Molecular and Biological Characterization of a Cervidpoxvirus Isolated From Moose with Necrotizing Dermatitis.

Cervidpoxvirus is one of the more recently designated genera within the subfamily Chordopoxvirinae, with Deerpox virus (DPV) as the only recognized species to date. In this study, the authors describe spontaneous disease and infection in the North American moose (Alces americanus) by a novel Cervidpoxvirus, here named Moosepox virus (MPV). Three 4-month-old moose calves developed a multifocal subacute-to-chronic, necrotizing, suppurative-to-granulomatous dermatitis that affected the face and the extremities. Ultrastructurally, all stages of MPV morphogenesis-that is, crescents, spherical immature particles, mature particles, and enveloped mature virus-were observed in skin tissue. In vitro infection with MPV confirmed that its morphogenesis was similar to that of the prototype vaccinia virus. The entire coding region, including 170 putative genes of this MPV, was sequenced and annotated. The sequence length was 164,258 bp with 98.5% nucleotide identity with DPV (strain W-1170-84) based on the whole genome. The genome of the study virus was distinct from that of the reference strain (W-1170-84) in certain genes, including the CD30-like protein (83.9% nucleotide, 81.6% amino acid), the endothelin precursor (73.2% nucleotide including some indels, 51.4% amino acid), and major histocompatibility class (MHC) class I-like protein (81.0% nucleotide, 68.2% amino acid). This study provides biological characterization of a new Cervidpoxvirus attained through in vivo and in vitro ultrastructural analyses. It also demonstrates the importance of whole-genome sequencing in the molecular characterization of poxviruses identified in taxonomically related hosts.

Isolation and characterization of a Cervidpoxvirus from a goitered gazelle (Gazella subgutturosa) from a zoologic park in Minnesota.

Deerpox virus (DPV) is the sole member of the newly ratified Cervidpoxvirus genus in the subfamily Chordopoxvirinae. Presented here is the first diagnostic report of isolation of DPV from a goitered gazelle (Gazella subgutturosa). A tissue homogenate was submitted by a zoologic park to the Minnesota Veterinary Diagnostic Laboratory at the University of Minnesota for poxvirus diagnostic investigation and then referred to Plum Island Foreign Animal Disease Diagnostic Laboratory for confirmation. Poxviral infection was confirmed using electron microscopy. The virus was cultured in vero cells and subjected to further diagnoses for characterization. Polymerase chain reaction targeting the major envelope (B2L) protein and RNA polymerase of parapoxviruses, and the poly-A polymerase gene of capripoxviruses, were all negative. Degenerative pan-poxvirus primers that target the DNA polymerase (DNApol) and DNA topoisomerase (DNAtopo) genes, however, successfully amplified poxviral DNA fragments. Amplification of the DNApol and DNAtopo genes yielded fragments of 543 and 344 base pairs, respectively. DNA sequence and phylogenetic analysis of each gene fragment from the gazelle isolate showed >97% identity in BLAST searches with two DPV virus strains (W848-83 and W-1170-84) isolated from North American mule deer (Odocoileus hemionus) in 1983-1984. Neighbor-joining trees indicate that the isolate is a member of the Cervidpoxvirus genus and shows a more-distant relationship to other ruminant poxviruses, namely the Capripoxvirus genus consisting of lumpy skin disease, sheeppox, and goatpox viruses. This report documents the premiere finding of DPV, a recently characterized virus, in gazelles and demonstrates the need for broadened investigation when diagnosing poxvirus infections in ruminants.

Comparative susceptibility of eastern cottontails and New Zealand white rabbits to classical rabbit haemorrhagic disease virus (RHDV) and RHDV2.

