non-B HIV-1 Research Articles

Characterizing viral subtypes to assess patterns of HIV transmission

We characterized HIV-1 subtypes among 204 persons newly diagnosed with HIV in Ontario from 2003 to 2005 using samples from the Canadian HIV Strain and Drug Resistance Surveillance Program. We examined HIV-1 subtype by demographic characteristics and exposure category, and determined independent predictors of infection with a non-B HIV subtype using multivariate logistic regression. The distribution of HIV subtypes was: B 77.0%, C 10.3%, AG 4.9%, A 2.5%, AE 2.5% and others 3.0%. Overall, 23.0% were non-B, greater in women than in men (62.8% versus 12.4%, P < 0.0001) and persons under 35 years (31.1% versus 18.5% in those ≥35, P = 0.04). Non-B subtype was predominant (78.9%) among persons from HIV-endemic regions and considerable (28.6%) among other persons infected heterosexually. In multivariate modelling adjusted for gender, non-B subtype was significantly associated with birth in an HIV-endemic region (adjusted odds ratio [aOR] 59.2, P < 0.0001) and heterosexual exposure (aOR 6.3, P = 0.02). Additionally, compared with men who had sex with men, non-B subtype was greater among heterosexual women (aOR 17.8, P < 0.001) and women who injected drugs (injection drug use, aOR 13.4, P = 0.01). We found a non-negligible proportion of non-B subtypes among women infected heterosexually not from HIV-endemic countries, providing interesting insights into HIV transmission patterns.

Phenotyping methods for determining HIV tropism and applications in clinical settings

HIV-1 enters CD4-expressing cells via one or both of the chemokine receptors CCR5 and CXCR4. Specific CCR5 antagonists are now in clinical use, but only for CCR5-tropic viruses. Hence, several methods have been developed for assessing HIV-1 tropism in patients who are candidates for CCR5 antagonists. This article reviews current data on phenotypic assays of tropism. Phenotypic assays are still used as reference, although genotypic methods have improved. The main advantages of phenotypic assays are their great sensitivity for detecting minor CXCR4-using variants and their capacity to assess non-B subtypes of HIV-1. Clinical trials of maraviroc have, thus, relied on the phenotypic determination of HIV-1 tropism. However, new genotypic approaches that are more sensitive for minor CXCR4-using variants, notably ultra-deep pyrosequencing, are now challenging phenotypic assays. Nevertheless, phenotypic assays are essential for improving genotypic algorithms for determining HIV-1 tropism as well as for assessing the resistance of R5-tropic viruses to CCR5 antagonists. HIV-1 tropism should be determined before using CCR5 antagonists. Phenotypic recombinant assays are still the benchmark tests for characterizing HIV-1 tropism as their great sensitivity enables them to detect minor CXCR4-using variants of both B and non-B HIV-1 subtypes.

English

Forty samples collected from HIV-1 AIDS and asymptomatic patients in 2002 and 2003 before the initiation of&nbsp;antiretroviral treatment (ART), most in poorly investigated regions of Ghana were retrospectively examined. For baseline data on genetic diversity before therapy,&nbsp;env,&nbsp;gagand&nbsp;pol&nbsp;phylogenetic analysis was generated.&nbsp;pol&nbsp;was further analysed for potential resistance to antiretroviral drugs. Plasma viral load was also quantified. HIV-1 subtypes A1 and G constituted a minority, while simple and/or complex recombinant strains dominated by CRF02_AG and including CRF06_cpx, CRF09 and CRF10 was observed.&nbsp;Major mutations,protease (PR) L33F and&nbsp;reverse transcriptase (RT) M41L,&nbsp;associated with high levels of phenotypic resistance or clinical evidence for reduced virological response&nbsp;were identified. In the protease-coding domain, positions 10 and 11 were mutated in 21% (5/24) of patients examined, carrying putative resistance to PIs. In the reverse transcriptase-coding domain, seven mutations were detected in 6/35 patients (17%). Of these, 8.5% (3/35) had either double or single mutations possibly affecting NRTIs or NNRTIs respectively.&nbsp;Within PR and RT regions, the mutation rate ranged between 2.6 and 7.0%. Position 10 had 57% mutation frequency.&nbsp;The mutational profile of our non-B HIV-1 strains were identical with subtype B strains at documented DR-related positions. Therefore, drug resistance (DR) pathways of non-B strains were presumed similar to subtype B viruses. Although antiretroviral therapy was expected to be successful, patients presenting resistance-related mutations prior to the initiation of ART could develop clinically relevant resistance earlier than wild-type strains. &nbsp; Key words:&nbsp;HIV-1 genotyping, antiretroviral therapy, drug-naive HIV-1 patients, drug resistance.

