Screening of environmental stimuli for the positive regulation of stomatal aperture in centipedegrass.

Grasslands, the largest carbon pool in China, possess enormous potential for carbon sequestration. Increasing the stomatal aperture to increase the CO2 absorption capacity is a potential method to improve plant photosynthetic efficiency and ultimately enhance the carbon sequestration capacity of grass plants. Research on stomatal aperture regulation has focused mostly on Arabidopsis or crops, while research on grass plants in these areas is scarce, which seriously restricts the implementation of this grassland carbon sequestration strategy. Here, a widely used ecological grass, centipedegrass, was used as the experimental material. First, a convenient method for observing the stomatal aperture was developed. The leaves were floated in a potassium ion-containing open solution (67 mM KCl, pH 6.0) with the adaxial surface rather than the abaxial surface in contact with the solution and were cultivated under light for 1.5 h. Then, nail polish was applied on the adaxial surface, and a large number of open stomata were imprinted. Second, with the help of this improved method, the concentration‒response characteristics of the stomatal aperture to eleven environmental stimuli were tested. The stomatal aperture is dependent on these environmental stimuli in a concentration-dependent manner. The addition of 100 µM brassinolide led to the maximal stomatal aperture. This study provided a technical basis for manipulating stomatal opening to increase the carbon sequestration capacity of centipedegrass.

Physiological and Proteomic Analyses of mtn1 Mutant Reveal Key Players in Centipedegrass Tiller Development.

Tillering directly determines the seed production and propagation capacity of clonal plants. However, the molecular mechanisms involved in the tiller development of clonal plants are still not fully understood. In this study, we conducted a proteome comparison between the tiller buds and stem node of a multiple-tiller mutant mtn1 (more tillering number 1) and a wild type of centipedegrass. The results showed significant increases of 29.03% and 27.89% in the first and secondary tiller numbers, respectively, in the mtn1 mutant compared to the wild type. The photosynthetic rate increased by 31.44%, while the starch, soluble sugar, and sucrose contents in the tiller buds and stem node showed increases of 13.79%, 39.10%, 97.64%, 37.97%, 55.64%, and 7.68%, respectively, compared to the wild type. Two groups comprising 438 and 589 protein species, respectively, were differentially accumulated in the tiller buds and stem node in the mtn1 mutant. Consistent with the physiological characteristics, sucrose and starch metabolism as well as plant hormone signaling were found to be enriched with differentially abundant proteins (DAPs) in the mtn1 mutant. These results revealed that sugars and plant hormones may play important regulatory roles in the tiller development in centipedegrass. These results expanded our understanding of tiller development in clonal plants.

Selection of Suitable Reference Genes for RT-qPCR Gene Expression Analysis in Centipedegrass under Different Abiotic Stress.

As a C4 warm-season turfgrass, centipedegrass (Eremochloa ophiuroides (Munro) Hack.) is known for its exceptional resilience to intensive maintenance practices. In this research, the most stably expressed reference genes in the leaves of centipedegrass under different stress treatments, including salt, cold, drought, aluminum (Al), and herbicide, were screened by the quantitative real-time PCR (RT-qPCR) technique. The stability of 13 candidate reference genes was evaluated by software GeNorm V3.4, NormFinder V20, BestKeeper V1.0, and ReFinder V1.0. The results of this experiment demonstrated that the expression of the UBC (ubiquitin-conjugating enzyme) remained the most stable under cold and Al stress conditions. On the other hand, the MD (malate dehydrogenase) gene exhibited the best performance in leaf tissues subjected to salt and drought stresses. Under herbicide stress, the expression level of the RIP (60S ribosomal protein L2) gene ranked the highest. The expression levels of abiotic stress-associated genes such as PIP1, PAL, COR413, ALMT9, and BAR were assessed to validate the reliability of the selected reference genes. This study provides valuable information and reference points for gene expression under abiotic stress conditions in centipedegrass.

Molecular Mechanism of Cold Tolerance of Centipedegrass Based on the Transcriptome.

