Physiological and metabolomic responses of bermudagrass (Cynodon dactylon) to alkali stress

Bermudagrass (Cynodon dactylon) is a widely used warm‐season turfgrass species with superior stress tolerance except for cold. In this study, a comparative analysis of the responses to alkali stress in bermudagrass at the physiological and metabolomic levels were performed. Mild alkali with relatively low pH slightly inhibited growth of bermudagrass as evidenced by lower electrolyte leakage, more rapid growth and higher survival rate when compared to moderate and severe alkali treatments. Moreover, the amount of 37 metabolites including amino acids, organic acids, sugars and sugar alcohols were modulated by the alkali treatments. Among them, 15 metabolites were involved in carbon and amino acid metabolic pathways. Under mild alkali stress, bermudagrass possibly slowed down metabolisms to maintain basic growth. However, moderate and severe alkali‐stressed plants accumulated significantly higher amount of carbohydrates which might result in carbon starvation. Taken together, alkali stress had severely inhibitory effect partially due to combined ionic stress and high pH stress. These results suggested that bermudagrass employed different strategies in response to alkali stresses with different pH and ionic values.     

三峡库区狗牙根对深淹胁迫的生理响应

选取三峡库区自然消落带野生狗牙根(XC)和非消落带野生狗牙根(FC)为研究对象,采用三重复裂区设计试验,主区为两品种,副区为6个不同深度水淹处理(0,1,2,5,10m和15m),分析了各处理植株几种酶活性和碳水化合物含量的变化情况.结果显示:①不同生境狗牙根受深淹胁迫后,丙二醛(MDA)都呈递增趋势,表明狗牙根在深淹胁迫下受到了不同程度的膜脂过氧化伤害,且随胁迫程度的增加受伤害增大.②超氧化物歧化酶(SOD)、过氧化物酶(POD)、谷胱甘肽还原酶(GR)活性和醇脱氢酶(ADH)活性较对照都有所增加;XC处理POD和SOD的最大值出现在5m深水淹处理,而FC出现在2m深处理,且XC 5米深淹处理GR值较对照和其他处理明显大;各受淹处理地下茎可溶性糖含量和淀粉含量都保持在较高水平.③从抗氧化酶活性和ADH活性的变化规律,以及淹水胁迫下植株的能量代谢情况,初步得出XC与FC都具有一定的耐淹性,且XC较FC具更耐深淹的能力,表明狗牙根作为禾本科植物遗传上具有一定的耐淹性,自然消落带狗牙根因长期生长于水淹胁迫环境,耐淹能力得到了进一步强化.从狗牙根对深淹胁迫的生理响应上,证明狗牙根是宜用于三峡水库消落带植被恢复的物种.     

克隆整合有助于狗牙根抵御水淹

尽管国内外开展了大量的克隆整合对克隆植物抵御逆境能力影响的研究, 但整合对植物抵御水淹能力的影响研究仍比较缺乏。该文从克隆整合的角度探讨多年生草本植物狗牙根(Cynodon dactylon)对水淹胁迫的响应。试验模拟了先端分株(相对年幼的分株)分别处于0、5和15 cm三种水淹胁迫环境, 并在每个水淹梯度下实施先端分株与基端分株(相对年长的分株)之间匍匐茎连接或切断处理, 调查水淹一个月后基端分株和先端分株以及整个克隆片段在形态和生理上的表现。研究发现: 切断匍匐茎连接显著降低了狗牙根先端分株的生长, 表现在生物量下降、匍匐茎长度减短和分株数减少等方面; 水淹显著抑制了先端分株的生长, 但对基端分株的生长并未造成显著影响; 在5 cm水淹处理下, 匍匐茎保持连接时, 先端分株和整个克隆片段的生长显著增加; 连接或切断处理在不同水淹梯度下对匍匐茎平均节间长没有显著影响, 对先端分株或基端分株在光化学转化效率上也未表现显著性差异。结果表明: 克隆整合效应促进了狗牙根在水淹胁迫下分株的生长, 并有助于整个克隆片段抵御水淹胁迫。     

Metabolic acclimation of source and sink tissues to salinity stress in bermudagrass (Cynodon dactylon)

