Grass Eragrostis Curvula Research Articles

Differential Methylation Patterns in Apomictic vs. Sexual Genotypes of the Diplosporous Grass Eragrostis curvula.

DNA methylation is an epigenetic mechanism by which a methyl group is added to a cytosine or an adenine. When located in a gene/regulatory sequence it may repress or de-repress genes, depending on the context and species. Eragrostis curvula is an apomictic grass in which facultative genotypes increases the frequency of sexual pistils triggered by epigenetic mechanisms. The aim of the present study was to look for correlations between the reproductive mode and specific methylated genes or genomic regions. To do so, plants with contrasting reproductive modes were investigated through MCSeEd (Methylation Context Sensitive Enzyme ddRad) showing higher levels of DNA methylation in apomictic genotypes. Moreover, an increased proportion of differentially methylated positions over the regulatory regions were observed, suggesting its possible role in regulation of gene expression. Interestingly, the methylation pathway was also found to be self-regulated since two of the main genes (ROS1 and ROS4), involved in de-methylation, were found differentially methylated between genotypes with different reproductive behavior. Moreover, this work allowed us to detect several genes regulated by methylation that were previously found as differentially expressed in the comparisons between apomictic and sexual genotypes, linking DNA methylation to differences in reproductive mode.

Genes Modulating the Increase in Sexuality in the Facultative Diplosporous Grass Eragrostis curvula under Water Stress Conditions.

Eragrostis curvula presents mainly facultative genotypes that reproduce by diplosporous apomixis, retaining a percentage of sexual pistils that increase under drought and other stressful situations, indicating that some regulators activated by stress could be affecting the apomixis/sexual switch. Water stress experiments were performed in order to associate the increase in sexual embryo sacs with the differential expression of genes in a facultative apomictic cultivar using cytoembryology and RNA sequencing. The percentage of sexual embryo sacs increased from 4 to 24% and 501 out of the 201,011 transcripts were differentially expressed (DE) between control and stressed plants. DE transcripts were compared with previous transcriptomes where apomictic and sexual genotypes were contrasted. The results point as candidates to transcripts related to methylation, ubiquitination, hormone and signal transduction pathways, transcription regulation and cell wall biosynthesis, some acting as a general response to stress and some that are specific to the reproductive mode. We suggest that a DNA glycosylase EcROS1-like could be demethylating, thus de-repressing a gene or genes involved in the sexuality pathways. Many of the other DE transcripts could be part of a complex mechanism that regulates apomixis and sexuality in this grass, the ones in the intersection between control/stress and apo/sex being the strongest candidates.

A High-Density Linkage Map of the Forage Grass Eragrostis curvula and Localization of the Diplospory Locus.

Eragrostis curvula (Schrad.) Nees (weeping lovegrass) is an apomictic species native to Southern Africa that is used as forage grass in semiarid regions of Argentina. Apomixis is a mechanism for clonal propagation through seeds that involves the avoidance of meiosis to generate an unreduced embryo sac (apomeiosis), parthenogenesis, and viable endosperm formation in a fertilization-dependent or -independent manner. Here, we constructed the first saturated linkage map of tetraploid E. curvula using both traditional (AFLP and SSR) and high-throughput molecular markers (GBS-SNP) and identified the locus controlling diplospory. We also identified putative regulatory regions affecting the expressivity of this trait and syntenic relationships with genomes of other grass species. We obtained a tetraploid mapping population from a cross between a full sexual genotype (OTA-S) with a facultative apomictic individual of cv. Don Walter. Phenotypic characterization of F1 hybrids by cytoembryological analysis yielded a 1:1 ratio of apomictic vs. sexual plants (34:27, X2 = 0.37), which agrees with the model of inheritance of a single dominant genetic factor. The final number of markers was 1,114 for OTA-S and 2,019 for Don Walter. These markers were distributed into 40 linkage groups per parental genotype, which is consistent with the number of E. curvula chromosomes (containing 2 to 123 markers per linkage group). The total length of the OTA-S map was 1,335 cM, with an average marker density of 1.22 cM per marker. The Don Walter map was 1,976.2 cM, with an average marker density of 0.98 cM/marker. The locus responsible for diplospory was mapped on Don Walter linkage group 3, with other 65 markers. QTL analyses of the expressivity of diplospory in the F1 hybrids revealed the presence of two main QTLs, located 3.27 and 15 cM from the diplospory locus. Both QTLs explained 28.6% of phenotypic variation. Syntenic analysis allowed us to establish the groups of homologs/homeologs for each linkage map. The genetic linkage map reported in this study, the first such map for E. curvula, is the most saturated map for the genus Eragrostis and one of the most saturated maps for a polyploid forage grass species.

