Fernanda Silveira Flores Vogel

Distribution of bovine herpesvirus type 5 DNA in the central nervous systems of latently, experimentally infected calves.

ABSTRACT Bovine herpesvirus type 5 (BHV-5) is an alphaherpesvirus associated with meningoencephalitis, a disease highly prevalent in South America. In this study, we investigated the distribution of BHV-5 DNA in the brains of latently, experimentally infected calves by using a PCR for the glycoprotein B gene. Twelve calves inoculated intranasally with a Brazilian BHV-5 isolate were divided into two groups: group A calves (n = 4) were euthanized 55 days postinoculation (p.i.) for tissue collection; group B calves (n = 8) were submitted to dexamethasone administration at day 60 p.i. for reactivation of latent infection and were euthanized 50 days later. Latent infection was reactivated in all group B calves, as demonstrated by virus isolation from nasal secretions and/or seroconversion. Three calves developed neurological disease and died or were euthanized in extremis. For group A calves, viral DNA was consistently detected in the trigeminal ganglia (4/4), midbrain (4/4), thalamus (4/4), and olfactory cortex (4/4) and less frequently in the pons (3/4), cerebellum (3/4), anterior cerebral cortex (2/4), and olfactory bulb (2/4). For calves previously submitted to reactivation (group B), viral DNA was detected with roughly the same frequency in the same areas as for the group A calves. In addition, viral DNA was detected in the posterior (5/5) and dorso-lateral cortex (3/5). All DNA-positive tissues were negative for infectivity and viral antigens. These results demonstrated that latent BHV-5 DNA is present in several areas of the brain during latent infection and that virus reactivation may result in the establishment of latent infection in additional sites of the brain.

Multiple resistance of gastrointestinal nematodes to nine different drugs in a sheep flock in southern Brazil

Based on clinical observation of a flock of approximately 5000 sheep (breed Merino) from southern Brazil, the failure of anthelmintic treatments was suspected. The sensitivity of the gastrointestinal nematodes that infected these sheep to nine drugs (Levamisole, Moxidectin, Albendazole, Ivermectin, Nitroxynil, Disophenol, Trichlorphon, Closantel, and a combination of Ivermectin, Levamisole and Albendazole) was evaluated. None of the drugs reduced the count of nematode eggs per gram of feces (EPG) in the sheep to an adequate extent (defined as a 95% reduction in EPG). Therefore, we found multiple parasite resistance to all classes of anthelmintic drugs available for small ruminants in the Brazilian market. The genera Haemonchus spp., Trichostrongylus spp. and Ostertagia spp. had various degrees of resistance (or natural tolerance) to each of the drug treatments.

A infecção pelo vírus da diarréia viral bovina (BVDV) no Brasil: histórico, situação atual e perspectivas

