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1.
Vet Parasitol ; 276: 108962, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31704559

RESUMO

Sarcocystis neurona is the major cause of the equine protozoal myeloencephalitis (EPM) in the Americas and has opossums of the genus Didelphis as definitive hosts. Most isolates of Sarcocystis sp. shed by opossums in Brazil differ genetically from the known species of Sarcocystis. These Brazilian isolates behave similarly as Sarcocystis falcatula, which causes sarcocystosis in birds, and for this reason, have been classified as Sarcocystis falcatula-like. Genes coding for the immunodominant surface antigens SAG2, SAG3 and SAG4 of S. falcatula-like are similar to those from S. neurona. It is unknown the Sarcocystis species that causes EPM in Brazil, as S. neurona has never been genetically confirmed in Brazilian horses. All cases associated with EPM in Brazil were diagnosed by immunological tests, which are not specific for S. neurona infection. It is possible that S. falcatula-like may infect horses in Brazil. The aims of the current study were to test the susceptibility of gerbils (Meriones unguiculatus) to experimental infections with S. neurona and S. falcatula-like, and to investigate potential serologic cross-reactivity to these parasites by immunofluorescent antibody test (IFAT) and Western blot (WB). A total of 27 gerbils, distributed in five experimental groups (G1-G5), were employed in this work (G1: 4 negative controls; G2: 6 infected with S. neurona merozoites, G3: 6 infected with S. falcatula-like merozoites; G4 and G5 (5 and 6, respectively, infected with different doses of sporocysts). None of the 17 animals that seroconverted for the parasites in IFAT presented any visualized organism or Sarcocystis DNA in the examined tissues. No serologic cross-reactivity was observed using IFAT. However, sera from animals infected with S. falcatula-like and S. neurona presented the same pattern of antigenic recognition when S. neurona merozoites were used as antigen in WB, including reactivity to proteins of 30 and 16 kDa, regarded as specific markers for S. neurona-infected animals. Gerbils did not sustain infection by these parasites, although produced antibodies after inoculation. These results are suggestive that other animal species that are exposed to S. falcatula-like, including horses, may present serologic cross-reactivity to S. neurona in WB. IFAT was demonstrated to be more specific that WB for the detection of antibodies to S. falcatula-like and S. neurona in the experimental conditions of this study.


Assuntos
Antígenos de Protozoários/imunologia , Sarcocystis/imunologia , Sarcocistose/imunologia , Animais , Antígenos de Superfície/imunologia , Western Blotting/veterinária , Linhagem Celular , Galinhas , Reações Cruzadas , Didelphis/parasitologia , Encefalomielite/imunologia , Encefalomielite/parasitologia , Encefalomielite/veterinária , Feminino , Imunofluorescência/veterinária , Gerbillinae , Epitopos Imunodominantes/imunologia , Reação em Cadeia da Polimerase , Sarcocistose/parasitologia , Sarcocistose/patologia , Células Vero
2.
Vet Parasitol ; 274: 108921, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31536867

RESUMO

Dogs are the main domestic reservoir of Leishmania infantum, and in cases of uncontrolled infection, a strong humoral immune response is elicited, which is inefficient against the parasites. Previous studies have suggested that an adequate antigen/antibody ratio, with a moderate prevalence of antigens with respect to the antibodies, could result in the formation of circulating immune complexes (CIC) in canine leishmaniosis (CanL). Deposition of these complexes in tissues has been associated with vasculitis, uveitis, arthritis, dermatitis and especially glomerulonephritis and renal failure. However, little is known about the relationship between the presence of CIC and disease progression. The aim of this study was to evaluate serum CIC level and its correlation with disease severity in infected dogs with different stages of disease and non-infected animals as a control. A total of 60 dogs were included in the study, classified according to the proposed LeishVet classification criteria: healthy non-infected (n = 13); healthy infected (n = 12); sick stage I (n = 9); sick stage II (n = 17); sick stage III (n = 8); and sick stage IV (n = 1). CIC were isolated from serum samples using a modified polyethylene glycol precipitation method, and their levels measured by ELISA and bicinchoninic acid protein assay. A nanoparticle tracking analysis was performed to investigate the relationship between the molecular size distribution of the CIC and disease progression. In conclusion, the results confirmed a positive association between CIC levels, their molecular size and disease progression that suggests a potential use of CIC as biomarkers of CanL.


Assuntos
Complexo Antígeno-Anticorpo/sangue , Doenças do Cão/sangue , Imunofluorescência/veterinária , Leishmaniose/veterinária , Animais , Anticorpos Antiprotozoários/sangue , Biomarcadores/sangue , Doenças do Cão/diagnóstico , Doenças do Cão/imunologia , Doenças do Cão/parasitologia , Cães , Leishmaniose/imunologia , Leishmaniose/patologia
3.
Vet J ; 249: 33-40, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31239162

RESUMO

Feline iris melanoma, the most common feline intraocular tumour, has a reported metastatic rate of 19-63%. However, there is a lack of knowledge about its molecular biology. Previous studies have reported that feline iris melanomas do not harbour mutations comparable to common mutations found in their human counterpart. Nevertheless, there are differences in the gene expression patterns. The aim of this study was to investigate the protein expression of B-RAF oncogene serine/threonine kinase (BRAF), G protein subunit alpha q (GNAQ) and 11 (GNA11), KIT proto-oncogene receptor tyrosine kinase (KIT), and Ras association family member 1 (RASSF1) in feline iris melanomas. Fifty-seven formalin-fixed paraffin embedded (FFPE) iris melanomas and 25 FFPE eyes without ocular abnormalities were stained with antibodies against the respective proteins using immunofluorescence. Averaged pixel intensities/µm2 and percentage of stained area from total tissue area were measured and the results were compared. Compared to the control group, iris melanomas showed overexpression of BRAF, GNAQ, GNA11 and KIT. The higher expression of BRAF, GNAQ, GNA11 and KIT in feline iris melanomas suggest that these proteins may play a key role in the development of feline iris melanomas and KIT may present a possible target for future therapies in cats with feline iris melanomas.