Comparative Analysis of Drug Resistance Among B and the Most Prevalent Non-B HIV Type 1 Subtypes (C, F, and CRF02_AG) in Italy

In recent years, increasing numbers of patients infected with HIV-1 non-B subtypes have been treated with modern antiretroviral regimens. Therefore, a better knowledge of HIV drug resistance in non-B strains is crucial. Thus, we compared the mutational pathways involved in drug resistance among the most common non-B subtypes in Italy (F, C, and CRF02_AG) and the B subtype. In total, 2234 pol sequences from 1231 virologically failing patients from Central Italy were analyzed. The prevalence of resistance mutations in protease and reverse transcriptase between non-B and B subtypes has been evaluated. Among patients treated with nucleoside/nucleotide reverse transcriptase inhibitors (NRTI) and with thymidine analogues (TA) experience, TAMs1 M41L and L210W were less prevalent in CRF02_AG, while TAMs2 T215F and K219E were more prevalent in the F subtype. In NRTI-treated patients having experience with abacavir, didanosine, tenofovir, or stavudine the K65R mutation was mostly prevalent in the C subtype. In non-NRTI (NNRTI)-treated patients infected by the C subtype the prevalence of K103N was lower than in patients infected with other subtypes, while the prevalence of Y181C and Y188L was higher compared to subtype B. The prevalence of Y181C was higher also in subtype F as compared to subtype B. In patients treated with protease inhibitors, L89V was predominantly found in CRF02_AG, while the TPV resistance mutation T74P was predominantly found in the C subtype. Some differences in the genotypic drug resistance have been found among patients infected with B, C, F, and CRF02_AG subtypes in relationship to treatment. These results may be useful for the therapeutic management of individuals infected with HIV-1 non-B strains.

Virological failure rates and HIV‐1 drug resistance patterns in patients on first‐line antiretroviral treatment in semirural and rural Gabon

IntroductionAs antiretroviral treatment (ART) continues to expand in resource-limited countries, the emergence of HIV drug resistance mutations (DRMs) is challenging in these settings. In Gabon (central Africa), no study has yet reported the virological effectiveness of initial ART given through routine HIV care.MethodsFollowing the World Health Organization (WHO) recommendations, a cross-sectional study with a one-time HIV-1 RNA viral load (VL) measurement was conducted in Gabon to assess virological failure (VF) defined by a VL result ≥1000 copies/ml and DRMs among adult patients living with non-B HIV-1 strains and receiving first-line non-nucleoside reverse transcriptase inhibitor (NNRTI)-based antiretroviral therapy for at least 12 months. Risk factors associated with VF and DRMs were assessed.ResultsBetween March 2010 and March 2011, a total of 375 patients were consecutively enrolled from two decentralized (one semirural and one rural) HIV care centres. Median time on ART was 33.6 months (range, 12–107). Overall, the rate of VF was 41.3% (36.4–46.4). Among viremic patients, 56.7% (80/141) had at least one DRM and 37.6% had dual-class resistance to nucleoside reverse transcriptase inhibitors (NRTIs) and NNRTIs. The most frequent DRMs were K103N/S (46.1%) and M184V/I (37.6%). Thymidine analogue mutations were found in 10.6%. Independent risk factors associated with VF were being followed up at the semirural centre (P=0.033), having experienced unstructured treatment interruptions (P=0.0044), and having low CD4+ counts at enrolment (P<0.0001). A longer time on ART (P=0.0008) and being followed up at the rural centre (P=0.021) were risk factors for DRMs.ConclusionsThis is the first study conducted in Gabon providing VF rates and DRM patterns in adult patients receiving first-line ART. In sub-Saharan Africa, where NNRTI-based regimens are recommended as the standard for first-line ART, strengthening virological monitoring together with preventing unplanned treatment interruptions are a global public health priority.

