Determining the best sectioning method and intestinal segment for morphometric analysis in broilers

Brazilian poultry production is very efficient and demands maximum broiler performance. Therefore, digestive system pathologies have a relevant role. Considering it is difficult to obtain consistent information on intestinal morphometric analysis, this study aimed at establishing essential and clear criteria for the collection of intestinal segments for morphometric analysis. Fifteen 13-d-old broilers were sacrificed and three intestinal segments were collected per bird. Two 3-cm long sections were obtained from each of the intestinal segments. Samples were collected open or closed. The closed samples were transversely, hemicylindrically, or longitudinally sectioned. Samples were processed and stained with hematoxylin and eosin. The number of microscopic fields in each section was counted. Villi presenting the base clearly embedded in the submucosa, no damage or folds, and simple columnar epithelium at the tip were considered adequate for measurements. These villi were counted in each sample. The results shows that hemicylindrical sections presented the highest number of observation fields, with an average of 9.76 fields. Jejunum samples were among the three highest average villi counts, with 18.23 in longitudinal sections and 15.61 in hemicylindrical sections. The results of the present study indicate that hemicylindrical sectioning and jejunal samples were, respectively, the best sectioning method and the best intestinal segment for the morphometric analysis of the intestines of broilers

Lesões Ulcerativas em Frangos de Corte Abatidos no RGS: Estudo Sobre a Cama dos Aviários: Material Utilizado e Número de Reutilizações

Os queratoacantomas (QA) e as dermatites granulomatosas por corpo estranho (DGCE) causam lesões ulcerativas cutâneas em frangos. O diagnóstico diferencial depende de exame histopatológico. 363 amostras de pele com a lesão foram colhidas e examinadas microscopicamente. O número de casos de DGCE foi superior ao de QA.  Lesões únicas sugerem DGCE, enquanto que as múltiplas, casos de QA. Camas de maravalha ou mistas com casca de arroz produziram mais casos de DGCE, enquanto que camas com serragem ou serragem com maravalha produziram mais QA. Ambas as doenças ocorreram tanto em aves criadas em camas novas como em reutilizadas. Há necessidade de estudos envolvendo um número maior de amostras e dados de manejo

Classification of Avian Pathogenic Escherichia coli by a Novel Pathogenicity Index Based on an Animal Model

Background: Avian Pathogenic Escherichia coli is the main agent of colibacillosis, a systemic disease that causes considerable economic losses to the poultry industry. In vivo experiments are used to measure the ability of E. coli to be pathogenic. Generally, these experiments have proposed different criteria for results interpretation and did not take into account the death time. The aim of this study was to propose a new methodology for the classification of E. coli pathogenicity by the establishment of a pathogenicity index based in the lethality, death time and the ability of the strain to cause colibacillosis lesions in challenged animals.Materials, Methods & Results: A total of 293 isolates of E. coli were randomly selected to this study. The strains were isolated from cellulitis lesions, broiler bedding material or respiratory diseases and were previously confirmed through biochemical profile. The bacterial isolates were kept frozen at -20°C. The strains were retrieved from stocks and cultured in brain-heart infusion broth overnight at 37°C to obtain a final concentration of 109 UFC/mL. A total of 2940 one-dayold chicks from commercial breeding hens were randomly assigned to groups containing 10 animals and each group was subcutaneously inoculated in the abdominal region with 0.1 mL of the standard inoculum solution containing each of the strains. A control group of 10 broilers were inoculated with 0.1 mL of brain-heart infusion broth by the same route. The chicks were kept for seven days. They were observed at intervals of 6, 12 and 24 h post-inoculation during the first days. From the second day on, the chicks were observed at intervals of 12 h. According to the death time and to the scores of each lesion (aerosaculitis, pericarditis, perihepatitis, peritonitis and cellulitis), a formula to determine the Individual Pathogenicity Index was established. A value of 10 was established as the maximum pathogenicity rate for an inoculated bird. From this rate, 5 points corresponded to scores for gross lesions present at necropsy. For each lesion present, it represents 1 point. The remaining 5 points corresponded to the death time. To obtain the death time value, an index of 1, corresponding to the maximum value assigned to a death on the first day, was divided by the number of days that the birds were evaluated, resulting in a value of 0.1428, which corresponded to a survival bonus factor. It was possible to classify E. coli strains into four pathogenicity groups according to the pathogenicity index: high pathogenicity (pathogenicity index ranging from 7 to 10), intermediate pathogenicity (pathogenicity index ranging from 4 to 6.99), low pathogenicity (pathogenicity index ranging from 1 to 3.99) and apathogenic (pathogenicity index ranging from 0 to 0.99). The analysis of the strains according to their origin revealed that isolates from broiler bedding material presented a lower pathogenicity index.Discussion: It is possible that the source of isolation implies in different results, depending on the criteria adopted. This data reinforces the importance of use a more accurate mathematical model to represents the biological phenomenon. In the study, all avian pathogenic Escherichia coli strains were classified based on a pathogenicity index and the concept of the death time represents an interesting parameter to measure the ability of the strain to promote acute and septicemic manifestation. The use of a support method for poultry veterinary diagnostic accompanying the fluctuation of the bacteria pathogenicity inside the farms may indicate a rational use of antimicrobial in poultry industry