Assuntos
Doenças do Gato/metabolismo , Neoplasias da Íris/veterinária , Melanoma/veterinária , Animais , Gatos , Feminino , Imunofluorescência/veterinária , Subunidades alfa de Proteínas de Ligação ao GTP/biossíntese , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/biossíntese , Neoplasias da Íris/metabolismo , Melanoma/metabolismo , Biossíntese de Proteínas , Proteínas Proto-Oncogênicas B-raf/biossíntese , Proteínas Proto-Oncogênicas c-kit/biossíntese , Proteínas Supressoras de Tumor/biossíntese
4.
Res Vet Sci ; 125: 82-88, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31174167

RESUMO

Strangles is a highly prevalent, extremely contagious, and occasionally lethal infectious disease affecting horses worldwide. Prophylactic antibiotics are ineffective in prevention of disease but are recommended for exposed horses at the first sign of fever and any horse obviously ill from strangles or with complications and there is an urgent need of a cost-effective, safe, efficacious vaccine. In the present study, we sought to develop effective vaccines by fusing the Streptococcus equi subspecies equi (S. equi) antigen SeM with the flagellin of Salmonella abortus equi FljB. We also explored the immunogenicity and efficacy of this candidate vaccine in mice and horses by intramuscular injection. Mice and horses immunized with FljB-SeM DNA vaccine showed high levels of specific antibody and increased production of IFN-γ and IL-4. This confirmed that both Th1 and Th2 type responses were induced. The mice survival rate was significantly higher after immunization with FljB-SeM than with SeM alone. The FljB-SeM DNA could strengthen both the Th1 and Th2 immune responses compared to SeM and could provide better protection against S. equi. This technique could help develop a candidate vaccine for S. equi infection.


Assuntos
Anticorpos Antibacterianos/imunologia , Vacinas Bacterianas/imunologia , Doenças dos Cavalos/prevenção & controle , Infecções Estreptocócicas/veterinária , Streptococcus equi/imunologia , Vacinas de DNA/imunologia , Animais , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Feminino , Flagelina/imunologia , Imunofluorescência/veterinária , Doenças dos Cavalos/imunologia , Cavalos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Injeções Intramusculares/veterinária , Interferon gama/sangue , Interleucina-4/sangue , Cinética , Camundongos , Camundongos Endogâmicos C57BL , Plasmídeos , Organismos Livres de Patógenos Específicos , Infecções Estreptocócicas/mortalidade , Infecções Estreptocócicas/prevenção & controle , Streptococcus equi/genética , Taxa de Sobrevida , Vacinação/veterinária
5.
BMC Vet Res ; 15(1): 112, 2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30975151

RESUMO

BACKGROUND: Duck Tembusu virus (DTMUV) is a novel member of Flavivirus. The isolated and purified DTMUV strain XZ-2012 was used as a strain model, to intramuscularly inject the six-month egg-laying shelducks with the infective dose of 104TCID50. The dynamic distribution of the virus in spleen at different time post-infection (pi) was studied using RT-PCR, RT-qPCR, ELISA, immunofluorescence and transmission electron microscopy (TEM). RESULT: The results showed that the virus occurred in the spleen after 2 hpi and lasted up to 18 dpi. The registered viral load increased from 2 hpi to 3 dpi, and then it diminished from 6 dpi to 18 dpi with a slight rise at 12 dpi. From 2 hpi to 6 dpi the DTMUV particles were mostly distributed in the periellipsoidal lymphatic sheath (PELS) of spleen white pulp, few being found in the sheathed capillary. From 9 dpi to 18 dpi, the DTMUV particles were migrating into periarterial lymphatic sheaths (PALS) around the central artery through the red pulp. Under TEM, the virus particles could be observed mostly in lymphocytes and macrophages. CONCLUSION: It was suggested that DTMUV invaded lymphocytes and macrophages of the spleen at 2 hpi and replicated significantly from 1 dpi to 3 dpi, being eliminated from 9 dpi to 18 dpi. This is the first study on the dynamic distribution of DTMUV from invasion to elimination in duck spleen conducted by molecular and morphological methods. It could provide theoretical basis for the occurrence, development and detoxification of the virus in the organs of the immune system.