Does Genetic Diversity of HIV-1 Non-B Subtypes Differentially Impact Disease Progression in Treatment-Naive HIV-1–Infected Individuals? A Systematic Review of Evidence

With 88% of HIV-1-infected individuals living in areas of high prevalence of non-B subtypes and with expanded global access to antiretroviral treatment (ART), studying disease progression amongst non-B subtypes gains relevance. Optimized clinical management is a possibility with knowledge of non-B subtype profiles at baseline, which is currently not possible due to lack of subtype-specific point-of-care assays. In a systematic review, we synthesized global evidence on differential disease progression amongst non-B subtypes in ART-naive individuals. Due to lack of consistent effect measures, we avoided pooling data and inferred patterns with respect to disease progression outcomes (ie, AIDS, Death, CD4, viral load changes). Subtypes C and D were more aggressive, followed by G, AE, and AG, and A being the least aggressive of all HIV-1 subtypes. Evidence of greater rates of disease progression in globally prevalent C and D subtypes highlight the importance of expanding early HIV detection, and determining subtype profile at baseline with CD4 staging to optimize the quality of ART delivery and care in global settings.

Poorer Immunologic Outcome on Treatment Among Patients Infected With HIV-1 Non-B Subtypes Compared With Subtype B in Singapore

Poorer Immunologic Outcome on Treatment Among Patients Infected With HIV-1 Non-B Subtypes Compared With Subtype B in Singapore

Molecular Characterization of Non-B HIV Type 1 Isolates from Patients of a Department of Infectious Diseases, University Hospital of Bordeaux, France, 1989–2009

The molecular characterization of non-B HIV type 1 subtypes and the sociodemographic baseline characteristics have been studied for 114 non-B HIV-1-infected patients followed at the University Hospital of Bordeaux, France, and diagnosed as HIV infected between 1989 and 2009. Individuals enrolled in this study were mainly women with heterosexual transmission in West and Central Africa and who have been discovered to be HIV positive during pregnancy. Nevertheless, HIV acquisition among individuals born in France was significantly increasing. Recombinant form CRF02_AG was the most frequent subtype (38%) among a highly diverse viral background since 19 subtypes and CRFs have been characterized with a maximal diversity observed in the past decade.

Effect of Natural Polymorphisms in the HIV-1 CRF02_AG Protease on Protease Inhibitor Hypersusceptibility

Hypersusceptibility (HS) to inhibition by different antiretroviral drugs (ARVs) among diverse HIV-infected individuals may be a misnomer because clinical response to treatment is evaluated in relation to subtype B infections while drug susceptibility of the infecting virus, regardless of subtype, is compared to a subtype B HIV-1 laboratory strain (NL4-3 or IIIB). Mounting evidence suggests that HS to different ARVs may result in better treatment outcome just as drug resistance leads to treatment failure. We have identified key amino acid polymorphisms in the protease coding region of a non-B HIV-1 subtype linked to protease inhibitor HS, namely, 17E and 64M in CRF02_AG. These HS-linked polymorphisms were introduced in the BD6-15 CRF02_AG molecular clone and tested for inhibition using a panel of protease inhibitors. In general, suspected HS-linked polymorphisms did increase susceptibility to specific protease inhibitors such as amprenavir and atazanavir, but the combination of the 17E/64M polymorphisms showed greater HS. These two mutations were found at low frequencies but linked in a sequence database of over 700 protease sequences of CRF02_AG. In direct head-to-head virus competitions, CRF02_AG harboring the 17E/64M polymorphisms also had higher replicative fitness than did the 17E or the 64M polymorphism in the CFR02_AG clone. These findings suggest that subtype-specific, linked polymorphisms can result in hypersusceptibility to ARVs. Considering the potential benefit of HS to treatment outcome, screening for potential HS-linked polymorphisms as well as preexisting drug resistance mutations in treatment-naïve patients may guide the choice of ARVs for the best treatment outcome.

