Effect of crude protein reduction in blood, performance, immunological, and intestinal histological parameters of broiler chickens.

We aimed to evaluate the effects of the reduction in dietary crude protein (CP) on blood urea, uric acid, performance, immunity, and intestinal histology of broilers. Four diets were formulated with 22.50%, 21.50%, 20.50%, and 19.50% of CP (1 to 21 days) and 19.20%, 18.20%, 17.20%, and 16.20% of CP (22 to 42 days), meeting the requirements of essential amino acids in all diets. A total of 800 male Ross chicks were randomly allocated to 32 pens, with 25 birds each (n = 8). Blood and intestines had been collected for analysis. Uric acid decreased and urea increased with the reduction of CP (p < 0.05). Reduction in performance and intestinal parameters (villus, crypt, and goblet cells) was observed with the reduction of CP (p < 0.05). Lower levels of CP resulted in alteration (p < 0.05) in CD4 and CD8 lineages (21 and 42 days). Broken-line models estimated (p < 0.05) the CP requirement for growth between 21% and 21.3% (1 to 21 days) and between 17.2% and 17.4% (22 to 42 days) and CP requirements between 17.2% and 18.2% for maximum response of immune cells (42 days). Reduction in dietary CP has a negative impact on performance, immune response, and intestinal histology of broilers, even with adequate levels of essential amino acids.

Histomorphological structure of the Bursa cloacalis (Bursa of Fabricius) in young Leiothrix lutea: a comparative study using light microscopy and transmission electron microscopy / Estructura histomorfológica de la Bursa cloacalis (Bursa de Fabricius) en jóvenes Leiothrix lutea: un estudio comparativo utilizando microscopía óptica y microscopía electrónica de transmisión

SUMMARY: This study aimed to investigate the microstructure and ultrastructure of the Bursa cloacalis (Bursa of Fabricius) (BC) in young Leiothrix lutea at various days of age (a few days after hatching) using light microscopy and transmission electron microscopy. The bird BC was sampled at 1, 5, 7, and 9 days of age. Immediately after dissection, the structure and integrity of the BC (not degenerative) were retained and the specific temporal features could be visualized precisely. After hematoxylin-eosin staining and uranyl acetate/lead citrate staining, the microstructure and ultrastructure of the BC, respectively, could be observed clearly. The microscopic observations revealed the following: in addition to change in the size of BC or lymphoid follicles, many cavities were found in the BC; the distribution of the lymphoid follicles in Leiothrix lutea was different from that in other birds; and the segregating line between the bursal cortex and medulla became increasingly clear as the age increased. In conclusion, the structural data obtained in this study provides a better understanding of the specific immunological function of the BC in Leiothrix lutea.

RESUMEN: Este estudio tuvo como objetivo investigar la microestructura y ultraestructura de la Bursa cloacalis (BC) en Leiothrix lutea joven unos días después de la eclosión, utilizando microscopía óptica y microscopía electrónica de transmisión. La BC se muestreó a los 1, 5, 7 y 9 días de edad del Leiothrix lutea. Inmediatamente después de la disección, se observó la estructura y la integridad de la CB (no degenerativa) y se pudo visualizar con precisión las características temporales específicas. Después de la tinción con hematoxilina-eosina y con acetato de uranilo /citrato de plomo, pudimos observar claramente la microestructura y ultraestructura de la BC. Las observaciones microscópicas revelaron el cambio en el tamaño de la CB o de los folículos linfoides y además, se encontraron numerosas cavidades en la CB; la distribución de los folículos linfoides en Leiothrix lutea era diferente a la de otras aves; y la línea de segregación entre la corteza bursal y la médula se hizo cada vez más clara a medida que aumentaba la edad. En conclusión, los datos estructurales obtenidos en este estudio proporcionan una mejor comprensión de la función inmunológica específica de la Bursa cloacalis en Leiothrix lutea.

Thyrotropin-Releasing Hormone (TRH) and Somatostatin (SST), but not Growth Hormone-Releasing Hormone (GHRH) nor Ghrelin (GHRL), Regulate Expression and Release of Immune Growth Hormone (GH) from Chicken Bursal B-Lymphocyte Cultures.

