Origin and spread of HIV-1 in persons who inject drugs in Bulgaria

Abstract

Increased HIV transmission in persons who inject drugs (PWIDs) has led to subepidemics and outbreaks in several countries in Europe, including Bulgaria. In this study in Bulgaria, we investigate the origin and spatiotemporal evolutionary history of HIV-1 infections in PWIDs and the distribution of antiretroviral resistance mutations and hepatitis co-infections in these populations. We analyzed HIV-1 polymerase sequences available from 117 of 359 PWIDs diagnosed with HIV/AIDS from 1999 to 2011. Of these, 50 (42.7%) were classified as CRF02_AG, 41 (35.0%) CRF01_AE, 12 (10.3%) URFs, ten (8.5%) subtype B, two (1.7%) subtype F1 and two (1.7%) CRF14_BG. Most recent common ancestor dating suggests that CRF01_AE was likely first introduced from Southeast Asia into persons reporting heterosexual infection in Bulgaria in 1992 and spread subsequently to PWIDs in the capital city of Sofia around 2003. Conversely, CRF02_AG in Bulgaria was likely first introduced into PWID from Germany in 2000 and later entered heterosexual populations around 2009. The overall prevalence of resistance mutations was 6.8% (8/117), of which 5.1% (5/117) was observed in patients on antiretroviral therapy and 1.7% (2/117) was from transmitted drug resistance mutations in drug-naïve individuals. 189/204 (92.6%) PWIDs were also co-infected with hepatitis C (HCV) and 31/183 (16.9%) were co-infected with hepatitis B (HBV). Our study provides valuable molecular epidemiological information on the introduction and distribution of the main HIV-1 subtypes, resistance mutations and hepatitis co-infections among PWIDs with HIV-1 in Bulgaria which can be used to target prevention efforts.

Keywords: Drug resistance; HIV; Hepatitis; Injection drug use; Molecular epidemiology; Subtype.

Fig. 1.

Geographic distribution of HIV-1 subtypes among persons who inject drugs (PWIDs) in Bulgaria. Pie charts show subtype distribution within Sofia and Plovdiv. Numbers of each subtype are in parentheses for the smaller towns. Cities where HIV-1-infected PWIDs reported living and that were included in our study are shown in red text.

Fig. 2.

Global maximum likelihood phylogenetic tree of CRF01_AE. The phylogenetic analysis is based on an alignment of 721 HIV-1 CRF01_AE polymerase (pol) nucleotides inferred with 97 pol sequences from Bulgaria, 41 from persons who inject drugs (PWIDs) and 56 heterosexual (HET) individuals and 4680 pol sequences of the respective CRF available in GenBank using FastTree. Confidence values of tree branches and clusters were assessed by using the Shimodaira-Hasegawa test and are given as probabilities. Black and blue branches indicate globally distributed and Bulgarian pol sequences, respectively. Solid and dashed purple arrows indicate positions of Bulgarian PWID and HET only pol sequences in the tree, respectively.

Fig 3.

Global maximum likelihood phylogenetic tree of CRF02_AG. The phylogenetic analysis is based on an alignment of 774 HIV-1 CRF02_AG polymerase (pol) nucleotidesmferred with 69 pol sequences from Bulgaria, 50 from persons who inject drugs (PWIDs) and 19 heterosexual (HET) individuals and 2933 pol sequences of the respective CRF available in GenBank using FastTree. Confidence values of tree branches and clusters were assessed by using the Shimodaira-Hasegawa test and are given as probabilities. Black and blue branches indicate globally distributed and Bulgarian pol sequences, respectively. Solid and dashed purple arrows indicate positions of Bulgarian PWID and HET only pol sequences in the tree, respectively.

Fig. 4.

Phylodynamics and phylogeography of HIV-1 CRF01_AE in Bulgaria inferred by Bayesian analysis. (A) Bayesian skyride plot showing the inferred growth of the CRF01_AE epidemic in Bulgaria. Inferred effective population sizes (Ne) over time in years are on the y- and x-axes, respectively. (B) Phylogenetic tree showing the most recent common ancestor and geographic spread of CRF01_AE. Analysis included 70 polymerase (pol) sequences from Bulgaria, 41 sequences from persons who inject drugs (PWIDs) and 29 heterosexual (HET) individuals and 56 pol sequences from other countries. Filled circles and diamonds at branch tips indicate Bulgarian HETs and PWIDs, respectively. Time scale in years provided on x-axis and country or city origin of the sequences provided in color and defined in key.

Fig. 5.

Phylodynamics and phylogeography of HIV-1 CRF02_AG in Bulgaria inferred by Bayesian analysis. (A) Bayesian skyride plot showing the inferred growth of the CRF02_AG epidemic in Bulgaria. Inferred effective population sizes (Ne) over time in years are on the y- and x-axes, respectively. (B) Phylogenetic tree showing the most recent common ancestor and geographic spread of CRF02_AG. Analysis included 69 polymerase (pol) sequences from Bulgaria, 50 sequences from persons who inject drugs (PWIDs), 19 heterosexual (HET) individuals and 22 pol sequences from other countries. Filled circles and diamonds at branch tips indicate Bulgarian HET and PWIDs, respectively. Time scale in years provided on x-axis and country or city origin of the sequences provided in color and defined in key.