Development and evaluation of an assay for HIV-1 protease and reverse transcriptase drug resistance genotyping of all major group-M subtypes

BackgroundHigh cost and varying sensitivity for non-B HIV-1 subtypes limits application of current commercial kits for HIV-1 drug resistance genotyping of all major HIV-1 group-M subtypes. ObjectivesOur research aimed to develop and validate an assay specific for all major HIV-1 group-M subtypes for use as an alternative to commercial assays for HIV-1 protease (PR) and reverse transcriptase (RT) drug resistance genotyping. Study designA nested RT-PCR encompassing the entire PR and RT up to amino acid 321 of HIV-1 was designed to detect HIV-1 group-M subtypes. Primers compatible with group-M subtypes were defined and analytical sensitivity of the assay evaluated using a panel of reference viruses for subtypes A–H and CRF01_AE. The assay was subsequently evaluated on 246 plasma samples from HIV-1 infected individuals harboring various group-M subtypes and viral loads (VLs). ResultsAll major group-M HIV-1 subtypes were detected with an overall analytical sensitivity of 1.00E+03 RNA copies/ml. Application of the genotyping assay on 246 primarily African clinical samples comprising subtypes A (n=52; 21.7%), B (n=12; 5.0%), C (n=127; 52.9%), D (n=25; 10.4%), CRF01_AE (n=10; 4.2%), and CRF02_AG (n=10; 4.2%), and unassigned variants (n=10; 4.2%), VL range 4.32E+02–8.63E+06 (median 2.66E+04) RNA copies/ml, was ∼98% successful. ConclusionsA group-M subtype-independent genotyping assay for detection of HIV-1 drug resistance was developed. The described assay can serve as an alternative to commercial assays for HIV-1 drug resistance genotyping in routine diagnostics, and for surveillance and monitoring of drug resistance in resource-limited settings (RLS).

Evaluation of a manual DNA extraction protocol and an isothermal amplification assay for detecting HIV-1 DNA from dried blood spots for use in resource-limited settings

BackgroundIn resource-limited settings (RLS) dried blood spots (DBS) are collected on infants and transported through provincial laboratories to a central facility where HIV-1 DNA PCR testing is performed using specialized equipment. Implementing a simpler approach not requiring such equipment or skilled personnel could allow the more numerous provincial laboratories to offer testing, improving turn-around-time to identify and treat infected infants sooner. ObjectivesAssess performances of a manual DNA extraction method and helicase-dependent amplification (HDA) assay for detecting HIV-1 DNA from DBS. Study design60 HIV-1 infected adults were enrolled, blood samples taken and DBS made. DBS extracts were assessed for DNA concentration and beta globin amplification using PCR and melt-curve analysis. These same extracts were then tested for HIV-1 DNA using HDA and compared to results generated by PCR and pyrosequencing. Finally, HDA limit of detection (LOD) studies were performed using DBS extracts prepared with known numbers of 8E5 cells. ResultsThe manual extraction protocol consistently yielded high concentrations of amplifiable DNA from DBS. LOD assessment demonstrated HDA detected ∼470copies/ml of HIV-1 DNA extracts in 4/4 replicates. No statistical difference was found using the McNemar's test when comparing HDA to PCR for detecting HIV-1 DNA from DBS. ConclusionsUsing just a magnet, heat block and pipettes, the manual extraction protocol and HDA assay detected HIV-1 DNA from DBS at levels that would be useful for early infant diagnosis. Next steps will include assessing HDA for non-B HIV-1 subtypes recognition and comparison to Roche HIV-1 DNA v1.5 PCR assay.

Drug Resistance Pathways and Impact of Protease Mutation L10I/V in HIV-1 Non-B Subtypes

