HIV-1 non-B Subtypes Research Articles

Relative resistance of patient-derived envelope sequences to SERINC5-mediated restriction of HIV-1 infectivity.

Pathogenesis of HIV-1 is enhanced through several viral-encoded proteins that counteract a range of host restriction molecules. HIV-1 Nef counteracts the cell membrane protein SERINC5 by downregulating it from the cell surface, thereby enhancing virion infectivity. Some subtype B reference Envelope sequences have shown the ability to bypass SERINC5 infectivity restriction independent of Nef. However, it is not clear if and to what extent circulating HIV-1 strains can exhibit resistance to SERINC5 restriction. Using a panel of Envelope sequences isolated from 50 Tanzanians infected with non-B HIV-1 subtypes, we show that the lentiviral reporters pseudotyped with patient-derived Envelopes have reduced sensitivity to SERINC5 and that this sensitivity differed among viral subtypes. Moreover, we found that SERINC5 sensitivity within patient-derived Envelopes can be modulated by separate regions, highlighting the complexity of viral/host interactions.

Limited emergence of resistance to integrase strand transfer inhibitors (INSTIs) in ART-experienced participants failing dolutegravir-based antiretroviral therapy: a cross-sectional analysis of a Northeast Nigerian cohort.

Due to the high prevalence of resistance to NNRTI-based ART since 2018, consolidated recommendations from the WHO have indicated dolutegravir as the preferred drug of choice for HIV treatment globally. There is a paucity of resistance outcome data from HIV-1 non-B subtypes circulating across West Africa. We characterized the mutational profiles of persons living with HIV from a cross-sectional cohort in North-East Nigeria failing a dolutegravir-based ART regimen. WGS of plasma samples collected from 61 HIV-1-infected participants following virological failure of dolutegravir-based ART were sequenced using the Illumina platform. Sequencing was successfully completed for samples from 55 participants. Following quality control, 33 full genomes were analysed from participants with a median age of 40 years and median time on ART of 9 years. HIV-1 subtyping was performed using SNAPPy. Most participants had mutational profiles reflective of exposure to previous first- and second-line ART regimens comprised NRTIs and NNRTIs. More than half of participants had one or more drug resistance-associated mutations (DRMs) affecting susceptibility to NRTIs (17/33; 52%) and NNRTIs (24/33; 73%). Almost a quarter of participants (8/33; 24.4%) had one or more DRMs affecting tenofovir susceptibility. Only one participant, infected with HIV-1 subtype G, had evidence of DRMs affecting dolutegravir susceptibility-this was characterized by the T66A, G118R, E138K and R263K mutations. This study found a low prevalence of resistance to dolutegravir; the data are therefore supportive of the continual rollout of dolutegravir as the primary first-line regimen for ART-naive participants and the preferred switch to second-line ART across the region. However, population-level, longer-term data collection on dolutegravir outcomes are required to further guide implementation and policy action across the region.

Attenuated HIV-1 Nef But Not Vpu Function in a Cohort of Rwandan Long-Term Survivors.

HIV-1 accessory proteins Nef and Vpu enhance viral pathogenesis through partially overlapping immune evasion activities. Attenuated Nef or Vpu functions have been reported in individuals who display slower disease progression, but few studies have assessed the relative impact of these proteins in non-B HIV-1 subtypes or examined paired proteins from the same individuals. Here, we examined the sequence and function of matched Nef and Vpu clones isolated from 29 long-term survivors (LTS) from Rwanda living with HIV-1 subtype A and compared our results to those of 104 Nef and 62 Vpu clones isolated from individuals living with chronic untreated HIV-1 subtype A from the same geographic area. Nef and vpu coding regions were amplified from plasma HIV RNA and cloned. The function of one intact, phylogenetically-validated Nef and Vpu clone per individual was then quantified by flow cytometry following transient expression in an immortalized CD4+ T-cell line. We measured the ability of each Nef clone to downregulate CD4 and HLA class I, and of each Vpu clone to downregulate CD4 and Tetherin, from the cell surface. Results were normalized to reference clones (Nef-SF2 and Vpu-NL4.3). We observed that Nef-mediated CD4 and HLA downregulation functions were lower in LTS compared to the control cohort (Mann-Whitney p=0.03 and p<0.0001, respectively). Moreover, we found a positive correlation between Nef-mediated CD4 downregulation function and plasma viral load in LTS and controls (Spearman ρ= 0.59, p=0.03 and ρ=0.30, p=0.005, respectively). In contrast, Vpu-mediated functions were similar between groups and did not correlate with clinical markers. Further analyses identified polymorphisms at Nef codon 184 and Vpu codons 60-62 that were associated with function, which were confirmed through mutagenesis. Overall, our results support attenuated function of Nef, but not Vpu, as a contributor to slower disease progression in this cohort of long-term survivors with HIV-1 subtype A.

