O1. Near full-length HIV-1 genome sequencing reveals higher intra-country spread of unique recombinant forms (URFs) among patients infected in Sweden

Ujjwal Neogi1, Abu B. Siddik1, Mohammed M. Morshed1, Anders Sönnerborg1,
Affiliates: 1Karolinska Institutet

Background
The Swedish HIV-1 epidemic represents one of the most diverse epidemics with increase of non-B subtypes due to human migrations. Based on the subtyping analysis of the pol gene, we hypothesized that this viral heterogeneity may become the hotspot for development of more complex and unique recombinant forms (URFs) if the epidemics converge. As the smaller gene fragments (e.g. pol) underrepresent the recombinant forms, we performed the near full-length HIV-1 genome sequencing (HIV-NFLG) to characterize the epidemic.

Method
HIV-NFLG sequencing (HXB2: 790 to 9454) was perform amplifying the genome in two fragments followed by sequencing either by Sanger Sequencing with 16 primers or by next generation sequencing in Illumina HiSeq2500 followed by de novo assembly in Iterative Virus Assembler (IVA). Subtyping was performed in four automated tools (RIPv3, REGA v3, COMET-HIV and jpHMM) followed by maximum likelihood phylogenetic analysis in RAxML with the general time reversible model with inverse gamma rates (GTR+I+G). Self-reported country of infection was considered as origin of the infection.

Results
Total 104 HIV-NFLG was successful (71%) attempting 147 samples. Among the 104 samples 43% (45/104) were infected in Sweden while rest (57%) outside the country, according to self-reports, that represents 22 countries. Subtyping identified presence of HIV-1C (50%), B (21%), A1 (5%), D (1%) and CRFs (01_AE, 02_AG, 63_02A1, 11_cpx) (12%) (Fig 1A). URFs identified in 10% (11/104) of the samples with A1C (n=4), A1D (n=3), BF1, 01B, CF1, CB (n=1 each) (Fig 1A). Higher proportion of URFs was identified among the patients infected in Sweden (18% vs 5%; p=0.05) (Fig 1B). Among the 45 patients infected in Sweden with URFs, all were observed in patients who infected recently (after 2010; p=0.01) (Fig 1C). One large cluster of eight HIV-1B strains from patients infected in Sweden was observed with high bootstrap support (>95%). Apart from that the URF A1D has also spread in three individuals most probably from the same source within the country.

Conclusions
Our study indicates a high proportion of URFs, identified by HIV-NFLG, that is spreading among the HIV-1 infected patients in the country. This indicates an intermixing of strains within the country, evolving the more complex recombinant forms which are spread within the country. Biological (replication capacity and pathogenicity) and clinical (in vivo and in vitro) characterization are presently ongoing to understand the effect of the newly emerging recombinants at individual patient level as well as in the population level.

O1-Ujjwal