It is known that growth hormone (GH) is expressed in immune cells, where it exerts immunomodulatory effects. However, the mechanisms of expression and release of GH in the immune system remain unclear. We analyzed the effect of growth hormone-releasing hormone (GHRH), thyrotropin-releasing hormone (TRH), ghrelin (GHRL), and somatostatin (SST) upon GH mRNA expression, intracellular and released GH, Ser133-phosphorylation of CREB (pCREBS133), intracellular Ca2+ levels, as well as B-cell activating factor (BAFF) mRNA expression in bursal B-lymphocytes (BBLs) cell cultures since several GH secretagogues, as well as their corresponding receptors (-R), are expressed in B-lymphocytes of several species. The expression of TRH/TRH-R, ghrelin/GHS-R1a, and SST/SST-Rs (Subtypes 1 to 5) was observed in BBLs by RT-PCR and immunocytochemistry (ICC), whereas GHRH/GHRH-R were absent in these cells. We found that TRH treatment significantly increased local GH mRNA expression and CREB phosphorylation. Conversely, SST decreased GH mRNA expression. Additionally, when added together, SST prevented TRH-induced GH mRNA expression, but no changes were observed in pCREBS133 levels. Furthermore, TRH stimulated GH release to the culture media, while SST increased the intracellular content of this hormone. Interestingly, SST inhibited TRH-induced GH release in a dose-dependent manner. The coaddition of TRH and SST decreased the intracellular content of GH. After 10 min. of incubation with either TRH or SST, the intracellular calcium levels significantly decreased, but they were increased at 60 min. However, the combined treatment with both peptides maintained the Ca2+ levels reduced up to 60-min. of incubation. On the other hand, BAFF cytokine mRNA expression was significantly increased by TRH administration. Altogether, our results suggest that TRH and SST are implicated in the regulation of GH expression and release in BBL cultures, which also involve changes in pCREBS133 and intracellular Ca2+ concentration. It is likely that TRH, SST, and GH exert autocrine/paracrine immunomodulatory actions and participate in the maturation of chicken BBLs.

Identification of genogroups of Infectious Bursal Disease Virus in poultry farms at Colombia / Identificación de genogrupos del virus de la enfermedad de gumboro en granjas avícolas en colombia

RESUMEN El virus de la enfermedad de Gumboro (IBDV) es un avibirnavirus con genoma dsARN que presenta altas tasas de mutación y recombinación. A pesar del efecto inmunosupresor en aves y la frecuencia con que ocurre la infección por este agente en el país son pocos los estudios que caracterizan los cuadros clínicos y se desconoce cuáles son los genogrupos circulantes. Esta investigación tuvo como objetivo determinar la frecuencia de lesiones histopatológicas en órganos del sistema inmune e identificar los genogrupos del IBDV en aves comerciales de Colombia. Para determinar la frecuencia de presentación de lesiones en órganos del sistema inmune se analizaron 381 casos clínicos de las bases de datos del Laboratorio de Patología Aviar (LPA) de la Universidad Nacional de Colombia, sede Bogotá (periodo 2016-2018). Asimismo, se secuenciaron los productos de RT-PCR del gen que codifica para la proteína viral VP2 provenientes de 35 muestras de bursas de Fabricio. Como resultado se encontró evidencia de lesiones microscópicas compatibles con procesos de inmunodepresión en órganos del sistema inmune (bursa de Fabricio, timo, bazo y médula ósea) en el 25 % (97) de los casos analizados y se identificaron los genogrupos 1, 2 y 4 en la siguiente proporción: genogrupo 1-69 % (virus clásicos), genogrupo 2-25 % (variantes) y genogrupo 4-6 % (identificado en Suramérica). Estos hallazgos demuestran la presencia de lesiones en órganos del sistema inmune y la existencia de los genogrupos 1, 2 y 3 del IBDV circulando en aves comerciales en Colombia. Esta es la primera investigación en el país con este sistema de clasificación que permite evidenciar con mayor precisión los cambios en el genoma del IBDV. Lo anterior señala la necesidad de continuar con este tipo de estudios para tener una mejor comprensión de la infección en campo y orientar el diseño e implementación de estrategias de control.

ABSTRACT Infectious Bursal Disease Virus (IBDV) is an avibirnavirus with a dsRNA genome, which has a high mutation and recombination rates. Despite the immunosuppressive effect in poultry and the frequency of infections by this agent, few studies in Colombia that characterize the clinical signs and identify the circulating genogroups have been reported. This study aimed to determine the frequency of histopathological lesions in immune organs and to identify IBDV genogroups in poultry from Colombia. In this way, the Laboratorio de Patología Aviar (LPA) databases from the Universidad Nacional de Colombia (period 2016-2018) were analyzed in order to identify the frequency of lesions present in immune organs. 35 samples of the bursa of Fabricius, positive by RT-PCR, were sequenced to the gene that codes for the VP2 protein. 97 (25 %) cases showed microscopic lesions in the immune organs. The genogroups identified and the frequencies were: genogroup 1-69 % (classical viruses), genogroup 2-25 % (antigenic variants), and genogroup 4-6 % (identified in South America). These findings demonstrate the lesions of immune organs and the presence of different genogroups circulating in commercial birds in Colombia. This indicates the need to continue with these studies in order to have a better understanding of the infection in the field and to guide the design and implementation of control strategies.

