The presence of transmission clusters (TCs) and their epidemiological characteristics in a treatment-naive cohort of HIV-1 patients in southern Spain over a decade (2004)(2005)(2006)(2007)(2008)(2009)(2010)(2011)(2012)(2013)(2014)(2015) were evaluated. Protease and reverse transcriptase sequences provided by each genotype test were used in the phylogenetic study, performed first by the neighbor-joining method and then confirmed by Bayesian analysis. We collected clinical, immunovirological, and demographic data for all patients included. Our cohort comprised 757 patients, 428 (56.5%) belonging to a TC. Overall, we found 123 TCs, 21 of them comprising five or more individuals and three with ‡10 sequences. Forty-three TCs (35.0%) remained active. The clustered patients were mainly men (92.8%) who had sex with men (MSM) (81.5%), Spanish (80.6%), and young adults (median age at diagnosis of 32.6 years).They had lower percentages of late diagnosis and AIDS cases (42.1% and 13.6%, respectively), whereas the presence of recent seroconverters (31.1%), HIV-1 B subtypes (79.4%), and transmission drug resistance (20.3%) increased within TCs, with regard to not-clustered individuals. Among the TCs of non-B variants, circulating recombinant forms (CRF) were predominant (87.5%), with the highest frequencies for CRF19_cpx (17.0% of non-B subtype sequences in TCs); CRF02_AG (15.9%); and CRF01_AE (9.1%). In conclusion, over half of our cohort was included within a TC. More than a third of TCs found could be considered active transmission events. Belonging to a TC was related to MSM, Spanish origin, recent seroconversion, high prevalence of resistance mutations, and B HIV subtype. Among the non-B genetic forms in TCs, we found a high prevalence of CRF19_cpx, CRF02_AG, and CRF01_AE variants.

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.