Temporal and spatial expression of genes involved in DNA methylation during reproductive development of sexual and apomictic Eragrostis curvula

Recent reports in model plant species have highlighted a role for DNA methylation pathways in the regulation of the somatic-to-reproductive transition in the ovule, suggesting that apomixis (asexual reproduction through seeds) likely relies on RdDM downregulation. Our aim was therefore to explore this hypothesis by characterizing genes involved in DNA methylation in the apomictic grass Eragrostis curvula. We explored floral transcriptomes to identify homologs of three candidate genes, for which mutations in Arabidopsis and maize mimic apomixis (AtAGO9/ZmAGO104, AtCMT3/ZmDMT102/ZmDMT105, and AtDDM1/ZmCHR106), and compared both their spatial and temporal expression patterns during reproduction in sexual and apomictic genotypes. Quantitative expression analyses revealed contrasting expression patterns for the three genes in apomictic vs sexual plants. In situ hybridization corroborated these results for two candidates, EcAGO104 and EcDMT102, and revealed an unexpected ectopic pattern for the AGO gene during germ line differentiation in apomicts. Although our data partially support previous results obtained in sexual plant models, they suggest that rather than an RdDM breakdown in the ovule, altered localization of AtAGO9/ZmAGO104 expression is required for achieving diplospory in E. curvula. The differences in the RdDM machinery acquired during plant evolution might have promoted the emergence of the numerous apomictic paths observed in plants.

Identification of Candidate Genes Related to Polyploidy and/or Apomixis in <i>Eragrostis curvula</i>

This work was aimed at identifying genes that show altered expression profiles in response to changes in ploidy and/or reproductive mode (from sexual to apomictic) in the African grass Eragrostis curvula. A differential display analysis was performed on leaf and flower transcriptomes from a series of genetically related euploid plants, including tetraploid apomictic, diploid sexual, and tetraploid sexual plants. More than 100 primer combinations were used to generate 11,864 total markers, yielding 1293 differential bands. Of these bands, 11.84% to 6.74% were related to ploidy and 0.71% to 2.17% to the reproductive mode, depending on the tissue. A small percentage of bands showed similar expressions between the tetraploid apomictic and the diploid sexual plants. Expression-based similarity dendrograms were constructed. Our data suggested that ploidy is more decisive than tissue type in defining the transcriptome structure. Out of 102 fragments sequenced, 50 showed strong homology to known genes. The differentially expressed genes were mapped in silico onto maize chromosomes. Several candidates mapped within the linkage group syntenic to the Tripsacum dactyloides diplospory-governing region. The evidence indicates that expression of genes located around the diplospory-associated region may be strongly influenced by ploidy and may be silenced in the apomictic genotype. These findings are discussed in the context of diplospory molecular control and its connection with ploidy.

Novel genotypes of the subtropical grass Eragrostis curvula for the study of apomixis (diplospory)

The aim of this work was to obtain a series of genetically related lines of Eragrostis curvula with different ploidy levels and reproductive modes, which can be used for the discovery of genes associated to diplospory. E. curvula is a widely cultivated forage grass native to Southern Africa that reproduces naturally by obligate diplosporous apomixis (asexual reproduction by seeds) of the Antennaria type. In vitro culture of immature inflorescences of the apomictic cultivar Tanganyika led to the regeneration of a diploid plant with a particular morphology (plant UNST1122). Embryo sac analysis and progeny tests determined that plant UNST1122 reproduces by sexuality. UNST1122 was cloned by dividing its tillers and a R1 generation was obtained from seeds after open pollination among the diploid clones. A set of 500 seeds from a diploid R1 plant was treated with colchicine and two plants showing a duplicated chromosome number (2n = 4x = 40) were obtained (plants UNST1112 and UNST1131). Progeny tests using RAPDs indicated that these plants reproduce also by sexuality. The availability of sexual tetraploid genotypes of E. curvula will allow the production of hybrids with new interesting combinations of agronomic traits and the developing of mapping populations segregating for diplospory. Besides, these genetically related plants with different reproduction modes constitutes an excellent model for the identification of gen(es) involved in diplosporous apomixis by mRNA profiling as well as to study the genomic rearrangements and gene expression alterations associated to changes at ploidy levels.

