Chiyu Zhang

On-going generation of multiple forms of HIV-1 intersubtype recombinants in the Yunnan Province of China

ObjectivesTo investigate the molecular epidemiology of HIV in Chinas Yunnan Province, where the initial HIV-1 outbreak among injecting drug users (IDU) occurred in 1989, and to analyse the genesis and interrelationship of the epidemic with that in surrounding areas. DesignA molecular epidemiological investigation was conducted among IDU in three prefectures in Yunnan Province, including Wenshan (east), Honghe (southeast) and Dehong (west). MethodsThirty-nine specimens were collected from consenting IDU in 2000–2001. The nucleotide sequences of 2.6 kb gag-RT and 340 base pair (bp) env (C2/V3) regions were determined. Phylogenetic tree and recombination breakpoint analyses were performed. ResultsThe circulating recombinant form (CRF), CRF08_BC, predominated in east Yunnan near Guangxi Province (89% in Wenshan and 81% in Honghe), whereas it was not detected in Dehong (0/14) in the west. In contrast, 71% (10/14) of the Dehong isolates were unique recombinant forms (URF), mostly between subtypes B′ (Thailand variant of subtype B) and C, with distinct profiles of recombination breakpoints. The subtype B′ accounts for the remaining 29% (4/14) of Dehong isolates. Interestingly, two Honghe isolates (2/16) shared some of the precise B′/C recombination breakpoints with CRF07_BC. ConclusionNew recombinant strains are arising continually in west Yunnan near the Myanmar border. Some appeared to be secondary recombinants derived from CRF07_BC that had further recombined with other strains. The uneven distribution of subtypes, CRF and URF, suggests the presence of independent transmission networks and clusters among IDU in Yunnan.

High prevalence of HIV-1 and hepatitis C virus coinfection among injection drug users in the southeastern region of Yunnan, China.

&NA; The southeastern region of Yunnan province is a key site for drug trafficking and HIV‐1 infection spread from the west of Yunnan and Laos to southeastern China. To investigate the prevalence of HIV‐1 infection and hepatitis C virus (HCV) coinfection among injection drug users (IDUs) in southeastern Yunnan, three cohorts of 285 addicts, including 242 IDUs and 43 oral drug users, living in the cities of Gejiu and Kaiyuan and the county of Yanshan were studied. HIV‐1 and HCV infections were detected by enzyme‐linked immunosorbent assay and/or polymerase chain reaction. Data on the age, sex, risk behavior, drug use history, employment, ethnic background, and marriage status were obtained by interview. The overall prevalence of HIV‐1 infection was 71.9%. The rate of HCV coinfection among 138 HIV‐1‐infected IDUs was 99.3%. Most HIV‐infected IDUs were 20 to 35 years old (86.7%) and were ethnic Han (75.9%), suggesting that the epidemic in Yunnan is no longer confined to non‐Han ethnic minorities. HIV prevalence in female IDUs (81.2%) was significantly higher than in male IDUs (68.2%) (p < .05). The prevalence of HIV infection reached 68.4% after 1 year of injection drug use. Needle/syringe sharing is the major high risk factor for the spread of HIV‐1 and HCV infections. Large‐scale educational campaigns are urgently needed to reduce the spread of HIV and HCV infection in these regions.

Rapidly increasing prevalence of HIV and syphilis and HIV-1 subtype characterization among men who have sex with men in Jiangsu, China.

