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Recent progress in fluorescent protein development has generated a large diversity of near-infrared fluorescent proteins (NIR FPs), which are rapidly becoming popular probes for a variety of imaging applications. However, the diversity of NIR FPs poses a challenge for end-users in choosing the optimal one for a given application. Here we conducted a systematic and quantitative assessment of intracellular brightness, photostability, oligomeric state, chemical stability and cytotoxicity of 22 NIR FPs in cultured mammalian cells and primary mouse neurons and identified a set of top-performing FPs including emiRFP670, miRFP680, miRFP713 and miRFP720, which can cover a majority of imaging applications. The top-performing proteins were further validated for in vivo imaging of neurons in Caenorhabditis elegans, zebrafish, and mice as well as in mice liver. We also assessed the applicability of the selected NIR FPs for multicolor imaging of fusions, expansion microscopy and two-photon imaging.
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Introduction: The cerebellum is organized into functional regions each dedicated to process different motor or sensory inputs for controlling different locomotor behaviors. This functional regionalization is prominent in the evolutionary conserved single-cell layered Purkinje cell (PC) population. Fragmented gene expression domains suggest a genetic organization of PC layer regionalization during cerebellum development. However, the establishment of such functionally specific domains during PC differentiation remained elusive. Methods and results: We show the progressive emergence of functional regionalization of PCs from broad responses to spatially restricted regions in zebrafish by means of in vivo Ca2+-imaging during stereotypic locomotive behavior. Moreover, we reveal that formation of new dendritic spines during cerebellar development using in vivo imaging parallels the time course of functional domain development. Pharmacological as well as cell-type specific optogenetic inhibition of PC neuronal activity results in reduced PC dendritic spine density and an altered stagnant pattern of functional domain formation in the PC layer. Discussion: Hence, our study suggests that functional regionalization of the PC layer is driven by physiological activity of maturing PCs themselves.
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Spectrally diverse fluorescent proteins (FPs) provide straightforward means for multiplexed imaging of biological systems. Among FPs fitting standard color channels, blue FPs (BFPs) are characterized by lower brightness compared to other spectral counterparts. Furthermore, available BFPs were not systematically characterized for imaging in cultured mammalian cells and common model organisms. Here we introduce a pair of new BFPs, named Electra1 and Electra2, developed through hierarchical screening in bacterial and mammalian cells using a novel dual-expression vector. We performed systematic benchmarking of Electras against state-of-art BFPs in cultured mammalian cells and demonstrated their utility as fluorescent tags for structural proteins. The Electras variants were validated for multicolor neuroimaging in Caenorhabditis elegans, zebrafish larvae, and mice in comparison with one of the best in the class BFP mTagBFP2 using one-photon and two-photon microscopy. The developed BFPs are suitable for multicolor imaging of cultured cells and model organisms in vivo. We believe that the described dual-expression vector has a great potential to be adopted by protein engineers for directed molecular evolution of FPs.
Assuntos
Evolução Molecular Direcionada , Peixe-Zebra , Animais , Linhagem Celular , Diagnóstico por Imagem , Proteínas de Fluorescência Verde/química , Proteínas de Fluorescência Verde/genética , Proteínas Luminescentes/química , Proteínas Luminescentes/genética , Mamíferos , Camundongos , Peixe-Zebra/genéticaRESUMO
In vivo imaging of model organisms is heavily reliant on fluorescent proteins with high intracellular brightness. Here we describe a practical method for rapid optimization of fluorescent proteins via directed molecular evolution in cultured mammalian cells. Using this method, we were able to perform screening of large gene libraries containing up to 2 × 107 independent random genes of fluorescent proteins expressed in HEK cells, completing one iteration of directed evolution in a course of 8 days. We employed this approach to develop a set of green and near-infrared fluorescent proteins with enhanced intracellular brightness. The developed near-infrared fluorescent proteins demonstrated high performance for fluorescent labeling of neurons in culture and in vivo in model organisms such as Caenorhabditis elegans, Drosophila, zebrafish, and mice. Spectral properties of the optimized near-infrared fluorescent proteins enabled crosstalk-free multicolor imaging in combination with common green and red fluorescent proteins, as well as dual-color near-infrared fluorescence imaging. The described method has a great potential to be adopted by protein engineers due to its simplicity and practicality. We also believe that the new enhanced fluorescent proteins will find wide application for in vivo multicolor imaging of small model organisms.
