About the necessity of including HoBi-like pestiviruses in bovine respiratory and reproductive viral vacines / Sobre a necessidade de incluir pestivírus HoBi-like em vacinas virais respiratórias e reprodutivas para bovinos

HoBi-like pestiviruses (HoBiPeV) constitute a novel group of bovine pestiviruses, genetically and antigenically related to bovine viral diarrhea virus 1 (BVDV-1) and BVDV-2. Recent data shows that HoBiPeV are endemic among Brazilian cattle, yet bovine reproductive/respiratory vaccines contain only BVDV-1 and BVDV-2 strains. The present study investigated the neutralizing antibody response against these pestiviruses induced by two commercial vaccines (VA = attenuated, VI = inactivated) and by three experimental, replicative, vaccine formulations (VAC1 = monovalent, BVDV-1; VAC2 = bivalent, BVDV-1 + BVDV-2; VAC3 = trivalent, BVDV-1 + BVDV-2 and HoBiPeV). Seronegative beef calves were immunized once (replicative vaccines) or twice (inactivated vaccine) and serum samples were tested by virus-neutralization (VN) 30 days after vaccination (dpv) (replicative vaccines) or 30 days after the second dose (VI). We considered a threshold VN titer of ≥60 indicative of protection against clinical disease. At 30 dpv, VA induced protective titers against BVDV-2 in 7/7 animals (GMT=289.8) and against BVDV-1 and HoBiPeV in 5/7 animals (GMTs=97.5 and 80, respectively). VI induced protective titers against BVDV-1 in 1/7 animal (GMT=16.4), 2/7 animals against BVDV-2 (GMT=53.8) and in none of the calves against HoBiPeV (GMT=12.2). When a pool of sera of each vaccine group was tested against individual Brazilian isolates, VA induced protective titers against 3/7 BVDV-1 isolates, to 9/10 (BVDV-2) and 1/8 (HoBiPeV); VI induced protective titers against 1/7 (BVDV-1), 1/10 (BVDV-2) and none (0/8) HoBiPeV isolates. The experimental vaccine VAC1 induced protective titers against BVDV-1 in 9/9 animals (GMT=320) but in no animal against BVDV-2 or HoBiPeV (GMT<10). VAC2 induced protective titers to BVDV-1 and BVDV-2 in 9/9 animals (GMTs=160 and 640, respectively), and against HoBiPeV in 7/9 animals (GMT=108.5). Finally, VAC3 induced protective titers in all animals against BVDV-1 (GMT=234.3), BVDV-2 (294.9) and HoBiPeV (201.1). Testing the pool of sera against pestivirus isolates, VAC1 induced titers ≥ 60 against 4/7 BVDV-1 but to none BVDV-2/HoBiPeV isolate; VAC2 induced protective titers against 4/7 BVDV-1; 10/10 BVDV-2 and 2/8 HoBiPeV; VAC3 induced protective titers against all BVDV-1, BVDV-2 and HoBiPeV isolates. These results indicate that vaccines composed by BVDV-1+BVDV-2, especially those containing inactivated virus, may not induce serological response against a variety of HoBiPeV isolates. Thus, the need of inclusion of HoBiPeV in vaccine formulations should be considered.(AU)

