M. Resende

Feline Immunodeficiency Virus Subtype B in Domestic Cats in Minas Gerais, Brazil

Feline immunodeficiency virus (FIV) was isolated in 1986 from a feline leukaemia virusnegative cat with chronic opportunistic infections (Pedersen et al., 1987). Clinical signs most frequently observed in FIV infection are stomatitis, gingivitis, anaemia and neurological dysfunctions (Ishida et al., 1989; Yamamoto et al., 1989). Epidemiological surveys conducted in several parts of the world revealed that the prevalence of FIV infection in cats varies from 1% to 15% in healthy cats and from 3% to 44% in sick cats (Sukura et al., 1992; Ueland and Lutz, 1992). FIV strains isolated from domestic cats have been classified into five subtypes, designated A, B, C, D and E, by comparing nucleotide sequences of region V3–V5 of the env gene (Sodora et al., 1994; Kakinuma et al., 1995; Pecoraro et al., 1996), and similar results were also obtained when the nucleotide sequences of the gag gene were analysed (Kakinuma et al., 1995). The aim of this survey was to detect and subtype of FIV strains circulating in Minas Gerais (MG), Brazil.

Genotyping of Infectious Bursal Disease Virus Strains by Restriction Fragment Length Polymorphism Analysis of the VP1, VP2, and VP3 Genes

Abstract This study aimed to genotype infectious bursal disease virus (IBDV) isolates from the Minas Gerais state poultry industry. RNA was extracted from bursae obtained from field cases without passage or commercial vaccines. Genetic subtyping of IBDV isolates and vaccine strains was carried out by the reverse transcriptase–polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP) analysis. A 588-bp fragment in the VP1 gene, an 847-bp fragment in the VP2 gene, and a 320-bp fragment in the VP3 gene were amplified by PCR and digested with restriction enzymes PstI and ScaI (VP1); BamHI, BstEII, and PstI (VP2); and NcoI, ScaI, and XbaI (VP3). Our work shows that complementing the clinical history of the outbreaks with RT-PCR followed by RFLP analysis using PstI for VP1, BamHI for VP2, and XbaI for VP3 allowed an accurate classification of a causative agent as a very virulent IBDV.

A Labelled Avidin–Biotin ELISA to Detect Antibodies to Caprine Arthritis-encephalitis Virus in Goats' Sera

A labelled avidin–biotin ELISA (lab-ELISA) was developed and compared with indirect ELISA (i-ELISA) and agar-gel immunodiffusion assay (AGID) for its efficacy in detecting antibodies against caprine arthritis-encephalitis virus (CAEV) in goat sera. The enzyme immunoassays were standardized using 113 sera from CAEV-negative goat flocks. The tests were compared using the results from 339 serum samples. The lab-ELISA showed the greatest number of positive results (94/339) as compared with AGID (51) and i-ELISA (64). The comparison of the other two tests with the lab-ELISA showed an agreement of 87.3% with AGID and 90.6% with i-ELISA. The lab-ELISA may be useful for screening large populations for CAEV antibodies, in epidemiological surveys and in the control of caprine arthritis-encephalitis.

Naturally occurring feline leukemia virus subgroup A and B infections in urban domestic cats

A nested-PCR (n-PCR) was used to detect feline leukemia virus (FeLV) proviral DNA in blood samples from 464 sick and 608 healthy domestic cats (Felis catus) selected by convenience, and a significantly high prevalence of FeLV infection was observed. n-PCR results revealed the presence of FeLV proviral DNA in 47.2 % of sick cats and 47.4 % of healthy cats. Phylogenetic analysis revealed that FeLV samples from healthy or sick cats were grouped into separate clades. We determined FeLV subgroups by an n-PCR based on the envelope (env) gene. The partial env gene of FeLV Minas Gerais (MG) samples were compared to various exogenous FeLV isolates and endogenous (enFeLV) provirus from the same region. FeLV-B MG samples were more similar to endogenous sequences and to natural FeLV-B isolates than to either FeLV-A or FeLV-C. The results revealed the circulation of FeLV-B in large populations of urban domestic cats in Brazil.

Survey of Feline Leukemia Virus and Feline Coronaviruses in Captive Neotropical Wild Felids from Southern Brazil

Abstract A total of 57 captive neotropical felids (one Leopardus geoffroyi, 14 Leopardus pardalis, 17 Leopardus wiedii, 22 Leopardus tigrinus, and three Puma yagouaroundi) from the Itaipu Binacional Wildlife Research Center (Refúgio Bela Vista, Southern Brazil) were anesthetized for blood collection. Feces samples were available for 44 animals, including one L. geoffroyi, eight L. pardalis, 14 L. wiedii, 20 L. tigrinus, and one P. yagouaroundi. Total DNA and RNA were extracted from blood and feces, respectively, using commercial kits. Blood DNA samples were evaluated by polymerase chain reaction (PCR) for feline leukemia virus (FeLV) proviral DNA, whereas reverse transcriptase–PCR was run on fecal samples for detection of coronavirus RNA. None of the samples were positive for coronaviruses. A male L. pardalis and a female L. tigrinus were positive for FeLV proviral DNA, and identities of PCR products were confirmed by sequencing. This is the first evidence of FeLV proviral DNA in these species in Southern Brazil.

Use of an ELISA system for detection of equine herpesvirus 1 (EHV-1) antibodies in non-symptomatic pregnant mares and neonatal foals

An ELISA system was applied to study the prevalence of EHV-1 in vaccinated and non-vaccinated Brazilian horses. Serum samples were taken from non-symptomatic pregnant mares, before and after the parturition, and from their neonatal foals. The antigen was prepared from a purified virus fraction (HVE-1 AB1 strain). The ELISA absorbance of 0.3516 was determined as the threshold of negative values. This cut-off level was defined from a selected interval, which determined a sensibility of 100% and a specificity of 94.7%, from a serum collection obtained from pregnant mares and fetuses. In this study, a strong correlation was found between the ELISA and serum neutralization for specific EHV-1 antibody detection.

