ABSTRACT
We report the coding-complete genome sequence of human alphaherpesvirus 1 (HHV1) isolated from a previously healthy 64-year-old male with fulminant hepatitis, a rare presentation of a common viral agent. The sequence is highly similar to previously described HHV1 sequences. Additional sequence data for fulminant hepatitis cases are required.
KEYWORDS: hepatitis, human alphaherpes virus 1, herpesviruses
ANNOUNCEMENT
Human alphaherpesvirus 1 (HHV1) belongs to the family Herpesviridae and is typically associated with mucocutaneous vesicular lesions, and disseminated disease is reported to be rare (1). HHV1 DNA was detected in the serum of a 64-year-old, previously healthy, male by qPCR confirming a diagnosis of fulminant herpetic hepatitis. Ethics approval was obtained from the Human Research Ethics Committee (Medical) at the University of the Witwatersrand (reference number: R14/49). Virus was isolated from serum (stored at −80°C) in Vero cells (ATCC CRL-1586), which were monitored for characteristic cytopathic effect (2). Cells were harvested after the second passage and concentrated from cleared supernatant using an Ultra-0.5 centrifugal filter with a 100-kDa cutoff (Amicon) in preparation for downstream processing for short- and long-read sequencing. For short-read sequencing, DNA was extracted using the Quick-DNA/RNA Viral Kit (Zymo Research), quantified using a Qubit fluorometer (Thermo Fisher Scientific), paired-end libraries prepared using the Nextera XT Library Prep kit (Illumina) for sequencing on the MiSeq (Illumina) platform (2 × 300 bp). For long-read sequencing, virus was purified through a 10% sucrose cushion, extracted using a genomic tip 20 /G (Qiagen), quantified using a Qubit fluorometer (Thermo Fisher Scientific), libraries prepared using the SMRTbell Express Template Prep kit 2.0 (Pacific BioSciences), DNA sheared using the Megaruptor 3 (Diagenode), and size selected using Ampure PB beads (Pacific BioSciences) for sequencing on the Sequel IIe system (Pacific BioSciences) using the Sequel II binding kit 2.2 and sequencing plate V2 (Pacific BioSciences).
FASTQ files were uploaded to the Galaxy Web platform and data analyzed using the server at http://usegalaxy.org (3). All tools were run using the default parameters, and short-read data were run for paired-end reads unless otherwise noted. Reads were quality trimmed (qualified quality phred, 20; length required for short reads, 40; length required for long reads, 100) using fastp V0.19.5 (4). For de novo assembly, reads were filtered against the reference sequence, assembled using MEGAHIT V1.2.9 (5) and evaluated using QUAST V5.2.0 (6). In addition, short and long reads were mapped on the reference sequence (GenBank accession number: NC001806.1 available at https://www.ncbi.nlm.nih.gov/nuccore/NC_001806) using Bowtie2 V2.4.5 (7) and Minimap2 V2.24 (8), respectively, and consensus sequence determined using iVar V1.3.1 (Minimum depth, 2) (9). Draft genomes obtained were aligned using MAFFT (10) and were in agreement. The most complete draft genome was obtained by long-read reference-based mapping, which was manually curated at ambiguous positions using Integrative Genomics Viewer V2.13.0 (11). The sequencing depth was determined using the SAMtools suite (12).
The genome is 152,063 bp in length (GC content 68.22%). Genome assembly metrics are provided in Table 1. The genome contains a 110 bp gap (position 117,349–117,458) in an inverted repeat region, which could not be resolved. Genome annotation was performed using NCBI ORF finder (13), DIAMOND (14), and the NCBI protein database (15). BLAST (16) analysis suggested that this sequence had a nucleotide identity of 98.75% with the reference sequence. Additional sequence data from HHV1 fulminant hepatitis cases are required to aid in studying potential virus-related factors that may play a role in the development of severe disease.
TABLE 1.
Genome assembly metrics for short- and long-read data
| Short-read data (MiSeq, Illumina) |
Long-read data (Sequel II, PacBio) |
|
|---|---|---|
| Total reads | 2.8 million | 14.1 thousand |
| Average read length | 224 bp | 3,675 bp |
| Genome coverage distribution for reference-based assembly | 151,379 bp | 152,159 bp |
| Average sequencing depth across genome | 485× | 12× |
| Genome coverage distribution with de novo assembly | 134,520 bp | 136,968 bp |
| Number of contigs for de novo assembly | 11 | 9 |
| N50 for de novo assembly | 44,644 bp | 45,163 bp |
ACKNOWLEDGMENTS
This work was supported by the National Research Foundation (NRF) of South Africa [116729 to N.V.], the NRF South African Research Chairs Initiative [U98346 to F.J.B], and operational funding at the NICD/NHLS. The authors acknowledge the technical support offered by the Core Sequencing Facility of the NICD/NHLS in performing MiSeq sequencing.
The funders had no role in the study design, data collection and analysis, decision to publish or the preparation of the manuscript.
Contributor Information
Jacqueline Weyer, Email: jacquelinew@nicd.ac.za.