Low temperature is an important limiting factor in the environment that affects the distribution, growth and development of warm-season grasses. Transcriptome sequencing has been widely used to mine candidate genes under low-temperature stress and other abiotic stresses. However, the molecular mechanism of centipedegrass in response to low-temperature stress was rarely reported. To understand the molecular mechanism of centipedegrass in response to low-temperature stress, we measured physiological indicators and sequenced the transcriptome of centipedegrass under different stress durations. Under cold stress, the SS content and APX activity of centipedegrass increased while the SOD activity decreased; the CAT activity, POD activity and flavonoid content first increased and then decreased; and the GSH-Px activity first decreased and then increased. Using full-length transcriptome and second-generation sequencing, we obtained 38.76 G subreads. These reads were integrated into 177,178 isoforms, and 885 differentially expressed transcripts were obtained. The expression of AUX_IAA and WRKY transcription factors and HSF transcription-influencing factors increased during cold stress. Through KEGG enrichment analysis, we determined that arginine and proline metabolism, plant circadian rhythm, plant hormone signal transduction and the flavonoid biosynthesis pathways played important roles in the cold stress resistance of centipedegrass. In addition, by using weighted gene coexpression network analysis (WGCNA), we determined that the turquoise module was significantly correlated with SS content and APX activity, while the blue module was significantly negatively correlated with POD and CAT activity. This paper is the first to report the response of centipedegrass to cold stress at the transcriptome level. Our results help to clarify the molecular mechanisms underlying the cold tolerance of warm-season grasses.

An appropriate ammonium: nitrate ratio promotes the growth of centipedegrass: insight from physiological and micromorphological analyses.

Reasonable nitrogen fertilizer application is an important strategy to maintain optimal growth of grasslands, thereby enabling them to better fulfil their ecological functions while reducing environmental pollution caused by high nitrogen fertilizer production and application. Optimizing the ammonium (NH4 +):nitrate (NO3 -) ratio is a common approach for growth promotion in crops and vegetables, but research on this topic in grass plants has not received sufficient attention. Centipedegrass, which is widely used in landscaping and ecological protection, was used as the experimental material. Different NH4 +:NO3 - ratios (0: 100, 25:75, 50:50, 75:25, 100:0) were used as the experimental treatments under hydroponic conditions. By monitoring the physiological and morphological changes under each treatment, the appropriate NH4 +:NO3 - ratio for growth and its underlying mechanism were determined. As the proportion of ammonium increased, the growth showed a "bell-shaped" response, with the maximum biomass and total carbon and nitrogen accumulation achieved with the NH4 +:NO3 - ratio of 50:50 treatment. Compared with the situation where nitrate was supplied alone, increasing the ammonium proportion increased the whole plant biomass by 93.2%, 139.7%, 59.0%, and 30.5%, the whole plant nitrogen accumulation by 44.9%, 94.6%, 32.8%, and 54.8%, and the whole plant carbon accumulation by 90.4%, 139.9%, 58.7%, and 26.6% in order. As a gateway for nitrogen input, the roots treated with an NH4 +:NO3 - ratio of 50:50 exhibited the highest ammonium and nitrate uptake rate, which may be related to the maximum total root length, root surface area, average root diameter, root volume, and largest root xylem vessel. As a gateway for carbon input, leaves treated with an NH4 +:NO3 - ratio of 50:50 exhibited the highest stomatal aperture, stomatal conductance, photosynthetic rate, transpiration rate, and photosynthetic products. The NH4 +:NO3 - ratio of 50:50 treatment had the largest stem xylem vessel area. This structure and force caused by transpiration may synergistically facilitate root-to-shoot nutrient translocation. Notably, the change in stomatal opening occurred in the early stage (4 hours) of the NH4 +:NO3 - ratio treatments, indicating that stomates are structures that are involved in the response to changes in the root NH4 +:NO3 - ratio. In summary, we recommend 50:50 as the appropriate NH4 +:NO3 - ratio for the growth of centipedegrass, which not only improves the nitrogen use efficiency but also enhances the carbon sequestration capacity.

