Resultado da pesquisa (25)
Termo utilizado na pesquisa antibody
#1 - Distribution of nesfatin-1 expression in Bactrian camels and its impact on adipocyte glucose metabolism
Abstract in English:
The study aimed to investigate the distribution and expression of nesfatin-1 in Bactrian camels and its impact on adipocyte glucose metabolism. Polyclonal antibodies against a single antigenic epitope of NUCB2/nesfatin-1 protein were prepared. Nesfatin-1 expression was detected in various adipose tissues, including the hypothalamus, humps, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, pancreas, liver, and abdomen, using Western blot and qRT-PCR. Glucose uptake levels, pyruvic acid content, and hexokinase and phosphofructokinase activities were measured in cultured 3T3-L1 preadipocytes treated with nesfatin-1 in a high glucose state using a non-radiofluorimetric assay. The results showed that nesfatin-1 was found in all Bactrian camel tissues, with higher adipose tissue and pancreas expression. However, qRT-PCR analysis revealed higher expression of nesfatin-1 mRNA in the abdominal fat and liver. After nesfatin-1 treatment, a significant decrease (p<0.05) was observed in glucose uptake levels, hexokinase, and phosphofructokinase activities in 3T3-L1 cells, with a significant increase (p<0.05) in their pyruvic acid content. These findings suggest that nesfatin-1 is expressed at high levels in the abdominal fat of Bactrian camels and significantly affects adipocyte glucose metabolism by regulating blood glucose levels, inhibiting adipocyte differentiation, and promoting lipid droplet hydrolysis to provide energy for the organism, which may relate to the Bactrian camel’s ability to adapt to harsh environments.
Abstract in Portuguese:
The study aimed to investigate the distribution and expression of nesfatin-1 in Bactrian camels and its impact on adipocyte glucose metabolism. Polyclonal antibodies against a single antigenic epitope of NUCB2/nesfatin-1 protein were prepared. Nesfatin-1 expression was detected in various adipose tissues, including the hypothalamus, humps, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, pancreas, liver, and abdomen, using Western blot and qRT-PCR. Glucose uptake levels, pyruvic acid content, and hexokinase and phosphofructokinase activities were measured in cultured 3T3-L1 preadipocytes treated with nesfatin-1 in a high glucose state using a non-radiofluorimetric assay. The results showed that nesfatin-1 was found in all Bactrian camel tissues, with higher adipose tissue and pancreas expression. However, qRT-PCR analysis revealed higher expression of nesfatin-1 mRNA in the abdominal fat and liver. After nesfatin-1 treatment, a significant decrease (p<0.05) was observed in glucose uptake levels, hexokinase, and phosphofructokinase activities in 3T3-L1 cells, with a significant increase (p<0.05) in their pyruvic acid content. These findings suggest that nesfatin-1 is expressed at high levels in the abdominal fat of Bactrian camels and significantly affects adipocyte glucose metabolism by regulating blood glucose levels, inhibiting adipocyte differentiation, and promoting lipid droplet hydrolysis to provide energy for the organism, which may relate to the Bactrian camel’s ability to adapt to harsh environments.
#2 - Bovine neosporosis in Rio Grande do Sul, Brazil: Elevated antibody detection rate in comparison to previous decades
Abstract in English:
This study aimed to determine the frequency of anti-Neospora caninum antibody detection in three samples and sampling methods: A prevalence study, routine diagnostic laboratory samples, and fetal bovine serum samples. These samples were collected from cattle in Rio Grande do Sul (RS), southern Brazil, and analyzed using the indirect immunofluorescence reaction technique. For each sampling method, a historical study was used as a reference for comparison. In the prevalence study, 1,248 serum samples were collected from 2020 to 2022. The prevalence of N. caninum in the RS state was 22.8% (285/1248). This figure was statistically different compared to previous studies conducted in 2002, which reported a prevalence of 11.2% (p<0.001). In the routine diagnostic samples, an average rate of 29.95% (985/3289) of anti-N. caninum antibodies were detected. This rate was statistically higher than that of a previous study conducted in 2003, which reported a rate of 20% (p=0.01). Similar data were found in the fetal bovine serum samples, which showed an increase compared to previous studies conducted in 2010 that reported a rate of 15% (p=0.003). The increase in the detection rate of N. caninum antibodies underscores the need for measures to control and prevent bovine neosporosis.
