Gonzalo Yebra

Geographic and Temporal Trends in the Molecular Epidemiology and Genetic Mechanisms of Transmitted HIV-1 Drug Resistance: An Individual-Patient- and Sequence-Level Meta-Analysis

Background Regional and subtype-specific mutational patterns of HIV-1 transmitted drug resistance (TDR) are essential for informing first-line antiretroviral (ARV) therapy guidelines and designing diagnostic assays for use in regions where standard genotypic resistance testing is not affordable. We sought to understand the molecular epidemiology of TDR and to identify the HIV-1 drug-resistance mutations responsible for TDR in different regions and virus subtypes. Methods and Findings We reviewed all GenBank submissions of HIV-1 reverse transcriptase sequences with or without protease and identified 287 studies published between March 1, 2000, and December 31, 2013, with more than 25 recently or chronically infected ARV-naïve individuals. These studies comprised 50,870 individuals from 111 countries. Each set of study sequences was analyzed for phylogenetic clustering and the presence of 93 surveillance drug-resistance mutations (SDRMs). The median overall TDR prevalence in sub-Saharan Africa (SSA), south/southeast Asia (SSEA), upper-income Asian countries, Latin America/Caribbean, Europe, and North America was 2.8%, 2.9%, 5.6%, 7.6%, 9.4%, and 11.5%, respectively. In SSA, there was a yearly 1.09-fold (95% CI: 1.05–1.14) increase in odds of TDR since national ARV scale-up attributable to an increase in non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance. The odds of NNRTI-associated TDR also increased in Latin America/Caribbean (odds ratio [OR] = 1.16; 95% CI: 1.06–1.25), North America (OR = 1.19; 95% CI: 1.12–1.26), Europe (OR = 1.07; 95% CI: 1.01–1.13), and upper-income Asian countries (OR = 1.33; 95% CI: 1.12–1.55). In SSEA, there was no significant change in the odds of TDR since national ARV scale-up (OR = 0.97; 95% CI: 0.92–1.02). An analysis limited to sequences with mixtures at less than 0.5% of their nucleotide positions—a proxy for recent infection—yielded trends comparable to those obtained using the complete dataset. Four NNRTI SDRMs—K101E, K103N, Y181C, and G190A—accounted for >80% of NNRTI-associated TDR in all regions and subtypes. Sixteen nucleoside reverse transcriptase inhibitor (NRTI) SDRMs accounted for >69% of NRTI-associated TDR in all regions and subtypes. In SSA and SSEA, 89% of NNRTI SDRMs were associated with high-level resistance to nevirapine or efavirenz, whereas only 27% of NRTI SDRMs were associated with high-level resistance to zidovudine, lamivudine, tenofovir, or abacavir. Of 763 viruses with TDR in SSA and SSEA, 725 (95%) were genetically dissimilar; 38 (5%) formed 19 sequence pairs. Inherent limitations of this study are that some cohorts may not represent the broader regional population and that studies were heterogeneous with respect to duration of infection prior to sampling. Conclusions Most TDR strains in SSA and SSEA arose independently, suggesting that ARV regimens with a high genetic barrier to resistance combined with improved patient adherence may mitigate TDR increases by reducing the generation of new ARV-resistant strains. A small number of NNRTI-resistance mutations were responsible for most cases of high-level resistance, suggesting that inexpensive point-mutation assays to detect these mutations may be useful for pre-therapy screening in regions with high levels of TDR. In the context of a public health approach to ARV therapy, a reliable point-of-care genotypic resistance test could identify which patients should receive standard first-line therapy and which should receive a protease-inhibitor-containing regimen.

