Introduction

Human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) continues to be among the most important threats to public health in China. Because of the large influence of traditional culture on society, homosexuality is stigmatized and people who are homosexual experience discrimination in China1,2. Men who have sex with men (MSM) are inclined to hide their sexual identity and engage in sexual behaviors in other regions rather than in their hometowns, where they may be easily recognized by acquaintances1,2,3. Thus, methods in addition to surveillance and other traditional epidemiological tools are needed to understand the characteristics of HIV-1 transmission among MSM. Previous studies showed that molecular analyses can be used to reliably estimate probable HIV-1 transmission networks4,5,6.

Guangxi, southwest China and adjacent to Vietnam, Yunnan, and Guangdong, is an area heavily affected by HIV-1 in China19, and gradually formed a large-scale epidemic in the area in recent years. In 2013, CRF01_AE (62.0%), CRF07_BC (25.0%), and CRF08_BC (6.5%) remained as the major strains20, with CRF07_BC accounting for 50.4% of cases in MSM in this study. These results suggest that CRF07_BC spread much easier among MSM and became a predominant strain in the region. CRF55_01B was first identified in Chinese MSM in 201221, and a recent paper has reported that CRF55_01B was identified in the MSM in 7 provinces with prevalence rates of 1.5–12.5%22. In that study, CRF55_01B accounted for 11.4% in MSM, with CRF55_01B widely disseminated among the MSM in the region. In addition to the major CRFs, we identified 2 CRFs (CRF67_01B, CRF68_01B) that were imported from outside of the province and 2 URFs (01_AE/CRF07_BC, CRF01_AE/C). A previous study reported that higher rates of dual-variant and multiple-variant HIV infection were found in MSM compared to in heterosexual individuals in the same populations23,24, increasing the probability of HIV-1 recombination. There is a growing concern that URFs may be prone to be formed in MSM because of the high-risk behaviour features of this population, including multiple sex partners, low rates of condom use, and anal intercourse.

It is challenging to trace the most probable infection route. We used novel molecular methods to explore the probable HIV-1 transmission network and risk associated with potential transmission. We showed that most HIV-1 infections were associated with cases outside of the province, and the main infection route was MSM. Moreover, the HIV-1 transmission network is concentrated; 3 networks contained more than 100 individuals, with the largest network containing 704 individuals. The finding reflects the existence of an HIV-1 super transmission network in the MSM in Guangxi and the characteristics of HIV-1 agglutinative transmission, which differs from those in other provinces of China9,17. Further analysis showed that 88.9% of individuals had more than 2°, meaning each subject was linked to at least two subjects; 451 (73.8%) subjects had more than 4° and were considered high-risk spreaders and more likely to spread HIV-1. One individual had 508° and was considered a super spreader. The finding reveals the existence of HIV-1 multiple partners and a super transmission network among MSM in Guangxi. Previous studies showed that individuals with more links in the network have a higher probability of spreading the virus to others because of their high viral load and high rate of partner change; therefore, these individuals may function as super-spreaders25,26. Chi-square test shows that marital status, subtype, and infection time were factors associated with potential transmission in networks, with a single status, CRF55_01B, and recent infection more likely to be associated with the virus spreading. This finding reveals why CRF55_01B has been widely disseminated among the MSM in the region in recent years. Thus, additional molecular epidemiology surveillance of CRF55_01B is required.

It is very important to analyze the geographic dimension of HIV-1 transmission among MSM within the province and throughout the country. Guangxi is located in southwest China and is adjacent to Vietnam, Yunnan, and Guangdong. This area of China is heavily affected by HIV-17. Nanning, the capital and largest city of Guangxi, is the provincial center in terms of the economy, culture, science, education, and tourism. Our findings revealed that Nanning is a major region of HIV-1 transmission, and the virus has spread to 13 other cities in Guangxi. Further analysis revealed that Nanning accounted for the dominant proportion of infections, not only in terms of local transmission (74.4%) but also in terms of cross-regional HIV-1 transmission (73.5%) based on the provincial transmission network, highlighting its remarkable role in the intra-provincial spread of HIV-1. The major province of cross-regional HIV-1 transmission with Guangxi is Guangdong, accounting for 57.5% of cases. Guangdong, a prosperous Chinese province adjacent to Guangxi, and it takes approximately 3 h to travel between Guangxi and Guangdong by high-speed railway. In the future, Guangdong is expected to become a major source of HIV-1 transmission. We also found that although most recent infections were associated with long-term infections, recent infections were closely related to potential transmission among networks, which is consistent with the results of previous studies18,27. This suggests that early HIV-1 infection plays an important role in transmission, emphasizing the need for early diagnosis and timely antiretroviral treatment.

Our findings revealed an estimated prevalence rate of drug-resistant HIV-1 strains of 4.8% among antiretroviral treatment-naïve MSM in Guangxi, representing a low level of TDR, which is consistent with the results of a previous study in the region28 and those of the studies in other regions of China9,29,30. However, the rate of drug resistance remains close to a moderate level according to the World Health Organization categorization method. Although it is not necessary to make large adjustments to current antiretroviral therapy regimens, further studies are essential for monitoring TDR.

This study has several limitations. First, most of the sequences from the other provinces were downloaded from the internet, which may not fully reflect the actual situation in the corresponding provinces. However, the results provide direction, which may strengthen regional cooperation. This is especially important in Guangdong, which is the province with the most cross-regional HIV-1 transmission with Guangxi. Second, although our findings show that MSM is the largest factor in HIV transmission, the sample sizes for other routes of infection, including among heterosexuals, were low. Third, a potential sampling bias; we could analyze only the samples that had been diagnosed, but those that had been infected but not diagnosed could not be included in the analysis. Another limitation is that LAg-Avidity cannot be completely accurate to distinguish recent infection and long-term infection, including viral load will be good to define recent infection. In the future, more samples will be included in the genetic transmission network.

We showed that network analysis based on a molecular approach can be used to trace the most probable HIV-1 transmission routes and infer additional features of an HIV-1 epidemic. Additionally, we determined the characteristics and risk of HIV-1 transmission among the MSM in Guangxi. These results can be used to design and implement evidence-based interventions. Further studies are required to determine how to combine the results of network analysis with public health approaches to enable effective HIV-1 intervention.