Assuntos
Patos/virologia , Infecções por Flavivirus/veterinária , Flavivirus/fisiologia , Doenças das Aves Domésticas/virologia , Baço/virologia , Animais , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Infecções por Flavivirus/virologia , Imunofluorescência/veterinária , Microscopia Eletrônica de Transmissão/veterinária , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Carga Viral/veterinária
6.
Parasit Vectors ; 12(1): 133, 2019 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-30909952

RESUMO

BACKGROUND: Feline leishmaniosis caused by Leishmania infantum is considered a rare disease in endemic areas, whereas subclinical infections are common. Immune response plays a key role in driving the course of L. infantum infection in other host species; however, the feline cell-mediated immune response to L. infantum infection has not yet been investigated. The aim of this study was to determine the cell-mediated immune response specific to L. infantum by means of interferon (IFN)-γ release in whole blood assay from cats living in endemic areas (66 in Sicily and 113 in Catalonia) and to compare with antibody levels to L. infantum [enzyme-linked immunosorbent assay (ELISA) and immunofluorescence antibody test (IFAT)], blood parasite load and retroviral infections. RESULTS: Most cats (n = 140) were L. infantum antibody negative and only 22% (n = 39) were positive. Only 9 and 2% of tested cats had a feline immunodeficency virus (FIV) infection or a feline leukemia virus (FeLV) infection, respectively. Thirty-two cats out of 179 (18%) produced IFN-γ after stimulation with L. infantum soluble antigen (LSA) while the majority of cats (93%) produced IFN-γ after stimulation with concanavalin A (ConA). Six LSA-IFN-γ-producer cats were seropositive (three to ELISA and five to IFAT) but they were polymerase chain reaction (PCR) negative, while only one cat was antibody- and PCR-positive. Significant positive correlations were found between IFN-γ concentrations after stimulation with LSA and ConA, and between serology and PCR testing. No association was found between FIV status and LSA or ConA-IFN-γ production. Combining PCR, serology and specific IFN-γ concentration results, we found that 36% of cats studied were exposed to L. infantum. CONCLUSIONS: As expected, cats from endemic areas produce IFN-γ after ex vivo blood stimulation with LSA and therefore are able to activate a cell-mediated adaptive immune response against the parasite that is variably associated with antibody or blood PCR positivity. The association of this assay to serological and molecular tests provides a better estimate of cat exposure to L. infantum.


Assuntos
Doenças do Gato/parasitologia , Doenças do Cão/parasitologia , Interferon gama/sangue , Leishmania infantum/imunologia , Leishmaniose Visceral/veterinária , Animais , Anticorpos Antiprotozoários/sangue , Anticorpos Antiprotozoários/imunologia , Doenças do Gato/imunologia , Gatos , Doenças do Cão/epidemiologia , Cães , Doenças Endêmicas/veterinária , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Imunofluorescência/veterinária , Imunidade Celular , Leishmaniose Visceral/imunologia , Masculino , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Sicília/epidemiologia , Espanha/epidemiologia
7.
Prev Vet Med ; 165: 1-7, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30851922

RESUMO

Senecavirus A (SVA) is a single-stranded RNA virus in the family Picornaviridae. Recently, SVA has been associated with idiopathic vesicular disease and increased neonate mortality outbreaks in the United States, Brazil, China, Colombia, and Thailand, with increasing incidence since 2014. Indirect detection by antibody detection methods, including indirect immunofluorescence assay (IFA), virus neutralization assay, and competitive or indirect enzyme-linked immunosorbent assays (ELISAs), have been reported in clinical and experimental trials. The objective of this study was to determine the seroprevalence of SVA in nonclinical affected herds in the United States. Individual samples were collected from 3654 and 2433 clinically healthy grower-finisher pigs and sows, respectively, from 219 unique commercial swine production sites. SVA seroprevalence was evaluated by SVA rVP1 ELISA and SVA IFA. The estimated seroprevalence for grower-finisher pigs and sows was 12.2% and 34.0%, respectively. The herd prevalence was 42.7% for grower-finisher farms and 75.8% for sow farms. The SVA rVP1 ELISA and SVA IFA exhibited a fair (sows) and moderate (grower-finisher) agreement at the herd level, while a fair agreement was observed at the individual level for both pig categories evaluated. The McNemar's test was significant at the individual and herd level (p < 0.05). In this study, we demonstrated the presence of SVA IgG antibodies in pigs from clinically healthy grower-finisher and sow herds. These results suggest that SVA is circulating subclinically in sow farms and grower-finisher pig farms in major swine producing-states in the United States.


Assuntos
Infecções por Picornaviridae/veterinária , Picornaviridae , Doenças dos Suínos/epidemiologia , Animais , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Imunofluorescência/veterinária , Infecções por Picornaviridae/diagnóstico , Infecções por Picornaviridae/epidemiologia , Estudos Soroepidemiológicos , Suínos , Doenças dos Suínos/diagnóstico , Doenças dos Suínos/virologia , Estados Unidos/epidemiologia
8.
Vet Res ; 50(1): 13, 2019 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-30777128

RESUMO

Equine herpesvirus type 5 (EHV5) is a ubiquitous, yet obscure pathogen in the horse population and is commonly associated with fatal equine multinodular pulmonary fibrosis (EMPF). To date, little is known about the precise pathogenesis of EHV5. Here, we evaluated the dynamics of EHV5 infection in representative ex vivo and in vitro equine models, using immunofluorescence staining and virus titration. EHV5 was unable to infect epithelial cells lining the mucosa of nasal and tracheal explants. Similarly, primary equine respiratory epithelial cells (EREC) were not susceptible to EHV5 following inoculation at the apical or basolateral surfaces. Upon direct delivery of EHV5 particles to lung explants, few EHV5-positive cell clusters were observed at 72 hours post-inoculation (hpi). These EHV5-positive cells were identified as cytokeratin-positive alveolar cells. Next, we examined the potential of EHV5 to infect three distinct equine PBMC populations (CD172a+ monocytes, CD3+ T lymphocytes and Ig light chain+ B lymphocytes). Monocytes did not support EHV5 replication. In contrast, up to 10% of inoculated equine T and B lymphocytes synthetized intracellular viral antigens 24 hpi and 72 hpi, respectively. Still, the production of mature virus particles was hampered, as we did not observe an increase in extracellular virus titer. After reaching a peak, the percentage of infected T and B lymphocytes decayed, which was partly due to the onset of apoptosis, but not necrosis. Based on these findings, we propose a model for EHV5 pathogenesis in the horse. Uncovering EHV5 pathogenesis is the corner step to finally contain or even eradicate the virus.