Assuntos
Evolução Molecular Direcionada , Peixe-Zebra , Animais , Linhagem Celular , Corantes Fluorescentes/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Mamíferos/genética , Camundongos , Neurônios/metabolismo , Imagem Óptica , Peixe-Zebra/genética , Peixe-Zebra/metabolismoRESUMO
Dominant spinocerebellar ataxias (SCAs) are progredient neurodegenerative diseases commonly affecting the survival of Purkinje cells (PCs) in the human cerebellum. Spinocerebellar ataxia type 1 (SCA1) is caused by the mutated ataxin1 (Atx1) gene product, in which a polyglutamine stretch encoded by CAG repeats is extended in affected SCA1 patients. As a monogenetic disease with the Atx1-polyQ protein exerting a gain of function, SCA1 can be genetically modelled in animals by cell type-specific overexpression. We have established a transgenic PC-specific SCA1 model in zebrafish coexpressing the fluorescent reporter protein mScarlet together with either human wild type Atx1[30Q] as control or SCA1 patient-derived Atx1[82Q]. SCA1 zebrafish display an age-dependent PC degeneration starting at larval stages around six weeks postfertilization, which continuously progresses during further juvenile and young adult stages. Interestingly, PC degeneration is observed more severely in rostral than in caudal regions of the PC population. Although such a neuropathology resulted in no gross locomotor control deficits, SCA1-fish with advanced PC loss display a reduced exploratory behaviour. In vivo imaging in this SCA1 model may help to better understand such patterned PC death known from PC neurodegeneration diseases, to elucidate disease mechanisms and to provide access to neuroprotective compound characterization in vivo.
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Ataxina-1/genética , Modelos Animais de Doenças , Ataxias Espinocerebelares/genética , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/genética , Animais , Animais Geneticamente Modificados , Ataxina-1/fisiologia , Morte Celular , Progressão da Doença , Comportamento Exploratório , Genes Reporter , Humanos , Larva , Proteínas Luminescentes/genética , Células de Purkinje/patologia , Transgenes , Expansão das Repetições de Trinucleotídeos , Peixe-Zebra/crescimento & desenvolvimento , Proteínas de Peixe-Zebra/fisiologia , Proteína Vermelha FluorescenteRESUMO
Clostridioides difficile infection (CDI) is considered to be one of the most common healthcare-associated gastrointestinal infections in the United States. The innate immune response against C. difficile has been described, but the exact roles of neutrophils and macrophages in CDI are less understood. In the current study, Danio rerio (zebrafish) larvae are used to establish a C. difficile infection model for imaging the behavior and cooperation of these innate immune cells in vivo. To monitor C. difficile, a labeling protocol using a fluorescent dye has been established. A localized infection is achieved by microinjecting labeled C. difficile, which actively grows in the zebrafish intestinal tract and mimics the intestinal epithelial damage in CDI. However, this direct infection protocol is invasive and causes microscopic wounds, which can affect experimental results. Hence, a more noninvasive microgavage protocol is described here. The method involves delivery of C. difficile cells directly into the intestine of zebrafish larvae by intubation through the open mouth. This infection method closely mimics the natural infection route of C. difficile.
