RESUMO
Escherichia coli is an important cause of septicemia (SEPEC) and neonatal meningitis (NMEC) in dairy calves. However, the diversity of virulence profiles, phylogroups, antimicrobial resistance patterns, carriage of integron structures, and fluoroquinolone (FQ) resistance mechanisms have not been fully investigated. Also, there is a paucity of knowledge about the virulence profiles and frequency of potential SEPEC in feces from calves with or without diarrhea. This study aimed to characterize the virulence potential, phylogroups, antimicrobial susceptibility, integron content, and FQ-resistance mechanisms in Escherichia coli isolated from calves with meningitis and septicemia. Additionally, the virulence genes (VGs) and profiles of E. coli isolated from diarrheic and non-diarrheic calves were compared between them and together with NMEC and SEPEC in order to identify shared profiles. Tissue and fluid samples from eight dairy calves with septicemia, four of which had concurrent meningitis, were processed for bacteriology and histopathology. Typing of VGs was assessed in 166 isolates from diverse samples of each calf. Selected isolates were evaluated for antimicrobial susceptibility by the disk diffusion test. Phylogroups, integron gene cassettes cartography, and FQ-resistance determinants were analyzed by PCR, sequencing, and bioinformatic tools. Furthermore, 109 fecal samples and 700 fecal isolates from dairy calves with or without diarrhea were evaluated to detect 19 VGs by uniplex PCR. Highly diverse VG profiles were characterized among NMEC and SEPEC isolates, but iucD was the predominant virulence marker. Histologic lesions in all calves supported their pathogenicity. Selected isolates mainly belonged to phylogroups A and C and showed multidrug resistance. Classic (dfrA17 and arr3-dfrA27) and complex (dfrA17-aadA5::ISCR1::blaCTX-M-2) class 1 integrons were identified. Target-site mutations in GyrA (S83L and D87N) and ParC (S80I) encoding genes were associated with FQ resistance. The VGs detected more frequently in fecal samples included f17G (50%), papC (30%), iucD (20%), clpG (19%), eae (16%), and afaE-8 (13%). Fecal isolates displaying the profiles of f17 or potential SEPEC were found in 25% of calves with and without diarrhea. The frequency of E. coli VGs and profiles did not differ between both groups (p > 0.05) and were identical or similar to those found in NMEC and SEPEC. Overall, multidrug-resistant E. coli isolates with diverse VG profiles and belonging to phylogroups A and C can be implicated in natural cases of meningitis and septicemia. Their resistance phenotypes can be partially explained by class 1 integron gene cassettes and target-site mutations in gyrA and parC. These results highlight the value of antimicrobial resistance surveillance in pathogenic bacteria isolated from food-producing animals. Besides, calves frequently shed potential SEPEC in their feces as commensals ("Trojan horse"). Thus, these bacteria may be disseminated in the farm environment, causing septicemia and meningitis under predisposing factors.
Assuntos
Infecções por Escherichia coli , Meningite , Sepse , Animais , Antibacterianos/farmacologia , Bovinos , Farmacorresistência Bacteriana Múltipla/genética , Escherichia coli/genética , Infecções por Escherichia coli/veterinária , Integrons , Sepse/veterináriaRESUMO
In-vitro fertilization is a routine livestock-breeding technique widely used around the world. Several studies have reported the interaction of bovine viral-diarrhea virus (BVDV) with gametes and in-vitro-produced (IVP) bovine embryos. Since, gene expression in BVDV-infected IVP bovine embryos is scarcely addressed. The aim of this work was to evaluate the differential expression of genes involved in immune and inflammatory response. Groups of 20-25 embryos on Day 6 (morula stage) were exposed (infected) or not (control) to an NCP-BVDV strain in SOF medium. After 24 h, embryos that reached expanded blastocyst stage were washed. Total RNA of each embryo group was extracted to determine the transcription levels of 9 specific transcripts related with antiviral and inflammatory response by SYBR Green real time quantitative (RT-qPCR). Culture media and an aliquot of the last embryos wash on Day 7 were analyzed by titration and virus isolation, respectively. A conventional PCR confirmed BVDV presence in IVP embryos. A significantly higher expression of interferon-α was observed in blastocysts exposed to NCP-BVDV compared to the controls (p < 0.05). In this study, the upregulation of INFα and TLR7 genes involved in inflammatory and immune response in BVDV-infected IVP bovine embryos is a new finding in this field. This differential expression suggest that embryonic cells could function in a manner like immune cells by recognizing and responding early to interaction with viral pathogens. These results provide new insights into the action of BVDV on the complex molecular pathways controlling bovine early embryonic development.