The gene ontology differs in bursa of Fabricius between two breeds of ducks post hatching by enriching the differentially expressed genes

The bursa of Fabricius (BF) is the central humoral immune organ unique to birds. The present study investigated the possible difference on a molecular level between two duck breeds. The digital gene expression profiling (DGE) technology was used to enrich the differentially expressed genes (DEGs) in BF between the Jianchang and Nonghua-P strains of ducks. DGE data identified 195 DEGs in the bursa. Gene Ontology (GO) analysis suggested that DEGs were mainly enriched in the metabolic pathways and ribosome components. Pathways analysis identified the spliceosome, RNA transport, RNA degradation process, Jak-STAT signaling pathway, TNF signaling pathway and B cell receptor signaling pathway. The results indicated that the main difference in the BF between the two duck strains was in the capabilities of protein formation and B cell development. These data have revealed the main divergence in the BF on a molecular level between genetically different duck breeds and may help to perform molecular breeding programs in poultry in the future.(AU)

The gene ontology differs in bursa of Fabricius between two breeds of ducks post hatching by enriching the differentially expressed genes

The bursa of Fabricius (BF) is the central humoral immune organ unique to birds. The present study investigated the possible difference on a molecular level between two duck breeds. The digital gene expression profiling (DGE) technology was used to enrich the differentially expressed genes (DEGs) in BF between the Jianchang and Nonghua-P strains of ducks. DGE data identified 195 DEGs in the bursa. Gene Ontology (GO) analysis suggested that DEGs were mainly enriched in the metabolic pathways and ribosome components. Pathways analysis identified the spliceosome, RNA transport, RNA degradation process, Jak-STAT signaling pathway, TNF signaling pathway and B cell receptor signaling pathway. The results indicated that the main difference in the BF between the two duck strains was in the capabilities of protein formation and B cell development. These data have revealed the main divergence in the BF on a molecular level between genetically different duck breeds and may help to perform molecular breeding programs in poultry in the future.

Induction of apoptosis in the bursa of Fabricius by vaccination against Gumboro disease.

Infectious bursal disease is a severe acute viral disease of young chickens, affecting mainly the B-lymphocytes in the bursa of Fabricius, leading to severe immunosuppression as a result of the death of lymphoid cells. In the bursa infected with infectious bursal disease virus, viral replication is associated with apoptosis of lymphoid cells, inflammatory change and atrophy. Vaccination has appeared to be a crucial factor for control, with live attenuated vaccines being the most used. However, the apoptotic effect of these vaccines on the bursa has not been tested. We determined the apoptotic effect caused by the most used vaccines in local production on the bursa of Fabricius cells and the correlation with histological changes. In this study, it was demonstrated that apoptosis levels in the vaccinated groups were higher than those observed in the non-vaccinated birds leading to the conclusion that the action of the live virus vaccine strains modifies the boundary of the bursa and shapes processes of cell death by apoptosis. In contrast to other studies, the vaccine strains used did not show the phenomenon of bursal atrophy during the experimental period.

Relationship between virus replication and apoptosis events in IgM + cells from chicken spleen and bursa of Fabricius infected with Malaysia strain of very virulent infectious bursal disease virus

Background: Infection of IBDV was reported to be endemic in worldwide including Malaysia and can be spread orally thru polluted fodder and water source, thus causing economic losses especially in commercial poultry industry. The infection resulted in depletion of B lymphocytes and subsequently destruction of the bursa which leaded to immunosuppression of the bird and it was postulated that the depletion of cells in the bursa was due to induction of apoptosis. In the current study, the infection of Malaysia isolated very virulent IBDV UPM0081 on IgM bearing B lymphocytes (IgM+ cells) from chicken spleen and bursa was compared. Materials, Methods & Results: A total of sixty eggs were obtained and raised until the age of 3 weeks old. The birds were divided into two groups (n = 30), which one of them served as control while IBDV strain UPM0081 was used to infect another group of birds at the concentration of 103 ELD50. The birds were observed and sacrificed at day 2, 4 and 5 post infections. Spleen and bursa of Fabricius were harvested and subjected to IgM+ cell enrichment using microbeads. The cell viability of enriched cells was assayed using MTT and cell cycle was analyzed using propidium iodide. Annexin V FITC and acridine orange/propidium iodide double stain assays were used to determine the event of apoptosis in the enriched IgM+ cells. Also, the IBDV viral load was also [...]