Artificial neural networks on eggs production data management

Background: Eggs have acquired a greater importance as an inexpensive and high-quality protein. The Brazilian egg industry has been characterized by a constant production expansion in the last decade, increasing the number of housed animals and facilitating the spread of many diseases. In order to reduce the sanitary and financial risks, decisions regard¬ing the production and the health status of the flock must be made based on objective criteria. The use of Artificial Neural Networks (ANN) is a valuable tool to reduce the subjectivity of the analysis. In this context, the aim of this study was at validating the ANNs as viable tool to be employed in the prediction and management of commercial egg production flocks. Materials, Methods & Results: Data from 42 flocks of commercial layer hens from a poultry company were selected. The data refer to the period between 2010 and 2018 and it represents a total of 600,000 layers. Six parameters were selected as “output” data (number of dead birds per week, feed consumption, number of eggs, weekly weight, weekly egg produc¬tion and flock uniformity) and a total of 13 parameters were selected as “input” data (flock age, flock identification, total hens in the flock, weekly weight, flock uniformity, lineage, weekly mortality, absolute number of dead birds, eggs/hen, weekly egg production, feed consumption, flock location, creation phase). ANNs were elaborated by software programs NeuroShell Predictor and NeuroShell Classifier. The programs identified input variables for the assembly of the networks seeking the prediction of the variables called outgoing that are subsequently validated. This validation goes through the comparison between the predictions and the real data present in the database that was the basis for the work. Validation of each ANN is expressed by the specific statistical parameters multiple determination (R2) and Mean Squared Error (MSE). For instance, R2 above 0.70 expresses a good validation. ANN developed for the output variable “number of dead birds per week” presented R2= 0.9533 and MSE= 256.88. For “feed consumption”, the results were R2= 0.7382 and MSE= 274.56. For “number of eggs (eggs/hen)”, the results were R2= 0.9901 and MSE= 172.26. For “weekly weight”, R2= 0.9712 and MSE= 11154.41. For “weekly egg production”, R2= 0.8015 and MSE= 72.60. For “flock uniformity”, R2= -2.9955 and MSE= 431.82. Discussion: From the six ANN designed in this study, in five it was possible to validate the predictions by comparing predictions with the real data. In one output parameter (“flock uniformity”), it was not possible to have adequate validation due to insufficient data in our database. For “number of dead birds per week”, “feed consumption”, “weekly weight” and “uniformity”, the most important variable was “flock age” (27.5%, 52.5%, 55.2% and 37.9%, respectively). For “number of eggs (eggs/hen)”, “uniformity” (52.1%) was the most relevant variable for prediction. For “weekly egg production”, “flock age” and “number of eggs (eggs/hen)” were the most important zootechnical parameters, both with a relative contribution of 38.2%. The results showed that even with the use of a robust tool such as ANNs, it is necessary to have well-noted and clear information that expresses the reality of the flocks. In any case, the results presented allow us to state that ANNs are capable for the management of data generated in a commercial egg production facility. The process of evaluation of these data would be improved if ANNs were routinely used by the professionals linked to this activity

Avaliação da capacidade de formação de biofilme por cepas de Pasteurella multocida isoladas de casos de cólera aviária e de pulmões de suínos e sua relação com a patogenicidade