Assuntos
Doença das Mucosas por Vírus da Diarreia Viral Bovina , Bovinos , Vírus da Diarreia Viral Bovina/imunologia , Desenvolvimento Embrionário/imunologia , Expressão Gênica/imunologia , Interferon-alfa , Animais , Doença das Mucosas por Vírus da Diarreia Viral Bovina/embriologia , Doença das Mucosas por Vírus da Diarreia Viral Bovina/imunologia , Doença das Mucosas por Vírus da Diarreia Viral Bovina/virologia , Bovinos/embriologia , Bovinos/imunologia , Vírus da Diarreia Viral Bovina/isolamento & purificação , Embrião de Mamíferos/imunologia , Embrião de Mamíferos/virologia , Feminino , Fertilização in vitro , Interferon-alfa/imunologia , Receptor 7 Toll-Like/imunologiaRESUMO
The aim of the present study was to characterize the specific immune response in prepubertal female calves inoculated with Neospora caninum. Forty-eight N. caninum-seronegative 6-month-old Angus female calves were randomly allocated into two groups: group A calves were inoculated subcutaneously (sc) with 1 × 106 tachyzoites of the low virulence NC-Argentina LP1 isolate in sterile phosphate-buffered saline (PBS); group B calves were mock inoculated sc with sterile PBS. Calves from group A developed a specific immune response characterized by the production of IgG antibodies and the expression of IFN-γ and TNF-α cytokines. Animals did not present any febrile reaction or reactions at the site of inoculation. Although chronic N. caninum infection was developed in 50% of calves of group A after inoculation, according to the presence of antibodies against rNc-SAG4, antigen characteristic of bradyzoites, N. caninum antibodies dropped below the cut-off of ELISA from day 210 post-inoculation onwards. Future trials using the same group of inoculated animals will allow the characterization of the evolution of the immune response during pregnancy and to determine whether the immunization with the local isolate is able to prevent congenital transmission and to protect against heterologous challenges.
Assuntos
Anticorpos Antiprotozoários/sangue , Antígenos de Protozoários/imunologia , Doenças dos Bovinos/imunologia , Coccidiose/veterinária , Neospora/imunologia , Animais , Bovinos , Doenças dos Bovinos/parasitologia , Coccidiose/imunologia , Coccidiose/parasitologia , Citocinas/metabolismo , Feminino , Imunização/veterinária , Neospora/patogenicidade , Distribuição AleatóriaRESUMO
Bovine herpesvirus 4 (BoHV-4) has been isolated from cattle with respiratory infections, vulvovaginitis, mastitis, abortions, endometritis and from apparently healthy animals throughout the world. Although it has not yet been established as causal agent of a specific disease entity, it is primarily associated with reproductive disorders of cattle. This virus can infect a wide range of species, either in vivo or in vitro. Two groups of prototype strains were originated from the first isolates: the DN599-type strains (American group) and the Movar-type strains (European group). In Argentina, BoHV-4 was isolated and characterized in 2007 from vaginal discharge samples taken from cows that had aborted. So far, more than 40 isolates, mainly associated with aborting bovine females have been registered in our country.
Assuntos
Aborto Animal/virologia , Doenças dos Bovinos/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 4/isolamento & purificação , Infecções Tumorais por Vírus/veterinária , Aborto Animal/epidemiologia , Animais , Anticorpos Antivirais/sangue , Apoptose , Argentina/epidemiologia , Bovinos , Doenças dos Bovinos/epidemiologia , Causalidade , Efeito Citopatogênico Viral , Endométrio/virologia , Feminino , Genoma Viral , Infecções por Herpesviridae/diagnóstico , Infecções por Herpesviridae/epidemiologia , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 4/classificação , Herpesvirus Bovino 4/patogenicidade , Herpesvirus Bovino 4/fisiologia , Especificidade de Hospedeiro , Interações Hospedeiro-Patógeno , Transtornos Puerperais/veterinária , Transtornos Puerperais/virologia , Infecções Tumorais por Vírus/diagnóstico , Infecções Tumorais por Vírus/epidemiologia , Infecções Tumorais por Vírus/virologia , Tropismo Viral , Virulência , Ativação ViralRESUMO
The aim of this study was to describe two events in which Neospora caninum was involved in bovine abortions in beef cattle. In the first event, 11 abortions in 57 heifers were recorded in 45 days. One aborted heifer was 5 times more likely to be seropositive than a non-aborted heifer (OR=4.9; IC 1.2-19.9) (p<0.05). In the second event, no association between serological results and abortions were observed (OR=0,69; 0,06-7,31) (p>0.05). Neither antibodies nor isolation of other pathogens were achieved in any case. On the contrary, antibodies and pathognomonic histopathological lesions were observed in the four fetuses from both cases. Interestingly, the findings in the first event suggest the epidemic behavior of the disease. In contrast, in the second event it appears that few abortions were due to N. caninum, suggesting the presence of endemic neosporosis.