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Clostridioides difficile/fisiologia , Peixe-Zebra/microbiologia , Animais , Clostridioides difficile/crescimento & desenvolvimento , Infecções por Clostridium/microbiologia , Modelos Animais de Doenças , Corantes Fluorescentes/química , Vida Livre de Germes , Intestinos/microbiologia , Intestinos/patologia , Larva/microbiologia , Esporos Bacterianos/fisiologiaRESUMO
The cerebellum integrates sensory information and motor actions. Increasing experimental evidence has revealed that these functions as well as the cerebellar cytoarchitecture are highly conserved in zebrafish compared with mammals. However, the potential of zebrafish for modelling human cerebellar diseases remains to be addressed. Spinocerebellar ataxias (SCAs) represent a group of genetically inherited cerebellar diseases leading to motor discoordination that is most often caused by affected cerebellar Purkinje cells (PCs). Towards modelling SCAs in zebrafish we identified a small-sized PC-specific regulatory element that was used to develop coexpression vectors with tunable expression strength. These vectors allow for in vivo imaging of SCA-affected PCs by high-resolution fluorescence imaging. Next, zebrafish with SCA type 13 (SCA13) transgene expression were established, revealing that SCA13-induced cell-autonomous PC degeneration results in eye movement deficits. Thus, SCA13 zebrafish mimic the neuropathology of an SCA-affected brain as well as the involved loss of motor control and hence provide a powerful approach to unravel SCA13-induced cell biological pathogenic and cytotoxic mechanisms.
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Purkinje cells (PCs) are primarily affected in neurodegenerative spinocerebellar ataxias (SCAs). For generating animal models for SCAs, genetic regulatory elements specifically targeting PCs are required, thereby linking pathological molecular effects with impaired function and organismic behavior. Because cerebellar anatomy and function are evolutionary conserved, zebrafish represent an excellent model to study SCAs in vivo We have isolated a 258 bp cross-species PC-specific enhancer element that can be used in a bidirectional manner for bioimaging of transgene-expressing PCs in zebrafish (both sexes) with variable copy numbers for tuning expression strength. Emerging ectopic expression at high copy numbers can be further eliminated by repurposing microRNA-mediated posttranslational mRNA regulation.Subsequently, we generated a transgenic SCA type 13 (SCA13) model, using a zebrafish-variant mimicking a human pathological SCA13R420H mutation, resulting in cell-autonomous progressive PC degeneration linked to cerebellum-driven eye-movement deficits as observed in SCA patients. This underscores that investigating PC-specific cerebellar neuropathologies in zebrafish allows for interconnecting bioimaging of disease mechanisms with behavioral analysis suitable for therapeutic compound testing.SIGNIFICANCE STATEMENT SCA13 patients carrying a KCNC3R420H allele have been shown to display mid-onset progressive cerebellar atrophy, but genetic modeling of SCA13 by expressing this pathogenic mutant in different animal models has not resulted in neuronal degeneration so far; likely because the transgene was expressed in heterologous cell types. We developed a genetic system for tunable PC-specific coexpression of several transgenes to manipulate and simultaneously monitor cerebellar PCs. We modeled a SCA13 zebrafish accessible for bioimaging to investigate disease progression, revealing robust PC degeneration, resulting in impaired eye movement. Our transgenic zebrafish mimicking both neuropathological and behavioral changes manifested in SCA-affected patients will be suitable for investigating causes of cerebellar diseases in vivo from the molecular to the behavioral level.
Assuntos
Cerebelo/metabolismo , Modelos Animais de Doenças , Células de Purkinje/metabolismo , Ataxias Espinocerebelares/congênito , Animais , Animais Geneticamente Modificados , Cerebelo/crescimento & desenvolvimento , Cerebelo/fisiopatologia , Feminino , Regulação da Expressão Gênica , Masculino , RNA Mensageiro/metabolismo , Elementos Reguladores de Transcrição , Canais de Potássio Shaw/genética , Ataxias Espinocerebelares/genética , Ataxias Espinocerebelares/metabolismo , Peixe-Zebra , Proteínas de Peixe-Zebra/genéticaRESUMO
Parkinson's disease (PD) is a neurodegenerative disease characterized by α-synuclein-positive inclusion bodies and loss of neurons, including dopaminergic neurons. Difficulty in replicating PD phenotypes using animal models partly limits the understanding of PD and the therapy required. Although PD is strongly associated with aging, most experimental animals may not exhibit age-related symptoms. Herein, we demonstrate that Nothobranchius furzeri, a rapidly aging teleost with a short life span, exhibits age-dependent degeneration of dopaminergic and noradrenergic neurons and progression of α-synuclein pathologies. These pathological phenotypes are similar to those observed in human patients with PD. Amelioration of the cell loss by genetic depletion of α-synuclein suggests that α-synuclein is not a bystander but a causative protein of neurodegeneration. N. furzeri can reveal mechanisms underlying PD, especially of the idiopathic form that affects a majority of patients with PD, including α-synuclein-dependent neurodegeneration, age-dependent phenotypes, and progression of α-synuclein pathology.