Physiological and Comparative Transcriptome Analyses of the High-Tillering Mutant mtn1 Reveal Regulatory Mechanisms in the Tillering of Centipedegrass (Eremochloa ophiuroides (Munro) Hack.).

Tillering is a key factor that determines the reproductive yields of centipedegrass, which is an important perennial warm-season turfgrass. However, the regulatory mechanism of tillering in perennial plants is poorly understood, especially in perennial turfgrasses. In this study, we created and characterised a cold plasma-mutagenised centipedegrass mutant, mtn1 (more tillering number 1). Phenotypic analysis showed that the mtn1 mutant exhibited high tillering, short internodes, long seeds and a heavy 1000-seed weight. Then, a comparative transcriptomic analysis of the mtn1 mutant and wild-type was performed to explore the molecular mechanisms of centipedegrass tillering. The results revealed that plant hormone signalling pathways, as well as starch and sucrose metabolism, might play important roles in centipedegrass tillering. Hormone and soluble sugar content measurements and exogenous treatment results validated that plant hormones and sugars play important roles in centipedegrass tiller development. In particular, the overexpression of the auxin transporter ATP-binding cassette B 11 (EoABCB11) in Arabidopsis resulted in more branches. Single nucleotide polymorphisms (SNPs) were also identified, which will provide a useful resource for molecular marker-assisted breeding in centipedegrass. According to the physiological characteristics and transcriptional expression levels of the related genes, the regulatory mechanism of centipedegrass tillering was systematically revealed. This research provides a new breeding resource for further studies into the molecular mechanism that regulates tillering in perennial plants and for breeding high-tillering centipedegrass varieties.

Low-molecular-weight organic acid-mediated tolerance and Pb accumulation in centipedegrass under Pb stress.

Lead (Pb) is one of the most harmful, toxic pollutants to the ecological environment and humans. Centipedegrass, a fast-growing warm-season turfgrass, is excellent for Pb pollution remediation. Exogenous low-molecular-weight organic acid (LMWOA) treatment is a promising approach for assisted phytoremediation. However, the effects of this treatment on the tolerance and Pb accumulation of centipedegrass are unclear. This study investigated these effects on the physiological growth response and Pb accumulation distribution characteristics of centipedegrass. Applications of 400 µM citric acid (CA), malic acid (MA) and tartaric acid (TA) significantly reduced membrane lipid peroxidation levels of leaves and improved biomass production of Pb-stressed plants. These treatments mainly increased peroxidase (POD), catalase (CAT) and ascorbate peroxidase (APX) activities and enhanced free protein (Pro), ascorbic acid (AsA) and phytochelatins (PCs) contents, ultimately improving the Pb tolerance of centipedegrass. Their promoting effects decreased as follows: TA>CA>MA. All the treatments decreased root Pb concentrations and increased stem and leaf Pb concentrations, thus increasing total Pb accumulation and TF values. MA had the best and worst effects on Pb accumulation and Pb transportation, respectively. CA had the best and worst effects on Pb transportation and Pb accumulation, respectively. TA exhibited strong effects on both Pb accumulation and transport. Furthermore, all treatments changed the subcellular Pb distribution patterns and distribution models of the chemical forms of Pb in each tissue. The root Pb concentration was more highly correlated with the Pb subcellular fraction distribution pattern, while the stem and leaf Pb concentrations were more highly correlated with the distribution models of the chemical forms of Pb. Overall, TA improved plant Pb tolerance best and promoted both Pb absorption and transportation well and is considered the best candidate for Pb-contaminated soil remediation with centipedegrass. This study provides a new idea for Pb-contaminated soil remediation with centipedegrass combined with LMWOAs.

NTA-assisted mineral element and lead transportation in Eremochloa ophiuroides (Munro) Hack.