Pasteurella multocida is a Gram-negative bacillus that causes economic losses due to the development of respiratory diseases in several animal species. Among the mechanisms of virulence, the formation of biofilms is an important factor for bacterial survival in hostile environments. Studies of biofilm formation by P. multocida are needed because P. multocida is an important pathogen involved in respiratory infections. However, in contrast to other microorganisms, few studies of biofilm formation have examined P. multocida. Studies comparing the pathogenicity of microbial strains as a function of their biofilm production capacity are also rare. Consequently, the aim of this study was to evaluate the biofilm formation capacity of 94 P. multocida strains isolated from cases of fowl cholera and from swine lungs on polystyrene plates. The associations of the biofilm formation capacity with the pathogenicity index (PI) in vivo and with the presence of four genes (screened by PCR) of the tad locus (tadB, tadD, tadE and tadG), described as adhesion markers, were also determined. Strains from both animal origins were able to form biofilms. However, most of the specimens (52.13%) were classified as weak producers, and more than 40% of the strains of P. multocida (40.42%) did not produce biofilms There was no significant difference (p>0.05) in the degree of biofilm production between the two sources of isolation. Of the analyzed strains, 56.52% contained all four genes (tadB, tadD, tadE and tadG). The PI arithmetic mean of the strains classified as non-biofilm producers was significantly different (p0.05) with the production of biofilms and with the origin of a given strain. Finally, low virulence strains may suggest a higher biofilm formation capacity on polystyrene plates.Pasteurella multocida é um bacilo Gram negativo que ocasiona perdas econômicas, geralmente associadas a doenças respiratórias em diversas espécies animais. Entre os mecanismos de virulência existentes, a formação de biofilmes demonstra ser um importante fator para a proteção e para a sobrevivência bacteriana em ambientes hostis. Estudos relacionados à formação de biofilmes por P. multocida são necessários, uma vez que este é um importante patógeno envolvido em infecções respiratórias. Entretanto, ainda são poucos os estudos desenvolvidos nesta área, quando comparados com aqueles envolvendo outros microrganismos. Também são os raros os estudos que comparam a patogenicidade das cepas com a sua capacidade de produção de biofilme. Neste contexto, o objetivo deste estudo foi avaliar a capacidade de formação de biofilme em placas de poliestireno de 94 cepas de P. multocida isoladas de casos de cólera aviária e de pulmões de suínos, associando-se com o índice de patogenicidade (IP) in vivo e com a presença de quatro genes do locus tad (tadB, tadD, tadE e tadG), descritos como marcadores de adesão e pesquisados através de PCR. As cepas de ambas as origens foram capazes de formar biofilme. Contudo, a maioria dos exemplares (52,12%) foi classificada como fracamente produtora e mais de 40% das cepas de P. multocida (40,42%) não produziram biofilme Não foi observada diferença estatística (p>0,05) quanto ao grau de produção de biofilme entre as duas origens de isolamento. 56,52% das cepas analisadas apresentaram os quatro genes (tadB, tadD, tadE e tadG) concomitantemente. O IP médio das cepas classificadas como não produtoras de biofilme apresentou diferença estatística (p˂0,05) em relação ao IP das cepas moderadamente produtoras. Os exemplares classificados como fracamente produtores de biofilme diferiram significativamente (p˂0,05) do grupo de cepas moderadamente produtoras. Os resultados obtidos indicaram que, apesar de as cepas de P. multocida isoladas de casos de cólera aviária e do pulmão de suínos apresentarem capacidade de formar biofilme em superfícies de poliestireno, a adesão ocorreu geralmente de forma fraca. Os genes tadB, tadD, tadE e tadG, pertencentes ao locus tad, não apresentaram associação significativa com a produção de biofilme e nem com a origem de isolamento da cepa. Por fim, observou-se que as cepas de menor patogenicidade apresentaram uma maior capacidade de formação de biofilme em placas de poliestireno

Prevalence and distribution of pathogenic genes in Campylobacter jejuni isolated from poultry and human sources

Introduction: Campylobacter jejuni is one of the most common bacterial causes of human gastroenteritis. Despite its public health importance, the virulence factors and mechanisms underlying C. jejuni pathogenesis remain poorly understood and the relationships between these genes and the sources of the strains are not clear. We aimed to determine the virulence profiles of C. jejuni isolated from poultry and human cases of Campylobacteriosis. Methodology: A total of 50 strains of C. jejuni isolated from poultry and human cases of Campylobacteriosis were screened by polymerase chain reaction (PCR) for the presence of six pathogenic genes (flaA, iam, wlaN, cdtA, cdtB, cdtC). Results: A total of 40% (10/25) of the human isolates presented only one virulence marker. In contrast, 64% (16/25) of the poultry-derived strains showed four or five virulence markers. cdtA and flaA occurred more frequently in poultry-derived strains than in human strains. Ten different virulence profiles were observed among the human isolates and 11 among the poultry strains. Only four profiles were common to both sources: profiles 3 (flaA, cdtA, cdtB, and cdtC), 5 (cdtA and cdtB), 7 (flaA and cdtB), and 10 (iam, flaA, cdtA, cdtB, and cdtC). The human-derived strains had a higher Shannon diversity index (1.9396) and Simpson index (0.8367), indicating a more diversified population than found in poultry (1.7742 and 0.7333, respectively). Conclusions: We found variations in the genetic profiles of the circulating strains based on the isolation source and genes that are potentially pathogenic to humans were detected in poultry-derived strains