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Dopamina/metabolismo , Neurônios/metabolismo , Doença de Parkinson/genética , alfa-Sinucleína/metabolismo , Animais , Modelos Animais de Doenças , Peixes , Fundulidae , HumanosRESUMO
The zebrafish is a well-established model organism in which to study in vivo mechanisms of cell communication, differentiation and function. Existing cell ablation methods are either invasive or they rely on the cellular expression of prokaryotic enzymes and the use of antibiotic drugs as cell death-inducing compounds. We have recently established a novel inducible genetic cell ablation system based on tamoxifen-inducible Caspase 8 activity, thereby exploiting mechanisms of cell death intrinsic to most cell types. Here, we prove its suitability in vivo by monitoring the ablation of cerebellar Purkinje cells (PCs) in transgenic zebrafish that co-express the inducible caspase and a fluorescent reporter. Incubation of larvae in tamoxifen for 8â h activated endogenous Caspase 3 and cell death, whereas incubation for 16â h led to the near-complete loss of PCs by apoptosis. We observed synchronous cell death autonomous to the PC population and phagocytosing microglia in the cerebellum, reminiscent of developmental apoptosis in the forebrain. Thus, induction of apoptosis through targeted activation of caspase by tamoxifen (ATTACTM) further expands the repertoire of genetic tools for conditional interrogation of cellular functions.
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Apoptose/genética , Caspase 8/genética , Cerebelo/citologia , Genes Transgênicos Suicidas , Células de Purkinje/fisiologia , Peixe-Zebra , Animais , Animais Geneticamente Modificados , Apoptose/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Sobrevivência Celular/genética , Cerebelo/efeitos dos fármacos , Cerebelo/metabolismo , Genes Reporter/efeitos dos fármacos , Genes Transgênicos Suicidas/efeitos dos fármacos , Fagocitose/genética , Células de Purkinje/efeitos dos fármacos , Tamoxifeno/farmacologia , Peixe-Zebra/embriologia , Peixe-Zebra/genéticaRESUMO
There has been accumulating evidence for a regionalized organization of the cerebellum, which was mostly deduced from anatomical mapping of axonal projections of cerebellar afferents. A likewise regionalization of the cerebellar output has been suggested from lesion studies and dye-tracer experiments, but its physiological targets as well as the functional relevance of such an output regionalization are less clear. Ideally, such functional regionalization should be proven noninvasively in vivo. We here provide evidence for such a regionalization of the output from the cerebellar cortex by genetically encoded transneuronal mapping of efferent circuits of zebrafish Purkinje neurons. These identified circuits correspond to distinct regionalized Purkinje cell activity patterns in freely behaving zebrafish larvae during the performance of cerebellar-dependent behaviors. Furthermore, optogenetic interrogation of selected Purkinje cell regions during animal behavior confirms the functional regionalization of Purkinje cell efferents and reveals their contribution to behavior control as well as their function in controlling lateralized behavioral output. Our findings reveal how brain compartments serve to fulfill a multitude of functions by dedicating specialized efferent circuits to distinct behavioral tasks.
