The Implications of HIV-1 Subtypes on Pathogenesis, Reservoirs and Cure Strategies.

What role does HIV-1 subtype play in transmission and pathogenesis? An epidemiological perspective.

A recent study demonstrated a greater propensity for HIV-1 subtype C to develop a K65R mutation under in-vitro selection with tenofovir compared with subtype B or other non-B subtypes [1]. The mechanistic basis for this in-vitro observation was not clear because the single nucleotide change is identical for the K65R substitution in either subtype B or subtype C. For subtype B, a switch from AAA to AGA occurs. For subtype C, a switch from AAG to AGG occurs. The third codon position, however, is different for subtype C, reflecting redundancy in the genetic code for lysine and a natural variation among different HIV-1 subtypes. The authors speculate that this third codon position difference or other genetic changes may influence the propensity for the development of K65R in subtype C HIV-1. We evaluated patients with subtype C, subtype B, and other non-B HIV-1 subtypes for virological failure and the development of resistance in two phase III clinical trials of tenofovir disoproxil fumarate (DF). These studies enrolled patients primarily from the United States, Europe and South America. Within these studies, approximately 7% of the 1200 patients enrolled were infected with non-B HIV-1 subtypes. Study 903 assessed the combination of tenofovir DF with lamivudine and efavirenz, and study 934 assessed the combination of tenofovir DF with emtricitabine and efavirenz. Although neither study was statistically powered to address efficacy in patients with non-B subtypes, the virological failure rates were similar between patients with subtype B or non-B HIV-1 subtypes (15.5 versus 19%, respectively, P = 0.75 for study 903 at week 144; 19 versus 13%, respectively, P = 1.0 for study 934 at week 48) [2,3]. Moreover, among the 10 patients with subtype C HIV-1 treated with tenofovir DF in both studies, only one patient was classified as a virological failure, and this patient did not develop a K65R mutation or any other resistance mutations. One patient developed K65R with a non-B subtype, a subtype AG patient from study 903. This patient did not show the AAG polymorphism at codon 65, but rather the AAA K65 codon typically found among subtype B patients. In summary, in highly suppressive antiretroviral regimens that include tenofovir DF, efavirenz and either emtricitabine or lamivudine, there was no evidence of a higher rate of virological failure or the development of K65R among patients with non-B HIV-1 subtypes or in particular among patients with subtype C HIV-1. The studies were limited by the number of patients enrolled with non-B subtypes. A larger study of tenofovir DF with zidovudine and lamivudine in African patients infected with subtypes A, C and D, however, concluded similarly that patients with subtypes A, C and D respond effectively to that drug combination with a low incidence of the development of K65R (three out of 20 virological failures; 15%) [4]. Another study [5], conducted exclusively among subtype C-infected patients in Botswana, demonstrated a higher rate of K65R development among patients treated with didanosine, stavudine, and efavirenz (seven out of 23 virological failures; 30%). The rate of thymidine analogue mutation development was also high in that trial, with eight out of 13 (62%) virological failure patients who were taking combivir developing thymidine analogue mutations, suggesting an overall higher rate of resistance development in that study. Overall, additional studies with the prospective enrollment of patients with subtype C and other comparator HIV-1 subtypes will be necessary to rule out any differential propensity for the development of K65R among subtype C patients in the clinical setting. Currently available clinical data, however, do not suggest a higher rate of K65R development among patients with subtype C.