Background: Molecular pathways to drug resistance have yet to be fully characterized in HIV-1 non-B subtypes. Furthermore, polymorphisms such as protease L10I/V are ubiquitous in non-B subtypes, but their biological implications are still unknown. We evaluated resistance pathways emerging at treatment failure in a cohort of HIV infected individuals in Mali, and characterized in vitro the role of L10I/V. Methods: Genotypic resistance testing was performed on plasma obtained from 132 HIV-infected individuals from Mali before and after 9 months of treatment using population sequencing. CRF02_AG chimeric viruses containing 10I/V mutants CRF02_AG were constructed using site directed mutagenesis and susceptibility to protease inhibitors (PI) as well as replicative capacity were determined in a PBMC culture assay. Results: At treatment initiation, 11/132 (8.3%; 95% CI 3.6-13.0%) patients harboured resistance mutations to NRTI (D67N, T69N, L210W, K219E and T215A) or NNRTI (K103N, V108I and V179E). Among these 11 patients, 5 with NNRTI mutations were in virological failure after 9 months of treatment. Six others with one Thymidine Analog Mutations (TAM) did not show complete resistance. Overall, 18/132 (14.0%; 95% CI 8.1-19.9%) patients failed at 9 months and resistance mutations to NRTI or NNRTI could be identified in 8 (6.10%; 95% CI 2.0-10.2%). NRTI mutation M184V was the most commonly observed, followed by NNRTI mutations Y181C and K103N. Polymorphisms in protease such as L10I/V were observed frequently. Their role was evaluated in vitro. CRF02_AGwt_10L showed a slight increase in IC50 for darunavir, lopinavir and nelfinavir compared to subtype BHXB2_10L with 1.2, 1.3 and 1.5 Fold-Changes (FC) respectively. Mutant’s viruses CRF02_AGL10I and CRF02_AGL10V showed a slight increase in IC50 for indinavir with 1.30 and 1.20 FC and a slight decrease in IC50 for lopinavir with 0.78 FC and 0.75 FC respectively compared to CRF02_AGwt_10L. We did not observe any difference in replicative capacity between CRF02_AGwt_10L and HXB2. However, compared to CRF02_AGwt_10L, mutants, viruses CRF02_AGL10I, and CRF02_AGL10V showed a significant reduction in replication capacity by 10% (p<0.03) and 12% (p<0.02) respectively. Conclusion: Primary resistance to NRTI and NNRTI impacts response to treatment. The presence of a single TAM mutation may have limited impact on first line treatment in CRF02_AG. A common polymorphism in non-B subtypes, L10V, may affect susceptibility of certain PIs. In the context of large-scale use of antiretroviral, monitoring the emergence of resistance in non-B subtypes is important to preserve treatment options.

Potency of Stampidine Against Multi-Nucleoside Reverse Transcriptase Inhibitor Resistant Human Immunodeficiency Viruses

The in vitro potency of stampidine (STAMP, DDE-113, HI-113, N-[p-(4-bromophenyl)-2',3'-didehydro-3'-deoxy-5'-thymidylyl]-L-alanine methyl ester, CAS 217178-62-6) was examined against 8 clinical non-B subtype HIV-1 isolates with resistance to stavudine (STV, d4T), adefovir and tenofovir, 19 clinical zidovudine-resistant HIV-1 isolates, and 6 recombinant HIV-1 clones with multi-resistance against nucleoside reverse transcriptase inhibitors. Stampidine exhibited potent anti-HIV activity against each one of these 33 HIV-1 isolates with subnanomolar to nanomolar IC50 values.

HIV-1 clade a infection and viral control: an immunological perspective on a case of underquantification

Although a vast majority of HIV-1-positive patients in the UK are infected with clade B virus, a large number of newly diagnosed cases of heterosexually transmitted HIV-1 are acquired abroad, in countries where non-B clade HIV-1 predominates. Since the development of the viral load assay in 1988, assessment of HIV-1 plasma viraemia has become an integral part of HIV clinical care; however, the contemporary viral load assay was developed and optimized for clade B HIV-1. Here we report the underquantification of viraemia in an individual infected with clade A virus, and the consequent initial classification of the patient as an HIV controller (HIC). Immunological investigations of interferon (IFN)-γ and lymphoproliferative responses to HIV-1 clade B antigens and peptides, in parallel with mitogenic stimulation, were performed. Subsequent comparison with responses observed within clade B-infected HIC led to viral sequencing, confirmation of infecting clade and recommendation of antiretroviral therapy initiation. We emphasize the growing need for awareness of possible limitations of the commonly used viral load assays, which cannot be relied upon unreservedly in a clinical setting. Furthermore, this case highlights the increasing need for more detailed investigation into both viral genetics and fitness when defining patients as HIC.