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Cerebelo/fisiologia , Peixe-Zebra/fisiologia , Animais , Animais Geneticamente Modificados , Comportamento Animal/fisiologia , Mapeamento Encefálico , Sinalização do Cálcio , Córtex Cerebelar/anatomia & histologia , Córtex Cerebelar/crescimento & desenvolvimento , Córtex Cerebelar/fisiologia , Cerebelo/anatomia & histologia , Cerebelo/crescimento & desenvolvimento , Vias Eferentes/anatomia & histologia , Vias Eferentes/fisiologia , Optogenética , Células de Purkinje/fisiologia , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimentoRESUMO
A lethargic household dog was referred to a private hospital in Japan. Diagnosis was carried out by the polymerase chain reaction (PCR) method developed for human Orientia tsutsugamushi infection using the dog's anticoagulated peripheral blood. Karp, Kato and Kuroki-type genomes were detected and the dog was diagnosed with O. tsutsugamushi infection. These findings demonstrate that dogs can act as a host for O. tsutsugamushi and the PCR method developed for human beings can be used for the diagnosis of canine O. tsutsugamushi infection. A concurrent epidemiological study examined 10 asymptomatic dogs that were fed in the same area as the sick dog. Kuroki-type genome in all dogs, Gilliam-type genome in 6 dogs and Kawasaki-type genome in 3 dogs were detected. These results provide further evidence that dogs can be naturally infected with O. tsutsugamushi outdoors and that dogs play a role as a host in the lifecycle of O. tsutsugamushi.
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Doenças do Cão/diagnóstico , Doenças do Cão/epidemiologia , Orientia tsutsugamushi/isolamento & purificação , Tifo por Ácaros/diagnóstico , Tifo por Ácaros/epidemiologia , Tifo por Ácaros/veterinária , Animais , Doenças do Cão/genética , Cães , Genoma Bacteriano , Japão/epidemiologia , Orientia tsutsugamushi/genética , Reação em Cadeia da Polimerase , Tifo por Ácaros/genética , Tifo por Ácaros/microbiologiaRESUMO
The red nucleus is located in the rostral midbrain of the vertebrate brain and controls motor coordination during locomotion. It receives input from the cerebellum and sends its output to the spinal cord. The presence of the red nucleus is well established in tetrapods, and its existence has also been suggested in teleosts but its presence and position has still been under discussion. By using wheat germ agglutinin (WGA) as a genetically encoded anterograde tracer, we recently identified contralateral projections from the cerebellum to a putative red nucleus in the zebrafish midbrain tegmentum. In this report we further revealed red nucleus derived from this contralateral afferent from the cerebellum using WGA and contralateral projections to the hindbrain-spinal cord junction site using DiI-mediated retrograde tracing. Thus the structure that we have identified by anterograde and retrograde tracing fulfills the anatomical demands for the red nucleus: the location in the midbrain tegmentum, contralateral afferent from the cerebellum (cerebello-ruber projection) and contralateral efferent to the spinal cord (rubro-spinal projection).
RESUMO
Pancreatitis-associated protein (PAP)-III, also known as regenerating gene/regenerating islet-derived (Reg)-IIIγ, is a small secretory protein whose expression is substantially induced in injured nerves. Here, we found that PAP-III protein underwent proteolytic N-terminal processing by trypsin-like protease(s) in injured sciatic nerves after axotomy. In vitro studies demonstrated that the N terminus-truncated PAP-III (ΔN-PAP-III) polymerized into a filament with a relatively uniform diameter of 10-20 nm, and the filaments formed higher order structures in a Na(+) concentration-dependent manner. When the ΔN-PAP-III fibers were added to the culture media, the ΔN-PAP-III fibers were tightly attached to neurites and somata of primary cortical neurons in vitro. In contrast, little association with glial cells was observed. When dense matrices of ΔN-PAP-III fibers were sheeted on a culture dish, neurites preferentially adhered to the fibers, and neurite extension was enhanced. This neurite outgrowth activity was significantly suppressed by preincubation with antibodies against PAP-III. These results imply that the released PAP-III might be cleaved and forms ΔN-PAP-III fibers at the nerve injury sites. Consequently, these resulting fibers would provide regenerating axons with a platform for extension.