Os pestivírus HoBi-like (HoBiPeV) compõe um grupo novo de pestivírus de bovinos, genética e antigenicamente relacionados com os vírus da diarreia viral bovina 1 e 2 (BVDV-1, BVDV2). Dados recentes indicam que os HoBiPeV são endêmicos na população bovina do Brasil, mas as vacinas respiratórias e reprodutivas bovinas contêm apenas cepas de BVDV-1 e BVDV-2. O presente estudo investigou a atividade neutralizante contra estes pestivírus induzidas por duas vacinas comerciais (VA = atenuada, VI = inativada) e por três vacinas experimentais replicativas (VAC1 = monovalente, BVDV-1; VAC2 = bivalente, BVDV-1 + BVDV-2; VAC3 = trivalente, BVDV-1 + BVDV-2 e HoBiPeV). Bezerros soronegativos foram imunizados uma vez (vacinas replicativas) ou duas (vacina inativada) e amostras de soro foram testadas por vírus-neutralização (VN) 30 dias após a vacinação (dpv) (vacinas replicativas) ou 30 dias após a segunda dose (VI). Títulos neutralizantes ≥60 foram considerados indicativos de proteção contra doença clínica. Nesta data, a VA induziu títulos protetivos contra o BVDV-2 em 7/7 animais (GMT=289,8) e contra BVDV-1 e HoBiPeV em 5/7 animals (GMTs=97,5 e 80, respectivamente). VI induziu títulos protetores contra BVDV-1 em 1/7 animal (GMT=16,4), em 2/7 animais contra BVDV-2 (GMT=53,8) e em nenhum contra HoBiPeV (GMT=12,2). Quando um pool de soro de cada grupo vacinal foi testado frente a isolados Brasileiros, a VA induziu títulos protetores contra 3/7 isolados de BVDV-1, 9/10 (BVDV-2) e 1/8 (HoBiPeV); VI induziu títulos protetores em 1/7 contra BVDV-1, 1/10 (BVDV-2) e em nenhum (0/8) contra isolados de HoBiPeV. A VAC1 induziu títulos protetores contra BVDV-1 em 9/9 animais (GMT=320) mas em nenhum animal contra BVDV-2 ou HoBiPeV (GMT<10). VAC2 induziu títulos protetores contra BVDV-1e BVDV-2 em 9/9 animais (GMTs=160 e 640, respectivamente),e contra HoBiPeV em 7/9 animais (GMT=108,5). Finalmente, VAC3 induziu títulos protetores em todos os animais contra BVDV-1 (GMT=234,3), BVDV-2 (294,9) e HoBiPeV (201,1). No teste de pool de soro contra isolados de pestivírus, VAC1 induziu títulos ≥60 contra 4/7 BVDV-1 mas contra nenhum isolado de BVDV-2/HoBiPeV; VAC2 induziu títulos protetores contra 4/7 BVDV-1; 10/10 BVDV-2 e 2/8 HoBiPeV; VAC3 induziu títulos protetores contra todos BVDV-1, BVDV-2 e HoBiPeV. Esses resultados indicam que vacinas contendo apenas BVDV-1 BVDV-2, especialmente aquelas inativadas, podem não conferir resposta sorológica protetora contra vários isolados de HoBiPeV. Portanto, a necessidade de se incluir cepas de HoBiPeV nas vacinas deve ser considerada.(AU)

Identification and characterization of atypical porcine pestivirus genomes in newborn piglets with congenital tremor in China

Recently, a novel atypical porcine pestivirus (APPV) in pig was reported. In this study, two APPV strains, APPV-China/GZ01/2016 (GZ01) and APPV-China/GD-SD/2016 (GD-SD), were identified in two newborn piglet herds with congenital tremor from China. The open reading frame of the two strains shared an 83.5% nucleotide identity. Phylogenetically, the APPV strains were placed into two groups: GZ01 belonged to group I and GD-SD belonged to group II. A high viral load was detected in the cerebellum (quantification cycles < 26). Further studies should be carried out to thoroughly elucidate the development of congenital tremors caused by APPV.

Detecção do virus 'HoBi'-like (BVDV-3) em bovino no semiárido do Estado da Paraíba / Detection of 'HoBi'-like (BVDV-3) vírus in a cow in the semiarid of Paraíba, Northeastern Brazil