HIV clinics in Canada provide care to an increasing number of patients born outside of Canada with HIV-1 non-B subtype infections. Because the Easy Q HIV-1 v1.2 assay (EQ; bioMérieux) failed to detect some non-B subtype infections, a multiassay HIV-1 viral load (VL) study was conducted with patients with diverse HIV subtype infections. Patients were enrolled from the Southern Alberta HIV Clinic (SAC), Calgary, Alberta, Canada (n = 349) and the McGill HIV Clinic (MHC), Montreal, Quebec, Canada (n = 20) and had four or five tubes of blood drawn for testing by EQ and three other commercial HIV VL assays: (i) the Versant 3.0 HIV-1 test, with the Versant 440 instrument (branched DNA [bDNA]; Siemens), (ii) the RealTime HIV-1 test, with the m2000rt instrument (m2000rt; Abbott Molecular Diagnostics), and (iii) the COBAS AmpliPrep TaqMan HIV-1 48 test (CAP-CTM; Roche Molecular Diagnostics). Blood was processed according to the individual manufacturer's requirements and stored frozen at -86°C. The HIV subtype was known for patients who had undergone HIV genotypic resistance testing (Virco, Belgium). Data analyses were done using standard statistical methods within Stata 9.0 (StataCorp, College Station, TX). A total of 371 samples were tested on 369 patients, of whom 291 (81%) had a Virco genotype result of B (195; 53%) or non-B (96; 26%) subtypes A to D and F to K, as well as circulating recombinant forms (CRFs) (i.e., CRF01_AE and CRF02_AG). Most (58/78; 74%) patients of unknown subtype were recent African emigrants who likely have non-subtype B infection. Overall bias was small in pairwise Bland-Altman plots, but the limits of agreement between assays were wide. Discordant viral load results occurred for 98 samples and were due to missing values, false negatives, and significant underquantification that varied by HIV subtype. Results were obtained for all 371 samples with m2000rt, but for only 357 (97%) with CAP-CTM, 338 (92%) with EQ, and 276 (75%) with bDNA due to errors/equipment failures. False-negative results (nondetection of viral RNA versus other assay results) occurred for all platforms, as follows: for m2000rt, 8 (2%) [B(4) and non-B(4) subtypes], CAP-CTM, 9 (2.5%) [B(6) and non-B(3) subtypes]; EQ, 20 (6%) [B(7) and non-B(13) subtypes]; bDNA, 5 (2%) [B(1) and C(4)]. EQ and bDNA had the highest rates of underquantification by ≥ 1.0 log(10) copies/ml, mainly for HIV non-B subtypes. Performance significantly varied between HIV VL platforms according to subtype. HIV viral diversity in the population being tested must be considered in selection of the viral load platform.

Summary: The HIV-1 subtype distribution was determined in 41 HIV-positive women (˜8% of all HIV-infected women in Denmark) belonging to different risk groups. HIV p17 gag and env gene subtypes were determined by DNA sequence analysis. Five different HIV subtypes were detected across all patients. Most HIV-1-positive women of Danish origin carried subtype B viruses, and a minority had virus belonging to subtypes A or C. All injecting drug users (IDUs) were infected with HIV subtype B viruses, whereas all non-B subtypes were present in cases linked to heterosexual transmission. In contrast, all women of African origin carried non-B HIV subtypes (subtypes A, C, D, or G) regardless of transmission mode. Of these women, 21% infected with non-B HIV subtypes appeared to be infected by subtype chimeric viruses and 7% were jointly infected with viruses belonging to two different subtypes (A and C). Data demonstrate a preferential representation of non-B HIV subtypes in women infected through heterosexual contact, as well as a high degree of recombination between viruses derived from endemic areas in which several HIV subtypes predominate. Combined with the increased incidence of heterosexual transmission of HIV, the results imply that an increased subtype diversity can be anticipated in newly infected individuals.

The HIV-1 subtype distribution was determined in 41 HIV-positive women (-8% of all HIV-infected women in Denmark) belonging to different risk groups. HIV p17 gag and env gene subtypes were determined by DNA sequence analysis. Five different HIV subtypes were detected across all patients. Most HIV-1-positive women of Danish origin carried subtype B viruses, and a minority had virus belonging to subtypes A or C. All injecting drug users (IDUs) were infected with HIV subtype B viruses, whereas all non-B subtypes were present in cases linked to heterosexual transmission. In contrast, all women of African origin carried non-B HIV subtypes (subtypes A, C, D, or G) regardless of transmission mode. Of these women, 21% infected with non-B HIV subtypes appeared to be infected by subtype chimeric viruses and 7% were jointly infected with viruses belonging to two different subtypes (A and C). Data demonstrate a preferential representation of non-B HIV subtypes in women infected through heterosexual contact, as well as a high degree of recombination between viruses derived from endemic areas in which several HIV subtypes predominate. Combined with the increased incidence of heterosexual transmission of HIV, the results imply that an increased subtype diversity can be anticipated in newly infected individuals.