Rabbit haemorrhagic disease virus (RHDV) is associated with high morbidity and mortality in the European rabbit (Oryctolagus cuniculus). In 2010, a genetically distinct RHDV named RHDV2 emerged in Europe and spread to many other regions, including North America in 2016. Prior to this study it was unknown if eastern cottontails (ECT(s); Sylvilagus floridanus), one of the most common wild lagomorphs in the United States, were susceptible to RHDV2. In this study, 10 wild-caught ECTs and 10 New Zealand white rabbits (NZWR(s); O. cuniculus) were each inoculated orally with either RHDV (RHDVa/GI.1a; n = 5 per species) or RHDV2 (a recombinant GI.1bP-GI.2; n = 5 per species) and monitored for the development of disease. Three of the five ECTs that were infected with RHDV2 developed disease consistent with RHD and died at 4 and 6 days post-inoculation (DPI). The RHDV major capsid protein/antigen (VP60) was detected in the livers of three ECTs infected with RHDV2, but none was detected in the ECTs infected with RHDV. Additionally, RHD viral RNA was detected in the liver, spleen, intestine and blood of ECTs infected with RHDV2, but not in the ECTs infected with RHDV. RHD viral RNA was detected in urine, oral swabs and rectal swabs in at least two of five ECTs infected with RHDV2. One ECT inoculated with RHDV2 seroconverted and developed a high antibody titre by the end of the experimental period (21 DPI). ECTs inoculated with the classic RHDV did not seroconvert. In comparison, NZWRs inoculated with RHDV2 exhibited high mortality (five of five) at 2 DPI and four of five NZWRs inoculated with RHDV either died or were euthanized at 2 DPI indicating both of these viruses were highly pathogenic to this species. This experiment indicates that ECTs are susceptible to RHDV2 and can shed viral RNA, thereby suggesting this species could be involved in the epidemiology of this virus.

Phylodynamics of parapoxvirus genus in Mexico (2007-2011).

In this study we report for the first time the phylodynamics of the parapoxvirus (PPV) genus in Mexico. Based on the analysis by PCR of 124 epithelial samples collected between 2007 and 2011 from naturally infected goats, sheep and cows in Mexico, we found that different PPV were present in 21 out of the 24 states sampled during this study. Our phylogenetic analysis confirmed the presence of different PPV species in Mexico, and their phylogenetic relationship with other PPV circulating in the US and Canada. Furthermore, we describe the existence of two different ORFV phylogenetic groups that are clearly host associated (sheep or goat). Evidence of directional selection at five specific amino acid residues in the enveloped glycoprotein B2L might help to support this host predilection. Collectively, the results generated in this study highlight the importance of PPV genus in Mexico and open the possibility for future studies describing with more detail the importance of this genus in North America.

Real-Time Reverse Transcription PCR Assay for Detection of Senecavirus A in Swine Vesicular Diagnostic Specimens.

Senecavirus A (SV-A), formerly, Seneca Valley virus (SVV), has been detected in swine with vesicular lesions and is thought to be associated with swine idiopathic vesicular disease (SIVD), a vesicular disease syndrome that lacks a defined causative agent. The clinical presentation of SIVD resembles that of other more contagious and economically devastating vesicular diseases, such as foot-and-mouth disease (FMD), swine vesicular disease (SVD), and vesicular stomatitis (VS), that typically require immediate rule out diagnostics to lift restrictions on animal quarantine, movement, and trade. This study presents the development of a sensitive, SYBR Green RT-qPCR assay suitable for detection of SV-A in diagnostic swine specimens. After testing 50 pigs with clinical signs consistent with vesicular disease, 44 (88%) were found to be positive for SV-A by RT-qPCR as compared to none from a negative cohort of 35 animals without vesicular disease, indicating that the assay is able to successfully detect the virus in an endemic population. SV-A RNA was also detectable at a low level in sera from a subset of pigs that presented with (18%) or without (6%) vesicular signs. In 2015, there has been an increase in the occurrence of SV-A in the US, and over 200 specimens submitted to our laboratory for vesicular investigation have tested positive for the virus using this method. SV-A RNA was detectable in all common types of vesicular specimens including swabs and tissue from hoof lesions, oral and snout epithelium, oral swabs, scabs, and internal organ tissues such as liver and lymph node. Genome sequencing analysis from recent virus isolates was performed to confirm target amplicon specificity and was aligned to previous isolates.

