Ancestors of white clover ( Trifolium repens L.), as revealed by isozyme polymorphisms

Isozymes were used to study the putative ancestors of white clover ( Trifolium repens L.). Ten enzymes were examined, and 18 loci were resolved via starch-gel electrophoresis for accessions representing T. repens, Trifolium isthmocarpum, Trifolium nigrescens, Trifolium occidentale and Trifolium uniflorum, in addition to two more distantly related species, Trifolium alpinum and Trifolium purseglovei. Nei's genetic identities indicate that T. uniflorum and T. nigrescens are the closest relatives of T. repens. The isozyme data thus support a hypothesis that the two genomes of the tetraploid T. repens could have been derived from hybridization between T. nigrescens and T. uniflorum. This conclusion is further supported by shared alleles between T. repens, T. nigrescens and T. uniflorum. However, the origin of T. repens is somewhat obscured by the presence of shared alleles between T. repens and both T. occidentale and T. isthmocarpum, suggesting that introgression of genes from the latter two species into T. repens may also have taken place. High values of genetic identity are shared between T. occidentale with T. nigrescens and T. uniflorum, also indicative of introgression. Alternatively the presence of shared alleles among the five species may reflect their recent common ancestry.     

On the origin of the cyanogenic polymorphism in Trifolium repens L.

Trifolium repens L. and Trifolium nigrescens Viv. are two of the approximately six cyanogenic species known in the genus Trifolium. The two species are closely related: T. nigrescens is considered to be one of the diploid ancestors of the amphidiploid T. repens . We studied morphology, meiosis and the cyanogenic system in T. repens (amphidiploid), T. nigrescens (diploid) and their reciprocal hybrids. A comparison of the enzyme linamarase in the species and hybrids shows that there is a general resemblance between their linamarases. Immunological studies indicate that the linamarases must have a somewhat different three-dimensional structure. These facts are consistent with the view that T. nigrescens (or an ancestral form of this species) has donated the Li gene of T. repens . The other putative parent, T. occidentale has probably not donated an active Li gene. The hypothesis of the origin of the Li gene does not explain its polymorphism in European populations of T. repens , as T. nigrescens is monomorphic for cyanogenesis and amphidiploids do not segregate for genes which are homozygous dominant in one of the parents. Segregation for Li could be caused by a gene mutation or a small exchange between homeologous chromosomes. The latter event is more probable. A nigrescens-repens exchange would give rise to a chromosomal region with reduced homology to both parental chromosomes. The genes in the region of exchange will be tightly linked due to diminished cross-over frequency. It has been known for years that Li has effects on the vegetative and reproductive characters in T. repens and we have recently shown that these effects must be the result of genes linked to Li . As the associated characters influence the fitness of the cyanotypes, not only the origin but also the maintenance of the cyanogenic polymorphism is closely related to the evolutionary history of T. repens .     

VARIATION IN ALCOHOL DEHYDROGENASE ACTIVITY AND FLOOD TOLERANCE IN WHITE CLOVER,TRIFOLIUM REPENS

Flooding results in induction of anaerobic metabolism in many higher plants. As an important component of anaerobic energy production, alcohol dehydrogenase (ADH) activity increases markedly in response to flooding in white clover, Trifolium repens. Significant inter-individual variation in flood-induced ADH activity exists in natural populations of T. repens. The genetic basis of this variation was analyzed by offspring-midparent regression of data from 75 greenhouse reared families; the estimated heritability of flood-induced ADH activity was 0.55 (±0.13). Genetic variation in flood-induced ADH activity has pronounced effects on physiological response and flood tolerance in this species. ADH activity is positively correlated with the rate of ethanol production, indicating that observed in vitro activity differences are manifested in in vivo physiological function. T. repens plants with higher ADH activities during flooding have greater flood tolerance (measured as growth rate when flooded/unflooded growth rate). Variation in ADH activity during flooding accounts for more than 79% of the variance in flood tolerance. On the basis of a limited field survey of populations occupying three sites differing in exposure to flooding conditions, individuals from site C, the most frequently flooded site, expressed significantly higher average ADH activity when flooded than individuals from site A, a site with no history of flooding. Since ADH activity levels are not correlated with electrophoretic mobility variation in T. repens, this work supports previous suggestions that regulatory variation in enzyme activity may play a central role in biochemical adaptations to environmental stress.     

