Elsevier

Virus Research

Volume 116, Issues 1–2, March 2006, Pages 146-158
Virus Research

Comparative analysis of genome sequences of three isolates of Orf virus reveals unexpected sequence variation

https://doi.org/10.1016/j.virusres.2005.09.011Get rights and content

Abstract

Orf virus (ORFV) is the type species of the Parapoxvirus genus. Here, we present the genomic sequence of the most well studied ORFV isolate, strain NZ2. The NZ2 genome is 138 kbp and contains 132 putative genes, 88 of which are present in all analyzed chordopoxviruses. Comparison of the NZ2 genome with the genomes of 2 other fully sequenced isolates of ORFV revealed that all 3 genomes carry each of the 132 genes, but there are substantial sequence variations between isolates in a significant number of genes, including 9 with inter-isolate amino acid sequence identity of only 38–79%. Each genome has an average of 64% G + C but each has a distinctive pattern of substantial deviation from the average within particular regions of the genome. The same pattern of variation was also seen in the genome of another parapoxvirus species and was clearly unlike the uniform patterns of G + C content seen in all other genera of chordopoxviruses. The availability of genomic sequences of three orf virus isolates allowed us to more accurately assess likely coding regions and thereby revise published data for 24 genes and to predict two previously unrecognized genes.

Introduction

Poxviruses are large DNA viruses, the most notorious of which is Variola virus (VARV), the causative agent of smallpox. The family is divided into entomopoxviruses and chordopoxviruses with the latter further subdivided into eight genera (Moyer et al., 2000). The genomes of the chordopoxviruses range in size from 135 to 365 kbp but show conservation of both genomic organization and content. The central regions of the genomes contain 88 genes which are present in all chordopoxviruses and which mostly occur in the same order and orientation (Delhon et al., 2004, Upton et al., 2003). In contrast, the terminal regions are variable in genetic content. Genes in these near-terminal regions of the genome are frequently not essential for growth in cultured cells but often encode factors with important roles in viral–host interactions including modulating host responses to infection and determining host range.

The established species in the genus Parapoxvirus are Bovine papular stomatitis virus (BPSV) and Pseudocowpoxvirus (PCPV) which are maintained in cattle, Parapoxvirus of red deer in New Zealand (PVNZ) and the type species, Orf virus (ORFV), which is maintained in sheep and goats (Mercer and Haig, 1999). BPSV, PCPV and ORFV have all been shown to infect humans. Infection of animals or humans by ORFV occurs via broken or scarified skin giving rise to pustular lesions. Viral replication is confined to the epidermis. In immune competent individuals, lesions resolve after a few weeks but severe progressive lesions can occur in immunosuppressed individuals. Despite an apparently normal host immune response to ORFV infection, the virus can repeatedly infect previously exposed animals, albeit with reductions in both the size of the lesions and the time to resolution (Haig and McInnes, 2002, Haig and Mercer, 1998).

The genomes of parapoxviruses are among the smallest of the chordopoxviruses and despite sharing the genome architecture typical of all chordopoxviruses, differences in G + C content, virion morphology and the presence of a substantial number of genes unique to parapoxviruses indicate a significant divergence from other poxvirus genera (Delhon et al., 2004, Mercer and Haig, 1999, Tikkanen et al., 2004). We present here the genome sequence of the NZ2 strain of ORFV, the most extensively studied strain of parapoxvirus.

Section snippets

Viruses

ORFV strain NZ2 was isolated in New Zealand from sheep scab material and plaque purified twice in primary bovine testis cells (Robinson et al., 1982). Sheep were then inoculated with the virus and the scabs that formed used as the source of viral DNA (Mercer et al., 1987). Other poxviruses used in sequence comparisons are listed in Table 1. These include ORFV strain SA00 isolated in Texas, USA, from a goat kid and propagated in Madin–Darby ovine kidney cells (Guo et al., 2003) and ORFV strain

DNA sequence of the ORFV NZ2 genome

ORFV strain NZ2 sequences were assembled into a contiguous sequence of 137,820 bp, which is in agreement with previous restriction enzyme-based size estimates of 139 kbp (Mercer et al., 1987). This assembly does not include the terminal hairpin loops but is bounded by near-terminal BamHI sites. “Snap-back” analysis of BamHI-digested NZ2 genomic DNA has revealed terminal hairpin loops of close to 0.1 kbp (Mercer et al., 1987) indicating a full genome size of 138.0 kbp. Using an approach that

Acknowledgments

This work was supported in part by the Health Research Council of New Zealand. We are grateful for the excellent technical assistance of Ellena Whelan and Catherine McCaughan.

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