Objetivou-se descrever os aspectos clínicos e anatomopatológicos, e a identificação viral de um caso de infecção pelo vírus 'Hobi'-like (BVDV-3) em bovino do semiárido paraibano, Nordeste do Brasil. Um bovino, fêmea, três meses de idade, foi levado ao Hospital Veterinário da UFCG apresentando salivação, dificuldade de apreensão do teto, falta de apetite, fezes escuras e em pouca quantidade. Diante da piora do quadro clínico optou-se por sua eutanásia in extremis, seguida da realização da necropsia e coleta de material para histopatologia. Histologicamente, nas mucosas do trato digestivo, havia edema, degeneração balonosa, necrose e infiltrado inflamatório, que foi observado na face dorsal da língua e no seu epitélio mais profundo. A imunohistoquímica de amostras de extremidade de pavilhão auricular demonstrou marcação antigênica positiva e pela RT-PCR foi possível detectar RNA viral do BVDV no soro sanguíneo, cujo efeito citopático em células epiteliais de rim bovino da linhagem "Madin Darby bovine kidney" (MDBK) não foi observado. O sequenciamento do gene 5'NCR demonstrou que o vírus isolado estaria mais relacionado ao 'Hobi'-like (BVDV-3). Após a confirmação do diagnóstico foram coletadas amostras de soro dos 23 animais do rebanho para sorologia por ELISA indireto, sendo constatada 69,6% (16/23) de soropositividade. A identificação deste novo caso de infecção por 'Hobi'-like na Paraíba reafirma a necessidade de um monitoramento regular para BVDV na região para detecção precoce da infecção dos rebanhos e adoção de medidas eficazes de prevenção e controle.(AU)

The aim of this study was to describe the clinical and anatomopathological aspects, and the viral identification of a case of 'Hobi'-like (BVDV-3) vírus infection in cattle from the semiarid of Paraiba State, Northeastern Brazil. A female bovine, three months old, was sent to the UFCG's Veterinary Hospital presenting salivation, difficulty of ceiling seizure, lack of appetite, dark feces and in small amounts. Due to worsening of symptoms it was decided to in extremis euthanasia, followed by the necropsy and collection of material for histopathology. Histologically, in the mucous membranes of the digestive tract there were edema, ballooning degeneration, necrosis and inflammatory infiltrate, which was observed on the dorsal surface of the tongue and in its deepest epithelium. The immunohistochemical of skin biopsies of the extremity of the ear (ear notches) showed positive antigenic marking and by RT-PCR it was possible to detect viral RNA of BVDV in the serum, whose cytopathic effect in epithelial cells of bovine kidney lineage "Madin Darby bovine kidney" (MDBK)was not observed. The sequencing of the 5'NCR gene showed that the virus isolated was more related to 'Hobi'-like (BVDV-3). After confirming the diagnosis serum samples were collected from 23 animals for serology by indirect ELISA, and a seropositivity of 69.6% (16/23) was found. The identification of this new case of 'Hobi'-like infection in Paraiba reaffirms the need for regular BVDV monitoring in the region to early detection of infection in the herds and adoption of effective preventive and control measures.(AU)

In vitro inhibition of the bovine viral diarrhoea virus by the essential oil of Ocimum basilicum (basil) and monoterpenes

The bovine viral diarrhoea virus (BVDV) is suggested as a model for antiviral studies of the hepatitis C virus (HCV). The antiviral activity of the essential oil of Ocimum basilicum and the monoterpenes camphor, thymol and 1,8-cineole against BVDV was investigated. The cytotoxicities of the compounds were measured by the MTT (3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide) test, and the antiviral activities were tested by the plaque reduction assay. The oil or compounds were added to the assay in three different time points: a) pre-treatment of the virus (virucidal assay); b) pre-treatment of the cells; or c) post-treatment of the cells (after virus inoculation). The percentage of plaques inhibition for each compound was determined based on the number of plaques in the viral control. The results were expressed by CC50 (50% cytotoxic concentration), IC50 (inhibitory concentration for 50% of plaques) and SI (selectivity index = CC50/IC50). Camphor (CC50 = 4420.12 µgmL-1) and 1,8-cineole (CC50 = 2996.10 µgmL-1) showed the lowest cytotoxicities and the best antiviral activities (camphor SI = 13.88 and 1,8-cineol SI = 9.05) in the virucidal assay. The higher activities achieved by the monoterpenes in the virucidal assay suggest that these compounds act directly on the viral particle.

