Genetic characterization of HIV-1 epidemic in Anhui Province, China.

BACKGROUND: Anhui Province in China is facing a severe HIV epidemic with an increasing number of newly diagnosed cases. METHODS: In this study, HIV genetic characteristics in the province were investigated. Newly reported HIV-positive individuals from 15 districts of Anhui Province were enrolled and interviewed. Total viral RNA was extracted from plasma isolated from blood samples. We amplified and sequenced an HIV pol fragment of the 1062 bp. The sequences were used for determination of HIV subtypes and the presence of drug resistance mutations. Transmission networks were constructed to explore possible relationships. And all of assembled partial pol genes were submitted to the Stanford HIV Drug Resistance Database website to find the transmitted drug resistance. RESULTS: Partial pol gene sequences were obtained from 486 cases. The results showed that MSM was the most dominant transmission route (253, 52.06%), followed by heterosexual transmission (210, 43.21%) and blood-borne transmission (1, 0.21%). Many subtypes were identified, including CRF01_AE (226, 46.50%), CRF07_BC (151, 31.07%), subtype B (28, 5.76%), CRF08_BC (20, 4.12%), CRF55_01B (15, 3.09%), CRF68_01B (7, 1.44%), CRF67_01B (3, 0.62%), CRF57_BC (2, 0.41%), CRF59_01B (2, 0.41%), CRF79_0107 (2, 0.41%), subtype C (2, 0.41%), CRF64_BC (1, 0.21%), and circulating recombinant forms (URFs) (27, 5.55%). Four transmission subnetworks containing high transmission risk individuals (with degree ≥4) were identified based on CRF01_AE and CRF07_BC sequences, including two CRF01_AE transmission subnetworks constituted by elderly people with average ages of 67.9 and 61.5 years. Infection occurred most likely through heterosexual transmission, while the other two CRF07_BC transmission subnetworks consist mainly of MSMs with average ages of 31.73 and 34.15. The level of HIV-transmitted drug resistance is 3.09%. CONCLUSIONS: The simultaneous spread of multiple HIV subtypes in Anhui province underscores that close surveillance of the local HIV epidemic is necessary. Furthermore, the elderly people were frequently involved, arguing for behaviour intervention in this specific population besides the MSM risk group.

The inference of HIV-1 transmission direction between HIV-1 positive couples based on the sequences of HIV-1 quasi-species.

BACKGROUND: To infer transmission direction of a HIV transmission chain is helpful not only in legal jurisdiction but also in precise intervention to prevent HIV spread. Recently, the direction of transmission is inferred by whether paraphyletic-monophyletic (PM) or a combination of paraphyletic and polyphyletic (PP) topologies is observed or not between the sequences of source and recipient in the phylogenetic tree. However, paraphyly between them often declines over time and may disappear between spouses due to bidirectional transmission after primary infection. In this study, our aim is to test the reliability of inferring HIV transmission direction between epidemiologically linked HIV-1 positive couples using whether or not paraphyly is observed in phylogenetic tree. METHODS: HIV quasi-species were sequenced using PCR product clones, and then Bayesian analysis of molecular sequences with MCMC was employed to construct phylogenetic relationship of env, gag, pol gene fragments of HIV-1 positive couples using BEAST software. RESULTS: Our results showed that all sequences of seven couples except pol sequences of couple 12 and 13 form their own monophyletic cluster in phylogenetic tree including the closest control sequences from GenBank or other studies on local samples, which are supported by significant Bayesian posterior probabilities more than 0.9932. Of seven couples, paraphyly is only observed in phylogenetic tree constructed with env and pol gene sequences of three couples and gag gene sequences of four couples. Paraphyly is not observed in half of HIV positive couples. Pol sequences of couple 13 is separated by Blast selected controls; pol sequences of couple 12 in phylogenetic tree is supported by a lower Bayesian posterior value. CONCLUSION: Paraphyly relationship between sequences of donator and recipient is only observed among partial HIV-1 positive couples with epidemiological link. Phylogenetic relationship is not always the same when various gene regions of HIV are used to conduct phylogenetic analysis. The combination of phylogenetic analysis based on various gene regions of HIV and enough epidemiology investigation is essential when inferring transmission direction of HIV in a transmission chain or in one couple. However, while observed paraphyly can be used to infer transmission direction in HIV-1 positive couple, no observed paraphyly cannot deny it.

The prevalence of transmitted HIV drug resistance among MSM in Anhui province, China.