Increase of Non-B Subtypes and Recombinants Among Newly Diagnosed HIV-1 Native Spaniards and Immigrants in Spain

Although HIV-1 clade B variants are predominant in Western Europe, non-B subtypes are rapidly spreading, mainly due to immigration from endemic regions. All newly diagnosed HIV-1-infected individuals at a HIV/AIDS clinic in Madrid from 2000 to 2007 were identified. Subtype assignment was based on phylogenetic analysis of pol sequences from plasma specimens collected at first visit. A total of 1,430 newly diagnosed HIV-1 individuals were identified: 902 Spaniards, 232 South Americans, and 162 Africans, among others. The proportion of South-Americans and Africans among diagnosed HIV-1 patients increased from 2000 to 2007 (from 17% to 22% and from 4% to 21%, respectively). Half of diagnosis of HIV-1 in 2007 was in foreigners whereas in previous years Spaniards were predominant. Non-B variants were found in 157 (24%) of the 649 subjects who could be subtyped: 11A, 6C, 2D, 1F2, 13G, 4H, 1J, 3CRF01_AE, 64CRF02_AG, 2CRF03_AB, 3CRF06_cpx, 3CRF10_CD, 7CRF11_cpx, 9CRF12_BF, 9CRF14_BG, 1CRF18_cpx, 1CRF19_cpx, 2CRF31_BC, 10 URF and 5 outgroups. They represented 93%, 14% and 4% of newly-diagnosed HIV-1 Africans, South-Americans and native Spaniards, respectively. Non-B subtypes increased from 9% in 2000 to 32% in 2007, specially among South-Americans (from 11% to 20%) and native Spaniards (from 4% to 10%). Most (75%) were recombinant viruses. The highest number and diversity of HIV-1 variants among natives was observed in 2007. HIV-1 non-B subtypes are increasingly present among newly diagnosed HIV-1 individuals in Madrid, representing a third of cases in 2007, whereas 10% of newly diagnosed HIV-1 native Spaniards had non-B viruses.

Most HIV Type 1 Non-B Infections in the Spanish Cohort of Antiretroviral Treatment-Naïve HIV-Infected Patients (CoRIS) Are Due to Recombinant Viruses

ABSTRACT HIV-1 group M is classified into 9 subtypes, as well as recombinants favored by coinfection and superinfection events with different variants. Although HIV-1 subtype B is predominant in Europe, intersubtype recombinants are increasing in prevalence and complexity. In this study, phylogenetic analyses of pol sequences were performed to detect the HIV-1 circulating and unique recombinant forms (CRFs and URFs, respectively) in a Spanish cohort of antiretroviral treatment-naïve HIV-infected patients included in the Research Network on HIV/AIDS (CoRIS). Bootscanning and other methods were used to define complex recombinants not assigned to any subtype or CRF. A total of 670 available HIV-1 pol sequences from different patients were collected, of which 588 (87.8%) were assigned to HIV-1 subtype B and 82 (12.2%) to HIV-1 non-B variants. Recombinants caused the majority (71.9%) of HIV-1 non-B infections and were found in 8.8% of CoRIS patients. Eleven URFs (accounting for 13.4% of HIV-1 non-B infections), presenting complex mosaic patterns, were detected. Among them, 10 harbored subtype B fragments. Four of the 11 URFs were found in Spanish natives. A cluster of three B/CRF02_AG recombinants was detected. We conclude that complex variants, including unique recombinant forms, are being introduced into Spain through both immigrants and natives. An increase in the frequency of mosaic viruses, reflecting the increasing heterogeneity of the HIV epidemic in our country, is expected.

Phylogenetic and demographic characterization of HIV-1 transmission in Madrid, Spain

BACKGROUNDnThe combination of phylogenetic analyses of HIV sequences with patients demographic data allows us to understand local HIV transmission, a necessary knowledge for designing prevention strategies. The Community of Madrid represents a challenge for the control of HIV epidemic in Spain given its high HIV prevalence and increasing proportion of immigrant people among HIV-infected population.nnnMETHODSnWe applied maximum likelihood methods and Bayesian Markov chain Monte Carlo (MCMC) inference using the program BEAST to a set of HIV-1 pol sequences from 1293 patients diagnosed in 1995-2010 in Madrid, Spain.nnnRESULTSnTwo-hundred and thirty six patients (18.2% of the cohort) were included in 100 transmission chains using phylogenetic criteria, 67 (67%) belonging to HIV-1 subtype B and 33 (33%) to 11 different non-B strains, especially BG and BF recombinants. Most networks involved transmission between MSM (48/100). Half of non-B clusters (15/33) included at least one Spaniard. Sub-Saharan African patients presented a low linkage rate (9%) in contrast to Spanish (21%) and Latin American (25%) patients. Three clusters involving treatment-independent transmission of drug-resistance mutations were found.nnnCONCLUSIONSnOne out of five HIV-infected patients in our cohort in Madrid was epidemically linked, mainly by transmission pairs. The inclusion in transmission networks was more likely for MSM, Spaniards and patients from Latin America. We found no evidence of self-sustained non-B epidemics due to the absence of large transmission chains with the exception of Cuban BG recombinants and CRF47_BF. However, the differences in transmission across variants are probably determined by the patient profile, especially the infection route.

