Identification of carboxymethyl (CM)-binding proteins derived from Lolium multiflorum pollen extract and antibody reactivity in Brazilian allergic patients.

Lolium multiflorum grass is the major pollen allergen source in the southern region of Brazil, but most of its allergens remain poorly characterized. The aim of this study was to investigate antibody reactivity to L. multiflorum crude and carboxymethyl-ligand extracts in allergic patients and healthy individuals. Ion exchange carboxymethyl (CM) chromatography (CM-Sepharose) was used to isolate proteins (S2) from L. multiflorum crude extract (S1), which were assessed by SDS-PAGE. S1- and S2-specific IgE and IgG4 levels were measured by ELISA using sera from 55 atopic and 16 non-atopic subjects. Reactive polypeptide bands in S1 and S2 were detected by immunoblotting, and the most prominent bands in S2 were analyzed by mass spectrometry (MS-MS). Similar IgE and IgG4 levels were observed to both S1 (IgE median absorbance: 1.22; IgG4 median absorbance: 0.68) and S2 (IgE median absorbance: 1.26; IgG4 median absorbance: 0.85) in atopic subjects. S1 and S2 had positive correlations for IgE and IgG4 (IgE: r=0.9567; IgG4: r=0.9229; P<0.0001) levels. Homology between S1 and S2 was confirmed by IgE (84%) and IgG4 (83%) inhibition. Immunoblotting revealed that the 29-32 kDa band was recognized by 100% of atopic subjects in both S1 and S2. MS-MS analysis identified similarity profile to groups 1 and 5 grass allergens. This study revealed that carboxymethyl-ligand fraction played an important role for pollen allergy diagnosis by containing clinically relevant allergens and constituted a promising candidate for allergen-specific immunotherapy.

Identification of carboxymethyl (CM)-binding proteins derived from Lolium multiflorum pollen extract and antibody reactivity in Brazilian allergic patients

Lolium multiflorum grass is the major pollen allergen source in the southern region of Brazil, but most of its allergens remain poorly characterized. The aim of this study was to investigate antibody reactivity to L. multiflorum crude and carboxymethyl-ligand extracts in allergic patients and healthy individuals. Ion exchange carboxymethyl (CM) chromatography (CM-Sepharose) was used to isolate proteins (S2) from L. multiflorum crude extract (S1), which were assessed by SDS-PAGE. S1- and S2-specific IgE and IgG4 levels were measured by ELISA using sera from 55 atopic and 16 non-atopic subjects. Reactive polypeptide bands in S1 and S2 were detected by immunoblotting, and the most prominent bands in S2 were analyzed by mass spectrometry (MS-MS). Similar IgE and IgG4 levels were observed to both S1 (IgE median absorbance: 1.22; IgG4 median absorbance: 0.68) and S2 (IgE median absorbance: 1.26; IgG4 median absorbance: 0.85) in atopic subjects. S1 and S2 had positive correlations for IgE and IgG4 (IgE: r=0.9567; IgG4: r=0.9229; P<0.0001) levels. Homology between S1 and S2 was confirmed by IgE (84%) and IgG4 (83%) inhibition. Immunoblotting revealed that the 29-32 kDa band was recognized by 100% of atopic subjects in both S1 and S2. MS-MS analysis identified similarity profile to groups 1 and 5 grass allergens. This study revealed that carboxymethyl-ligand fraction played an important role for pollen allergy diagnosis by containing clinically relevant allergens and constituted a promising candidate for allergen-specific immunotherapy.

Mating system and intrapopulational genetic diversity of Copernicia prunifera (Arecaceae): a native palm from Brazilian semiarid.

The aim of this study was to investigate the mating system and genetic diversity of the palm Copernicia prunifera using inter-simple sequence repeat markers. We found that the C. prunifera has multiple inflorescences with hermaphroditic flowers and pollen viability of 62%. Outcrossing rates at the population level (N = 267) produced a multilocus outcrossing rate (tm) of 0.878 and a single-locus outcrossing rate (ts) of 0.738, indicating that C. prunifera has a mixed mating system that is preferentially allogamous. The rate of mating among relatives (tm - ts) was low, indicating limited outcrossing between closely related individuals. The fixation index between seed trees (F) was negative (0.200), suggesting an absence of inbreeding, while the correlation of selfing (rs) was high (0.914). The values of the diversity indices among adults and progenies did not differ statistically (HE = 0.319 and I = 0.470; HE = 0.337 and I = 0.505, respectively). In testing for the presence of genetic bottlenecks using the infinite allele model and the stepwise mutation model, we observed a reduction in the effective population, as well as a deficit in heterozygosity (P < 0.0001). The results of this study inform management strategies for the conservation and genetic improvement of the C. prunifera palm.

