Estimation of repeatability and genotypic superiority of elephant grass half-sib families for energy purposes using mixed models

The mixed-model methodology is an alternative to select genotypes for traits highly influenced by the environment. In addition, this method allows FOR estimating the repeatability coefficient and predicting the number of assessments needed for a selection process to increase reliability. This study aimed to determine the minimum number of evaluations necessary for a reliable selection process and to estimate the variance components used for predicting genetic gains between and within half-sib families of elephant grass ( Cenchrus purpureus (Schumach.) Morrone ) using the mixed-model methodology. Half-sib families were generated using genotypes from the Active Germplasm Bank of Elephant Grass. The experiment was performed in a randomized block design with nine half-sib families, three replicates, and eight plants per plot. We evaluated 216 genotypes (individual plants) of elephant grass. The deviance analysis was carried out, genetic parameters were estimated, gains between and within families were predicted, and repeatability coefficients were obtained using Selegen software. There was genetic variability for selection within the families evaluated. The reliability values found above 60 % for plant height and number of tillers and above 80 % for dry matter yield suggest that only two evaluations are required to select superior genotypes with outstanding reliability. Sixteen genotypes were identified and selected for their productive potential, which can be used as parents in elephant grass breeding programs for bioenergy production.(AU)

Repeatability estimates in longitudinal data on guava trees

The use of longitudinal measurements is an essential practice both in Psidium guajava L. breeding and in other perennial crops in which covariance structures can be introduced to explain the form of dependence between measurements. Hence, this study aimed to analyze six covariance structures to identify one that best described the correlation between the repeated measurements in time in traits of guava full-sib families. The repeatability coefficient for each trait was estimated and the minimum number of evaluations required for estimates representing the population was determined. The work was performed based on average data of three yield-related variables from nine harvests of a guava tree population evaluated from 2011 to 2018. The best model was chosen based on the Akaike and Schwarz Bayesian information criterion. The autoregressive covariance structure best represented the dependencies among families between crops for all traits. The number of variables of fruits and total yield per plant presented repeatability estimates higher than 0.5 and may be essential traits for indirect selection of others, such as fruit mass, which had an estimated repeatability of 0.24, proving low regularity in the repetition of the character from one cycle to another. It was also possible to define four harvests as the minimum acceptable number of observations necessary on the same individual for these traits; therefore, the repetitions represented the individuals.(AU)

An RT-rtPCR assay for detection of rabies virus in bovine specimens / Um ensaio RT-rtPCR para detecção de vírus da raiva em espécimes bovinos

ABSTRACT: Bovine rabies is endemic in most Brazilian States, including Rio Grande do Sul (RS), which has faced an unprecedented rabies outbreak between 2011 and 2018. We described a real-time reverse transcription quantitative PCR (RT-rtPCR) for detection of rabies virus (RABV) in bovine samples. The primers were designed targeting a highly conserved region of the nucleoprotein (N) gene of RABV obtained from cattle. The detection limit corresponded to 13 DNA copies and the intra- and inter-run repeatability was adequate (CV<9%) in all dilutions tested. Amplification of other pathogens associated with neurological disease in cattle or cross-contamination was not observed. Brain samples from cattle suspicious of rabies (n=21) were tested in triplicate by the RT-rtPCR and by the gold-standard direct fluorescent antibody test (DFAT), resulting in 100% of sensitivity and specificity of the RT-rtPCR. Testing of additional 41 bovine brain samples submitted to the routine DFAT testing yielded 37 (90.2%) concordant results (30 positive/7 negative) and 4 (9.7%) inconclusive in DFAT and RT-rtPCR positive. These results showed a good concordance between the tests and a higher sensitivity of the RT-rtPCR. This assay represents an alternative for RABV detection, either as a confirmatory test or for large-scale diagnosis in endemic regions.