Assuntos
Gammaherpesvirinae/fisiologia , Infecções por Herpesviridae/veterinária , Doenças dos Cavalos/virologia , Infecções Tumorais por Vírus/veterinária , Animais , Linhagem Celular , Células Epiteliais , Imunofluorescência/veterinária , Infecções por Herpesviridae/virologia , Cavalos , Técnicas In Vitro , Infecções Tumorais por Vírus/virologia , Carga Viral/veterinária
9.
Jpn J Infect Dis ; 72(3): 168-172, 2019 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-30700657

RESUMO

We evaluated the prevalence of Anaplasma infection in 332 dogs from Ibaraki, Japan, using serological and molecular methods. An immunofluorescence antibody assay against Anaplasma phagocytophilum indicated that 7 of the 328 serum samples tested (2.1%) were positive for A. phagocytophilum. Screening by polymerase chain reaction (PCR) analysis demonstrated that 8 of the 331 peripheral blood samples tested (2.4%) were positive for Anaplasmataceae. Phylogenetic analysis of the partial 16S rRNA sequence of the PCR amplicons revealed that 6 sequences were most similar to the 16S rRNA sequence of a Wolbachia sp., and the remaining 2 to A. bovis. Further analysis by A. phagocytophilum-specific nested PCR demonstrated that 1 dog infected with A. bovis was also positive for A. phagocytophilum. This is the first study to report the dual infection of a dog in Japan with A. bovis and A. phagocytophilum.


Assuntos
Anaplasma/genética , Anaplasmose/epidemiologia , Doenças do Cão/epidemiologia , Doenças do Cão/microbiologia , Anaplasma/isolamento & purificação , Anaplasma phagocytophilum/isolamento & purificação , Anaplasmose/sangue , Animais , Anticorpos Antibacterianos/isolamento & purificação , DNA Bacteriano/genética , Doenças do Cão/sangue , Cães , Feminino , Imunofluorescência/veterinária , Japão/epidemiologia , Masculino , Reação em Cadeia da Polimerase/veterinária , Prevalência
10.
Vet Ophthalmol ; 22(3): 345-352, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30701644

RESUMO

OBJECTIVE: To provide a complete nerve architecture and neuropeptide distribution in the cat cornea. ANIMALS STUDIED: Two adult domestic cats. PROCEDURE: The cat corneas were stained with protein gene product (PGP) 9.5 antibody-a pan marker for nerve fibers-and then divided into four quarters and double labeled with calcitonin gene-related peptide (CGRP) or substance P (SP) antibodies. Relative corneal nerve fiber densities and nerve terminals were evaluated in whole mount images by computer-assisted analysis. RESULTS: An average of 21.5 ± 2.1 thick stromal nerves enters the cornea around the limbus where they split into many branches going up to the anterior stroma. Some branches link to each other, but most of them penetrate the basement membrane in the periphery to give origin to subbasal bundles, which run centripetally and merge to form a whirl-like structure (vortex) at the center. These nerve bundles send out many fine terminals that innervate the epithelial cells. Subbasal nerve density and nerve terminals were greater in the center than in the periphery of the cornea. Additionally, CGRP-positive central epithelial nerve fibers and terminals were more abundant than SP-positive nerves and terminals. CONCLUSION: The architecture of cat corneal nerves shows similarities to human and mouse cornea innervation. This study provides useful data for researchers who use the cat model to assess corneal nerve pathological alterations, as well as in the veterinary field where corneal opacities, ulcerations, and infections damage the nerves and decrease sensitivity.


Assuntos
Gatos/anatomia & histologia , Córnea/inervação , Nervo Oftálmico/anatomia & histologia , Animais , Feminino , Imunofluorescência/veterinária , Fibras Nervosas
11.
Parasitology ; 146(2): 246-252, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30058514

RESUMO

Strongyloidiosis by Strongyloides stercoralis is a disease of increasing interest in human and animal medicine. The scientific knowledge on canine strongyloidiosis is hindered by the poor diagnostics available. To assess the most sensitive and specific diagnostic method, feces and blood from 100 shelter dogs were screened for S. stercoralis by coprological, molecular and serological tests. Thirty-six dogs (36%) scored positive to S. stercoralis by coprology (22.3% to Baermann) and/or 30% to real time-polymerase chain reaction (rt-PCR). According to two composite reference standards (CRS) based on all coprological methods and rt-PCR (first CRS) or in combination with serology (second CRS), the most sensitive test was IFAT (93.8%; CI 82.8-98.7), followed by rt-PCR (80.6%; 95% CI 64-91.8) and Baermann (60.6%; 95% CI 42.1-77.1). The inconsistent shedding of L1 during the 4-week follow-up in infected dogs suggests the importance of multiple faecal collections for a reliable diagnosis. A combination of serological and coprological tests is recommended for the surveillance and diagnosis of S. stercoralis infection in dogs.


