Retrospective study of poxviruses diagnosed in cattle from Goiás State, Brazil (2010-2018) / Estudo retrospectivo de poxviroses diagnosticadas em bovinos no estado de Goiás (2010-2018)

A retrospective study of poxvirus infections diagnosed in cattle from Goiás state (GO), Brazil, from 2010 to 2018, was performed. All cases have been investigated by the GO Official Veterinary Service (Agrodefesa), from which technical forms and protocols of veterinary diagnosis laboratories were reviewed. In most cases, samples of oral or cutaneous tissues and/or swabs were submitted for virological diagnosis by polymerase chain reaction (PCR) and/or virus isolation. Thirty seven outbreaks/cases of vesicular disease were notified in cattle of 25 counties; in 33 cases the animals presented lesions clinically compatible with poxviruses. The etiology of 25 out of 33 outbreaks/cases was confirmed as poxviruses by PCR and/or viral isolation: 13 as bovine vaccinia virus (VACV), six as pseudocowpox virus (PCPV), five as bovine papular stomatitis virus (BPSV) and one coinfection (VACV and an Orf virus-like parapoxvirus). The laboratory confirmed that cases occurred mainly in dairy cattle (19/25) and during the dry season (22/25). In adult cattle, gross changes were observed mainly in the teats and udder and included vesicles, ulcers, crusts, papules and scars and varied of type, severity and affected region, depending on the poxvirus species. In calves, the main lesions were ulcers in the mouth and muzzle. Zoonotic lesions compatible with poxvirus infections were observed for all diagnosed poxviruses, affecting especially the hands of milkers and other farm workers. Our data demonstrate the sanitary and economic relevance of these diseases and the wide circulation of different poxviruses in cattle from GO.(AU)

Foi realizado um estudo retrospectivo das infecções por poxvírus diagnosticadas em bovinos do estado de Goiás (GO), entre 2010 e 2018. Todos os casos foram investigados pela Agência Goiana de Defesa Agropecuária (Agrodefesa). Foram revisados formulários técnicos e protocolos de laboratórios de diagnóstico veterinário. Na maioria dos casos, amostras de tecidos orais ou cutâneos e/ou swabs foram encaminhadas para diagnóstico virológico. Foram notificados 37 surtos/casos de doença vesicular em bovinos em 25 municípios; em 33 casos os animais apresentavam lesões clinicamente compatíveis com poxvírus. A etiologia de 25 de 33 surtos/casos foi confirmada como poxvírus por PCR e/ou isolamento viral: 13 como vírus vaccínia (VACV), seis como vírus pseudocowpox (PCPV), cinco como vírus da estomatite papular bovina (BPSV) e um caso de coinfecção (VACV e um parapoxvírus semelhante ao Orf vírus). Os casos confirmados laboratorialmente ocorreram principalmente em bovinos leiteiros (19/25) e durante a estação seca (22/25). Em bovinos adultos, alterações macroscópicas foram observadas principalmente nas tetas e úbere e incluíram vesículas, úlceras, crostas, pápulas e cicatrizes e variaram quanto ao tipo, gravidade e região afetada, dependendo da espécie do poxvírus. Em bezerros, as principais lesões foram úlceras na boca e focinho. Lesões zoonóticas compatíveis com infecção por poxvírus foram observadas em todas as poxviroses diagnosticadas, afetando principalmente as mãos dos ordenhadores e outros trabalhadores rurais. Nossos dados demonstram a relevância sanitária e econômica dessas doenças e a ampla circulação de diferentes poxvírus em bovinos de GO.(AU)

Retrospective study of poxviruses diagnosed in cattle from Goiás State, Brazil (2010-2018) / Estudo retrospectivo de poxviroses diagnosticadas em bovinos no estado de Goiás (2010-2018)