Lead (Pb) is one of the most toxic and harmful pollutants to the environment and human health. Centipedegrass (Eremochloa ophiuroides (Munro) Hack.), an excellent ground cover plant for urban plant communities, exhibits the outstanding lead tolerance and accumulation. Nitrilotriacetic acid (NTA) is an environmentally friendly chelating agent that strengthens phytoremediation. This study explored the effects of different NTA concentrations on the absorption and transportation of mineral elements and Pb in centipedegrass. Following exposure to Pb (500 µM) for 7 days in hydroponic nutrient solution, NTA increased root Mg, K, and Ca concentrations and shoot Fe, Cu, and Mg concentrations and significantly enhanced the translocation factors of mineral elements to the shoot. Although NTA notably decreased root Pb absorption and accumulation, it significantly enhanced Pb translocation factors, and the Pb TF value was the highest in the 2.0 mM NTA treatment. Furthermore, the shoot translocation of Pb and mineral elements was synergistic. NTA can support mineral element homeostasis and improve Pb translocation efficiency in centipedegrass. Regarding root radial transport, NTA (2.0 mM) significantly promoted Pb transport by the symplastic pathway under the treatments with low-temperature and metabolic inhibitors. Meanwhile, NTA increased apoplastic Pb transport at medium and high Pb concentrations (200-800 µM). NTA also enhanced the Pb radial transport efficiency in roots and thus assisted Pb translocation. The results of this study elucidate the effects of NTA on the absorption and transportation of mineral elements and Pb in plants and provide a theoretical basis for the practical application of the biodegradable chelating agent NTA in soil Pb remediation.

Use of Transcriptomic Analyses to Elucidate the Mechanism Governing Nodal Root Development in Eremochloa ophiuroides (Munro) Hack.

Centipedegrass [Eremochloa ophiuroides (Munro) Hack.] is a perennial warm-season grass that originated in China, and its speed of nodal rooting is important for lawn establishment. In our study, centipedegrass nodal rooting ability was limited by node aging. Transcriptome sequencing of nodal roots after 0, 2, 4, and 8 days of water culture was performed to investigate the molecular mechanisms of root development. GO enrichment and KEGG pathway analyses of DEGs indicated that plant hormone signal transduction and transcription factors might play important roles in centipedegrass nodal root growth. Among them, E3 ubiquitin-protein ligases participated in multiple hormone signal transduction pathways and interacted with transcription factors. Furthermore, an E3 ubiquitin protein ligase EoSINAT5 overexpressed in rice resulted in longer roots and more numerous root tips, while knockout of LOC_Os07g46560 (the homologous gene of EoSINAT5 in rice) resulted in shorter roots and fewer root tips. These results indicated that EoSINAT5 and its homologous gene are able to promote nodal root development. This research presents the transcriptomic analyses of centipedegrass nodal roots, and may contribute to elucidating the mechanism governing the development of nodal roots and facilitates the use of molecular breeding in improving rooting ability.

Radioprotective effects of centipedegrass extract on NIH-3T3 fibroblasts via anti-oxidative activity.

Centipedegrass originates from China and South America, and has been reported to contain several C-glycosyl flavones and phenolic compounds, including maysin and luteolin. The present study aimed to investigate the radioprotective activity of centipedegrass extract (CGE) in radiation exposed-fibroblasts and to assess the affected molecular pathway. The radioprotective effects of CGE were determined in NIH-3T3 cells using Cell Counting Kit-8 and morphological changes were observed. Reactive oxygen species (ROS) levels and the apoptotic profile of NIH-3T3 cells were also measured. The expression levels of B-cell lymphoma-2 (Bcl-2) family proteins [Bcl-2, Bcl-2 like protein 4 (Bax), Bcl-2-associated death promoter (Bad), caspase-3, poly(ADP-ribose) polymerase (PARP)], AKT and MAPK family proteins (ERK, p38 and JNK) were measured in vitro. The results demonstrated that when 3T3 fibroblasts pretreated with CGE were subjected to H2O2-induced cell damage, their viability was significantly decreased. Additionally, CGE pretreatment decreased ROS levels and the protein expression levels of cleaved PARP upon H2O2 treatment, indicating that CGE induced cytoprotective effects against H2O2-induced oxidative stress. Moreover, significant protective effects of CGE against intracellular ROS, induced upon exposure to ionizing radiation (IR), were observed. The protective effects of CGE pretreatment were also determined by morphological observation of NIH-3T3 cells following exposure to IR. CGE pretreatment increased the expression levels of anti-apoptotic signals (Bcl-2, p-BAD) and decreased the levels of pro-apoptotic signals (Bax, Bad), and led to cleavage of PARP and caspase-3 proteins. Additionally, in cells pretreated with CGE, the phosphorylation of AKT and ERK was increased and that of p38 and JNK was decreased compared with in cells subjected only to IR. These results indicated that CGE may act as a radioprotector due to its anti-oxidative activity, restoring cell homeostasis and redox balance in radiation-exposed fibroblast cells. Therefore, it could be suggested that CGE may be an effective candidate in the treatment of oxidative stress-related diseases and in radioprotection.