Assuntos
Aborto Animal/epidemiologia , Doenças dos Bovinos/epidemiologia , Coccidiose/veterinária , Doenças Endêmicas , Neospora , Animais , Bovinos , Coccidiose/epidemiologiaRESUMO
Bovine viral diarrhea virus (BVDV) genome consists of a single-stranded, positive-sense RNA with high genetic diversity. In the last years, significant progress has been achieved in BVDV knowledge evolution through phylodynamic analysis based on the partial 5'UTR sequences, whereas a few studies have used other genes or the complete coding sequence (CDS). However, no research has evaluated and compared BVDV evolutionary history based on the complete genome (CG), CDS, and individual genes. In this study, phylodynamic analyses were carried out with BVDV-1 (Pestivirus A) and BVDV-2 (Pestivirus B) CG sequences available on the GenBank database and each genomic region: CDS, UTRs, and individual genes. In comparison to the CG, the estimations for both BVDV species varied according to the dataset used, pointing out the importance of considering the analyzed genomic region when concluding. This study may provide new insight into BVDV evolution history while highlighting the need to increase the available BVDV CG sequences to perform more comprehensive phylodynamic studies in the future.
Assuntos
Doença das Mucosas por Vírus da Diarreia Viral Bovina , Vírus da Diarreia Viral Bovina , Animais , Bovinos , Genótipo , Vírus da Diarreia Viral Bovina/genética , Genômica , Filogenia , Evolução Molecular , Diarreia , Genoma ViralRESUMO
Rotavirus A (RVA) causes diarrhea in calves and frequently possesses the G6 and P[5]/P[11] genotypes, whereas G8 is less common. We aimed to compare RVA infections and G/P genotypes in beef and dairy calves from major livestock regions of Argentina, elucidate the evolutionary origin of a G8 strain and analyze the G8 lineages, infer the phylogenetic relationship of RVA field strains, and investigate the evolution and spatio-temporal dynamics of the main G6 lineages in American countries. Fecal samples (n = 422) from diarrheic (beef, 104; dairy, 137) and non-diarrheic (beef, 78; dairy, 103) calves were analyzed by ELISA and semi-nested multiplex RT-PCR. Sequencing, phylogenetic, phylodynamic, and phylogeographic analyses were performed. RVA infections were more frequent in beef (22.0%) than in dairy (14.2%) calves. Prevalent genotypes and G6 lineages were G6(IV)P[5] in beef (90.9%) and G6(III)P[11] (41.2%) or mixed genotypes (23.5%) in dairy calves. The only G8 strain was phylogenetically related to bovine and artiodactyl bovine-like strains. Re-analyses inside the G8 genotype identified G8(I) to G8(VIII) lineages. Of all G6 strains characterized, the G6(IV)P[5](I) strains from "Cuenca del Salado" (Argentina) and Uruguay clustered together. According to farm location, a clustering pattern for G6(IV)P[5] strains of beef farms was observed. Both G6 lineage strains together revealed an evolutionary rate of 1.24 × 10-3 substitutions/site/year, and the time to the most recent common ancestor was dated in 1853. The most probable ancestral locations were Argentina in 1981 for G6(III) strains and the USA in 1940 for G6(IV) strains. The highest migration rates for both G6 lineages together were from Argentina to Brazil and Uruguay. Altogether, the epidemiology, genetic diversity, and phylogeny of RVA in calves can differ according to the production system and farm location. We provide novel knowledge about the evolutionary origin of a bovine G8P[11] strain. Finally, bovine G6 strains from American countries would have originated in the USA nearly a century before its first description.