Salinity is one of the major environmental factors affecting plant growth and survival by modifying source and sink relationships at physiological and metabolic levels. Individual metabolite levels and/or ratios in sink and source tissues may reflect the complex interplay of metabolic activities in sink and source tissues at the whole‐plant level. We used a non‐targeted gas chromatography–mass spectrometry (GC‐MS) approach to study sink and source tissue‐specific metabolite levels and ratios from bermudagrass under salinity stress. Shoot growth rate decreased while root growth rate increased which lead to an increased root/shoot growth rate ratio under salt stress. A clear shift in soluble sugars (sucrose, glucose and fructose) and metabolites linked to nitrogen metabolism (glutamate, aspartate and asparagine) in favor of sink roots was observed, when compared with sink and source leaves. The higher shifts in soluble sugars and metabolites linked to nitrogen metabolism in favor of sink roots may contribute to the root sink strength maintenance that facilitated the recovery of the functional equilibrium between shoot and root, allowing the roots to increase competitive ability for below‐ground resource capture. This trait could be considered in breeding programs for increasing salt tolerance, which would help maintain root functioning (i.e. water and nutrient absorption, Na⁺ exclusion) and adaptation to stress.     

狗牙根花序发育过程的形态学观察

对狗牙根(Cynodon dactylon‘C299’)花序发育过程中的形态学变化进行了观察。结果表明,‘C299’花序的整个发育过程可分为8个阶段,即营养生长期、穗轴发生期、苞叶原基分化期、小穗原基分化期、小穗分化期、小花分化期、颖片和内外稃发育期及花药和柱头形成期。其中,穗轴发生期(直立茎上有6~9片叶)是抑制花序形成和决定种子产量的关键时期。     

Physiological integration enhanced the tolerance of Cynodon dactylon to flooding

Many flooding‐tolerant species are clonal plants; however, the effects of physiological integration on plant responses to flooding have received limited attention. We hypothesise that flooding can trigger changes in metabolism of carbohydrates and ROS (reactive oxygen species) in clonal plants, and that physiological integration can ameliorate the adverse effects of stress, subsequently restoring the growth of flooded ramets. In the present study, we conducted a factorial experiment combining flooding to apical ramets and stolon severing (preventing physiological integration) between apical and basal ramets of Cynodon dactylon, which is a stoloniferous perennial grass with considerable flooding tolerance. Flooding‐induced responses including decreased root biomass, accumulation of soluble sugar and starch, as well as increased activity of superoxide dismutase (SOD) and ascorbate peroxidase (APX) in apical ramets. Physiological integration relieved growth inhibition, carbohydrate accumulation and induction of antioxidant enzyme activity in stressed ramets, as expected, without any observable cost in unstressed ramets. We speculate that relief of flooding stress in clonal plants may rely on oxidising power and electron acceptors transferred between ramets through physiological integration.     

基于SRAP和TRAP标记的河北野生狗牙根种质遗传多样性分析

以SRAP和TRAP 2种标记技术对36份狗牙根材料的遗传多样性及亲缘关系进行了分析,其中包含34份河北省野生狗牙根种质资源。分别由238对SRAP和85对TRAP引物组合中筛选获得具有多态性的SRAP和TRAP引物组合各10对,PCR扩增总条带分别为186和161条,多态性条带156和132条,平均每对引物扩增出多态性条带各15.6和13.2条,多态性位点比率分别为83.4%和81.0%。2种标记合并进行聚类分析,所有供试的36份狗牙根材料遗传相似系数GS=0.519~0.983,平均为0.7。当GS=0.68时,可将36份供试材料分为4个类群。本研究结果表明河北野生狗牙根种质资源存在较丰富的遗传多样性,可为种质资源保护和选育优良狗牙根新品种提供科学依据。     

Nitrogen fertilization has minimal influence on rhizosphere effects of smooth crabgrass (Digitaria ischaemum) and bermudagrass (Cynodon dactylon)

Aims Plants generally respond to nitrogen (N) fertilization with increased growth, but N addition can also suppress rhizosphere effects, which consequently alters soil processes. We quantified the influence of N addition on rhizosphere effects of two C₄ grasses: smooth crabgrass (Digitaria ischaemum) and bermudagrass (Cynodon dactylon).Methods Plants were grown in nutrient-poor soil for 80 days with either 20 or 120 μg NH 4 NO 3 -N g dry soil^-1. N mineralization rates, microbial biomass, extracellular enzyme activities and bacterial community structure were measured on both rhizosphere and bulk (unplanted) soils after plant harvest.Important findings Fertilization showed nominal differences in net N mineralization, extracellular enzyme activity and microbial biomass between the rhizosphere and bulk soils, indicating minimal influence of N on rhizosphere effects. Instead, the presence of plant roots showed the strongest impact (up to 80%) on rates of net N mineralization and activities of three soil enzymes indicative of N release from organic matter. Principal component analysis of terminal restriction fragment length polymorphism (T-RFLP) also reflected these trends by highlighting the importance of plant roots in structuring the soil bacterial community, followed by plant species and N fertilization (to a minor extent). Overall, the results indicate minor contributions of short-term N fertilization to changes in the magnitude of rhizosphere effects for both grass species.     