Relationship between virus replication and apoptosis events in IgM + cells from chicken spleen and bursa of Fabricius infected with Malaysia strain of very virulent infectious bursal disease virus

Background: Infection of IBDV was reported to be endemic in worldwide including Malaysia and can be spread orally thru polluted fodder and water source, thus causing economic losses especially in commercial poultry industry. The infection resulted in depletion of B lymphocytes and subsequently destruction of the bursa which leaded to immunosuppression of the bird and it was postulated that the depletion of cells in the bursa was due to induction of apoptosis. In the current study, the infection of Malaysia isolated very virulent IBDV UPM0081 on IgM bearing B lymphocytes (IgM+ cells) from chicken spleen and bursa was compared. Materials, Methods & Results: A total of sixty eggs were obtained and raised until the age of 3 weeks old. The birds were divided into two groups (n = 30), which one of them served as control while IBDV strain UPM0081 was used to infect another group of birds at the concentration of 103 ELD50. The birds were observed and sacrificed at day 2, 4 and 5 post infections. Spleen and bursa of Fabricius were harvested and subjected to IgM+ cell enrichment using microbeads. The cell viability of enriched cells was assayed using MTT and cell cycle was analyzed using propidium iodide. Annexin V FITC and acridine orange/propidium iodide double stain assays were used to determine the event of apoptosis in the enriched IgM+ cells. Also, the IBDV viral load was also [...](AU)

Direct antiapoptotic effects of growth hormone are mediated by PI3K/Akt pathway in the chicken bursa of Fabricius.

Growth hormone (GH) is expressed in several extra-pituitary tissues, including the primary and secondary lymphoid organs of the immune system. In birds, GH mRNA and protein expression show a specific developmental distribution pattern in the bursa of Fabricius (BF), particularly in epithelial and B cells. Changes in the bursal concentration and distribution of locally produced GH during ontogeny suggest it is involved in B cell differentiation and maturation, as well as in a functional survival role in this organ, which may be mediated by paracrine/autocrine mechanisms. Here, we analyzed the anti-apoptotic effect of GH in BF and the intracellular signaling pathways involved in this activity. Also, we studied if this effect was exerted directly by GH or mediated indirectly by IGF-I. Bursal cell cultures showed an important loss of their viability after 4h of incubation and a significant increase in apoptosis. However, treatment with 10nM GH or 40 nM IGF-I significantly increased B cell viability (16.7 ± 0.67% and 13.4 ± 1.12%, respectively) when compared with the untreated controls. In addition, the presence of apoptotic bodies (TUNEL) dramatically decreased (5.5-fold) after GH and IGF-I treatments, whereas co-incubation with anti-GH or anti-IGF-I, respectively, blocked their anti-apoptotic effect. Likewise, both GH and IGF-I significantly inhibited caspase-3 activity (by 40 ± 2.0%) in these cultures. However, the use of anti-IGF-I could not reverse the GH anti-apoptotic effects, thus indicating that these were exerted directly. The addition of 100 nM wortmannin (a PI3K/Akt inhibitor) blocked the GH protective effects. Also, GH stimulated (3-fold) the phosphorylation of Akt in bursal cells, and adding wortmannin or an anti-GH antibody inhibited this effect. Furthermore, GH was capable to stimulate (7-fold) the expression of Bcl-2. Taken together, these results indicate that the direct anti-apoptotic activity of GH observed in the chicken bursal B cell cultures might be mediated through the PI3K/Akt pathway.

Cryptosporidium spp. in bursa of Fabricius of broiler chickens from Uruguay / Cryptosporidium spp. em bursa de Fabrícius de frangos de corte no Uruguai

The presence of Cryptosporidium spp. was detected in broiler chickens of 32-40 days of age, colonizing the bursa of Fabricius. The histologic study with haematoxylin and eosin staining was done on specimens of trachea, complete intestinal tract and bursa. Samples of intestinal content were also studied using Kinyoun staining technique. All birds with signs of illness were positive to the presence of parasite, showing a hypertrophic bursa of Fabricius, filled with caseous content. The bursal epithelial cells were full of different developmental stages of Cryptosporidium spp. This note describing the presence of Cryptosporidium spp. in industrial flocks of broiler chickens, is the first report of this parasite in Uruguay.(AU)

A presença de Cryptosporidium spp. foi detectada em frangos de corte de 32-40 dias de idade, colonizando a bursa de Fabrícius. O estudo histológico com hematoxilina e eosina foi feito em amostras de traqueia, trato intestinal completo e bursa. As amostras de conteúdo intestinal também foram estudadas utilizando a técnica de coloração de Kinyoun. Todas as aves com sinais de doença foram positivas para a presença do parasita, mostrando uma bursa de Fabrícius hipertrófica com grande quantidade de conteúdo caseoso. As células epiteliais bursais estavam repletas de diferentes estágios de desenvolvimento de Cryptosporidium spp. Esta nota, que descreve a presença de Cryptosporidium spp. em lotes industriais de frangos de corte, é o primeiro relato deste parasita no Uruguai.(AU)