Characteristics and Outcome of a Cohort of HIV-1 Non-B Subtype-Infected Patients After a 10-Year Follow-up Period: A Single Centre Experience.

To describe the demographics, risk behaviors, and HIV-1 subtypes in a large cohort of recently HIV-infected military personnel. Descriptive, cross-sectional study. US military personnel with recent HIV seroconversion from six medical referral centers were enrolled with a self-administered questionnaire, CD4 cell counts, syphilis and hepatitis B serologies, plasma viral RNA levels, and HIV-1 subtype nucleic acid sequencing. Between February 1997 and May 2000, 520 patients were enrolled. Most [488 (94.3%)] were infected with HIV-1 subtype B. The most prevalent non-B subtype was a circulating recombinant form (CRF01_AE) [17 (61%)]; however, two pure subtypes (C and D), as well as CRF02_AG, CRF09_cpx and a BE recombinant were identified. The likely area of HIV-1 acquisition was the United States for 70% of the volunteers. At least three non-B subtype infections (two subtype C, one subtype CRF01_AE) were apparently acquired domestically. Risk behaviors and comorbid sexually transmitted diseases were reported during the seroconversion period. Volunteers with non-B subtype HIV infection were more likely to report heterosexual contacts [92% vs. 39%; odds ratio (OR), 10.0], including contacts with commercial sex workers (41% vs. 13%; OR, 4.9). The Roche Amplicor version 1.0 assay was less sensitive for non-B subtype infections than the Roche Amplicor version 1.5 assay. There is a high prevalence and diversity of non-B HIV subtypes in this large cohort. Efficient diagnosis of acute primary HIV-1 infection was identified as a goal for prevention programs. Modifiable risk behaviors and target populations for intervention were identified.

Phylogenetic inferences on HIV-1 transmission

The objective of this study was to investigate factors influencing mother to child transmission of HIV-1 in Thailand, where HIV-1 CRF01_AE, the major subtype in Southeast Asia, predominates. Samples from 84 HIV-1 infected, anti-retroviral treatment-naïve, non-breast feeding mothers, 28 who transmitted HIV-1 to their babies (transmitters) and 56 who did not (non-transmitters), were studied for maternal humoral immune response and virus characteristics. Maternal humoral immune response was measured by lymphocyte phenotyping; neutralizing antibodies to laboratory HIV-1 MN strain and two clinical isolates; peptide binding antibody to gp41 and V3 from strains CRF01_AE, B, and MN; autologous antibodies; and quasispecies diversity. Virus characteristics studied were viral load, co-receptor usage, and viral replication capacity. No significant difference between transmitters and non-transmitters was found for any parameter of maternal humoral immune response. However, viral load and viral replication capacity were significantly higher in transmitters versus non-transmitters and were not correlated with each other. This suggests that viral replication capacity may be a transmission factor independent of viral load, which is already well established as a risk factor for transmission of HIV-1. All except four viral isolates used the CCR5 co-receptor. This is one of few studies of vertical transmission in a population where HIV-1 CRF01_AE predominates. The data suggest that in this population the maternal humoral immune response was not important in preventing transmission at parturition, but that virus characteristics were key factors, and that viral replication capacity may contribute to birth-associated mother to child transmission of HIV-1.

Various studies have demonstrated the increasing prevalence of non-B HIV-1 subtypes in Western Europe. In contrast, knowledge about the molecular epidemiology of HIV-1 in Central and Eastern Europe is limited. The objective of present study was to investigate the HIV-1 molecular diversity as well as time trends in HIV-1 subtype distribution in Slovenia. A retrospective molecular epidemiological survey was conducted on a cohort representing 88% (131/149) of all HIV-1 infected patients diagnosed between January 1996 and June 2005. The study revealed that subtype B is a predominant HIV-1 subtype in Slovenia (110/131; 84%), although a relatively high proportion (21/131; 16%) of non-B subtypes was found. Among them, a high proportion of recombinant (10/21; 48%) and different unclassified strains (8/21; 38%) were identified. Non-B subtype viruses were predominant among heterosexuals (19/21; 90%) and subtype B viruses among men who have sex with men (84/110; 76%). Importantly, 86% (18/21) of patients infected with non-B subtypes were of Slovenian nationality. In contrast to Western European countries, a significant increase (P = 0.015) in the proportion of men who have sex with men was observed recently among newly diagnosed HIV-1 infected patients in Slovenia.