Analysis of Natural Variation in Bermudagrass (Cynodon dactylon) Reveals Physiological Responses Underlying Drought Tolerance

Bermudagrass (Cynodon dactylon) is a widely used warm-season turfgrass and one of the most drought tolerant species. Dissecting the natural variation in drought tolerance and physiological responses will bring us powerful basis and novel insight for plant breeding. In the present study, we evaluated the natural variation of drought tolerance among nine bermudagrass varieties by measuring physiological responses after drought stress treatment through withholding water. Three groups differing in drought tolerance were identified, including two tolerant, five moderately tolerant and two susceptible varieties. Under drought stress condition, drought sensitive variety (Yukon) showed relative higher water loss, more severe cell membrane damage (EL), and more accumulation of hydrogen peroxide (H₂O₂) and malondialdehyde (MDA), while drought tolerant variety (Tifgreen) exhibited significantly higher antioxidant enzymes activities. Further results indicated that drought induced cell injury in different varieties (Yukon, SR9554 and Tifgreen) exhibited liner correlation with leaf water content (LWC), H₂O₂ content, MDA content and antioxidant enzyme activities. Additionally, Tifgreen plants had significantly higher levels of osmolytes (proline level and soluble sugars) when compared with Yukon and SR9554 under drought stress condition. Taken together, our results indicated that natural variation of drought stress tolerance in bermudagrass varieties might be largely related to the induced changes of water status, osmolyte accumulation and antioxidant defense system.     

水淹对狗牙根营养繁殖植株的生理生态学效应

通过控制实验,测定了经过水淹处理的狗牙根营养繁殖体在恢复阶段的光合作用及其相关的生理生化指标的变化。结果显示,水淹时间对恢复阶段营养繁殖体的蒸腾作用和叶片温度的影响达到显著水平,水淹深度对该时期营养繁殖体的光合作用、气孔导度、胞间二氧化碳浓度和叶片温度有显著影响。水淹还导致了恢复期间植株叶片光合色素含量的显著变化。经过水淹的植株的各类光合色素含量以及色素总含量都显著高于对照植株,其中全淹处理的植株显著高于半淹处理的植株,叶绿素a与叶绿素b的比例也是全淹处理的植株显著高于半淹处理的植株。结果表明狗牙根营养繁殖体具有较强的恢复生长和生理活动的能力,是一种适宜于水电工程库区消落带生态恢复的物种。     

Effects of temperature and photoperiod on postponing bermudagrass (Cynodon dactylon [L.] Pers.) turf dormancy

Growth chamber and field experiments were carried out to determine the effects of extended photoperiod under low and freezing temperatures on bermudagrass turf dormancy at Bajgah, in the southern part of Iran. The experiment in the growth chamber was conducted with four temperature regimes (15, 7.5, 0 and -7.5°C) and three light durations (8, 12 and 16h) in a completely randomized design with four replications. The field study was conducted in two consecutive years (2008-2009) with three light durations (8, 12 and 16h) in months with natural short day length and arranged in a randomized complete blocks design with three replications. Results in both experiments showed that decreasing temperature and photoperiod decreased verdure fresh and dry weight, shoot height, tiller density, leaf area and chlorophyll and relative water contents (RWC). However, rooting depth and fresh weight of roots increased in the growth chamber. Decreasing the temperature and light duration increased electrolyte leakage and proline content. Reducing sugars increased with decreasing temperature and declined with lowering light duration in both shoots and roots. Starch content of both shoots and roots showed an adverse trend compared to reducing sugars; starch content increased in both shoots and roots in all treatments by shortening the photoperiod. Practically, the problem of bermudagrass turf's dormancy could be solved via increasing the photoperiod in months with short day lengths. This treatment would be efficient and useful for turfgrass managers to apply in landscapes and stadiums.     