Assuntos
Doenças do Cão/parasitologia , Strongyloides stercoralis , Estrongiloidíase/veterinária , Animais , Anticorpos Anti-Helmínticos/sangue , Estudos de Coortes , DNA de Helmintos/análise , Doenças do Cão/diagnóstico , Doenças do Cão/epidemiologia , Cães , Ensaio de Imunoadsorção Enzimática/veterinária , Fezes/parasitologia , Feminino , Imunofluorescência/veterinária , Masculino , Curva ROC , Reação em Cadeia da Polimerase em Tempo Real , Sensibilidade e Especificidade , Strongyloides stercoralis/genética , Strongyloides stercoralis/imunologia , Estrongiloidíase/diagnóstico , Estrongiloidíase/epidemiologia
12.
Vet Pathol ; 56(2): 200-207, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30131013

RESUMO

Canine prostatic carcinoma is a relevant model for human prostatic carcinoma. Survivin is proposed as a biomarker of malignancy in human prostatic cancer. Sox9 is a stem cell marker required for prostate development and expressed in several adult tissues. The aims of the present study were to evaluate the patterns and expression levels of 2 putative stem cell markers, survivin and Sox9, in canine benign prostatic hyperplasia (BPH) and prostatic carcinoma to investigate their potential as stem cell markers. Immunohistochemistry with specific antibodies was performed on 3 samples of normal prostate gland, 18 samples of canine BPH, and 16 samples of prostatic carcinoma. The basal cell layer of normal and hyperplastic prostatic lobules had nuclear Sox9 immunolabeling and nuclear and rarely cytoplasmic survivin immunostaining, identifying them as potential stem cell markers. Significantly more frequent survivin and Sox9 expression (≥10% of nuclei) was observed in prostatic carcinoma as compared with BPH. The potential coexpression of survivin with Sox9, androgen receptor, and p63 was also investigated in selected BPH and prostatic carcinoma cases with immunofluorescence, and a partial colocalization was observed. Results indicate that Sox9 and survivin could be considered markers of stemness in canine prostate cells. Given its role in proliferation, cells in the basal cell layer with nuclear survivin expression are likely to be transit-amplifying cells that maintain some stem cell proprieties.


Assuntos
Doenças do Cão/metabolismo , Próstata/metabolismo , Neoplasias da Próstata/veterinária , Fatores de Transcrição SOX9/metabolismo , Células-Tronco/metabolismo , Survivina/metabolismo , Animais , Biomarcadores Tumorais/metabolismo , Doenças do Cão/diagnóstico , Doenças do Cão/patologia , Cães , Imunofluorescência/veterinária , Masculino , Próstata/patologia , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia
13.
J Vet Sci ; 20(1): 51-57, 2019 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-30481981

RESUMO

Monoclonal antibodies (MAbs) are widely applied in disease diagnoses. Herein, we report a MAb, WF-4, against Influenza A virus nucleoprotein (NP), its broad response with Influenza A virus, and its application in an immunohistochemistry (IHC) assay. WF-4 was screened by immunofluorescence assay (IFA). The results showed that its reactivity with baculovirus-expressed full-length recombinant NP (rNP) in Western blot (WB), indicating its IHC applicability. Fifteen Influenza A virus (reference subtypes H1 to H15) infected chicken embryonated chorioallantoic membranes (CAM), fixed by formalin, were all detectable in the WF-4-based IHC assay. Also, the reactivity of the IHC test with NP from experimentally inoculated H6N1 and from all recent outbreaks of H5 subtype avian Influenza A virus (AIV) field cases in Taiwan showed positive results. Our data indicate that CAM, a by-product of Influenza A virus preparation, is helpful for Influenza A virus-specific MAb characterization, and that the WF-4 MAb recognizes conserved and linear epitopes of Influenza A virus NP. Therefore, WF-4 is capable of detecting NP antigens via IHC and may be suitable for developing various tests for diagnosis of Influenza A virus and, especially, AIV infection.


Assuntos
Galinhas , Imuno-Histoquímica/veterinária , Vírus da Influenza A Subtipo H5N2/imunologia , Vírus da Influenza A/imunologia , Proteínas de Ligação a RNA/imunologia , Proteínas do Core Viral/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Membrana Corioalantoide/imunologia , Imunofluorescência/veterinária , Taiwan
14.
J Vet Med Sci ; 81(2): 314-320, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30584200

RESUMO

Duck Tembusu virus disease, caused by the duck Tembusu virus (DTMUV), can lead to a severe reduction in egg production and growth retardation in laying ducks and ducklings, respectively. In this study, we engineered a novel recombinant adenovirus expressing the E protein of DTMUV (rAd-E) in AAV-293 cells (analyzed by western blot and indirect immunofluorescence assays). Intramuscular immunization of Cherry Valley ducks with rAd-E was performed to evaluate host cellular and humoral immune responses. Compared to the phosphate-buffered saline administered group and the negative control wild-type adenovirus (wtAd) group, the rAd-E vaccinated group showed increased cellular and humoral responses. The results from the cytokine release and lymphocyte proliferation assays showed that rAd-E induced a stronger cellular immune response than the control group (P<0.01), 4 weeks after primary immunization. The results of enzyme-linked immunosorbent and virus neutralization assays showed that rAd-E induced higher titers of specific neutralizing antibodies, 2 weeks after primary immunization. The DTMUV challenge experiment showed a higher survival rate (80%) of ducks in the rAd-E group, when challenged with 0.5 ml (ELD50=10-2.67/0.2 ml) of the DTMUV strain AH-F10. These results indicate that rAd-E effectively protects ducks against DTMUV infection. Therefore, rAd-E could be a vaccine candidate to provide an effective and safe method for prevention and control of DTMUV infection.


