Hyaluronidase from the venom of the social wasp Polybia paulista (Hymenoptera, Vespidae): Cloning, structural modeling, purification, and immunological analysis

In this study, we describe the cDNA cloning, sequencing, and 3-D structure of the allergen hyaluronidase from Polybia paulista venom (Pp-Hyal). Using a proteomic approach, the native form of Pp-Hyal was purified to homogeneity and used to produce a Pp-specific polyclonal antibody. The results revealed that Pp-Hyal can be classified as a glycosyl hydrolase and that the full-length Pp-Hyal cDNA (1315 bp; GI: 302201582) is similar (80-90%) to hyaluronidase from the venoms of endemic Northern wasp species. The isolated mature protein is comprised of 338 amino acids, with a theoretical pI of 8.77 and a molecular mass of 39,648.8 Da versus a pI of 8.13 and 43,277.0 Da indicated by MS. The Pp-Hyal 3D-structural model revealed a central core (α/β)7 barrel, two sulfide bonds (Cys 19-308 and Cys 185-197), and three putative glycosylation sites (Asn79, Asn187, and Asn325), two of which are also found in the rVes v 2 protein. Based on the model, residues Ser299, Asp107, and Glu109 interact with the substrate and potential epitopes (five conformational and seven linear) located at surface-exposed regions of the structure. Purified native Pp-Hyal showed high similarity (97%) with hyaluronidase from Polistes annularis venom (Q9U6V9). Immunoblotting analysis confirmed the specificity of the Pp-Hyal-specific antibody as it recognized the Pp-Hyal protein in both the purified fraction and P. paulista crude venom. No reaction was observed with the venoms of Apis mellifera, Solenopsis invicta, Agelaia pallipes pallipes, and Polistes lanio lanio, with the exception of immune cross-reactivity with venoms of the genus Polybia (sericea and ignobilis). Our results demonstrate cross-reactivity only between wasp venoms from the genus Polybia. The absence of cross-reactivity between the venoms of wasps and bees observed here is important because it allows identification of the insect responsible for sensitization, or at least of the phylogenetically closest insect, in order to facilitate effective immunotherapy in allergic patients. © 2013 Elsevier Ltd.

Humoral antibody immune response of newcastle disease vaccination in lovebirds (Agapornis roseicollis)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Broiler breeder feeding programs and trace minerals on maternal antibody transfer and broiler humoral immune response

Breeder feed restriction may negatively affect broiler progeny immunity. Sources of trace minerals (TM) with higher bioavailability in breeder diets have been reported to enhance humoral and cellular immunity in broiler progeny. An experiment was conducted to examine the effects of breeder feeding programs and TM dietary sources on maternal antibody transfer and humoral immune response of progeny to a live vaccine against Newcastle disease virus (NDV). Cobb 500 breeders were fed according to 2 feed allocation programs, sigmoid late fast and sigmoid late slow, from 14 to 29 wk of age. From 56 to 62 wk of age, breeders were fed with either inorganic TM or an organic source (OTM) to replace 30% of Cu, Zn, and Mn. Progeny broilers were vaccinated intraocularly with La Sota NDV vaccine at 7 d of age. Blood samples were collected at hatching, 4, and 14 d postvaccination. Serum antibody levels against NDV were assessed by ELISA and cytokine expression by real time PCR. At hatching, late slow breeder progeny fed diets with 30% OTM had higher antibody titers as compared with progeny of breeders fed 100% inorganic TM. Similar results were observed 2 wk postvaccination. Breeder feeding programs and TM sources affected the expression level of IL-4 in NDV vaccinated broiler progeny. It was concluded that breeder feeding programs influenced humoral immune response to NDV vaccine in the broiler progeny, and 30% OTM may increase these responses. © 2013 Poultry Science Association, Inc.

Lack of host specificity of Colletotrichum spp. isolates associated with anthracnose symptoms on mango in Brazil

The aim of the present study was to analyse the genetic and pathogenic variability of Colletotrichum spp. isolates from various organs and cultivars of mango with anthracnose symptoms, collected from different municipalities of São Paulo State, Brazil. Colletotrichum gloeosporioides isolates from symptomless citrus leaves and C. acutatum isolates from citrus flowers with post-bloom fruit drop symptoms were included as controls. Sequencing of the ITS region allowed the identification of 183 C. gloeosporioides isolates from mango; only one isolate was identified as C. acutatum. amova analysis of ITS sequences showed larger genetic variability among isolates from the same municipality than among those from different populations. fAFLP markers indicated high levels of genetic variability among the C. gloeosporioides isolates from mango and no correlation between genetic variability and isolate source. Only one C. gloeosporioides mango isolate had the same genotype as the C. gloeosporioides isolates from citrus leaves, as determined by ITS sequencing and fAFLP analysis. Pathogenicity tests revealed that C. gloeosporioides and C. acutatum isolates from either mango or citrus can cause anthracnose symptoms on leaves of mango cvs Palmer and Tommy Atkins and blossom blight symptoms in citrus flowers. These outcomes indicate a lack of host specificity of the Colletotrichum species and suggest the possibility of host migration. © 2012 British Society for Plant Pathology.

