PESQUISA VETERINÁRIA BRASILEIRA

Abstract in English:

ABSTRACT.- Almeida S.R., Diel D.G., Rissi D.R., Weiblen R. & Flores E.F. 2008. [Clinic and pathological characterization of acute mammillitis in lactating ewes inoculated with bovine herpesvirus 2.] Caracterização clinicopatológica da mamilite aguda em ovelhas lactantes inoculadas experimentalmente com o herpesvírus bovino 2. Pesquisa Veterinária Brasileira 28(1):87-94. Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, 97105-900, Santa Maria, RS, Brazil. E-mail: flores@ccr.ufsm.br Mammillitis caused by bovine herpesvirus type 2 (BoHV-2) is an important disease in dairy herds yet its pathogenesis remains largely unknown. This report describes the reproduction and characterization of acute mammillitis in lactating ewes inoculated with BoHV-2 in the skin of the udder and teats. Five out of eight inoculated ewes developed large plaques, with focal necrosis, small vesicles and crust formation in the inoculated areas. The lesions were first observed on day 4 post-inoculation (pi), progressed in size and severity up to days 7-8pi and subsided progressively thereafter. Infectious virus was isolated from the lesions at days 7 and 8pi. Viral antigens and herpesvirus-like particles were demonstrated by electron microscopy in lesions examined at days 5, 6 and 10pi. Histological findings included epithelial necrosis, erosions and ulcers, and formation of syncytial cells. Intranuclear inclusions bodies in epithelial, syncytial and inflammatory cells and lymphoplasmacytic inflammatory infiltrate in the dermis were also observed. In a second experiment, seven out of ten lambs inoculated into the nostrils and muzzle developed nasal hyperemia and discharge, vesicles, and erosions in the nose. Infectious virus was isolated from lesions during up to three days and all lambs seroconverted to BoHV-2. Attempts to reactivate the latent infection by dexamethasone administration on day 40pi failed, since virus shedding, clinical recrudescence or seroconversion were not observed. The reproduction of acute infection and mammillitis resembling that occurring in cattle paves the way for the use of sheep to study several aspects of the biology of BoHV-2 infection.

• Bovine herpesvirus-2 BoHV-2 mammillitis sheep experimental infection

Abstract in Portuguese:

ABSTRACT.- Almeida S.R., Diel D.G., Rissi D.R., Weiblen R. & Flores E.F. 2008. [Clinic and pathological characterization of acute mammillitis in lactating ewes inoculated with bovine herpesvirus 2.] Caracterização clinicopatológica da mamilite aguda em ovelhas lactantes inoculadas experimentalmente com o herpesvírus bovino 2. Pesquisa Veterinária Brasileira 28(1):87-94. Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, 97105-900, Santa Maria, RS, Brazil. E-mail: flores@ccr.ufsm.br Mammillitis caused by bovine herpesvirus type 2 (BoHV-2) is an important disease in dairy herds yet its pathogenesis remains largely unknown. This report describes the reproduction and characterization of acute mammillitis in lactating ewes inoculated with BoHV-2 in the skin of the udder and teats. Five out of eight inoculated ewes developed large plaques, with focal necrosis, small vesicles and crust formation in the inoculated areas. The lesions were first observed on day 4 post-inoculation (pi), progressed in size and severity up to days 7-8pi and subsided progressively thereafter. Infectious virus was isolated from the lesions at days 7 and 8pi. Viral antigens and herpesvirus-like particles were demonstrated by electron microscopy in lesions examined at days 5, 6 and 10pi. Histological findings included epithelial necrosis, erosions and ulcers, and formation of syncytial cells. Intranuclear inclusions bodies in epithelial, syncytial and inflammatory cells and lymphoplasmacytic inflammatory infiltrate in the dermis were also observed. In a second experiment, seven out of ten lambs inoculated into the nostrils and muzzle developed nasal hyperemia and discharge, vesicles, and erosions in the nose. Infectious virus was isolated from lesions during up to three days and all lambs seroconverted to BoHV-2. Attempts to reactivate the latent infection by dexamethasone administration on day 40pi failed, since virus shedding, clinical recrudescence or seroconversion were not observed. The reproduction of acute infection and mammillitis resembling that occurring in cattle paves the way for the use of sheep to study several aspects of the biology of BoHV-2 infection.

Abstract in English:

ABSTRACT.- Brandão P.E., Laura Y. B. Villarreal L.Y.B., F.Gregori F., Souza S.L.P., Lopes M.A.E., Gomes C.R., Sforsin A.J., Sanches A.A., Rosales C.A.R., Richtzenhain L.J., Ferreira A.J.P. & Jerez J.A. 2007. On the etiology of an outbreak of winter dysentery in dairy cows in Brazil. Pesquisa Veterinária Brasileira 27(10):398-402. Faculdade de Medicina Veterinária e Zootecnia, Universidade de São Paulo, Av. Prof. Dr. Orlando Marques de Paiva 87, São Paulo, SP 05508-270, Brazil. E-mail: paulo7926@yahoo.com Winter dysentery (WD) is a seasonal infectious disease described worldwide that causes a marked decrease in milk production in dairy cows. In the Northern hemisphere, where the disease is classically recognized, bovine coronavirus (BCoV) has been assigned as a major etiologic agent of the disease. Nonetheless, in the Southern hemisphere, an in-deep etiological survey on WD cases had not been carried out. This study aimed to survey for BCoV by nested-RT-PCR, rotavirus by polyacrylamide gel electrophoresis (PAGE) and ELISA, bacteria by classical bacteriological methods and PCR for virulence factors and parasites by sugar flotation test on fecal samples of 21 cows from a farm during an outbreak of WD in São Paulo state, Southeastern Brazil. BCoV was detected in all 21 samples, while rotavirus was detected in two symptomatic cows. Escherichia coli, Yersinia intermedia, Providencia rustigianii Proteus penneri, Klebsiella terrigena and Enterobacter aglomerans were detected in samples from both asymptomatic and healthy cows in different associations. The study of E. coli virulence factors revealed that the strains isolated were all apathogenic. Cysts of Eimeria sp. and eggs of Strongyloidea were detected at low numbers in four of the symptomatic cows, with one co-infestation. These results suggest BCoV as the main etiologic agent of the cases of WD in Brazil, a conclusion that, with the clinical and epidemiological patterns of the disease studied herein, match those already described elsewhere. These findings give basis to the development of preventive measures and contribute to the understanding of the etiology of WD.

• Bovine coronavirus etiology winter dysentery

Abstract in Portuguese:

ABSTRACT.- Brandão P.E., Laura Y. B. Villarreal L.Y.B., F.Gregori F., Souza S.L.P., Lopes M.A.E., Gomes C.R., Sforsin A.J., Sanches A.A., Rosales C.A.R., Richtzenhain L.J., Ferreira A.J.P. & Jerez J.A. 2007. On the etiology of an outbreak of winter dysentery in dairy cows in Brazil. Pesquisa Veterinária Brasileira 27(10):398-402. Faculdade de Medicina Veterinária e Zootecnia, Universidade de São Paulo, Av. Prof. Dr. Orlando Marques de Paiva 87, São Paulo, SP 05508-270, Brazil. E-mail: paulo7926@yahoo.com Winter dysentery (WD) is a seasonal infectious disease described worldwide that causes a marked decrease in milk production in dairy cows. In the Northern hemisphere, where the disease is classically recognized, bovine coronavirus (BCoV) has been assigned as a major etiologic agent of the disease. Nonetheless, in the Southern hemisphere, an in-deep etiological survey on WD cases had not been carried out. This study aimed to survey for BCoV by nested-RT-PCR, rotavirus by polyacrylamide gel electrophoresis (PAGE) and ELISA, bacteria by classical bacteriological methods and PCR for virulence factors and parasites by sugar flotation test on fecal samples of 21 cows from a farm during an outbreak of WD in São Paulo state, Southeastern Brazil. BCoV was detected in all 21 samples, while rotavirus was detected in two symptomatic cows. Escherichia coli, Yersinia intermedia, Providencia rustigianii Proteus penneri, Klebsiella terrigena and Enterobacter aglomerans were detected in samples from both asymptomatic and healthy cows in different associations. The study of E. coli virulence factors revealed that the strains isolated were all apathogenic. Cysts of Eimeria sp. and eggs of Strongyloidea were detected at low numbers in four of the symptomatic cows, with one co-infestation. These results suggest BCoV as the main etiologic agent of the cases of WD in Brazil, a conclusion that, with the clinical and epidemiological patterns of the disease studied herein, match those already described elsewhere. These findings give basis to the development of preventive measures and contribute to the understanding of the etiology of WD.

Abstract in English:

ABSTRACT.- Silva M.S., Brum M.C.S., Weiblen R. & Flores E.F. 2007. [Identification and differentiation of herpesvirus types 1 and 5 isolated from clinical samples in central-southern Brazil, Argentina and Uruguay (1987-2006).] Identificação e diferenciação de herpesvírus bovino tipos 1 e 5 isolados de amostras clínicas no Centro-Sul do Brasil, Argentina e Uruguai (1987-2006). Pesquisa Veterinária Brasileira 27(10):403-408. Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, 97105-900 Santa Maria, RS, Brazil. E-mail: flores@ccr.ufsm.br Bovine herpesviruses types 1 and 5 (BoHV-1; BoHV-5) are genetically and antigenically closely related such they can not be distinguished by routine diagnostic tests. As BoHV-1 has been historically associated with respiratory and genital disease, herpesviruses isolated from these clinical syndromes have been tentatively – and sometimes definitively - diagnosed as BoHV-1. Likewise, cases of herpetic neurological infection in cattle have been generally attributed to BoHV-5. This study reports the identification of 40 herpesvirus isolates from different clinical specimens and syndromes in central-southern Brazil, Argentina and Uruguay (1987-2006) by the use of a PCR able to differentiate between BoHV-1 and BoHV-5. BoHV-1 isolates (n=16) were identified in cases of respiratory disease (n=3), vulvovaginitis and/or balanoposthitis (n=3), in semen of healthy bulls (n=5) and in cases of neurological disease (n=5). Viruses identified as BoHV-5 (n=24) were isolated predominantly from cases of neurological disease (n=21), but also from semen of healthy bulls (n=2) and from a spleen of a calf with systemic disease (n=1). These results show that both BoHV-1 and BoHV-5 are not strictly associated with their respective diseases; yet are frequently involved in clinical conditions otherwise attributed to the other virus. These findings also reinforce the need of correctly identifying the herpesvirus isolates as to better understand their pathogenesis and epidemiology.