Anthropization as a determinant factor in the genetic structure of Copernicia prunifera (Arecaceae).

This study uses ISSR molecular markers to characterize the demographic pattern, and spatial genetic structure (SGS) at different life stages of development (cohorts) in a natural population of Copernicia prunifera in the Rio Grande do Norte State, Brazil. All individuals were sampled and georeferenced in a 0.55-ha plot. The demographic analyses showed a clustered pattern in the first-distance classes and a random or segregated pattern at higher distance classes for all cohorts. Among the three studied life stages, juveniles showed the greatest value for Nei's genetic diversity index (HE = 0.369), while the lowest was found among reproductive adults (HE = 0.341). Greater genetic differentiation was found within life stages (98.61%) than among stages (1.39%). The total population (N = 161) showed positive and significant kinship in the first-distance class (12.3 m). The juveniles showed significant kinship up to 10.5 m. Non-reproductive adults had a positive kinship in the first-distance class (11.0 m) and a random distribution of genotypes in the remaining classes, while reproductive adults showed a random spatial distribution of genotypes. Tests for genetic bottleneck showed that the number of loci with excess observed heterozygosity was greater than expected. The SGS results reflected the restricted seed dispersion of the species, and the genetic bottleneck reflected the reduction of genotypes as a result of the anthropization of C. prunifera natural environments.

Morfologia e dinâmica testicular em cutias (Dasyprocta prymnolopha) adultas / Testicular morphology and dynamic in adult agoutis (Dasyprocta prymnolopha)

Este trabalho objetivou apresentar a caracterização da morfologia do testículo de cutia (Dasyproctaprymnolopha) macho, com o intuito de colaborar com o conhecimento da morfofisiologia reprodutiva da espécie. Foram utilizados testículos de 47 animais, com idade entre um e dois anos, pesos homogêneos (2,08 ± 0,23kg), oriundos do Núcleo de Estudos e Preservação de Animais Silvestres do Centro de Ciências Agrárias da Universidade Federal do Piauí. As estruturas foram dissecadas, descritas, e fragmentos foram processados para a microscopia de luz, sendo, posteriormente avaliada a atividade gonadal. Observou-se que os testículos são órgãos elipsoides alongados, podendo ser encontrados na região inguinal ou na cavidade abdominal, não apresentando um escroto bem delimitado. Verificou-se também parênquima com característica histológica padrão para o órgão em mamíferos, com a identificação de oito tipos de associações celulares, caracterizando os estádios do ciclo do epitélio seminífero, com menor e maior frequência dos estádios 3 e 5, respectivamente.(AU)

This study meant to characterize the morphology of the testicle from (Dasyprocta prymnolopha) agouti males, in order to collaborate with the knowledge of reproductive morphophysiology of the specie. Testicles were used from 47 animals aged between 1 and 2 years, homogeneous weight (2.08±0.23kg), coming from the Centre for the Study and Conservation of Wild Animals of Agricultural Sciences Center of the Federal University of Piauí. The structures were dissected, described and fragments were processed for light microscopy, and, subsequently, gonadal activity was evaluated. Testes were observed to be elongated ellipsoidal bodies that can be found in the groin or in the abdominal cavity, not having a clearly defined scrotum. We also could see parenchymal with standard histological characteristic for the mammalian body, with the identification of eight types of cell associations, characterized epithelium Seminiferous stages of the cycle, with lower and higher frequency of stages 3 and 5, respectively.(AU)

Increased relatedness among the neighboring plants from seedling to adult stages in carnaúba wax palm.