Assuntos
Doenças dos Bovinos , Infecções por Rotavirus , Rotavirus , Animais , Bovinos , Rotavirus/genética , Epidemiologia Molecular , Filogenia , Infecções por Rotavirus/epidemiologia , Infecções por Rotavirus/veterinária , Diarreia/epidemiologia , Diarreia/veterinária , Genótipo , Fezes , Doenças dos Bovinos/epidemiologiaRESUMO
Bovine alphaherpesvirus type 1 (BoAHV-1) is associated with respiratory and reproductive syndromes. Until present the immunologic mechanisms involved in BoAHV-1 abortion are partially known. We studied key elements of the innate immune response in the placentas and fetal lungs from cattle experimentally-inoculated with BoAHV-1. These tissues were analyzed by histopathology. Furthermore, virus identification was performed by qPCR and the expression of the inflammatory cytokines such as tumor necrosis factor-alpha, interleukin 1-alpha and inflammatory mediators like inducible nitric oxide synthase and cyclooxeganse-2 was evaluated by immunohistochemistry. The viral transplacental infection was confirmed by the detection of BoAHV-1 by qPCR in the placenta and fetal organs, which revealed mild inflammatory lesions. Inducible nitric oxide synthase immunolabelling was high in the lungs of infected fetuses and placentas, as well as for tumor necrosis factor-alpha in the pulmonary parenchyma and cyclooxeganse-2 in fetal annexes. However, the expression of interleukin 1-alpha was weak in these organs. To our knowledge, this is the first study that provides strong evidence of an early immune response to BoAHV-1 infection in the conceptus. Advances in the knowledge of the complex immunological interactions at the feto-maternal unit during BoAHV-1 infection are needed to clarify the pathogenesis of abortion.
Assuntos
Citocinas , Fator de Necrose Tumoral alfa , Gravidez , Feminino , Bovinos , Animais , Citocinas/genética , Citocinas/metabolismo , Ciclo-Oxigenase 2/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Placenta , Pulmão/patologia , Interleucina-1/metabolismoRESUMO
Neosporosis, a disease caused by the obligate intracellular protozoan Neospora caninum, produces abortions in cattle. The severe economic losses in cattle industry justify the need to develop control measures for preventing bovine abortion. Apicomplexan parasitic resistance is associated with T helper 1 immune response mediated by CD4 cytotoxic T lymphocytes, the production of interferon-gamma, interleukin-12, tumor necrosis factor and immunoglobulin G2. The reduction of vertical transmission in subsequent pregnancies and the low levels of abortion repetition suggests the existence of protective immune mechanisms. Inoculation with live tachyzoites before mating protects against infection and abortion. Antecedents of the development of live vaccines against other protozoa stimulate research to develop a live vaccine against N. caninum. On the other hand, an inactivated vaccine with low efficacy against neosporosis is useful in the prevention of abortion in farms with epizootic disease. A neosporosis vaccine should avoid abortion, transplacental transmission and infection persistence. In the present work, advances in vaccine development including lysate of tachyzoites, live parasites, recombinant antigens and vaccine vectors are reviewed.
Assuntos
Doenças dos Bovinos/prevenção & controle , Coccidiose/veterinária , Neospora/imunologia , Vacinas Protozoárias , Aborto Animal/etiologia , Aborto Animal/parasitologia , Aborto Animal/prevenção & controle , Adjuvantes Imunológicos , Animais , Antígenos de Protozoários/imunologia , Antígenos de Superfície/imunologia , Apicomplexa/imunologia , Bovinos , Coccidiose/imunologia , Coccidiose/prevenção & controle , Coccidiose/transmissão , Feminino , Vetores Genéticos/imunologia , Camundongos , Neospora/crescimento & desenvolvimento , Gravidez , Vacinas Protozoárias/imunologia , Vacinas Atenuadas/imunologia , Vacinas de DNA/imunologia , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
Bovine viral diarrhea virus (BVDV) is a worldwide distributed pathogen of livestock classified into three species, BVDV-1 (Pestivirus A), BVDV-2 (Pestivirus B), and HoBi-like pestivirus (HoBiPeV; Pestivirus H). Despite being considered endemic in several regions of the Americas, the spatiotemporal distribution of BVDV is scarcely known. This study aimed to reconstruct the population dynamics of BVDV in American countries. The analyses performed with the partial 5´UTR gene showed that BVDV-1 and -2 would have started their diversification in the 1670s and 1790s in the United States, whereas HoBiPeV probably emerged in the 1980s in Brazil. No evident geographic clustering was observed in the Bayesian trees, which may indicate that multiple introductions events would have occurred following the first introduction. This study provides new insights into BVDV dynamics, although further analyses including sequences from other American countries and continents will help to expand the knowledge of BVDV evolution and transmission.