A retrospective study of poxvirus infections diagnosed in cattle from Goiás state (GO), Brazil, from 2010 to 2018, was performed. All cases have been investigated by the GO Official Veterinary Service (Agrodefesa), from which technical forms and protocols of veterinary diagnosis laboratories were reviewed. In most cases, samples of oral or cutaneous tissues and/or swabs were submitted for virological diagnosis by polymerase chain reaction (PCR) and/or virus isolation. Thirty seven outbreaks/cases of vesicular disease were notified in cattle of 25 counties; in 33 cases the animals presented lesions clinically compatible with poxviruses. The etiology of 25 out of 33 outbreaks/cases was confirmed as poxviruses by PCR and/or viral isolation: 13 as bovine vaccinia virus (VACV), six as pseudocowpox virus (PCPV), five as bovine papular stomatitis virus (BPSV) and one coinfection (VACV and an Orf virus-like parapoxvirus). The laboratory confirmed that cases occurred mainly in dairy cattle (19/25) and during the dry season (22/25). In adult cattle, gross changes were observed mainly in the teats and udder and included vesicles, ulcers, crusts, papules and scars and varied of type, severity and affected region, depending on the poxvirus species. In calves, the main lesions were ulcers in the mouth and muzzle. Zoonotic lesions compatible with poxvirus infections were observed for all diagnosed poxviruses, affecting especially the hands of milkers and other farm workers. Our data demonstrate the sanitary and economic relevance of these diseases and the wide circulation of different poxviruses in cattle from GO.

Foi realizado um estudo retrospectivo das infecções por poxvírus diagnosticadas em bovinos do estado de Goiás (GO), entre 2010 e 2018. Todos os casos foram investigados pela Agência Goiana de Defesa Agropecuária (Agrodefesa). Foram revisados formulários técnicos e protocolos de laboratórios de diagnóstico veterinário. Na maioria dos casos, amostras de tecidos orais ou cutâneos e/ou swabs foram encaminhadas para diagnóstico virológico. Foram notificados 37 surtos/casos de doença vesicular em bovinos em 25 municípios; em 33 casos os animais apresentavam lesões clinicamente compatíveis com poxvírus. A etiologia de 25 de 33 surtos/casos foi confirmada como poxvírus por PCR e/ou isolamento viral: 13 como vírus vaccínia (VACV), seis como vírus pseudocowpox (PCPV), cinco como vírus da estomatite papular bovina (BPSV) e um caso de coinfecção (VACV e um parapoxvírus semelhante ao Orf vírus). Os casos confirmados laboratorialmente ocorreram principalmente em bovinos leiteiros (19/25) e durante a estação seca (22/25). Em bovinos adultos, alterações macroscópicas foram observadas principalmente nas tetas e úbere e incluíram vesículas, úlceras, crostas, pápulas e cicatrizes e variaram quanto ao tipo, gravidade e região afetada, dependendo da espécie do poxvírus. Em bezerros, as principais lesões foram úlceras na boca e focinho. Lesões zoonóticas compatíveis com infecção por poxvírus foram observadas em todas as poxviroses diagnosticadas, afetando principalmente as mãos dos ordenhadores e outros trabalhadores rurais. Nossos dados demonstram a relevância sanitária e econômica dessas doenças e a ampla circulação de diferentes poxvírus em bovinos de GO.

First molecular characterization of poxviruses in cattle, sheep, and goats in Botswana.