Emerging animal viruses: real threats or simple bystanders? / Vírus emergentes de animais: verdadeiras ameaças ou meros espectadores?

The list of animal viruses has been frequently added of new members raising permanent concerns to virologists and veterinarians. The pathogenic potential and association with disease have been clearly demonstrated for some, but not for all of these emerging viruses. This review describes recent discoveries of animal viruses and their potential relevance for veterinary practice. Dogs were considered refractory to influenza viruses until 2004, when an influenza A virus subtype H3N8 was transmitted from horses and produced severe respiratory disease in racing greyhounds in Florida/USA. The novel virus, named canine influenza virus (CIV), is considered now a separate virus lineage and has spread among urban canine population in the USA. A new pestivirus (Flaviviridae), tentatively called HoBi-like pestivirus, was identified in 2004 in commercial fetal bovine serum from Brazil. Hobi-like viruses are genetically and antigenically related to bovine viral diarrhea virus (BVDV) and induce similar clinical manifestations. These novel viruses seem to be widespread in Brazilian herds and have also been detected in Southeast Asia and Europe. In 2011, a novel mosquito-borne orthobunyavirus, named Schmallenberg virus (SBV), was associated with fever, drop in milk production, abortion and newborn malformation in cattle and sheep in Germany. Subsequently, the virus disseminated over several European countries and currently represents a real treat for animal health. [...] Finally, the long time and intensive search for animal relatives of human hepatitis C virus (HCV) has led to the identification of novel hepaciviruses in dogs (canine hepacivirus [CHV]), horses (non-primate hepaciviruses [NPHV] or Theiler's disease associated virus [TDAV]) and rodents. For these, a clear and definitive association with disease is still lacking and only time and investigation will tell whether they are real disease agents or simple spectators.

O número de vírus animais cresce continuamente, causando preocupação permanente a virologistas e veterinários. O potencial patogênico e associação com doença tem sido claramente demonstrado para alguns - mas não para todos - vírus emergentes. Esse artigo apresenta uma breve revisão das recentes descobertas de vírus animais e a sua potencial relevância para saúde animal. Cães eram considerados refratários aos vírus da influenza até 2004, quando um vírus influenza A subtipo H3N8 foi transmitido de equinos e causou doença respiratória severa em cães galgos na Flórida/EUA. O novo vírus, denominado vírus da influenza canina (CIV), agora considerado uma linhagem distinta do vírus da influenza equina, disseminou-se na população canina urbana dos EUA. Um novo Pestivirus (Flaviviridae) - provisoriamente denominado pestivírus Hobi-like - foi identificado em 2004 em soro fetal bovino importado do Brasil. Os vírus Hobi-like são genética e antigenicamente relacionados com o vírus da diarreia viral bovina (BVDV) e induzem manifestações clínicas semelhantes. A sua origem e distribuição são desconhecidas, mas estão aparentemente disseminados no rebanho brasileiro e já foram identificados no sudeste asiático e na Europa. Em 2011, um novo buniavírus transmitido por mosquitos, denominado vírus Schmallemberg (SBV), foi associado com febre, redução da produção de leite, abortos e malformações fetais em bovinos e ovinos da Alemanha. [...] Finalmente, a longa e intensiva busca por vírus animais relacionados ao vírus da hepatite C humana (HCV) tem levado a identificação de "novos" pestivírus em cães (canine hepacivirus [CHV]), equinos (hepacivirus de não-primatas [NPHV] ou vírus associado à doença de Theiler [TDAV]) e em roedores. Para estes, uma associação clara e definitiva com doença ainda não foi demonstrada e apenas tempo e investigação irão dizer se são patógenos reais ou apenas espectadores.

