Predominance of CXCR4 tropism in HIV-1 CRF14_BG strains from newly diagnosed infections.

OBJECTIVES: R5-tropic viruses are associated with HIV-1 transmission and predominate during the early stages of infection. X4-tropic populations have been detected in ~50% of patients with late-stage disease infected with subtype B viruses. In this study, we compared the frequency of X4 tropism in individuals infected with HIV-1 CRF14_BG viruses, which have a V3 loop of subtype B, with a control group of individuals infected with subtype B viruses. METHODS: Sixty-three individuals infected with HIV-1 CRF14_BG (n = 31) or subtype B (n = 32) were studied. Similar proportions of newly diagnosed and chronically infected individuals were included in the subtype B and CRF14_BG groups. V3 sequences were obtained and coreceptor tropism was predicted using the Geno2pheno[coreceptor] algorithm. V3 net charge and 11/25 rules were also used for coreceptor prediction. RESULTS: Overall, X4 tropism was more frequent among individuals infected with CRF14_BG viruses (87.1%) than subtype B viruses (34.3%), a difference that was statistically highly significant (P = 0.00001). Importantly, the frequencies among newly diagnosed individuals were 90% and 13.3%, respectively (P = 0.0007). Characteristic amino acids in the V3 loop (T13, M14, V19 and W20) were identified at higher frequencies in CRF14_BG viruses (54%) than subtype B viruses (0%; P < 0.000001). CONCLUSIONS: CRF14_BG is the genetic form with the highest proportion of X4-tropic viruses reported to date in newly diagnosed and chronic infections. This suggests high pathogenicity for CRF14_BG viruses, potentially leading to rapid disease progression. CCR5 antagonists will be ineffective in most CRF14_BG-infected patients, even at early stages of infection.

Improvement of HIV-1 coreceptor tropism prediction by employing selected nucleotide positions of the env gene in a Bayesian network classifier.

OBJECTIVES: This study aimed to develop a genotypic method to predict HIV-1 coreceptor usage by employing the nucleotide sequence of the env gene in a tree-augmented naive Bayes (TAN) classifier, and to evaluate its accuracy in prediction compared with other available tools. METHODS: A wrapper data-mining strategy interleaved with a TAN algorithm was employed to evaluate the predictor value of every single-nucleotide position throughout the HIV-1 env gene. Based on these results, different nucleotide positions were selected to develop a TAN classifier, which was employed to predict the coreceptor tropism of all the full-length env gene sequences with information on coreceptor tropism currently available at the Los Alamos HIV Sequence Database. RESULTS: Employing 26 nucleotide positions in the TAN classifier, an accuracy of 95.6%, a specificity (identification of CCR5-tropic viruses) of 99.4% and a sensitivity (identification of CXCR4/dual-tropic viruses) of 80.5% were achieved for the in silico cross-validation. Compared with the phenotypic determination of coreceptor usage, the TAN algorithm achieved more accurate predictions than WebPSSM and Geno2pheno [coreceptor] (P<0.05). CONCLUSIONS: The use of the methodology presented in this work constitutes a robust strategy to identify genetic patterns throughout the HIV-1 env gene differently present in CCR5-tropic and CXCR4/dual-tropic viruses. Moreover, the TAN classifier can be used as a genotypic tool to predict the coreceptor usage of HIV-1 isolates reaching more accurate predictions than with other widely used genotypic tools. The use of this algorithm could improve the correct prescribing of CCR5 antagonist drugs to HIV-1-infected patients.

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

OBJECTIVES: 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. METHODS: 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). RESULTS: 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. CONCLUSIONS: 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.

Identification of CRF89_BF, a new member of an HIV-1 circulating BF intersubtype recombinant form family widely spread in South America.