Cryptosporidium spp. in bursa of Fabricius of broiler chickens from Uruguay / Cryptosporidium spp. em bursa de Fabrícius de frangos de corte no Uruguai

The presence of Cryptosporidium spp. was detected in broiler chickens of 32-40 days of age, colonizing the bursa of Fabricius. The histologic study with haematoxylin and eosin staining was done on specimens of trachea, complete intestinal tract and bursa. Samples of intestinal content were also studied using Kinyoun staining technique. All birds with signs of illness were positive to the presence of parasite, showing a hypertrophic bursa of Fabricius, filled with caseous content. The bursal epithelial cells were full of different developmental stages of Cryptosporidium spp. This note describing the presence of Cryptosporidium spp. in industrial flocks of broiler chickens, is the first report of this parasite in Uruguay.

A presença de Cryptosporidium spp. foi detectada em frangos de corte de 32-40 dias de idade, colonizando a bursa de Fabrícius. O estudo histológico com hematoxilina e eosina foi feito em amostras de traqueia, trato intestinal completo e bursa. As amostras de conteúdo intestinal também foram estudadas utilizando a técnica de coloração de Kinyoun. Todas as aves com sinais de doença foram positivas para a presença do parasita, mostrando uma bursa de Fabrícius hipertrófica com grande quantidade de conteúdo caseoso. As células epiteliais bursais estavam repletas de diferentes estágios de desenvolvimento de Cryptosporidium spp. Esta nota, que descreve a presença de Cryptosporidium spp. em lotes industriais de frangos de corte, é o primeiro relato deste parasita no Uruguai.

Desempenho e resposta imune de frangos de corte alimentados com dietas suplementadas com cromo na forma orgânica / Immune response and performance by broiler chickens dietarysupplemented with chromium in organic form

O experimento foi conduzido com o objetivo de avaliar o efeito da suplementação de cromo-orgânico, sobre o desempenho, biometria dos órgãos digestivos e linfoides, histomoformetria da bolsa cloacal e rendimento de carcaça e cortes nobres de frangos de corte de 1 a 41 dias de idade, naturalmente submetidos a estresse por calor cíclico. Foram utilizados 400 pintos de corte da linhagem Cobb, em um delineamento experimental inteiramente casualizado, com cinco tratamentos: 0; 350; 700; 1050 e 1400 ppb de cromo na ração. Os valores do índice de temperatura de globo e umidade foram respectivamente, 83,39 e 80, para fase de crescimento e terminação. A suplementação do cromo orgânico influenciou (P 0,01) de forma quadrática a conversão alimentar dos animais segundo a equação Y=1,67-0,00016x+0,00000012x² (r² 0,976), na fase de 1 a 21 dias de idade. Os níveis de suplementação do cromo orgânico proporcionaram melhor índice de eficiência produtiva e maior percentual de parênquima cortical nas aves aos 21 dias. Na fase de 1 a 41 dias de idade, verificou-se que o peso de penas foi influenciado positivamente com os níveis de cromo orgânico e que não houve efeito para o desempenho dos frangos. Conclui-se que a suplementação da ração com cromo orgânico melhora desempenho e a resposta imune de frangos de corte frente a condições de estresse por calor.(AU)

This study was conducted to evaluate the effect of adding organic chromium dietary supplementation on performance, digestiveand lymphoid organs biometry and histomorphometry of the clocal bursa and on carcass and prime cuts performance of broilersfrom 1 to 41 days old, naturally stressed by heat cyclic. We used 400 chicks Cobb, the experimental design adopted was acompletely randomized, with five treatments: 0; 350; 700; 1050 and 1400 ppb chromium in diet. The supplementation withorganic chromium influenced (p<0,01) quadratic form on alimentary conversion the animals according to the equation Y = 1.67to 0.00016 x 0.00000012 x ² (r ² 0.976) in the period 1 to 21. The levels of supplementation of organic chromium improvementproductive efficiency and percentage of cortical parenchyma of broilers to 21 days. In phase to 1 from 41 days old, it was observedimprovement the feathers weight with levels of organic chromium, and there wasnt influence on performance of broilers. Thechromium in the organic form improves the performance of broiler chickens and immune response by broiler chickens in cyclic heat stress.(AU)

Desempenho e resposta imune de frangos de corte alimentados com dietas suplementadas com cromo na forma orgânica / Immune response and performance by broiler chickens dietarysupplemented with chromium in organic form