Selection of a Less Pathogenic BVDV Strain for the Construction of Avirulent Chimeric Pestivirus

To select a less pathogenic bovine viral diarrhea virus (BVDV) strain for the construction of chimeric pestivirus harboring classical swine fever virus (CSFV) E2 gene, five Korean BVDV isolates (four type 1 isolates and a type 2 isolate) were evaluated for their pathological and biological properties with respect to porcine infection. Each of five groups of 100-day-old pigs was inoculated intranasally with one of the five BVDV isolates. No clinical sign or leukopenia was observed in any pig throughout the duration of the experiment, but viruses were detected in blood, nasal discharges and postmortem samples using RT-PCR. These results indicated that although the five BVD viruses could infect pigs, they did not cause clinically apparent symptoms. Because of its proper infection dynamics shown in this preliminary animal study and its fast growth rate and quick CPE in cell culture, one isolate (KD26-1) was chosen among the five isolates to test its virulence and immunogenic properties in 40-day-old piglets. Neither clinical sign nor pathological lesion was observed in 40-day-old piglets during the course of infection of isolate KD26-1. The first neutralizing antibodies were detectable 14 days post-inoculation (PI) and increased to 1:128~1:256 28 days PI. A BVDV specific gene was detectable by RT-PCR in tonsil, spleen, inguinal lymph node and brain until 14 days PI. According to this study, it can be concluded that isolate KD26-1 has little pathological effect in pigs and is a candidate for construction of chimeric pestivirus harboring CSFV E2 gene.

Selection of a Less Pathogenic BVDV Strain for the Construction of Avirulent Chimeric Pestivirus

To select a less pathogenic bovine viral diarrhea virus (BVDV) strain for the construction of chimeric pestivirus harboring classical swine fever virus (CSFV) E2 gene, five Korean BVDV isolates (four type 1 isolates and a type 2 isolate) were evaluated for their pathological and biological properties with respect to porcine infection. Each of five groups of 100-day-old pigs was inoculated intranasally with one of the five BVDV isolates. No clinical sign or leukopenia was observed in any pig throughout the duration of the experiment, but viruses were detected in blood, nasal discharges and postmortem samples using RT-PCR. These results indicated that although the five BVD viruses could infect pigs, they did not cause clinically apparent symptoms. Because of its proper infection dynamics shown in this preliminary animal study and its fast growth rate and quick CPE in cell culture, one isolate (KD26-1) was chosen among the five isolates to test its virulence and immunogenic properties in 40-day-old piglets. Neither clinical sign nor pathological lesion was observed in 40-day-old piglets during the course of infection of isolate KD26-1. The first neutralizing antibodies were detectable 14 days post-inoculation (PI) and increased to 1:128~1:256 28 days PI. A BVDV specific gene was detectable by RT-PCR in tonsil, spleen, inguinal lymph node and brain until 14 days PI. According to this study, it can be concluded that isolate KD26-1 has little pathological effect in pigs and is a candidate for construction of chimeric pestivirus harboring CSFV E2 gene.

Genetic characterization of Brazilian bovine viral diarrhea virus isolates by partial nucleotide sequencing of the 5'-UTR region

Nineteen isolates of bovine viral diarrhea virus (BVDV) from Brazil were genetically characterized through partial nucleotide sequencing and analysis of the 5'UTR region. The isolates were grouped as BVDV-1 (11/19), BVDV-2 (6/19) or "atypical" pestivirus (2/19). Among the BVDV-1, eight isolates were classified as subgenotype BVDV-1a, whereas most (4 out of 6) BVDV-2 belonged to subgenotype 2b. Two isolates from aborted fetuses were not classified into any genetic group, being considered atypical BVDVs. Genetic diversity among Brazilian BVDV isolates may be responsible for vaccination and diag-nostic failure and therefore may influence the control strategies for BVDV infection in the country.

Dezenove amostras do vírus da diarréia viral bovina (BVDV) foram caracterizadas geneticamente através do seqüenciamento parcial de nucleotídeos da Região 5'UTR. As amostras foram agrupadas em BVDV-1 (11/19), BVDV-2 (6/19) e num terceiro grupo de amostras denominadas "atípicas" (2/19). Das onze amostras genotipadas como BVDV-1, oito amostras foram sub-genotipadas como BVDV-1a, enquanto que a maioria (4/6) das amostras de BVDV-2 foi agrupada como BVDV-2b. Duas amostras provenientes de fetos bovinos abortados foram classificadas como atípicas, não BVDV-1 e 2. A presença da diversidade genética de BVDV detectada nas amostras estudadas pode ser responsável por falhas vacinais e de diagnóstico e deve influenciar nas estratégias de controle do BVDV aplicadas nas diferentes regiões brasileiras.