Soluble CD14 is subtype-dependent in serum but not in cerebrospinal fluid in people with HIV

Monocytes and macrophages activation are crucial in human immunodeficiency virus (HIV) central nervous system (CNS) infection and HIV associated neurocognitive disorders (HAND) pathogenesis. The soluble form of CD14 (sCD14) is a marker of monocyte activation. We hypothesized that sCD14 levels would be lower in people with HIV-1 subtype C (HIV-1C) than in HIV-1B owing to a variant Tat cysteine dimotif (C30S31) with reduced chemotactic activity. A total of 68 paired cerebrospinal fluid (CSF) and blood samples from people with HIV (PWH); 27 samples of the HIV-1B subtype and 40 of the non-B HIV-1 subtypes (including 26,HIV-1C), and 18 HIV-negative controls were included. sCD14 levels were quantified using a high-sensitivity enzyme-linked immunosorbent assay. sCD14 increase in serum, but not in CSF, was higher in samples from HIV-1B than HIV-1C (p = 0.002; Cohen's d, 0.7). CSF or serum sCD14 values were not correlated with global deficit score or specific cognitive domains. The impact of HIV-1 on monocyte stimulation biomarkers evaluated by sCD14 in serum was subtype-dependent, higher in HIV-1B than HIV-1C, consistent with reduced chemotactic activity as hypothesized.

Detection of Gag C-terminal mutations among HIV-1 non-B subtypes in a subset of Cameroonian patients

Response to ritonavir-boosted-protease inhibitors (PI/r)-based regimen is associated with some Gag mutations among HIV-1 B-clade. There is limited data on Gag mutations and their covariation with mutations in protease among HIV-1 non-B-clades at PI/r-based treatment failure. Thus, we characterized Gag mutations present in isolates from HIV-1 infected individuals treated with a PI/r-regimen (n = 143) and compared them with those obtained from individuals not treated with PI/r (ART-naïve [n = 101] or reverse transcriptase inhibitors (RTI) treated [n = 118]). The most frequent HIV-1 subtypes were CRF02_AG (54.69%), A (13.53%), D (6.35%) and G (4.69%). Eighteen Gag mutations showed a significantly higher prevalence in PI/r-treated isolates compared to ART-naïve (p < 0.05): Group 1 (prevalence < 1% in drug-naïve): L449F, D480N, L483Q, Y484P, T487V; group 2 (prevalence 1–5% in drug-naïve): S462L, I479G, I479K, D480E; group 3 (prevalence ≥ 5% in drug-naïve): P453L, E460A, R464G, S465F, V467E, Q474P, I479R, E482G, T487A. Five Gag mutations (L449F, P453L, D480E, S465F, Y484P) positively correlated (Phi ≥ 0.2, p < 0.05) with protease-resistance mutations. At PI/r-failure, no significant difference was observed between patients with and without these associated Gag mutations in term of viremia or CD4 count. This analysis suggests that some Gag mutations show an increased frequency in patients failing PIs among HIV-1 non-B clades.

Increasing Frequency and Transmission of HIV-1 Non-B Subtypes Among Men Who Have Sex With Men in the Swiss HIV Cohort Study.