High Drug Resistance Prevalence among Vertically HIV-Infected Patients Transferred from Pediatric Care to Adult Units in Spain

Background Antiretroviral treatment (ART) has contributed to increased life expectancy of HIV-1 infected children. In developed countries, an increasing number of children reaching adulthood are transferred to adult units. The objectives were to describe the demographic and clinical features, ART history, antiviral drug resistance and drug susceptibility in HIV-1 perinatally infected adolescents transferred to adult care units in Spain from the Madrid Cohort of HIV-1 infected children. Methods Clinical, virological and immunological features of HIV-1 vertically infected patients in the Madrid Cohort of HIV-infected children were analyzed at the time of transfer. Pol sequences from each patient were recovered before transfer. Resistance mutations according to the InternationaI AIDS Society 2011 list were identified and interpreted using the Stanford algorithm. Results were compared to the non-transferred HIV-1 infected pediatric cohort from Madrid. Results One hundred twelve infected patients were transferred to adult units between 1997 and 2011. They were mainly perinatally infected (93.7%), with a mean nadir CD4+-T-cells count of 10% and presented moderate or severe clinical symptoms (75%). By the time of transfer, the mean age was 18.9 years, the mean CD4+T-cells count was 627.5 cells/ml, 64.2% presented more than 350 CD4+T-cells/ml and 47.3% had ≤200 RNA-copies/ml. Most (97.3%) were ART experienced receiving Highly Active ART (HAART) (84.8%). Resistance prevalence among pretreated was 50.9%, 76.9% and 36.5% for Protease Inhibitors (PI), Nucleoside Reverse Transcriptase Inhibitors (NRTI) and Non-NRTI (NNRTI), respectively. Resistance mutations were significantly higher among transferred patients compared to non-transferred for the PI+NRTI combination (19% vs. 8.4%). Triple resistance was similar to non-transferred pediatric patients (17.3% vs. 17.6%). Conclusion Despite a good immunological and virological control before transfer, we found high levels of resistance to PI, NRTI and triple drug resistance in HIV-1 infected adolescents transferred to adult units.

Clinical Differences and Viral Diversity between Newly HIV Type 1-Diagnosed African and Non-African Patients in Spain (2005–2007)

Abstract The diagnosis of HIV-1 is increasing in African-born persons residing in Europe. They present a high prevalence of HIV-1 non-B variant infections and of parasitic infections, both of which are infrequent in Western countries. Immigration favors their presence in nonendemic countries. In this study, all newly HIV-diagnosed individuals at an HIV/AIDS and Tropical Medicine reference center in Madrid from 2005 through 2007 were retrospectively studied. HIV-1 subtyping was performed in gag, pol, and gp41 coding regions by phylogenetic analyses. The presence of other pathogens was also evaluated. Furthermore, all HIV-1-infected Africans were screened for parasitic infections. Newly diagnosed HIV-1 subjects included 90 sub-Saharan Africans and 188 non-Africans (116 Spaniards, 13 other Europeans, and 59 Latin Americans). Significantly higher numbers of HIV-1-infected Africans than non-Africans were females, acquired HIV-1 by heterosexual contact, and presented a more advanced clinical CDC stage and criteria for starting antiretroviral therapy in the first clinical visit. They predominantly carried non-B subtype infections, mainly intersubtype recombinants. Half of HIV-1-infected Africans had parasitic infections. CD4(+) T cell counts were lower among Africans than Europeans at the time of HIV-1 diagnosis. At 12 months of follow-up after starting antiretroviral treatment, a significantly lower proportion of Africans than non-Africans achieved undetectable viremia due to their higher loss to follow-up. However, CD4(+) T cell recovery and virological failure rates were similar. Therefore, the profile of African HIV-1-infected immigrants varies widely with respect to Spanish HIV-infected individuals. More advanced immunodeficiency and the coexistence of parasitic diseases and infections with a large diversity of HIV-1 non-B and recombinant variants are expected.