Abstract in Portuguese:
Este estudo teve como objetivo determinar a frequência de detecção de anticorpos anti-Neospora caninum em três diferentes amostras e métodos de coleta: um estudo de prevalência, amostras de laboratório de diagnóstico de rotina e amostras de soro fetal bovino. Essas amostras foram coletadas de bovinos no Rio Grande do Sul (RS), região Sul do Brasil, e analisadas usando a técnica de reação de imunofluorescência indireta. Para cada método de coleta, um estudo histórico foi usado como referência para comparação. No estudo de prevalência, foram coletadas 1.248 amostras de soro entre 2020 e 2022. A prevalência de N. caninum no estado do RS foi de 22,8% (285/1248). Esse valor foi estatisticamente diferente quando comparado a estudos anteriores realizados em 2002, que relataram uma prevalência de 11,2% (p<0,001). Nas amostras de diagnóstico de rotina, foi detectada uma taxa média de 29,95% (985/3289) de anticorpos anti-N. caninum. Essa taxa foi estatisticamente maior do que a de um estudo anterior realizado em 2003, que relatou uma taxa de 20% (p=0,01). Dados semelhantes foram encontrados nas amostras de soro fetal bovino, que mostraram um aumento em comparação com estudos anteriores realizados em 2010 que relataram uma taxa de 15% (p=0,003). O aumento na taxa de detecção de anticorpos anti-N. caninum destaca a necessidade de medidas para controlar e prevenir a neosporose bovina.
#3 - Sarcocystis spp. detection in cattle using different diagnostic methods
Abstract in English:
Cattle are considered intermediate hosts of Sarcocystis, which can cause clinical signs and lower performance in the acute phase of infection. Sarcocystis spp. are usually not visible to the naked eye during the post mortem inspection. Moreover, fresh microscopic examination and transmission electron microscopy techniques are difficult to apply to large samples. Therefore, extensive studies on Sarcocystis infection in cattle using molecular and serological methods are required. Here, we investigated Sarcocystis spp. infection in cattle using fresh microscopic examination and polymerase chain reaction of myocardium samples and compared the results with the presence of antibodies against Sarcocystis spp. in corresponding serum samples detected using indirect fluorescent antibody test. Microscopic Sarcocystis were observed in 100% of the myocardial samples, and Sarcocystis DNA was present in 86% (43/50) of these samples. Antibodies against Sarcocystis spp. were detected in 96% (48/50) and 80% (40/50) of the serum samples at 1:25 and 1:200 dilutions, respectively. The three associated methods (fresh microscopic examination, PCR and serology) showed good sensitivity and detection for Sarcocystis spp. compared with fresh microscopic examination (only), and they may facilitate diagnosis in live animals on a large scale as well as monitoring of the herd status.
Abstract in Portuguese:
Os bovinos são considerados hospedeiros intermediários de Sarcocystis, podendo causar sinais clínicos e menor desempenho na fase aguda da infecção. Sarcocystis spp. geralmente não são visíveis a olho nu durante a inspeção post mortem. Além disso, o exame microscópico a fresco e as técnicas de microscopia eletrônica de transmissão são difíceis de aplicar a uma amostras de grande tamanho. Portanto, são necessários extensos estudos sobre a infecção por Sarcocystis em bovinos usando métodos moleculares e sorológicos. Aqui, investigamos a infecção de Sarcocystis spp. em bovinos por meio de exame microscópico a fresco e reação em cadeia da polimerase de amostras de miocárdio e comparado os resultados com a presença de anticorpos contra Sarcocystis spp. em amostras de soro correspondentes detectadas usando o teste de anticorpos fluorescentes indiretos. Sarcocistos microscópicos foram observados em 100% das amostras de miocárdio, e o DNA de Sarcocystis estava presente em 86% (43/50) dessas amostras. Anticorpos contra Sarcocystis spp. foram detectados em 96% (48/50) e 80% (40/50) das amostras de soro nas diluições 1:25 e 1:200, respectivamente. Os três métodos associados (exame microscópico a fresco, PCR e sorologia) mostraram boa sensibilidade e detecção para Sarcocystis spp. em comparação com o exame microscópico fresco (apenas) e podem facilitar o diagnóstico em animais vivos em larga escala, bem como o monitoramento do status do rebanho.
#4 - Use of oral fluids to detect anti Lawsonia intracellularis antibodies in experimentally infected pigs
Abstract in English:
Several pathogens and antibodies derived from serum or produced in tissues associated with the oral cavity are present in the oral fluid (OF). Considering the applicability of this alternative sample, recent studies in veterinary medicine have tested OF as a replacement for serum in diagnostic assays. The aim of this study was to standardize the immunoperoxidase monolayer assay (IPMA) to detect anti-Lawsonia intracellularis immunoglobulin A (IgA) and immunoglobulin G (IgG) in OF samples from experimentally infected pigs. Sixty-two pigs were divided into two groups: control (T1, n=30) and inoculated with L. intracellularis (T2, n=32). Blood, OF and fecal samples were collected at 0, 7, 14, 21, 28 and 42 days post-inoculation (dpi). Some adaptations of the standard technique for serum were made to IPMA for the detection of IgA and IgG in OF. The IPMA showed high specificity and sensitivity for serum samples and high specificity and moderate sensitivity for the detection of IgA and IgG in OF. There was high agreement between the results of serum IgG and OF IgA and IgG. Based on our results, oral fluid samples may be used for the evaluation and determination of anti-L. intracellularis antibodies in pigs, but not for individual diagnosis of swine proliferative enteropathy.