Genetic consequences of a single-founder population bottleneck in Trifolium amoenum (Fabaceae)

We investigated the genetic consequences of a single-founder bottleneck in a population of showy Indian clover (Trifolium amoenum), a species presumed to be extinct until rediscovered near Occidental, California, in 1993. Electrophoretic variation was evaluated in the bottlenecked population and in a larger population (Dillon Beach) discovered during the course of this study, as well as in populations of two closely related species, T. albopurpureum var. dichotomum and T. macraei. We found a surprisingly high amount of polymorphism in the single-founder T. amoenum population from Occidental (15% of loci polymorphic; an average of 1.1 alleles per locus). However, this represents a 53% reduction in number of polymorphic loci and a 20% reduction in average number of alleles per locus compared to three Trifolium populations with putatively similar mating systems (the Dillon Beach T. amoenum population and both populations of T. albopurpureum var. dichotomum). Expanding the genetic base of the Occidental T. amoenum population is a priority due to concerns about loss of evolutionary potential and the possibility of deleterious effects associated with inbreeding. However, using seed from the Dillon Beach T. amoenum population may not be beneficial due to distinct, presumably adaptive differences between plants from the two populations and concerns about outbreeding depression.     

Morphological response to competition for light in the clonal Trifolium repens (Fabaceae)

? Premise of the study: Plant communities in temperate zones are dominated by clonal plants that can plastically modify their growth characteristics in response to competition. Given that plants compete with one another, and the implications this has for species coexistence, we conducted a study to assess how clonal species morphologically respond to competition for light depending on its intensity and heterogeneity, which are determined by the competitor species. ? Methods: We assessed the morphological response to competition for light of the clonal species Trifolium repens L. by measuring its growth performance, and vertical and horizontal growth traits. We used five competitive environments, i.e., one without competitor and four differing by their competitor species creating different conditions of competition intensity and heterogeneity. ? Key results: The morphological response of Trifolium repens to competition for light depended on the competitor identity. Competition intensity and heterogeneity, determined by competitor identity, had an interactive effect on most traits. The increase in petiole elongation and specific leaf area due to increased competition intensity was observed only at low to intermediate competition heterogeneity. Competition heterogeneity promoted the elongation of clone connections allowing space exploration. ? Conclusions: Our results demonstrated that the intensity and heterogeneity of competition, which depended on competitor identity, are of primary importance in determining the plastic response of Trifolium repens. This emphasizes that it is important to consider the fine-scale spatial distribution of individuals when studying their interactions within plant communities.     

Identification of homologous, homoeologous and paralogous sequence variants in an outbreeding allopolyploid species based on comparison with progenitor taxa

The combination of homologous, homoeologous and paralogous classes of sequence variation presents major challenges for SNP discovery in outbreeding allopolyploid species. Previous in vitro gene-associated SNP discovery studies in the allotetraploid forage legume white clover (Trifolium repens L.) were vulnerable to such effects, leading to prohibitive levels of attrition during SNP validation. Identification of T. occidentale and T. pallescens as the putative diploid progenitors of white clover has permitted discrimination of the different sequence variant categories. Amplicons from selected abiotic stress tolerance-related genes were obtained using mapping family parents and individuals from each diploid species. Following cloning, progenitor comparison allowed tentative assignment of individual haplotypes to one or other sub-genome, as well as to gene copies within sub-genomes. A high degree of coincidence and identity between SNPs and HSVs was observed. Close similarity was observed between the genome of T. occidentale and one white clover sub-genome, but the affinity between T. pallescens and the other sub-genome was weaker, suggesting that a currently uncharacterised taxon may be the true second progenitor. Selected validated SNPs were attributed to individual sub-genomes by assignment to and naming of homoeologous linkage groups, providing the basis for improved genetic trait-dissection studies. The approach described in this study is broadly applicable to a range of allopolyploid taxa of equivocal ancestry.     

