Immunohistochemical identification of a malignant catarrhal fever virus in cattle with renal diseases from Paraná state, Southern Brazil: a retrospective epidemiological study.

Malignant catarrhal fever (MCF) is a viral infectious disease caused by specific members of the Macavirus genus that are referred to as the MCF virus (MCFV) complex group. This study determined the prevalence of MCFV-associated infections in cattle within the mesoregions of the state of Paraná, Southern Brazil, by analyzing the histopathologic patterns of renal lesions in association with positive immunoreactivity to intralesional antigens of MCFV. Intracytoplasmic MCFV antigens were identified in 41.7% (48/115) of the kidneys of cattle evaluated. Lymphocytic interstitial nephritis, vascular degeneration, and ballooning degeneration of the renal tubules were the principal histopathological findings associated with positive immunoreactivity to MCFV. The results indicate that MCFV infections are endemic within the state of Paraná and suggest that the kidney can be of diagnostic value in suspected cases of MCF-associated infections in cattle. Furthermore, the utilization of an in situ diagnostic technique resulted in the detection of a greater number of cases of infections by MCFV than previously identified using other diagnostic methods. Additionally, degenerative vascular lesions of the kidney should be considered during the establishment of a histological diagnosis of MCFV-induced infections in cattle in the absence of fibrinoid change or necrotizing vasculitis.

The Participation of a Malignant Catarrhal Fever Virus and Mycoplasma bovis in the Development of Single and Mixed Infections in Beef and Dairy Cattle With Bovine Respiratory Disease.

The bovine respiratory disease (BRD) complex is a multietiological and multifactorial disease associated with a wide range of viral and bacterial pathogens. This study evaluated the contribution of specific infectious disease agents in the development of BRD in cattle from Brazil and determined if a virus within the malignant catarrhal fever virus (MCFV) group and Mycoplasma bovis, acting individually or in conjunction, can be associated with the development of BRD. Formalin-fixed paraffin-embedded pulmonary sections were used in immunohistochemical assays to determine the intralesional presence of six antigens associated with BRD: bovine alphaherpesvirus 1 (BoHV-1), bovine parainfluenza virus 3 (BPIV-3), bovine viral diarrhea virus (BVDV), bovine respiratory syncytial virus (BRSV), MCFV, and M. bovis. Pneumonia was diagnosed in 82.7% (120/145) of all cattle evaluated. Interstitial pneumonia (60%, 72/120) and suppurative bronchopneumonia (25.8%, 31/120) were the most frequent patterns of pneumonia identified. Intralesional antigens of MCFV (53.3%, 64/120) were the most frequently associated with BRD, followed by M. bovis (47.5%, 57/120), BVDV (42.5%, 51/120), BoHV-1 (28.3%, 34/120), BRSV (24.2%, 29/120), and BPIV-3 (8.3%, 10/120). Additionally, antigens of BVDV, MCFV, and M. bovis were the most frequently identified agents associated with singular and concomitant infections. The MCFV identified during this study is more likely to be ovine gammaherpesvirus 2 (OvHV-2), since OvHV-2 is the only MCFV identified within the geographical region of this study. Interstitial pneumonia with proliferative vascular lesions may be a useful histologic feature to differentiate MCFV-induced pneumonia from other viral pneumonias of cattle. These results demonstrate that MCFV and M. bovis, in single or mixed infections, can produce pneumonia in cattle and should therefore be considered as primary agents in the development of BRD.

A review of the epidemiological, clinical, and pathological aspects of malignant catarrhal fever in Brazil.

Sheep-associated malignant catarrhal fever (SA-MCF), the form of MCF that occurs in Brazil, is a severe, frequently fatal, infectious disease caused by ovine gammaherpesvirus-2 (OvHV-2), in which sheep are the asymptomatic hosts and cattle and other cloven-hoofed animals are the accidental hosts. This review provides a critical analysis of the historical, epidemiological aspects and the estimated economic impacts associated with SA-MCF in Brazil. Moreover, the clinical manifestations and pathological lesions associated with SA-MCF in cattle are reviewed and discussed and the phylogenetic distribution of OvHV-2 in Brazil is presented. OvHV-2 is the only MCF virus identified in animals from Brazil. It is recommended that a histopathologic diagnosis of SA-MCF be based on all aspects of vascular disease in the affected animal and not only lymphocytic/necrotizing vasculitis and/or fibrinoid change. Conformation of the intralesional participation of OvHV-2 in these alterations can be achieved by immunohistochemistry and/or in situ hybridization assays. Additionally, it is proposed that OvHV-2 should be considered as a possible infectious disease agent associated with the development of bovine respiratory disease in cattle. Furthermore, the possible role of the small intestine in the dissemination of OvHV-2 is discussed.