Circulating recombinant forms (CRFs) contribute substantially to the HIV-1 pandemic. Among 105 CRFs described in the literature, 16 are BF intersubtype recombinants, most of South American origin, of which CRF12_BF is the most widely spread. A BF recombinant cluster identified in Bolivia was suggested to represent a new CRF_BF. Here we find that it belongs to a larger cluster incorporating 39 viruses collected in 7 countries from 3 continents, 22 of them in Spain, most from Bolivian or Peruvian individuals, and 12 in South America (Bolivia, Argentina, and Peru). This BF cluster comprises three major subclusters, two associated with Bolivian and one with Peruvian individuals. Near full-length genome sequence analyses of nine viruses, collected in Spain, Bolivia, and Peru, revealed coincident BF mosaic structures, with 13 breakpoints, 6 and 7 of which coincided with CRF12_BF and CRF17_BF, respectively. In a phylogenetic tree, they grouped in a clade closely related to these CRFs, and more distantly to CRF38_BF and CRF44_BF, all circulating in South America. These results allowed to identify a new HIV-1 CRF, designated CRF89_BF. Through phylodynamic analyses, CRF89_BF emergence was estimated in Bolivia around 1986. CRF89_BF is the fifth CRF member of the HIV-1 recombinant family related to CRF12_BF.

HIV-1 reverse transcriptase thumb subdomain polymorphisms associated with virological failure to nucleoside drug combinations.

OBJECTIVES: The aim of this study was to identify mutations within the fingers, palm and thumb subdomains of the HIV-1 reverse transcriptase (RT) associated with therapy failure to various combinations of two nucleoside analogues. METHODS: RT nucleotide sequences of 1893 HIV-1 isolates from 1680 persons with different treatment histories (including naive and treated patients) were analysed. chi(2) contingency tests were performed to detect mutations within positions 1-333 of HIV-1 RT, associated with therapy failure, and correlated mutations were identified using statistical methods. RESULTS: Thymidine analogue resistance mutations were strongly associated with therapy failure to all nucleoside analogue combinations analysed. Previously identified accessory mutations at positions 35, 39, 43, 90, 98, 101, 122, 178, 196, 203, 208, 221, 223 and 228 were associated with therapy failure with at least one of the drug combinations studied. Interestingly, several mutations affecting RT thumb subdomain polymorphisms were strongly associated (P < 10(-3)) with therapy failure to abacavir/stavudine, stavudine/didanosine and abacavir/stavudine/didanosine. Mutations A272P, K277R and V293I were more prevalent in patients failing treatment with abacavir/stavudine than in naive individuals, while mutation frequencies of T286A, V292I and to a lesser extent E291D and E297K decreased in the treated population. These effects were also detected for A272P, T286A and V292I, when analysing didanosine/stavudine therapy failure, although statistical significance was higher for T286A. CONCLUSIONS: RT thumb subdomain polymorphisms are strongly associated with therapy failure to nucleoside analogues. Based on their structural location, we suggest a role for those residues in controlling the balance between RNase H degradation and nucleotide excision during DNA polymerization.

Diverse Large HIV-1 Non-subtype B Clusters Are Spreading Among Men Who Have Sex With Men in Spain.