The objective of this study was to assess the spatial genetic structure (SGS) at different life stages (cohorts) in a remnant population (N = 101) of Copernicia prunifera in the semiarid region of northeastern Brazil. Using seven inter-simple sequence repeat molecular markers, we were able to analyze 93 loci with 100% polymorphism. Seedlings had the highest level of genetic diversity (HE = 0.411, HO = 0.599), followed by juveniles (HE = 0.394, HO = 0.579) and adults (HE = 0.267, HO = 0.427). Based on analysis of molecular variance, the majority of genetic variations were observed to occur within the life stages (93.42%) rather than between the life stages (6.58%). We found a recent reduction in the population size (bottleneck) based on the number of loci with heterozygosity excess for the two models used (infinite allele = 92 and stepwise = 91). All the life stages showed significant SGS, with positive and significant kinship values. Sp values were 0.040 for seedlings, 0.093 for juveniles, 0.156 for adults, and 0.053 for the total population. We found an increase in SGS from the seedling to adult stages, indicating that the plants were from related adult progenitors. Data from this study can be used in designing effective management and conservation strategies for the species.

Naturally fragmented but not genetically isolated populations of Podocarpus sellowii Klotzsch (Podocarpaceae) in southeast Brazil.

In southeastern Brazil, the majority of the riparian ecosystems are fragmented and degraded mainly due to human activities. The perennial gymnosperm Podocarpus sellowii Klotzch is a typical tree species from a gallery forest in the rupestrian area. Ten alloenzymatic loci were used to estimate the allelic frequency of 232 individuals distributed in eight subpopulations naturally divided by rock outcrops. The results indicated high genetic variability for the species in all subpopulations, with HO varying from 0.593 to 0.658, and HE, from 0.484 to 0.502. No endogamy was observed within (f = -0.292) and for the population set (f = -0.264). Genetic divergence of the species between subpopulations was 2.1%. Historic gene flow was low between subpopulations located in different water streams, corroborating the positive and significant correlation between genetic and geographical distance (rm = 0.496, P = 0.022). Co-ancestry revealed that only population A presents continuous distribution of the genotypes up to 94 m. Sp statistics did not indicate significant spatial genetic structure in the populations. In all the subpopulations, values of effective sizes were higher than the numbers of sampled individuals. The balance between mutation and drift was not verified, indicating the occurrence of a recent population bottleneck. These data can be used to determine the most effective strategies for the genetic conservation of this species.

Divergent responsiveness of the dentary and vertebral bone to a selective estrogen-receptor modulator (SERM) in the teleost Sparus auratus.

In teleosts the regulation of skeletal homeostasis and turnover by estrogen is poorly understood. For this reason raloxifene, a selective estrogen-receptor modulator (SERM), was administered to sea bream (Sparus auratus) and its effect on plasma calcium balance and transcript expression in dentary (dermal bone) and vertebra (perichondral bone) was studied. The concentration of total calcium or phosphorus in plasma was unchanged by raloxifene treatment for 6days. The activity of alkaline phosphatase (ALP) in dentary bone of raloxifene treated fish was significantly (p<0.05) higher than control fish but it was not changed in vertebral bone. Transcripts for estrogen receptor (ER) α were in very low abundance in the sea bream dentary and vertebra and were unchanged by raloxifene treatment. In contrast, raloxifene caused significant (p<0.05) up-regulation of the duplicate ERß transcripts in the dentary but did not affect specific transcripts for osteoclast (TRAP), osteoblast (ALP, Runx2, osteonectin) or cartilage (IGF1, CILP2, FN1a). In the vertebra ERßb was not changed by raloxifene but ERßa was significantly (p<0.05) down-regulated as was the skeletal specific transcripts, TRAP, ALP, CILP2, FN1a. In summary, ERßs regulate estrogen sensitivity of the skeleton in sea bream, which responds in a non uniform manner. In common with mammals raloxifene appears to have an anti-resorptive role (in sea bream vertebra), but also an osteoblast stimulatory role, inducing ALP activity in the dentary of sea bream. Overall, the results indicate bone specific responsiveness to raloxifene in the sea bream. Further work will be required to understand the basis of bone responsiveness and the role of E(2) and ERs in teleost bone homeostasis.

Genetic diversity of Annona crassiflora (Annonaceae) in northern Minas Gerais State.

