Estudo do efeito de dois intervalos de crescimento e seis níveis de azoto na produção de matéria seca e na qualidade de uma pastagem de Lolium perenne, em consociação com Trifolium repens e Trifolium pratense

Dissertação de Mestrado, Engenharia Zootécnica, 13 de Janeiro de 2016 , Universidade dos Açores.

Determination of the impact of continuous defoliation of Lolium perenne and Trifolium repens on bacterial and fungal community structure in rhizosphere soil

To determine the effects of defoliation on microbial community structure, rhizosphere soil samples were taken pre-, and post-defoliation from the root tip and mature root regions of Trifolium repens L. and Lolium perenne L. Microbial DNA isolated from samples was used to generate polymerase chain reaction-denaturing gradient gel electrophoresis molecular profiles of bacterial and fungal communities. Bacterial plate counts were also obtained. Neither plant species nor defoliation affected the bacterial and fungal community structures in both the root tip and mature root regions, but there were significant differences in the bacterial and fungal community profiles between the two root regions for each plant. Prior to defoliation, there was no difference between plants for bacterial plate counts of soils from the root tip regions; however, counts were greater in the mature root region of L. perenne than T. repens. Bacterial plate counts for T. repens were higher in the root tip than the mature root region. After defoliation, there was no effect of plant type, position along the root or defoliation status on bacterial plate counts, although there were significant increases in bacterial plate counts with time. The results indicate that a general effect existed during maturation in the root regions of each plant, which had a greater impact on microbial community structure than either plant type or the effect of defoliation. In addition there were no generic consequences with regard to microbial populations in the rhizosphere as a response to plant defoliation.

Protein precipitation behaviour of condensed tannins from Lotus pedunculatus and Trifolium repens with different mean degrees of polymerization

The precipitation of bovine serum albumin (BSA), lysozyme (LYS) and alfalfa leaf protein (ALF) by two large- and two medium-sized condensed tannin (CT) fractions of similar flavan-3-ol subunit composition is described. CT fractions isolated from white clover flowers and big trefoil leaves exhibited high purity profiles by 1D/2D NMR and purities >90% (determined by thiolysis). At pH 6.5, large CTs with a mean degree of polymerization (mDP) of ~18 exhibited similar protein precipitation behaviors and were significantly more effective than medium CTs (mDP ~9). Medium CTs exhibited similar capacities to precipitate ALF or BSA, but showed small but significant differences in their capacity to precipitate LYS. All CTs precipitated ALF more effectively than BSA or LYS. Aggregation of CT-protein complexes likely aided precipitation of ALF and BSA, but not LYS. This study, one of the first to use CTs of confirmed high purity, demonstrates that mDP of CTs influences protein precipitation efficacy.

Caracterização morfológica e molecular da coleção básica de trevo-branco (Trifolium repens L.)

O objetivo deste trabalhofoi avaliarcaracterísticas moleculares e morfológicasde 79 acessos pertencentes à coleção básica de trevo-branco obtida do Departamento de Agricultura dos Estados Unidos(USDA), a fim de caracterizar a variabilidade genética existente.

Efeito de sistemas de pastejo sobre o desempenho e o comportamento de suínos em crescimento e terminação, mantidos em pastagem de trevo-branco (Trifolium repens L.)

O presente trabalho foi conduzido com o objetivo de avaliar o efeito de diferentes sistemas de pastejo (contínuo, alternado e rotativo) sobre o desempenho e o comportamento de suínos, assim como, sobre a disponibilidade e a composição química do trevo-branco (Trifolium repens L.). Foram utilizados 48 suínos machos, castrados, com peso médio inicial de 27 kg, sendo doze mantidos em sistema confinado, os quais foram utilizados como tratamento testemunha. Os animais foram distribuídos em um delineamento de blocos casualizados, com três tratamentos e três repetições. Não foram observadas diferenças significativas (P>0,05) entre os sistemas de pastejo nas características de desempenho e de disponibilidade. No entanto, foram observadas diferenças significativas (P<0,05) da composição química da pastagem entre os sistemas de pastejo estudados. Verificou-se que os suínos em sistemas de pastejo contínuo, alternado e rotativo consumiram de 13% a 17% a menos de ração (P<0,05) e apresentaram menores ganhos de peso (P<0,05) e espessura de toucinho (P<0,05) do que os suínos criados em confinamento. Não foram observadas diferenças significativas (P>0,05) na conversão alimentar entre os suínos mantidos em confinamento e sobre pastagem. Deitar, beber água, ingerir ração, pastejar e caminhar foram as atividades que apresentaram maiores freqüências de ocorrência, independente dos sistemas de pastejo estudados. Houve efeito significativo dos sistemas de pastejo sobre a atividade de ingerir ração (P=0,0186) e interação de sistema x dia de observação para as atividades de deitar (P=0,0345) e pastejar (P=0,0678).

