Two draft Canadian monkeypox virus (MPXV) genomes, June 2022

Matthew Croxen1,2,3, Ashwin Deo1, Paul Dieu4, Xiaoli Dong4, Christina Ferrato4, Kara Gill4, David Granger4, Vanipriyadarsini Ikkurti4, Jamil Kanji4,5, Petya Koleva1,6, Vincent Li4, Colin Lloyd1,6, Tarah Lynch4,7, Raymond Ma4, Kanti Pabbaraju4, Silas Rotich1, Hilary Sergeant1, Steven Shideler4, Todd Skitsko1, Sandy Shokoples1, Graham Tipples1,2,3, Johanna Thayer4,6, Anita Wong4

  1. Alberta Precision Laboratories, Public Health Laboratory, Edmonton, Alberta, Canada
  2. Department of Laboratory Medicine and Pathology, University of Alberta, Edmonton, Alberta, Canada
  3. Li Ka Shing Institute for Virology, University of Alberta, Edmonton, Canada
  4. Alberta Precision Laboratories, Public Health Laboratory, Calgary, Alberta, Canada
  5. Department of Medicine, University of Calgary, Calgary, Alberta, Canada
  6. National Microbiology Laboratory, Public Health Agency of Canada
  7. Department of Pathology and Laboratory Medicine, University of Calgary, Calgary, Alberta, Canada

Cases of monkeypox (MPXV) have been identified in Canada as part of an on-going international outbreak. As of June 17, 2022 (10:00 am EDT), there have been 168 confirmed MPXV cases reported by the Public Health Agency of Canada (PHAC) (1), 4 of which were identified in the province of Alberta. Here we report on two MPXV genomes sequenced at the Alberta Precision Laboratories, Public Health Laboratory. Both genomes are available at NCBI (ON736420.2, ON754989.2), and GISAID (EPI_ISL_13194516, EPI_ISL_13269478). These genomes will help contribute to the global epidemiology of the MPXV clade 3 outbreak.

All aerosol generating procedures such as specimen manipulation were performed in a Class II A2 biosafety cabinet with enhanced personal protective equipment. The patient specimen was inoculated into the Nuclisens easyMAG Lysis Buffer (BioMerieux) and incubated for one hour for complete viral inactivation. Viral DNA was extracted from the inactivated specimen using the easyMAG (BioMerieux), and prepared as an Illumina-compatible library using Illumina DNA Prep. Each sample, along with a negative control, was sequenced on an Illumina MiniSeq using a 300 cycle Mid Output Reagent Kit. Raw sequences were adapter-trimmed and quality-filtered using fastp 0.23.2 (2). Non-MPXV reads were filtered using ReadItAndKeep 0.30.0 (3). The dehosted and quality filtered reads were mapped to MPXV_USA_2022_MA001 (NCBI Accession: ON563414.3) using bwa mem 0.7.17-r1188 (4), sorted and filtered with samtools 1.15 (5), and iVar 1.3.1 (6) was used to generate a consensus genome with options “-t 0.75 -m 1”.

Alignment of our consensus genomes with select clade 3 genomes from mpoxSPECTRUM (7) was performed using MAFFT 7.490 (8), and a maximum-likelihood tree was generated with IQ-TREE v2.2.0.3 (9) using 1000 ultrafast bootstraps (10), 1000 SH-like approximate likelihood ratio test (SH-aLRT) (11), and ModelFinder (12). The best fitting model was HKY+F+I. The final tree was visualized with FigTree 1.4.3 (13). This placed our two genomes as lineage B.1 within the international outbreak clade 3 (14) (Figure 1).

Figure 1. Maximum likelihood tree of 37 MPXV genomes, including two MPXV genomes sequenced from Alberta, Canada. Both Alberta genomes (ON736420.2 and ON754989.2) are indicated in red and cluster within the MPXV clade 3 as the current outbreak lineage, B.1 (14). MPXV isolates from 2021 are highlighted in blue.

Comparison of single nucleotide variants (SNVs) relative to MPXV_USA_2022_MA001 show that the first genome (ON736420.2; EPI_ISL_13194516) differs by 1 nucleotide, while the second genome has 8 nucleotide differences (ON754989.2; EPI_ISL_13269478) (Figure 2).