BACKGROUND: Poxviruses within the Capripoxvirus, Orthopoxvirus, and Parapoxvirus genera can infect livestock, with the two former having zoonotic importance. In addition, they induce similar clinical symptoms in common host species, creating a challenge for diagnosis. Although endemic in the country, poxvirus infections of small ruminants and cattle have received little attention in Botswana, with no prior use of molecular tools to diagnose and characterize the pathogens. METHODS: A high-resolution melting (HRM) assay was used to detect and differentiate poxviruses in skin biopsy and skin scab samples from four cattle, one sheep, and one goat. Molecular characterization of capripoxviruses and parapoxviruses was undertaken by sequence analysis of RPO30 and GPCR genes. RESULTS: The HRM assay revealed lumpy skin disease virus (LSDV) in three cattle samples, pseudocowpox virus (PCPV) in one cattle sample, and orf virus (ORFV) in one goat and one sheep sample. The phylogenetic analyses, based on the RPO30 and GPCR multiple sequence alignments showed that the LSDV sequences of Botswana were similar to common LSDV field isolates encountered in Africa, Asia, and Europe. The Botswana PCPV presented unique features and clustered between camel and cattle PCPV isolates. The Botswana ORFV sequence isolated from goat differed from the ORFV sequence isolated from sheep. CONCLUSIONS: This study is the first report on the genetic characterization of poxvirus diseases circulating in cattle, goats, and sheep in Botswana. It shows the importance of molecular methods to differentially diagnose poxvirus diseases of ruminants.

An unusual presentation of pseudocowpox associated with an outbreak of pustular ulcerative vulvovaginitis in a Swedish dairy herd.

Species Pseudocowpox virus (PCPV; family Poxviridae) is known to cause pustular cutaneous disease in cattle. We describe an outbreak of pseudocowpox with an unusual clinical picture in a free-stall dairy herd of ~80 cows. Approximately 90% of the cows had vesicles, erosions, papules, and scabs on the vulva and vaginal mucosa. Histologic analysis of biopsy tissues indicated a primary, although not specified, viral infection. Transmission electron microscopy revealed parapoxvirus particles in both tissue and vesicular materials. Deep sequencing analysis of extracted DNA from swabbed vesicle areas gave a contig of nearly 120,000 nucleotides, matching the PCPV strain VR 634 with 100% identity. Analyses confirmed the absence of other potential causes of pustular vulvovaginitis such as bovine herpesvirus 1 and Ureaplasma diversum. A rolling cow brush was suspected to be the fomite.

First isolation and genetic characterization of pseudocowpox virus from cattle in Japan.

BACKGROUND: Pseudocowpox virus (PCPV) infects cattle worldwide with zoonotic potential but has not been isolated in Japan. Thus, the epidemiological status of PCPV infection in cattle is undetermined. RESULTS: In May 2016, a cattle in a farm in Yamaguchi Prefecture showed white vesicles and hyperemia in the mucosa under the tongue surface, but not on the teats and coronary cushions. A parapoxvirus was isolated from the oral lesion swab and was genetically characterized based on the full-length sequence of B2L gene encoding viral envelope. Phylogenetic analysis showed that the isolated virus was classified into PCPV. CONCLUSION: This case indicates its potential spread in Japan. This is the first report of isolation of PCPV in Japan.

Milker's nodules: classic histological findings.

Milker's nodule is an occupational dermatovirose caused by Parapoxvirus, which is self-limited and, due to the lack of information of health professionals, may lead to underdiagnosis. We present two cases with exuberant manifestations and classic histopathologic findings. Case 1: Male, 19 years of age, milker, presented nodules and blisters on his palm for 15 days. Case 2: Male, 33 years of age, administrative assistant, presented erythematous nodules on his palms with lymphangitis for 5 days. He had milked a cow one week before the onset of the lesions. In both cases, the histopathology was representative and coincident with the clinical hypothesis. The lesions have presented complete involution. Milker's nodule diagnosis is based on clinical presentation, epidemiology, and histopathology. The knowledge of this disease is essential for its correct diagnosis, as well as to guide the implementation of public health measures and the appropriate treatment of sick cattle.

Detection of pseudocowpox virus in water buffalo (Bubalus bubalis) with vesicular disease in the state of São Paulo, Brazil, in 2016.