In Western Europe, the HIV-1 epidemic among men who have sex with men (MSM) is dominated by subtype B. However, recently, other genetic forms have been reported to circulate in this population, as evidenced by their grouping in clusters predominantly comprising European individuals. Here we describe four large HIV-1 non-subtype B clusters spreading among MSM in Spain. Samples were collected in 9 regions. A pol fragment was amplified from plasma RNA or blood-extracted DNA. Phylogenetic analyses were performed via maximum likelihood, including database sequences of the same genetic forms as the identified clusters. Times and locations of the most recent common ancestors (MRCA) of clusters were estimated with a Bayesian method. Five large non-subtype B clusters associated with MSM were identified. The largest one, of F1 subtype, was reported previously. The other four were of CRF02_AG (CRF02_1; n = 115) and subtypes A1 (A1_1; n = 66), F1 (F1_3; n = 36), and C (C_7; n = 17). Most individuals belonging to them had been diagnosed of HIV-1 infection in the last 10 years. Each cluster comprised viruses from 3 to 8 Spanish regions and also comprised or was related to viruses from other countries: CRF02_1 comprised a Japanese subcluster and viruses from 8 other countries from Western Europe, Asia, and South America; A1_1 comprised viruses from Portugal, United Kingom, and United States, and was related to the A1 strain circulating in Greece, Albania and Cyprus; F1_3 was related to viruses from Romania; and C_7 comprised viruses from Portugal and was related to a virus from Mozambique. A subcluster within CRF02_1 was associated with heterosexual transmission. Near full-length genomes of each cluster were of uniform genetic form. Times of MRCAs of CRF02_1, A1_1, F1_3, and C_7 were estimated around 1986, 1989, 2013, and 1983, respectively. MRCA locations for CRF02_1 and A1_1 were uncertain (however initial expansions in Spain in Madrid and Vigo, respectively, were estimated) and were most probable in Bilbao, Spain, for F1_3 and Portugal for C_7. These results show that the HIV-1 epidemic among MSM in Spain is becoming increasingly diverse through the expansion of diverse non-subtype B clusters, comprising or related to viruses circulating in other countries.

HIV type 1 intersubtype recombinants during the evolution of a dual infection with subtypes B and G.

ABSTRACT The aim of this study was to characterize the HIV-1 intersubtype recombinant forms generated during the follow-up of a dual natural infection with subtypes B and G. Near full-length sequences from plasma and peripheral blood mononuclear cell (PBMC) compartments were analyzed and the biological characteristics of their derived primary isolates studied. Different mutations were detected in V1, V2, and V3 sequences from primary isolates but not in sequences from plasma RNA or PBMC DNA. The HIV-1 near full-length sequence from the first collected plasma was of subtype G and the presence of subpopulations of subtypes B and G was observed with subtype-specific primers for protease and reverse transcriptase segments. Subsequent sequences from plasma, PBMCs, and primary isolates were obtained during a follow-up of 6 years; all of them were BG recombinants and showed identical intersubtype breakpoints between subtypes B and G in pol and nef. The env sequence from all primary isolates harbored a unique insert of subtype B. Specific primers for the V3 loop identified fluctuating subtype B and/or subtype G sequences either from plasma RNA or PBMC DNA.

Genome-scale analysis of evolutionary rate and selection in a fast-expanding Spanish cluster of HIV-1 subtype F1.

This work is aimed at assessing the presence of positive selection and/or shifts of the evolutionary rate in a fast-expanding HIV-1 subtype F1 transmission cluster affecting men who have sex with men in Spain. We applied Bayesian coalescent phylogenetics and selection analyses to 23 full-coding region sequences from patients belonging to that cluster, along with other 19 F1 epidemiologically-unrelated sequences. A shift in the overall evolutionary rate of the virus, explained by positively selected sites in the cluster, was detected. We also found one substitution in Nef (H89F) that was specific to the cluster and experienced positive selection. These results suggest that fast transmission could have been facilitated by some inherent genetic properties of this HIV-1 variant.

HIV-1 Genetic Diversity in Recently Diagnosed Infections in Moscow: Predominance of AFSU, Frequent Branching in Clusters, and Circulation of the Iberian Subtype G Variant.

HIV-1 protease-reverse transcriptase sequences from 62 HIV-1-infected individuals recently diagnosed in Moscow were analyzed. Subtype A former Soviet Union (FSU) (AFSU) variant was the predominant clade (62.9%), followed by subtype B (22.6%), unique recombinants (6.5%), subtype G (6.5%), and CRF01_AE (1.6%). AFSU predominated among people who inject drugs (88.9%) and heterosexually acquired infections (77.8%), while subtype B was the most prevalent genetic form among men who have sex with men (44%), although AFSU was also frequent in this population (36%). Forty-eight (77.4%) viruses branched within intrasubtype clusters, three of which, of subtype B, had a majority of viruses collected outside of FSU. The four subtype G viruses identified in this study belonged to the Portuguese-Spanish (Iberian) variant and, together with three from databases, formed a Russian cluster closely related to viruses from Denmark. This is the first report of the circulation of the Iberian subtype G variant in Russia.