Assuntos
Adenoviridae/imunologia , Patos/virologia , Infecções por Flavivirus/veterinária , Flavivirus/imunologia , Doenças das Aves Domésticas/virologia , Vacinas Sintéticas/genética , Vacinas Virais/genética , Adenoviridae/genética , Animais , Western Blotting/veterinária , Patos/imunologia , Infecções por Flavivirus/imunologia , Infecções por Flavivirus/virologia , Imunofluorescência/veterinária , Imunidade Celular/imunologia , Imunidade Humoral/imunologia , Testes de Neutralização/veterinária , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/virologia , Vacinas Virais/imunologia
15.
Vet Immunol Immunopathol ; 207: 1-9, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30593344

RESUMO

Since CD8+ T cells play an important role in resistance to infection with heartwater, effective vaccines against this disease will likely require identification of antigens that contain CD8+ T cell epitopes responsible for cytotoxic T lymphocyte (CTL) responses. With the use of the fluorescent antigen-transfected target cell (FATT)-CTL assay, IFN-γ ELISPOT and flow cytometry, peptides that induce CTL, proliferation of CD8 + T cells and IFN-γ production were identified as possible target antigens for vaccine development. Of particular relevance was the finding that different peptides from different antigens were able to elicit varied cytotoxic activities by immune peripheral blood mononuclear cells (PBMC) from heartwater immune tick-infected sheep. Several peptides derived from Erum0660, Erum2330, Erum2540, Erum2580 and Erum5000 induced CTL in immune sheep PBMC. Peptide Erum2540-6 was the only peptide that induced significant CTL, CD8+CD45RO+ and CD8+IFN-γ+ by PBMC from all three sheep, and Erum2540 and p2540-20 induced the highest % CTL response in all three outbred sheep. These results suggest that these epitopes may be of major importance in heartwater recombinant vaccine development.


Assuntos
Antígenos de Bactérias/imunologia , Ehrlichia ruminantium/imunologia , Peptídeos/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Vacinas Bacterianas/imunologia , Epitopos/imunologia , Feminino , Imunofluorescência/veterinária , Hidropericárdio/imunologia , Hidropericárdio/microbiologia , Hidropericárdio/prevenção & controle , Técnicas In Vitro , Ativação Linfocitária/imunologia , Masculino , Reação em Cadeia da Polimerase/veterinária , Ovinos/imunologia , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/microbiologia , Doenças dos Ovinos/prevenção & controle
16.
Braz. J. Vet. Res. Anim. Sci. (Online) ; 56(4): e158601, Dezembro 03, 2019. mapas, graf
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1048085

RESUMO

In the Guarapiranga dam region located in the metropolitan area of São Paulo, human cases have been reported of Brazilian spotted fever (BSF), a tick-borne disease caused by the bacterium Rickettsia rickettsii. In this area, R. rickettsii is known to be transmitted to humans by Amblyomma aureolatum, a typical dog tick that is not associated with horses. In other BSF-endemic areas, R. rickettsii transmission is associated with Amblyomma sculptum, a tick species that typically infest capybaras and horses. The Guarapiranga Dam bears abundant populations of capybaras and horses. However, since nothing is known about a possible cycle of transmission of R. rickettsii by A. sculptum in this area, this study evaluated such transmission by performing a serosurvey of horses living in the Guarapiranga Dam region. A total of 206 equids living in the margins of the Guarapiranga Dam were serologically tested for antibodies reactive to five Rickettsia species, four of the spotted fever group (R. rickettsii, R. parkeri, R. amblyommatis, R. rhipicephali) and one basal group species, R. bellii. Overall, 171 (83%) equids reacted positively to at least one Rickettsia species. A total of 160 (78%), 123 (60%), 80 (39%), 72 (35%), and 71 (34%), equid sera reacted to R. bellii, R. rickettsii, R. parkeri, R. rhipicephali, and R. amblyommatis, respectively, with endpoint titers ranging from 64 to 1024 for R. bellii, and 64 to 512 for the remaining four Rickettsia species. Endpoint titers to R. bellii (median: 256) was significantly higher (P<0.05) than the endpoint titers to the other four Rickettsia species, for which the median values varied from 64 to 128. A total of 65 (32%) equid sera showed endpoint titers to R. bellii at least 4-fold higher than those to any of the other four antigens, indicating that they have been exposed to R. bellii or a very closely related species. Our results provide serological evidence that the sampled equids were not frequently exposed to R. rickettsii-infected ticks. Since horses are a highly suitable sentinel for R. rickettsii transmission by A. sculptum, we conclude that this tick species has no epidemiological role in the transmission of R. rickettsii in the BSF-endemic area of the Guarapiranga Dam in the metropolitan area of São Paulo.(AU)