BACKGROUND: Parapoxviruses are zoonotic viruses that infect cattle, goats and sheep; there have also been reports of infections in camels, domestic cats and seals. OBJECTIVE: The objective of this report was to describe a case of vesicular disease caused by pseudocowpox virus (PCPV) in water buffalo (Bubalus bubalis) in Brazil. ANIMALS: Sixty buffalo less than 6 months old exhibited ulcers and widespread peeling of the tongue epithelium. There were no cases of vesicular disease in pigs or horses on the same property. METHODS: Samples were analysed by PCR and sequencing. Phylogenetic analysis in MEGA 7.01 was reconstructed using major envelope protein (B2L) by the Tamura three-parameter nucleotide substitution model and the maximum likelihood and neighbor joining models, both with 1000 bootstrap replicates. The genetic distance between the groups was analysed in MEGA using the maximum composite likelihood model. The rate variation among sites was modeled using gamma distribution. RESULTS: The presence of PCPV in the buffalo herd could be demonstrated in epithelium and serum. The minimum genetic distance between the isolated PCPV strain (262-2016) and orf virus and bovine papular stomatitis virus was 6.7% and 18.4%, respectively. The maximum genetic distance calculated was 4.6% when compared with a PCPV detected in a camel. Conclusions/Clinical Importance: The peculiar position of the isolated strain in the phylogenetic trees does not necessarily indicate a different kind of PCPV that infects buffalo. More samples from cattle and buffalo in Brazil must be sequenced and compared to verify if PCPV from buffalo are genetically different from samples derived from cattle.

Milker's nodule--Case report.

Milker's nodule is an occupational viral skin disease of universal distribution, caused by the Paravaccinia virus and that occurs in individuals who deal with dairy cattle herds. We describe a case acquired due to lack of use of PPE (Personal Protective Equipment) and perform a literature review.

Milker's nodule - Case report

Abstract Milker's nodule is an occupational viral skin disease of universal distribution, caused by the Paravaccinia virus and that occurs in individuals who deal with dairy cattle herds. We describe a case acquired due to lack of use of PPE (Personal Protective Equipment) and perform a literature review.

Characterization of GM-CSF-inhibitory factor and Uracil DNA glycosylase encoding genes from camel pseudocowpoxvirus.

The present study describes the PCR amplification of GM-CSF-inhibitory factor (GIF) and Uracil DNA glycosylase (UDG) encoding genes of pseudocowpoxvirus (PCPV) from the Indian Dromedaries (Camelus dromedarius) infected with contagious ecthyma using the primers based on the corresponding gene sequences of human PCPV and reindeer PCPV, respectively. The length of GIF gene of PCPV obtained from camel is 795 bp and due to the addition of one cytosine residue at position 374 and one adenine residue at position 516, the open reading frame (ORF) got altered, resulting in the production of truncated polypeptide. The ORF of UDG encoding gene of camel PCPV is 696 bp encoding a polypeptide of 26.0 kDa. Comparison of amino acid sequence homologies of GIF and UDG of camel PCPV revealed that the camel PCPV is closer to ORFV and PCPV (reference stains of both human and reindeer), respectively.

Comparison of virokine from camel pseudocowpoxvirus (PCPV) with interleukin 10 of the Dromedary camel (Camelus dromedarius).

Cellular interleukin-10 (IL-10) gene from the peripheral blood mononuclear cells of the healthy Dromedary camel (Camelus dromedarius) and viral IL-10 (vIL-10) from the skin scabs of the Dromedary camels infected with contagious ecthyma (a parapoxviral infection in the camels) were amplified by polymerase chain reaction, cloned and characterized. Sequence analysis revealed that the open reading frame (ORF) of dromedarian camel IL-10 is 537 bp in length, encoding 178 amino acid polypeptide while open reading frame of vIL-10 from camel is 561 bp, encoding 187 amino acid polypeptide. The Dromedary camel IL-10 exhibited 62.6% and 68.5% sequence identity at the nucleotide and amino acid level, respectively, with vIL-10 from camel. Sequence analysis also revealed that the Dromedary camel IL-10 shared 99.4% and 98.3% identity at the nucleotide and amino acid level, respectively, with the Bactrian camel (Camelus bactrianus). But vIL-10 from camel shared 84.7% and 83.4% sequence identity at the nucleotide and amino acid level, respectively, with vIL-10 from reindeer (Rangifer tarandus), which is a ruminant species belonging to the order Artiodactyla. The present study was conducted to evaluate the evolutionary origin of the camel parapoxvirus with parapoxviruses of cattle and sheep and the resultant sequence analysis revealed that camel parapoxvirus is closely related to cattle parapoxvirus than sheep parapoxvirus (Orf virus).