Bovine viral diarrhea virus (BVDV) is one of the most important pathogens of cattle worldwide. BVDV infection and associated diseases have been reported in Brazil since the late 1960ties. Several serological, virological, clinical and pathological reports demonstrate the widespread distribution of BVDV infection among Brazilian cattle. In addition to variable levels of positive serology in beef and dairy cattle, BVDV antibodies have been occasionally detected in swine, wild boars, goats, cervids and water buffaloes. BVDV infection has been diagnosed in aborted fetuses, buffy coats of persistently infected (PI) animals, clinical specimens from animals suffering from different clinical syndromes, semen of bulls of artificial insemination (AI) centers, in healthy fetuses and in commercial fetal bovine serum and/or cultured cells. About 50 isolates have been genetically and/or antigenically characterized up to date, whilst roughly an equivalent number of isolates awaits characterization. Most of the characterized isolates belong to BVDV-1 genotype, non-cytopathic (NCP) biotype, yet some BVDV-2 (and some CP BVDV) have been identified as well. Brazilian BVDV isolates display a high antigenic variability and are markedly different from North American vaccine strains. A few inactivated, polyvalent vaccines are currently licensed in the country, yet vaccination is still incipient in many regions: only about 2.5 million doses were marketed in 2003. The low serological cross-reactivity between vaccine strains and field isolates has recently stimulated national industries to develop vaccines containing Brazilian BVDV-1 and BVDV-2 strains. The overall knowledge about BVDV infection in Brazil has grown considerably in the last years, due to an increasing number of laboratories performing diagnosis and research. Studies on the pathogenesis, serological and molecular epidemiology and production of reagents for diagnosis have contributed decisively for the recent growing knowledge on BVDV infections in the country.Bovine viral diarrhea virus (BVDV) is one of the most important pathogens of cattle worldwide. BVDV infection and associated diseases have been reported in Brazil since the late 1960ties. Several serological, virological, clinical and pathological reports demonstrate the widespread distribution of BVDV infection among Brazilian cattle. In addition to variable levels of positive serology in beef and dairy cattle, BVDV antibodies have been occasionally detected in swine, wild boars, goats, cervids and water buffaloes. BVDV infection has been diagnosed in aborted fetuses, buffy coats of persistently infected (PI) animals, clinical specimens from animals suffering from different clinical syndromes, semen of bulls of artificial insemination (AI) centers, in healthy fetuses and in commercial fetal bovine serum and/or cultured cells. About 50 isolates have been genetically and/or antigenically characterized up to date, whilst roughly an equivalent number of isolates awaits characterization. Most of the characterized isolates belong to BVDV-1 genotype, non-cytopathic (NCP) biotype, yet some BVDV-2 (and some CP BVDV) have been identified as well. Brazilian BVDV isolates display a high antigenic variability and are markedly different from North American vaccine strains. A few inactivated, polyvalent vaccines are currently licensed in the country, yet vaccination is still incipient in many regions: only about 2.5 million doses were marketed in 2003. The low serological cross-reactivity between vaccine strains and field isolates has recently stimulated national industries to develop vaccines containing Brazilian BVDV-1 and BVDV-2 strains. The overall knowledge about BVDV infection in Brazil has grown considerably in the last years, due to an increasing number of laboratories performing diagnosis and research. Studies on the pathogenesis, serological and molecular epidemiology and production of reagents for diagnosis have contributed decisively for the recent growing knowledge on BVDV infections in the country.

Óleo de citronela no controle do carrapato de bovinos

This research was aimed at evaluating in vitro and in vivo effects of citronella (Cymbopogon nardus (L.) Rendle) oil on catle ticks. The volatile oil was obtained from de fresh leaves of citronella grass by steam destilation and the yelds was 0.7%. Seven (0; 0.5; 1.0; 10.0; 25.0; 50.0; 100.0%) and nine (0.1; 0.25; 0.5; 1.0; 2.0; 10.0; 25.0; 50.0; 100.0%) concentrations of citonella oil were used on in vitro trials with engorged female ticks. The efficacy control ticks was 0; 44.2; 92.1; 85.6; 87.8; 87.0; 88.9 and 0; 0.7; 2.8; 51.6; 79.3; 81.0; 87.1; 86.7 89.5 %, respectively. These sugest that the citronella oil can be practical used for controlling of catle ticks.

Anticorpos anti-Neospora caninum em bovinos, ovinos e bubalinos no Estado do Rio Grande do Sul

The infection by Neospora caninum is distributed worldwide and has been considered an important cause of abortion in cattle, which are intermediate hosts of the parasite. The present article reports an serological survey of the N.caninum infection in 1024 serum samples of cattle, sheep and water buffalo from 55 herds in 16 counties of the state of Rio Grande do Sul (RS). Antibodies to the agent were detected by ELISA in 11.4% (89/781) bovine samples, in 14.6% (24/164) water buffalo and in 3.2% (2/62) sheep sera. Positive cattle were detected in all tested counties. These results demonstrate that N. caninum infection is widespread among bovine and other ruminants in the state. Taken together with previous clinical and pathological reports, these results are indicative of the importance of the parasite as the etiological agent of reproductive failure in cattle in RS.