Objectives: To investigate the prevalence of HIV, hepatitis B (HBV), hepatitis C (HCV), and syphilis among men who have sex with men (MSM) in 2 cities of Jiangsu, China, and to characterize the HIV-1 subtypes prevalent among this population. Methods: During September 2006 and July 2007, 296 and 173 MSM were recruited from Nanjing and Yangzhou, respectively. Sera samples were collected and tested for HIV, HBV, HCV, and syphilis infections. The nucleotide sequences of p17 and C2V3 regions were determined by RT-nested-PCR and sequencing. HIV-1 subtypes were characterized by phylogenetic analysis. Results: The prevalence of HIV, HBV, HCV, and syphilis infections among MSM was 5.8%, 11.1%, 0.7%, and 27.7%, respectively. The prevalence of HIV and syphilis was significantly higher in 2006–2007 than in 2003 (P ≤0.0013) in Jiangsu than in other regions of China (P ≤0.003). In contrast, there was no significant difference in HBV and HCV prevalence between present and 2003 studies (P >0.05). The phylogenetic tree of p17 showed that HIV-1 subtypes B, CRF01_AE, and CRF07_BC accounted for 35.7%, 35.7%, and 28.6%, respectively. The result of C2V3 showed that 45.5%, 36.4%, and 18.2% sequences belonged to HIV-1 subtype B, CRF01_AE, and BC recombinants, respectively. The subtype characterization in Jiangsu was significantly different from those in Beijing (P <0.05). Furthermore, Jiangsu HIV-1 B strains were different from majority of China B′ strains and originated from Beijing. Conclusions: The rapidly increasing prevalence and complex subtypes of HIV-1 suggest that effective prevention and intervention strategies are urgently needed for MSM in Jiangsu.

Epidemiology of hand, foot, and mouth disease and genotype characterization of Enterovirus 71 in Jiangsu, China

BACKGROUND In the spring of 2008, an EV71-caused hand, foot, and mouth disease (HFMD) outbreak occurred in Fuyang city, Anhui Province, China. Jiangsu Province that borders Auhui to the east is presumed as a key station for the spread of EV71 to other regions of the Yangtze River Delta. OBJECTIVES To investigate the HFMD prevalence in Zhenjiang city of Jiangsu from May 2008 to October 2009, and the epidemic origin of EV71 circulating in Jiangsu. STUDY DESIGN During May 2008 and October 2009, a total of 6324 HFMD cases in Zhenjiang, Jiangsu, were investigated. Sixty throat specimens were randomly selected from different patients, and 28 nucleotide sequences of EV71 VP1 regions were successfully determined by RT-nested-PCR and sequencing. EV71 genotypes were characterized by phylogenetic analyses. RESULTS The incidence rate of HFMD was highest in the period of March-July and in the 1-4 years old age groups. Intriguingly, there was a slight predominance for boys and for children living in rural areas in HFMD infection. Phylogenetic analyses indicated that all Jiangsu EV71 strains and most China strains belonged to subgenotype C4a. CONCLUSION The C4a was the most prominent EV71 subgenotype circulating in China. Routine HFMD surveillance should be focused on the period of March-July, and more prevention efforts should be aimed at 1-4 years old children. Moreover, government efforts are urgently needed to improve public health condition and medical service quality in rural areas.

Identification and Characterization of a New Class of Human Immunodeficiency Virus Type 1 Recombinants Comprised of Two Circulating Recombinant Forms, CRF07_BC and CRF08_BC, in China

ABSTRACT We identified a new class of human immunodeficiency virus type 1 (HIV-1) recombinants (00CN-HH069 and 00CN-HH086) in which further recombination occurred between two established circulating recombinant forms (CRFs). These two isolates were found among 57 HIV-1 samples from a cohort of injecting drug users in eastern Yunnan Province of China. Informative-site analysis in conjunction with bootscanning plots and exploratory tree analysis revealed that these two strains were closely related mosaics comprised of CRF07_BC and CRF08_BC, which are found in China. The genotype screening based on gag-reverse transcriptase sequences of 57 samples from eastern Yunnan identified 47 CRF08_BC specimens (82.5%), 5 CRF07_BC specimens (8.8%), and 3 additional specimens with the novel recombinant structure. These new “second-generation” recombinants thus constitute a substantial proportion (5 of 57; 8.8%) of HIV-1 strains in this population and may belong to a new but yet-undefined class of CRF. This might be the first example of CRFs recombining with each other, leading to the evolution of second-generation inter-CRF recombinants.

Adaptive evolution of the spike gene of SARS coronavirus: changes in positively selected sites in different epidemic groups.