RESUMO: A raiva bovina é endêmica na maioria dos estados brasileiros, inclusive no Rio Grande do Sul (RS), que enfrentou um surto de raiva sem precedentes entre 2011 e 2018. Descrevemos um PCR quantitativo de transcrição reversa em tempo real (RT-rtPCR) para detecção do vírus da raiva (RABV) em bovinos. Os primers foram desenhados visando uma região altamente conservada do gene da nucleoproteína (N) de RABV obtido de bovinos. O limite de detecção correspondeu a 13 cópias de DNA e a repetibilidade intra e inter-ensaios foi adequada (CV <9%) em todas as diluições testadas. Não foi observada amplificação de outros patógenos associados a doenças neurológicas em bovinos ou contaminação cruzada. Amostras de cérebro de bovinos com suspeita de raiva (n = 21) foram testadas em triplicata no RT-rtPCR e pelo teste de anticorpo fluorescente padrão ouro (DFAT), resultando em 100% de sensibilidade e especificidade do RT-rtPCR. O teste de 41 amostras de cérebro bovino adicionais submetidas ao teste de DFAT de rotina rendeu 37 (90,2%) resultados concordantes (30 positivos / sete negativos) e quatro (9,7%) inconclusivos em DFAT e RT-rtPCR positivo. Esses resultados mostraram boa concordância entre os testes e maior sensibilidade do RT-rtPCR. Este ensaio representa uma alternativa para a detecção do vírus da raiva, seja como teste confirmatório ou para diagnóstico em larga escala em regiões endêmicas.

An RT-rtPCR assay for detection of rabies virus in bovine specimens / Um ensaio RT-rtPCR para detecção de vírus da raiva em espécimes bovinos

Bovine rabies is endemic in most Brazilian States, including Rio Grande do Sul (RS), which has faced an unprecedented rabies outbreak between 2011 and 2018. We described a real-time reverse transcription quantitative PCR (RT-rtPCR) for detection of rabies virus (RABV) in bovine samples. The primers were designed targeting a highly conserved region of the nucleoprotein (N) gene of RABV obtained from cattle. The detection limit corresponded to 13 DNA copies and the intra- and inter-run repeatability was adequate (CV<9%) in all dilutions tested. Amplification of other pathogens associated with neurological disease in cattle or cross-contamination was not observed. Brain samples from cattle suspicious of rabies (n=21) were tested in triplicate by the RT-rtPCR and by the gold-standard direct fluorescent antibody test (DFAT), resulting in 100% of sensitivity and specificity of the RT-rtPCR. Testing of additional 41 bovine brain samples submitted to the routine DFAT testing yielded 37 (90.2%) concordant results (30 positive/7 negative) and 4 (9.7%) inconclusive in DFAT and RT-rtPCR positive. These results showed a good concordance between the tests and a higher sensitivity of the RT-rtPCR. This assay represents an alternative for RABV detection, either as a confirmatory test or for large-scale diagnosis in endemic regions.

A raiva bovina é endêmica na maioria dos estados brasileiros, inclusive no Rio Grande do Sul (RS), que enfrentou um surto de raiva sem precedentes entre 2011 e 2018. Descrevemos um PCR quantitativo de transcrição reversa em tempo real (RT-rtPCR) para detecção do vírus da raiva (RABV) em bovinos. Os primers foram desenhados visando uma região altamente conservada do gene da nucleoproteína (N) de RABV obtido de bovinos. O limite de detecção correspondeu a 13 cópias de DNA e a repetibilidade intra e inter-ensaios foi adequada (CV <9%) em todas as diluições testadas. Não foi observada amplificação de outros patógenos associados a doenças neurológicas em bovinos ou contaminação cruzada. Amostras de cérebro de bovinos com suspeita de raiva (n = 21) foram testadas em triplicata no RT-rtPCR e pelo teste de anticorpo fluorescente padrão ouro (DFAT), resultando em 100% de sensibilidade e especificidade do RT-rtPCR. O teste de 41 amostras de cérebro bovino adicionais submetidas ao teste de DFAT de rotina rendeu 37 (90,2%) resultados concordantes (30 positivos / sete negativos) e quatro (9,7%) inconclusivos em DFAT e RT-rtPCR positivo. Esses resultados mostraram boa concordância entre os testes e maior sensibilidade do RT-rtPCR. Este ensaio representa uma alternativa para a detecção do vírus da raiva, seja como teste confirmatório ou para diagnóstico em larga escala em regiões endêmicas.