Genetic diversity analysis of Cynodon dactylon (bermudagrass) accessions and cultivars from different countries based on ISSR and SSR markers

ISSR and SSR markers were used to evaluate genetic diversity among 33 Cynodon dactylon accessions and 22 cultivars from four different countries in order to provide information on how to improve the utilization of bermudagrass germplasms. Eighty eight bands were amplified by nine SSR primer combinations and 236 bands were observed from 23 ISSR primers. The results showed that 97.7% of the SSR primers and 86.9% of the ISSR primers were polymorphic. The genetic similarity coefficients (GSC), gene diversity (He) and Shannon index (I) were 0.58–0.97, 0.27 and 0.41, respectively, for ISSR and 0.52–0.97, 0.29, and 0.43 for SSR. The UPGMA analysis clustered the 55 accessions (cultivars) into three groups. The cluster results produced by the ISSR data were close to the SSR data results. Analysis based on the combined ISSR and SSR data was more closely related to the geographical distribution of the tested germplasm.     

Growth promotion effect of Fusarium spp. PPF1 from bermudagrass (Cynodon dactylon) rhizosphere on Indian spinach (Basella alba) seedlings are linked to root colonisation

A rhizosphere fungus was isolated from roots of bermudagrass (Cynodon dactylon) and identified as Fusarium spp. PPF1. A pot experiment was conducted to test its ability to promote the vegetative growth of Indian spinach seedlings (Basella alba). Indian spinach seeds were sown in sterilised field soil amended with wheat grain inoculum of PPF1 at the rate of 0.5 and 1.0% w/w, and plants were grown for 21?days in a net house. Significantly, higher germination percentage and vigour index were observed due to application of PPF1 in the potting soil. Treatment with PPF1 also significantly increased shoot length, shoot fresh weight, shoot dry weight, root length, root fresh weight, root dry weight, leaf area and leaf chlorophyll content of cucumber plants compared to non-treated control. The growth promotion rate increased with the increasing concentration of inoculum of PPF1 applied to the soil. The fungus was re-isolated from the roots of cucumber plants at higher frequencies, while a positive co-relation was found between the root colonisation ability and the plant growth enhancement by the isolate. These results suggest that growth promotion effect of Fusarium spp. PPF1 on Indian spinach (B. alba) are linked to root colonisation ability of the fungus.     

Comparative proteomic and metabolomic analyses reveal mechanisms of improved cold stress tolerance in bermudagrass (Cynodon dactylon (L.) Pers.) by exogenous calcium

As an important second messenger, calcium is involved in plant cold stress response, including chilling (<20 °C) and freezing (<0 °C). In this study, exogenous application of calcium chloride (CaCl₂) improved both chilling and freezing stress tolerances, while ethylene glycol‐bis‐(β‐aminoethyl) ether‐N,N,N,N‐tetraacetic acid (EGTA) reversed CaCl₂ effects in bermudagrass (Cynodon dactylon (L.) Pers.). Physiological analyses showed that CaCl₂ treatment alleviated the reactive oxygen species (ROS) burst and cell damage triggered by chilling stress, via activating antioxidant enzymes, non‐enzymatic glutathione antioxidant pool, while EGTA treatment had the opposite effects. Additionally, comparative proteomic analysis identified 51 differentially expressed proteins that were enriched in redox, tricarboxylicacid cycle, glycolysis, photosynthesis, oxidative pentose phosphate pathway, and amino acid metabolisms. Consistently, 42 metabolites including amino acids, organic acids, sugars, and sugar alcohols were regulated by CaCl₂ treatment under control and cold stress conditions, further confirming the common modulation of CaCl₂ treatment in carbon metabolites and amino acid metabolism. Taken together, this study reported first evidence of the essential and protective roles of endogenous and exogenous calcium in bermudagrass response to cold stress, partially via activation of the antioxidants and modulation of several differentially expressed proteins and metabolic homeostasis in the process of cold acclimation.     

Herbicide Tolerance and Polyploidy in Cynodon dactylon (L.) Pers. (Gramineae)

Responses to treatment with the herbicide dalapon have beendetermined and the karyotypes studied in four chromosome racesof Cynodon dactylon. No relationship was found between toleranceto dalapon and the level of ploidy. At the dalapon concentrationwhere differences in response were most apparent (2·5mM), the difference in mean value of the two diploid cloneswas greater than any other pair of values. No positive correlationwas found between tolerance and the morphology of the leaf surface.Karyotype studies have not resolved the conflicting evidencerelating to the origin and inter-relationships of the chromosomeraces of this species.     