Abstract in Portuguese:
Vários patógenos e anticorpos derivados do soro ou produzidos em tecidos associados a cavidade oral estão presentes no fluido oral (FO). Considerando a aplicabilidade dessa amostra alternativa, estudos recentes em medicina veterinária têm testado o FO como substituto do soro para testes diagnósticos. O objetivo desse estudo foi padronizar a imunoperoxidase em monocamada de célula (IPMC) para a detecção de imunoglobulina A e imunoglobulina G anti-Lawsonia intracellularis em amostras de FO de suínos experimentalmente infectados. Um total de 62 suínos foram divididos em dois grupos: controle (T1, n=30) e inoculados com L. intracellularis (T2, n=32). Sangue, FO e amostras de fezes foram coletados aos 0, 7,14, 21, 28 e 42 dias após a inoculação (dpi). Algumas adaptações da técnica foram realizadas na técnica padrão da IPMC para a detecção de IgA e IgG. A IPMC demostrou alta especificidade e sensibilidade para amostras de soro e alta especificidade de moderada sensibilidade para a detecção de IgA e IgG em FO. Houve alta concordância entre resultados de detecção de IgG em soro com a IgA e IgG em amostras de FO. Baseado em nossos resultados, amostras de fluido oral podem ser usadas em avaliações e detecção de anticorpos anti-L. intracellularis em suínos, porém não de forma individual.
#5 - Occurrence of anti-Toxoplasma gondii antibody and evaluation of risk infection factors in goats raised in Sergipe state, Brazil
Abstract in English:
The objective was to determine, through indirect immunofluorescence reaction (RIFI, 1:64), the occurrence of IgG antibodies to Toxoplasma gondii and the risk factors associated with infection in goats in the state of Sergipe. To this study were used 675 samples of blood of animals from 41 farms of the three state mesoregions from 2013 to 2014. The occurrence of seropositive goats was 30.07%, with 90.24% of farms with seropositive animals. The distribution of titers obtained was 37.93%, 11.82%, 17.24%, 18.22%, and 17.77% for 64, 128, 512 and 1024 respectively. The risk factors observed were farms that did not have facilities (p=0.000, OR=2.30, CI 95%=1.41-3.74), with flooded soils (p=0.011, OR=2.94, CI 95%=1.27-6.79), which provided feed on the ground (p=0.032, OR=1.69, CI 95%=1.04-2.74), in uncovered cages (p=0.032, OR=1.69, CI 95%=1.04-2.74), pasture-based feed (p=0.003, OR=3.52, CI 95%=1.53-8.09), with access from cats to (p=0.031, OR=1.45, CI 95%=1.03-2.04) and introduced new breeders in the last five years (p=0.036, OR=1.58, CI 95%=1.02-2.74).
Abstract in Portuguese:
O objetivo desse estudo foi determinar, através da reação de imunofluorescência indireta (RIFI, 1:64), a ocorrência de anticorpos IgG para o Toxoplasma gondii e os fatores de risco associados à infecção em rebanhos caprinos do estado de Sergipe, a partir de 675 amostras de sangue de animais de 41 propriedades das três mesorregiões do estado entre os anos de 2013 e 2014. A ocorrência de caprinos soropositivos foi de 30,07% com 90,24% de propriedades com animais soropositivos. A distribuição dos títulos obtidos foi de 37,93%, 11,82%, 17,24%, 18,22% e 17,77% para 64, 128, 512 e 1024 respectivamente. Os fatores de riscos observados foram propriedades que não possuíam instalações (p=0,000, OR=2,30, IC 95%=1,41-3,74), com terrenos alagados (p=0,011, OR=2,94, IC 95%=1,27-6,79), que disponibilizam a ração no chão (p=0,032, OR=1,69, IC 95%=1,04-2,74), em cochos descobertos (p=0,000, OR=1,97, IC 95%=1,32-2,94), alimentação à base de pastagem (p=0,003, OR=3,52, IC 95%=1,53-8,09), com acesso de gatos à fonte de água (p=0,031, OR=1,45, IC 95%=1,03-2,04) e que introduziram novos reprodutores nos últimos cinco anos (p=0,036, OR=1,58, IC 95%=1,02-2,74).
