In the last decade, next-generation sequencing (NGS) technology, which is characterized by being high-throughput, rapid, sensitive, and accurate, has developed rapidly. Main components of NGS are platforms: 454 sequencing; illumina sequencing; ion torrent sequencing; SOLID sequencing. NGS is used widely for the human immunodeficiency virus (HIV)-1. In this review, we focus on applications of the dynamics of HIV-1 quasispecies.
Animals ; HIV Infections ; virology ; HIV-1 ; classification ; genetics ; isolation & purification ; High-Throughput Nucleotide Sequencing ; methods ; Humans

This study aims to analyze the molecular epidemiological characteristics of HIV-1 strains prevailing among men who have sex with men (MSM) in Beijing, China. The pol gene fragments from 250 newly diagnosed HIV-1-infected MSM individuals during 2006-2010 in Beijing were amplified by RT-nested PCR, sequenced, and phylogenetically analyzed. HIV-1 pol gene from 189 individuals were amplified and analyzed; 81 (42. 9%), 3 (1. 6%), 2 (1.0%), 88 (46. 6%), and 15 (7.9%) individuals were infected with HIV-1 subtypes B, B', C, CRF01_AE, and CRF07_BC, respectively. The subtypes B and CRF01_AE could both be grouped into two clusters, and CRFO7_BC strains shared high homology and were presumed to originate from a common ancestor. The HIV-1 circulating in MSM in Beijing had a lower genetic diversity than in heterosexuals. The HIV-1 epidemic (2006-2010) in MSM in Beijing was actually a rapid spread of HIV-1 CRF01 AE and B, or rather native strains of the two viruses.
Adult ; China ; epidemiology ; Epidemics ; Genetic Variation ; HIV Infections ; diagnosis ; epidemiology ; virology ; HIV-1 ; classification ; genetics ; isolation & purification ; Homosexuality, Male ; statistics & numerical data ; Humans ; Male ; Middle Aged ; Molecular Epidemiology ; Molecular Sequence Data ; Phylogeny ; Young Adult

OBJECTIVEIn order to evaluate the distribution of genetic subtypes and epidemiological feature of HIV-1 circulating strains in Fujian province.

METHODSBlood samples and epidemiological data were collected from 104 newly infected patients who were distinguished by BED-CEIA methodology, during 2011-2012. Viral sequences(n = 81) of HIV-1 gag, env, and pol segments were amplified by nested PCR.

RESULTSSubtypes B and four Circulating Recombinant Forms, (CRF01_AE, CRF07_BC, CRF08_BC and CRF55_01B) were found in the samples, CRF01_AE(45.68%)and CRF07_BC(35.80%) were the two main HIV-1 strains in Fujian province. Compared with previous data, the proportion of CRF07_BC rose significantly while it gradually decreased in CRF01_AE. Heterosexual contact was still the principal transmission route in Fujian province, but the number of infection among men-who-have-sex-with- men grew rapidly.

CONCLUSIONResults from this study suggested that different subtypes of HIV-1 strain existed in Fujian province. The distribution of subtypes and the mode of transmission were changing with the progress of epidemic. Dynamic monitoring of the molecular epidemiology trends of HIV-1 infection should be enhanced.

Adult ; China ; epidemiology ; Female ; Genotype ; HIV Infections ; epidemiology ; HIV-1 ; classification ; genetics ; Humans ; Male ; Molecular Epidemiology ; Young Adult

To explore the genetic characteristics of viral quasispecies in HIV-1 CRF07_BC infections among intravenous drug users (IDU), the gp120 fragments of HIV-1 env gene were amplified from plasma samples collected from 6 CRF07_BC infected persons using single genome amplification and sequencing (SGA/ SGS) method, and 11 to 28 sequences were obtained from these samples, respectively, A neighbor-joining phylogenetic tree was reconstructed to describe the genetic characteristics of viral quasispecies. The Simplot, segments' phylogenetic trees and diversity plots based on average pairwise distance (APD) were used to identify the recombination events between quasispecies. The SGA sequences derived from single specimen formed a large monophyletic cluster in the neighbor-joining phylogenetic tree and showed the complex topologic structures of viral quasispecies. Of the 6 CRF07_BC infected patients, only one possessed the high genetic homogeneity, whereas the other five individuals showed high heterogeneity, with two to four subclusters inside the monophyletic cluster for each specimen. In addition, the recombinant events were identified among viral quasispecies from 3 cases. The results show SGA technique and phylogenetic analyses are useful tool to investigate the intrahost CRF07_BC gp120 complex quasispecies variation and high genetic diversity.
Adult ; Drug Users ; Female ; HIV Infections ; virology ; HIV-1 ; classification ; genetics ; isolation & purification ; Humans ; Male ; Molecular Sequence Data ; Phylogeny ; Substance Abuse, Intravenous ; virology ; Young Adult

