014 to November 2015. Convenience sampling methods were used to recruit the male clients for this study. Self-reported information on social-demographic characteristics, with sexual and drug behavior patterns, was gathered. Both blood and urine samples were collected for HIV, with for opiate testing. Multivariate logistic regression and Exhaustive CHAID method were used to determine the correlated factors associated with HIV infection. Statistical analysis was used by SPSS 22.0 software and Clementine 12.0 software. Results: The overall HIV prevalence of male clients was 2.06% (16/776). Male clients who keep using condom with female sex worker was estimated as 68.81% (534/776). The last commercial sexual partner of Vietnamese male clients was all Vietnamese female sex workers. Compared with Chinese male clients, Vietnamese male clients have a higher rate of morphine positive. Factors as: age ≥50 years vs. age <30 years (OR=8.11, 95%CI: 1.26-52.16) and testing for morphine positive vs. morphine negative (OR=7.35, 95%CI: 1.42-38.06) were significantly associated with HIV infection through multiple logistic regression analysis. Through Exhaustive CHAID, it confirmed that age was the primary factor that associated with HIV infection of male clients. Conclusions Relationship between morphine and HIV infection indicated that HIV prevalence of male clients in Hekou county was influenced by the combined effect of both illegal drug use and commercial sexual behavior. Special attention should be paid to male clients over 50 years of age, on HIV intervention.