New insights into the origin of the HIV type 1 subtype A epidemic in former Soviet Union's countries derived from sequence analyses of preepidemically transmitted viruses.

The HIV-1 subtype A epidemic affecting injecting drug users (IDU) in former Soviet Union (FSU) countries started dramatically in Odessa, southern Ukraine, in 1995, and is caused by a variant of monophyletic origin, often designated IDU-A. We phylogenetically analyzed one near full-length genome and two partial sequences of three HIV-1 subtype A viruses collected in St. Petersburg, Russia, heterosexually transmitted in 1992-1994. The sequences branched basally to the IDU-A clade, together with eight viruses from Odessa collected in 1993, all presumably acquired heterosexually, and two viruses from the Democratic Republic of Congo. Of all other FSU sequences in databases, only those from three recently collected viruses, one from Ukraine and two from northwestern Russia, at least one of them acquired heterosexually, branched basally to the IDU-A cluster. The results indicate that the FSU IDU-A variant derives from a strain that initially propagated heterosexually in Ukraine and originated in central Africa.

Genetic Diversity of HIV-1 in Tunisia.

In this study, the genetic diversity of HIV-1 in Tunisia was analyzed. For this, 193 samples were collected in different regions of Tunisia between 2012 and 2015. A protease and reverse transcriptase fragment were amplified and sequenced. Phylogenetic analyses were performed through maximum likelihood and recombination was analyzed by bootscanning. Six HIV-1 subtypes (B, A1, G, D, C, and F2), 5 circulating recombinant forms (CRF02_AG, CRF25_cpx, CRF43_02G, CRF06_cpx, and CRF19_cpx), and 11 unique recombinant forms were identified. Subtype B (46.4%) and CRF02_AG (39.4%) were the predominant genetic forms. A group of 44 CRF02_AG sequences formed a distinct Tunisian cluster, which also included four viruses from western Europe. Nine viruses were closely related to isolates collected in other African or in European countries. In conclusion, a high HIV-1 genetic diversity is observed in Tunisia and the local spread of CRF02_AG is first documented in this country.

Transmission dynamics of HIV-1 subtype B in the Basque Country, Spain.

This work was aimed to study the HIV-1 subtype B epidemics in the Basque Country, Spain. 1727 HIV-1 subtype B sequences comprising protease and reverse transcriptase (PR/RT) coding regions, sampled between 2001 and 2008, were analyzed. 156 transmission clusters were detected by means of phylogenetic analyses. Most of them comprised less than 4 individuals and, in total, they included 441 patients. Six clusters comprised 10 or more patients and were further analyzed in order to study their origin and diversification. Four clusters included men who had unprotected homosexual sex (MSM), one group was formed by intravenous drug users (IDUs), and another included both IDUs and people infected through unprotected heterosexual sex (HTs). Most of these clusters originated from the mid-1980s to the mid-1990s. Only one cluster, formed by MSM, originated after 2000. The time between infections was significantly lower in MSM groups than in those containing IDUs (P-value <0.0001). Nucleoside RT and non-nucleoside RT inhibitor (NRTI and NNRTI)-resistance mutations to antiretroviral treatment were found in these six clusters except the most recent MSM group, but only the IDU clusters presented protease inhibitor (PI)-resistance mutations. The most prevalent mutations for each inhibitor class were PI L90M, NRTI T215D/Y/F, and NNRTI K103N, which were also among the most prevalent resistant variants in the whole dataset. In conclusion, while most infections occur as isolated introductions into the population, the number of infections found to be epidemiologically related within the Basque Country is significant. Public health control measures should be reinforced to prevent the further expansion of transmission clusters and resistant mutations occurring within them.