This study aims to compare the influence of different genes to the results of HIV-1 subtype B' phylogenetic analyses. We first split 47 near full-length genome sequences of subtype B' into different regions (gag, pol, vif, vpr, vpu, env, nef), which derived from various risk populations and geographic regions from Thailand, Myanmar and China from published studies. Phylogenetic analyses were performed to each region obtained. The phylogenetic results of different regions were compared to that of the near full-length genome sequences. The pol gene was found to have the lowest diversity and evolutionary rate, and could repeat the phylogenetic results by using near full-length genome sequences. Although the env gene has the highest diversity and evolutionary rate, it could not achieve the similar results. This study compared the influence to the results of HIV-1 subtype B' phylogenetic analyses by using different genes and laid foundation for further molecular survey and analyses of the transmission of subtype B' in China.
Amino Acid Sequence ; China ; epidemiology ; Cluster Analysis ; Genes, Viral ; genetics ; HIV-1 ; classification ; genetics ; Molecular Epidemiology ; Molecular Sequence Data ; Phylogeny ; pol Gene Products, Human Immunodeficiency Virus ; chemistry ; genetics

To study the CTL antigen epitopes and drug resistance mutations of HIV-1 gag and pol genes through analyzing gag and pol gene sequences. The HIV-1 gag and pol gene fragments were amplified using nested polymerase chain reaction. A total of 23 PCR sequences, 449 cloned gag sequences and 402 cloned pol sequences were obtained. Sequence analyses showed the 23 samples were subtype B or B'. A total of 4 in 8 CTL antigen epitopes appeared 8 mutations in consensus sequence of subtype B and B'. There were no mutations found in the PCR sequences, whereas a few mutations were found in clone sequences (9.80%) in 5 antigen epitopes in p24 region. Eighteen PIs-related mutations and 24 RTIs-related mutations were found in PCR sequences and clone sequences in pol gene region, in which 17 (94.44%) PIs-related mutations and 15 (62.50%) RTIs-related mutations were found only in the clone sequences, respectively. The results showed that the prevalence of HIV-1 drug resistance strains in this study was at a higher level (17.39%), suggesting that some samples were resistant.to existing antiviral drugs.
Antigens, Viral ; immunology ; DNA Mutational Analysis ; Drug Resistance, Viral ; genetics ; Epitopes ; immunology ; HIV-1 ; classification ; drug effects ; genetics ; immunology ; Human Immunodeficiency Virus Proteins ; genetics ; Mutation ; Phylogeny ; T-Lymphocytes, Cytotoxic ; immunology ; gag Gene Products, Human Immunodeficiency Virus ; genetics ; pol Gene Products, Human Immunodeficiency Virus ; genetics

To investigate the subtype distribution of human immunodeficiency virus-1(HIV-1) infection among men who have sex with men (MSM) in Zhengzhou, Henan Province, forty blood samples were collected from HIV-1 carriers, who acknowledged to have sex with men. The complete gag gene was amplified by RT-PCR and nested-PCR and sequenced. All sequences were edited by BioEdit and subtyped by genotyping software. Phylogenetic analysis of gag gene were then performed using the MEGA 3.1 software, the gene distances were calculated by Distance program. There were three different HIV-1 subtypes including B, CRF01-AE and CRF07-BC present among twenty four MSMs in Zhengzhou. Genotyping results showed that 33.33% (8/24) were B, 41.67% (10/24) were CRF01-AE and 25% (6/24) were CRF07-BC, and subtype CRF01-AE had become the most prevalent HIV-1 subtype in Zhengzhou, Henan province. In conclusion, recombinant HIV-1 strains are circulating in Henan province and the epidemiology is complicated.
Adult ; China ; HIV-1 ; classification ; genetics ; isolation & purification ; Homosexuality, Male ; Humans ; Male ; Middle Aged ; Phylogeny ; Sequence Analysis, DNA ; Young Adult ; gag Gene Products, Human Immunodeficiency Virus ; genetics

OBJECTIVETo analyze the molecular epidemiological characteristics of HIV-1 subtype CRF01_AE strains being prevailed among HIV/AIDS in Zhejiang province in 2009.

METHODSA total of 303 subjects were identified by stratified random sampling among HIV infected individuals in Zhejiang province in 2009. Gag fragments of the HIV-1 strains were amplified by nested polymerase chain reaction from the DNA extracted from whole blood of HIV-1 infected individuals. PCR products were sequenced and analyzed by phylogenetic method.

RESULTSA total of 132 HIV-1 subtype CRF01_AE sequences were identified from the 225 samples that sequenced successfully, accounting for 58.67% (132/225). A total of 90.91% (120/132) CRF01_AE strains infected HIV/AIDS were transmitted mainly by sexual contacts. A total of 65.91% (87/132) of the cases infected by heterosexual route and 25.00% (33/132) by homosexual route. There were three main clusters in the phylogenetic tree. Pairwise DNA distance within three groups was 0.037 ± 0.011, 0.034 ± 0.008 and 0.047 ± 0.010, which has statistical significance (P < 0.05). Distribution of the sequence of homosexual behavior infected individuals was relatively concentrated in clusters one (96.97%, 32/33), and crossed with heterosexual behavior infectors, and presented the close relations with strains from Jiangsu province, Zhengzhou of Henan province, Liaoning province, Shijiazhuang of Hebei province.