In Switzerland, HIV-1 transmission among men who have sex with men (MSM) has been dominated by subtype B, whilst non-B subtypes are commonly attributed to infections acquired abroad among heterosexuals. Here, we evaluated the temporal trends of non-B subtypes and the characteristics of molecular transmission clusters (MTCs) among MSM. Sociodemographic and clinical data and partial pol sequences were obtained from participants enrolled in the Swiss HIV Cohort Study. For non-B subtypes, maximum likelihood trees were constructed, from which Swiss MTCs were identified and analyzed by transmission group. Non-B subtypes were identified in 8.1% (416/5116) of MSM participants. CRF01_AE was the most prevalent strain (3.5%), followed by subtype A (1.2%), F (1.1%), CRF02_AG (1.1%), C (0.9%), and G (0.3%). Between 1990 and 2019, an increase in the proportion of newly diagnosed individuals (0/123 [0%] to 11/32 [34%]) with non-B subtypes in MSM was found. Across all non-B subtypes, the majority of MSM MTCs were European. Larger MTCs were observed for MSM than heterosexuals. We found a substantial increase in HIV-1 non-B subtypes among MSM in Switzerland and the occurrence of large MTCs, highlighting the importance of molecular surveillance in guiding public health strategies targeting the HIV-1 epidemic.

Phenotypic and Genotypic Co-receptor Tropism Testing in HIV-1 Epidemic Region of Tanzania Where Multiple Non-B Subtypes Co-circulate.

HIV human immunodeficiency virus type I (HIV-1) entry inhibitor potency is dependent on viral co-receptor tropisms and thereby tropism determination is clinically important. However, phenotypic tropisms of HIV-1 non-B subtypes have been poorly investigated and the genotypic prediction algorithms remain insufficiently validated. To clarify this issue, we recruited 52 treatment-naïve, HIV-1-infected patients in Tanzania, where multiple HIV-1 non-B subtypes co-circulate. Sequence analysis of 93 infectious envelope clones isolated from their plasma viral RNA revealed the co-circulation of subtypes A1, C, D, and inter-subtype recombinant forms (isRFs). Phenotypic tropism assays revealed that lentivirus reporters pseudotyped with 75 (80.6%) and 5 (5.4%) envelope clones could establish infection toward U87.CD4 cells expressing CCR5 (R5) and CXCR4 (X4), respectively; whereas the remaining 13 (14%) clones could infect both cells. Genotypic analyses by widely used algorithms including V3 net charge, Geno2pheno, WebPSSM, and PhenoSeq showed that almost all phenotypic X4-tropic clones and only 15 of 75 phenotypic R5-tropic clones were concordantly predicted. However, the remaining 60 phenotypic R5-tropic clones were discordantly predicted by at least one algorithm. In particular, 2 phenotypic R5-tropic clones were discordantly predicted by all algorithms tested. Taken together, the results demonstrate the limitation of currently available genotypic algorithms for predicting co-receptor inference among co-circulating multiple non-B subtypes and emerging isRFs. Also, the phenotypic tropism dataset presented here could be valuable for retraining of the widely used genotypic prediction algorithms to enhance their performance.

Presence of HIV-1 G-to-A mutations linked to APOBEC editing is more prevalent in non-B HIV-1 subtypes and is associated with lower HIV-1 reservoir.

APOBEC3 editing activity contributes to sequences variation and viral diversification. We aimed to characterize virological and clinical factors associated with G-to-A mutations and stop codons in the HIV-1 reservoir, markers of APOBEC3 footprints, in order to better understand HIV-1 diversity among virologically suppressed HIV-1-infected patients. Immuno-virological and clinical factors were compared between 92 patients harbouring G-to-A mutations and stop codons (APOBEC+) in the reverse transcriptase gene and 92 patients without G-to-A mutations (APOBEC-) and stop codons in their DNA genotypes. Patients were predominantly men (74.5%) and were mostly infected by B-subtype (69.0%), with 44.1% and 55.9% in APOBEC+ and APOBEC- groups, respectively. At time of HIV DNA genotypes, the total cell-associated HIV-1 DNA load was 2.34 log10 copies/106 cells (IQR 1.85-2.67) and 33.2% of them had a detectable ultrasensitive plasma viral load. Hypermutated sequences were identified in 28.2% of the APOBEC+ group. The median total cell-associated HIV-1 DNA level was significantly lower in APOBEC+ than APOBEC- group: 2.13 log10 copies/106 cells (IQR 1.60-2.60) versus 2.52 log10 copies/106 cells (IQR 2.19-2.71) (P < 0.001), respectively. Presence of G-to-A mutations and stop codon was independently associated with HIV-1 subtype non-B (P = 0.017). These results show an independent association between the presence of G-to-A mutations and stop codons with HIV-1 subtype non-B and low proviral DNA that could be explained by the APOBEC3 footprints and restriction of DNA synthesis and integration. However, further investigations are needed to study the contribution of Vif amino acid variability among HIV-1 subtypes.