Eficiência in vitro de acaricidas sobre carrapatos de bovinos no Estado do Rio Grande do Sul, Brasil

Infestation by ticks is responsible for significant economic losses to the cattle industry in several regions, including Rio Grande do Sul (RS) state. Losses may be derived from animal stress, weight loss and skin damage, as well as from costs with treatment. The indiscriminated use of pesticides against these parasites has resulted in high levels of genetic resistance to several drugs, representing a problem for disease control. Thus, the objective of this research was to evaluate the susceptibility of field specimens of Rhipcephalus (Boophilus) microplus to the acaricides used in the control of the parasite. Female ticks obtained from 42 farms in different counties of RS were submitted to the engorgeted female immersion test, to estimate the resistance of the parasite to the drugs. Amitraz in combination with chlorpyriphos was the most efficient drug, acting effectively in ticks from 100% of the tested farms (11/11). Associations of cypermethrin-chlorpyriphos-citronelol were efficient in ticks from 61% of the farms (25/41) and cypermethrin-ethion in 37% (10/27). Cypermethrin alone was efficient in 20,7% (6/29) and Amitraz. one of the most used products, was efficient in ticks from only 14,2% (6/42) of the farms. These results demonstrate that most drugs used in the control of R. B. microplus in the state present low efficacy, as measured by in vitro tests. The low drug efficacy is probably derived from the genetic resistance developed by the parasites due to the indiscriminated and inadequate use of these acaricides in the field.

Atividade neutralizante anti-herpesvírus bovino tipos 1 (BHV-1) e 5 (BHV-5) no soro de bovinos imunizados com vacinas contra o BHV-1

Calves were immunized with two vaccines against bovine herspevirus type 1 (BHV-1; vaccine A, n=28; B, n=28) or with a vaccine containing BHV-1 and BHV-5 antigens (vaccine C, n=32) and the serum neutralizing activity against BHV-1 and BHV-5 was measured after three vaccine administrations. Neutralizing activity to BHV-1 was detected in sera of 100% (n=88) of the animals (geometric mean titers [GMT] of 13.1; 14.8 and 34.3 for vaccines A, B and C, respectively) whereas 82 sera (93.2%) reacted to BHV-5 (GMTs: 10.6; 11.5 and 29.8). In all three groups, the mean BHV-1 titers did not differ from BHV-5 titers. However, comparison among the vaccines demonstrated that the mean titers to BHV-1 and BHV-5 were higher in animals receiving vaccine C (p<0.01). This vaccine also induced a higher proportion of reagents to BHV-5 (96.9%; against 85.7% for vaccine A and 92.9% for vaccine B). These results demonstrate that vaccine C induced higher neutralizing antibody titers against both viruses; and that antibodies induced by BHV-1 antigens (vaccines A and B) cross-reacted and displayed relevant neutralizing activity against BHV-5.

Dexamethasone-induced reactivation of bovine herpesvirus type 5 latent infection in experimentally infected rabbits results in a broader distribution of latent viral DNA in the brain

Bovine herpesvirus type 5 (BHV-5) is a major agent of meningoencephalitis in cattle and establishes latent infections mainly in sensory nerve ganglia. The distribution of latent BHV-5 DNA in the brain of rabbits prior to and after virus reactivation was studied using a nested PCR. Fifteen rabbits inoculated intranasally with BHV-5 were euthanized 60 days post-inoculation (group A, N = 8) or submitted to dexamethasone treatment (2.6 mg kg(-1) day(-1), im, for 5 days) and euthanized 60 days later (group B, N = 7) for tissue examination. Two groups of BHV-1-infected rabbits (C, N = 3 and D, N = 3) submitted to each treatment were used as controls. Viral DNA of group A rabbits was consistently detected in trigeminal ganglia (8/8), frequently in cerebellum (5/8), anterior cerebral cortex and pons-medulla (3/8) and occasionally in dorsolateral (2/8), ventrolateral and posterior cerebral cortices, midbrain and thalamus (1/8). Viral DNA of group B rabbits showed a broader distribution, being detected at higher frequency in ventrolateral (6/7) and posterior cerebral cortices (5/7), pons-medulla (6/7), thalamus (4/7), and midbrain (3/7). In contrast, rabbits inoculated with BHV-1 harbored viral DNA almost completely restricted to trigeminal ganglia and the distribution did not change post-reactivation. These results demonstrate that latency by BHV-5 is established in several areas of the rabbits brain and that virus reactivation leads to a broader distribution of latent viral DNA. Spread of virus from trigeminal ganglia and other areas of the brain likely contributes to this dissemination and may contribute to the recrudescence of neurological disease frequently observed upon BHV-5 reactivation.

