CRF01_AE, which has led a new epidemic in many provinces in China and has displayed complex characteristics, has now evolved into multiple clusters in China. Some clusters often circulate in specific regions or among specific risk populations in China. To better determine the characteristics of CRF01_AE circulating in Anhui Province, we analyzed CRF01_AE based on gag and pol sequences. Our results showed that CRF01_AE circulating in Anhui Province was clearly divided into three clusters. Cluster 1 covered 90% of the sequences in all CRF01_AE. Among Cluster 1, the sequences from men who have sex with men (MSM) and heterosexuals were interwoven. It is suggested that MSM may play a bridge role in transmitting HIV-1 among the different risk groups.