RESUMO
Bovine viral diarrhea virus (BVDV) is an important pathogen of ruminants worldwide and is characterized by high genetic diversity and a wide range of clinical presentations. In Argentina, several studies have evaluated the genetic diversity of BVDV but no phylodynamic study has been published yet. In this study, a comprehensive compilation and update of Argentinean BVDV sequences were performed, and the evolutionary history of BVDV was characterized by phylodynamic analyses based on the 5´UTR. Although BVDV-1b and BVDV-1a were the most frequent subtypes, novel subtypes for Argentina, 1e and 1i, were identified. The phylodynamic analysis suggested that BVDV started its diversification in the mid-1650s with an exponential increase in viral diversity since the late 1990s, possibly related to the livestock expansion and intensification in the country. Evolutionary rate in the 5´UTR was faster for BVDV-1a than for BVDV-1b, and both subtypes presented an endemic nature according to the demographic reconstructions. The current study contributes to clarify the evolutionary history of BVDV in the main cattle region of the country and provides useful information about the epidemiology and future development of diagnostic and control tools in Argentina.
Assuntos
Doença das Mucosas por Vírus da Diarreia Viral Bovina/epidemiologia , Vírus da Diarreia Viral Bovina/genética , Variação Genética , Genoma Viral , Animais , Argentina/epidemiologia , Doença das Mucosas por Vírus da Diarreia Viral Bovina/virologia , Bovinos , Vírus da Diarreia Viral Bovina Tipo 1/genética , Vírus da Diarreia Viral Bovina Tipo 2/genética , FilogeniaRESUMO
Bovine viral diarrhea (BVD) is a major worldwide disease with negative economic impact on cattle production. Successful control programs of BVD require the identification and culling of persistently infected (PI) animals with bovine viral diarrhea virus (BVDV). A variety of diagnostic tests are available to detect BVDV, but no comparison has been performed among those tests in Argentina. Sera collected from 2864 cattle, belonging to 55 herds from three Argentinean provinces, were analyzed by nested RT-PCR (RT-nPCR) to detect BVDV for diagnostic purposes. Additionally, this study evaluated the agreement of the RT-nPCR along with virus isolation, antigen-capture ELISA, and real-time RT-PCR for BVDV detection in archived bovine serum samples (n = 90). The RT-nPCR was useful for BVDV detection in pooled and individual serum samples. BVDV was detected in 1% (29/2864) of the cattle and in 20% (11/55) of the herds. The proportion of BVDV-positive sera was not statistically different among the tests. In addition, comparisons showed high agreement levels, with the highest values between both RT-PCR protocols. The frequency of BVDV infection at individual and herd level was lower than the reported values worldwide. Since follow-up testing was not performed, the frequency of PI cattle was unknown. Also, this study demonstrated that the four diagnostic tests can be used reliably for BVDV identification in individual serum samples. Further epidemiologically designed studies that address prevalence, risk factors, and economic impact of BVDV in Argentina will be necessary to implement effective control programs.
Assuntos
Doença das Mucosas por Vírus da Diarreia Viral Bovina/sangue , Doença das Mucosas por Vírus da Diarreia Viral Bovina/diagnóstico , Vírus da Diarreia Viral Bovina/imunologia , Técnicas de Diagnóstico Molecular/normas , Técnicas de Diagnóstico Molecular/veterinária , Testes Sorológicos/normas , Testes Sorológicos/veterinária , Animais , Argentina , Bovinos , Vírus da Diarreia Viral Bovina/genética , Vírus da Diarreia Viral Bovina/isolamento & purificação , Limite de Detecção , Técnicas de Diagnóstico Molecular/métodos , Testes Sorológicos/métodos , SoroRESUMO
Detection of bovine viral diarrhea virus (BVDV) in aborted fetus samples is often difficult due to tissue autolysis and inappropriate sampling. Studies assessing different methods for BVDV identification in fetal specimens are scarce. The present study evaluated the agreement between different diagnostic techniques to detect BVDV infections in specimens from a large number of bovine aborted fetuses and neonatal deaths over a period of 22 years. Additionally, genetic, serological, and pathological analyses were conducted in order to characterize BVDV strains of fetal origin. Samples from 95 selected cases from 1997 to 2018 were analyzed by antigen-capture ELISA (AgELISA), nested RT-PCR (RT-nPCR), and real-time RT-PCR (RT-qPCR). In addition, amplification and sequencing of the 5'UTR region were performed for phylogenetic purposes. Virus neutralization tests against the BVDV-1a, BVDV-1b, and BVDV-2b subtypes were conducted on 60 fetal fluids of the selected cases. Furthermore, the frequency and severity of histopathological lesions were evaluated in BVDV-positive cases. This study demonstrated that RT-nPCR and RT-qPCR were more suitable than AgELISA for BVDV detection in fetal specimens. However, the agreement between the two RT-PCR methods was moderate. The BVDV-1b subtype was more frequently detected than the BVDV-1a and BVDV-2b subtypes. Neutralizing antibodies to any of the three subtypes evaluated were present in 94% of the fetal fluids. Microscopically, half of the BVDV-positive cases showed a mild non-suppurative inflammatory response. These results emphasize the need to consider different methods for a diagnostic approach of BVDV associated to reproductive losses.