Centipedegrass extract enhances radiosensitivity in melanoma cells by inducing G2/M cell cycle phase arrest.

Melanoma is aggressive, highly metastatic, and potentially fatal. In the case of patients with advanced melanoma, it is difficult to expect a good prognosis, since this cancer has low sensitivity to chemotherapy and radiation therapy. The use of natural ingredients may enhance existing therapies. Centipedegrass extract (CGE) which contains phenolic structures and C-glycosyl flavones, has been shown to have anti-inflammatory effects and anti-cancer effects. The purpose of this study was to evaluate the radio sensitizing effects of CGE in combination with ionizing radiation (IR). Two melanoma cell lines were exposed to IR after treatment with CGE at concentrations that were not toxic alone. The effects of CGE + IR on cell survival, cell cycle, and apoptotic cell death were examined using MTT and Muse® Cell Analyzer, and fluorescence microscopy. Molecular signaling mechanisms were explored by western blots. Our findings showed that co-treatment of CGE + IR reduced the survival of melanoma cells more than IR alone. Also, cell cycle arrest in CGE-treated cells was enhanced and these cells became more radiosensitive. CGE + IR increased apoptotic cell death more than IR alone. Western blot results showed that the effect of CGE + IR involved MAPKs (ERK1/2, p38, and JNK) pathway. Our study suggests that CGE + IR treatment enhanced radio-sensitization and cell death of melanoma cells via cell cycle arrest and the MAPKs pathway.

Physiological responses and tolerance mechanisms of seashore paspalum and centipedegrass exposed to osmotic and iso-osmotic salt stresses.

Osmotic stresses caused by reduced water availability or the accumulation of salts in the soil can be highly damaging to plants. The objective of this study was to investigate physiological responses and tolerance mechanisms of two turfgrass species (seashore paspalum and centipedegrass) with distinct differences in salinity tolerance exposed to osmotic and iso-osmotic salt stresses. Three turfgrass genotypes including seashore paspalums 'Seastar' and 'UGP113', and centipedegrass 'TifBlair' were grown in ½ strength Hoagland's solution with three different treatment conditions; control (no external addition), salt stress (-0.4 MPa by adding NaCl) and osmotic stress [-0.4 MPa by adding polyethylene glycol (PEG)]. Osmotic stress damages were more severe with greater reductions in turf quality, photochemical efficiency (Fv/Fm), relative water content (RWC) and leaf water potential (Ψw) compared to iso-osmotic salt stress in both seashore paspalum and centipedegrass. Greater osmotic adjustment (OA) with greater accumulation of metabolically inexpensive inorganic osmolytes (Na+) helped turfgrasses to lessen damages in salt stress compared to osmotic stress. However, such accumulation of Na+ resulted ion-toxicity and triggered some damages in terms of increased electrolyte leakage (EL) and reduced total protein in salt-sensitive centipedegrass. Seashore paspalum had better ion regulation and also maintained greater antioxidant enzyme activities compared to centipedegrass; therefore it was able to avoid ion-specific damages under salt stress. Differences in the utilization of specific solutes for osmotic adjustment and antioxidant metabolism are partially responsible for the differences in salt versus osmotic stress responses in these species; the regulation of these defense mechanisms requires further investigation.