Factors Influencing HIV Drug Resistance among Pregnant Women in Luanda, Angola: Findings from a Cross-Sectional Study.

The increase in HIV infection and drug-resistant strains is an important public health concern, especially in resource-limited settings. However, the identification of factors related to the propagation of infectious diseases represents a crucial target offering an opportunity to reduce health care costs as well as deepening the focus on preventing infection in high-risk groups. In this study, we investigate the factors related to drug resistance among HIV-infected pregnant women in Luanda, the capital city of Angola. This was a part of a cross-sectional study conducted with 42 HIV-positive pregnant women. A blood sample was collected, and HIV-1 genotyping was carried out using an in-house method. Multivariate analyses were performed to determine the interaction between sociodemographic characteristics and drug resistance. HIV drug resistance was detected in 44.1% of the studied population. High probabilities of drug resistance were observed for HIV-infected pregnant women living in rural areas (AOR: 2.73; 95% CI: 0.50–14.9) with high educational level (AOR: 6.27; 95% CI: 0.77–51.2) and comorbidities (AOR: 5.47; 95% CI: 0.28–106) and infected with a HIV-1 non-B subtype other than subtype C (AOR: 1.60; 95% CI: 0.25–10.3). The present study reports high HIV drug resistance. Furthermore, older-age, rural areas, high educational levels, unemployed status, having comorbidities, and HIV-1 subtypes were factors related to drug resistance. These factors impact on drug susceptibility and need to be urgently addressed in order to promote health education campaigns able to prevent the spread of drug-resistant HIV strains in Angola.

Pre-treatment integrase inhibitor resistance is uncommon in antiretroviral therapy-naive individuals with HIV-1 subtype A1 and D infections in Uganda.

Dolutegravir (DTG) is now a preferred component of first-line antiretroviral therapy (ART). However, prevalence data on natural resistance to integrase inhibitors [integrase strand transfer inhibitors (INSTIs)] in circulating non-subtype B HIV-1 in sub-Saharan Africa is scarce. Our objective is to report prevalence of pre-treatment integrase polymorphisms associated with resistance to INSTIs in an ART-naive cohort with diverse HIV-1 subtypes. We retrospectively examined HIV-1 integrase sequences from Uganda. Plasma samples were derived from the Uganda AIDS Rural Treatment Outcomes (UARTO) cohort, reflecting enrollment from 2002 to 2010, prior to initiation of ART. HIV-1 integrase was amplified using nested-PCR and Sanger-sequenced (HXB2 4230-5093). Stanford HIVdb v8.8 was used to infer clinically significant INSTI-associated mutations. Human leukocyte antigen (HLA) typing was performed for all study participants. Plasma samples from 511 ART-naive individuals (subtype: 48% A1, 39% D) yielded HIV-1 integrase genotyping results. Six out of 511 participants (1.2%) had any major INSTI-associated mutations. Of these, two had E138T (subtype A1), three had E138E/K (subtype D), and one had T66T/I (subtype D). No participants had mutations traditionally associated with high levels of INSTI resistance. HLA genotypes A∗02:01/05/14, B∗44:15, and C∗04:07 predicted the presence of L74I, a mutation recently observed in association with long-acting INSTI cabotegravir virologic failure. We detected no HIV-1 polymorphisms associated with high levels of DTG resistance in Uganda in the pre-DTG era. Our results support widespread implementation of DTG but careful monitoring of patients on INSTI with virologic failure is warranted to determine if unique mutations predict failure for non-B subtypes of HIV-1.

Accumulation of integrase strand transfer inhibitor resistance mutations confers high-level resistance to dolutegravir in non-B subtype HIV-1 strains from patients failing raltegravir in Uganda.