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Antígenos de Neoplasias/metabolismo , Axônios/metabolismo , Biomarcadores Tumorais/metabolismo , Lectinas Tipo C/metabolismo , Regeneração Nervosa/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Neuritos/metabolismo , Proteólise , Animais , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Células Cultivadas , Lectinas Tipo C/genética , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas Associadas a Pancreatite , Ratos , Ratos Wistar , Nervo Isquiático/lesões , Nervo Isquiático/metabolismo , Sódio/metabolismoRESUMO
Metal complexes with N-heterocyclic carbene (NHC) ligands have been widely used in catalytic chemistry and are now increasingly considered for the development of new chemical tools and metal based drugs. Ruthenium complexes of the type (p-cymene)(NHC)RuCl(2) interacted with biologically relevant thiols and selenols, which resulted in the inhibition of enzymes such as thioredoxin reductase or cathepsin B. Pronounced antiproliferative effects could be obtained provided that an appropriate cellular uptake was achieved. Inhibition of tumor cell growth was accompanied by a perturbation of metabolic parameters such as cellular respiration.
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Complexos de Coordenação/química , Metano/análogos & derivados , Rutênio/química , Compostos de Selênio/química , Compostos de Sulfidrila/química , Animais , Catepsina B/antagonistas & inibidores , Catepsina B/metabolismo , Proliferação de Células/efeitos dos fármacos , Complexos de Coordenação/síntese química , Complexos de Coordenação/toxicidade , DNA/metabolismo , Embrião não Mamífero/efeitos dos fármacos , Glutationa Redutase/antagonistas & inibidores , Glutationa Redutase/metabolismo , Células HT29 , Humanos , Células MCF-7 , Metano/química , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Tiorredoxina Dissulfeto Redutase/metabolismo , Tripsina/química , Tripsina/metabolismo , Peixe-Zebra/crescimento & desenvolvimentoRESUMO
Gingival mass lesions developed when cyclosporine was administered for 600 days to a female, 7-year-old, longhaired dachshund diagnosed with intractable immune-mediated hemolytic anemia (IMHA). Histopathology indicated hyperplastic suppurative gingivitis. As the anemia improved, the dosage of cyclosporine A (CsA) was markedly decreased, and the mass lesions decreased in size and disappeared, thus suggesting that the mass lesions were an adverse reaction to CsA.
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Ciclosporina/efeitos adversos , Doenças do Cão/induzido quimicamente , Hiperplasia Gengival/veterinária , Imunossupressores/efeitos adversos , Anemia Hemolítica/tratamento farmacológico , Anemia Hemolítica/veterinária , Animais , Ciclosporina/uso terapêutico , Cães , Feminino , Hiperplasia Gengival/induzido quimicamente , Imunossupressores/uso terapêuticoRESUMO
Domesticated adult dogs with antibody titer classified as below 'high' to one or more of canine distemper virus (CDV), canine parvovirus type-2 (CPV-2) and canine adenovirus type-1 (CAdV-1) were then given an additional inoculation, and the effectiveness of this booster evaluated 2 months later. Consequently, CDV and CAdV-1 antibody titer experienced a significant increase, but the same effect was not observed in the antibody titer of CPV-2. These findings suggest that with additional inoculation, a booster effect may be expected in increasing antibody titers for CDV and CAdV-1, but it is unlikely to give an increase in CPV-2 antibody titer.
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Adenovirus Caninos/imunologia , Vírus da Cinomose Canina/imunologia , Imunização Secundária , Parvovirus Canino/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Cães , Vacinas Combinadas/administração & dosagem , Vacinas Combinadas/imunologia , Vacinas Virais/administração & dosagemRESUMO
In the quest for the identification of catalytic transformations to be used in chemical biology and medicinal chemistry, we identified iron(III) meso-tetraarylporphines as efficient catalysts for the reduction of aromatic azides to their amines. The reaction uses thiols as reducing agents and tolerates water, air, and other biological components. A caged fluorophore was employed to demonstrate that the reduction can be performed even in living mammalian cells. However, in vivo experiments in nematodes (Caenorhabditis elegans) and zebrafish (Danio rerio) revealed a limitation to this method: the metabolic reduction of aromatic azides.