Evaluation of genotypic tropism prediction tests compared with in vitro co-receptor usage in HIV-1 primary isolates of diverse subtypes

To evaluate the sensitivity and specificity of genotypic methods for predicting the co-receptor usage of subtypes B and non-B HIV-1 primary isolates, using as gold standard the infectivity of each primary isolate in GHOST cells stably expressing HIV-1 co-receptors. Primary isolates were obtained by co-culturing either patient's peripheral blood mononuclear cells (PBMCs) or ultracentrifuged plasma with donor-activated PBMCs. In vitro co-receptor usage was determined by infecting GHOST cells. Tropism prediction, based on V3 sequences, was determined with simple rules and bioinformatic tools (Geno2pheno[coreceptor] and WebPSSM). This study includes 102 HIV-1 primary isolates; 23 (22.5%) subtype B and 79 (77.5%) non-B genetic forms. V3 sequences were classified into six subtypes (A-G), although 32 (31.4%) were circulating recombinant forms and 21 (20.6%) were unique recombinant forms. Sixty-nine isolates were R5, 27 R5X4 and 6 X4. The highest levels of sensitivity and specificity for the detection of X4 strains among V3 sequences, between 91% and 100%, were obtained by using PSSM(x4r5), PSSM(si/nsi) and the 11/25 rule for sequences of subtypes A, B and G, but not for subtype F. Establishing the recommended cut-off for clinical settings of a 10% false positive rate for Geno2pheno, we obtained 93% specificity and 97% sensitivity. Comparing genotypic assays for HIV-1 co-receptor use with a cell-culture phenotypic assay could provide more reliable results of sensitivity and specificity for the detection of X4 strains than comparing them with recombinant assays, considered as gold standard. In general, except for subtype F isolates, there is a good correlation for tropism prediction.

Polymorphism in Gag Gene Cleavage Sites of HIV-1 Non-B Subtype and Virological Outcome of a First-Line Lopinavir/Ritonavir Single Drug Regimen

Virological failure on a boosted-protease inhibitor (PI/r) first-line triple combination is usually not associated with the detection of resistance mutations in the protease gene. Thus, other resistance pathways are being investigated. First-line PI/r monotherapy is the best model to investigate in vivo if the presence of mutations in the cleavage sites (CS) of gag gene prior to any antiretroviral treatment might influence PI/r efficacy. 83 patients were assigned to initiate antiretroviral treatment with first-line lopinavir/r monotherapy in the randomised Monark trial. We compared baseline sequence of gag CS between patients harbouring B or non-B HIV-1 subtype, and between those who achieved viral suppression and those who experienced virological failure while on LPV/r monotherapy up to Week 96. Baseline sequence of gag CS was available for 82/83 isolates; 81/82 carried at least one substitution in gag CS compared to HXB2 sequence. At baseline, non-B subtype isolates were significantly more likely to harbour mutations in gag CS than B subtype isolates (p<0.0001). Twenty-three patients experienced virological failure while on lopinavir/r monotherapy. The presence of more than two substitutions in p2/NC site at baseline significantly predicted virological failure (p = 0.0479), non-B subtype isolates being more likely to harbour more than two substitutions in this specific site. In conclusion, gag cleavage site was highly polymorphic in antiretroviral-naive patients harbouring a non-B HIV-1 strain. We show that pre-therapy mutations in gag cleavage site sequence were significantly associated with the virological outcome of a first-line LPV/r single drug regimen in the Monark trial.

Prevalence and resistance mutations of non-B HIV-1 subtypes among immigrants in Southern Spain along the decade 2000-2010

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.

Molecular Surveillance of HIV-1 in Madrid, Spain: a Phylogeographic Analysis

The molecular epidemiology of HIV-1 is constantly changing, mainly as a result of human migratory flows and the high adaptive ability of the virus. In recent years, Spain has become one of Europe's main destinations for immigrants and one of the western European countries with the highest rates of HIV-positive patients. Using a phylogeographic approach, we have analyzed the relationship between HIV-1 variants detected in immigrant and native populations of the urban area of Madrid. Our project was based on two coincidental facts. First, resistance tests were extended to naïve and newly diagnosed patients, and second, the Spanish government legislated the provision of legal status to many immigrants. This allowed us to obtain a large data set (n = 2,792) from 11 Madrid hospitals of viral pol sequences from the two populations, and with this unique material, we explored the impact of immigration in the epidemiological trends of HIV-1 variants circulating in the largest Spanish city. The prevalence of infections by non-B HIV-1 variants in the studied cohort was 9%, rising to 25% among native Spanish patients. Multiple transmission events involving different lineages and subsubtypes were observed in all the subtypes and recombinant forms studied. Our results also revealed strong social clustering among the most recent immigrant groups, such as Russians and Romanians, but not in those groups who have lived in Madrid for many years. Additionally, we document for the first time the presence of CRF47_BF and CRF38_BF in Europe, and a new BG recombinant form found in Spaniards and Africans is tentatively proposed. These results suggest that the HIV-1 epidemic will evolve toward a more complex epidemiological landscape.