Na região da represa de Guarapiranga localizada na área metropolitana de São Paulo, têm sido relatados casos humanos de Febre Maculosa Brasileira (FMB), uma doença transmitida por carrapatos causada pela bactéria Rickettsia rickettsii. Nesta área de estudo, R. rickettsii é conhecida por ser transmitida aos seres humanos pelo Amblyomma aureolatum, um carrapato de cão que não está associado a cavalos. Em outras áreas endêmicas da FMB, a transmissão de R. rickettsii está associada ao Amblyomma sculptum, uma espécie de carrapato que normalmente infesta capivaras e cavalos. A represa de Guarapiranga possui populações abundantes de capivaras e cavalos; no entanto, como nada se sabe sobre um possível ciclo de transmissão de R. rickettsii por A. sculptum nessa área, este estudo avaliou essa transmissão realizando um levantamento sorológico em cavalos que vivem na região da represa de Guarapiranga. Um total de 206 equídeos que vivem nas margens da represa de Guarapiranga foram testados sorologicamente para cinco espécies de Rickettsia, sendo quatro do grupo da FMB (R. rickettsii, R. parkeri, R. amblyommatis, R. rhipicephali) e um do grupo basal (R. bellii). No geral, 171 (83%) equídeos reagiram positivamente a pelo menos uma espécie de Rickettsia. Um total de 160 (78%), 123 (60%), 80 (39%), 72 (35%) e 71 (34%), reagiram a R. bellii, R. rickettsii, R. parkeri, R. rhipicephali e R. amblyommatis, respectivamente, com títulos finais variando de 64 a 1024 para R. bellii e 64 a 512 para as quatro espécies restantes de Rickettsia. Os títulos finais para R. bellii (mediana: 256) foram significativamente maiores (P <0,05) do que os títulos para as outras quatro espécies de Rickettsia, para os quais os valores medianos variaram de 64 a 128. Um total de 65 (32%) equideos, os soros mostraram títulos finais para R. bellii pelo menos quatro vezes maior que os de qualquer um dos outros quatro antígenos, indicando que eles foram expostos a R. bellii ou a uma espécie muito próxima. Os resultados obtidos fornecem evidências sorológicas de que os equídeos amostrados não eram frequentemente expostos a carrapatos infectados por R. rickettsii. Como os cavalos são um sentinela altamente adequado para a transmissão de R. rickettsiipor A. sculptum, a conclusão obtida foi que essa espécie de carrapato não tem papel epidemiológico na transmissão da bactéria na área endêmica de FMB da represa de Guarapiranga na região metropolitana de São Paulo.(AU)


Assuntos
Animais , Imunofluorescência/veterinária , Cavalos/parasitologia , Febre Maculosa das Montanhas Rochosas/diagnóstico , Imunofluorescência/métodos
17.
BMC Vet Res ; 14(1): 391, 2018 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-30526618

RESUMO

BACKGROUND: Infectious bronchitis virus (IBV) is one of the leading causes of mortality and morbidity in chickens. There are numerous serotypes and variants, which do not confer cross protection resulting in failure of currently used IBV vaccines. Although variant IBV isolates with major genetic differences have been subjected to comparative studies, it is unknown whether minor genetic differences in IBV variants within a serotype are different in terms of pathogenesis and eliciting host responses. Two Massachusetts (Mass) variant IBV isolates recovered from commercial layer flocks in the Western Canadian provinces of Alberta (AB) and Saskatchewan (SK) were compared genetically and evaluated for their pathogenicity, tissue distribution and ability to recruit and replicate in macrophages. RESULTS: Although whole genome sequencing of these two Mass IBV isolates showed low similarity with the M41 vaccinal strain, they had an identical nucleotide sequence at open reading frames (ORFs) 3a, 3b, envelop (E), matrix (M), 5a and 5b. The rest of the ORFs of these 2 IBV isolates showed 99.9% nucleotide similarity. However, upon experimental infection, we found that the IBV isolate originating from AB was different to the one that originated in SK due to higher tracheal lesion scores and lower lung viral replication and lower genome loads in cecal tonsils. Nevertheless, both IBV isolates elicited host responses characterized by significant macrophage recruitment to the respiratory tract and there was evidence that both IBV isolates replicated within tracheal and lung macrophages. CONCLUSIONS: Overall, this study shows that Mass variant IBV isolates, although possessing minor genetic variations, can lead to significant differences in pathogenicity in young chickens. Further studies are required to investigate the pathogenicity of these two Mass variant IBV isolates in laying hens.


Assuntos
Infecções por Coronavirus/veterinária , Vírus da Bronquite Infecciosa , Doenças das Aves Domésticas/patologia , Alberta/epidemiologia , Animais , Sequência de Bases , Galinhas/virologia , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Feminino , Imunofluorescência/veterinária , Genoma Viral/genética , Vírus da Bronquite Infecciosa/genética , Masculino , Massachusetts , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/virologia , Saskatchewan/epidemiologia
18.
J Vet Intern Med ; 32(6): 1958-1964, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30307643