O experimento foi conduzido com o objetivo de avaliar o efeito da suplementação de cromo-orgânico, sobre o desempenho, biometria dos órgãos digestivos e linfoides, histomoformetria da bolsa cloacal e rendimento de carcaça e cortes nobres de frangos de corte de 1 a 41 dias de idade, naturalmente submetidos a estresse por calor cíclico. Foram utilizados 400 pintos de corte da linhagem Cobb, em um delineamento experimental inteiramente casualizado, com cinco tratamentos: 0; 350; 700; 1050 e 1400 ppb de cromo na ração. Os valores do índice de temperatura de globo e umidade foram respectivamente, 83,39 e 80, para fase de crescimento e terminação. A suplementação do cromo orgânico influenciou (P 0,01) de forma quadrática a conversão alimentar dos animais segundo a equação Y=1,67-0,00016x+0,00000012x² (r² 0,976), na fase de 1 a 21 dias de idade. Os níveis de suplementação do cromo orgânico proporcionaram melhor índice de eficiência produtiva e maior percentual de parênquima cortical nas aves aos 21 dias. Na fase de 1 a 41 dias de idade, verificou-se que o peso de penas foi influenciado positivamente com os níveis de cromo orgânico e que não houve efeito para o desempenho dos frangos. Conclui-se que a suplementação da ração com cromo orgânico melhora desempenho e a resposta imune de frangos de corte frente a condições de estresse por calor.

This study was conducted to evaluate the effect of adding organic chromium dietary supplementation on performance, digestiveand lymphoid organs biometry and histomorphometry of the clocal bursa and on carcass and prime cuts performance of broilersfrom 1 to 41 days old, naturally stressed by heat cyclic. We used 400 chicks Cobb, the experimental design adopted was acompletely randomized, with five treatments: 0; 350; 700; 1050 and 1400 ppb chromium in diet. The supplementation withorganic chromium influenced (p<0,01) quadratic form on alimentary conversion the animals according to the equation Y = 1.67to 0.00016 x 0.00000012 x ² (r ² 0.976) in the period 1 to 21. The levels of supplementation of organic chromium improvementproductive efficiency and percentage of cortical parenchyma of broilers to 21 days. In phase to 1 from 41 days old, it was observedimprovement the feathers weight with levels of organic chromium, and there wasn’t influence on performance of broilers. Thechromium in the organic form improves the performance of broiler chickens and immune response by broiler chickens in cyclic heat stress.

Expression and function of chicken bursal growth hormone (GH).

Growth hormone (GH) has several effects on the immune system. Our group has shown that GH is produced in the chicken bursa of Fabricius (BF) where it may act as an autocrine/paracrine modulator that participates in B-cell differentiation and maturation. The time course of GH mRNA and protein expression in the BF suggests that GH may be involved in development and involution of the BF, since GH is known to be present mainly in B lymphocytes and epithelial cells. In addition, as GH is anti-apoptotic in other tissues, we assessed the possibility that GH promotes cell survival in the BF. This work focused on determining the mechanism by which GH can inhibit apoptosis of B cells and if the PI3K/Akt pathway is activated. Bursal cell cultures were treated with a range of GH concentrations (0.1-100nM). The addition of 10nM GH significantly increased viability (16.7±0.6%) compared with the control and decreased caspase-3 activity to 40.6±6.5% of the control. Together, these data indicate that GH is produced locally in the BF and that the presence of exogenous GH in B cell cultures has antiapoptotic effects and increases B cell survival, probably through the PI3k/Akt pathway.

Evaluation of follicular lymphoid depletion in the Bursa of Fabricius: an alternative methodology using digital image analysis and artificial neural networks / Avaliação da depleção linfóide folicular da Bursa de Fabricius: uma metodologia alternativa utilizando análise digital de imagem e redes neurais artificiais

Fifty Bursa of Fabricius (BF) were examined by conventional optical microscopy and digital images were acquired and processed using Matlab® 6.5 software. The Artificial Neuronal Network (ANN) was generated using Neuroshell® Classifier software and the optical and digital data were compared. The ANN was able to make a comparable classification of digital and optical scores. The use of ANN was able to classify correctly the majority of the follicles, reaching sensibility and specificity of 89 percent and 96 percent, respectively. When the follicles were scored and grouped in a binary fashion the sensibility increased to 90 percent and obtained the maximum value for the specificity of 92 percent. These results demonstrate that the use of digital image analysis and ANN is a useful tool for the pathological classification of the BF lymphoid depletion. In addition it provides objective results that allow measuring the dimension of the error in the diagnosis and classification therefore making comparison between databases feasible.(AU)

