Herpes simplex virus serotype 2 (HSV-2) is a ubiquitous human pathogen that causes recurrent genital infections and ulcerations. Many HSV-2 strains with different biological properties have been identified, but only the genomes of HSV-2 strains HG52, SD90e and 333 have been reported as complete and fully characterized sequences. We de novo assembled, annotated and manually curated the complete genome sequence of HSV-2 strain MS, a highly neurovirulent strain, originally isolated from a multiple sclerosis patient. We resolved both DNA ends, as well as the complex inverted repeats regions present in HSV genomes, usually undisclosed in previous published partial herpesvirus genomes, using long reads from Pacific Biosciences (PacBio) technology. Additionally, we identified isomeric genomes by determining the alternative relative orientation of unique fragments in the genome of the sequenced viral population. Illumina short-read sequencing was crucial to examine genetic variability, such as nucleotide polymorphisms, insertion/deletions and sequence determinants of strain-specific virulence factors. We used Illumina data to fix two disrupted open reading frames found in coding homopolymers after PacBio assembly. These results support the combination of long- and short-read sequencing technologies as a precise and effective approach for the accurate de novo assembly and curation of complex microbial genomes.

中文翻译：

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长读长和短读长相结合完全解析单纯疱疹病毒2株MS完整基因组的复杂重复

单纯疱疹病毒血清型 2 (HSV-2) 是一种普遍存在的人类病原体，可导致复发性生殖器感染和溃疡。已鉴定出许多具有不同生物学特性的 HSV-2 菌株，但只有 HSV-2 菌株 HG52、SD90e 和 333 的基因组被报告为完整且完全表征的序列。我们从头组装、注释和手动管理 HSV-2 MS 毒株的完整基因组序列，这是一种高度神经毒力毒株，最初从一名多发性硬化症患者中分离出来。我们使用 Pacific Biosciences (PacBio) 技术的长读长解析了两个 DNA 末端，以及 HSV 基因组中存在的复杂反向重复区域，这些区域通常在之前发表的部分疱疹病毒基因组中未公开。此外，我们通过确定已测序病毒种群基因组中独特片段的替代相对方向来鉴定异构基因组。Illumina 短读长测序对于检查遗传变异性至关重要，例如核苷酸多态性、插入/缺失和菌株特异性毒力因子的序列决定因素。我们使用 Illumina 数据修复了 PacBio 组装后编码均聚物中发现的两个中断的开放阅读框。这些结果支持长读长和短读长测序技术的组合作为一种精确有效的方法复杂微生物基因组的从头组装和管理。