Auxotrophic complementation as a selectable marker for stable expression of foreign antigens in Mycobacterium bovis BCG.

Mycobacterium bovis BCG has the potential to be an effective live vector for multivalent vaccines. However, most mycobacterial cloning vectors rely on antibiotic resistance genes as selectable markers, which would be undesirable in any practical vaccine. Here we report the use of auxotrophic complementation as a selectable marker that would be suitable for use in a recombinant vaccine. A BCG auxotrophic for the amino acid leucine was constructed by knocking out the leuD gene by unmarked homologous recombination. Expression of leuD on a plasmid not only allowed complementation, but also acted as a selectable marker. Removal of the kanamycin resistance gene, which remained necessary for plasmid manipulations in Escherichia coli, was accomplished by two different methods: restriction enzyme digestion followed by re-ligation before BCG transformation, or by Cre-loxP in vitro recombination mediated by the bacteriophage P1 Cre Recombinase. Stability of the plasmid was evaluated during in vitro and in vivo growth of the recombinant BCG in comparison to selection by antibiotic resistance. The new system was highly stable even during in vivo growth, as the selective pressure is maintained, whereas the conventional vector was unstable in the absence of selective pressure. This new system will now allow the construction of potential recombinante vaccine strains using stable multicopy plasmid vectors without the inclusion of antibiotic resistance markers.

Immunogenicity of Mycobacterium bovis BCG expressing Anaplasma marginale MSP1a antigen.

Humoral and cellular immune responses of mice inoculated with recombinant Mycobacterium bovis BCG expressing the MSP1a antigen of Anaplasma marginale were evaluated. The msp1a gene was amplified by PCR and cloned into the mycobacterial expression vectors pUS2000 and pMIP12. Immunization of isogenic BALB/c mice with the rBCG/pUS2000-msp1a construct induced significant seroconversion to MSP1a (p<0.001), which was 26 times above pre-immunization levels at day 63 post-initial immunization and which remained stable for the duration of the experiment (6 months). In contrast, rBCG/pMIP12-msp1a induced seroconversion at a level of 6 times above pre-immunization values, which peaked at day 63. Western blot analysis showed that sera derived from mice vaccinated with either rBCG construct recognized both native and recombinant forms of A. marginale MSP1a. In contrast to the humoral response data, immunization with rBCG/pMIP12-msp1a was found to induce a markedly stronger cellular response than that recorded for BCG/pUS2000-msp1a. These observations clearly demonstrated the immunogenicity of recombinant BCG expressing the MSP1a antigen and suggested that the immune responses were influenced by the level of antigen expression. The results of this research warrant studies of recombinant M. bovis BCG expressing MSP1a in cattle to test for protective antibody production for control of bovine anaplasmosis.

Alterações hematológicas e sorológicas em eqüinos experimentalmente infectados com Babesia equi / Haematological and serological changes in horses experimentally infected with Babesia equi

No presente estudo säo relatadas alteraçöes hematológicas e sorológicas apresentadas por eqüinos, infectados experimentalmente com B. equi, em diferentes estágios da infecçäo e após esterilizaçäo química do parasito. Dez eqüinos, clinicamente sadios e sorologicamente negativos para Babesia spp, foram inoculados com B. equi e tratados com drogas babesicidas durante o pico de parasitemia. Após o tratamento os animais foram divididos em dois grupos: portadores, que desenvolveram a fase crônica da enfermidade e esterilizados, nos quais o parasito foi eliminado. Durante todo o experimento o hematócrito, a parasitemia e o título de anticorpos foram acompanhados, de forma a caracterizar sua dinâmica na fase aguda e na fase crônica da enfermidade, assim como após a eliminaçäo do parasito.