Artificial neural networks to predict egg-production traits in commercial laying breeder hens

In recent years, egg production has had an intense growth in Brazil, and Brazilian egg consumption per capita has significantly increased in the last decade. To reduce sanitary and financial risks, decisions regarding the production and health status of the flock must be made based on objective criteria. Our aim was to determine the main “input” variables for the prediction of egg production performance in commercial laying breeder flocks using an ANN model. The software NeuroShellClassifier and NeuroShell Predictor were used to build the ANN. A total of 26 egg-production traits were selected as input variables and eight as output variables. A database of 44,120 Excel cells was generated. For the training and validation of the models, 74.9% and 25.1% of the data were used, respectively. The accuracy of the ANN models was calculated and compared using the analysis of coefficient of multiple determination (R2), mean squared error (MSE), and an assessment of uniform scatter in the residual plots. The models for the outputs “weekly egg production,” “weekly incubated egg,”, “accumulated commercial egg,” and “viability” showed an R2 greater than 0.8. Other models yielded R2 values lower than 0.8. The ANN predicts adequately eight egg-production traits in the breeders of commercial laying hens. The method is an option for data management analysis in the egg industry, providing estimates of the relative contribution of each input variable to the outputs

Characterization of immune and enteric systems of broilers after imunosupression with dexamethasone

Bursa of Fabricius (BF) and the thymus are primary lymphoid organs of poultry and play a major role in avian immunity. Enteric system is also involved in immunity. Several pathologic conditions directly impact BF and thymus size, and also affect intestinal parameters. Besides, there are several immune system depressor agents which affect birds. The selection of glucocorticoid as inducer of immunosuppression is applied in many experiments; however there are few studies that are applied to the reality in the field. In this context, the aim of this study was to evaluate the effects of dexamethasone as an inducer of immunosuppression on lymphoid organs and microscopic structures of the jejunum. Materials, Methods & Results: One-day-old chicks were used as a control group (n = 8) and the treated group (n = 25) received intramuscular dexamethasone on 21, 23, 24 and 26 day-old. Control birds and treated birds were euthanized 8, 16, 24, 32 and 40 h after inoculation; four control birds and six treated birds were euthanized on the eighth day after the last inoculation. Thymus, BF and jejunum were collected during the necropsy. The selected organs were processed, stained with hematoxylin and eosin and photographed. The BF and thymus cuts were evaluated by three histopathologists to determine the depletion score. Ten villi of each jejunum were evaluated for width and length of villi, depth crypt, microvillus length, enterocyte length of each villus, and wall thickness. Treated birds presented a mean weight lower than control group during all the experiment. The mean weight and the relative weight of the BF and thymus of control birds were significantly higher than treated ones. The lymphocyte depletion in BF and thymus scores differed significantly between groups, being higher in the group challenged with dexamethasone. There were no significant differences between groups for depth of crypt, height of core and height of microvilli. The intestines of the control group had higher mean values for length of villi, width of villi, height of enterocyte and thickness of wall. Significant correlation between weight of thymus and weight of BF, between weight of thymus and weight of bird and between weight of BF and bird weight were found. Discussion: Easy administration, low cost and the absence of suffering during inoculation make the use of glucocorticoids more advantageous to mimic immunosupression in poultry. Treatment with dexamethasone interfered directly in the weight of the birds. Evaluation of immune response of birds can be performed by the ratio of the BF weight in relation to the weight of the bird, as observed in the present study. The relative weight of the BF of untreated animals varied at all ages between 0.21% and 0.29%. It was also observed that the relative weight of BF and thymus in immunosuppressed birds tended to increase according to the end of the treatment. Depletion scores were higher in thymus, suggesting that glucocorticoid promoted more harmful effects on this organ. Dexamethasone had negative influence on length of villus, once mean value was 13% lower than those of control group. Width of villus and height of enterocyte core of treatment group were lower than those observed in control animals. Coefficient of determination greater than 70% was observed for almost all established relationships, except for the length of villi and weight of the thymus. Administration of dexamethasone promotes significant and negative effects on the gain of weight, length of villi, width of villi and height of enterocytes. These finding are useful for future experiments with controlled immunosuppression induction, once it provides significant information of the secondary effects of glucocorticoids administration in poultry

Avian pathogenic Escherichia coli (APEC) and uropathogenic Escherichia coli (UPEC) : characterization and comparison