CONCLUSIONThe CRF01_AE strains were the dominant subtypes among HIV infected individuals. The majority of the CRF01_AE infected cases had high risk sexual behavior. The heterosexual infected cases were more than homosexual cases. The circulating status of CRF01_AE strains in homosexual population was relatively independent, but also had evidence of transmission from man who have sex with man to heterosexual population.

Acquired Immunodeficiency Syndrome ; epidemiology ; virology ; Adolescent ; Adult ; China ; epidemiology ; Female ; Genetic Variation ; HIV-1 ; classification ; genetics ; Humans ; Male ; Middle Aged ; Molecular Epidemiology ; Phylogeny ; Young Adult

The purpose is to screen and identify the specific H-2d restricted T-cell epitopes. These epitopes are used to investigate the cellular immune response of BALB/c (H-2d) mice immunized with a HIV-1 vaccine which expresses six antigens of gp160, gag, pol, rev, tat and nef of HIV subtype B'/C. A replicating DNA vaccine and a non-replicating recombinant vaccinia virus vector, both expressing the six antigens mentioned above, were used to immune BALB/c (H-2d) mice in a prime-boost regiment. The six peptide libraries of HIV B'/C corresponding respectively to the six complete antigens were pooled according to a designed matrix format and used to test for IFN-gamma production from splenocytes of immunized mice by an enzyme-linked immunospot (IFN-gamma ELISPOT) assay. The ELISPOT data indicated that two of seven Gag-specific T-cell epitope peptides were identified to be the novel epitopes. One of three Pol-specific T-cell epitope is unreported. One novel epitope was confirmed in two gp160-specific T-cell epitope peptides. One Nef-specific T-cell epitope was identified. Three Tat-specific T-cell epitope peptides were continuous sequences in Tat peptide library and all contained either complete or partial sequence reported. Rev-specific T-cell epitope was not be found. The specific T-cell epitopes (H-2d restricted) were identified by IFN-7 ELISPOT assay, which could be used to detect the cellular immune response of BALB/c mice immunized with the HIV-1 vaccine expressing six antigens of gp160, gag, pol, rev, tat and nef of HIV subtype B'/C.
Amino Acid Sequence ; Animals ; Enzyme-Linked Immunospot Assay ; methods ; Epitopes, T-Lymphocyte ; chemistry ; genetics ; immunology ; Female ; H-2 Antigens ; chemistry ; genetics ; immunology ; HIV Antigens ; chemistry ; genetics ; immunology ; HIV Infections ; immunology ; virology ; HIV-1 ; classification ; genetics ; immunology ; Histocompatibility Antigen H-2D ; Humans ; Mice ; Mice, Inbred BALB C ; Molecular Sequence Data ; Peptide Mapping ; methods

OBJECTIVETo study the prevalent status of CRF01_AE strains of recombinant HIV-1 in Shenzhen and their source of infection in order to predict the epidemic trend and evolution.

METHODSA total of 489 samples of HIV-1 positive plasma were collected from 1992 to 2008 in Shenzhen. HIV-1 Env genes were amplified by nested-PCR from RNA. Subtype analysis were performed on the nucleotide sequence data. CRF01_AE sequences were analyzed by phylogenetic methods and characterized by calculating the genetic distance.

RESULTSA total of 300 CRF01_AE strain sequences were amplified, accounting for 64.5% of all genotyped samples of all the 465 samples. The CRF01_AE strains of recombinant in 1992 - 1999, 2000 - 2005 and 2006 - 2008 accounted for 56.8% (21/37), 68.4% (78/114) and 64.0% (201/314) of genotyped samples, respectively. And in these three periods of times, 52.4% (11/21), 43.6% (34/78) and 45.8% (92/201) were heterosexually acquired adults; 4.8% (1/21), 0.0% (0/78) and 22.4% (45/201) were homosexually acquired adults; 19.0% (4/21), 51.3% (40/78), 30.8% (62/210) were intravenous drug users, respectively.Phylogenetic tree analysis revealed that HIV samples from different period of time showed distinct aggregation in time and transmission as well as cross infection. The gene divergence rate of CRF01_AE strains in the three different periods of time were (8.783 ± 4.717)%, (11.054 ± 7.141)%, and (13.218 ± 4.080)%, respectively.

CONCLUSIONCRF01_AE is the major epidemic strains in Shenzhen, which is transmitted through heterosexual contact, MSM and intravenous drug users. The gene variation increased gradually as time goes by.

Acquired Immunodeficiency Syndrome ; epidemiology ; virology ; China ; epidemiology ; Evolution, Molecular ; Female ; Genotype ; HIV-1 ; classification ; genetics ; isolation & purification ; Humans ; Male ; Molecular Epidemiology ; Polymerase Chain Reaction ; Recombination, Genetic