Genomic Sequencing and Analysis of a Novel Human Cowpox Virus With Mosaic Sequences From North America and Old World Orthopoxvirus

Abstract

Orthopoxviruses (OPXVs) not only infect their natural hosts, but some OPXVs can also cause disease in humans. Previously, we partially characterized an OPXV isolated from an 18-year-old male living in Northern Norway. Restriction enzyme analysis and partial genome sequencing characterized this virus as an atypical cowpox virus (CPXV), which we named CPXV-No-H2. In this study, we determined the complete genome sequence of CPXV-No-H2 using Illumina and Nanopore sequencing. Our results showed that the whole CPXV-No-H2 genome is 220,276 base pairs (bp) in length, with inverted terminal repeat regions of approximately 7 kbp, containing 217 predicted genes. Seventeen predicted CPXV-No-H2 proteins were most similar to OPXV proteins from the Old World, including Ectromelia virus (ECTV) and Vaccinia virus, and North America, Alaskapox virus (AKPV). CPXV-No-H2 has a mosaic genome with genes most similar to other OPXV genes, and seven potential recombination events were identified. The phylogenetic analysis showed that CPXV-No-H2 formed a separate clade with the German CPXV isolates CPXV_GerMygEK938_17 and CPXV_Ger2010_MKY, sharing 96.4 and 96.3% nucleotide identity, respectively, and this clade clustered closely with the ECTV-OPXV Abatino clade. CPXV-No-H2 is a mosaic virus that may have arisen out of several recombination events between OPXVs, and its phylogenetic clustering suggests that ECTV-Abatino-like cowpox viruses form a distinct, new clade of cowpox viruses.

Keywords: Fennoscandian; Norway; phylogenetics; poxvirus; recombination.

FIGURE 1

Genome map of CPXV-No-H2. Localization of 217 predicted coding sequences (CDS) and nine putative recombination events in the CPXV-No-H2 genome. Green blocks represent the putative recombination events. Other colors were used to visualize the amino acid sequence similarity between translated CDS to other OPXV proteins: blue blocks represent CDS with a higher similarity to Alaskapox virus proteins, yellow blocks represent CDS with a higher similarity to Ectromelia virus (ECTV) proteins, orange blocks represent CDS with a higher similarity to Vaccinia virus (VACV) proteins, and fuchsia block represents the CDS with a higher similarity to ECTV, VACV, and Horsepox virus proteins.

FIGURE 2

Bayesian inference phylogenetic tree based on 76 orthopoxvirus whole genomes. Posterior probabilities are shown on the right side of each node, and only posterior probabilities above 0.6 are shown. The cowpox virus strains were grouped into different clades: CPXV-like 1, CPXV-like 2, and VARV-like (Franke et al., 2017). The scale bar represents the expected substitutions per site.

FIGURE 3

Bayesian inference phylogenetic tree based on 76 orthopoxvirus core genomes. Posterior probabilities are shown on the right side of each node, and only posterior probabilities above 0.6 are shown. The cowpox virus (CPXV) strains were grouped into different clades: CPXV-like 1, CPXV-like 2, and VARV-like (Franke et al., 2017). The scale bar represents the expected substitutions per site.

FIGURE 4

Bayesian inference phylogenetic tree based on 134 orthopoxvirus orthologous genes. Posterior probabilities are shown on the right side of each node, and only posterior probabilities above 0.6 are shown. The cowpox virus (CPXV) strains were grouped into different clades: CPXV-like 1, CPXV-like 2, and VARV-like (Franke et al., 2017). The scale bar represents the expected substitutions per site.