Genetic diversity analyses of tropical tree species are relevant to landscape management, plant genetic resource inventory, and biological conservation of threatened species. Annona crassiflora is an endangered fruit tree native to the Cerrado biome that is threatened by reduction of natural populations and fruit extraction. We examined the intra- and interpopulational genetic diversity of this species in the northern region of Minas Gerais State. Seventy-two individuals from four natural populations were genotyped using RAPD markers. We found moderate genetic diversity among populations, with Shannon's I index varying between 0.31 and 0.44, and Nei's genetic diversity (H(E)) for the population set equal to 0.31. AMOVA indicated a greater genetic variation within (77.38%) rather than among populations (22.62%), tending towards isolation by distance (Mantel's r = 0.914; P = 0.089). Nei's genetic identity estimates among populations revealed a hierarchical pattern of genetic similarity of form [(CA1, CA2), MC], [(GM)], corroborating the high genetic differentiation between spatially isolated populations.

IgE cross-reactivity between Lolium multiflorum and commercial grass pollen allergen extracts in Brazilian patients with pollinosis

Lolium multiflorum (Lm) grass pollen is the major cause of pollinosis in Southern Brazil. The objectives of this study were to investigate immunodominant components of Lm pollen allergens and the cross-reactivity of IgE with commercial grass pollen allergen extracts. Thirty-eight serum samples from patients with seasonal allergic rhinitis (SAR), 35 serum samples from patients with perennial allergic rhinitis (PAR) and 30 serum samples from non-atopic subjects were analyzed. Allergen sensitization was evaluated using skin prick test and serum IgE levels against Lm pollen extract were determined by ELISA. Inhibition ELISA and immunoblot were used to evaluate the cross-reactivity of IgE between allergens from Lm and commercial grass pollen extracts, including L. perenne (Lp), grass mix I (GI) and II (GII) extracts. IgE antibodies against Lm were detected in 100 percent of SAR patients and 8.6 percent of PAR patients. Inhibition ELISA demonstrated IgE cross-reactivity between homologous (Lm) and heterologous (Lp or GII) grass pollen extracts, but not for the GI extract. Fifteen IgE-binding Lm components were detected and immunoblot bands of 26, 28-30, and 32-35 kDa showed >90 percent recognition. Lm, Lp and GII extracts significantly inhibited IgE binding to the most immunodominant Lm components, particularly the 55 kDa band. The 26 kDa and 90-114 kDa bands presented the lowest amount of heterologous inhibition. We demonstrated that Lm extract contains both Lm-specific and cross-reactive IgE-binding components and therefore it is suitable for measuring quantitative IgE levels for diagnostic and therapeutic purposes in patients with pollinosis sensitized to Lm grass pollen rather than other phylogenetically related grass pollen extracts.

IgE cross-reactivity between Lolium multiflorum and commercial grass pollen allergen extracts in Brazilian patients with pollinosis.

Lolium multiflorum (Lm) grass pollen is the major cause of pollinosis in Southern Brazil. The objectives of this study were to investigate immunodominant components of Lm pollen allergens and the cross-reactivity of IgE with commercial grass pollen allergen extracts. Thirty-eight serum samples from patients with seasonal allergic rhinitis (SAR), 35 serum samples from patients with perennial allergic rhinitis (PAR) and 30 serum samples from non-atopic subjects were analyzed. Allergen sensitization was evaluated using skin prick test and serum IgE levels against Lm pollen extract were determined by ELISA. Inhibition ELISA and immunoblot were used to evaluate the cross-reactivity of IgE between allergens from Lm and commercial grass pollen extracts, including L. perenne (Lp), grass mix I (GI) and II (GII) extracts. IgE antibodies against Lm were detected in 100% of SAR patients and 8.6% of PAR patients. Inhibition ELISA demonstrated IgE cross-reactivity between homologous (Lm) and heterologous (Lp or GII) grass pollen extracts, but not for the GI extract. Fifteen IgE-binding Lm components were detected and immunoblot bands of 26, 28-30, and 32-35 kDa showed >90% recognition. Lm, Lp and GII extracts significantly inhibited IgE binding to the most immunodominant Lm components, particularly the 55 kDa band. The 26 kDa and 90-114 kDa bands presented the lowest amount of heterologous inhibition. We demonstrated that Lm extract contains both Lm-specific and cross-reactive IgE-binding components and therefore it is suitable for measuring quantitative IgE levels for diagnostic and therapeutic purposes in patients with pollinosis sensitized to Lm grass pollen rather than other phylogenetically related grass pollen extracts.