Negative effects of habitat degradation and fragmentation on the declining grassland plant Trifolium montanum

Changes in land use have resulted in a strong decline in the plant diversity of nutrient-poor grasslands, but little is known about the combined effects of habitat degradation and fragmentation on populations of individual species. We studied these effects on stage structure, recruitment, reproduction and offspring fitness in populations of the declining perennial grassland plant Trifolium montanum in central Germany. Density and survival probability of juvenile plants decreased with light competition, measured as leaf area index (LAI) above T. montanum plants, resulting in aged populations with few juvenile plants at unmanaged sites with higher LAI. Reproduction of T. montanum was not related to LAI, but increased strongly with local density, suggesting pollinator limitation in fragmented populations with a low density of flowering plants. In the common garden, the survival of sown offspring increased with mean seed size, whereas seed production of offspring decreased with isolation, and in strong contrast to previous studies, also decreased with size and density of the population of origin. This could be due to increased inbreeding because of pollination between closely related neighbouring plants in dense and large populations. Our results indicate that both habitat degradation and fragmentation have negative effects on populations of T. montanum, but affect different phases of the life cycle. In the short term, the effects of habitat degradation are more important than those of fragmentation, and populations of T. montanum are primarily threatened by an increase in light competition in unmanaged sites, which rapidly affects the dynamics of the populations. The observed opposite effects of habitat fragmentation on reproduction and offspring fitness indicate that the effects of population size, density and isolation on plant fitness and population viability may be complex.     

狗牙根、白三叶草对淡剑纹灰翅夜蛾生长发育和繁殖的影响

利用室内饲喂法,以高羊茅分别为对照,研究狗牙根(Cynodon dacyton)、白三叶草(Trifolium repens)对淡剑纹灰翅夜蛾Spodoptera depravata(Butler)存活、生长发育及繁殖的影响。结果表明,用狗牙根、白三叶草饲喂淡剑纹灰翅夜蛾的化蛹率、幼虫成蛾率均显著低于对照组。幼虫历期显著长于对照组。用狗牙根、白三叶草饲喂幼虫时,淡剑纹灰翅夜蛾的蛹期、产卵前期、产卵期、产卵量、♀成虫寿命与对照组均无显著差异,但幼虫龄期明显增加。狗牙根、白三叶草饲喂的淡剑纹灰翅夜蛾种群增长参数与对照组相比存在明显差异:(1)世代净生殖率(R0);(2)内禀增长率(rm);(3)周限增长率(λ);(4)种群数量趋势指数(I)明显降低;世代平均周期(T)和种群加倍时间(DT)明显延长。由此可见,狗牙根和白三叶草并不是淡剑纹灰翅夜蛾喜好的寄主植物。     

Self-thinning in Trifolium subterraneum not affected by cultivar shape

As it was thought that self-thinning in crowded plant populations might be influenced by the shape of the plants involved, self-thinning was examined in five cultivars of Trifolium subterraneum. The height of these cultivars is fixed after early growth, being determined mainly by petiole length which itself varies between cultivars. No detectable differences between cultivars were found, and it is argued that only wore considerable differences in shape than those reported here would have detectable effects. When plants can only grow radially but not in height, it is to be expected that log-log self-thinning curves would have a slope of -1, but these Trifolium populations had the – 3/2 slope usually found for species which exhibit continuous growth in height. This finding remains unexplained.     

Detection of selection in populations of white clover (Trifolium repens L.)

Seed populations of white clover polymorphic for the presence/absence of both ovariogenic glucosides and the hydrolysing enzyme linamarase, were introduced into three natural populations. Over the first six months of life a significant increase in the frequency of linamarase containing individuals occurred. Estimated selection coefficients against plants lacking linamarase were in the region of 0.3. This result may have been due to selection at the enzyme locus alone, or to selection favouring cyanogenic individuals which possess both cyanogenic glucosides and enzyme.     

小麦—白三叶草复合群体中种群关系的初步研究

小麦和白三叶草（Ｔｒｉｆｏｌｉｕｍｒｅｐｅｎｓ）复合共生,是一种新型的农田人工群落。其功能表现为种养（养地与养殖）的有效结合,并通过小麦与三叶草的群体优势抑制农田杂草种群。因此,在理论上,这种复合群体是多功能效益,而且是相对稳定的农田生态系。在生产实...     