HIV-1 CRF01_AE coreceptor usage prediction using kernel methods based logistic model trees

Human immunodeficiency virus type 1 (HIV-1) tropism can be assessed using phenotypic assays, but this is quite laborious, expensive, and time-consuming and can be made only in sophisticated laboratories. More accessible albeit reliable tools for testing of HIV-1 tropism are needed in view of the prompt introduction of CCR5 antagonists in clinical practice. Bioinformatics tools based on V3 sequences might help to predict HIV-1 tropism; however, most of these methods have been designed by taking only genetic information derived from HIV-1 subtype B into consideration. The aim of this study was to evaluate the performances of several genotypic tools to predict HIV-1 tropism in non-B subtypes, as data on this issue are scarce. Plasma samples were tested using a new phenotypic tropism assay (Phenoscript-tropism; Eurofins), and results were compared with estimates of coreceptor usage using eight different genotypic predictor softwares (Support Vector Machine [SVM], C4.5, C4.5 with positions 8 to 12 only, PART, Charge Rule, geno2pheno coreceptor, Position-Specific Scoring Matrix X4R5 [PSSM(X4R5)], and PSSM(sinsi)). A total of 150 samples were tested, with 115 belonging to patients infected with non-B subtypes and 35 drawn from subtype B-infected patients, which were taken as controls. When non-B subtypes were tested, the concordances between the results obtained using the phenotypic assay and distinct genotypic tools were as follows: 78.8% for SVM, 77.5% for C4.5, 82.5% for C4.5 with positions 8 to 12 only, 82.5% for PART, 82.5% for Charge Rule, 82.5% for PSSM(X4R5), 83.8% for PSSM(sinsi), and 71.3% for geno2pheno. When clade B viruses were tested, the best concordances were seen for PSSM(X4R5) (91.4%), PSSM(sinsi) (88.6%), and geno2pheno (88.6%). The sensitivity for detecting X4 variants was lower for non-B than for B viruses, especially in the case of PSSM(sinsi) (38.4% versus 100%, respectively), SVM(wetcat) (46% versus 100%, respectively), and PART (30% versus 90%, respectively). In summary, while inferences of HIV-1 coreceptor usage using genotypic tools seem to be reliable for clade B viruses, their performances are poor for non-B subtypes, in which they particularly fail to detect X4 variants.

The effectiveness of antiretroviral treatment (ART) was compared in 416 naive patients from a French clinical cohort infected with B and non-B HIV-1 subtypes. Time to HIV viral load (VL) undetectability was calculated for each subtype group. Three other parameters were estimated 3, 6 and 12 months after enrolment: clinical progression (that is, AIDS-defining events or death), changes in CD4 cell counts from baseline and proportion of patients achieving an undetectable VL (<400 HIV-RNA copies/ml). In this cohort, 317 patients (76%) were infected with a B subtype and 99 (24%) with a non-B subtype. Median time to VL undetectability was similar in the B subtype group [147 days, 95% confidence interval (CI) 119-165] and non-B subtype group (168 days, 95% CI: 105-234; P=0.16). After adjusting for AIDS-defining events at enrolment, baseline CD4 cell counts and VL, and for the treatment on which patients were initiated, no association was found between HIV subtypes and time to VL undetectability (B subtype vs non-B subtype: hazard ratio=0.80, 95% CI: 0.62-1.02, P=0.07). In the 3, 6 and 12 months after enrolment, subtype had no impact on clinical progression, CD4 cell count or VL responses to ART. This suggests that B and non-B subtypes do not affect first-line therapy efficacy, which is encouraging in view of the worldwide spread of non-B HIV-1 subtypes and the increasing availability of ART in developing countries. However, in this study we did not take into account individual non-B subtype species, therefore further studies should be designed to evaluate the efficacy of these regimens in patients with particular non-B subtypes.