Avaliação do potencial de produção de sementes de acessos de trevo branco (Trifolium repens L)

A produção de sementes de qualidade é um dos pontos chave na difusão do trevo branco em nosso meio. Esta leguminosa é uma espécie de boa produção de sementes, entretanto, como as demais espécies de Trifolium, é conhecida por flutuações no rendimento de sementes. Com o objetivo de avaliar o potencial de produção de sementes de 27 acessos da coleção básica de trevo branco conduziu-se o presente trabalho na região da Depressão Central do RS, EEA/UFRGS, Eldorado do Sul, no período de outubro de 2003 a março de 2004. As variáveis estudadas através de amostragens semanais realizadas entre 06/11 e 10/03 foram: número de inflorescências por planta, número de flores por inflorescência, número de inflorescências maduras, número de legumes maduros por inflorescência, peso de 1000 sementes e rendimento de sementes por planta. Foram separados três grupos distintos quanto à produção de sementes. O grupo superior com o acesso 53 (Peru), outro intermediário com os acessos 2 (Israel), 3 (EUA), 7 (Etiópia), 13 (Espanha), 19 (Costa Rica), 20 (Uruguai), 28 (Austrália), 33 (Irlanda), 50 (África do Sul), 58 (Índia), 64 (Suíça), 68 (Jamaica) e 75 (Uruguai) e um grupo inferior com os acessos 15 (Nova Zelândia), 22 (Hungria), 23 (Bélgica), 29 (França), 31 (EUA), 38 (Irã), 54 (Nova Zelândia), 59 (Líbano), 79 (Nova Zelândia) e 80 (EUA). O número de inflorescências, número de inflorescências maduras e peso de 1000 sementes apresentaram correlação positiva e altamente significativa em relação ao rendimento de sementes. O acesso 20 apresentou superioridade no número de inflorescências por planta e número de inflorescências maduras por planta, no entanto, o acesso 53 foi o de maior potencial de produção de sementes, em função de sua superioridade no peso de 1000 sementes. Os acessos 27 (Canadá), 65 (Dinamarca) e 73 (Polônia) não produziram sementes nas condições locais.

Identification and expression of different dehydrin subclasses involved in the drought response of Trifolium repens

Reverse transcribed RNAs coding for YnKn, YnSKn, SKn, and KS dehydrin types in drought-stressed white clover (Trifolium repens) were identified and characterized. The nucleotide analyses revealed the complex nature of dehydrin-coding sequences, often featured with alternative start and stop codons within the open reading frames, which could be a prerequisite for high variability among the transcripts originating from a single gene. For some dehydrin sequences, the existence of natural antisense transcripts was predicted. The differential distribution of dehydrin homologues in roots and leaves from a single white clover stolon under normal and drought conditions was evaluated by semi-quantitative RT-PCR and immunoblots with antibodies against the conserved K-, Y- and S-segments. The data suggest that different dehydrin classes have distinct roles in the drought stress response and vegetative development, demonstrating some specific characteristic features. Substantial levels of YSK-type proteins with different molecular weights were immunodetected in the non-stressed developing leaves. The acidic SK2 and KS dehydrin transcripts exhibited some developmental gradient in leaves. A strong increase of YK transcripts was documented in the fully expanded leaves and roots of drought-stressed individuals. The immunodetected drought-induced signals imply that Y- and K-segment containing dehydrins could be the major inducible Late Embryogenesis Abundant class 2 proteins (LEA 2) that accumulate predominantly under drought.