Poxvirus infection of Steller sea lions (Eumetopias jubatus) in Alaska.

Lesions suggestive of poxvirus infection were observed in two Steller sea lions (Eumetopias jubatus) in Alaska during live capture-and-release studies during 2000 and 2001. Both of these animals, female pups in poor body condition, were from Prince William Sound; this population is part of the declining western stock. Umbilicated, typically ulcerated dermal nodules were present, primarily on the fore flippers in one case, and over most of the body in the second case. Histologically, there were discrete masses in the superficial dermis composed of epithelial cells, some of which contained eosinophilic intracytoplasmic inclusion bodies. Negative staining of skin biopsy homogenates demonstrated the presence of orthopoxvirus-like particles. Total DNA extracted from skin biopsies were analyzed by polymerase chain reaction (PCR) using primers that targeted the DNA polymerase and DNA topoisomerase genes. These primers directed the amplification of fragments 543 base pairs (bp) and 344 bp, respectively, whose deduced amino acid sequences indicated the presence of a novel poxvirus within the Chordopoxvirinae subfamily. Comparison of these amino acid sequences with homologous sequences from members of the Chordopoxvirinae indicated highest identity with orthopoxviruses.

A partial deletion in non-structural protein 3A can attenuate foot-and-mouth disease virus in cattle.

The role of non-structural protein 3A of foot-and-mouth disease virus (FMDV) on the virulence in cattle has received significant attention. Particularly, a characteristic 10-20 amino acid deletion has been implicated as responsible for virus attenuation in cattle: a 10 amino acid deletion in the naturally occurring, porcinophilic FMDV O1 Taiwanese strain, and an approximately 20 amino acid deletion found in egg-adapted derivatives of FMDV serotypes O1 and C3. Previous reports using chimeric viruses linked the presence of these deletions to an attenuated phenotype in cattle although results were not conclusive. We report here the construction of a FMDV O1Campos variant differing exclusively from the highly virulent parental virus in a 20 amino acid deletion between 3A residues 87-106, and its characterization in vitro and in vivo. We describe a direct link between a deletion in the FMDV 3A protein and disease attenuation in cattle.

Diagnosis of Porcine teschovirus encephalomyelitis in the Republic of Haiti.

In February and March 2009, approximately 1,500 backyard pigs of variable age became sick, and approximately 700 of them died or were euthanized in the Lower Artibonite Valley and the Lower Plateau of the Republic of Haiti. The main clinical sign was posterior ataxia followed by paresis and/or paralysis on the second or third day of illness. No gross lesions were observed at postmortem examinations. The morbidity and mortality were approximately 60% and 40%, respectively. Diagnostic samples (whole blood, brain, tonsil, lymph nodes, spleen, and lung) were negative for Classical swine fever virus and African swine fever virus. Porcine teschovirus type 1 was detected by reverse transcription polymerase chain reactions in brain samples. Results of virus isolation, electron microscopy of virus particles, histopathological analysis on brain tissues, nucleic acid sequencing, and phylogenetic analysis of the viral isolate supported the diagnosis of teschovirus encephalomyelitis. The outbreak of the disease in Haiti is the first appearance of the severe form of teschovirus encephalomyelitis in the Americas. This disease poses a potential threat to the swine industries in other Caribbean countries, as well as to Central and North American countries.

Discovery of swine as a host for the Reston ebolavirus.

Since the discovery of the Marburg and Ebola species of filovirus, seemingly random, sporadic fatal outbreaks of disease in humans and nonhuman primates have given impetus to identification of host tropisms and potential reservoirs. Domestic swine in the Philippines, experiencing unusually severe outbreaks of porcine reproductive and respiratory disease syndrome, have now been discovered to host Reston ebolavirus (REBOV). Although REBOV is the only member of Filoviridae that has not been associated with disease in humans, its emergence in the human food chain is of concern. REBOV isolates were found to be more divergent from each other than from the original virus isolated in 1989, indicating polyphyletic origins and that REBOV has been circulating since, and possibly before, the initial discovery of REBOV in monkeys.