Functional and genomic characterization of a wound- and methyl jasmonate-inducible chalcone isomerase in Eremochloa ophiuroides [Munro] Hack.

Eremochloa ophiuroides, a perennial warm-season lawn grass, has a characteristic phenotype of red pigmentation in tissues during maturation. The putative gene families associated with the red coloration were previously identified in E. ophiuroides. These genes encode chalcone synthases, flavonol 3-hydroxylases, and flavonol 3'-hydroxylases, acting on the early flavonoid-biosynthesis pathway. Here, a type-I chalcone isomerase (CHI) gene was isolated from E. ophiuroides based on leaf-transcriptome data, and the corresponding enzyme was functionally characterized in vitro and in planta. Complementation of Arabidopsis tt5 mutants by overexpressing EoCHI recapitulated the wild-type seed coat color. Wounding and methyl jasmonate treatments significantly elevated the transcript level of EoCHI and total anthocyanin content in shoots. Confocal microscopy indicated the localization of EoCHI to the endoplasmic reticulum. The genomic EoCHI sequence contained two introns with a novel pattern of exon‒intron organization. Further examinations on genomic structures of CHI family from ancient to advanced plant lineages should be of interests to decipher evolutionary pathways of extant plant CHI genes.

Transcriptome-guided identification and functional characterization of key terpene synthases involved in constitutive and methyl jasmonate-inducible volatile terpene formation in Eremochloa ophiuroides (Munro) Hack.

Centipedegrass (Eremochloa ophiuroides [Munro] Hack.) is a warm-season turfgrass, widely planted in residential lawns and recreational fields. Here, we uncovered three major terpenes released from the shoots of Eo: (E)-ß-ocimene (6%), α-muurolene (87.8%), and eremophilene (6.2%). Methyl jasmonate (MeJA) treatment increased the emission of monoterpenes, including (E)- and (Z)-ß-ocimene, limonene, and myrcene, as well as sesquiterpene blends of (E)-caryophyllene, α-copaene, (+)-cyclosativene, and α-farnesene. RNA sequencing analysis predicted 14 putative Eo terpene synthase (EoTPS) genes, and two full-length EoTPS were successfully amplified: Eo7816 (1722 bp) and Eo6039 (1701 bp). Phylogenetic analysis revealed that Eo7816 and Eo6039 belonged to the clades of TPS-b and TPS-a, respectively. The Arabidopsis transgenic plants overexpressing Eo7816 exclusively released (E)-ß-ocimene (96%) with (Z)-ß-ocimene and myrcene. In contrast, Eo6039-overexpressing Arabidopsis plants emitted significant amounts of α-muurolene (69.4%) and eremophilene (21.8%). Together, we demonstrated that the two TPSs play roles in producing major volatile terpenes in Eo.

Centipedegrass extracts regulate LPS-mediated aberrant immune responses by inhibiting Janus kinase.

BACKGROUND: Centipedegrass extract (CGE) is rich in several polyphenolic compounds including C-glycosylflavonoids, such as maysin and its derivatives, and exerts antioxidant, anti-adipogenic and anticancer effects. However, the effect of CGE on the immune system is unclear. PURPOSE: CGE might inhibit NO production induced by lipopolysaccharide (LPS). In this study, we propose a molecular mechanism for regulation of aberrant immune responses by CGE in LPS-stimulated RAW264.7 cells. STUDY DESIGN: We will preparation of Centipedegrass extract and purify partially in rich of maysin and its derivatives. And examine the effect of the CGE on immune system using LPS-induced RAW cells and animals. METHODS: LPS-induced nitric oxide (NO) and interleukin-6 levels were measured by enzyme-linked immunosorbent assay. The mRNA and protein levels of immune mediators were analyzed by reverse-transcription polymerase chain reaction and immunoblotting, respectively. RESULTS: CGE inhibited LPS-induced NO production in a concentration-dependent manner by suppressing inducible nitric oxide synthase (iNOS) expression in LPS-stimulated cells; this effect was mediated by inhibition of the JAK/STAT pathway. However, CGE did not regulate the expression of other factors, including phosphorylated p38, c-jun N-terminal kinase, or extracellular signal-regulated kinase 1/2. In addition, CGE increased T cells percentage in peripheral blood after oral administration. CONCLUSION: These results indicate that CGE suppresses LPS-induced production of NO and expression of iNOS by directly inhibiting JAK2 kinase activity and enhancing effects on the immune system in mice.