Differentiation of bermudagrass (Cynodon spp.) genotypes by AFLP analyses

 Bermudagrasses (Cynodon spp.) are major turfgrasses for home lawns, public parks, golf courses and sport fields, and are widely adapted to tropical and warmer temperate climates. Morphological and physiological characteristics are not sufficient to differentiate some bermudagrass genotypes because the differences between them are often subtle and subject to environmental influence. In this study, a DNA-typing technique, amplified fragment length polymorphism (AFLP), was used to differentiate bermudagrass genotypes and to explore their genetic relationships. Twenty seven bermudagrass cultivars and introductions, mostly from the Coastal Plain Experiment Station in Tifton, Ga., were assayed by the radioactive (³²P) and the fluorescence-labeled AFLP methods. The AFLP technique produced enough polymorphism to differentiate all 27 bermudagrass genotypes, even the closely related ones. An average of 48–74 bands in the 30–600-bp size range was detected by the ³²P-labeled AFLP method. The results indicated that most of the 14 primer combinations tested in this study could be used to distinguish bermudagrass genotypes, and that some single primer-pairs could differentiate all 27 of them. To test the reliability and reproducibility of the AFLP procedure, three DNA isolations (replications) of the 27 bermudagrass genotypes were assayed using five primer pairs. Only 0.6% of the bands were evaluated differently among the three replications. One replication of one genotype (which was most likely a planting contaminant) was grouped in an unexpected cluster using the Unweighted Pair Group Mean Average (UPGMA) method. A one- or two-band difference in scoring did not change the clustering of genotypes or the replications within genotypes. The 27 genotypes were grouped into three major clusters, many of which were in agreement with known pedigrees. Trees constructed with different primer combinations using ³²P- and fluorescence-labelling formed similar major groupings. The semi-automated fluorescence-based AFLP technique offered significant improvements on fragment sizing and data handling. It was also more accurate for detection and more efficient than the radioactive labelling method. This study shows that the AFLP technique is a reliable tool for differentiating bermudagrass genotypes and for determining genetic relationships among them. Received: 28 July 1998 / Accepted: 3 November 1998     

AFLP analysis of Cynodon dactylon (L.) Pers. var. dactylon genetic variation.

Cynodon dactylon (L.) Pers. var. dactylon (common bermudagrass) is geographically widely distributed between about lat 45 degrees N and lat 45 degrees S, penetrating to about lat 53 degrees N in Europe. The extensive variation of morphological and adaptive characteristics of the taxon is substantially documented, but information is lacking on DNA molecular variation in geographically disparate forms. Accordingly, this study was conducted to assess molecular genetic variation and genetic relatedness among 28 C. dactylon var. dactylon accessions originating from 11 countries on 4 continents (Africa, Asia, Australia, and Europe). A fluorescence-labeled amplified fragment length polymorphism (AFLP) DNA profiling method was used to detect the genetic diversity and relatedness. On the basis of 443 polymorphic AFLP fragments from 8 primer combinations, the accessions were grouped into clusters and subclusters associating with their geographic origins. Genetic similarity coefficients (SC) for the 28 accessions ranged from 0.53 to 0.98. Accessions originating from Africa, Australia, Asia, and Europe formed major groupings as indicated by cluster and principal coordinate analysis. Accessions from Australia and Asia, though separately clustered, were relatively closely related and most distantly related to accessions of European origin. African accessions formed two distant clusters and had the greatest variation in genetic relatedness relative to accessions from other geographic regions. Sampling the full extent of genetic variation in C. dactylon var. dactylon would require extensive germplasm collection in the major geographic regions of its distributional range.     

Chromosome Number in Cynodon dactylon in Relation to Ecological Conditions

Populations of Cynodon dactylon (L.) Pers. were collected fromthree habitat types (roadsides, lawns and paddy-fields) withineach of five climatic regions (arid, dry, intermediate, wetand hill country) in Sri Lanka. Further populations were collectedfrom forests and grasslands in the hill country. Most populations contained only tetraploid plants (2n = 36),but populations from roadsides and lawns in the wet region,and from forests in the hill country, contained only diploidplants (2n = 18). Populations from paddy-fields in the wet regioncontained both tetraploid and diploid plants. No triploid plantswere found in any environment, despite the fact that C. dactylonpredominantly reproduces vegetatively by both stolons and rhizomes.Soil acidity seems to be the main factor determining the distributionof the two cytotypes.Copyright 1995, 1999 Academic Press Cynodon dactylon (L.) Pers., chromosome number, ecology, habitat, climatic region