#6 - Neospora caninum IgG antibody research and evaluation of the risk factors associated to the infection in ovine, State of Sergipe, Brazil, 37(8):813-819
Abstract in English:
ABSTRACT.- Rizzo H., De Jesus T.K.S., Gaeta N.C., Carvalho J.S., Pinheiro Júnior J.W., Gregory L., Gennari S.M. & Villalobos E.M.C. 2017. [Neospora caninum IgG antibody research and evaluation of the risk factors associated to the infection in ovine, State of Sergipe, Brazil.] Pesquisa de anticorpos IgG para Neospora caninum e avaliação dos fatores de risco em ovinos do Estado de Sergipe. Pesquisa Veterinária Brasileira 37(8):813-819. Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Campus Dois Irmãos, Rua Manoel de Medeiros s/n, Recife, PE 52171-900, Brazil. E-mail: hubervet@gmail.com
The aim of this study was to determine the occurrence of antibodies IgG against Neospora caninum and evaluate the risk factors associated with the infection in ovine herds, State of Sergipe, Brazil. Blood samples (n=1200) were collected from sheep raised in 60 sheep run located in the three mesoregions of the State of Sergipe, between 2011 e 2012. Antibodies were investigated by Indirect Immunofluorescence Antibody Test of which cutoff point was 50 and positive samples were diluted in base 2 until the last positive titer. Data from 15 variables was obtained from questionnaires given to farmers. Absolute and relative frequencies were determined and the risk factors were analyzed by Pearson’s Qui-Square test (p≤0,05). The occurrences of serum reactive sheep were 39.83% (478/1200). The occurrences of positive sheep and sheep run were 55.88% (380/680) and 88.24% (30/34) in the Eastern region; 21.42% (60/280) e 42.85% (6/14) in dry region and 15.83% (38/240) e 41.67% (5/12) in the backwoods respectively. Antibody titers ranged from 50 (n=459), represented 96.02% (459/478) of seropositive samples to 6400 (1/478). Among the significant variables in the multivariate analysis were considered risk factors for infection with N. caninum were, sheep run located in Eastern region (p=0.000, OR=4.64, CI95%=3.36-6.41); standing and running water sources (p=0.000 OR=2.03, CI95%=1.41-2.92), absence of quarantine (p=0.000, OR=2.71, CI95%=2.08-3.53), absence of dunghill (p=0.000, OR=3.14, CI95%=2.45-4.02), presence of dogs (p=0.000, OR=2.74, CI95%=1.73-433), presence of wild animals (p=0.000, OR=3.45, CI95%=2.44-4.87) and subsistence (p=0.000, OR=4.99, CI95%=3.15-7.92) or reproduction (p=0.002, OR=1.74, CI95%=1.22-2.49) livestock were important risk factors. Our results highlight the occurrence of N. caninum in the ovine herds from State of Sergipe. Management and location of sheep runs were important risk factors associated to the infection.
Abstract in Portuguese:
RESUMO.- Rizzo H., De Jesus T.K.S., Gaeta N.C., Carvalho J.S., Pinheiro Júnior J.W., Gregory L., Gennari S.M. & Villalobos E.M.C. 2017. [Neospora caninum IgG antibody research and evaluation of the risk factors associated to the infection in ovine, State of Sergipe, Brazil.] Pesquisa de anticorpos IgG para Neospora caninum e avaliação dos fatores de risco em ovinos do Estado de Sergipe. Pesquisa Veterinária Brasileira 37(8):813-819. Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Campus Dois Irmãos, Rua Manoel de Medeiros s/n, Recife, PE 52171-900, Brazil. E-mail: hubervet@gmail.com
O objetivo desse trabalho foi determinar a ocorrência de anticorpos IgG para Neospora caninum bem como avaliar os fatores de risco associados à infecção em rebanhos ovinos do estado de Sergipe, Brasil. Foram coletadas, nos anos de 2011 e 2012, 1200 amostras de sangue de ovinos oriundos de sessenta propriedades distribuídas em três mesorregiões do estado para pesquisa de anticorpos para N. caninum pela Reação de Imunofluorescência Indireta (RIFI) utilizando-se como ponto de corte de 50 e as amostras diluídas na base 2. Os dados de 34 variáveis estudadas foram obtidos a partir de questionários aplicados aos proprietários e analisados para se determinar a frequências absolutas e relativas e análise dos fatores de risco pelo teste Qui-quadrado de Pearson (p≤0,05). A ocorrência de ovinos soropositivos para N. caninum foi de 39,83% (478/1200). Em relação às mesorregiões a ocorrência de animais e propriedades positivas foi de, respectivamente, 55,88% (380/680) e 88,24% (30/34) na Leste; 21,42% (60/280) e 42,85% (6/14) no Agreste e 15,83% (38/240) e 41,67% (5/12) no Sertão. Os títulos de anticorpos variaram de 50, representando 96,02% (459/478) das amostras soropositivas, a 6400 (1/478). Dentre as variáveis significantes, na analise multivariada, que foram consideradas com fatores de risco para a infecção pelo N. caninum estavam propriedades localizadas na mesorregião Leste (p=0,000, OR=4,64, IC95%=3,36-6,41), presença de fonte de água parada e corrente (p=0,000 OR=2,03, IC95%=1,41-2,92), ausência de quarentena (p=0,000 OR=2,71, IC95%=2,08-3,53), não utilização de esterqueiras (p=0,000 OR=3,14, IC95%=2,45-4,02), criações com finalidade de subsistência (p=0,000 OR=4,99, IC95%=3,15-7,92), de reprodução (p=0,002, OR=1,74, IC95%=1,22-2,49), presença de cães (p=0,000 OR=2,74, IC95%=1,73-433) e circulação de animais silvestres nos rebanhos (p=0,000 OR=3,45, IC95%=2,44-4,87). Os resultados evidenciam a ocorrência de N. caninum em rebanhos ovinos sergipanos, demonstrando o manejo e a localização dos rebanhos no estado como importantes fatores de risco.
