Analysis of the Origin and Dissemination of HIV-1 Subtype C in Bulgaria

Abstract

HIV-1 subtype C is the most abundant strain of HIV-1 infections worldwide and was found in the first known patients diagnosed with HIV/AIDS in Bulgaria in 1986. However, there is limited information on the molecular-epidemiological characteristics of this strain in the epidemic of the country. In this study, we analyze the evolutionary history of the introduction and dissemination of HIV-1 subtype C in Bulgaria using global phylogenetic analysis, Bayesian coalescent-based approach, and molecular clock methods. All available samples with HIV-1 subtype C from individuals diagnosed with HIV/AIDS between 1986 and 2017 were analyzed. Men and women were equally represented, and 24.3% of patients reported being infected abroad. The global phylogenetic analysis indicated multiple introductions of HIV-1 subtype C from various countries of the world. The reconstruction of a Bayesian time-scaled phylogenies showed that several Bulgarian strains segregated together in clusters, while others were intermixed in larger clades containing strains isolated from both European and non-European countries. The time-scale of HIV-1 subtype C introductions in Bulgaria demonstrates the early introduction of these viruses in the country. Our in-depth phylogenetic and phylogeographic analyses are compatible with a scenario of multiple early introductions in the country followed by limited local distribution in the subsequent years. HIV-1 subtype C was introduced in the early years of the epidemic, originating from different countries of the world. Due to the comprehensive measures for prevention and control in the early years of the epidemic in Bulgaria, HIV-1 subtype C was not widely disseminated among the general population of the country.

Keywords: Bulgaria; HIV; molecular epidemiology; subtype C.

Figure 1

Distribution of HIV-1 subtype C in Bulgaria. (A) Number of individuals with HIV-1 subtype C by year of diagnosis. (B) Distribution of HIV-1 subtype C in different regions of the country. Population % represents the percentage of the population in the respective district of the country.

Figure 2

Global maximum likelihood phylogenetic tree of HIV-1 subtype C in Bulgaria. The tree reconstruction was inferred using Dataset-1 containing 37 Bulgarian sequences and 1731 specially selected HIV-1 subtype C sequences from the global search. A total of 1532 sequences originated from Africa, 98 from Europe, 67 from Asia, and 34 from North America and South America. The red dots on the circle indicate the position of the Bulgarian sequences on the phylogenetic tree. The colors of the branches of the phylogenetic tree correspond to the color chart on the relevant continent.

Figure 3

Bayesian molecular clock analysis of HIV-1 subtype C in Bulgaria. Bayesian maximum clade credibility tree and the time of the most recent common ancestor (tMRCA) estimation. The tree reconstruction was inferred using Dataset-2 consisting of 300 sequences, 37 of which were Bulgarian, and 263 were selected from the global search. Clades with posterior probabilities ≥90% are indicated by black dots. Red dots at the end of the red lines indicate Bulgarian sequences. Scale years are reported at the bottom of the figure. Some branches are collapsed for better visualization.

Figure 4

Statistically significant clusters with posterior probability >90% in the Bayesian maximum clade credibility tree. (A) Clusters involving sequences isolated in Bulgaria and (B) mixed clusters consisting of Bulgarian and non-Bulgarian sequences. Sequences from Bulgaria (BG) are represented in red color, South Africa (ZA) pink, Romania (RO) light blue, Zambia (ZM) orange, and Sweden (SE) dark blue.

Figure 5

The introduction and dissemination of HIV-1 subtype C in Bulgaria. The blue arrows represent the introduction of strains from abroad into Bulgaria, and the red arrows represent the spreading of strains within the country. Country codes: ZM, Zambia; ZA, South Africa; TZ, Tanzania; SE, Sweden; MW, Malawi; BR, Brazil; RO, Romania. n, the number of the introduced strains in Bulgaria.

Figure 6

Bayesian skyline plot showing the inferred growth of the HIV-1 subtype C. Inferred effective population sizes (Ne) over time in years are on the y- and x-axes, respectively.