Work

1224Defining herpes simplex virus type 2 sequence heterogeneity using next generation sequencing

Year: 2014

Type: article

Abstract: Background. Vaccines are most successful for pathogens in which wild-type infection leads to protection against homologous re-infection. For HSV-2, we have little information concerning natural protection from re-infection or the variability between strains that could modulate protection from challenge. Understanding differences in HSV-2 strains worldwide, and the potential for reinfection, is essential in development of an HSV-2 vaccine. Methods. DNA from genital HSV-2 lesions containing ≥7 log10 copies HSV DNA/swab were sequenced using the Illumina next generation sequencing (NGS) platform. Human and highly repetitive sequences were subtracted, and remaining reads were aligned to the reference HSV-2 genome. Prevalent SNPs were defined as present in >10% and <90% of the sequenced strains. Informative SNPs were identified using FastTagger. Bayesian computations were used to estimate probabilities of identifying super-infection. Results. Fifty-six samples from genital herpes lesions were sequenced. Of 49 sequences from 39 persons with adequate coverage, the HSV genome had a median of 73.9 fold coverage (range 9.8-771 fold). Sixteen persons (41%) were from Africa, 12 (31%) from Peru, and 11 (28%) from the US; one-third were HIV infected. Of 2854 loci with heterogeneity, prevalent SNPs were found at 456 (16%) loci. Using genotyping at the 6 most informative SNPs to distinguish strains, we estimate a 90% probability of correctly identifying super-infection or lack thereof. Of 10 persons with paired specimens collected a median of 2.1 years apart, 2 (20%) had super-infection; both were HIV-infected. In each case, the strains differed at 186 loci. Six of 10 remaining pairs differed at 0 loci, and 2 differed at only 1 locus, indicating minimal within host evolution and low sequencing error. Conclusion. High quality HSV-2 sequence data can be obtained directly from genital swabs, removing the need for culture which may introduce mutations. The initial phase of our study has identified super-infection in 20% of persons investigated. Further research will determine whether host immune status, gender, and sexual exposure history are correlates of super-infection. Disclosures. C. Johnston, AiCuris: Investigator, Research support; Genocea: Investigator, Research support; Agenus: Investigator, Research support D. M. Koelle, Sanofi Pasteur: Research Contractor, Research support A. Wald, Aicuris: Consultant, Consulting fee; Genocea: Investigator, Research support; Agenus: Investigator, Research support; Vical: Investigator, Research support

Source: Open Forum Infectious Diseases

DOI: 10.1093/ofid/ofu051.112