Figure 2. Comparison of single nucleotide variants (SNV) between two MPXV genomes sequenced from Alberta, Canada (ON736420.2 and ON754989.2) and MPXV_USA_2022_MA001 (ON563414.3). Visualization of SNVs generated with snipit (15)


  1. (last accessed June 21, 2022)
  2. Chen S, Zhou Y, Chen Y, Gu J. fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics. 2018 Sep 1;34(17):i884-i890. doi: 10.1093/bioinformatics/bty560. PMID: 30423086; PMCID: PMC6129281.
  3. Hunt M, Swann J, Constantinides B, Fowler PW, Iqbal Z. ReadItAndKeep: rapid decontamination of SARS-CoV-2 sequencing reads. Bioinformatics. 2022 May 13:btac311. doi: 10.1093/bioinformatics/btac311. Epub ahead of print. PMID: 35551365.
  4. Ki H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. 2013. arXiv:1303.3997v2. doi: 10.48550/arXiv.1303.3997
  5. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R; 1000 Genome Project Data Processing Subgroup. The Sequence Alignment/Map format and SAMtools. Bioinformatics. 2009 Aug 15;25(16):2078-9. doi: 10.1093/bioinformatics/btp352. Epub 2009 Jun 8. PMID: 19505943; PMCID: PMC2723002.
  6. Grubaugh ND, Gangavarapu K, Quick J, Matteson NL, De Jesus JG, Main BJ, Tan AL, Paul LM, Brackney DE, Grewal S, Gurfield N, Van Rompay KKA, Isern S, Michael SF, Coffey LL, Loman NJ, Andersen KG. An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar. Genome Biol. 2019 Jan 8;20(1):8. doi: 10.1186/s13059-018-1618-7. PMID: 30621750; PMCID: PMC6325816.
  8. Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013 Apr;30(4):772-80. doi: 10.1093/molbev/mst010. Epub 2013 Jan 16. PMID: 23329690; PMCID: PMC3603318.
  9. Nguyen LT, Schmidt HA, von Haeseler A, Minh BQ. IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol Biol Evol. 2015 Jan;32(1):268-74. doi: 10.1093/molbev/msu300. Epub 2014 Nov 3. PMID: 25371430; PMCID: PMC4271533.
  10. Minh BQ, Nguyen MA, von Haeseler A. Ultrafast approximation for phylogenetic bootstrap. Mol Biol Evol. 2013 May;30(5):1188-95. doi: 10.1093/molbev/mst024. Epub 2013 Feb 15. PMID: 23418397; PMCID: PMC3670741.
  11. Guindon S, Dufayard JF, Lefort V, Anisimova M, Hordijk W, Gascuel O. New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst Biol. 2010 May;59(3):307-21. doi: 10.1093/sysbio/syq010. Epub 2010 Mar 29. PMID: 20525638.
  12. Kalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nat Methods. 2017 Jun;14(6):587-589. doi: 10.1038/nmeth.4285. Epub 2017 May 8. PMID: 28481363; PMCID: PMC5453245.
  13. GitHub - rambaut/figtree: Automatically exported from
  14. Happi C, Adetifa I, Mbala P, Njouom R, Nakoune E, Happi A, Ndodo N, Ayansola O, Mboowa G, Bedford T, Neher RA, Roemer C, Hodcroft E, Tegally H, O’Toole Á, Rambaut A, Pybus O, Kraemer MUG, Wilkinson E, Isidro J, Borges V, Pinto M, Paulo Gomes J, Baxter C, Lessells R, Ogwell AE, Kebede T, Tessema SK, de Oliveira T. Urgent need for a non-discriminatory and non-stigmatizing nomenclature for monkeypox virus (Urgent need for a non-discriminatory and non-stigmatizing nomenclature for monkeypox virus)
  15. GitHub - aineniamh/snipit: snipit: summarise snps relative to your reference sequence

We want to thank the entire Alberta Precision Laboratories Public Health Laboratory staff for their tireless efforts in confirming and sequencing these MPXV genomes.

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