Different trends of transmitted HIV-1 drug resistance in Madrid, Spain, among risk groups in the last decade

Abstract The presence of transmitted HIV drug resistance (TDR) threatens the efficacy of antiretroviral treatment. We aimed to assess the changes in TDR prevalence over the last decade in Madrid, Spain, to verify the role of the patients’ risk groups in these changes. We analysed the trends of TDR between 2000 and 2011 in a cohort of 1,022 naïve HIV-infected patients in Madrid, Spain, whose pol sequences were available. They included, among others, 369 heterosexuals, 340 men who have sex with men (MSM), and 90 injection drug users (IDUs). TDR was reported following the WHO mutation list. The TDR rate in the whole cohort was 8.3xa0%, being the highest in MSM (9.1xa0%) and the lowest in IDUs (4.4xa0%). Over time, this rate decreased significantly (to 5.4xa0% in 2009-2011) since the period 2004-2006, when it peaked (10.7xa0%). Heterosexuals and IDUs showed similar trends, but in the 2009-2011 period, the TDR rate among MSM rebounded to 9.0xa0% (being absent among IDUs). TDR stabilized in the last years (2007-2011) for nucleoside reverse transcriptase inhibitors. The risk group also determined differences in the mutational profile, sex distribution, proportion of immigrants, and viral variants. In conclusion, the risk group caused different HIV sub-epidemics, determined by the patients’ profiles. Despite the general decreasing trend in TDR, we observed a non-significant increase in TDR rate among MSM, a tendency that needs confirmation. Periodic TDR surveillance is important to prevent the widespread distribution of resistance, especially in MSM, given the growing HIV/AIDS epidemic in this high-risk population.

Drug resistance prevalence and HIV-1 variant characterization in the naive and pretreated HIV-1-infected paediatric population in Madrid, Spain

BACKGROUNDnDrug resistance mutations affect antiretroviral therapy (ART) effectiveness in HIV-1-infected children, compromising long-term therapy. HIV-1 variants and drug resistance mutations were identified in HIV-infected children from Madrid, Spain.nnnMETHODSnPatients from the Madrid cohort of HIV-infected children (1993-2009) with available pol sequences or infected samples stored at the Spanish HIV-1 BioBank were selected. Specimens were used to perform new pol sequences when not available. HIV-1 variants were characterized by phylogenetic analysis. Resistance mutations were identified according to the International AIDS Society-USA list (2009).nnnRESULTSnIn 198 patients, pol sequences were recovered from routine resistance testing (nu200a=u200a98) or newly performed using stored plasma, lymphocytes or DNA (nu200a=u200a100). Patients were mostly Europeans (90%), with moderate to severe AIDS symptoms (65%), on ART (85%) when the specimen was sequenced and infected by subtype B (90%). Among the 19 HIV-1 non-B variants found, 58% were recombinants (8CRF02_AG, 1CRF08_BC, 1CRF12_BF and 1CRF13_cpx) and the rest were pure non-B subtypes (1A2, 2C, 2D, 1F1, 1G and 1H). Transmitted drug resistance (TDR) mutations were detected in 13% of naive children; 4%, 7% and 10% for protease inhibitors (PIs), nucleoside reverse transcriptase inhibitors (NRTIs) and non-nucleoside reverse transcriptase inhibitors (NNRTIs), respectively. Global resistance prevalence was higher (66%) among ART-exposed children; 37% for PIs, 54% for NRTIs and 35% for NNRTIs.nnnCONCLUSIONSnHIV-1 non-B variants infected 10% of the cohort during 1993-2009. Resistant viruses were present in 26.5% and 66% of naive and pretreated children, respectively. Our data suggest that TDR prevalence in children could be higher than that reported in adults in Spain. The provided data will help to improve clinical management of HIV-infected children in Spain.