Increasing first-line treatment failures in low- and middle-income countries (LMICs) have led to increased use of integrase strand transfer inhibitors (INSTIs) such as dolutegravir. However, HIV-1 susceptibility to INSTIs in LMICs, especially with previous raltegravir exposure, is poorly understood due to infrequent reporting of INSTI failures and testing for INSTI drug resistance mutations (DRMs). A total of 51 non-subtype B HIV-1 infected patients failing third-line (raltegravir-based) therapy in Uganda were initially selected for the study. DRMs were detected using Sanger and deep sequencing. HIV integrase genes of 13 patients were cloned and replication capacities (RCs) and phenotypic susceptibilities to dolutegravir, raltegravir and elvitegravir were determined with TZM-bl cells. Spearman's correlation coefficient was used to determine cross-resistance between INSTIs. INSTI DRMs were detected in 47% of patients. HIV integrase-recombinant virus carrying one primary INSTI DRM (N155H or Y143R/S) was susceptible to dolutegravir but highly resistant to raltegravir and elvitegravir (>50-fold change). Two patients, one with E138A/G140A/Q148R/G163R and one with E138K/G140A/S147G/Q148K, displayed the highest reported resistance to raltegravir, elvitegravir and even dolutegravir. The former multi-DRM virus had WT RC whereas the latter had lower RCs than WT. In HIV-1 subtype A- and D-infected patients failing raltegravir and harbouring INSTI DRMs, there is high-level resistance to elvitegravir and raltegravir. More routine monitoring of INSTI treatment may be advised in LMICs, considering that multiple INSTI DRMs may have accumulated during prolonged exposure to raltegravir during virological failure, leading to high-level INSTI resistance, including dolutegravir resistance.

Evaluation of HIV Transmission Clusters among Natives and Foreigners Living in Italy.

We aimed at evaluating the characteristics of HIV-1 molecular transmission clusters (MTCs) among natives and migrants living in Italy, diagnosed between 1998 and 2018. Phylogenetic analyses were performed on HIV-1 polymerase (pol) sequences to characterise subtypes and identify MTCs, divided into small (SMTCs, 2–3 sequences), medium (MMTCs, 4–9 sequences) and large (LMTCs, ≥10 sequences). Among 3499 drug-naïve individuals enrolled in the Italian Cohort Naive Antiretroviral (ICONA) cohort (2804 natives; 695 migrants), 726 (20.8%; 644 natives, 82 migrants) were involved in 228 MTCs (6 LMTCs, 36 MMTCs, 186 SMTCs). Migrants contributed 14.4% to SMTCs, 7.6% to MMTCs and 7.1% to LMTCs, respectively. HIV-1 non-B subtypes were found in 51 MTCs; noteworthy was that non-B infections involved in MTCs were more commonly found in natives (n = 47) than in migrants (n = 4). Factors such as Italian origin, being men who have sex with men (MSM), younger age, more recent diagnosis and a higher CD4 count were significantly associated with MTCs. Our findings show that HIV-1 clustering transmission among newly diagnosed individuals living in Italy is prevalently driven by natives, mainly MSM, with a more recent diagnosis and frequently infected with HIV-1 non-B subtypes. These results can contribute to monitoring of the HIV epidemic and guiding the public health response to prevent new HIV infections.

High prevalence of integrase mutation L74I in West African HIV-1 subtypes prior to integrase inhibitor treatment.

ObjectivesHIV-1 integrase inhibitors are recommended as first-line therapy by WHO, though efficacy and resistance data for non-B subtypes are limited. Two recent trials have identified the integrase L74I mutation to be associated with integrase inhibitor treatment failure in HIV-1 non-B subtypes. We sought to define the prevalence of integrase resistance mutations, including L74I, in West Africa.MethodsWe studied a Nigerian cohort of recipients prior to and during receipt of second-line PI-based therapy, who were integrase inhibitor-naive. Illumina next-generation sequencing with target enrichment was used on stored plasma samples. Drug resistance was interpreted using the Stanford Resistance Database and the IAS-USA 2019 mutation lists.ResultsOf 115 individuals, 59.1% harboured CRF02_AG HIV-1 and 40.9% harboured subtype G HIV-1. Four participants had major IAS-USA integrase resistance-associated mutations detected at low levels (2%–5% frequency). Two had Q148K minority variants and two had R263K (one of whom also had L74I). L74I was detected in plasma samples at over 2% frequency in 40% (46/115). Twelve (26.1%) had low-level minority variants of between 2% and 20% of the viral population sampled. The remaining 34 (73.9%) had L74I present at >20% frequency. L74I was more common among those with subtype G infection (55.3%, 26/47) than those with CRF02_AG infection (29.4%, 20/68) (P = 0.005).ConclusionsHIV-1 subtypes circulating in West Africa appear to have very low prevalence of major integrase mutations, but significant prevalence of L74I. A combination of in vitro and clinical studies is warranted to understand the potential implications.