Label free redox capacitive biosensing

A surface confined redox group contributes to an interfacial charging (quantifiable by redox capacitance) that can be sensitively probed by impedance derived capacitance spectroscopy. In generating mixed molecular films comprising such redox groups, together with specific recognition elements (here antibodies), this charging signal is able to sensitively transduce the recognition and binding of specific analytes. This novel transduction method, exemplified here with C-reactive protein, an important biomarker of cardiac status and general trauma, is equally applicable to any suitably prepared interfacial combination of redox reporter and receptor. The assays are label free, ultrasensitive, highly specific and accompanied by a good linear range. © 2013 Elsevier B.V.

Validação de anticorpo monoclonal anti-CD45 obtido in house para utilização em citometria de fluxo e imuno-histoquímica

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Avaliação diagnóstica para Leishmania spp. e Trypanosoma cruzi em gatos domésticos procedentes da associação protetora dos animais do município de Ilha Solteira, SP, Brasil

Pós-graduação em Doenças Tropicais - FMB

Comparação entre os métodos de ELISA, imunofluorescência indireta e imunocromatografia para o diagnóstico da leishmaniose visceral canina

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Resposta sorológica de bezerras da raça Tabapuã recém-vacinadas com amostra B19 de Brucella abortus

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Desenvolvimento de imunossensores para aflatoxina B1

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Elucidação diagnóstica da leishmaniose visceral canina pra a vigilância epidemiológica e controle desta zoonose

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Epidemiologia da leishmaniose visceral canina no município do Poxoréo-MT

Pós-graduação em Medicina Veterinária - FMVZ

Infecção pelo HCV em uma unidade de diálise em Belém-Pará: desempenho de testes imunoenzimáticos em relação à técnica de biologia molecular

O presente estudo objetivou comparar o desempenho de quatro kits para detecção do anti-HCV, por enzimaimunoensaio, frente à detecção do HCV-RNA por meio de reação em cadeia pela polimerase (PCR), em 19 funcionários (FC) e 66 pacientes, sendo 54 submetidos à hemodiálise (HD) e 12 a diálise peritoneal (DP), em uma unidade de diálise em Belém, Pará. A pesquisa do anti-HCV foi realizada com kits de 3ª geração do Ortho, Ubi, BioChem e Abbott. A amplificação e quantificação de HCV-RNA foram realizadas utilizando-se kits do Roche. A concordância de resultados entre os quatro kits imunoenzimáticos foi de 89,5% entre os funcionários e 81,8% entre os pacientes. A prevalência do anti-HCV variou de 34,1 a 37,5% entre a população estudada, e de 42,4 a 47% entre os pacientes. O HCV-RNA foi detectado em 25 (37,9%) dos pacientes, dos quais 24 eram submetidos à hemodiálise. Não foi detectada infecção pelo HCV entre os funcionários. Não houve diferença estatística à análise dos índices de sensibilidade, especificidade e valores preditivos, entre os quatro kits em relação ao PCR, contudo, deixou-se de identificar, na dependência do kit utilizado, de 12,5 a 20% de indivíduos infectados pelo HCV. Não se observou correlação entre infecção pelo HCV e aumento nas aminotransferases. Foi possível mostrar correlação estatística entre a infecção pelo HCV e o tratamento por hemodiálise, tempo de tratamento superior a seis meses e idade dos pacientes superior a cinqüenta anos. Além disso, houve alta prevalência de portadores do HBV (13%) entre os hemodiálisados.

Avaliação de seis testes sorológicos no diagnóstico da brucelose bubalina

ABSTRACT: Four hundred and forty buffalo sera, selected from about 1,200 blood samples of another study, were examined. The samples were tested by six serological methods: two of agglutination, two of indirect ELISA and two of competitive ELISA. To determine the relative sensitivity and specificity of different tests, animals with a positive result to competitive ELISA of the FAO/IAEA were considered as infected. The relative sensitivity of competitive ELISA, indirect ELISA with conjugate anti-bovine light chain monoclonal antibody labelled with HRPO, indirect ELISA with anti-bovine IgG conjugate, rose Bengal test and rapid slide agglutination test was 100%, 98.57%, 97.14%, 91.42% and 79.28%, and the relative specificity 99.33%, 97.33%, 95.66%, 94.00% and 86.33%, respectively. The value of the different serological tests for the diagnosis of brucellosis is discussed.

Diagnóstico sorológico e molecular de Chlamydophila felis em gatos (Felis catus)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)