#7 - Effects of polysaccharide from Ophiopogon japonicus on immune response to Newcastle disease vaccine in chicken, 36(12):1155-1159
Abstract in English:
ABSTRACT.- Song X., Cao M., Yin Z., Jia R., Zou Y., Li L., Yue G., Liang X., Yin L. & He C. 2016. Effects of polysaccharide from Ophiopogon japonicus on immune response to Newcastle disease vaccine in chicken. Pesquisa Veterinária Brasileira 36(12):1155-1159. Natural Medicine Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Chengdu, 611130, China. E-mail: yinzhongq@163.com
In order to investigate the immune enhancement effects of Ophiopogon japonicus polysaccharide Ophiopogon japonicus (OJPS) on Newcastle disease (ND) live vaccine, chickens vaccinated against ND live vaccine was orally administered with the OJPS at high, medium and low concentrations respectively. In negative control group, chickens were given orally equal volume of physiological saline. On day 14, 21 and 28, the serum antibody titer, erythrocyte-C3b receptor rosette rate (E-C3bRR), erythrocyte-C3b immune complex rosette rate (E-ICRR) and peripheral lymphocyte proliferation were measured. The results showed that at most time points, the antibody titer, peripheral lymphocyte proliferation, E-C3bRR and elimination rate of immune complex of three OJPS administrating groups were significantly higher (P<0.05) than those in negative control group. It indicated that OJPS could significantly improve the immune efficacy of Newcastle disease live vaccine, Ophiopogon japonicus polysaccharide possessed synergistical immunoenhancement.
Abstract in Portuguese:
ABSTRACT.- Song X., Cao M., Yin Z., Jia R., Zou Y., Li L., Yue G., Liang X., Yin L. & He C. 2016. Effects of polysaccharide from Ophiopogon japonicus on immune response to Newcastle disease vaccine in chicken. Pesquisa Veterinária Brasileira 36(12):1155-1159. Natural Medicine Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Chengdu, 611130, China. E-mail: yinzhongq@163.com
In order to investigate the immune enhancement effects of Ophiopogon japonicus polysaccharide Ophiopogon japonicus (OJPS) on Newcastle disease (ND) live vaccine, chickens vaccinated against ND live vaccine was orally administered with the OJPS at high, medium and low concentrations respectively. In negative control group, chickens were given orally equal volume of physiological saline. On day 14, 21 and 28, the serum antibody titer, erythrocyte-C3b receptor rosette rate (E-C3bRR), erythrocyte-C3b immune complex rosette rate (E-ICRR) and peripheral lymphocyte proliferation were measured. The results showed that at most time points, the antibody titer, peripheral lymphocyte proliferation, E-C3bRR and elimination rate of immune complex of three OJPS administrating groups were significantly higher (P<0.05) than those in negative control group. It indicated that OJPS could significantly improve the immune efficacy of Newcastle disease live vaccine, Ophiopogon japonicus polysaccharide possessed synergistical immunoenhancement.