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Azidas/química , Compostos Férricos/química , Metaloporfirinas/química , Aminas/química , Animais , Caenorhabditis elegans , Catálise , Meio Ambiente , Células HeLa , Humanos , Oxirredução , Peixe-ZebraRESUMO
The objective of this study was to determine whether post-vaccination antibody titers vary according to body weight in adult dogs. Antibody titers against canine parvovirus type 2 (CPV-2), canine distemper virus (CDV), and canine adenovirus type 1 (CAdV-1) were measured for 978 domestic adult dogs from 2 to 6 y of age. The dogs had been vaccinated approximately 12 mo earlier with a commercial combination vaccine. The dogs were divided into groups according to their weight. It was found that mean antibody titers in all weight groups were sufficient to prevent infection. Intergroup comparison, however, revealed that CPV-2 antibody titers were significantly higher in the Super Light (< 5 kg) group than in the Medium (10 to 19.9 kg) and Heavy (> 20 kg) groups and were also significantly higher in the Light (5 to 9.9 kg) group than in the Heavy group. Antibody titers against CDV were significantly higher in the Super Light, Light, and Medium groups than in the Heavy group. There were no significant differences among the groups for the CAdV-1 antibody titers.
Pour vérifier que les taux d'anticorps chez des chiens vaccinés changeaient en fonction de leur poids après la vaccination par un vaccin commercial combiné, on a mesuré les anticorps antivirus de la parvovirose canine (CPV-2), de la maladie de Carré (CDV) et de l'encéphalite de Rubarth type-1 (CAdV-1) chez 978 chiens de compagnie agés de 2 à 6 ans, un an après leur vaccination. Par nos mesures, nous observons dans tous les groupes un taux satisfaisant d' immunisation moyen des animaux. Mais en comparant les groupes de poids, on s'aperçoit que pour la parvovirose canine CPV-2, le groupe des super-légers (< 5 kg) est significativement plus protégé en anticorps que les groupes de poids moyen (de 10 à 19,9 kg) et de poids le plus lourd (> 20 kg). De même les poids légers (de 5 à 9,9 kg) sont significativement mieux protégés que les poids lourds. Pour la maladie de Carré (CDV), les super-légers, les poids légers ou les groupes de poids moyen ont un taux d'anticorps significativement plus élevé que les plus lourds. Par contre pour l'Encéphalite de Rubarth (CAdV-1) aucune différence des taux d'anticorps dans les groupes de poids n'a été observée.(Traduit par les auteurs).
Assuntos
Adenovirus Caninos/imunologia , Anticorpos Antivirais/sangue , Vírus da Cinomose Canina/imunologia , Doenças do Cão/virologia , Parvovirus Canino/imunologia , Vacinas Virais/imunologia , Animais , Peso Corporal/imunologia , Doenças do Cão/imunologia , Doenças do Cão/prevenção & controle , Cães , Feminino , Masculino , Estatísticas não Paramétricas , Vacinação/veterinária , Vacinas Virais/administração & dosagemRESUMO
We determined the molecular characteristics of four proteins, BgP32, BgP45, BgP47, and BgP50, of Babesia gibsoni. Localization by subcellular fractionations followed by Western blotting revealed that the corresponding native proteins belong to merozoite surface protein family of B. gibsoni (BgMSP). Moreover, antisera against either rBgP45 or rBgP47 cross-reacted with all the proteins of the BgMSP family on ELISA and IFAT analyses. Of the four candidate antigens, ELISA with rBgP45 yielded high sensitivity, and ELISA with rBgP32 resulted in high specificity and in concordance with IFAT results.