Divergence and recombination of clinical herpes simplex virus type 2 isolates

Abstract

Herpes simplex virus type 2 (HSV-2) infects the genital mucosa and is one of the most common sexually transmitted viruses. Here we sequenced a segment comprising 3.5% of the HSV-2 genome, including genes coding for glycoproteins G, I, and E, from 27 clinical isolates from Tanzania, 10 isolates from Norway, and 10 isolates from Sweden. The sequence variation was low compared to that described for clinical HSV-1 isolates, with an overall similarity of 99.6% between the two most distant HSV-2 isolates. Phylogenetic analysis revealed a divergence into at least two genogroups arbitrarily designated A and B, supported by high bootstrap values and evolutionarily separated at the root. Genogroup A contained isolates collected in Tanzania, and genogroup B contained isolates collected in Tanzania and Scandinavia, implying that the genetic variability of HSV-2 is higher in Tanzania than in Scandinavia. Recombination network analysis and bootscan analysis revealed a complex pattern of phylogenetically conflicting informative sites in the sequence alignments. These signals were present in synonymous and nonsynonymous sites in all three genes and were not accumulated in specific regions, observations arguing against positive selection. Since the PHI test applied solely to synonymous sites revealed a high statistical probability of recombination, we suggest as a novel finding that homologous recombination is, as reported earlier for HSV-1 and varicella-zoster virus, a prominent feature in the evolution of HSV-2.

FIG. 1.

Sequence analysis of the US4 gene for 27 Tanzanian (red), 10 Swedish (green), and 10 Norwegian (blue) clinical HSV-2 isolates and the two laboratory strains HG52 and B4327. (A) Recombination networks, including all sequences, were first constructed by using the SplitsTree program. (B) The isolates inferring phylogenetically conflicting signals (recombinant candidates) were removed, and new phylogenetic networks were constructed based solely on nonrecombinants. (C) Trees based on all isolates using the maximum-likelihood algorithm. The recombinant isolates are connected with dotted branches and the nonrecombinant isolates with solid branches. The bootstrap values are derived from a consensus tree based on 100 bootstrap replicates, including solely nonrecombinant isolates. Only bootstrap values above 60 are shown. (D) Recombinant candidates were analyzed by using the bootscan method implemented in the SimPlot program. Results from analyses of the US4 gene segment in two isolates are shown.

FIG. 2.

Sequence analysis of the US7-to-US8 segment. See the legend to Fig. 1 for details.

FIG. 3.

Recombination network based solely on silent mutations. The network includes all isolates and is based on the concatenated genes US4, US7, and US8. The Tanzanian, Swedish, and Norwegian isolates are shown in red, green, and blue, respectively.

FIG. 4.

Results from the bootscan analysis of three recombinant candidates based on concatenations of the US4 and US7-to US8 segments.