Cinquenta Bursa de Fabrícius (BF) foram examinadas através de microscopia óptica convencional e imagens digitais foram obtidas e processadas através do software Matlab® 6.5. Redes Neurais Artificiais (ANN) foram geradas com a utilização do software Neuroshell® Classifier, e os dados das análises óptica e digital foram comparados. A ANN classificou corretamente a maioria dos folículos, atingindo sensibilidade e especificidade de 89 por cento e 96 por cento, respectivamente. Quando os folículos foram agrupados de forma binária houve um aumento da sensibilidade para 90 por cento e obteve-se um valor máximo para a especificidade de 92 por cento. Estes resultados demonstram que o uso da análise digital de imagem associada à ANNé uma ferramenta bastante útil para a classificação patológica da depleção linfóide da BF. Além disso, fornece resultados objetivos que permitem medir a dimensão do erro classificatório, tornando possível a comparação entre distintos bancos de dados.(AU)

Evaluation of follicular lymphoid depletion in the Bursa of Fabricius: an alternative methodology using digital image analysis and artificial neural networks / Avaliação da depleção linfóide folicular da Bursa de Fabricius: uma metodologia alternativa utilizando análise digital de imagem e redes neurais artificiais

Fifty Bursa of Fabricius (BF) were examined by conventional optical microscopy and digital images were acquired and processed using Matlab® 6.5 software. The Artificial Neuronal Network (ANN) was generated using Neuroshell® Classifier software and the optical and digital data were compared. The ANN was able to make a comparable classification of digital and optical scores. The use of ANN was able to classify correctly the majority of the follicles, reaching sensibility and specificity of 89 percent and 96 percent, respectively. When the follicles were scored and grouped in a binary fashion the sensibility increased to 90 percent and obtained the maximum value for the specificity of 92 percent. These results demonstrate that the use of digital image analysis and ANN is a useful tool for the pathological classification of the BF lymphoid depletion. In addition it provides objective results that allow measuring the dimension of the error in the diagnosis and classification therefore making comparison between databases feasible.

Cinquenta Bursa de Fabrícius (BF) foram examinadas através de microscopia óptica convencional e imagens digitais foram obtidas e processadas através do software Matlab® 6.5. Redes Neurais Artificiais (ANN) foram geradas com a utilização do software Neuroshell® Classifier, e os dados das análises óptica e digital foram comparados. A ANN classificou corretamente a maioria dos folículos, atingindo sensibilidade e especificidade de 89 por cento e 96 por cento, respectivamente. Quando os folículos foram agrupados de forma binária houve um aumento da sensibilidade para 90 por cento e obteve-se um valor máximo para a especificidade de 92 por cento. Estes resultados demonstram que o uso da análise digital de imagem associada à ANNé uma ferramenta bastante útil para a classificação patológica da depleção linfóide da BF. Além disso, fornece resultados objetivos que permitem medir a dimensão do erro classificatório, tornando possível a comparação entre distintos bancos de dados.

Apoptose e expressão de VP2 e GAPDH na infecção precoce pelo vírus da doença infecciosa da bursa de Fabricius em pintos SPF / Apoptosis and expression of VP2 and GADPH in an experimental infectious bursal disease in SPF chicks

Vinte e nove pintos SPF de um dia foram inoculados com o vírus da doença infecciosa da bursa de Fabricius (VDIB) para avaliar a ocorrência precoce de apoptose e a expressão da proteína viral 2 (VP2) e da enzima gliceraldeído fosfato dehidrogenase (GAPDH). Os animais foram distribuídos em cinco grupos: 1-controle; e 2 a 5- com 24, 48, 72 e 96 horas pós-inoculação, respectivamente. Fragmentos da bursa de Fabricius foram colhidos para processamento histológico e extração de RNA. Lâminas coradas em HE e TUNEL (marcação in situ da fragmentação do genoma com transferase terminal de deoxinucleotídeo) foram utilizadas na morfometria do índice apoptótico. Amostras de mRNA foram testadas para a expressão dos genes VP2 e GAPDH utilizando-se transcrição reversa e RT-PCR. Utilizou-se um kit SYBR GREEN PCR, e a reação foi desenvolvida em ABI Prism 7000 SDS. Os índices apoptóticos cresceram progressivamente indicando uma relação na atrofia bursal causada pelo VDIB. Paralelamente, os resultados da PCR em tempo real demonstraram queda da carga viral nas células linfóides da bursa nos diferentes intervalos de tempo do experimento. Esses resultados sugerem um papel protetor da apoptose na diminuição da replicação viral.(AU)