BackgroundIt is believed that animal-to-human transmission of severe acute respiratory syndrome (SARS) coronavirus (CoV) is the cause of the SARS outbreak worldwide. The spike (S) protein is one of the best characterized proteins of SARS-CoV, which plays a key role in SARS-CoV overcoming species barrier and accomplishing interspecies transmission from animals to humans, suggesting that it may be the major target of selective pressure. However, the process of adaptive evolution of S protein and the exact positively selected sites associated with this process remain unknown.ResultsBy investigating the adaptive evolution of S protein, we identified twelve amino acid sites (75, 239, 244, 311, 479, 609, 613, 743, 765, 778, 1148, and 1163) in the S protein under positive selective pressure. Based on phylogenetic tree and epidemiological investigation, SARS outbreak was divided into three epidemic groups: 02–04 interspecies, 03-early-mid, and 03-late epidemic groups in the present study. Positive selection was detected in the first two groups, which represent the course of SARS-CoV interspecies transmission and of viral adaptation to human host, respectively. In contrast, purifying selection was detected in 03-late group. These indicate that S protein experiences variable positive selective pressures before reaching stabilization. A total of 25 sites in 02–04 interspecies epidemic group and 16 sites in 03-early-mid epidemic group were identified under positive selection. The identified sites were different between these two groups except for site 239, which suggests that positively selected sites are changeable between groups. Moreover, it was showed that a larger proportion (24%) of positively selected sites was located in receptor-binding domain (RBD) than in heptad repeat (HR)1-HR2 region in 02–04 interspecies epidemic group (p = 0.0208), and a greater percentage (25%) of these sites occurred in HR1–HR2 region than in RBD in 03-early-mid epidemic group (p = 0.0721). These suggest that functionally different domains of S protein may not experience same positive selection in each epidemic group. In addition, three specific replacements (F360S, T487S and L665S) were only found between 03-human SARS-CoVs and strains from 02–04 interspecies epidemic group, which reveals that selective sweep may also force the evolution of S genes before the jump of SARS-CoVs into human hosts. Since certain residues at these positively selected sites are associated with receptor recognition and/or membrane fusion, they are likely to be the crucial residues for animal-to-human transmission of SARS-CoVs, and subsequent adaptation to human hosts.ConclusionThe variation of positive selective pressures and positively selected sites are likely to contribute to the adaptive evolution of S protein from animals to humans.

Evolutionary Dynamics of the Interferon-Induced Transmembrane Gene Family in Vertebrates

Vertebrate interferon-induced transmembrane (IFITM) genes have been demonstrated to have extensive and diverse functions, playing important roles in the evolution of vertebrates. Despite observance of their functionality, the evolutionary dynamics of this gene family are complex and currently unknown. Here, we performed detailed evolutionary analyses to unravel the evolutionary history of the vertebrate IFITM family. A total of 174 IFITM orthologous genes and 112 pseudogenes were identified from 27 vertebrate genome sequences. The vertebrate IFITM family can be divided into immunity-related IFITM (IR-IFITM), IFITM5 and IFITM10 sub-families in phylogeny, implying origins from three different progenitors. In general, vertebrate IFITM genes are located in two loci, one containing the IFITM10 gene, and the other locus containing IFITM5 and various numbers of IR-IFITM genes. Conservation of evolutionary synteny was observed in these IFITM genes. Significant functional divergence was detected among the three IFITM sub-families. No gene duplication or positive selection was found in IFITM5 sub-family, implying the functional conservation of IFITM5 in vertebrate evolution, which is involved in bone formation. No IFITM5 locus was identified in the marmoset genome, suggesting a potential association with the tiny size of this monkey. The IFITM10 sub-family was divided into two groups: aquatic and terrestrial types. Functional divergence was detected between the two groups, and five IFITM10-like genes from frog were dispersed into the two groups. Both gene duplication and positive selection were observed in aquatic vertebrate IFITM10-like genes, indicating that IFITM10 might be associated with the adaptation to aquatic environments. A large number of lineage- and species-specific gene duplications were observed in IR-IFITM sub-family and positive selection was detected in IR-IFITM of primates and rodents. Because primates have experienced a long history of viral infection, such rapid expansion and positive selection suggests that the evolution of primate IR-IFITM genes is associated with broad-spectrum antiviral activity.