Analysis of the history and spread of HIV-1 in Uganda using phylodynamics

HIV prevalence has decreased in Uganda since the 1990s, but remains substantial within high-risk groups. Here, we reconstruct the history and spread of HIV subtypes A1 and D in Uganda and explore the transmission dynamics in high-risk populations. We analysed HIV pol sequences from female sex workers in Kampala (nu200a=u200a42), Lake Victoria fisher-folk (nu200a=u200a46) and a rural clinical cohort (nu200a=u200a74), together with publicly available sequences from adjacent regions in Uganda (nu200a=u200a412) and newly generated sequences from samples taken in Kampala in 1986 (nu200a=u200a12). Of the sequences from the three Ugandan populations, 60 (37.1u200a%) were classified as subtype D, 54 (33.3u200a%) as subtype A1, 31 (19.1u200a%) as A1/D recombinants, six (3.7u200a%) as subtype C, one (0.6u200a%) as subtype G and 10 (6.2u200a%) as other recombinants. Among the A1/D recombinants we identified a new candidate circulating recombinant form. Phylodynamic and phylogeographic analyses using BEAST indicated that the Ugandan epidemics originated in 1960 (1950–1968) for subtype A1 and 1973 (1970–1977) for D, in rural south-western Uganda with subsequent spread to Kampala. They also showed extensive interconnection with adjacent countries. The sequence analysis shows both epidemics grew exponentially during the 1970s–1980s and decreased from 1992, which agrees with HIV prevalence reports in Uganda. Inclusion of sequences from the 1980s indicated the origin of both epidemics was more recent than expected and substantially narrowed the confidence intervals in comparison to previous estimates. We identified three transmission clusters and ten pairs, none of them including patients from different populations, suggesting active transmission within a structured transmission network.

Trends in drug resistance prevalence in HIV-1-infected children in Madrid: 1993 to 2010 analysis.

Background: Drug resistance mutations compromise antiretroviral treatment (ART) effectiveness in HIV-1–infected children. Trends in drug resistance prevalence have not been previously evaluated in HIV-infected children in Spain. Methods: HIV-1 variants, drug resistance prevalence dynamics and drug susceptibility were analyzed from 1993 to 2010 in HIV-infected children with available pol sequence, sample or drug resistance profile. HIV-1 variants were characterized by phylogenetic analysis. Resistance mutations in pretreated and naive patients were identified according to International AIDS Society-2010 and the World Health Organization list, respectively. Results: In 232 patients, genotypic resistance profiles (n = 11) or pol sequences (n = 128) were recovered or newly generated from infected samples (n = 93). Patients were mainly in care at pediatric units (63%), were mostly Europeans (84%), with moderate AIDS symptoms (65%), on ART (91%) and infected by HIV-1 subtype B (89%). Transmitted major drug resistance mutations were selected in 6 (13.6%) of the 44 ART-naive children: 4.8%, 9.3% and 11.6%, for protease inhibitors, nucleoside reverse transcriptase inhibitors and nonnucleoside reverse transcriptase inhibitors, respectively. Overall resistance prevalence was higher (71.8%) among ART-exposed children: 39.9%, 66.5% and 35.3% for protease inhibitors, nucleoside reverse transcriptase inhibitors and nonnucleoside reverse transcriptase inhibitors, respectively. Resistance prevalence among ART-exposed children was higher in 2009 to 2010 relative to 1993 to1999 for nonnucleoside reverse transcriptase inhibitors (42% versus 6%; P = 0.006), protease inhibitors (39% versus 13%; P = 0.004) and nucleoside reverse transcriptase inhibitors (63% versus 44%; P = NS). Susceptibility to each drug in resistant viruses was predicted. The rate of non-B infections increased in the last years, mainly caused by recombinant viruses. Conclusions: The increasing resistance prevalence among the HIV-infected pediatric population in Spain highlights the importance of specific drug resistance and drug susceptibility surveillance in long-term pretreated children to optimize treatment regimens.