Genetic evaluation of oocyte and embryo production in dairy Gir cattle using repeatability and random regression models

The objective of this work is to estimate genetic parameters and breeding values to improve embryo and oocyte production, using repeatability and random regression models (RRM) for Gir dairy cattle. We used 11,398 records of ovum pick-up from 1,747 dairy Gir donors and evaluated sixteen different models: the traditional repeatability model and fifteen RRM, each of which considered a different combination of Legendre polynomial regressors to describe the additive genetic and permanent environment effects. The 4G1P model (four regressors for the genetic effect and one regressor for the permanent environment effect) is the most suitable model to analyze the number of viable and total oocytes, while the 3G1P is the best model to analyze the number of cleaved and viable embryos, according to the values of the Akaike information criterion (AIC) and the Bayesian information criterion (BIC). The heritability estimated with the RRM was higher than that estimated with the repeatability model. The high repeatability reported for oocyte and embryo count traits indicates that donors, which had high oocyte and embryo counts in the first ovum pick-up, should maintain this result in the next ovum pick-up. Genetic correlations between adjacent ages were high and positive, while genetic correlations between extreme ages were weak. We observed a reranking of the top sires and females (heifers and cows) over the period evaluated. The reliability of the estimated breeding values by RRM showed changes across age, and the expected genetic gains by RRM are larger. This shows that RRM is most suitable alternative for the evaluation and selection of oocyte and embryo count traits.(AU)

Estimation of genetic parameters in dairy production in girolando cattle / Estimação de parâmetros genéticos na produção leiteira em bovinos girolando

Milk production is an important economic activity in Brazil. Dairy farmers would benefit from animal breeding programs that aid in identification and selection of animals with the best cost/benefit ratio to maximize productivity, and additionally provide advice on disposal of less productive animals. This study aims to estimate the heritability and repeatability of milk production corrected for 305 days (PL305) in a herd of Girolando cattle. We analyzed 528 lactations in 251 cows. For the analysis, uniform a priori distribution was defined for systematic effects. Gaussian and inverted Wishart distributions were defined as a priori distributions for random effects. The variance components were estimated based on Bayesian inference using the MCMCglmm function available in the MCMCglmm package of the R software. Convergence was verifed with the Geweke test available in the R software. The heritability and repeatability were estimated from the variance component results. Heritability was at 0.28, suggesting that selection for the milk production trait leads to efficient genetic progress in the herd. Phenotypic variance was mainly due to environmental variance; therefore, the phenotype of individuals should not be considered as indicator for additive genetic variance. Repeatability was at 0.93, indicating that the first performance of the animals based on milk production average is a good indicator of the second, and the data could be used for disposal decisions.(AU)

A produção de leite é uma das atividades econômicas mais importantes da agropecuária brasileira. Produtores podem usufruir de programas de melhoramento genético que permitem a identificação dos melhores animais e sua seleção para maximizar a produtividade com a melhor relação custo/benefício, além do aconselhamento do descarte de animais menos produtivos. Objetivou-se estimar a herdabilidade e repetibilidade da produção de leite corrigida para 305 dias (PL305) de um rebanho de bovinos da raça Girolando. Foram analisadas 528 lactações de 251 vacas. Para análise foi definida a distribuição uniforme a priori para efeitos sistemáticos. As distribuições de Wishart gaussiana e invertida foram definidas como distribuições a priori para efeitos aleatórios. Os componentes de variância foram estimados utilizando inferência bayesiana pela função MCMCglmm disponível no pacote MCMCglmm do software R. A convergência foi verificada pelo teste de Geweke disponível no software R. Após a obtenção dos componentes de variância foram estimados a herdabilidade e repetibilidade. A herdabilidade observada foi 0,28, o que sugere que a seleção para esta característica resultará em progresso genético eficiente no rebanho. A maior parte da variância fenotípica é devido a variância ambiental, com isso, o fenótipo dos indivíduos não é um bom indicador da variância genética aditiva. A repetibilidade foi de 0,93, indicando que o primeiro desempenho dos animais é considerado um bom indicador do segundo, podendo ser utilizadas em decisões de descarte.(AU)

Validation of a one-tube nested real-time PCR assay for the detection of Cryptosporidium spp. in avian fecal samples / Validação da nested PCR em tempo real em um tubo para detecção de Cryptosporidium spp. em amostras fecais de aves