BACKGROUND: To optimize treatment regimens, we assessed human immunodeficiency virus (HIV) diversity and the prevalence of transmitted drug resistance (TDR) among men who have sex with men (MSM) in Anhui province, China. METHODS: A total of 139 MSM who were newly diagnosed and antiretroviral treatment-naive were enrolled in Anhui in 2011. A partial pol fragment was amplified and sequenced, and HIV subtypes were determined by phylogenetic analyses. Surveillance/transmitted drug resistance mutations (SDRMs) were identified according to the 2009 World Health Organization (WHO) list. RESULTS: A total of 133 (95.7%) samples were successfully amplified and sequenced. Based on phylogenetic analyses of the pol fragment, CRF01_AE accounted for 55.6% (74/133) of the infections, followed by CRF07_BC with 32.3% (43/133), B with 5.3% (7/133), and unique recombinant forms with 6.8% (9/133). A total of 3.0% (4/133) of the subjects were found to harbor HIV variants with SDRMs, including 1.5% with NRTI-related mutations and 1.5% with NNRTI-related mutations. PI-related mutations were absent. The SDRMs included L210W (1.5%), Y181C (0.8%), and G190A (0.8%). CONCLUSIONS: In Anhui, CRF01_AE strains contributed to most of the HIV infections among MSM, and the prevalence of TDR was relatively low in this population. Further studies should be performed to evaluate the trend of TDR among MSM in Anhui and to inform first-line antiretroviral treatment.

The characteristics of HIV-1 subtype B on phylogenetic dynamic and molecular transmission network in Fuyang City, China, 2011 to 2019.

Introduction: HIV-1 subtype B, as once one of the earliest strains introduced into mainland China rapidly spread in commercial plasma donors and heterosexuals in 1990s. Here, we aim to investigate the origin and evolutionary history of HIV-1 subtype B in Fuyang city, China. Methods: We collected sequences tested from Fuyang in the east of China where higher prevalence of HIV-1 among commercial plasma donors and heterosexuals to construct a phylogenetic tree using the Markov chain Monte Carlo (MCMC) algorithm, infer molecular transmission network using TN93 model and visualize it with Cytoscape software. Results and discussion: Our results showed that >99% of subtype B sequences belonged to Thai B. The sequences from Fuyang often cluster closer to those from other its adjacent cities, which clustered together and formed a monophyletic cluster. HIV-1 B circulating in Fuyang dates back to approximately 1990. Among the 1,437 sequences, 166 clustered at a genetic distance of ≤1.2%, resulting in 73 clusters. The degree of clustering with at least one other person was 11.55%. Among the transmission clusters, 50 (80.65%) comprised two individuals. Most clusters consisted of both heterosexual transmission routes and men who have sex with men. Phylogenetic and molecular network analyses revealed a common origin with neighboring regions in mainland China, local onwards transmission after its introduction, and a limited clustering degree. However, at least two co-existing transmission routes in most transmission clusters imply a greater challenge in controlling the spread of HIV-1. Our findings highlight the value on tailoring prevention interventions by combination of molecular surveillance and epidemiology.

Natural Killer Cells Induce CD8+ T Cell Dysfunction via Galectin-9/TIM-3 in Chronic Hepatitis B Virus Infection.

The antiviral response of natural killer (NK) cells and CD8+ T cells is weak in patients with chronic hepatitis B (CHB) infection. However, the specific characteristics of these cells and the association between NK cells and CD8+ T cell dysfunction is not well known. In this study, higher galectin-9 (Gal-9) expression was observed in circulating NK cells from CHB patients than from healthy controls and was found to contribute to NK cell dysfunction. In addition, circulating CD8+ T cells showed obvious dysfunction and overexpressed TIM-3, the natural receptor of Gal-9, during active CHB infection. Gal-9+ and Gal-9- NK cells from active CHB patients were sorted and cocultured with autologous CD8+ T cells. The proportion of tetramer+CD8+ T cells and the cytokines production of CD8+ T cells were lower after cocultivation with Gal-9+ than with Gal-9- NK cells. We showed that in vitro depletion of NK cells increased circulating hepatitis B virus (HBV)-specific CD8+ T cell responses in patients with active CHB infection. Because Gal-9 is increased in the serum of CHB patients, CD8+ T cells were sorted and cultured with exogenous Gal-9, resulting in lower IFN-γ, TNF-α, CD107a, and granzyme B levels, decreased expression of the activation receptor CD69, increased expression of TIM-3, and a high percentage of early apoptotic CD8+ T cells. Blocking Gal-9 or TIM-3 in vitro in a culture of peripheral blood mononuclear cells (PBMCs) stimulated with HBV peptide from active CHB patients restored CD8+ T cell function. However, blocking Gal-9 in vitro after removal of NK cells from PBMCs did not rescue CD8+ T cells exhaustion. Furthermore, NK and CD8+ T cells from active CHB patients were sorted and cocultured in vitro, and the exhaustion of CD8+ T cells were alleviated after blocking Gal-9 or TIM-3. In summary, overexpression of Gal-9 on NK cells, which interacts with TIM-3+CD8+ T cells and likely contributes to antiviral CD8+ T cell dysfunction, may be a potential target for the treatment of CHB patients.

The predominant cluster of CRF01_AE circulating among newly diagnosed HIV-1-positive people in Anhui Province, China.

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.