Aiming to explore universal HIV testing, and to understand exact HIV prevalence in older general population, we conducted a community-based cross-sectional epidemiological investigation in two counties of Zhejiang province. Using census strategy and convenience sampling method, those participants who were older than 50 years and meet certain eligibility were enrolled, and HIV prevalence was presented by crude infection rate. A total of 215,441 (64.82%) were enrolled into this study, HIV testing was added into their health exam plan and 18 were confirmed as HIV infection, with a crude rate of 0.84/10,000. HIV prevalence was higher among men than among women in all age groups (p < 0.05). Unlike previous research, 14 cases (77.78%) were still with a relative high value of CD4 + count; 17 cases (94.44%) were infected by sexual transmission, based upon the epidemiological investigation two female cases were likely to be infected from their husbands. Active large-scale HIV screening by integrating into routine public health care can be an effective strategy to find people living with HIV at relative early progression.