Abstract The aim of this study was to validate a one-tube nested real-time PCR assay followed by genetic sequencing to detect and identify Cryptosporidium species and genotypes in birds. A total of 443 genomic DNA extracted from avian fecal samples were analyzed by one-tube nested real-time PCR and conventional nested PCR. By one-tube nested real-time PCR, 90/443 (20.3%) samples were positive for Cryptosporidium spp. In contrast, 36/443 (8.1%) samples were positive for Cryptosporidium spp. by conventional nested PCR. The analytical sensitivity test showed that one-tube nested real-time PCR detects approximately 0.5 oocyst (2 sporozoites) per reaction. An evaluation of analytical specificity did not reveal amplification of microorganisms that commonly present nonspecific amplification with primers used for the diagnosis of Cryptosporidium spp. The repeatability analysis showed the same result in 27 out of 30 samples (90%). As for the reproducibility of one-tube nested real-time PCR, 24 of the 30 samples examined (80%) showed the same result. All the 90 samples amplified by one-tube real-time nested PCR were successfully sequenced, leading to the identification of C. baileyi, C. galli, C. meleagridis, C. proventriculi, and Cryptosporidium avian genotype I. Genetic sequencing of conventional nested PCR amplicons was successful in 10/36 (27.8%) of positive samples.

Resumo O objetivo deste trabalho foi validar um protocolo de nested PCR em tempo real em um tubo (nPCR-TR-1T) seguida de sequenciamento genético para detectar e caracterizar as espécies e genótipos de Cryptosporidium em aves. Um total de 443 amostras de DNA genômico, extraído de amostras fecais de aves, foi analisado pela nPCR-TR-1T e pela nested PCR convencional. Pela nPCR-TR-1T, foi observada positividade para Cryptosporidium spp. de 20,3% (90/443), em contraste com a nested PCR convencional, que apresentou positividade de 8,1% (36/443). O teste de sensibilidade analítica mostrou que a nPCR-TR-1T detecta aproximadamente 0,5 oocisto (2 esporozoítos) por reação. A avaliação da especificidade analítica não revelou amplificação de microrganismos que comumente apresentam amplificação inespecífica com primers utilizados para o diagnóstico de Cryptosporidium spp. O cálculo da repetibilidade evidenciou o mesmo resultado em 27 de 30 amostras (90%). Em relação à reprodutibilidade da nPCR-TR-1T, foi observado o mesmo resultado em 80% (24/30) das amostras examinadas. Foi possível realizar o sequenciamento em todas as 90 amostras amplificadas pela nPCR-TR-1T, com identificação de C. baileyi, C. galli, C. meleagridis, C. proventriculi e Cryptosporidium genótipo I em aves. O sequenciamento dos fragmentos amplificados pela nested PCR convencional foi possível em 10/36 (27,8%) das amostras positivas.

Diagrammatic scales for the estimation of black node disease severity in common bean / Escalas diagramáticas para a estimativa da gravidade da doença do nó negro no feijão comum

The objective of this study was to design two diagrammatic scales for the evaluation of black node disease (Stagonosporopsis hortensis and Boeremia spp.) in common bean (Phaseolus vulgaris L.). The developed scales corresponded to logarithmic intervals (SADL) of seven levels and equal intervals (SADE) of nine levels. These scales were compared to the scale developed by the International Center for Tropical Agriculture (CIAT), which corresponds to a descriptive scale (NoSAD). In the scale validation the accuracy, precision, and concordance of repeatability were determined using Lins concordance correlation coefficient, whereas inter-rater reproducibility was determined by the overall concordance correlation coefficient (OCCC). It was observed that with the use of the proposed scales, reproducibility and repeatability were significantly improved for inexperienced raters, and reproducibility was improved in the case of experienced raters. Thus, the designed standard area diagrams with equal and logarithmic intervals are a useful tool for estimating severity under field and experimental conditions as part of the study of this patho-system.(AU)

O objetivo deste estudo foi delinear duas escalas diagramáticas para avaliação da doença do nó preto (Stagonosporopsis hortensis e Boeremia spp.) em feijoeiro (Phaseolus vulgaris L.). As escalas desenvolvidas corresponderam a intervalos logarítmicos (SADL) de sete níveis e intervalos iguais (SADE) de nove níveis. Essas escalas foram comparadas com a escala desenvolvida pelo Centro Internacional de Agricultura Tropical (CIAT), que corresponde a uma escala descritiva (NoSAD). Na validação da escala, a exatidão, precisão e concordância da repetibilidade foram determinadas usando o coeficiente de correlação de concordância de Lin, enquanto a reprodutibilidade entre avaliadores foi determinada pelo coeficiente de correlação de concordância geral (OCCC). Observou-se que, com o uso das escalas propostas, a reprodutibilidade e a repetibilidade foram significativamente melhoradas para os avaliadores inexperientes, e a reprodutibilidade foi melhorada no caso dos avaliadores experientes. Assim, as escalas diagramáticas projetadas são uma ferramenta útil para estimar a severidade em condições experimentais e de campo como parte do estudo desse sistema patológico.(AU)