Natural antisense transcripts of Trifolium repens dehydrins

The recently described complex nature of some dehydrin-coding sequences in Trifolium repens could explain the considerable variability among transcripts originating from a single gene.1 For some of the sequences the existence of natural antisense transcripts (NAT s), which could form sense-antisense (SAS) pairs, was predicted. The present study demonstrates that cis-natural antisense transcripts of 2 dehydrin types (YnKn and YnSKn) accumulate in white clover plants subjected to treatments with polyethylene glycol (PEG), abscisic acid (ABA), and high salt concentration. The isolated YnKn cis-NAT s mapped to sequence site enriched in alternative start codons. Some of the sense-antisense pairs exhibited inverse expression with differing profiles which depended on the applied stress. A natural antisense transcript coding for an ABC F family protein (a trans-NAT) which shares short sequence homology with YnSKn dehydrin was identified in plants subjected to salt stress. Forthcoming experiments will evaluate the impact of NAT s on transcript abundances, elucidating the role of transcriptional and post-transcriptional interferences in the regulation of dehydrin levels under various abiotic stresses.

Transgenerational effects of land use on offspring performance and growth in Trifolium repens

entral European grasslands vary widely in productivity and in mowing and grazing regimes. The resulting differences in competition and heterogeneity among grasslands might have direct effects on plants, but might also affect the growth and morphology of their offspring through maternal effects or adaptive evolution. To test for such transgenerational effects, we grew plants of the clonal herb Trifolium repens from seeds collected in 58 grassland sites differing in productivity and mowing and grazing intensities in different treatments: without competition, with homogeneous competition, and with heterogeneous competition. In the competition-free treatment, T. repens from more productive, less frequently mown, and less intensively grazed sites produced more vegetative offspring, but this was not the case in the other treatments. When grown among or in close proximity to competitors, T. repens plants did not show preferential growth towards open spaces (i.e., no horizontal foraging), but did show strong vertical foraging by petiole elongation. In the homogeneous competition treatment, petiole length increased with the productivity of the parental site, but this was not the case in the heterogeneous competition treatment. Moreover, petiole length increased with mowing frequency and grazing intensity of the parental site in all but the homogeneous competition treatment. In summary, although the expression of differences between plants from sites with different productivities and land-use intensities depended on the experimental treatment, our findings imply that there are transgenerational effects of land use on the morphology and performance of T. repens.

Timpanismo espumoso em bovinos leiteiros em pastagens de Trifolium spp. (Leg.Caesalpinoideae)

O pastejo de plantas leguminosas que provocam fermentação excessiva pode causar surtos de timpanismo e mortes em ruminantes. Em uma propriedade no município de São Francisco de Paula, Rio Grande do Sul, oito bovinos de um total de 66 morreram subitamente, ao haverem sido transferidos de um potreiro de campo nativo, para outro cuja pastagem era composta por Trifolium repens e Trifolium pratense. Os animais foram encontrados mortos no amanhecer do dia seguinte à transferência, não foram observados sinais clínicos prévios. Os principais achados macroscópicos incluíram aumento de volume abdominal, protrusão de vagina e língua, distensão ruminal, fígado de coloração pálida e aumento do baço. Na histologia, havia congestão e edema pulmonares e hiperplasia linfóide difusa e acentuada no baço. A evidência de ingestão das leguminosas associada aos achados patológicos e à ausência de microrganismos no exame bacteriológico confirmaram o diagnóstico de timpanismo.

Caracterização isoenzimática de oito espécies do gênero Trifolium L., ocorrentes no Rio Grande do Sul

O presente estudo tem como objetivos: contribuir para o desenvolvimento de oito espécies do gênero Trifolium L., Trifolium incamatum L., Trifolium polymorphum Poir, Trifolium pratense L., Trifolium repens L., trifolium resupinatum L., Trifolium riograndense Burkart, Trifolium subterraneum L. e Trifolium vesiculosum Savi, através da caracterização da variabilidade isoenzimática presente em quatro sistemas enzimáticos, Fosfoglicoisomerase (PGI), Esterase (EST), Malato desidrogenase (MDH) e Superóxido dismutase (SOD) e fornecer subsídios para futuros trabalhos de melhoramento de plantas forrageiras.