HIV-1 subtype variability and transmitted drug resistance in a culturally diverse population in Western Sydney, New South Wales, Australia.

A clinic database was examined for all newly diagnosed treatment-naïve HIV-1 patients and information on their HIV-1 resistance and subtype, demographics including country of birth and diagnosis of a bacterial sexually transmissible infection was collected. Data were available from 74/79 individuals (62 cis-male, 16 cis-female and 1 transgender woman). Of the 74 genotypes, the prevalence of non-B subtypes and TDR was 43/74 (58%; 95%CI = 46.9-69.3) and 14/74 (19%; 95%CI = 10.0 to 27.8). It was also found that 30/79 (38%) had a concurrent bacterial STI. TDR was associated with subtype B infection (OR 3.53; 95%CI = 1.41-8.82; P = 0.007) and being born in Australia (OR 12.0; 95%CI = 2.45-58.86; P = 0.002). The relative prevalence of non-B HIV-1 subtypes and TDR is higher in Western Sydney than in the rest of Australia. TDR is associated with subtype B HIV-1 and being Australian born, suggesting ongoing local transmission. This highlights the diversity of the HIV epidemic locally and the need for interventions to prevent ongoing HIV transmission.

Molecular epidemiology of HIV-1 infection in immigrant population in northern Italy.

Human immunodeficiency virus-1 (HIV-1) is characterised by a vast genetic diversity classified into distinct phylogenetic strains and recombinant forms. We describe the HIV-1 molecular epidemiology and evolution of 129 consecutive HIV-1 positive migrants living in Milan (northern Italy). Polymerase gene sequences of 116 HIV-1 subtype-B positive patients were aligned with HIV-1 reference sequences (https://www.ncbi.nlm.nih.gov/) by using MAFFT alignment and edited by using Bioedit software. A maximum likelihood (ML) phylogenetic tree was performed by MEGA7 and was visualised by using FigTree v1.4.3. Of 129 migrants, 35 were born in Europe (28 in Eastern Europe), 70 in the Americas (67 in South America), 15 in Africa and nine in Asia; 76.4% were men who have sex with men (MSM). The serotype HIV-1-B prevailed (89.9%), followed by -C, -F1, -D and -A. Compared with 116 HIV-B patients, the 13 with HIV-non-B showed lower Nadir of CD4+ cell/mmc (P = 0.043), more frequently had sub Saharan origin (38.5 vs. 1.72%, P = 0.0001) and less frequently were MSM (40 vs. 74.5%, P = 0.02). The ML phylogenetic tree of the 116 HIV-1 subtype-B positive patients showed 13 statistically supported nodes (bootstrap > 70%). Most of the sequences included in these nodes have been isolated from male patients from the Americas and the most common risk factor was MSM. The low number of HIV-1 non-B subtype patients did not allow to perform this analysis. These results suggest a shift of HIV-1 prevention projects' focus and a continuous monitoring of HIV-1 molecular epidemiology among entry populations. Prevention efforts based on HIV molecular epidemiology may improve public health surveillance setting.

Prevalence of Non-B HIV-1 Subtypes in North Italy and Analysis of Transmission Clusters Based on Sequence Data Analysis.