Identification of an HIV-1 BG Intersubtype Recombinant Form (CRF73_BG), Partially Related to CRF14_BG, Which Is Circulating in Portugal and Spain.

HIV-1 exhibits a characteristically high genetic diversity, with the M group, responsible for the pandemic, being classified into nine subtypes, 72 circulating recombinant forms (CRFs) and numerous unique recombinant forms (URFs). Here we characterize the near full-length genome sequence of an HIV-1 BG intersubtype recombinant virus (X3208) collected in Galicia (Northwest Spain) which exhibits a mosaic structure coincident with that of a previously characterized BG recombinant virus (9601_01), collected in Germany and epidemiologically linked to Portugal, and different from currently defined CRFs. Similar recombination patterns were found in partial genome sequences from three other BG recombinant viruses, one newly derived, from a virus collected in Spain, and two retrieved from databases, collected in France and Portugal, respectively. Breakpoint coincidence and clustering in phylogenetic trees of these epidemiologically-unlinked viruses allow to define a new HIV-1 CRF (CRF73_BG). CRF73_BG shares one breakpoint in the envelope with CRF14_BG, which circulates in Portugal and Spain, and groups with it in a subtype B envelope fragment, but the greatest part of its genome does not appear to derive from CRF14_BG, although both CRFs share as parental strain the subtype G variant circulating in the Iberian Peninsula. Phylogenetic clustering of partial pol and env segments from viruses collected in Portugal and Spain with X3208 and 9691_01 indicates that CRF73_BG is circulating in both countries, with proportions of around 2-3% Portuguese database HIV-1 isolates clustering with CRF73_BG. The fact that an HIV-1 recombinant virus characterized ten years ago as a URF has been shown to represent a CRF suggests that the number of HIV-1 CRFs may be much greater than currently known.

The analysis of near full-length genome sequences of human immunodeficiency virus type 1 BF intersubtype recombinant viruses from Chile, Venezuela and Spain reveals their relationship to diverse lineages of recombinant viruses related to CRF12_BF.

Human immunodeficiency virus type 1 (HIV-1) BF intersubtype recombinant viruses are common in Argentina and Uruguay, where CRF12_BF and related recombinants are frequently found, and, in a lower proportion, in Brazil. Full-length genome sequences have been characterized in several of these recombinant viruses. Here, we analyze six newly derived near full-length genome sequences of BF recombinant viruses, three from Chile, one from Venezuela and two from Spain. Five of them had known epidemiological links to Argentina. Genomes were amplified by PCR from plasma RNA or from peripheral blood mononuclear cells' DNA. Mosaic structures and phylogenetic relationships were analyzed by bootscanning, neighbour-joining phylogenetic trees and by examination of subtype signature nucleotides. One virus from Spain had a mosaic structure fully coincident with CRF12_BF. The others had unique mosaic structures, except the viruses from two Chilean sisters infected vertically from the same mother, who showed identical recombination patterns. Each of the unique recombinants had one to six breakpoints coincident with CRF12_BF and three also had two or three breakpoints coincident with a previously characterized unique recombinant from Argentina (A025) related to CRF12_BF. A phylogenetic tree of concatenated subtype F segments supported the relationship of five recombinants with CRF12_BF. In trees of partial subtype F and B segments, four recombinants clustered with A025. The examination of CRF12_BF signature amino acids and nucleotides supported the common ancestry of all the analyzed viruses. Based on these results, a model of generation of HIV-1 BF recombinants of Argentinean ancestry by successive rounds of recombination along diverse lineages deriving from a common BF recombinant ancestor related to CRF12_BF is proposed.

Analysis of discrepancies in the interpretation of antiretroviral drug resistance results in HIV-1 infected patients of Basque Country, Spain.