Conservation and variation of the parapoxvirus GM-CSF-inhibitory factor (GIF) proteins.

The GIF protein of orf virus (ORFV) binds and inhibits the ovine cytokines granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-2 (IL-2). An equivalent protein has so far not been found in any of the other poxvirus genera and we therefore investigated whether it was conserved in the parapoxviruses. The corresponding genes from both the bovine-specific pseudocowpox virus (PCPV) and bovine papular stomatitis virus (BPSV) were cloned and sequenced. The predicted amino acid sequences of the PCPV and BPSV proteins shared 88 and 37 % identity, respectively, with the ORFV protein. Both retained the six cysteine residues and the WSXWS-like motif that are required for biological activity of the ORFV protein. However, an analysis of the biological activity of the two recombinant proteins revealed that, whilst the PCPV GIF protein bound to both ovine and bovine GM-CSF and IL-2 with very similar binding affinities to the ORFV GIF protein, no GM-CSF- or IL-2-binding activity was found for the BPSV protein.

[Screening of HIV-1 replication inhibitors by using pseudotyped virus system].

This study is to establish a cell-based pharmacological model targeting HIV-1 replication for compounds screening and to screen compounds randomly selected from compounds library by using this pseudotyped viral system. The cell-based HIV-1 replication pharmacological model was set up by HIV-1 core packed with vesicular stomatitis virus glycoprotein. The level of HIV-1 replication was presented by reporter genes expression (luciferase activity or percentage of GFP positive cells). When a compound has inhibitory effect on VSVG/HIV model, VSVG/MLV model would be used to test for specificity. Vesicular stomatitis virus glycoprotein can efficiently mediate HIV core into a wide range of host cells. Expression level of reporter genes showed dose-dependent manner with virion dilution. Among 500 compounds, three compounds dose-dependently inhibit HIV-1 replication, but not MLV replication. VSVG/HIV pseudotyped viral system can be used as a pharmacological model for HIV-1 replication inhibitor screening. Compounds 2-methylthio-5-(4-methylbenzo)amido-l,3,4-thiadiazole, N-(3-hydroxyphenyl)-2-(4-isobutylphenyl) propionamide, and N-(4-picolyl)-4-methylbenzenesulfonamide can specifically inhibit HIV-1 replication with IC50 of 1.92, 5.38, and 3.39 micromol L(-1) respectively.

Development of methodology based on commercialized SERS-active substrates for rapid discrimination of Poxviridae virions.

Surface-enhanced Raman spectroscopy (SERS) can be made an attractive approach for identification of Raman-active compounds and biological materials (i.e., toxins, viruses, or intact bacterial cells/spores) through development of reproducible, spatially uniform SERS-active substrates. Recently, reproducible (from substrate-to-substrate), spatially homogeneous (over large areas) SERS-active substrates have been commercialized and are now available in the marketplace. We have utilized these patterned surfaces to acquire SERS spectral signatures of intact bovine papular stomatitis, pseudocowpox, and Yaba monkey tumor viruses. Salient spectral signature features make it possible to discriminate among these genetically distinct Poxviridae-Chordopoxvirinae virions. In addition, partial least-squares, a multivariate calibration method, has been used to develop personal computer-borne algorithms useful for classification of unknown Parapoxvirus (e.g., bovine papular stomatitis virus and pseudocowpox virus) samples based solely on SERS spectral signatures. To our knowledge, this is the first report detailing application of these commercial-off-the-shelf (COTS) SERS-active substrates to identification of intact poxviruses.