Rhizodeposits of Trifolium pratense and Lolium perenne: Their comparative effects on 2,4-D mineralization in two contrasting soils

Rhizosphere enhanced biodegradation of organic pollutants has been reported frequently and a stimulatory role for specific components of rhizodeposits postulated. As rhizodeposit composition is a function of plant species and soil type, we compared the effect of Lolium perenne and Trifolium pratense grown in two different soils (a sandy silt loam: pH 4, 2.8% OC, no previous 2,4-D exposure and a silt loam: pH 6.5, 4.3% OC, previous 2,4-D exposure) on the mineralization of the herbicide 2,4-D (2,4-dichlorophenoxyacetic acid). We investigated the relationship of mineralization kinetics to dehydrogenase activity, most probable number of 2,4-D degraders (MPN2,4-D) and 2,4-D degrader composition (using sequence analysis of the gene encoding alpha-ketoglutarate/2,4-D dioxygenase (tfdA)). There were significant (P < 0.01) plant-soil interaction effects on MPN2,4-D and 2,4-D mineralization kinetics (e.g. T pratense rhizodeposits enhanced the maximum mineralization rate by 30% in the acid sandy silt loam soil, but not in the neutral silt loam soil). Differences in mineralization kinetics could not be ascribed to 2,4-D degrader composition as both soils had tfdA sequences which clustered with tfdAs representative of two distinct classes of 2,4-D degrader: canonical R. eutropha JMP134-like and oligotrophic alpha-proteobacterial-like. Other explanations for the differential rhizodeposit effect between soils and plants (e.g. nutrient competition effects) are discussed. Our findings stress that complexity of soil-plant-microbe interactions in the rhizosphere make the occurrence and extent of rhizosphere-enhanced xenobiotic degradation difficult to predict.

Cyanogenic polymorphism in Eucalyptus polyanthemos Schauer subsp vestita L. Johnson and K. Hill (Myrtaceae)

Plant cyanogenesis, the release of cyanide from endogenous cyanide-containing compounds, is an effective herbivore deterrent. This paper characterises cyanogenesis in the Australian tree Eucalyptus polyanthemos Schauer subsp. vestita L. Johnson and K. Hill for the first time. The cyanogenic glucoside prunasin ((R)-mandelonitrile beta-D-glucoside) was determined to be the only cyanogenic compound in E. polyanthemos foliage. Two natural populations of E. polyanthernos showed quantitative variation in foliar prumasin concentration, varying from zero (i.e. acyanogenic) to 2.07 mg CN g(-1) dry weight in one population and from 0.17 to 1.98 mg CN g(-1) dry weight in the other. No significant difference was detected between the populations with respect to the mean prunasin concentration or the degree of variation in foliar prunasin, despite significant differences in foliar nitrogen. Variation between individuals was also observed with respect to the capacity of foliage to catabolise prunasin to form cyanide. Moreover, variation in this capacity generally correlated with the amount of prunasin in the tissue, suggesting genetic linkage between prunasin and beta-glucosidase. (C) 2002 Elsevier Science Ltd. All rights reserved.

Identification of research to improve the efficiency of breeding strategies for white clover in Australia - a review

A major challenge faced by today's white clover breeder is how to manage resources within a breeding program. It is essential to utilise these resources with sufficient flexibility to build on past progress from conventional breeding strategies, but also take advantage of emerging opportunities from molecular breeding tools such as molecular markers and transformation. It is timely to review white clover breeding strategies. This background can then be used as a foundation for considering how to continue conventional plant improvement activities and complement them with molecular breeding opportunities. In this review, conventional white clover breeding strategies relevant to the Australian dryland target population environments are considered. Attention is given to: (i) availability of genetic variation, (ii) characterisation of germplasm collections, (iii) quantitative models for estimation of heritability, (iv) the role of multi-environment trials to accommodate genotype-by-environment interactions, (v) interdisciplinary research to understand adaptation to dryland environments, (vi) breeding and selection strategies, and (vii) cultivar structure. Current achievements in biotechnology with specific reference to white clover breeding in Australia are considered, and computer modelling of breeding programs is discussed as a useful integrative tool for the joint evaluation of conventional and molecular breeding strategies and optimisation of resource use in breeding programs. Four areas are identified as future research priorities: (i) capturing the potential genetic diversity among introduced accessions and ecotypes that are adapted to key constraints such as summer moisture stress and the use of molecular markers to assess the genetic diversity, (ii) understanding the underlying physiological/morphological root and shoot mechanisms involved in water use efficiency of white clover, with the objective of identifying appropriate selection criteria, (iii) estimation of quantitative genetic parameters of important morphological/physiological attributes to enable prediction of response to selection in target environments, and (iv) modelling white clover breeding strategies to evaluate the opportunities for integration of molecular breeding strategies with conventional breeding programs.

Projecto de um sistema de rega para uma exploração agrícola na Beira Gomes

Trabalho de Projecto de Natureza Científica para obtenção do grau de Mestre em Engenharia Civil