BACKGROUND: Genotypic and phenotypic analysis of HIV-1 resistance mutations constitute one important point for providing guidelines in the choice of antiretroviral regimens and to design lines of rescue treatment for patients holding HIV-1 drug-resistant variants. However, some levels of discordance among them has been described. OBJECTIVES: (i) To compare the genotypic analysis of resistance mutations to reverse transcriptase (RT) and protease (PR) inhibitors by Stanford HIVdb program (http://hivdb.stanford.edu) (St-HIVdb), and genotype with quantitative phenotypic analysis (Virtual Phenotype, VircoNET). (ii) To identify drug resistance mutations associated with discrepant results. STUDY DESIGN: Five hundred HIV-1 infected patients were included in this study. RNA was extracted from plasma. RT and PR regions were amplified and sequenced using ABI-Prism DNA sequencing system. Sequences were corrected and assembled with Seqman and Bioedit computer programs. The corrected sequences were submitted to the Stanford HIV-Seq program (http://hivdb.stanford.edu) and to Virtual Phenotype (VircoNET). RESULTS: Discrepant cases were considered if results were high or intermediate resistant by Stanford HIV-Seq program and susceptible by Virtual Phenotype, being detected as follows: (i) nucleoside RT inhibitors (NRT): 31.7% (ABC), 31% (d4T), 29.5% (ddC), 27.6% (ddI), 14.3% (TDF) and 11.3% (ZDV) and to PR inhibitors: 8.8% (SQV), 5% (APV), 3.8% (NFV) and 3.2% (IDV). These discrepant results were related to the presence of thymidine analogue mutations (TAMs) and also to key resistance mutations to NRT inhibitors: 65R, 69D/N, 74V/I, 184V/I and 215Y/F. (ii) PR inhibitors: 82A/F/T/S, 84I and 90M. Concordant results were considered when the interpretations by both programs were coincident, being higher than 96.7% for non-NRT inhibitors. CONCLUSIONS: The detection of discrepant results to NRT inhibitors and PR inhibitors, including the analysis of sequences with key resistant mutations to some drugs, means that further investigation is necessary in order to establish which is the best interpretation system as antiretroviral therapy guide.

Epidemiological Surveillance of HIV-1 Transmitted Drug Resistance in Spain in 2004-2012: Relevance of Transmission Clusters in the Propagation of Resistance Mutations.

Our objectives were to carry out an epidemiological surveillance study on transmitted drug resistance (TDR) among individuals newly diagnosed of HIV-1 infection during a nine year period in Spain and to assess the role of transmission clusters (TC) in the propagation of resistant strains. An overall of 1614 newly diagnosed individuals were included in the study from January 2004 through December 2012. Individuals come from two different Spanish regions: Galicia and the Basque Country. Resistance mutations to reverse transcriptase inhibitors (RTI) and protease inhibitors (PI) were analyzed according to mutations included in the surveillance drug-resistance mutations list updated in 2009. TC were defined as those comprising viruses from five or more individuals whose sequences clustered in maximum likelihood phylogenetic trees with a bootstrap value ≥90%. The overall prevalence of TDR to any drug was 9.9%: 4.9% to nucleoside RTIs (NRTIs), 3.6% to non-nucleoside RTIs (NNRTIs), and 2.7% to PIs. A significant decrease of TDR to NRTIs over time was observed [from 10% in 2004 to 2% in 2012 (p=0.01)]. Sixty eight (42.2%) of 161 sequences with TDR were included in 25 TC composed of 5 or more individuals. Of them, 9 clusters harbored TDR associated with high level resistance to antiretroviral drugs. T215D revertant mutation was transmitted in a large cluster comprising 25 individuals. The impact of epidemiological networks on TDR frequency may explain its persistence in newly diagnosed individuals. The knowledge of the populations involved in TC would facilitate the design of prevention programs and public health interventions.

Phylogeny and Phylogeography of a Recent HIV-1 Subtype F Outbreak among Men Who Have Sex with Men in Spain Deriving from a Cluster with a Wide Geographic Circulation in Western Europe.