HIV-1 diversity is increasing in European countries due to immigration flows, as well as travels and human mobility, leading to the circulation of both new viral subtypes and new recombinant forms, with important implications for public health. We analyzed 710 HIV-1 sequences comprising protease and reverse-transcriptase (PR/RT) coding regions, sampled from 2011 to 2017, from naive patients in Spedali Civili Hospital, Brescia, Italy. Subtyping was performed by using a combination of different tools; the phylogenetic analysis with a structured coalescence model and Makarov Chain Monte Carlo was used on the datasets, to determine clusters and evolution. We detected 304 (43%) patients infected with HIV-1 non-B variants, of which only 293 sequences were available, with four pure subtypes and five recombinant forms; subtype F1 (17%) and CRF02_AG (51.1%) were most common. Twenty-five transmission clusters were identified, three of which included >10 patients, belonging to subtype CRF02_AG and subtype F. Most cases of alleged transmission were between heterosexual couples. Probably due to strong migratory flows, we have identified different subtypes with particular patterns of recombination or, as in the case of the subtype G (18/293, 6.1%), to a complete lack of relationship between the sequenced strains, revealing that they are all singletons. Continued HIV molecular surveillance is most important to analyze the dynamics of the boost of transmission clusters in order to implement public health interventions aimed at controlling the HIV epidemic.

Treatment outcome in dually HIV-1 and HIV-2 coinfected patients living in Spain.

Whereas HIV-1 has spread globally, HIV-2 is mainly found in West Africa where dual HIV-1/HIV-2 coinfection is nowadays uncommon. Herein, we report the rate, main characteristics, and treatment outcomes of all dually infected patients living in Spain. We identified retrospectively all persons coinfected with HIV-1 recorded at the Spanish HIV-2 registry. Dual infection had been confirmed using PCR in plasma and/or cells, and/or using discriminatory serological tests. From a total of 373 individuals with HIV-2 recorded at the Spanish registry, 34 (9.1%) were coinfected with HIV-1. Compared with HIV-2 monoinfected persons, dually infected patients were more often male (67.6%), presented with lower median CD4 cell counts (204 cells/μl), and had developed more frequently AIDS events (26.5%). Although 61.7% came from West Africa, 6 (17.6%) were native Spaniards. HIV-1 non-B subtypes were recognized in 75% of coinfected patients, being the most prevalent CRF02_AG. At baseline, 45% of dually infected patients had undetectable plasma HIV-2 RNA. After a median follow-up of 32 (13-48) months on antiretroviral therapy, dually infected patients achieved undetectable viremia in 85% for HIV-1, in 80% for HIV-2; and in 70% for both viruses. Median CD4 cell counts reached up to 418 cells/μl. Roughly 9% of individuals with HIV-2 infection living in Spain are coinfected with HIV-1. Overall, 70% of dually infected patients achieved viral suppression for both viruses under antiretroviral therapy. Given the relatively large population of West Africans living in Spain and the continuous migration flow from HIV-2 endemic areas, HIV-1/HIV-2 coinfection should always be excluded at first diagnosis in all HIV-seroreactive persons.

Transmitted drug resistance mutations and trends of HIV-1 subtypes in treatment-naïve patients: A single-centre experience.

Transmitted drug resistance (TDR) and HIV-1 genetic diversity may affect treatment efficacy and clinical outcomes. Here we describe the circulating viral subtypes and estimate the prevalence of drug resistance among antiretroviral therapy (ART)-naïve patients attending Sapienza University Hospital (Rome, Italy) from 2006-2017. Genotypic resistance testing (GRT) was performed on 668 ART-naïve patients for integrase (n = 52), protease and reverse transcriptase (n = 668) sequences. Twenty-one different HIV-1 subtypes and circulating recombinant forms (CRFs) were identified. Subtype B was the most common (67.1%), followed by CRF02_AG (8.4%), and subtypes C and F (both 6.0%). A significantly increase in the proportion of non-B strains (P < 0.001) and the rate of non-Italian patients was observed over time. The overall prevalence of TDR was 9.4% (NRTI, 4.2%; NNRTI, 5.8%; and PI, 1.0%) and was higher in subtype B strains. Transmitted INSTI mutations (Q148H and G140S) responsible for high-level resistance to raltegravir and elvitegravir and intermediate resistance to dolutegravir and bictegravir were found, for the first time, in two individuals. Minor or accessory INSTI mutations were detected in 17.3% of patients. No significant decrease in the prevalence of TDR was documented over time. The significant increase in non-B subtypes suggests that the molecular epidemiology of HIV-1 is changing. Detection of a major INSTI mutation in two ART-naïve patients highlights the importance of performing GRT before commencing treatment. This finding and the lack of a significant reduction in TDRs underline the importance of continuous surveillance of resistance mutations.