Background: Human immunodeficiency viruses (HIV) are characterized by extremely high genetic variability. This extensive heterogeneity resulted from high error rate of reverse transcriptase enzyme in combination with fast turnover of virions among HIV-infected individuals. With geographical distribution of subtypes its evolution and unpredictable process and intermixing of HIV-1 variants is inevitable. Recombinant viruses contribute already substantially to the global pandemic especially in sub-Saharan African. East Africa studies has shown more complex mixture of HIV-1 Subtypes and associated recombinant virus especially in Tanzania. Objective: To review past and current literature on HIV-1 diversity and challenges emerging toward effective, safe and cheap HIV-1 Vaccine in Tanzania and gives out some recommendations that could help in tackling these challenges. Methods: A Pub med, HINARI, Springer online archives and Google Scholar literature of publication on HIV-1 Subtypes and Vaccine was performed to identify articles on ‘HIV-1 diversity’, ‘HIV-1 Subtypes’, ‘HIV-1 and vaccine’ , ‘HIV-1 Evolution’, ‘HIV-1 subtypes and vaccine’ and HIV-1 diversity and vaccine in Tanzania. Articles resulting from these searches in the articles were reviewed. Findings: The predominant HIV-1 subtypes across African continent are HIV-1 Subtype C with few cases of Subtypes A in east Africa and recombinant forms in West Africa. In Tanzania the predominant subtype was HIV-1 Subtype A in Northern side compared with HIV-1 subtype C in the western and central part of the country. We also reported few cases of HIV-1 Subtype D and B in the shore of lake Victoria near Uganda border. Conclusion: With regard to HIV infection, we are facing a unique challenge where by a concept of vaccine as a standalone prevention measure to end the epidemic is unlikely. Therefore it is a high time to join global effort to come up with a new paradigm of joining potentially promising ideas toward effective HIV/AIDS vaccine. In area like Tanzania where HIV-1 is very diverse can be taken as an opportunity to harness the possibility of finding broad neutralizing antibodies across all strains for a good candidate of HIV-1 vaccine.

BackgroundMost of the non-B HIV-1 subtypes are predominant in Sub-Saharan Africa and India although they have been found worldwide. In the last decade, immigration from these areas has increased considerably in Spain. The objective of this study was to evaluate the prevalence of non-B subtypes circulating in a cohort of HIV-1-infected immigrants in Seville, Southern Spain and to identify drug resistance-associated mutations.MethodsComplete protease and first 220 codons of the reverse transcriptase coding regions were amplified and sequenced by population sequencing. HIV-1 subtypes were determined using Stanford University Drug Resistance Database, and phylogenetic analysis was performed comparing multiple reported sequences. Drug resistance mutations were defined according to the International AIDS Society-USA.ResultsFrom 2000 to 2010 a total of 1,089 newly diagnosed HIV-1-infected patients were enrolled in our cohort. Of these, 121 were immigrants, of which 98 had ethical approval and informed consent to include in our study. Twenty-nine immigrants (29/98, 29.6%) were infected with non-B subtypes, of which 15/29 (51.7%) were CRF02-AG, mostly from Sub-Saharan Africa, and 2/29 (6.9%) were CRF01-AE from Eastern Europe. A, C, F, J and G subtypes from Eastern Europe, Central-South America and Sub-Saharan Africa were also present. Some others harboured recombinant forms CRF02-AG/CRF01-AE, CRF2-AG/G and F/B, B/C, and K/G, in PR and RT-coding regions. Patients infected with non-B subtypes showed a high frequency of minor protease inhibitor resistance mutations, M36I, L63P, and K20R/I. Only one patient, CRF02_AG, showed major resistance mutation L90M. Major RT inhibitor resistance mutations K70R and A98G were present in one patient with subtype G, L100I in one patient with CRF01_AE, and K103N in another patient with CRF01_AE. Three patients had other mutations such as V118I, E138A and V90I.ConclusionsThe circulation of non-B subtypes has significantly increased in Southern Spain during the last decade, with 29.6% prevalence, in association with demographic changes among immigrants. This could be an issue in the treatment and management of these patients. Resistance mutations have been detected in these patients with a prevalence of 7% among treatment-naïve patients compared with the 21% detected among patients under HAART or during treatment interruption.