Retention of mobile water during dehydration in the desiccation‐tolerant grass Eragrostis nindensis

Leaf tensile strength was measured for the drought‐tolerant grass Eragrostis curvula and the desiccation‐tolerant grass E. nindensis when fully hydrated, partially dehydrated, naturally air‐dried, and flash‐dried. Leaf tensile strength increased in intact, air‐dried leaves of E. curvula but not for similarly treated leaves of E. nindensis. Examination of leaf cross‐sections by light microscopy and histochemical staining for lignins failed to show any significant structural differences between the two species in the hydrated state. When leaves were flash‐dried, the tensile strength of E. curvula remained unchanged from leaves dried naturally, while there was a marked increase in the tensile strength of flash‐dried leaves of E. nindensis. Proton NMR indicated that the desiccation‐tolerant E. nindensis retained mobile water when leaf relative water content was less than 20% if dried naturally but not if flash‐dried, whereas no mobile water was detected in leaves of E. curvula when dried either naturally or with flash‐drying to below 20% relative water content. This behaviour suggests a fundamental difference in strategy for surviving water loss in vegetative tissues between desiccation‐tolerant species and drought‐tolerant species.

Development of plant regeneration and transformation protocols for the desiccation-sensitive weeping lovegrass Eragrostis curvula

A tissue culture protocol, suitable for transformation, was established for the pasture grass Eragrostis curvula. Callus was generated in the dark from leaf and seed tissues on a medium comprising MS salts supplemented with 2 mg/l 2,4-D, 0.01 mg/l BAP and 2% sucrose. Plant regeneration occurred via organogenesis on the same medium with 6% and 3% sucrose for shoot and root formation, respectively. Optimal regeneration (50 plantlets per callus) occurred when light of 45 micromol/m2 per s was used. The yeast Saccharomyces cerevisiae Hsp12 gene was cloned into the Sac1 of the pCAMBIAUbeeQ vector and callus was transformed by biolistic bombardment. Best transformation (30%) occurred when the target tissue was bombarded twice at a distance of 70 mm using a bombardment pressure of 9,100 kPa. Although successful transformation and transcription of the Hsp12 gene occurred, no Hsp12 protein was found present in tissue extracts of transformed grass.

Photosynthesis and PSII functionality of drought-resistant and drought-sensitive weeping lovegrass plants

World areas subject to drought are expected to increase under conditions of climate change. The purpose of this study is to clarify the response of grass species that can grow and produce under water stress. Therefore leaf photosynthesis, chlorophyll fluorescence and pigment content response to water stress were studied in two varieties of the C 4 grass Eragrostis curvula. Two-year-old plants of cv Ermelo and Consol were grown in plastic pots. Drought stress was imposed by withholding irrigation for 15 days and then rewatering for 5 days. During drought relative water content (RWC) decreased 65% in cv Ermelo, while lower reductions of RWC were observed in cv Consol. During the experiment in cv Ermelo increasing drought stress severity caused large decreases in photosynthetic rates, maximal PSII photochemical efficiency ( F V/ F M) and leaf pigment content. Cv Consol showed small variations in these parameters. Compared to cv Consol, after 15 days of drought, effective PSII quantum yield (Φ II) was significantly lower in cv Ermelo. Reductions of Φ II were related to significant reductions of open PSII energy capture efficiency ( F′ V/ F′ M). Photosynthetic response to increasing PPFD levels and to internal CO 2 concentration ( C i) were reduced by drought in cv Ermelo. Compared to well-watered control plants and to cv Consol, drought stressed plants of cv Ermelo showed also reductions of the initial slope of photosynthetic response to C i and in the photosynthetic rate measured at saturating C i. Moreover stomatal conductance ( g) of both cvs decreased during drought. However, g was lower in drought stressed plants of cv Consol than in cv Ermelo. Water stress caused large reductions in leaf chlorophyll and carotenoid content in cv Ermelo, and small reductions in cv Consol. In drought-sensitive cv Ermelo water stress reduced the capabilities to down regulate PSII functionality through thermal energy dissipation. Results suggest that drought resistance of cv Consol, can be attributed to a higher water use efficiency.

Invasion of alien grass Eragrostis curvula on the gravelly floodplains of the Kinu River and decrease of river endemics : Necessity of urgent measures

Invasion of alien grass Eragrostis curvula on the gravelly floodplains of the Kinu River and decrease of river endemics : Necessity of urgent measures

Dietary shifts: do grazers become browsers in the Thicket Biome?

The diet of buffalo (Syncerus coffer) and Burchell's zebra (Equus burchelli) was investigated in the Addo Elephant National Park to test the hypothesis that the shortage of grass associated with thicket vegetation results in grazers increasing the proportion of dicotyledonous plant species in their diet. Diet composition, estimated from faecal analysis, indicated that 19 and 21 plant species were identified in the faeces of zebra and buffalo, respectively. Buffalo proved to have a higher percentage dicotyledonous (28.1 vs. 8.5 ) and lower percentage monocotyledonous (71.9 vs. 91.5 ) species in their diet than zebra. The grass Eragrostis curvula was found to be the dominant food item for both buffalo (18 of diet) and zebra (39 of diet) during the period of study. The present study indicated that a great proportion of the buffalo diet does in fact consist of grass, thereby refuting the perception that the Addo buffalo are primarily browsers. Both buffalo and zebra are specialised grazers and unsuited to a diet of browse. The probability of dietary shift by these grazers in the grass limited thicket vegetation is therefore challenged.