Nódulo de los ordeñadores / Milker's nodule

El nódulo de los ordeñadores es una afección viral benigna que por lo general se presenta como un nódulo en la mano de personas que manipulan ganado bovino infectado. Reportamos el caso de una mujer de 36 años con infrecuente localización facial consecutivo a cornada de vaca. Los rasgos clínicos característicos, la autorresolución del cuadro, así como los cambios citopáticos virales en la biopsia fueron significativos para establecer el diagnóstico. Es una enfermedad que podemos encontrarla en zonas periurbanas de nuestro país y factible de condicionar error diagnóstico.

Pseudocowpox virus encodes a homolog of vascular endothelial growth factor.

We have identified a gene encoding a homolog of vascular endothelial growth factor (VEGF) in the Pseudocowpox virus (PCPV) genome. The predicted protein shows 27% amino acid identity to human VEGF-A. It also shows 41 and 61% amino acid identity to VEGFs encoded by orf virus (ORFV) strains NZ2 and NZ7, respectively. Assays of the expressed VEGF-like protein of PCPV (PCPV(VR634)VEGF) demonstrated that PCPV(VR634)VEGF is mitogenic for endothelial cells and is capable of inducing vascular permeability. PCPV(VR634)VEGF bound VEGF receptor-2 (VEGFR-2) but did not bind VEGFR-1 or VEGFR-3. These results indicate that PCPV(VR634)VEGF is a biologically active member of the VEGF family which shares with the ORFV-encoded VEGFs a receptor binding profile that differs from those of all cellular members of the VEGF family. It seems likely that the biological activities of PCPV(VR634)VEGF contribute to the proliferative and highly vascularized nature of PCPV lesions.

Nódulos dos ordenhadores: relato de dez casos / Milker's nodules: a report of ten cases

Os autores apresentam estudo de dez pacientes portadores de nódulos dos ordenhadores, com idades variando de 13 a 48 anos, sendo oito do sexo masculino e dois do feminino, todos com atividades relacionadas ao contato ou manuseio de animais ou derivados contaminados. O número de lesöes variou de um a três, todas localizadas nas mäos e nos dedos. Dois pacientes apresentaram fenômenos associados, febre em um e no outro linfangite e erupçäo micropapulosa do membro superior direito. Näo foram realizados tratamentos, excluído o sintomático, havendo regressäo espontânea das lesoes

Differentiation of parapoxviruses by application of orf virus-specific monoclonal antibodies against cell surface proteins.

Monoclonal antibodies were produced against orf virus-specified cell surface proteins in an attempt to develop reagents capable of differentiating between members of the Parapoxviridae. Two immunization protocols were used to induce an anti-orf response in BALB/c mice, one of which resulted in virus replication in the recipient. The monoclonal antibodies produced were tested for crossreactivity with bovine papular stomatitis virus (BPS) and milker's node virus (MNV) by indirect immunofluorescence assay (IFA) and immunoblotting. The results indicate that significant antigenic overlap exists between isolates of orf, MNV and BPS, even at the level of specificity provided by monoclonal antibodies. One monoclonal antibody reacted strongly in IFA with orf virus isolates, very weakly with MNV, and not at all with BPS. On immunoblots this same antibody recognized a 40-43 kDa protein in orf virus-infected cells, and also a 45-48 kDa protein in cells infected with MNV or BPS virus. The data suggest that it may be possible to define parapoxvirus strains on the basis of small variations in specific virus-directed cell surface proteins.

Giant parapox infection in a two year old child.

We report the case of a two year old boy who developed a large tumour on his face. The lesion resembled a pyogenic granuloma clinically and histologically. Viral studies indicated a parapox infection and a bovine source was supported on epidemiological grounds. The lesion was removed by shave excision and the area healed without significant scarring.