Diarrea viral bovina: patogénesis e inmunopatología / Bovine viral diarrhea: pathogenesis and inmunopathology

La diarrea viral bovina (DVB) representa un problema de ámbito mundial que causa considerables pérdidas tanto en ganado de carne como lechero, afectándolo de diversas formas las cuales están supeditadas a la edad del animal, estado inmunológico y momento de la gestación en el que se produce la infección. La DVB es causada por un virus ARN, género Pestivirus, familia Flaviviridae, el cual ha sido clasificado en 2 biotipos (citopático y no citopático) según su comportamiento en células de cultivo y en 2 genotipos (I y II) basados en su secuencia genética. Dependiendo de las cepas infectantes se presenta un cuadro clínico particular variando en severidad desde una forma subclínica, pasando por la forma clínica e incluso produciendo la fatal enfermedad de las mucosas o causando efectos deletéreos sobre el feto. A pesar de que en nuestro medio ya existen estudios sobre esta entidad, la implementación de metodologías diagnósticas constituye una limitante para el manejo de la misma. La presente revisión se enfoca en la patogenia e inmunopatología de la DVB.

The effect of infection of hatched blastocyst bovine embryos with border disease and bovine viral diarrhea viruses.

The effect of infection with teratogenic viruses at early stages of pregnancy is not fully understood. This study aimed to look at the effect of infection with teratogenic viruses such as bovine viral diarrhea virus (BVDV) and border disease virus (BDV), on early stage embryos at the hatched blastocyst stage. BVDV and BDV are known to cross the placenta of infected mothers and lead to congenital defects and death of developing fetuses. This study can be a good model for better understanding the effects of other teratogenic viruses such as Rubella virus in humans.

Establishment of Rapid Detection Methods for Bovine Viral Diarrhea Virus(Pestivirus) in Attenuated Animal Viral Vaccines by RT-PCR and Nested PCR

Bovine viral diarrhea virus (BVDV) of the genus Pestivirus is known as a common contaminant of cell culture-derived vaccines. Hog cholera virus (HCV), which is also of the genus Pestivirus and an important livestock disease in Korea, is recognized as a potential contaminant of vaccines produced in porcine cells. However, it is difficult for the National Biological Assays of korea to adequately detect contamination of these agents in biological products. For these reasons, we established rapid and sensitive methods for the detection of BVDV and HCV contamination in cell cultures and veterinary biologicals by using RT-PCR and nested PCR assays. We designed a Pestivirus primer amplifying 152 bp to detect both BVDV and HCV and a common primer amplifying 237 bp to detect only BVDV. Also, for the differentiation between BVDV type 1 and type 2, nested PCR was conducted using the amplified 237 bp PCR product, to amplify 179 bp in BVDV type 2 genome. The sensitivity of the PCR using common primer for the detection of BVDV was 400 TCID50/ml. All 6 strains of Korean BVDV isolates, 5 vaccines strains and the standard strain NADL could be detected. No reactions were observed when testing 5 types of viruses infecting pigs (HCV, TGEV, PEDV, JEV, PRRSV), 4 types infecting cattle (Akabane virus, BEFV, BCV, BRV) and 4 types infecting cats (FIP, FPL, FCV, FVR). Using this RT-PCR assay, commercial vaccines were tested and, 55 lots from 12 vaccine companies, were negative for BVDV contaminations. Same results were obtained by the immunoflourescence assay. The newly developed PCR or RT-PCR assays can be used as rapid, reliable, sensitive, and simple methods for the detection of BVDV (Pestivirus) in cell cultures, master seeds, and live viral vaccines.