#8 - A double-antibody sandwich ELISA based on the porcine circovirus type 2 (PCV2) propagated in cell culture for antibody detection, 36(12):1171-1177
Abstract in English:
ABSTRACT.- Cruz T.F., Kanashiro T.M., Castro A.M.M.G., Baldin C.M., Richtzenhain L.J. & Araujo Jr J.P. 2016. A double-antibody sandwich ELISA based on the porcine circovirus type 2 (PCV2) propagated in cell culture for antibody detection. Pesquisa Veterinária Brasileira 36(12):1171-1177. Departamento de Microbiologia e Imunologia, Instituto de Biociências, Universidade Estadual Paulista Júlio de Mesquita Filho, Botucatu, SP 18618-970, Brazil. E-mail: jpessoa@ibb.unesp.br, tfcruz@yahoo.com.br
Few studies have described enzyme-linked immunosorbent assays (ELISAs) for the detection of antibodies against porcine circovirus type 2 (PCV2) based on antigens produced in cell culture. Furthermore, few articles have described viral purification techniques for members of the family Circoviridae. This occurs because circoviruses are difficult to isolate, noncytopathogenic, and produce low viral titres in cell culture. Thus, for overcoming these difficulties in the cultivation of PCV2, this study aimed to develop a double-antibody sandwich ELISA based on the cell culture antigen PCV2b for the quantification of anti-PCV2 antibodies. A 20% and 50% discontinuous sucrose cushion was used for viral purification, which enabled the separation of cell culture proteins in the 20% sucrose cushion and a greater viral concentration in the 50% sucrose cushion. Following isopycnic centrifugation, PCV2 was concentrated in the band with density values from 1.330 to 1.395g/cm3. Viral purification was assessed using SDS-PAGE, indirect ELISA and electron microscopy. The standardised ELISA revealed a strong linear correlation (r= 0.826, p<0.001) when compared with a commercial ELISA kit. The assay exhibited low variability (inter-assay coefficient of variation of 4.24% and intra-assay of 1.80%) and excellent analytical specificity conferred by the capture antibody produced in rabbit. Thus, this ELISA is a rapid, specific and convenient method for the detection of antibodies against PCV2 in studies of experimental and natural infection, and in monitoring the response to vaccination on commercial farms.
Abstract in Portuguese:
RESUMO.- Cruz T.F., Kanashiro T.M., Castro A.M.M.G., Baldin C.M., Richtzenhain L.J. & Araujo Jr J.P. 2016. A double-antibody sandwich ELISA based on the porcine circovirus type 2 (PCV2) propagated in cell culture for antibody detection. [Sandwich ELISA com duplo anticorpo baseado no circovirus suíno tipo 2 (PCV2) produzido em cultivo celular para detecção de anticorpos.] Pesquisa Veterinária Brasileira 36(12):1171-1177. Departamento de Microbiologia e Imunologia, Instituto de Biociências, Universidade Estadual Paulista Júlio de Mesquita Filho, Botucatu, SP 18618-970, Brazil. E-mail: jpessoa@ibb.unesp.br, tfcruz@yahoo.com.br
Há poucos relatos na literatura de métodos de ELISA (Enzyme-linked immunosorbent assay), para a detecção de anticorpos contra o circovírus suíno tipo 2 (PCV2), baseados em antígenos produzidos em cultivo celular, bem como uma escassez de trabalhos descrevendo técnicas de purificação viral para os membros da família Circoviridae. Isso ocorre, pois os circovírus são de difícil isolamento, não causam efeito citopático e produzem um baixo título viral em cultivo celular. Assim, para superar essas dificuldades encontradas no cultivo do PCV2, este estudo objetivou desenvolver um sandwich ELISA com duplo anticorpo, baseado no antígeno de PCV2 produzido em cultivo celular, para a quantificação de anticorpos anti-PCV2. Um colchão de sacarose descontínuo a 20% e 50% foi utilizado para a purificação viral, o qual possibilitou a separação das proteínas oriundas do cultivo celular no colchão de sacarose a 20% e uma maior concentração viral no colchão de sacarose a 50%. Com a ultracentrifugação isopícnica, o PCV2 ficou mais concentrado na banda com valores de densidade de 1,330 a 1,395g/cm3. A purificação viral foi avaliada pelas técnicas de SDS-PAGE, ELISA indireto e microscopia eletrônica. Assim, o método de ELISA padronizado revelou uma forte correlação linear (r = 0,826, p <0,001) quando comparado com um kit de ELISA comercial. O ensaio demonstrou baixa variabilidade (coeficientes de variação inter-teste de 4,24% e intra-teste de 1,80%) e uma excelente especificidade analítica conferida pelo anticorpo de captura produzido em coelho. Portanto, o método de ELISA demonstrou ser rápido, específico e conveniente para a detecção de anticorpos contra o PCV2 em estudos de infecção natural e experimental, além da monitoria da resposta à vacinação contra o PCV2 em granjas comerciais.