We recently reported the rapid expansion of an HIV-1 subtype F cluster among men who have sex with men (MSM) in the region of Galicia, Northwest Spain. Here we update this outbreak, analyze near full-length genomes, determine phylogenetic relationships, and estimate its origin. For this study, we used sequences of HIV-1 protease-reverse transcriptase and env V3 region, and for 17 samples, near full-length genome sequences were obtained. Phylogenetic analyses were performed via maximum likelihood. Locations and times of most recent common ancestors were estimated using Bayesian inference. Among samples analyzed by us, 100 HIV-1 F1 subsubtype infections of monophyletic origin were diagnosed in Spain, including 88 in Galicia and 12 in four other regions. Most viruses (n = 90) grouped in a subcluster (Galician subcluster), while 7 from Valladolid (Central Spain) grouped in another subcluster. At least 94 individuals were sexually-infected males and at least 71 were MSM. Seventeen near full-length genomes were uniformly of F1 subsubtype. Through similarity searches and phylogenetic analyses, we identified 18 viruses from four other Western European countries [Switzerland (n = 8), Belgium (n = 5), France (n = 3), and United Kingdom (n = 2)] and one from Brazil, from samples collected in 2005-2011, which branched within the subtype F cluster, outside of both Spanish subclusters, most of them corresponding to recently infected individuals. The most probable geographic origin and age of the Galician subcluster was Ferrol, Northwest Galicia, around 2007, while the Western European cluster probably emerged in Switzerland around 2002. In conclusion, a recently expanded HIV-1 subtype F cluster, the largest non-subtype B cluster reported in Western Europe, continues to spread among MSM in Spain; this cluster is part of a larger cluster with a wide geographic circulation in diverse Western European countries.

Primary resistance mutations to fusion inhibitors and polymorphisms in gp41 sequences of HIV-1 non-B subtypes and recombinants.

Primary resistance mutations to fusion inhibitors and polymorphisms in gp41 sequences of non-B subtypes and recombinant HIV-1 isolates were analysed. L91H to RPR103611 was detected in one DGpol/Denv/Dgp41 recombinant; L9F and K144R, rarely reported previously, were frequent in the B region of CRF14_BG recombinants. V194I and V318A, not described in the G subtype, were detected in the G region of BG recombinants and in G subtype viruses that also show the rare mutations T115L, M118V and K90R.

High HIV-1 genetic diversity in Cuba.

BACKGROUND: HIV-1 subtype B is largely predominant in the Caribbean, although other subtypes have been recently identified in Cuba. OBJECTIVES: To examine HIV-1 genetic diversity in Cuba. METHODS: The study enrolled 105 HIV-1-infected individuals, 93 of whom had acquired the infection in Cuba. DNA from peripheral blood mononuclear cells was used for polymerase chain reaction amplification and sequencing of pol (protease-reverse transcriptase) and env (V3 region) segments. Phylogenetic trees were constructed using the neighbour-joining method. Intersubtype recombination was analysed by bootscanning. RESULTS: Of the samples, 50 (48%) were of subtype B and 55 (52%) of diverse non-B subtypes and recombinant forms. Among non-B viruses, 12 were non-recombinant, belonging to six subtypes (C, D, F1, G, H and J), the most frequent of which was subtype G (n = 5). The remaining 43 (78%) non-B viruses were recombinant, with 14 different forms, the two most common of which were Dpol/Aenv (n = 21) and U(unknown)pol/Henv (n = 7), which grouped in respective monophyletic clusters. Twelve recombinant viruses were mosaics of different genetic forms circulating in Cuba. Overall, 21 genetic forms were identified, with all known HIV-1 group M subtypes present in Cuba, either as non-recombinant viruses or as segments of recombinant forms. Non-B subtype viruses were predominant among heterosexuals (72%) and B subtype viruses among homo- or bisexuals (63%). CONCLUSION: An extraordinarily high diversity of HIV-1 genetic forms, unparalleled in the Americas and comparable to that found in Central Africa, is present in Cuba.