• Bovine herpesvirus BoHV-1 BoHV-5 differential diagnosis epidemiology

Abstract in Portuguese:

ABSTRACT.- Silva M.S., Brum M.C.S., Weiblen R. & Flores E.F. 2007. [Identification and differentiation of herpesvirus types 1 and 5 isolated from clinical samples in central-southern Brazil, Argentina and Uruguay (1987-2006).] Identificação e diferenciação de herpesvírus bovino tipos 1 e 5 isolados de amostras clínicas no Centro-Sul do Brasil, Argentina e Uruguai (1987-2006). Pesquisa Veterinária Brasileira 27(10):403-408. Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, 97105-900 Santa Maria, RS, Brazil. E-mail: flores@ccr.ufsm.br Bovine herpesviruses types 1 and 5 (BoHV-1; BoHV-5) are genetically and antigenically closely related such they can not be distinguished by routine diagnostic tests. As BoHV-1 has been historically associated with respiratory and genital disease, herpesviruses isolated from these clinical syndromes have been tentatively – and sometimes definitively - diagnosed as BoHV-1. Likewise, cases of herpetic neurological infection in cattle have been generally attributed to BoHV-5. This study reports the identification of 40 herpesvirus isolates from different clinical specimens and syndromes in central-southern Brazil, Argentina and Uruguay (1987-2006) by the use of a PCR able to differentiate between BoHV-1 and BoHV-5. BoHV-1 isolates (n=16) were identified in cases of respiratory disease (n=3), vulvovaginitis and/or balanoposthitis (n=3), in semen of healthy bulls (n=5) and in cases of neurological disease (n=5). Viruses identified as BoHV-5 (n=24) were isolated predominantly from cases of neurological disease (n=21), but also from semen of healthy bulls (n=2) and from a spleen of a calf with systemic disease (n=1). These results show that both BoHV-1 and BoHV-5 are not strictly associated with their respective diseases; yet are frequently involved in clinical conditions otherwise attributed to the other virus. These findings also reinforce the need of correctly identifying the herpesvirus isolates as to better understand their pathogenesis and epidemiology.

Abstract in English:

ABSTRACT.- David N., Hubner S.O., Riet-Correa F., Halfen D. & Lemos R.A. 2007. Reactivation of latent bovine herpesvirus type 5 in cattle with polioencephalomalacia induced by ammonium sulphate. Pesquisa Veterinária Brasileira 27(10):435-441. Hospital Veterinário, CSTR, Universidade Federal de Campina Grande, Campus de Patos, 58700-000 Patos, PB, Brazil. E-mail: franklin.riet@pesquisador.cnpq.br In the state Mato Grosso do Sul, Brazil, outbreaks of meningoencephalitis by BoHV-5 and polioencephalomalacia (PEM) display similar epidemiological features, suggesting that meningoencephalitis may be associated with reactivation of a latent BoHV-5 infection, during the development of PEM. To test this hypothesis, four 7-8 months old steers negative for BoHV-5 antibodies were inoculated intranasally with BoHV-5 and received amprolium from day 35 to day 105 after inoculation. Because PEM was not produced during this period, ammonium sulphate was given from day 114 to day 180 after inoculation. Two uninfected control steers received amprolium and ammonium sulphate for the same periods. All inoculated cattle developed antibodies against BoHV-5 after inoculation and the virus was isolated from nasal swabs, indicating that they were infected. Two inoculated steers had clinical signs of PEM after 118 and 146 days after virus inoculation. One was euthanized after a clinical manifestation period of seven days and had severe lesions of PEM and meningoencephalitis. BoHV-5 was isolated from the central nervous system of this animal. The other animal recovered but continued to manifest chronic signs of PEM and was euthanatized. On histological examination, the cerebral cortex, caudate nucleus and thalamus had multifocal areas of malacia and mild meningoencephalitis of the cortex. BoHV-5 was not isolated from the brain. One uninfected control steer had signs of neurological disease on day 158 and had lesions of PEM without meningoencephalitis at necropsy. The simultaneous production of PEM and diffuse meningoencephalitis, with isolation of BoHV-5, in one steer treated with ammonium sulphate, 118 days after BoHV-5 inoculation, suggests that latent BoHV-5 was reactivated in this animal submitted to experimental induction of PEM.

• Polioencephalomalacia bovine herpesvirus type 5 viral reactivation cattle

Abstract in Portuguese:

ABSTRACT.- David N., Hubner S.O., Riet-Correa F., Halfen D. & Lemos R.A. 2007. Reactivation of latent bovine herpesvirus type 5 in cattle with polioencephalomalacia induced by ammonium sulphate. Pesquisa Veterinária Brasileira 27(10):435-441. Hospital Veterinário, CSTR, Universidade Federal de Campina Grande, Campus de Patos, 58700-000 Patos, PB, Brazil. E-mail: franklin.riet@pesquisador.cnpq.br In the state Mato Grosso do Sul, Brazil, outbreaks of meningoencephalitis by BoHV-5 and polioencephalomalacia (PEM) display similar epidemiological features, suggesting that meningoencephalitis may be associated with reactivation of a latent BoHV-5 infection, during the development of PEM. To test this hypothesis, four 7-8 months old steers negative for BoHV-5 antibodies were inoculated intranasally with BoHV-5 and received amprolium from day 35 to day 105 after inoculation. Because PEM was not produced during this period, ammonium sulphate was given from day 114 to day 180 after inoculation. Two uninfected control steers received amprolium and ammonium sulphate for the same periods. All inoculated cattle developed antibodies against BoHV-5 after inoculation and the virus was isolated from nasal swabs, indicating that they were infected. Two inoculated steers had clinical signs of PEM after 118 and 146 days after virus inoculation. One was euthanized after a clinical manifestation period of seven days and had severe lesions of PEM and meningoencephalitis. BoHV-5 was isolated from the central nervous system of this animal. The other animal recovered but continued to manifest chronic signs of PEM and was euthanatized. On histological examination, the cerebral cortex, caudate nucleus and thalamus had multifocal areas of malacia and mild meningoencephalitis of the cortex. BoHV-5 was not isolated from the brain. One uninfected control steer had signs of neurological disease on day 158 and had lesions of PEM without meningoencephalitis at necropsy. The simultaneous production of PEM and diffuse meningoencephalitis, with isolation of BoHV-5, in one steer treated with ammonium sulphate, 118 days after BoHV-5 inoculation, suggests that latent BoHV-5 was reactivated in this animal submitted to experimental induction of PEM.

Abstract in English:

ABSTRACT.- Claus M.P., Vivian D., Lunardi M., Alfieri A.F. & Alfieri A.A. 2007. [Phylogenetic analysis of bovine papillomavirus associated with skin warts in cattle herds from the state of Paraná.] Análise filogenética do papilomavírus bovino associado a lesões cutâneas em rebanhos do Estado do Paraná. Pesquisa Veterinária Brasileira 27(7):314-318. Laboratório de Virologia Animal, Departamento de Medicina Veterinária Preventiva, Centro de Ciências Agrárias, Universidade Estadual de Londrina, Cx. Postal 6001, Campus Universitário, Londrina, PR 86051-990, Brazil. E-mail: alfieri@uel.br Bovine papillomavirus (BPV) infection causes hyperplastic lesions in the cutaneous epithelium of cattle. Six types of BPV were classified in two sub-groups, being correlated to the anatomical regions of the infection and morphologic characteristics of the lesions. The present study was carried out to identify the types of BPV present in skin warts of cattle from the state of Paraná, Brazil. The generic primers FAP59 and FAP64 were used for amplification of a 478 bp fragment of BPV L1 gene in nine cutaneous papilloma samples obtained from six animals in four herds. In all papillomas examined, a product with the expected molecular size was amplified. Phylogenetic analysis of the PCR products identified BPV-2 in three samples, BPV-1 in one, and BPV-6 in five papillomas. BPV-6 was detected in cutaneous papillomas of the teat and in other body parts as well. In one animal, from which more than one sample was collected, a concomitant infection by BPV-1 and BPV-2 was identified. The five positive BPV-6 samples showed a nucleotide identity of 100% with the sequence of the reference strain available in GenBank. However, differences among BPV-2 and BPV-1 Brazilian samples and the respective reference sequences deposited in GenBank were observed. Molecular comparison of the two BPV-2 strains identified showed the involvement of two viral variants. This study revealed the diversity of BPV types circulating in the state of Paraná.

• Bovine papillomavirus cutaneous papillomatosis phylogeny polymerase chain reaction

Abstract in Portuguese:

ABSTRACT.- Claus M.P., Vivian D., Lunardi M., Alfieri A.F. & Alfieri A.A. 2007. [Phylogenetic analysis of bovine papillomavirus associated with skin warts in cattle herds from the state of Paraná.] Análise filogenética do papilomavírus bovino associado a lesões cutâneas em rebanhos do Estado do Paraná. Pesquisa Veterinária Brasileira 27(7):314-318. Laboratório de Virologia Animal, Departamento de Medicina Veterinária Preventiva, Centro de Ciências Agrárias, Universidade Estadual de Londrina, Cx. Postal 6001, Campus Universitário, Londrina, PR 86051-990, Brazil. E-mail: alfieri@uel.br Bovine papillomavirus (BPV) infection causes hyperplastic lesions in the cutaneous epithelium of cattle. Six types of BPV were classified in two sub-groups, being correlated to the anatomical regions of the infection and morphologic characteristics of the lesions. The present study was carried out to identify the types of BPV present in skin warts of cattle from the state of Paraná, Brazil. The generic primers FAP59 and FAP64 were used for amplification of a 478 bp fragment of BPV L1 gene in nine cutaneous papilloma samples obtained from six animals in four herds. In all papillomas examined, a product with the expected molecular size was amplified. Phylogenetic analysis of the PCR products identified BPV-2 in three samples, BPV-1 in one, and BPV-6 in five papillomas. BPV-6 was detected in cutaneous papillomas of the teat and in other body parts as well. In one animal, from which more than one sample was collected, a concomitant infection by BPV-1 and BPV-2 was identified. The five positive BPV-6 samples showed a nucleotide identity of 100% with the sequence of the reference strain available in GenBank. However, differences among BPV-2 and BPV-1 Brazilian samples and the respective reference sequences deposited in GenBank were observed. Molecular comparison of the two BPV-2 strains identified showed the involvement of two viral variants. This study revealed the diversity of BPV types circulating in the state of Paraná.