MAGNITUDE, DURAÇÃO E ESPECIFICIDADE DA RESPOSTA SOROLÓGICA EM BOVINOS VACINADOS CONTRA O VÍRUS DA DIARRÉIA VIRAL BOVINA (BVDV)

The serological response induced by three inactivated commercial vaccines against bovine viral diarrhea virus (BVDV) was evaluated in calves immunized three times (days 0, 30 and 180) and examined at different intervals after vaccination. Thirty days after the second vaccination, 74.5% (70/94) of the calves had developed neutralizing antibodies titers against BVDV-1 and 52.1% (49/94) against BVDV-2. The geometric mean titers (GMT) and the number of animals with antibodies to BVDV-1 were 109.3(32/36); 54.6(22/28) and 25.5(16/30) for vaccines A, B e C, respectively and 19(27/36), 42.3(12/28) and 18.4(10/30) to BVDV-2. The antibody titers decreased by day 180, when 31.9%(30/94) of the calves showed no neutralizing activity against BVDV-1 and 63.8%(60/94) to BVDV-2. At that time, the GMTs and the number of animals seropositive to BVDV-1 were 28.3 (30/36), 28.3(20/28) and 16.1(14/30); and to BVDV-2 were 16.8(18/36), 21.6(10/28) and 28.3(6/30) for vaccines A, B and C, respectively. After the third dosis (day 180), the antibody titers to BVDV-1 increased significantly in all groups whereas a significant increase in BVDV-2 titers was observed only for vaccine A. At day 210, GMTs and the number of positive animals to BVDV-1 were 104.8(23/24), 50.3(24/26) and 43.7(24/28) and to BVDV-2 were 33.4(23/24), 23.3(22/26) and 15.7(22/28) for vaccines A, B and C. The neutralizing activity against BVDV-1 was higher than to BVDV-2 in all groups at day 210. Sera from five seropositive animals of each vaccine group were tested against four Brazilian BVDV-1 and two BVDV-2 isolates. In addition to the low and moderate titers, the cross-neutralization assays revealed a highly variable neutralizing activity against the Brazilian field isolates. These results demonstrated that the vaccination did not induce an adequate serological response in most animals, mainly due to the high antigenic variability of BVDV.

Neuropatogênese experimental da infecção pelo herpesvírus bovino tipo 5 em coelhos

Several aspects of the biology of bovine herpesvirus 5 (BoHV-5) have been studied in rabbits, which develop acute infection and neurological disease upon experimental inoculation. The acute infection is followed by the establishment of latent infection, which can be naturally or artificially reactivated. The first experiments in rabbits established a protocol for virus inoculation and monitoring the infection, and characterized the main virological, clinical and pathological aspects of the acute infection. The pathogenesis of acute infection, from the initial viral replication at site of inoculation, pathways and kinetics of viral transport to the brain, distribution and virus replication in the central nervous system (CNS), cellular and tissue tropism, clinical signs and CNS pathology have been extensively studied using this animal model. Subsequently, several biological and molecular aspects of latent BoHV-5 infection have also been elucidated upon inoculation of rabbits. Rabbits have also been used to investigate the phenotype (neuroinvasiveness, neurogrowth) of field isolates and recombinant vaccine candidates, protection by passive immunity, vaccine protection, the efficacy of anti-viral drugs and support therapies for neurological disease. This animal model was also used to investigate the origin and distribution of electric impulses involved in seizures - a hallmark of BoHV-5 induced neurological infection - and also to test the efficacy of anti-convulsivants. In spite of the possible differences between rabbits and cattle - the natural host of the virus - the observations taken from this experimental model have greatly contributed to the knowledge of the biology of BoHV-5 infection. The present article presents a review of the main published and unpublished results and observations by our group, comprising more than a decade of studies on the pathogenesis of BoHV-5 infection in the rabbit model.