Neurological disorder in cattle associated with bovine herpesvirus 4

A nested PCR assay was used to diagnose bovine encephalitis through herpesviruses including bovine herpesvirus 5 (BHV-5), bovine herpesvirus 1 (BHV-1), Aujeszkys disease virus (SHV-1), and ovine herpesvirus 2 (OHV-2) in 14 fragments of central nervous system (CNS) from cattle that died with neurological signs. In addition, as some samples of bovine herpesvirus type 4 (BHV-4) have been isolated from neural tissue, it was also tested by nested PCR. The cases of encephalitis occurred in isolation at different times of the year and did not present any seasonality. The duration of the clinical course ranged between 1 to 15 days, and in 64.3% of the cases it manifested between 1 to 2 days. The most frequently observed neurological signs were ataxia, recumbency, unsteadiness and inability to stand, opisthotonus, paddling movements, nystagmus and ptyalism. In the nested assay, there was no evidence of: BHV-1, SHV-1 or OHV-2 in the DNA obtained from the CNS in any of the samples. But the presence of BHV-4 was found in all fragments of the CNS in cattle which died presenting neurological signs. Moreover, BHV-5 was found in association with BHV-4 in two of these samples.

Molecular Characterization of Contaminating Infectious Anemia Virus of Chickens in Live Commercial Vaccines Produced in the 1990s

SUMMARY. The presence of infectious chicken anemia virus (CAV) was detected in a previous study by nested-PCR as a contaminant in seven commercial vaccines, produced in the 1990s by three different manufacturers, prepared against the most relevant virus etiologies. In order to phylogenetically characterize the genome and compare it to CAV isolates from Brazil and other parts of the world, sequences of approximately 675 bp of the gene encoding the hypervariable region of VP1 protein of three CAV vaccine contaminant strains were studied. The CAV genome in contaminated vaccines showed high similarity (>98.9%) with the Brazilian BR91/99 and Argentinian ArgA001028 (>99%) strains. However, the comparison with the Cuxhaven-1 vaccine strain showed a lower identity of between 96.8% and 97.7%, and comparing it with the CAV26P4 vaccine strain showed an identity between 97.2% and 98.2%; both are available in Brazil. Such differences might be relevant for the highly conserved CAV genome. CAV contaminants were positioned in the same genetic group (clusters) with the Brazilian strain BR91/99 and Argentinian strain ArgA001028. Results indicated that the contamination of live vaccines by CAV may have influenced CAV epidemiology in the Brazilian and Argentinian poultry industry. RESUMEN. Caracterización molecular de los virus de la anemia infecciosa del pollo detectados como contaminantes en vacunas comerciales producidas en la década de los noventas. En un estudio previo, se detectó mediante un método de PCR anidado la presencia del virus de la anemia infecciosa del pollo (CAV) como contaminantes en siete vacunas comerciales contra etiologías variadas, que fueron producidas en los años noventas por tres laboratorios diferentes. Con el objetivo de caracterizar filogenéticamente el genoma y compararlo con aislamientos del virus de la anemia de Brasil y de otras partes del mundo, se estudiaron las secuencias de aproximadamente 675 pares de bases que codificaban para la región hipervariable de la proteína VP1 de tres cepas del virus de la anemia que fueron contaminantes de vacunas. El genoma del virus de la anemia que se encontró en las vacunas contaminadas mostro una similitud alta (>98.9%), con la cepa brasileña BR91/99 y con la estirpe argentina ArgA001028 (>99%). Sin embargo, la comparación con la cepa vacunal Cuxhaven-1, mostró una similitud de entre 96.8 a 97.7% y de 97.2 a 98.2% con la estirpe vacunal CAV26P4, ambas cepas son usadas en Brasil para producir vacunas comerciales contra el virus de la anemia. Estas diferencias pueden ser relevantes debido a que el genoma del virus de la anemia infecciosa es altamente conservado. Los virus contaminantes de la anemia infecciosa se agruparon en el mismo grupo genético (clúster) con la cepa brasileña BR91/99 y de Argentina ArgA001028. Estos resultados indican que la contaminación de las vacunas vivas con el virus de la anemia infecciosa pudo haber influenciado la epidemiología del virus en la industria avícola de Brasil y Argentina.

Avaliação sorológica para Toxoplasma gondii pela imunofluorescência indireta e detecção do vírus da imunodeficiência felina pela nested PCR em felinos selvagens

Nineteen sera and blood samples from wild feline kept in captivity were tested for Toxoplasma gondii antibody and presence of feline immunodeficiency virus (FIV) DNA, respectively. Eighteen (94.7%) of the them were seropositive for toxoplasma. However, the only negative animal, a Leopardus pardalis, was the only FIV positve. These results suggest that the infection by FIV may have compromised its immune system and interfered with antibody production for toxoplasma.

Phylogenetic analysis of feline immunodeficiency virus strains from State of Minas Gerais, Brazil

A regiao p17-p24 do gene gag de 10 amostras do virus da imunodeficiencia felina detectadas no estado de Minas Gerais (Brasil) foi sequenciada com o objetivo de determinar a sua classificacao molecular e a sua relacao com sequencias de amostras previamente descritas. As amostras pertenciam ao subtipo B, entretanto foi possivel observar que a maioria delas encontra-se em um subgrupo dentro do subtipo B, o que indica presenca de um possivel ancestral comum entre elas.