RESUMO

BACKGROUND: Because of poor sensitivity and questionable specificity of immunofluorescent antibody assays (IFAs), serological diagnosis of Bartonella species infections in dogs remains challenging. Despite limitations, IFA testing is the historical "gold standard" for Bartonella serodiagnosis in animals and humans. Because most diagnostic laboratories test against only 1 or 2 Bartonella spp., testing against a broader panel of Bartonella antigens may enhance diagnostic sensitivity and specificity. OBJECTIVE: To evaluate the sensitivity and specificity of Bartonella IFA using 8 cell culture-grown Bartonella spp. isolates. ANIMALS: Archived serum samples from 34 Bartonella spp. naturally exposed, polymerase chain reaction (PCR)-positive dogs and from 26 PCR-negative and IFA-negative dogs. METHODS: Bartonella IFA sensitivity and specificity were assessed using cell culture-grown whole cell antigens derived from 3 Bartonella henselae (Bh) strains (Bh Houston 1, Bh San Antonio Type 2, Bh California 1), 3 Bartonella vinsonii subsp. berkhoffii genotypes (Bvb I, II, and III), Bartonella koehlerae (Bk), and Bartonella quintana (Bq). RESULTS: Only 62% of 34 Bartonella spp. PCR-positive dogs were seroreactive to any of the 8 Bartonella IFA antigens, indicating low IFA sensitivity. PCR-positive dogs were most often IFA seroreactive to Bq (n = 15), to Bvb II (n = 13), or to both (n = 9) antigens. Of the 26 previously IFA-negative/PCR-negative dogs, 4 (15%) were seroreactive using the expanded antigen panel. CONCLUSION AND CLINICAL IMPORTANCE: Despite IFA testing of dogs against 8 different Bartonella isolates, IFA sensitivity remained poor, and specificity was only 85%. Development of a reliable serological assay is needed to facilitate the diagnosis of Bartonella infection in dogs.


Assuntos
Antígenos de Bactérias/imunologia , Infecções por Bartonella/veterinária , Bartonella/imunologia , Doenças do Cão/diagnóstico , Animais , Infecções por Bartonella/diagnóstico , Bartonella henselae/imunologia , Bartonella quintana/imunologia , Células Cultivadas , Doenças do Cão/microbiologia , Cães , Imunofluorescência/métodos , Imunofluorescência/veterinária , Reação em Cadeia da Polimerase/veterinária , Sensibilidade e Especificidade , Testes Sorológicos/métodos , Testes Sorológicos/veterinária , Febre das Trincheiras/diagnóstico , Febre das Trincheiras/veterinária
19.
J Fish Dis ; 41(12): 1803-1809, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30320411

RESUMO

Tilapia lake virus (TiLV) is an emerging disease threatening tilapia culture in many parts of the world. A cell line from the brain of tilapia, which was named TiB, was established, characterized and subcultured with more than 100 passages. The TiB cell line was optimally maintained at 27°C using medium 199 (M199) supplemented with 10% foetal bovine serum (FBS). Chromosome analysis revealed that 60% of TiB cells at passage 5 maintained the modal chromosome number 2n = 44, while at passage 60, there were 43% of TiB cells with the diploid chromosome number 2n = 50. A significant cytopathic effect was observed in TiB cells after infection with tilapia lake virus (TiLV-2017A), and the viral replication in the cells was confirmed by transmission electron microscopy, immunofluorescence assays and viral titres, indicating the susceptibility of TiB cells to TiLV-2017A. The viral titres of TiLV-2017A in TiB cells reached 107.43 TCID50 /ml within 10 days. The stable growth and susceptibility to fish viruses make TiB cells a useful tool for fish virus-host cell interaction and for immune response of fish.


Assuntos
Doenças dos Peixes/virologia , Infecções por Vírus de RNA/veterinária , Vírus de RNA/isolamento & purificação , Vírus de RNA/fisiologia , Tilápia , Replicação Viral , Animais , Encéfalo/virologia , Técnicas de Cultura de Células , Linhagem Celular , Imunofluorescência/veterinária , Microscopia Eletrônica de Transmissão/veterinária , Infecções por Vírus de RNA/virologia , Carga Viral
20.
Arch Razi Inst ; 73(2): 87-93, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-30242799

RESUMO

Canine ehrlichiosis is a very important emerging disease in India. This study is the first attempt screening a large number of canines in India for the detection of canine monocytic ehrlichiosis. In the present study, 510 blood samples of dogs were screened for the presence of Ehrlichia canis and other variants of Anaplasmataceae family by serological and molecular methods.Out of the 510 serum samples, 293 (57.5%) cases were found positive for the presence of E. canis antibodies through enzyme-linked immunosorbent assay (ELISA). Furthermore, and 45 (8.8%) and 1 (0.2%) specimens were positive for E. canis and A. platys, respectively, based on the polymerase chain reaction (PCR). In the clinical samples of E. canis, the minimum detection limit for PCR was9 ng. In the immunofluorescence assay (IFA), the positive blood samples showed comparable results with those obtained from the commercially available dot ELISA kit (giving equivalent IFA titer). The results of sequencing were compared with other reported isolates in various regions of the world, and a phylogenetic relationship was established. The 16S rRNA region that was amplified and sequenced for E. canis and A. platys was highly conserved and so was another Vir B9 region.


Assuntos
Anaplasma/isolamento & purificação , Anaplasmose/epidemiologia , Doenças do Cão/epidemiologia , Ehrlichia canis/isolamento & purificação , Ehrlichiose/veterinária , Anaplasmose/microbiologia , Animais , Proteínas de Bactérias/análise , Doenças do Cão/microbiologia , Cães , Ehrlichiose/epidemiologia , Ehrlichiose/microbiologia , Ensaio de Imunoadsorção Enzimática/veterinária , Imunofluorescência/veterinária , Índia/epidemiologia , Monócitos/microbiologia , Reação em Cadeia da Polimerase/veterinária , Prevalência , RNA Ribossômico 16S/análise , Estudos Soroepidemiológicos
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