Epitope mapping from Mycobacterium leprae proteins: Convergent data from in silico and in vitro approaches for serodiagnosis of leprosy.

Knowledge of immunodominant B-cell epitopes is essential to design powerful diagnostic strategies aiming for antibody detection. Outstanding progress in computational prediction has achieved a significant contribution to the biomedical fields, including immunodiagnosis. In silico analysis may have an even more important role when information concerning antigens from etiologic agents of neglected diseases, such as leprosy, is scarce. The aim of this study was to provide mapping of B-cell epitopes from two Mycobacterium leprae-derived antigens (Ag85B and ML2055), confirm their antigenicity, and to assess the ability of in silico immunoinformatics tools to accurately predict them. Linear B-cell epitopes predicted by ABCpred and SVMTrip servers were compared to antigenic regions of synthetic overlapping peptides that exhibited reactivity to antibodies from patients with leprosy. Our in vitro results identified several immunodominant regions that had also been indicated by in silico prediction, providing agreement between experimental and simulated data. After chemical synthesis, we used enzyme-linked immunosorbent assays to determine the effectiveness of the first identified sequence (GTNVPAEFLENFVHG) which had 72 % sensitivity and 78 % specificity (AUC = 0.79) while the second one (PVSSEAQPGDPNAPS) had 72 % sensitivity and 93.8 % specificity (AUC = 0.85). Using dot blotting, an easy-to-read visual test, both peptides could distinguish sera from patients with leprosy from those with tuberculosis and from sera of healthy volunteers. Our findings suggest that these synthetic peptides, with some refinement, may be useful as serological diagnostic antigens for leprosy. In addition, it was displayed that immunoinformatics provides reliable information for mapping potential B-cell epitopes for development of peptide-based diagnostic assays for neglected diseases.

Análise dialélica e uso de marcadores microssatélites na avaliação de cultivares de trigo

The combining ability of eight wheat varieties was evaluated according to the fourth model of the Griffing's diallelic methodology. Studies on genetic dissimilarity based on microsatellite markers and genetic distance among genotypes from pedigree data were also performed. The experiment was carried out in a randomized block design with two replications. Eight traits were evaluated in the diallel. Genotypes were grouped according to the UPGMA and Tocher methods. Genetic variability among genotypes was evident. Varieties 'CD 108', 'CD 0542' and 'CD 104' were those who showed high values for general combining ability in several traits. Since the effects of specific combining ability were more important in those particularly heterozygous combinations obtained from varieties allocated in different clusters, field and molecular results coincided in a certain way. There was no a good coincidence between the dendrogram based on parentage coefficient and the one based on microssatellite markers. The very small association between standards of grouping was probably related to the intrinsic properties of each way of estimating the genetic distance, which can modify the interpretation and the distribution of the genetic variability in the evaluated genotypes.

Foi avaliada a capacidade combinatória de oito cultivares de trigo, por meio de um esquema dialélico analisado segundo o modelo quatro da metodologia de Griffing. Paralelamente, foi realizada uma análise de dissimilaridade com marcadores SSR, a partir de estimativas de distância genética baseadas em pedigree. Oito características foram avaliadas num experimento delineado em blocos ao acaso, com duas repetições. O agrupamento das cultivares a partir das distâncias genéticas foi efetuado com os métodos UPGMA e Tocher. Ficou evidenciada a variabilidade entre os genótipos de trigo. As cultivares 'CD 108', 'CD 0542' e 'CD 104' apresentaram grande capacidade geral de combinação para vários caracteres. Os maiores valores de capacidade específica de combinação foram detectados nos híbridos mais heterozigotos, formados pelo cruzamento de parentais integrantes de grupos diferentes. Os agrupamentos indicados pelo pedigree não coincidiram com os indicados a partir dos marcadores moleculares. As distâncias dos marcadores SSR provavelmente refletem melhor as relações entre as cultivares de trigo do que as distâncias medidas com base na genealogia. A falta de associação entre os padrões de agrupamento foi provavelmente devida às propriedades intrínsecas de cada forma de estimação das distâncias genéticas, as quais podem modificar a interpretação e a distribuição da variabilidade genética entre os genótipos avaliados.

Análise dialélica e uso de marcadores microssatélites na avaliação de cultivares de trigo

The combining ability of eight wheat varieties was evaluated according to the fourth model of the Griffing's diallelic methodology. Studies on genetic dissimilarity based on microsatellite markers and genetic distance among genotypes from pedigree data were also performed. The experiment was carried out in a randomized block design with two replications. Eight traits were evaluated in the diallel. Genotypes were grouped according to the UPGMA and Tocher methods. Genetic variability among genotypes was evident. Varieties 'CD 108', 'CD 0542' and 'CD 104' were those who showed high values for general combining ability in several traits. Since the effects of specific combining ability were more important in those particularly heterozygous combinations obtained from varieties allocated in different clusters, field and molecular results coincided in a certain way. There was no a good coincidence between the dendrogram based on parentage coefficient and the one based on microssatellite markers. The very small association between standards of grouping was probably related to the intrinsic properties of each way of estimating the genetic distance, which can modify the interpretation and the distribution of the genetic variability in the evaluated genotypes.

Foi avaliada a capacidade combinatória de oito cultivares de trigo, por meio de um esquema dialélico analisado segundo o modelo quatro da metodologia de Griffing. Paralelamente, foi realizada uma análise de dissimilaridade com marcadores SSR, a partir de estimativas de distância genética baseadas em pedigree. Oito características foram avaliadas num experimento delineado em blocos ao acaso, com duas repetições. O agrupamento das cultivares a partir das distâncias genéticas foi efetuado com os métodos UPGMA e Tocher. Ficou evidenciada a variabilidade entre os genótipos de trigo. As cultivares 'CD 108', 'CD 0542' e 'CD 104' apresentaram grande capacidade geral de combinação para vários caracteres. Os maiores valores de capacidade específica de combinação foram detectados nos híbridos mais heterozigotos, formados pelo cruzamento de parentais integrantes de grupos diferentes. Os agrupamentos indicados pelo pedigree não coincidiram com os indicados a partir dos marcadores moleculares. As distâncias dos marcadores SSR provavelmente refletem melhor as relações entre as cultivares de trigo do que as distâncias medidas com base na genealogia. A falta de associação entre os padrões de agrupamento foi provavelmente devida às propriedades intrínsecas de cada forma de estimação das distâncias genéticas, as quais podem modificar a interpretação e a distribuição da variabilidade genética entre os genótipos avaliados.