Composição bromatológica da silagem de quatro genótipos de girassol, ensilados em cinco diferentes idades de corte / Bromatological composition of si/age of four sunflower genotypes ensiled at different harvesting ages

Determinaram-se a composição bromatológica do material original e das silagens de quatro genótipos de girassol (M742, MG4, PM92007 e VDH483) utilizados para produção de grãos, ensilados em diferentes idades de corte após o plantio e aos 90, 97, 104, 111, e 118 dias, e o melhor momento de colheita desses cultivares para ensilagem. Os teores de matéria seca variaram entre as idades e cultivares, sendo observados valores de 12,7 a 42,2 no material original e 11,1 e 40,2 na silagem. Os valores de proteína bruta variaram de 8,3 a 12,6 no material original e de 9,4 a 12,9 na silagem. Foi observada diminuição (P<0,05) nos teores de carboidratos com a ensilagem. Os teores de extrato etéreo do material original e das silagens variaram de 2,2 a 13,3 no material original e de 3,0 a 17,9 na ensilagem. Houve diminuição dos teores de hemicelulose com a ensilagem, sugerindo que essa fração foi utilizada com substrato adicional para fermentação. Os teores de ácido lático diminuíram com o estágio de maturação da planta. Houve diminuição nos valores de digestibilidade in vitro da matéria seca com o avanço do estádio de maturação da planta. O melhor momento de colheita para os genótipos M742, MG4 e VDH483 foi aos 104 dias de idade e para o genótipo PM92007, aos 111 dias.

Estudo técnico e econômico de um sistema de produção de leite com gado mestiço F1 Holandês-Zebu / Technical and economic evaluation on of a milk production system for F1 Holstein-Zebu cattle

Avaliaram-se a viabilidade técnica e a rentabilidade de um sistema de produção de leite com gado mestiço, mediante análise de custos e receitas da atividade e avaliação da rentabilidade. Os dados foram obtidos de uma unidade experimental da EPAMIG, situada no município de Felixlândia, MG, e referem-se a dois períodos iguais de um ano cada. As variáveis analisadas foram: rentabilidade nos períodos de análise, custo de cria do bezerro e sua importância sobre a receita da atividade. A viabilidade do sistema foi demonstrada pela rentabilidade de 20,2 por cento no ano 1 e 37,6 por cento no ano 2; a venda dos bezerros à desmama representou 25 por cento da receita total.

Qualidade de silagens de sorgo com aditivos / Quality of sorghum silages with additives

Um delineamento inteiramente ao acaso, com três repetições, foi utilizado para avaliar os efeitos da adição de 0,5 por cento de uréia, de 0,5 por cento de carbonato de cálcio (CaCO3), de 0,5 por cento de uréia mais 0,5 por cento de CaCO3 e de inoculante bacteriano sobre o pH, teores de matéria seca (MS), proteína bruta (PB), fibra em detergente neutro (FDN), fibra em detergente ácido (FDA), hemicelulose, celulose, lignina, carboidratos solúveis, ácido lático e ácido acético sobre a digestibilidade in vitro da matéria seca (DIVMS) de silagens de quatro híbridos de sorgo (BR700, BR701, BR601 e AG2002). Os híbridos ensilados sem a adição de aditivos constituíram o grupo-controle. De cada genótipo foram coletadas três amostras do material original (MOR). Foram utilizados 60 silos de PVC com 10cm de diâmetro interno e 50cm de comprimento, sendo três por tratamento. A abertura dos silos ocorreu depois de 56 dias de fermentação. As silagens contendo CaCO3 e inoculante bacteriano apresentaram, em geral, características muito semelhantes às do controle. A adição de uréia e de uréia mais CaCO3 às silagens resultou em aumento (P<0,05) nos teores de PB. A adição conjunta de uréia e CaCO3 não propiciou aumento (P<0,05) superior ao obtido nas silagens contendo apenas uréia. Uréia pura, como aditivo, pode ser recomendada na ensilagem dos híbridos BR700, BR601 e AG2002.