Interaction between Some Pasture Species and 2

Possible control options are investigated for the introduced Hieracium weeds, particular problems in South Island high country, New Zealand. In a pot experiment regression of input to output ratios of above ground biomass over successive harvests, from binary mixtures was used to determine the competitive interaction between 13 pasture species and two Hieracium species, H. pilosella and H. praealtum, in a low fertility soil. Treatments also included a factorial of presence or absence of compartments separating root and shoots of species. Species differed in their mean growth rate, relative to Hieracium species. The ranking of mean growth rate relative to H. pilosella was Trifolium repens (best), Bromus inermis, Sanguisorba minor, Festuca novaezelandiae, F. rubra, Arrhenatherum elatius, Anthoxanthum odoratum, Lotus corniculatus, Hypochoeris radicata, Trifolium repens, T. hybridum, T. medium and Astragalus cicer. Shoot partitions decreased Hieracium's interaction with white clover while root partitions increased interaction with B. inermis. However, the rates were not related to the proportion of Hieracium in the mixtures, indicating a general lack of specific competitive effects against Hieracium.     

Response of Trifolium repens to UV-B radiation: morphological links to plant productivity and water availability

This study used nine populations of Trifolium repens L. (white clover) to investigate possible relationships between plant morphological attributes and responses to ultraviolet-B (UV-B) radiation. Plants were exposed to 0 or 13.3 kJ·m(-2) ·day(-1) UV-B for 12 weeks. Drought was applied in parallel to these treatments during the last 4 weeks of the experiment to test whether limited moisture availability would alter morphological UV-B responses. UV-B affected plant morphology under well-watered conditions, reducing leaf size by 15%, leaf number by 5% and stolon elongation by 19%. The number of leaf primordia in the apical bud was decreased by 4% under UV-B, and by 12% under drought. In drought-exposed plants, leaf size was reduced by 50%, leaf number by 30% and stolon elongation by 60%. In addition, drought reduced specific leaf area (SLA) by 33% and increased leaf percentage dry mass (PDM) by 40%. UV-B-induced reduction in plant biomass in the T. repens populations was associated with higher plant productivity and this was further linked to larger leaf size as well as to lower PDM. In conclusion, the findings suggest that morphological attributes conferring fast potential growth under productive conditions carry a cost in the form of lower biomass accumulation under UV-B.     

Developmental and Physiological Studies on the Cyanogenic Glucosides of White Clover, Trifolium repens L.

Cyanogenesis in Trifolium repens L. is under the control oftwo loci; Ac/ac and Li/Hi control cyanogenic glucoside and linamaraseproduction respectively. Results obtained show that neitherthe dominant allele (Ac) coding for cyanogenic glucoside productionnor the dominant allele (Li) coding for linamarase productionare expressed in roots, seeds or seedlings before shoot emergence.Both linamarase and cyanogenic glucoside are produced duringshoot growth and there is little turnover of cyanogenic glucosidein mature leaves. As the leaves senesce there is breakdown ofthe mechanism separating cyanogenic glucoside and linamarase,since cyanogenic glucoside is lost in plants of genotype AcAc Li Li but not in those of genotype Ac Ac Li Li. About 60%of the cyanogenic glucoside produced was lotaustralin, in shootsof plants which were fed with equal quantities of the precursoramino acids L-valine and L-isoleucine. In contrast, the proportionof cyanogenic glucoside as lotaustralin found in leaves of oneplant, was only 40%. Different plants were shown to producedifferent quantities of cyanogenic glucoside, and the amountproduced was dependent on temperature.     

Genetic analysis reveals human-mediated long-distance dispersal among war cemeteries in Trifolium micranthum

Many grassland plant species have limited capacity to disperse their seeds beyond local boundaries by natural means. Meanwhile, various forms of human transportation are observed to provide long-distance dispersal. However, the contribution of human-mediated dispersal to the spatial dynamics of established (meta)populations on the regional scale has only scarcely been addressed. Trifolium micranthum is a very small legume that is rare in western Belgium. It is found (i) inland, as a lawn weed on war cemeteries that are managed by the Commonwealth War Graves Commission, and (ii) in coastal grasslands that are managed for conservation. These stations lie over 30?km apart, though interestingly, a number of satellite war cemeteries are found within the vicinity of the coastal reserves, and these also harbor T. micranthum. We have sampled plants from these populations to study their affinities through genetic analysis (AFLP). Results indicated that the coastal cemetery populations were closely related to the inland cemeteries and not to the coastal reserves. This most likely is the result of exchange of seeds with composted lawn material among regions, whereas mowing machinery and livestock may effectuate frequent dispersal within regions. Assignment tests nonetheless indicated limited genetic admixture to have taken place at the coastal cemeteries. Thus, human management is responsible for the realization of gene flow among these remote gene pools. Human-mediated long-distance dispersal is likely to play a role in the local or regional dynamics of many other native plants, including more common and well-dispersing species.     