#9 - Cellulitis in Japanese quails fed oregano extract in the diet and inoculated with Escherichia coli, 36(9):831-836
Abstract in English:
ABSTRACT.- Sakamoto M.I., Esteves A.F., Reis C.A.C., Carregaro V.M.L., Fernandes N.L.M. & Fernandes J.I.M. 2016. [Cellulitis in Japanese quails fed oregano extract in the diet and inoculated with Escherichia coli.] Celulite em codornas japonesas alimentadas com extrato de orégano nas dietas e inoculadas com Escherichia coli. Pesquisa Veterinária Brasileira 36(9):831-836. Departamento de Produção Animal, Universidade Camilo Castelo Branco, Av. Hilário da Silva Passos 950, Parque Universitário, Descalvado, SP 13690-950, Brazil. E-mail: mizumiss@yahoo.com.br
The aim was to evaluate the use of oregano extract in Japanese quail (Coturnix coturnix japonica) diet, challenged with Escherichia coli strains, on performance, incidence of avian cellulitis and of antibody specific antigens against E. coli. Three hundred sixty Japanese quails with 90 days of age were distributed into galvanized wire cages in a conventional shed. The experimental design was completely randomized in factorial 5x2 design (oregano extract x challenged or not with E. coli), totaling ten treatments with six replicates of six birds per cage. Oregano extract levels were 0.00, 0.025, 0.050, 0.100 and 0.150%. Performance productive characteristics were evaluated, macroscopic lesions of cellulitis were measured after post-inoculation of the strains, and serum samples were collected for antibodies during experiment. The data were subjected to analysis of variance and averages compared by T test. Effect of E. coli was observed on all productive characteristics, regardless of the EO level evaluated, where challenged groups showed worse performance results. The macroscopic lesions, characteristic of cellulitis, observed only in birds of groups challenged with E. coli, were classified as mild and without bleeding. For specific antibodies, there was a higher number of birds challenged with E. coli strains in relation to unchallenged birds. It can be concluded that oregano extract supplemented in the diet was not effective against the challenge with E. coli in laying quails, and challenged birds with E. coli showed higher humoral and cellular immune response, compared with unchallenged birds, characterized by increased antibody titer and pectoral macroscopic lesion, regardless of the oregano extract levels evaluated.
Abstract in Portuguese:
RESUMO.- Sakamoto M.I., Esteves A.F., Reis C.A.C., Carregaro V.M.L., Fernandes N.L.M. & Fernandes J.I.M. 2016. [Cellulitis in Japanese quails fed oregano extract in the diet and inoculated with Escherichia coli.] Celulite em codornas japonesas alimentadas com extrato de orégano nas dietas e inoculadas com Escherichia coli. Pesquisa Veterinária Brasileira 36(9):831-836. Departamento de Produção Animal, Universidade Camilo Castelo Branco, Av. Hilário da Silva Passos 950, Parque Universitário, Descalvado, SP 13690-950, Brazil. E-mail: mizumiss@yahoo.com.br
Com o objetivo de avaliar a utilização do extrato de orégano nas dietas de codornas japonesas (Coturnix coturnix japonica) e desafiadas com cepas de Escherichia coli, sobre as características de desempenho, a incidência de celulite aviária e titulação de anticorpos específicos contra antígenos de E. coli, foram utilizadas 360 codornas japonesas, com 90 dias de idade, distribuídas em gaiolas de arame galvanizado em galpão convencional. O delineamento experimental foi inteiramente casualizado, em esquema fatorial 5x2 (extrato de orégano x desafiado ou não com E. oli), totalizando dez tratamentos com seis repetições de seis aves por gaiola. Os níveis do extrato de orégano (EO) avaliados foram: 0,00; 0,025; 0,050; 0,100 e 0,150%. Foram avaliadas características de desempenho produtivo, lesões macroscópicas da celulite após períodos pós-inoculação das cepas e amostras de soro foram colhidas para verificar a titulação de anticorpos nas aves. Os dados obtidos foram submetidos à análise de variância e as médias comparadas pelo Teste T. Foi observado efeito de E. coli sobre todas as características produtivas, independentemente dos níveis de EO avaliados, onde grupos desafiados apresentaram piores resultados de desempenho. As lesões macroscópicas, características da celulite, observadas somente nas aves desafiadas com E. coli foram classificadas como grau leve e sem presença de hemorragias. Para a titulação de anticorpos específicos, houve maior quantificação para aves desafiadas com as cepas de E. coli em relação às não desafiadas. Pode-se concluir que o extrato de orégano suplementado nas rações não se mostrou eficaz frente ao desafio com E. coli em codornas na fase de postura e as aves desafiadas com E. coli apresentaram maiores respostas imunes humoral e celular, em relação às não desafiadas, caracterizadas pelo aumento na titulação de anticorpos e pela lesão macroscópica peitoral, independentemente dos níveis de extrato de orégano avaliados.