Assuntos
Feto Abortado/virologia , Doença das Mucosas por Vírus da Diarreia Viral Bovina/diagnóstico , Vírus da Diarreia Viral Bovina/classificação , Filogenia , Regiões 5' não Traduzidas , Animais , Anticorpos Neutralizantes/imunologia , Bovinos , Vírus da Diarreia Viral Bovina/isolamento & purificação , Ensaio de Imunoadsorção Enzimática , Reação em Cadeia da Polimerase , Reação em Cadeia da Polimerase em Tempo Real , Fatores de TempoRESUMO
The aim of this study was to evaluate delayed type hypersensitivity (DTH) induced by the intradermal inoculation of a Neospora caninum tachyzoite soluble lysate in cattle previously exposed with the protozoa. Four experimental groups were selected according to the prior exposure to N. caninum antigen. All cows were intradermally injected with a N. caninum tachyzoite soluble lysate and skinfold thickness growth at the inoculation sites was measured at 0, 24, 48, 72 and 96 h post inoculation (hpi). Additionally, specific antibodies and IFN-γ production were assessed. Cows experimentally infected with live N. caninum tachyzoites and cows naturally exposed to N. caninum developed skin reactions compatible with DTH between 24 and 96 hpi (p < 0.05). Moreover, cows inoculated with an experimental N. caninum vaccine and cows without evidence of exposure to N. caninum did not show a significant increase in skin thickness (p > 0.05). Furthermore, serological status of the animals was not modified due to the intradermal inoculation. The highest IFN-γ production was observed at 15 days after intradermal inoculation (p < 0.05). Therefore, these results suggest that cattle previously exposed to N. caninum develop a reaction compatible with DTH which could be useful as in vivo cell mediated immunity parameter for assessed bovine neosporosis.
Assuntos
Antígenos de Protozoários/imunologia , Hipersensibilidade Tardia/veterinária , Neospora/imunologia , Animais , Antígenos de Protozoários/administração & dosagem , Bovinos/imunologia , Bovinos/parasitologia , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/parasitologia , Coccidiose/imunologia , Coccidiose/veterinária , Relação Dose-Resposta Imunológica , Hipersensibilidade Tardia/imunologia , Hipersensibilidade Tardia/parasitologia , Imunoglobulina G/imunologia , Injeções Intradérmicas/veterináriaRESUMO
Bovine herpesvirus (BoHV) types 1 and 5 are neuroinvasive. Cases of BoHV-1-induced encephalitis are not as frequent as those caused by BoHV-5. In this study, the capability of BoHV-5 to induce apoptosis in cell cultures and in the trigeminal ganglion during acute infection of experimentally-infected cattle was analyzed. Apoptotic changes in cell cultures agree with the ability of the viral strains to replicate in each cell line. Marked differences were observed between the in vitro induction of apoptosis by BoHV-1Cooper and BoHV-5 97/613 strains. Apoptotic neurons were clearly evident in the trigeminal ganglion of BoHV-1-infected calves. For BoHV-5 a fewer number of positive neurons was observed. There is an association between the magnitude of bovine herpesviruses replication and the induction of apoptosis in trigeminal ganglion. These findings suggest that the induction of apoptosis and the innate immune response orchestrate the final outcome of alpha herpesviruses infection of the bovine nervous system.
Assuntos
Doenças dos Bovinos/virologia , Infecções por Herpesviridae/veterinária , Herpesvirus Bovino 1/patogenicidade , Herpesvirus Bovino 5/patogenicidade , Neurônios/virologia , Gânglio Trigeminal/virologia , Animais , Apoptose , Bovinos , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/patologia , Linhagem Celular , Células Epiteliais/imunologia , Células Epiteliais/patologia , Células Epiteliais/virologia , Células HeLa , Infecções por Herpesviridae/imunologia , Infecções por Herpesviridae/patologia , Infecções por Herpesviridae/virologia , Herpesvirus Bovino 1/imunologia , Herpesvirus Bovino 5/imunologia , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata , Neurônios/imunologia , Neurônios/patologia , Especificidade da Espécie , Gânglio Trigeminal/imunologia , Gânglio Trigeminal/patologia , Replicação ViralRESUMO
Dual infections with both bovine viral diarrhea virus (BVDV)-1 and -2 seem to be unusual. The aim of this study was to describe an infection with both BVDV genotypes in a stillborn calf. Virus isolation and phylogenetic analyses of the 5´UTR and NS5B regions confirmed the presence of BVDV-1b and -2b in spleen and lung, whereas BVDV-2b was also detected in brain, heart, liver, kidney and, fluid of cavities. These results confirm that dual infections with both BVDV-1 and BVDV-2 species can occur naturally and their tissue distribution can be different.