Detection and identification of plasma bacterial and viral elements in HIV/AIDS patients in comparison to healthy adults

A low level of CD4+ lymphocyte cells makes end-stage HIV/AIDS patients highly susceptible to microbial infections. We have adopted the next generation sequencing method to identify the spectrum of bacterial plasma and viral elements that might be present in these patients. The HIV/AIDS plasma microbiome was dominated by bacterial elements in the taxonomical order Pseudomonadales, while healthy people carried fewer bacterial DNA in the plasma. We have found that many of the bacterial elements in HIV/AIDS plasma are similar to those of the microbes found in the human gut, suggesting potential acquisition of microbial elements from the gut. The HIV/AIDS and normal plasma DNA virome shared some similarities in the presence of common ubiquitous eukaryotic viruses. The normal DNA virome was mainly composed of viruses from Anelloviridae. In contrast, the HIV/AIDS DNA virome contained a large proportion of bacteriophages, endogenous retroviruses and a non-human virus. In addition, several sequences, which might belong to novel bacteria or endogenous retroviruses, were identified. Taken together, the use of high-throughput sequencing technology in unveiling microbial metagenomics may facilitate future research in combating HIV/AIDS and its associated microbial complications.

Extensive and complex HIV-1 recombination between B ', C and CRF01_AE among IDUs in south-east Asia

Objective:To investigate the subtype characterization of HIV-1 among IDUs in northern Myanmar. Design:A molecular epidemiological investigation was conducted among IDUs in Laza and Maizayang cities of northern Myanmar. Methods:A total of 83 HIV-1-positive serums were collected from consenting IDUs during June to August 2009. HIV-1 p17, pol, vif-env, C2V3 fragments were amplified and sequenced. Phylogenetic and bootscanning analyses were performed. Results:A very high proportion (86.1%) of HIV-1 intersubtype recombinants and very low proportion of subtypes B′ (3.8%), C (7.6%) and CRF01_AE (1.3%) were found in this HIV-infected IDUs cohort. These recombinants cover all four kinds of recombination forms formed among CRF01_AE, B and C. The B/C and CRF01_AE/B/C recombinants are the two most dominant recombinants, accounting for 54.4 and 42.6% of all cases, respectively, and indicating the ongoing generation of extensive and complex HIV-1 recombination among CRF01_AE, B′ and C in northern Myanmar. Intriguingly, most recombinants have different chimeric patterns from each other, forming 64 unique recombination forms (URFs) that are quite distinct from any previously identified circulating recombinant forms (CRFs) and URFs in Asia. Conclusion:The extremely high proportion of intersubtype recombinants, especially CRF01_AE/B′/C recombinants (42.6%), strongly suggests that northern Myanmar is a big forge for HIV-1 recombination among CRF01_AE, B′ and C.

Co-Evolution of Primate SAMHD1 and Lentivirus Vpx Leads to the Loss of the vpx Gene in HIV-1 Ancestor

Cross-species transmission and adaptation of simian immunodeficiency viruses (SIVs) to humans have given rise to human immunodeficiency viruses (HIVs). HIV type 1 (HIV-1) and type 2 (HIV-2) were derived from SIVs that infected chimpanzee (SIVcpz) and sooty mangabey (SIVsm), respectively. The HIV-1 restriction factor SAMHD1 inhibits HIV-1 infection in human myeloid cells and can be counteracted by the Vpx protein of HIV-2 and the SIVsm lineage. However, HIV-1 and its ancestor SIVcpz do not encode a Vpx protein and HIV-1 has not evolved a mechanism to overcome SAMHD1-mediated restriction. Here we show that the co-evolution of primate SAMHD1 and lentivirus Vpx leads to the loss of the vpx gene in SIVcpz and HIV-1. We found evidence for positive selection of SAMHD1 in orangutan, gibbon, rhesus macaque, and marmoset, but not in human, chimpanzee and gorilla that are natural hosts of Vpx-negative HIV-1, SIVcpz and SIVgor, respectively, indicating that vpx drives the evolution of primate SAMHD1. Ancestral host state reconstruction and temporal dynamic analyses suggest that the most recent common ancestor of SIVrcm, SIVmnd, SIVcpz, SIVgor and HIV-1 was a SIV that had a vpx gene; however, the vpx gene of SIVcpz was lost approximately 3643 to 2969 years ago during the infection of chimpanzees. Thus, HIV-1 could not inherit the lost vpx gene from its ancestor SIVcpz. The lack of Vpx in HIV-1 results in restricted infection in myeloid cells that are important for antiviral immunity, which could contribute to the AIDS pandemic by escaping the immune responses.