Genetic and phenotypic analysis of novel South African Avian poxviruses

Avian poxviruses are important pathogens of both wild and domestic birds and exhibit a large degree of intragenus diversity at a genomic level. These viruses are known to differ in growth characteristics (in vitro and in vivo), virulence, and cross-protection, with little known about the genomic contributions to these differences. Only six isolates from subclades A and B and one from proposed subclade E have had their genomes completely sequenced. These genomes have been shown to exhibit typical poxvirus genome characteristics with conserved central regions and more variable terminal regions, however all isolates exhibit major differences in defined central regions. This study aimed to analyze and characterize novel isolates from South Africa in terms of growth characteristics and phylogenetic relationships. It also added to the pool of genome sequences available for comparative genomic analyses to further investigate genome architecture. Poxvirus isolates from lesser flamingo (Phoenicopterus minor) and African penguin (Spheniscus demersus) were chosen for analysis from a larger pool of donated isolates by comparison of macroscopic growth characteristics on chorioallantoic membranes, membrane histology and phylogenetic analyses based on nucleotide alignment of partial P4b sequences. Flamingopox virus was shown to group in subclade A3, induce membrane thickening and mesodermal hyperplasia while Penguinpox virus grouped in subclade A2, and did not induce membrane thickening or hyperplasia. The genomes of the above isolates were sequenced and compared to other available avipoxvirus genomes. Dotplot comparisons revealed major differences in central regions that have traditionally been thought to be conserved. Further analysis revealed five regions of difference, of varying lengths, spread across the central regions of the various genomes. Although individual gene identities at the nucleotide level did not vary greatly, gene content and synteny between isolates/species at these identified regions were far more divergent than expected. The reasons for these large genomic rearrangements are yet to be elucidated and will need to be considered in future phylogenetic studies and vaccine vector design. Sequencing and analysis of further avian poxvirus genomes will help characterize this complex genus of poxviruses

Phylogenetic analysis of three genes of Penguinpox virus corresponding to Vaccinia virus G8R (VLTF-1), A3L (P4b) and H3L reveals that it is most closely related to Turkeypox virus, Ostrichpox virus and Pigeonpox virus

Phylogenetic analysis of three genes of Penguinpox virus, a novel Avipoxvirus isolated from African penguins, reveals its relationship to other poxviruses. The genes corresponding to Vaccinia virus G8R (VLTF-1), A3L (P4b) and H3L were sequenced and phylogenetic trees (Neighbour-Joining and UPGMA) constructed from MUSCLE nucleotide and amino acid alignments of the equivalent sequences from several different poxviruses. Based on this analysis, PEPV was confirmed to belong to the genus Avipoxvirus, specifically, clade A, subclade A2 and to be most closely related to Turkeypox virus (TKPV), Ostrichpox virus (OSPV)and Pigeonpox virus (PGPV)

Six host-range restricted poxviruses from three genera induce distinct gene expression profiles in an in vivo mouse model

BACKGROUND: Host-range restricted poxviruses make promising vaccine vectors due to their safety profile and immunogenicity. An understanding of the host innate immune responses produced by different poxvirus vectors would aid in the assessment, selection and rational design of improved vaccines for human and veterinary applications. Novel avipoxviruses are being assessed to determine if they are different from other poxvirus vectors. Analysis of the transcriptome induced in a mouse model would aid in determining if there were significant differences between different poxvirus vectors which may reflect different adjuvant potential as well as establish if they should be further evaluated as vaccine vectors. RESULTS: We compared host transcript abundance in the spleens of BALB/c mice twenty four hours after intravenous infection (10 5 pfu/mouse) with six host-restricted poxvirus species from three genera, namely Lumpy Skin Disease virus (LSDV), Canarypox virus (CNPV), Fowlpox virus (FWPV), modified vaccinia Ankara (MVA) and two novel South African avipoxviruses, Feral Pigeonpox virus (FeP2) and Penguinpox virus (PEPV). These six viruses produced qualitatively and quantitatively distinct host responses with LSDV, followed by MVA, inducing the greatest interferon (IFN) response. FeP2 and PEPV caused very little change to host transcript abundance compared to the other 4 viruses tested. CNPV and FWPV induced the up regulation of two immunoglobulin genes (Ighg and Ighg3 (IgG3)) with CNPV inducing a third, Ighm (IgM). HIV-1-specific IgG3 antibodies have been correlated with decreased risk of HIV-1 infection in the RV144 trial, which included a CNPV-based vector (Yates et al. (Sci Transl Med, 6(228) p228, 2014). Up regulation of IgG3 by CNPV and FWPV but not the other poxviruses tested in vivo, implies that these two avipoxvirus-vector backbones may be involved in stimulation of the clinically important IgG3 antibody subclass. Differential transcript abundance associated with the different poxviruses is further discussed with particular emphasis on responses related to immune responses. CONCLUSION: Six, genetically diverse host-restricted poxviruses produce different responses in a mouse model early after infection. These differences may affect the immune response induced to vaccine antigen in vectors based on these viruses. The two novel avipoxviruses were clearly distinguishable from the other viruses

Phylogenetic Analysis of South African Bovine Leukaemia Virus (BLV) Isolates

Bovine leukaemia virus (BLV) causes chronic lymphoproliferative disorder and fatal lymphosarcoma in cattle, leading to significant economic losses in the beef and dairy industries. BLV is endemic globally and eleven genotypes have been identified. To date, only Zambian isolates have been genotyped from Africa. Although high BLV prevalence has been reported in South Africa, there has been no molecular characterisation of South African BLV isolates. To characterise BLV isolates in South Africa for the first time, we investigated the phylogenetic relationships and compared the genetic variability of eight South African BLV isolates with BLV isolates representing the eleven known genotypes from different geographical regions worldwide. Phylogenetic analyses based on full-length and partial env sequences as well as full-length gag sequences revealed that at least two genotypes, genotypes 1 (G1) and 4 (G4), are present in cattle in South Africa, which is consistent with studies from Zambia. However, our analysis revealed that the G1 South African isolate is more similar to other G1 isolates than the G1 Zambian isolates whereas, the G4 South African isolates are more divergent from other G4 isolates but closely related to the G4 Zambian isolate. Lastly, amino acid sequence alignment identified genotype-specific as well as novel amino acid substitutions in the South African isolates. The detection of two genotypes (G1 and G4) in southern Africa highlights the urgent need for disease management and the development of an efficacious vaccine against local strains