#10 - Immunomagnetic method associated with MesenCult® medium to obtain bone marrow mononuclear cells from rabbits negative for CD45 monoclonal antibody, 36(4):339-344
Abstract in English:
ABSTRACT.- Souza L.A., Silva L.A.F., Oliveira B.J.N.A., Lacerda E.P.S., Beletti M.E., Lima A.P., Dias T.A. & Eurides D. 2016. [Immunomagnetic method associated with MesenCult® medium to obtain bone marrow mononuclear cells from rabbits negative for CD45 monoclonal antibody.] Método imunomagnético associado ao meio MesenCult® na obtenção de células mononucleares da medula óssea de coelhos negativas para o anticorpo monoclonal CD45. Pesquisa Veterinária Brasileira 36(4):339-344. Departamento de Medicina Veterinária, Escola de Veterinária e Zootecnia, Universidade Federal de Goiás, Campus Samambaia, Avenida Esperança s/n, Campus Samambaia, Goiânia, GO 74690-900, Brazil. E-mail: souza_vet@yahoo.com.br
The objective of this study was to describe guidelines for the isolation of bone marrow mononuclear cells from rabbits, followed by cell purification by negative depletion with CD45 monoclonal antibody, and further expansion in MesenCult® medium. Ten adult male New Zealand White rabbits, age average of 1.0±0.2 years and weighting 3.5±0.24kg, were used to obtain a standardized method. The mononuclear cells of the bone marrow were isolated with Ficoll-paque® density gradient centrifugation, and the cell purification and acquisition was completed by negative depletion with CD45 monoclonal antibody in immunomagnetic base. The cell population obtained was expanded in MesenCult® medium. Through isolation with Ficoll-paque® density gradient was possible to obtain an average yield of 7.31x106 cells/mL. After purification and acquisiton of potential mesenchymal stem cells by the immunomagnetic base, there was a yield decrease to 2.28x106 cells/mL; however the expansion process was increased in cell culture. The results indicated that cells obtained from the mononuclear fraction of bone marrow and cultivated in vitro were capable to generate adherent cells 24 hours after culture, with predominance of fibroblastoid cells suggestive of mesenchymal stem cells. It can be concluded that mesenchymal stem cells can be achieved with purified rabbit bone marrow mononuclear cells through the immunomagnetic method, as the MesenCult® medium provides a suitable environment for a quick in vitro expansion, and the number of passages exerts negative influence on the morphological characteristics.
Abstract in Portuguese:
RESUMO.- Souza L.A., Silva L.A.F., Oliveira B.J.N.A., Lacerda E.P.S., Beletti M.E., Lima A.P., Dias T.A. & Eurides D. 2016. [Immunomagnetic method associated with MesenCult® medium to obtain bone marrow mononuclear cells from rabbits negative for CD45 monoclonal antibody.] Método imunomagnético associado ao meio MesenCult® na obtenção de células mononucleares da medula óssea de coelhos negativas para o anticorpo monoclonal CD45. Pesquisa Veterinária Brasileira 36(4):339-344. Departamento de Medicina Veterinária, Escola de Veterinária e Zootecnia, Universidade Federal de Goiás, Campus Samambaia, Avenida Esperança s/n, Campus Samambaia, Goiânia, GO 74690-900, Brazil. E-mail: souza_vet@yahoo.com.br
O objetivo detse artigo é de descrever um protocolo de isolamento das células mononucleares da medula óssea de coelhos, seguido de purificação celular por depleção negativa com o anticorpo monoclonal CD45 e posterior expansão em meio de cultura MesenCult®. Dez coelhos machos adultos, da raça Nova Zelândia, com idade média de 1,0±0,2 anos e peso médio 3,5±0,24kg, foram utilizados para padronização da metodologia. O isolamento das células mononuclares da medula óssea foi realizado pelo gradiente de densidade Ficoll-paque® e a purificação e obtenção das células- pela depleção negativa com o anticorpo monoclonal CD45 em base imunomagnética. A população celular obtida foi expandida posteriormente em meio de cultura MesenCult®. No isolamento pelo gradiente de icoll-Paque® foi obtido um rendimento médio de 7,31x106 células/mL. Após purificação e obtenção das possíveis células-tronco mesenquimais pela base imunomagnética, houve um decréscimo do rendimento para 2,28x106 células/mL, mas o processo de expansão foi incrementado pelo cultivo celular. Os resultados indicaram que as células obtidas da fração mononuclear da medula óssea, cultivadas in vitro foram capazes de gerar células aderentes 24 horas após o cultivo, com predominância de células fibroblastóides sugestivas de células-tronco mesenquimais. Concluiu-se que a obtenção de células-tronco mesenquimais pode ser alcançada após purificação das células mononucleares da medula óssea de coelhos pelo método imunomagético, o meio de cultura MesenCult® proporciona um ambiente adequado para a rápida expansão in vitro e o número de passagens exerce influência negativa sobre as características morfológicas das células.