Rapid HIV Antigen-Antibody Assays and Detection of Acute HIV Infection in Sub-Saharan Africa.

Detection of acute HIV infection is a unique problem that fourth-generation HIV assays were expected to alleviate. In this commentary, we draw attention to the limitations and challenges with use of currently available rapid antigen-antibody (Ag/Ab) combination tests for detection of acute HIV infection in sub-Saharan Africa. Laboratory-based HIV-1 Ag/Ab immunoassays are complex, requiring specialized equipment and handling that are currently not affordable in many settings in Africa. The point-of-care Ag/Ab platform on the other hand is easier to deploy and potentially more accessible in resource-limited settings. However, available fourth-generation HIV-1 rapid diagnostic tests have demonstrated poor performance characteristics in field studies where non-B subtypes of HIV-1 dominate. The potential for point-of-care HIV-1 Ag/Ab diagnostics to significantly improve detection of acute HIV infection remains yet to be realized in sub-Saharan Africa. Assay platforms need to be optimized to identify local circulating subtypes, and optimal algorithms need to be determined.

A8 Epidemiological study of transmission clusters in a local HIV-1 cohort

Integration of molecular, clinical, and demographic data represents a powerful tool to understand the dynamics of local HIV-1 transmission chains (TCs). The aim of our study was the phylogenetic analysis of the TCs within a HIV-1 cohort and the description of the relevant patient data within a TC. We performed a phylogenetic analysis of 757 sequences from newly HIV-1 diagnosed patients in Málaga (Southern Spain) during the period 2004–15. We used partial pol gene sequences in a preliminary phylogenetic reconstruction using the Neighbour Joining method (MEGA v6.06 program). After eliminating branches with bootstrap values <80 per cent, we constructed a new phylogeny by Maximum likelihood method (FastTree program). We considered as TC any cluster with bootstrap values ≥ 90 per cent. Patient sequences within and outside TCs were compared. Resistance mutations in the protease (PR) and reverse transcriptase (RT) sequences were analyzed using the Stanford algorithm. Four hundred and fifty-one out of 757 patients (59.6%) were grouped into fifty-three TCs, seventeen of them with five or more subjects. The largest number of patients associated within a TC was ninety. Patients younger than 40 years [odds ratio (OR) 1.75, 95% confidence interval (95% CI) 1.2–2.4, P = 0.002], men who have sex with men (MSM) (OR 2.14, 95% CI 1.3–3.2, P < 0.0001), non-Spanish (OR 1.48, 95% CI 1.0–2.1, P = 0.038), with a non-B subtype HIV-1 (OR 3.12, 95% CI 2.0–4.8, P < 0.0001), and presenting primary resistance mutations (OR 14.1, 95% CI 3.1–62.6, P = 0.001) were more likely to be associated within a cluster. Ninety-four out of 118 patients (79.6%) with transmission resistance mutations were included in some TC. The most frequent mutations associated with clusters were T69D/N, L210W, and K219E/Q, for NRTIs, K103N, and G190A/S for NNRTIs, and the I54L/M and L90M mutations for PIs. The prevalence for resistance to NNRTIs in TCs was 13.7 per cent. There were two TCs of rarer non-B subtypes: CRF19_cpx, with twenty-one individuals, sixteen of them (76.2%) with mutation G190A; and CRF51_01B with thirty-nine patients, twenty of them with the K103N mutation. Approximately 60 per cent of newly HIV-1 diagnosed patients were included in a TC. Younger patients, MSM, non-Spanish, with non-B subtype HIV-1 and primary resistance mutations were more likely to belong to a cluster. NNRTI mutations were the most frequent ones among patients in TCs. We observed two TCs represented by infrequent non-B subtypes in our area—CRF19_cpx and CRF51_01B—both of which were associated to the transmission of primary resistance.