Rhizobium strains that nodulate Trifolium semipilosum Fres. are phylogenetically distinct

The genetic relationships of Rhizobium isolated from temperate and tropical perennial Trifolium species were investigated using PCR-based nucleotide sequence analysis of 16S and 23S rDNA regions. Comparative analysis of partial 23S rDNA sequences clustered Rhizobium isolates effective with T. semipilosum, T. repens, T. pratense, T. hybridum and T. fragiferum into two distinct groups. These groups were consistent with the pattern of symbiotic effectiveness observed in cross-inoculation experiments. Our data suggested that strains from T. semipilosum were more closely related phylogenetically to R. etli, indicating that these strains do not belong in the R. leguminosarum bv trifolii group. Further differentiation of Rhizobium strains effective on T. semipilosum was reflected in the broader metabolic profile observed using the BIOLOG MicroPlate ^TM system to evaluate carbon utilization. This revised version was published online in August 2006 with corrections to the Cover Date.     

豆链格孢菌对白车轴草生理代谢及活性氧清除系统酶的影响

通过盆栽实验,研究了接种豆链格孢菌对白车轴草叶组织细胞膜透性、色素含量、丙二醛(MDA)含量以及超氧化物歧化酶(SOD)、过氧化物酶(POD)和过氧化氢酶(CAT)活性的影响.结果表明,豆链格孢菌使白车轴草叶组织细胞膜透性上升,电导率显著升高,膜脂过氧化加强,MDA水平上升,色素含量下降.电导率和MDA含量均与感病时间呈显著正相关,而色素含量与感病时间呈显著负相关.豆链格孢菌刺激其寄主白车轴草叶片膜脂过氧化作用发生,积累大量的活性氧,使其超出防御酶的清除能力,致使活性氧清除系统遭到破坏,保护酶系统失衡,其中SOD和CAT活性显著下降,而POD活性明显上升.接种12d后,SOD和CAT分别降低了55.2%和37.8%,而POD比对照升高约1.6倍.     

茶园间作不同绿肥对节肢动物群落结构和多样性的影响

【目的】研究在茶园中合理间作绿肥对茶园节肢动物群落结构和多样性的影响。【方法】本文在茶园中分别设间作铺地木兰+罗顿豆、圆叶决明+白三叶、白三叶+平托花生3种不同绿肥组合处理,并设不间作绿肥、常规除草的茶园为对照。【结果】间作不同绿肥均能提高茶园节肢动物群落的物种丰富度和群落多样性,提高茶园捕食性天敌昆虫、蜘蛛和寄生性天敌在茶园冠层群落的比率;不同处理茶园冠层节肢动物Shannon-wiener多样性指数(H)、Simpson指数(D)和均匀度指数变化趋势基本一致;茶冠层的节肢动物群落多样性指数在4类不同处理茶园间差异显著;间作绿肥增加了茶园的生物多样性,4类不同处理茶园茶冠层群落相似性系数在0.741~0.892之间,表明4类不同处理茶园冠层群落在物种组成上有较高的相似性。4类不同处理茶园害虫与天敌两类群功能团的丰盛度之间成显著的正相关,表明天敌对害虫的空间数量跟随效应强,主要表现为二者数量间的相互作用。灰色关联度分析表明在4类不同处理茶园中的群落节肢动物物种数/个体数、天敌物种数/害虫物种数、害虫丰富度与多样性指数和均匀度指数的关联系数较高,反应了群落的多样性和稳定性。【结论】间作增强了茶园群落的生物多样性,提高害虫天敌的种类和个体数,有利有效发挥天敌对有害生物的生态控制。