Uso da fluoxetina no tratamento da tricotilomania felina / Use of fluoxetine in the treatment of feline psycogenic alopecia

A tricotilomania ou alopecia psicogênica felina é uma dermatopatia de origem psicogênica, decorrente da lambedura compulsiva do pelame, realizada pelo gato em situações de estresse. Tal distúrbio decorre de alterações neuro-hormonais e pode associar-se à introdução de novos animais e/ou crianças no ambiente. Além de mudanças de manejo e atitude para com o animal, sugere-se o emprego de ansiolíticos no tratamento da doença. A fluoxetina foi utilizada no tratamento de cinco gatos domésticos com tricotilomania, apresentando inibição do comportamento de lambedura, com repilação após dois a três meses de terapia.

Neurological disorder associated with pestivirus infection in sheep in Rio Grande do Sul, Brazil

Um cordeiro de dois meses de idade apresentando sinais severos de doença neurológica foi enviado ao Laboratório de Patologia Veterinária - Universidade Federal do Rio Grande do Sul. Os sinais clínicos consistiam de tremores musculares, hipermetria e incoordenação motora. Os principais achados de necropsia foram uma pronunciada redução do tamanho do cerebelo e dilatação bilateral dos ventrículos laterais. As alterações microscópicas consistiram de áreas de desorganização celular no córtex cerebelar, redução da camada granular do cerebelo associada com diminuição da densidade de células e a presença de grandes vacúolos citoplasmáticos na camada molecular. Cortes histológicos foram submetidos à imunoistoquímica, utilizando o anticorpo monoclonal 15C5 contra o vírus da Diarréia Viral Bovina (BVDV). Reação positiva foi detectada difusamente nos neurônios da substância cinzenta e em macrófagos dos linfonodos mesentéricos. Os achados clínicos, histopatológicos e imunoistoquímicos sugerem que este cordeiro tenha sido infectado por um pestivírus, provavelmente Border Disease Vírus (BDV) ou BVDV.

Isolation and Complete Nucleotide Analysis of a Noncytopathic BVDV Variant using a Dominant Selective Marker

Upon infection, bovine viral diarrhea virus (BVDV), one of the pestiviruses in the Flaviviridae family, is divided into two biotypes, cytopathic (cp) and noncytopathic (ncp). The mechanism of cytopathogenicity, however, is not elucidated yet. In this study, we have investigated viral genetic element affecting the cytopathogenicity of BVDV by using an infectious cDNA molecular clone containing a dominant selective marker, puromycin acetyltransferase gene (pNADL/pac). From the recombinant cDNA clone pNADL/pac, viral RNA was synthesized by T7 RNA polymerase in vitro. By selecting the MDBK cells transfected with in vitro transcribed cp NADL/pac viral RNA with puromycin, we obtained the selected MDBK cells harboring cp NADL/pac viral RNA, which did not show cytopathic effect and could be passaged. Little cytopathic effect was observed upon infection of naive MDBK cells with the viral particles released from the selected cp NADL/pac-transfected MDBK cells. Complete nucleotide analysis of viral particles released from cp NADL/pac-selected MDBK cells revealed 5 substitutions in the viral open reading frame, but not in the 5' and 3'NTRs. Interestingly, only two point mutations (G8147A and T11343C) changed their amino acid code (M2588V in NS4B and T2653M in NSSB), and the other 3 substitutions (C1627T, T3097C, and T3376C) was silent. Therefore, our results suggest that NS4B and NSSB proteins of BVDV may play a role in cytopathogenicity.

Isolamento e caracterizaçäo de cepa moderada virulência do vírus de peste suína clássica no Estado do Paraná, Brasil / Isolation and characterization of a moderate virulence strain of classical swine fever virus, in State of Paraná, Brazil