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Structural changes in the zona pellucida (ZP) of bovine oocytes seem to modulate their interaction with various viral agents, facilitating the viral infection in in vitro production systems. To evaluate the susceptibility of bovine oocytes to noncytopathogenic bovine viral diarrhea virus (ncp-BVDV), cumulus-oocyte complexes were exposed to 10(7) âtissue culture-infective doses (TCID50)/mL of an ncp-BVDV strain during IVM (in vitro maturation). After that, cumulus cells and the ZP were removed by hyaluronidase and pronase treatment, respectively, and the percentages of oocytes with polar body were analyzed as a sign of nuclear maturation. After passage through cell culture, the virus was isolated from granulosa cells, ZP-free mature oocytes, and ZP-intact mature oocytes. These results were confirmed by reverse transcription-polymerase chain reaction. After consecutive washes, the virus remained associated with ZP-free oocytes, maintaining its replication and infectivity in permissive cells. Based on these findings, it is concluded that the classical viral isolation procedure has a predictive value to detect BVDV associated with ZP-free oocytes and that it was novelty demonstrated that both washing and trypsin treatment of oocytes were ineffective to remove BVDV infection.
Assuntos
Bovinos , Células do Cúmulo/virologia , Vírus da Diarreia Viral Bovina/isolamento & purificação , Técnicas de Maturação in Vitro de Oócitos/veterinária , Oócitos/virologia , Animais , Membrana Celular , Corpos Polares , Zona PelúcidaRESUMO
Bovine herpesvirus 4 (BoHV-4) is increasingly considered as responsible for various problems of the reproductive tract. The virus infects mainly blood mononuclear cells and displays specific tropism for vascular endothelia, reproductive and fetal tissues. Epidemiological studies suggest its impact on reproductive performance, and its presence in various sites in the reproductive tract highlights its potential transmission in transfer-stage embryos. This work describes the biological and genetic characterization of BoHV-4 strains isolated from an in vitro bovine embryo production system. BoHV-4 strains were isolated in 2011 and 2013 from granulosa cells and bovine oocytes from ovary batches collected at a local abattoir, used as "starting material" for in vitro production of bovine embryos. Compatible BoHV-4-CPE was observed in the co-culture of granulosa cells and oocytes with MDBK cells. The identity of the isolates was confirmed by PCR assays targeting three ORFs of the viral genome. The phylogenetic analyses of the strains suggest that they were evolutionary unlinked. Therefore it is possible that BoHV-4 ovary infections occurred regularly along the evolution of the virus, at least in Argentina, which can have implications in the systems of in vitro embryo production. Thus, although BoHV-4 does not appear to be a frequent risk factor for in vitro embryo production, data are still limited. This study reveals the potential of BoHV-4 transmission via embryo transfer. Moreover, the high variability among the BoHV-4 strains isolated from aborted cows in Argentina highlights the importance of further research on the role of this virus as an agent with the potential to cause reproductive disease in cattle. The genetic characterization of the isolated strains provides data to better understand the pathogenesis of BoHV-4 infections. Furthermore, it will lead to fundamental insights into the molecular aspects of the virus and the means by which these strains circulate in the herds.
