Sequence Variability in Clinical and Laboratory Isolates of Herpes Simplex Virus 1 Reveals New Mutations

Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.
Journal of Virology (Impact Factor: 4.44). 03/2010; 84(10):5303-13. DOI: 10.1128/JVI.00312-10
Source: PubMed


Herpes simplex virus 1 (HSV-1) is a well-adapted human pathogen that can invade the peripheral nervous system and persist there as a lifelong latent infection. Despite their ubiquity, only one natural isolate of HSV-1 (strain 17) has been sequenced. Using Illumina high-throughput sequencing of viral DNA, we obtained the genome sequences of both a laboratory strain (F) and a low-passage clinical isolate (H129). These data demonstrated the extent of interstrain variation across the entire genome of HSV-1 in both coding and noncoding regions. We found many amino acid differences distributed across the proteome of the new strain F sequence and the previously known strain 17, demonstrating the spectrum of variability among wild-type HSV-1 proteins. The clinical isolate, strain H129, displays a unique anterograde spread phenotype for which the causal mutations were completely unknown. We have defined the sequence differences in H129 and propose a number of potentially causal genes, including the neurovirulence protein ICP34.5 (RL1). Further studies will be required to demonstrate which change(s) is sufficient to recapitulate the spread defect of strain H129. Unexpectedly, these data also revealed a frameshift mutation in the UL13 kinase in our strain F isolate, demonstrating how deep genome sequencing can reveal the full complement of background mutations in any given strain, particularly those passaged or plaque purified in a laboratory setting. These data increase our knowledge of sequence variation in large DNA viruses and demonstrate the potential of deep sequencing to yield insight into DNA genome evolution and the variation among different pathogen isolates.

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Available from: Lynn Enquist, Oct 07, 2015
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    • "Illumina sequencing of this sample generated $ 6000-fold sequence coverage. Assembly of Illumina data using de novo assembly programs gave a number of blocks of sequence, or contigs, as observed for HSV-1 (Szpara et al., 2010). To refine the assembly and fill in large gaps, we mapped the sequence reads to the individual U L , U S , R L and R S regions containing 200 bp of overlapping sequence based on the HG52 reference sequence. "
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    ABSTRACT: Herpes simplex virus 2 is an important human pathogen as the causative agent of genital herpes, neonatal herpes, and increased risk of HIV acquisition and transmission. Nevertheless, the only genomic sequence that has been completed is the attenuated HSV-2 HG52 laboratory strain. In this study we defined the genomic sequence of the HSV-2 SD90e low passage clinical isolate and a plaque-purified derivative, SD90-3P. We found minimal sequence differences between SD90e and SD90-3P. However, in comparisons with the HSV-2 HG52 reference genome sequence, the SD90e genome ORFs contained numerous point mutations, 13 insertions/deletions (indels), and 9 short compensatory frameshifts. The indels were true sequence differences, but the compensatory frameshifts were likely sequence errors in the original HG52 sequence. Because HG52 virus is less virulent than other HSV-2 strains and may not be representative of wildtype HSV-2 strains, we propose that the HSV-2 SD90e genome serve as the new HSV-2 reference genome.
    Virology 02/2014; s 450–451:140–145. DOI:10.1016/j.virol.2013.12.014 · 3.32 Impact Factor
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    • "More accurate information on genetic relationships requires the use of whole or nearly complete genomes. Recently, next-generation sequencing techniques have been used to sequence several HSV-1 genomes [23], [24], [25] with more being directly deposited into GenBank. Currently complete, or nearly complete genomic sequences are available from North America, Europe, East Asia and Eastern Africa. "
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    ABSTRACT: We compared 31 complete and nearly complete globally derived HSV-1 genomic sequences using HSV-2 HG52 as an outgroup to investigate their phylogenetic relationships and look for evidence of recombination. The sequences were retrieved from NCBI and were then aligned using Clustal W. The generation of a maximum likelihood tree resulted in a six clade structure that corresponded with the timing and routes of past human migration. The East African derived viruses contained the greatest amount of genetic diversity and formed four of the six clades. The East Asian and European/North American derived viruses formed separate clades. HSV-1 strains E07, E22 and E03 were highly divergent and may each represent an individual clade. Possible recombination was analyzed by partitioning the alignment into 5 kb segments, performing individual phylogenetic analysis on each partition and generating a.phylogenetic network from the results. However most evidence for recombination spread at the base of the tree suggesting that recombination did not significantly disrupt the clade structure. Examination of previous estimates of HSV-1 mutation rates in conjunction with the phylogenetic data presented here, suggests that the substitution rate for HSV-1 is approximately 1.38×10(-7) subs/site/year. In conclusion, this study expands the previously described HSV-1 three clade phylogenetic structures to a minimum of six and shows that the clade structure also mirrors global human migrations. Given that HSV-1 has co-evolved with its host, sequencing HSV-1 isolated from various populations could serve as a surrogate biomarker to study human population structure and migration patterns.
    PLoS ONE 10/2013; 8(10):e76267. DOI:10.1371/journal.pone.0076267 · 3.23 Impact Factor
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    • "UL49A has two aa differences at residues 28 and 51 with histidine and threonine for McKrae, arginine and threonine for 17, and histidine and alanine for the other strains. Among all the strains, UL49A was also found to contain variability unique to F strain (Szpara et al., 2010). No UL49A was annotated in HF10 strain; therefore, BSR is not calculated when HF10 strain was included in the comparison. "
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    ABSTRACT: Ocular infection by HSV-1 strain McKrae is neurovirulent in both mice and rabbits and causes fatal encephalitis in approximately 50% of animals. In addition, it spontaneously reactivates with high frequency relative to other HSV-1 strains in rabbits. We sequenced the McKrae strain genome and compared its coding protein sequences with those of six other HSV-1 strains. Most of the 74 predicted protein sequences are conserved; only eleven are less than 98% conserved. Eight proteins were identified to be unique for McKrae based on sequence homology bit score ratio (BSR). These include five proteins showing significant variations (RL1, RS1, UL49A, US7 and US11), two truncated proteins (UL36 and UL56) and one (US10) containing an extended open reading frame. The McKrae strain also has unique features in its 'a' sequence and non-coding sequences, such as LAT and miRNA. These data are indicative of strain variation but need further work to connect observed differences with phenotype effects.
    Virology 09/2012; 433(2):528-37. DOI:10.1016/j.virol.2012.08.043 · 3.32 Impact Factor
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