Trata o presente trabalho do isolamento viral realizado em outubro de 1997, no Laboratório de Virologia do Centro de Diagnóstico "Marcos Enrietti", de uma cepa de moderada virulência do vírus da Peste Suína Clássica (PSC), a partir de fragmentos de órgäos provenientes de granjas de suínos do município de Säo Pedro do Iguaçu, Paraná, Brasil. Foram empregadas técnicas de imunofluorescência e imunoperoxidase indiretas em cortes histológicos, ensaio imunoenzimático (ELISA) para detecçäo de antígeno viral e isolamento viral em cultivo celular da linhagem PK15 (Porcine Kidney, Sus scrofa). Os resultados positivos obtidos pelas diferentes técnicas empregadas mostraram grande precisäo para o diagnóstico da Peste Suína Clássica. Com base nesses resultados, o Serviço de Sanidade Animal recomendou o abate de cerca de 2.000 suínos em sete propriedades envolvidas no foco e a tomada de medidas zoosanitárias adequadas para prevenir a disseminaçäo da doença

Soroneutralizaçäo como teste sorológico diferencial entre infecçöes pelo vírus da peste suína clássica e outros pestivírus / Serum neutralization as a differential serological test for classical swine fever virus and other pestivirus infections

Testes de soroneutralizaçäo (SN) foram realizados para detecçäo de anticorpos contra o vírus da peste suína clássica (VPSC), da diarréia viral bovina (VDVB) e do vírus da doença da fronteira ou "border disease" dos ovinos (VBDO) em amostras de soros suínos coletadas para triagem de anticorpos antiVPSC, com o objetivo de determinar o valor da SN como prova sorológica diferencial. Noventa e nove soros de uma amostragem de 16.664 foram positivos em um teste de ELISA policlonal, incapaz de diferenciar anticorpos induzidos contra os diferentes pestivírus. Quando submetidos à SN, 81 apresentaram anticorpos somente contra o VPSC. Dezessete soros apresentaram anticorpos com reatividade cruzada contra os VPSC, VDVB ou VBDO, e na maioria deles (13/17) as diferenças entre os títulos obtidos pela SN näo permitiram inferir qual seria o vírus provável indutor dos anticorpos detectados. Concluiu-se que a prova requer cuidados ao ser usada com essa finalidade, pois quando amostras säo isoladas de soro säo examinadas a variaçäo entre os títulos de anticorpos pode ser insuficiente para permitir essa diferenciaçäo, necessitando uma amostragem que inclua um número significativo de animais dentro do criatório ou da granja suspeita

Presença de pestivírus e anticorpos contra pestivírus em soros e cultivos celulares / Presence of pestiviruses and antibodies to pestiviruses in sera and cell cultures

Cultivos celulares e soro utilizados como suplemento de meios de cultivo foram examinados em busca de pestivírus ou anticorpos contra pestivírus, por técnicas de isolamento de vírus e soro-neutralizaçäo, seguidas de revelaçäo por imunoperoxidase. Sete de um total de 13 partidas de linhagens celulares, incluindo células de origem bovina (MDBK, GBK, BT), ovina (FLK) e suína (SK6, PK15), apresentavam-se contaminadas com pestivírus. Uma linhagem celular de origem de rim de coelho (RK13) näo apresentou contaminaçäo. Duas de sete partidas de cultivos primários de testículos de neonatos bovinos preparados no local também se encontraram infectados. Soros de neonatos bovinos obtidos de abatedouros continham pestivírus em oito de 80 amostras e anticorpos anti-pestivírus em 15 de 80 amostras examinadas. Soro fetal bovino adquirido comercialmente de diferentes firmas apresentou anticorpos anti-pestivírus em quatro de oito amostras examinadas

Anticorpos contra pestivirus e herpesvirus em caprinos leiteiros no Estado de Pernambuco / Antibodies against pestivirus and herpesvirus in dairy goats in Pernambuco State, Brazil

A preliminar survey to detect antibodies against BVD/MD (NADL) and BHV1 (Los Angeles) was perfomed in five dairy goat flocks from four counties of Pernambuco State, Brazil. Two hundred and six serum samples of purebred (Saanen, Toggenburg and Alpine) and crossbred goats were tested by microplate serumneutralization test. Positive results (titre >/ 4) were recorded in 11.6 per cent and 6.8 per cent of samples test to BVD/MD and BHV 1, respectively