Introduction: Avian pathogenic E. coli (APEC) and uropathogenic E. coli (UPEC) are responsible for avian colibacillosis and human urinary tract infections, respectively. There are genetic similarities between the APEC and UPEC pathotypes, suggesting the APEC strains could be a potential reservoir of virulence and antimicrobial-resistance genes for the UPEC strains. This study aimed to characterize and compare APEC and UPEC strains regarding the phylogroup classification, pathogenicity and antimicrobial susceptibility. Methodology: A total of 238 APEC and 184 UPEC strains were selected and characterized. The strains were assayed for antimicrobial susceptibility and classified into phylogenetic groups using a multiplex-PCR protocol. In addition, the APEC strains had previously been classified according to their in vivo pathogenicity. Results: The results showed that both pathotypes had variation in their susceptibility to most of the antimicrobial agents evaluated, with few strains classified as multidrug resistant. The highest resistance rate for both pathotypes was to amoxicillin. Classifying the APEC and UPEC strains into phylogenetic groups determined that the most frequently frequencies were for groups D and B2, respectively. These results reflect the pathogenic potential of these strains, as all the UPEC strains were isolated from unhealthy patients, and most of the APEC strains were previously classified as pathogenic. Conclusions: The results indicate that distribution into phylogenetic groups provided, in part, similar classification to those of in vivo pathogenicity index, as it was possible to adequately differentiate most of the pathogenic and commensal or low-pathogenicity bacteria. However, no relationship could be found between the specific antimicrobial agents and pathogenicity or phylogenetic group for either pathotype

Characterization of Immune and Enteric Systems of Broilers after Imunosupression with Dexamethasone

Background: Bursa of Fabricius (BF) and the thymus are primary lymphoid organs of poultry and play a major role in avian immunity. Enteric system is also involved in immunity. Several pathologic conditions directly impact BF and thymus size, and also affect intestinal parameters. Besides, there are several immune system depressor agents which affect birds. The selection of glucocorticoid as inducer of immunosuppression is applied in many experiments; however there are few studies that are applied to the reality in the field. In this context, the aim of this study was to evaluate the effects of dexamethasone as an inducer of immunosuppression on lymphoid organs and microscopic structures of the jejunum.Materials, Methods & Results: One-day-old chicks were used as a control group (n = 8) and the treated group (n = 25) received intramuscular dexamethasone on 21, 23, 24 and 26 day-old. Control birds and treated birds were euthanized 8, 16, 24, 32 and 40 h after inoculation; four control birds and six treated birds were euthanized on the eighth day after the last inoculation. Thymus, BF and jejunum were collected during the necropsy. The selected organs were processed, stained with hematoxylin and eosin and photographed. The BF and thymus cuts were evaluated by three histopathologists to determine the depletion score. Ten villi of each jejunum were evaluated for width and length of villi, depth crypt, microvillus length, enterocyte length of each villus, and wall thickness. Treated birds presented a mean weight lower than control group during all the experiment. The mean weight and the relative weight of the BF and thymus of control birds were significantly higher than treated ones. The lymphocyte depletion in BF and thymus scores differed significantly between groups, being higher in the group challenged with dexamethasone. There were no significant differences between groups for depth of crypt, height of core and height of microvilli. The intestines of the control group had higher mean values for length of villi, width of villi, height of enterocyte and thickness of wall. Significant correlation between weight of thymus and weight of BF, between weight of thymus and weight of bird and between weight of BF and bird weight were found.Discussion: Easy administration, low cost and the absence of suffering during inoculation make the use of glucocorticoids more advantageous to mimic immunosupression in poultry. Treatment with dexamethasone interfered directly in the weight of the birds. Evaluation of immune response of birds can be performed by the ratio of the BF weight in relation to the weight of the bird, as observed in the present study. The relative weight of the BF of untreated animals varied at all ages between 0.21% and 0.29%. It was also observed that the relative weight of BF and thymus in immunosuppressed birds tended to increase according to the end of the treatment. Depletion scores were higher in thymus, suggesting that glucocorticoid promoted more harmful effects on this organ. Dexamethasone had negative influence on length of villus, once mean value was 13% lower than those of control group. Width of villus and height of enterocyte core of treatment group were lower than those observed in control animals. Coefficient of determination greater than 70% was observed for almost all established relationships, except for the length of villi and weight of the thymus. Administration of dexamethasone promotes significant and negative effects on the gain of weight, length of villi, width of villi and height of enterocytes. These finding are useful for future experiments with controlled immunosuppression induction, once it provides significant information of the secondary effects of glucocorticoids administration in poultry