Whole Plant Carbon Gain of an Endangered Herbaceous Species Aster kantoensis and the Influence of Shading by an Alien Grass Eragrostis curvula in its Gravelly Floodplain Habitat

Aster kantoensis, an endangered plant species, is endemic to gravelly floodplains of a few large rivers in central Japan. In recent years, competitive exclusion by alien perennial grasses in its natural habitat has been suspected to be one of the major factors threatening this species. In the River Kinu, increased shading by the perennial alien grass Eragrostis curvula reduces light availability for A. kantoensis. To reveal the influence of shading on the establishment and growth of A. kantoensis rosettes, the potential carbon gain of A. kantoensis in its natural habitat was estimated using microenvironmental data and whole plant photosynthetic and respiratory responses to light and temperature. Whole plant CO2exchange responses were measured with a specifically designed ‘double chamber’, which enabled measurement of the CO2gas exchange rates of the foliage (F) and the culm (C; stem and roots) separately. It was demonstrated that in a plant with average C/F ratio, positive carbon gain could be maintained only in the microsites where the relative PPFD (photosynthetically active photon flux density) was above 15 or 30% of unshaded conditions in early- or mid-summer, respectively. Increasing C/F ratio, caused by an increase in root biomass as an adaptive response to drought, resulted in a large reduction in the carbon gain irrespective of microsite, weather and season. The high light requirement of A. kantoensis is interpreted as a cost of the morphological responses necessary to avoid stresses characteristic of this gravelly floodplain habitat.

Tillering responses to red:far-red light ratio during different phenological stages in Eragrostis curvula

Tillering responses to light quality in different phenological stages of a perennial warm-season grass Eragrostis curvula were investigated in controlled environments. In vegetative plants, the tillering rate was greater ( P<0.01) in the high (1.1–1.3) than in the low (0.59–0.70) red:far-red ratio (R/FR) light regime. Tillering rates were higher in the low R/FR treatment when the plants in the high R/FR regime reached the reproductive stage, while the plants in the low R/FR regime remained vegetative. After the reproductive tillers were removed by defoliation, more tillers were produced in the defoliated plants grown in the high R/FR regime. When the plants in both light treatments entered the reproductive stage, the tillering rate under the two light regimes became similar, suggesting a significant interaction between tillering and inflorescence development. The more advanced inflorescence development in the high R/FR regime may have reduced assimilate availability to tiller growth and overshadowed the effect of high R/FR on tillering. Both tillering and inflorescence development appeared to be controlled by R/FR ratio. The higher rate of aerial tiller production on the reproductive culms during the post-anthesis period in the high R/FR regime suggested that high R/FR ratio stimulated not only basal tillering, but also aerial tillering.

Seed Production of Species in the Highveld Secondary Succession

After cessation of cropping in the Trachypogon-other species grassland of the South African Highveld, the usual secondary succession was described as follows by Rose-Innes (1939). (1) The ruderal or weed stage, dominated by species such as Erigeron canadensis.t Grasses such as Eleusine indica can dominate with high soil fertility (Davidson 1962). (2) The primary grass stage, usually dominant 2-3 years after abandonment. Characteristic species are the tussock grass Eragrostis curvula and the stoloniferous Cynodon dactylon. (3) The secondary grass stage, usually dominant some 10 years after abandonment. The characteristic species is the tussock grass Hyparrhenia hirta. (4) The sub-climax grassland. Rose-Innes suggested that after 30 or so years the seral areas would have a botanical composition similar to undisturbed Trachypogon-other species grassland. The undisturbed grassland will be referred to as sub-climax grassland, a term used by previous workers (e.g. Davidson 1962). Characteristic species are T. spicatus and Tristachya hispida. In almost all seral communities there are some species from each of the first three stages, although one particular stage may be dominant. Seral grasses are also found scattered throughout sub-climax grassland. Studies by Rose-Innes (1939) showed that seed viability of species in the Highveld succession declined as succession advanced, but no studies have been made of the seed output of different seral stages of the Highveld succession. Salisbury (1942) showed that in general seed output of dicotyledons decreased as species came from later seral stages, but no reports have been encountered which have followed seed output through succession in one particular locality. Investigations of seed production were made during the 1961-62 growing season on the Frankenwald Research Station of the University of the Witwatersrand. The station is on latitude 26?S and longitude 28?E. The soils are acid and light textured. The annual rainfall is approximately 30 in. (760 mm) per year and is virtually confined to the summer months.