Abstract in English:

ABSTRACT.- Rissi D.R., Rech R.R., Flores E.F., Kommers G.D. & Barros C.S.L. 2007. [Meningoencephalitis by bovine herpesvirus-5.] Meningoencefalite por herpesvírus bovino-5. Pesquisa Veterinária Brasileira 27(7):251-260. Departamento de Patologia, Universidade Federal de Santa Maria, 97105-900, Santa Maria, RS, Brazil. E-mail: claudioslbarros@uol.com.br Meningoencephalitis caused by bovine herpesvirus-5 (BoHV-5) is an often fatal, acute or subacute infectious disease that affects mainly young cattle under stressing conditions. The disease has been recognized in several Brazilian regions and in other parts of the world. BoHV-5 is a double stranded DNA virus member of the Herpesviridae family and subfamily Alphaherpesvirinae. The virus is characterized by rapid and lytic replication in cell cultures and by the ability to establish lifelong latent infection in sensory nerve ganglia of the host. BoHV-5 is transmitted mainly by direct and indirect contact and replicates acutely in the oral, nasal, oropharingeal or ocular mucosae. After primary replication, the virus invades nerve endings and is transported to the neuron cell bodies of the sensory ganglia where it replicates actively and/or establishes latency. Viral invasion of the brain may result in massive virus replication and production of neurological disease. Virtually all cattle developing neurological disease die of meningoencephalitis; yet the infection may be subclinical in some animals. These animals recover and become latently infected. Viral dissemination within a herd is facilitated by conditions such as crowding, introduction of cattle from other herds and weaning of calves in ages that coincide with decrease of passive immunity. Certain natural or induced conditions may reactivate the latent virus and favor its transmission and dissemination to other susceptible individuals. The disease may occur as outbreaks or as sporadic cases, with morbidity rates ranging of 0.05%-5%; lethality is almost always 100%. Clinical signs include depression, nasal and ocular discharge, grinding of teeth, circling, blindness, fever, paddling movements, disphagia, abdominal pain, nystagmus, muscle tremors, drooling, incoordinated gait, opisthotonus, head pressing, falls and convulsions. Clinical course is usually 1-15 days. Necropsy findings may be absent but often there is swollen of the rostral portions of the cerebral cortex and flattening of gyri, with softening and segmental yellow discoloration (malacia). As the disease progresses the affected areas become gelatinous and grey and, in advanced cases, there is segmental loss of the cerebral cortex of the frontal lobe of the brain (residual lesion). In several cases there is malacia of the basal nuclei and of the thalamus. Histologically, there is necrotizing non-suppurative meningoencephalitis affecting mainly the cerebral cortex of the frontal lobe associated with eosinophilic intranuclear inclusion bodies in neurons and astrocytes, although the frequency of the inclusion bodies is inconsistent. The diagnosis of meningoencephalitis by BoHV-5 should be based on epidemiology, clinical signs, necropsy and histological findings. The diagnosis should be confirmed by viral isolation in cell culture and/or by detection of viral antigens in brain sections or in exfoliated cells from nasal secretions. The identification and characterization of BoHV-5 can be done by the use of monoclonal antibodies, polymerase chain reaction (PCR) and/or by restriction enzyme analysis of the viral genome. There is no specific treatment for the disease. As BoHV-1 and BoHV-5 are antigenically related, vaccination using BoHV-1 vaccines may be recommended as a means of reducing the losses caused by BoHV-5 infection, mainly during outbreaks of neurologic disease. Additionally, measures such as serologic testing of new additions to the herd; and management practices to prevent stress and to reduce conditions for virus dissemination among animals may help in reducing the incidence and the consequences of BoHV-5 infection and disease.

• Bovine herpesvirus-5 BoHV-5 BHV-5 meningoencephalitis viral diseases diseases of cattle neuropathology

Abstract in Portuguese:

ABSTRACT.- Rissi D.R., Rech R.R., Flores E.F., Kommers G.D. & Barros C.S.L. 2007. [Meningoencephalitis by bovine herpesvirus-5.] Meningoencefalite por herpesvírus bovino-5. Pesquisa Veterinária Brasileira 27(7):251-260. Departamento de Patologia, Universidade Federal de Santa Maria, 97105-900, Santa Maria, RS, Brazil. E-mail: claudioslbarros@uol.com.br Meningoencephalitis caused by bovine herpesvirus-5 (BoHV-5) is an often fatal, acute or subacute infectious disease that affects mainly young cattle under stressing conditions. The disease has been recognized in several Brazilian regions and in other parts of the world. BoHV-5 is a double stranded DNA virus member of the Herpesviridae family and subfamily Alphaherpesvirinae. The virus is characterized by rapid and lytic replication in cell cultures and by the ability to establish lifelong latent infection in sensory nerve ganglia of the host. BoHV-5 is transmitted mainly by direct and indirect contact and replicates acutely in the oral, nasal, oropharingeal or ocular mucosae. After primary replication, the virus invades nerve endings and is transported to the neuron cell bodies of the sensory ganglia where it replicates actively and/or establishes latency. Viral invasion of the brain may result in massive virus replication and production of neurological disease. Virtually all cattle developing neurological disease die of meningoencephalitis; yet the infection may be subclinical in some animals. These animals recover and become latently infected. Viral dissemination within a herd is facilitated by conditions such as crowding, introduction of cattle from other herds and weaning of calves in ages that coincide with decrease of passive immunity. Certain natural or induced conditions may reactivate the latent virus and favor its transmission and dissemination to other susceptible individuals. The disease may occur as outbreaks or as sporadic cases, with morbidity rates ranging of 0.05%-5%; lethality is almost always 100%. Clinical signs include depression, nasal and ocular discharge, grinding of teeth, circling, blindness, fever, paddling movements, disphagia, abdominal pain, nystagmus, muscle tremors, drooling, incoordinated gait, opisthotonus, head pressing, falls and convulsions. Clinical course is usually 1-15 days. Necropsy findings may be absent but often there is swollen of the rostral portions of the cerebral cortex and flattening of gyri, with softening and segmental yellow discoloration (malacia). As the disease progresses the affected areas become gelatinous and grey and, in advanced cases, there is segmental loss of the cerebral cortex of the frontal lobe of the brain (residual lesion). In several cases there is malacia of the basal nuclei and of the thalamus. Histologically, there is necrotizing non-suppurative meningoencephalitis affecting mainly the cerebral cortex of the frontal lobe associated with eosinophilic intranuclear inclusion bodies in neurons and astrocytes, although the frequency of the inclusion bodies is inconsistent. The diagnosis of meningoencephalitis by BoHV-5 should be based on epidemiology, clinical signs, necropsy and histological findings. The diagnosis should be confirmed by viral isolation in cell culture and/or by detection of viral antigens in brain sections or in exfoliated cells from nasal secretions. The identification and characterization of BoHV-5 can be done by the use of monoclonal antibodies, polymerase chain reaction (PCR) and/or by restriction enzyme analysis of the viral genome. There is no specific treatment for the disease. As BoHV-1 and BoHV-5 are antigenically related, vaccination using BoHV-1 vaccines may be recommended as a means of reducing the losses caused by BoHV-5 infection, mainly during outbreaks of neurologic disease. Additionally, measures such as serologic testing of new additions to the herd; and management practices to prevent stress and to reduce conditions for virus dissemination among animals may help in reducing the incidence and the consequences of BoHV-5 infection and disease.

Abstract in English:

ABSTRACT.- Cazerta S.M.M., Miglino M.A., Marques R.S., Vulcano M. & Pereira F.T.V. 2007. [Characterization of hemophagous areas of the bovine placenta.] Caracterização das áreas hemófagas da placenta bovina. Pesquisa Veterinária Brasileira 26(6):229-235. Faculdade de Zootecnia, Campus Dracena, Unesp, Rod. Com. João Ribeiro de Barros, SP294 Km 651, Dracena, SP 17900-000, Brazil. E-mail: fverechia@dracena.unesp.br The specific region of maternal fetal interface needs to be clarified and corresponds to the “arcade zone” of sheep and goat placentomes. In small ruminants that area is also characterized by macroscopic blood extravasation (hemophagous areas). This occurs possibly because the iron is transferred to the embryo by trophoblastic erytrophagocytosis in these hemophagous placental areas. In order to investigate the hemophagous placental areas in cattle, placentomes of 34 pregnant Bos indicus cows (2-3, 4-6, 7-8 and 9 months of gestation) were analyzed. They were fixed by perfusion with 10% formaldehyde aqueous solution and 4% paraformoldehyde in PBS, pH 7.4, 0.1M, processed and stained for light microscopy and histochemistry. The methodology used allowed to observe placental haematomes between the uterine and trophoblastic epithelium since 3 months of pregnancy. Erythrocytes had been found in the trophoblastic cells, elucidating the erytrophagocytosis. Through Perl’s histochemical reaction it was possible to prove the existence of ferric iron in the trophoblastic cells. The PAS reaction was positive staining mucoid substance in the epithelial cells, especially in the binucleate cells of the fetal epithelium. Based on histology and histochemistry, it can be inferred that the hemophagous areas are important sites for iron transfer in the bovine placenta.

• Erythrophagocytosis hemophagous areas bovine placenta reproduction trophoblast

Abstract in Portuguese:

ABSTRACT.- Cazerta S.M.M., Miglino M.A., Marques R.S., Vulcano M. & Pereira F.T.V. 2007. [Characterization of hemophagous areas of the bovine placenta.] Caracterização das áreas hemófagas da placenta bovina. Pesquisa Veterinária Brasileira 26(6):229-235. Faculdade de Zootecnia, Campus Dracena, Unesp, Rod. Com. João Ribeiro de Barros, SP294 Km 651, Dracena, SP 17900-000, Brazil. E-mail: fverechia@dracena.unesp.br The specific region of maternal fetal interface needs to be clarified and corresponds to the “arcade zone” of sheep and goat placentomes. In small ruminants that area is also characterized by macroscopic blood extravasation (hemophagous areas). This occurs possibly because the iron is transferred to the embryo by trophoblastic erytrophagocytosis in these hemophagous placental areas. In order to investigate the hemophagous placental areas in cattle, placentomes of 34 pregnant Bos indicus cows (2-3, 4-6, 7-8 and 9 months of gestation) were analyzed. They were fixed by perfusion with 10% formaldehyde aqueous solution and 4% paraformoldehyde in PBS, pH 7.4, 0.1M, processed and stained for light microscopy and histochemistry. The methodology used allowed to observe placental haematomes between the uterine and trophoblastic epithelium since 3 months of pregnancy. Erythrocytes had been found in the trophoblastic cells, elucidating the erytrophagocytosis. Through Perl’s histochemical reaction it was possible to prove the existence of ferric iron in the trophoblastic cells. The PAS reaction was positive staining mucoid substance in the epithelial cells, especially in the binucleate cells of the fetal epithelium. Based on histology and histochemistry, it can be inferred that the hemophagous areas are important sites for iron transfer in the bovine placenta.