Assuntos
Embrião de Mamíferos/virologia , Células da Granulosa/virologia , Herpesvirus Bovino 4/genética , Oócitos/virologia , Animais , Argentina , Teorema de Bayes , Bovinos , Células Cultivadas , Técnicas de Cocultura , DNA Viral/análise , Cães , Feminino , Células da Granulosa/citologia , Herpesvirus Bovino 4/classificação , Herpesvirus Bovino 4/isolamento & purificação , Células Madin Darby de Rim Canino , Oócitos/citologia , Fases de Leitura Aberta/genética , Filogenia , Análise de Sequência de DNARESUMO
The objective of the present work is the description outbreaks caused by bovine viral diarrhea virus (BVDV) in commercial beef cattle ranches in Argentina. Genetic affiliation and their association with the clinical manifestation were carried out with five BVDV isolates from an outbreak of mucosal disease (MD) (Outbreak #1), acute enteritis (Outbreaks #2 and #3) and generalized dermatitis (Outbreaks #4 and #5). Upon genetic analysis CP BVDV isolate of Outbreak #1 clustered to closely to BVDV Oregon (Genotype 1). BVDV isolates from the outbreaks of generalized dermatitis (Outbreaks #4 and #5) were located close to BVDV Osloss within Genotype 1. The identification by immunohistochemistry of BVDV in exudative dermatitis indicates the epithelial cell tropism of the virus. Phylogenic characterization of BVDV from Outbreaks #2 and #3 locate them as BVDV-2. 5'UTR sequence of these viruses revealed a homology of 88 and 90% to BVDV-890 (Genotype 2) and a 77 and 75% to BVDV-SD1 (Genotype 1), respectively. The association of BVDV-2 with severe disease indicates the presence of highly virulent strains. Data from natural outbreaks where BVDV-1 and BVDV-2 were isolated revealed that pathology overlaps and not clearly allows the differentiation between genotypes based on gross or microscopic lesions. Thus, for a definitive diagnosis, further virology and molecular studies are necessary. Additionally, the results of this work focused on the origin and consequences of genetic variations of BVDV with regard to pathogenesis and suggest the association between genotype and a defined clinical syndrome.
Assuntos
Doença das Mucosas por Vírus da Diarreia Viral Bovina/virologia , Vírus da Diarreia Viral Bovina/genética , Surtos de Doenças/veterinária , Animais , Argentina/epidemiologia , Sequência de Bases , Doença das Mucosas por Vírus da Diarreia Viral Bovina/epidemiologia , Doença das Mucosas por Vírus da Diarreia Viral Bovina/patologia , Bovinos , DNA Viral/análise , DNA Viral/química , Vírus da Diarreia Viral Bovina/classificação , Feminino , Genótipo , Imuno-Histoquímica/veterinária , Masculino , Dados de Sequência Molecular , Testes de Neutralização/veterinária , Filogenia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Homologia de Sequência do Ácido NucleicoRESUMO
Bovine abortion is an important cause of significant economic losses in beef and dairy herds. This syndrome is usually difficult to diagnose. The aim of this study was to characterize bovine abortion causes in Argentina by standard diagnosis procedures (histology, bacterial and viral isolation) and other diagnostic tests like direct fluorescent antibody test (DFAT), fetal serology, immunohistochemistry (IHC), and PCR, showing their specific advantages and limitations. Necropsies were performed in 150 aborted bovine fetuses submitted to the diagnostic laboratories of Instituto Nacional de Tecnología Agropecuaria (INTA) Balcarce, Argentina. Etiological diagnosis was confirmed in 78 fetuses (52% of the cases). Most causes of abortion were of infectious origin, being Neospora caninum (14.67%), Campylobacter fetus sp. (9.33%), Leptospira spp. (7.33%) and Brucella abortus (6.65%) the main microorganisms identified. Bovine viral diarrhea virus (BVDV) and bovine herpes virus (BHV) were diagnosed in 2 (1.33%) and 3 (2%) cases, respectively. This study showed a better characterization of bovine abortion compared with previous researches done on this topic.(AU)
O aborto bovino é uma causa importante de perdas econômicas significativas em rebanhos bovinos e leiteiros. Esta síndrome é geralmente difícil de diagnosticar. O objetivo deste estudo foi caracterizar o aborto bovino na Argentina por procedimentos diagnósticos de rotina (histologia, isolamento viral e bacteriana) e outros testes diagnósticos como ensaio directo de anticorpos fluorescentes (DFAT), sorologia fetal, imuno-histoquica (IHC), e PCR; mostrando suas vantagens e limitações específicas. As necropsias foram realizadas em 150 fetos bovinos abortados submetidos aos laboratórios de diagnóstico do Instituto Nacional de Tecnologia Agropecuária (INTA) de Balcarce, na Argentina. O diagnóstico etiológico foi confirmado em 78 fetos (52% dos casos). A maioria das causas de aborto foram de origem infecciosa, sendo Neospora caninum (14,67%), Campylobacter fetus sp. (9,33%), Leptospira spp. (7,33%) e Brucella abortus (6,65%) os principais microrganismos identificados. O vírus da diarréia viral bovina (BVDV) e o herpesvírus bovino (BHV) foram diagnosticados em 2 (1,33%) e 3 (2%) casos, respectivamente. Este estudo mostrou uma melhor caracterização do aborto bovino em comparação com pesquisas anteriores feita sobre este tema.(AU)