Kenneth M. Stedman, Portland State University, Portland, Oregon, USA
DATA AVAILABILITY
This sequence has been deposited in GenBank under the accession number OQ102003.1 (https://www.ncbi.nlm.nih.gov/nuccore/OQ102003.1). The version described here is the first version. The raw reads were deposited in the NCBI Sequence Read Archive (SRA) under the accession number PRJNA913943 (https://www.ncbi.nlm.nih.gov/sra/SRX18774967 and https://www.ncbi.nlm.nih.gov/sra/SRX18774968).
REFERENCES
- 1. Norvell JP, Blei AT, Jovanovic BD, Levitsky J. 2007. Herpes simplex virus hepatitis: an analysis of the published literature and institutional cases. Liver Transpl 13:1428–1434. doi: 10.1002/lt.21250 [DOI] [PubMed] [Google Scholar]
- 2. World Health Organization . 2017. Use of cell culture in Virology for developing countries in the South-East Asia region [Google Scholar]
- 3. Schatz MC, Philippakis AA, Afgan E, Banks E, Carey VJ, Carroll RJ, Culotti A, Ellrott K, Goecks J, Grossman RL, Hall IM, Hansen KD, Lawson J, Leek JT, Luria AO, Mosher S, Morgan M, Nekrutenko A, O’Connor BD, Osborn K, Paten B, Patterson C, Tan FJ, Taylor CO, Vessio J, Waldron L, Wang T, Wuichet K. 2022. The Galaxy platform for accessible, reproducible and collaborative BIOMEDICAL analyses. Nucleic Acids Res 50 2:W345–W351. doi: 10.1016/j.xgen.2021.100085 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4. Chen S, Zhou Y, Chen Y, Gu J. 2018. Fastp: an ultra-fast all-in-one FASTQ Preprocessor. Bioinformatics 34:i884–i890. doi: 10.1093/bioinformatics/bty560 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5. Li D, Liu CM, Luo R, Sadakane K, Lam TW. 2015. MEGAHIT: an ultra-fast single-node solution for large and complex metagenomics assembly via succinct de Bruijn graph. Bioinformatics 31:1674–1676. doi: 10.1093/bioinformatics/btv033 [DOI] [PubMed] [Google Scholar]
- 6. Mikheenko A, Prjibelski A, Saveliev V, Antipov D, Gurevich A. 2018. Versatile genome assembly evaluation with QUAST-LG. Bioinformatics 34:i142–i150. doi: 10.1093/bioinformatics/bty266 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7. Langmead B, Salzberg SL. 2012. fast gapped-read alignment with Bowtie 2. Nat Methods 9:357–359. doi: 10.1038/nmeth.1923 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8. Li H. 2018. Minimap2: pairwise alignment for nucleotide sequences. Bioinformatics 34:3094–3100. doi: 10.1093/bioinformatics/bty191 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9. Castellano S, Cestari F, Faglioni G, Tenedini E, Marino M, Artuso L, Manfredini R, Luppi M, Trenti T, Tagliafico E. 2021. iVar, an interpretation-oriented tool to manage the update and revision of variant annotation and classification. Genes 12:384. doi: 10.3390/genes12030384 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10. Katoh K, Standley DM. 2013. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol 30:772–780. doi: 10.1093/molbev/mst010 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11. Thorvaldsdóttir H, Robinson JT, Mesirov JP. 2013. Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration. Brief Bioinform 14:178–192. doi: 10.1093/bib/bbs017 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12. Danecek P, Bonfield JK, Liddle J, Marshall J, Ohan V, Pollard MO, Whitwham A, Keane T, McCarthy SA, Davies RM, Li H. 2021. Twelve years of SAMtools and BCFtools. Gigascience 10:giab008. doi: 10.1093/gigascience/giab008 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13. Wheeler DL, Church DM, Federhen S, Lash AE, Madden TL, Pontius JU, Schuler GD, Schriml LM, Sequeira E, Tatusova TA, Wagner L. 2003. Database resources of the national center for biotechnology. Nucleic Acids Res 31:28–33. doi: 10.1093/nar/gkg033 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14. Buchfink B, Xie C, Huson DH. 2015. Fast and sensitive protein alignment using DIAMOND. Nat Methods 12:59–60. doi: 10.1038/nmeth.3176 [DOI] [PubMed] [Google Scholar]
- 15. Sayers EW, Bolton EE, Brister JR, Canese K, Chan J, Comeau DC, Connor R, Funk K, Kelly C, Kim S, Madej T, Marchler-Bauer A, Lanczycki C, Lathrop S, Lu Z, Thibaud-Nissen F, Murphy T, Phan L, Skripchenko Y, Tse T, Wang J, Williams R, Trawick BW, Pruitt KD, Sherry ST. 2022. Database resources of the national center for biotechnology information. Nucleic Acids Res 50:D20–D26. doi: 10.1093/nar/gkab1112 [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. 1990. J Mol Biolbasic local alignment search tool. J Mol Biol 215:403–410. doi: 10.1016/S0022-2836(05)80360-2 [DOI] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
This sequence has been deposited in GenBank under the accession number OQ102003.1 (https://www.ncbi.nlm.nih.gov/nuccore/OQ102003.1). The version described here is the first version. The raw reads were deposited in the NCBI Sequence Read Archive (SRA) under the accession number PRJNA913943 (https://www.ncbi.nlm.nih.gov/sra/SRX18774967 and https://www.ncbi.nlm.nih.gov/sra/SRX18774968).