Clinical efficacy of raltegravir against B and non-B subtype HIV-1 in phase III clinical studies

We evaluated the long-term efficacy of raltegravir according to HIV-1 subtype (B and non-B) using data from three phase III studies in treatment-experienced (BENCHMRK-1 and 2) and treatment-naive (STARTMRK) HIV-infected patients. HIV-1 subtypes were identified from baseline plasma specimens using genotypic data of the PhenoSense GT test (Monogram Biosciences, South San Francisco, California, USA). Non-B subtypes were combined for the current analyses due to small numbers of each specific subtype. An observed failure approach was used (only discontinuations due to lack of efficacy were treated as failures). Resistance evaluation was performed in patients with documented virologic failure. Seven hundred and forty-three patients received raltegravir and 519 received comparator (efavirenz in STARTMRK; optimized background therapy in BENCHMRK). Non-B subtype virus (A, A/C, A/D, A/G, A1, AE, AG, B/G, BF, C, D, D/F, F, F1, G, and complex) was isolated at baseline in 98 (13%) raltegravir recipients and 62 (12%) comparator recipients. Subtypes AE and C were most common, isolated in 41 and 43 patients, respectively. The proportion of raltegravir recipients achieving HIV RNA less than 50 copies/ml was similar between non-B and B subtypes (STARTMRK: 94.5 vs. 88.7%; BENCHMRK-1 and 2: 66.7 vs. 60.7%); change in CD4 cell count also was similar between non-B and B subtypes (STARTMRK: 243 vs. 221 cells/μl; BENCHMRK-1 and 2: 121 vs. 144 cells/μl). Phenotypic resistance to raltegravir in non-B virus was associated with integrase mutations observed previously in subtype B virus. In phase III studies in treatment-naive and treatment-experienced patients, raltegravir showed comparable and potent clinical efficacy against B and non-B HIV-1 subtypes.

Men Who Have Sex With Men, Risk Behavior, and HIV Infection: Integrative Analysis of Clinical, Epidemiological, and Laboratory Databases

Centralized data collection and analytic tools facilitate tracing HIV transmission trends at the patient-population level with increasing resolution, complementing behavioral studies while avoiding sampling biases. By several measures, the rate of HIV infection among men who have sex with men (MSM) in Israel increased in the past several years more rapidly than was expected. We describe features of the data that connect this increase to behavioral changes. We retrospectively analyzed data from the national HIV reference laboratory and the national HIV and sexually transmitted infections registries. We examined changes in selected epidemiologic and clinical parameters and in the pattern of drug-resistant virus transmission among MSM in Israel. In particular, virus isolates from 296 MSM (23.8% of all MSM who received a diagnosis) were genotyped, drug-resistance conferring mutations were characterized, and phylogenetic trees were constructed. Compared with earlier years, during 2007-2009 MSM were more often infected with drug-resistant virus before treatment initiation, were coinfected with syphilis, and received a diagnosis during acute retroviral syndrome. Phylogenetic analysis suggested frequent transmission of drug-resistant HIV by drug-treated individuals to >1 partner. Secondary transmission of resistant virus by drug-naive patients is also consistent with the phylogenetic patterns. In addition, non-B HIV subtypes began to appear among MSM. Together, our findings suggest that the sexual behavior of MSM, both HIV-infected and uninfected, has become riskier, contributing to the number of those seeking early clarification of status, to syphilis comorbidity, and to the spread of drug resistance. These findings call for action by public health planners and community-based organizations aimed at increasing awareness of the risks, bringing a change in attitude and establishing safe sex norms.