Abstract in English:

ABSTRACT.- Curci V.C.M., Dutra I.S., Döbereiner J. & Lucas Junior J. 2007. [The effect of composting cattle carcasses on botulinum spores and toxin content.] Pré-compostagem de cadáveres de bovinos acometidos pelo botulismo. Pesquisa Veterinária Brasileira 27(4):157-161. Unidade de Pesquisa e Desenvolvimento Tecnológico do Agronegócio, APTA Regional Extremo Oeste, Laboratório de Sanidade Animal, Av. Alcides Fagundes Chagas 122, Araçatuba, SP 15055-240, Brazil. E-mail: vlmcursi@aptaregional.sp.gov.br Five carcasses of cattle that died from botulism were composted in individual isolated beds with the purpose of monitoring the effect on spores of Clostridium botulinum and of botulinum toxin. The diagnosis of botulinum poisoning was based on clinical and pathological, epidemic and/or laboratorial characteristics. Of the five bovines with clinical signs of chronic botulism, three were affected by the type D toxin, one by the CD complex, and one was negative for toxin and spores of C. botulinum in the viscera using a mouse bioassay. The composting process was carried out in beds with carcasses cut up and mixed with sawdust as a carbon source. They were kept anaerobic under a plastic sheet for 50 days. The temperature of the beds was monitored during the period and oscillated from 40.5 to 52.4°C. After this time, soft tissue had complete decomposed with a significant loss of weight (26.5-44.5%). The remaining bones no longer contained spores or botulinum toxin in the 5 samples taken from each carcass. Only 2 of the 200 samples of homogenized remaining (40 for each carcass) contained spores of C. botulinum type C. All samples were negative for botulinum toxin using the mouse bioassay. Composting carcasses of cattle that died from botulism eliminated botulinum spores and toxin from the remains.

• Composting bovine carcasses botulism botulinum toxin C and D Clostridium botu-linum

Abstract in Portuguese:

ABSTRACT.- Curci V.C.M., Dutra I.S., Döbereiner J. & Lucas Junior J. 2007. [The effect of composting cattle carcasses on botulinum spores and toxin content.] Pré-compostagem de cadáveres de bovinos acometidos pelo botulismo. Pesquisa Veterinária Brasileira 27(4):157-161. Unidade de Pesquisa e Desenvolvimento Tecnológico do Agronegócio, APTA Regional Extremo Oeste, Laboratório de Sanidade Animal, Av. Alcides Fagundes Chagas 122, Araçatuba, SP 15055-240, Brazil. E-mail: vlmcursi@aptaregional.sp.gov.br Five carcasses of cattle that died from botulism were composted in individual isolated beds with the purpose of monitoring the effect on spores of Clostridium botulinum and of botulinum toxin. The diagnosis of botulinum poisoning was based on clinical and pathological, epidemic and/or laboratorial characteristics. Of the five bovines with clinical signs of chronic botulism, three were affected by the type D toxin, one by the CD complex, and one was negative for toxin and spores of C. botulinum in the viscera using a mouse bioassay. The composting process was carried out in beds with carcasses cut up and mixed with sawdust as a carbon source. They were kept anaerobic under a plastic sheet for 50 days. The temperature of the beds was monitored during the period and oscillated from 40.5 to 52.4°C. After this time, soft tissue had complete decomposed with a significant loss of weight (26.5-44.5%). The remaining bones no longer contained spores or botulinum toxin in the 5 samples taken from each carcass. Only 2 of the 200 samples of homogenized remaining (40 for each carcass) contained spores of C. botulinum type C. All samples were negative for botulinum toxin using the mouse bioassay. Composting carcasses of cattle that died from botulism eliminated botulinum spores and toxin from the remains.

Abstract in English:

ABSTRACT.- Mathias L.A., Meirelles R.B. & Buchala F.G. 2007. [Stability of Brucella abortus whole cell antigen for use in the serological diagnosis of bovine brucellosis by the complement fixation test.] Estabilidade do antígeno de célula total de Brucella abortus para uso no diagnóstico sorológico da brucelose bovina pela reação de fixação de complemento. Pesquisa Veterinária Brasileira 27(1):18-22. Departamento de Medicina Veterinária Preventiva e Reprodução Animal, Faculdade de Ciências Agrárias e Veterinárias (FCAV), Universidade Estadual Paulista (Unesp), Jaboticabal, SP 14884-900, Brazil. E-mail: lmathias@fcav.unesp.br The complement fixation test is used worldwide in the confirmatory diagnosis of bovine brucellosis. For this technique the antigen is the same as the one used in the tube agglutination test. However, literature is poor in information about the stability of the whole cell Brucella antigen for use in the complement fixation test to establish a time of validity of the antigen. Hence the aim of this investigation was to evaluate the stability of this antigen under refrigeration for use in the complement fixation test. Fourteen batches of antigen prepared with Brucella abortus strain 1119/3, produced from 9 months to 23 years and 11 months before, were analysed. One hundred and sixty-seven cattle sera with varying titres of antibodies to Brucella were tested through the warm complement fixation microtechnique with five 50% haemolytic units of complement. Sera with at least 25% of complement fixation in dilution 1:4 were considered positive. The results with 13 of the antigen batches were compared with the results obtained with the batch produced 9 months before by the McNemar c2 test and kappa statistic. The oldest antigen batch gave a higher proportion of sera titres which were exactly the same observed with the 9-month-batch (90.4%), and the antigen produced 4 years and 3 months before the test gave de lowest proportion of sera with the same titre of the 9-month-antigen (73.7%). The comparison of the results after being classified as positive and negative showed that the highest proportion of agreed results was observed with the antigen produced 21 years and 4 months before (98.8%, kappa 0.98). The antigen with the lowest proportion of agreed results was the one produced 3 years and 2 months before (91.6%, kappa 0.84). The results of the study show that most sera gave very similar results with all antigen batches evaluated, and that there was no relationship between the period of antigen production and the difference in test results.

• Bovine brucellosis serological diagnosis complement fixation test

Abstract in Portuguese:

ABSTRACT.- Mathias L.A., Meirelles R.B. & Buchala F.G. 2007. [Stability of Brucella abortus whole cell antigen for use in the serological diagnosis of bovine brucellosis by the complement fixation test.] Estabilidade do antígeno de célula total de Brucella abortus para uso no diagnóstico sorológico da brucelose bovina pela reação de fixação de complemento. Pesquisa Veterinária Brasileira 27(1):18-22. Departamento de Medicina Veterinária Preventiva e Reprodução Animal, Faculdade de Ciências Agrárias e Veterinárias (FCAV), Universidade Estadual Paulista (Unesp), Jaboticabal, SP 14884-900, Brazil. E-mail: lmathias@fcav.unesp.br The complement fixation test is used worldwide in the confirmatory diagnosis of bovine brucellosis. For this technique the antigen is the same as the one used in the tube agglutination test. However, literature is poor in information about the stability of the whole cell Brucella antigen for use in the complement fixation test to establish a time of validity of the antigen. Hence the aim of this investigation was to evaluate the stability of this antigen under refrigeration for use in the complement fixation test. Fourteen batches of antigen prepared with Brucella abortus strain 1119/3, produced from 9 months to 23 years and 11 months before, were analysed. One hundred and sixty-seven cattle sera with varying titres of antibodies to Brucella were tested through the warm complement fixation microtechnique with five 50% haemolytic units of complement. Sera with at least 25% of complement fixation in dilution 1:4 were considered positive. The results with 13 of the antigen batches were compared with the results obtained with the batch produced 9 months before by the McNemar c2 test and kappa statistic. The oldest antigen batch gave a higher proportion of sera titres which were exactly the same observed with the 9-month-batch (90.4%), and the antigen produced 4 years and 3 months before the test gave de lowest proportion of sera with the same titre of the 9-month-antigen (73.7%). The comparison of the results after being classified as positive and negative showed that the highest proportion of agreed results was observed with the antigen produced 21 years and 4 months before (98.8%, kappa 0.98). The antigen with the lowest proportion of agreed results was the one produced 3 years and 2 months before (91.6%, kappa 0.84). The results of the study show that most sera gave very similar results with all antigen batches evaluated, and that there was no relationship between the period of antigen production and the difference in test results.

Abstract in English:

ABSTRACT.- Cortez A., Heinemann M.B., Castro A.M.M.G., Soares M.S, Pinto A.M.V., Alfieri A.A., Flores E.F., Leite R.C. & Richtzenhain L.J. 2006. Genetic characterization of Brazilian bovine viral diarrhea virus isolates by partial nucleotide sequencing of the 5’-UTR region. Pesquisa Veterinária Brasileira 26(4):211-216. Faculdade de Medicina Veterinária e Zootecnia, Universidade de São Paulo, Av. Prof. Dr. Orlando Marques de Paiva 87, Cidade Universitária, São Paulo, SP 05508-000, Brazil. E-mail: leonardo@usp.br 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.

• BVDV-1 BVDV-2 Pestivirus phylogenetic analysis

Abstract in Portuguese:

ABSTRACT.- Cortez A., Heinemann M.B., Castro A.M.M.G., Soares M.S, Pinto A.M.V., Alfieri A.A., Flores E.F., Leite R.C. & Richtzenhain L.J. 2006. Genetic characterization of Brazilian bovine viral diarrhea virus isolates by partial nucleotide sequencing of the 5’-UTR region. Pesquisa Veterinária Brasileira 26(4):211-216. Faculdade de Medicina Veterinária e Zootecnia, Universidade de São Paulo, Av. Prof. Dr. Orlando Marques de Paiva 87, Cidade Universitária, São Paulo, SP 05508-000, Brazil. E-mail: leonardo@usp.br 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.