Standardization of blood meal production process at a rendering plant using design of experiments / Padronização do processo de produção de farinha de sangue em uma usina de processamento por meio do planejamento de experimentos

This research focuses on the standardization of the blood meal production process at a Colombian rendering plant through a design of experiments. Initially, 108 samples of blood meal were taken where only 23% achieved the moisture target (7.5% to 8.5%). Therefore, an analysis of the measurement system was performed using a repeatability and reproducibility (R&R) study. Results showed that 39.96% of the observed variability was caused by the measurement system that was out of control. So, it was necessary to improve the method of sampling reducing the participation of the measurement system in the variability of the process to only 3.79%. Later, several experiments were accomplished with a 2k factorial design. Each experiment consisted of a response variable (blood meal moisture), two controllable factors (drying chamber temperature and percentage of rotation of the metering screw), and an uncontrollable factor (initial blood meal moisture). Finally, experiments were carried out and validated observing that, with a drying chamber temperature of 160 °C and a percentage of screw rotation of 29%, more than 97% of the blood meal was according to the moisture target. In conclusion, is confirmed that the design of experiments is a tool that allows a clear path towards optimization and standardization of processes.(AU)

Esta pesquisa se concentra na padronização do processo de produção de farinha de sangue de uma usina de beneficiamento colombiana por meio de um planejamento de experimentos. Em primeiro lugar, foi realizado um estudo de 108 amostras de farinha de sangue onde apenas 23% atingiram a meta de umidade (7.5% a 8.5%). Portanto, uma análise do sistema de medição foi realizada usando um estudo de repetibilidade e reprodutibilidade (R&R). Os resultados mostraram que 39.96% da variabilidade observada foi causada pelo sistema. Assim, foi necessário aprimorar o método de amostragem, reduzindo a participação do sistema na variabilidade para apenas 3.79%. Posteriormente, vários experimentos foram realizados com um planejamento fatorial 2k. Cada experimento consistiu em uma variável de resposta (umidade da farinha de sangue), dois fatores controláveis (temperatura da câmara de secagem e porcentagem de rotação da rosca dosadora) e um fator incontrolável (umidade inicial da farinha de sangue). Finalmente, experimentos foram realizados e validados observando que, com uma temperatura da câmara de secagem de 160 ° C e um percentual de rotação da rosca de 29%, mais de 97% da farinha de sangue estava de acordo com a umidade desejada. Em conclusão, confirma-se que o planejamento de experimentos é uma ferramenta que permite um caminho claro para a otimização e padronização de processos.(AU)

Validation of Alternative Methods of Detection of Salmonella Spp. in Experimentally Contaminated Poultry Environmental Samples

The analysis of Salmonella in the feces and the birds environment is a way of monitoring the colonization in the flocks and verifying the need for the introduction of stricter controls, in such a way that the results of the tests should be known before being sent for slaughter. The polymerase chain reaction (PCR), as well as other rapid methods represent alternatives increasingly used to detect enteric pathogens, but they need proof of effectiveness for their wide use. The aim of this study was to evaluate the equivalence between the results obtained by the methods: real-time PCR (BAX® System), Modified Rappaport-Vassiliadis Semi-solid Medium (MSRV) (ISO 6579) and the traditional method of official reference in Brazil for research of S. Typhimurium and S. Enteritidis in poultry samples. Two hundred and fifty-two samples of disposable shoe covers (DSC) and 252 samples of feces were infected with an average of 2 to 3 log CFU/g of each serovar, and the same samples without fortification were evaluated by the three methods. Five hundred and four diagnoses were obtained with satisfactory results in terms of repeatability (greater than 80%), reproducibility (mean 83,1%), sensitivity (81% to 100%), specificity (95% to 100%), and accuracy (90% to 100%). The compliance test verified that there was not a significant difference between the alternative and the official methods, allowing us to state that the methodologies have had equivalent performances.(AU)