Twenty-nine SPF 1-day-old chicks were inoculated with infectious bursal disease virus (IBDV) to evaluate early apoptosis and the expression of viral protein 2 (VP2) and glyceraldehyde-3-phosphate dehydrogenease (GAPDH). Five groups were formed: G1-control -and G2 to G5, - 24, 48, 72 and 96 hours post inoculation, respectively. Half of each BF was fixed and processed by routine techniques. To quantify apoptosis, 5µm-thick sections were stained with HE and submitted to TUNEL (terminal transferase UDP nick end labeling) technique. mRNA was extracted from pooled samples of 3 animals/group and used for the expression of VP2 and GADPH genes using the reverse transcription and real-time polymerase chain reaction (RT-PCR). A SYBR GREEN PCR kit was used and the reaction was carried out in an ABI Prism 7000 SDS. Apoptotic indexes progressively increased indicating a role of IBDV in inducing hypotrophy of the BF. Also, it was showed that as long as apoptosis increased, viral protein expression decreased, which suggests that apoptosis plays a role as a defense mechanism against viral replication.(AU)

Influence of sex, age, and fasting on blood parameters and body, bursa, spleen and yolk sac weights of broiler chicks

The effects of water and feed fasting for 24, 48 and 72 hours post-hatching on blood parameters (mean corpuscular volume, MCV; red blood-cell, RBC; hematocrit, HCT; hemoglobin, HGB; plasma glucose, CGP; plasma total protein, PP, and differential leukocytes count), and on body, liver, spleen, bursa, and yolk sac weights were analyzed. Erythrogram data were obtained with a blood cell counter. Total plasma protein and plasma glucose were determined by using the Bradford method (1976) and a glucose PAP liquiform kit (Labtest, cat. n. 84), respectively. Specific leukocyte counts were carried out on blood smears stained with Rosenfeld solution. According to the obtained data, water and feed post-hatching fasting reduced MCV values, which also were lower in males than that in females. Fasting for 48 hours promoted an increase in PP, while fasting for 72 hours reduced HCT. Chicks submitted to fasting presented lower body weights as compared to fed chicks, but their liver weight did not increase between 48 and 72 hours of age. Fasting decreased spleen weight, but bursa and yolk sac weight were not affected. Data showed that female and male chicks react in a similar way to post-hatching fasting, which affects body weight, liver and spleen weight, and HCT and PP values. Moreover, 72 hours of fasting affected more intensely HCT and MCV values.

Evaluation of anatomopathological, serological, immunological responses and protection in broilers vaccinated with live infectious bursal disease vaccines

ABSTRACT The study was divided into three experiments. In the first one, broilers were distributed into six groups and vaccinated against infectious bursal disease at 14 days of age: T1-not vaccinated, T2-Lukert1 (intermediate), T3-Lukert2 (intermediate plus), T4-228E, T5-V877 and T6-Winterfield 2512 (hot strains). Bursas of Fabricius (BF) were collected at 17, 21, 28 and 35 days to measure BF relative weight (BFRW), diameter, histological examination and image processing analysis (IPA). At one, 14, 21, 28 and 35 days of age, samples of blood taken from eight birds from each group for serology analysis by ELISA test. Hot strains vaccines induced reduction of BFRW and BF diameter, higher histological score lesion degree, more lymphocyte depletion on the BF follicles and higher IBD antibody titer. In the second experiment, 16 birds from groups T1 to T6 were isolated and challenged with a very virulent strain of IBDV (vvIBDV) at 25 days of age. Only groups T4, T5 and T6 were totally protected against vvIBDV challenge. In the third experiment, the immunosuppressive potential of each vaccine was determined by examining the ability of IBDVvaccinated birds to respond to Newcastle disease (ND) vaccination and challenge. None of the vaccines was found to be immunodepressive.

RESUMO O estudo foi dividido em três experimentos. No primeiro, frangos foram distribuídos em seis grupos e vacinados contra a Doença Infecciosa da Bursa (IBD) aos 14 dias de idade: T1-não vacinados, T2-Lukert1 (intermediária), T3-Lukert2 (intermediária plus), T4-228E, T5-V877 e T6-Winterfield 2512 (cepa forte). As bursas de Fabrícius (BF) foram colhidas aos 17, 21, 28 e 35 dias para mensurar o seus pesos relativos da BF (BFRW), diâmetros, exames histológicos e processamento de imagem imagens(IPA). Ao primeiro, 14, 21, 28 e 35 dias de idade, foram colhidas amostras de sangue de oito aves de cada grupo para a realização de sorologia através do método ELISA indireto. As vacinas cepa forte induziram redução do BFRW peso e do diâmetro da BF, maior grau de lesão histológica e depleção linfocitária e títulos mais elevados de anticorpos. No segundo experimento, 16 aves dos grupos T1 ao T6 foram isoladas e desafiadas com o vírus muito virulento da IBD (vvIBDV) aos 25 dias de idade. Somente os grupos T4, T5 e T6 foram completamente protegidos do desafio com vvIBDV. No terceiro experimento, o potencial imunossupressor de cada vacina foi determinado através da capacidade das aves vacinadas responderem à vacinação contra a Doença de Newcastle (ND) e posterior desafio. Nenhuma vacina apresentou-se imunossupressora.