Characterization of a New Vaccinia virus Isolate Reveals the C23L Gene as a Putative Genetic Marker for Autochthonous Group 1 Brazilian Vaccinia virus

Abstract

Since 1999, several Vaccinia virus (VACV) isolates, the etiological agents of bovine vaccinia (BV), have been frequently isolated and characterized with various biological and molecular methods. The results from these approaches have grouped these VACV isolates into two different clusters. This dichotomy has elicited debates surrounding the origin of the Brazilian VACV and its epidemiological significance. To ascertain vital information to settle these debates, we and other research groups have made efforts to identify molecular markers to discriminate VACV from other viruses of the genus Orthopoxvirus (OPV) and other VACV-BR groups. In this way, some genes have been identified as useful markers to discriminate between the VACV-BR groups. However, new markers are needed to infer ancestry and to correlate each sample or group with its unique epidemiological and biological features. The aims of this work were to characterize a new VACV isolate (VACV DMTV-2005) molecularly and biologically using conserved and non-conserved gene analyses for phylogenetic inference and to search for new genes that would elucidate the VACV-BR dichotomy. The VACV DMTV-2005 isolate reported in this study is biologically and phylogenetically clustered with other strains of Group 1 VACV-BR, the most prevalent VACV group that was isolated during the bovine vaccinia outbreaks in Brazil. Sequence analysis of C23L, the gene that encodes for the CC-chemokine-binding protein, revealed a ten-nucleotide deletion, which is a new Group 1 Brazilian VACV genetic marker. This deletion in the C23L open reading frame produces a premature stop-codon that is shared by all Group 1 VACV-BR strains and may also reflect the VACV-BR dichotomy; the deletion can also be considered to be a putative genetic marker for non-virulent Brazilian VACV isolates and may be used for the detection and molecular characterization of new isolates.

From Lesions to Viral Clones: Biological and Molecular Diversity amongst Autochthonous Brazilian Vaccinia Virus

“…For the clones’ molecular characterization, DNA extractions were carried out using phenol-chloroform-isoamyl alcohol (PCI) [ 51 ] and used as template for Polymerase chain reaction (PCR) amplification of the viral genes A56R (hemagglutinin), C23L (chemokine binding protein) and A26L (A-type inclusion body). These genes are traditionally used for OPV phylogenetic studies, including Brazilian VACV strains [ 27 , 29 , 30 ].…”

“…Group 2 strains display higher plaque sizes and are virulent to mice, unlike Group 1 [ 24 , 25 , 26 , 27 , 28 ]. Moreover, polymorphisms observed in specific VACV genes, such as hemagglutinin gene (A56R), A-type inclusion body gene (A26L), and chemokine binding protein gene (C23L), have been used in phylogenetic studies and further confirmed the dichotomy between G1 and G2 VACV-BR [ 22 , 27 , 29 , 30 ].…”

Natural Vaccinia Virus Infection: Diagnosis, Isolation, and Characterization

“…To create the file containing nucleotide sequences to be analyzed, we suggest including sequences obtained from different species of Orthopoxvirus (Variola virus, Cowpox virus, Camelpox, Monkeypox virus, Rabbitpox virus, Horsepox virus, Buffalopox virus) and different sequences of VACV, including Br-VACV from groups 1 and 2 (see Table 14A .5.3;Drumond et al, 2008;Campos et al, 2011;Assis et al, 2012 Table 14A .5.3;Drumond et al, 2008;Campos et al, 2011;Assis et al, 2012). You can also retry sequences after a similarity search using each of your consensus sequences (after BLASTn or FASTAn analysis).…”

Dairy production practices and associated risks for bovine vaccinia exposure in cattle, Brazil

“…A genetically diverse set of vaccinia isolates has been recovered during these outbreaks and from related biological surveys aimed at virus discovery [5] , [6] , [7] , [8] , [9] , [10] , [11] , [12] , [13] , [14] , [15] , [16] . Some strains have been observed to persist in different regions across years and even decades [17] , suggesting that environmental conditions which favor the circulation and maintenance of these viruses exist across broad swaths of the country.…”

“…Speculation about the origin and emergence of VACV strains has pointed towards either the escape of so-called feral smallpox vaccine strains [3] , [20] or the emergence of resident ancestral strains [6] , [7] , [14] , [21] , [22] . Neither hypothesis has been wholly successful at explaining the range of lineages observed, some with genetic similarities to historic smallpox vaccine strains and others without these similarities.…”