De novo assembly and comparative transcriptome analysis reveals genes potentially involved in tissue-color changes in centipedegrass (Eremochloa ophiuroides [Munro] Hack.).

Turf color is the most important characteristics of visual quality for a turfgrass species with high ornamental value and wide application prospects. Centipedegrass is a well-adapted warm-season turfgrass species in tropical, subtropical and temperate regions, possessing many outstanding properties including uniform green color. However, quite a few centipedegrass accessions or cultivars produce stolons and spike tissues with red-purple color, thereby decreasing their aesthetic value. A research focus in centipedegrass is to develop high-quality cultivars with uniform green color. To explore the major genes associated with the color changes in certain organs/tissues contributes to understand the molecular mechanisms of the same tissues having different phenotypic characteristics. In the present study, two phenotypically distinct centipedegrass accessions, E092 being a wild-type (WT) with red-purple stolons and spike tissues and E092-1 being a mutant (MT) with uniform green stolons and spike tissues, were used. Using the Illumina sequencing platform, approximately 401.7 million high-quality paired-end reads were obtained. After de novo assembly and quantitative assessment, 352,513 transcript sequences corresponding to 293,033 unigenes were generated with an average length of 735 bp. A total of 145,032 (49.49%) unigenes were annotated by alignment with public protein databases. Of these unigenes, 329 differentially expressed genes (DEGs) were identified between WT and MT stolons, with 156 up-regulated and 173 down-regulated; and 829 DEGs were detected between WT and MT spike tissues, including 497 up-regulated and 332 down-regulated. The expression profile of 10 randomly selected DEGs was confirmed with RT-qPCR. Candidate genes involved in the flavonoid biosynthesis were identified showing significant transcript changes between WT and MT organs/tissues. And transcript abundances of these flavonoid biosynthetic pathway-related genes were positively correlated with the accumulation of total anthocyanin in respective organs/tissues. This assembled transcriptome of centipedegrass can be served as a global description of expressed genes of above-ground organs/tissues and provide more molecular resources for future functional characterization analysis of genomics in warm-season turfgrass. Identified genes related to centipedegrass organ/tissue changes will contribute to molecular improvement of turf quality through genetic manipulation.

Growth and physiological responses of two phenotypically distinct accessions of centipedegrass (Eremochloa ophiuroides (Munro) Hack.) to salt stress.

Salinity is one of the major abiotic environmental stress factors affecting plant growth and development. Centipedegrass (Eremochloa ophiuroides [Munro)] Hack.) is an important warm-season turfgrass species with low turf maintenance requirements, but is sensitive to salinity stress. To explore salt tolerant germplasms in centipedegrass and better understand the growth and physiological responses of centipedegrass to salinity, we conducted anatomic observation and phytochemical quantification, examined growth parameters, and investigated photosynthetic machinery and antioxidant system in two phenotypically distinct centipedegrass accessions under NaCl salt stress. The morphophenotypical difference of the stems in the two accessions mainly depends on whether or not a thickened epidermal horny layer with purple colour was formed, which was caused by anthocyanin accumulation in the tissue. Successive salinity treatment was found to result in an inhibition of leaf growth, a marked decrease in photosynthesis, chlorophyll contents, and the maximal photochemical efficiency of PSII (Fv/Fm). Under the same treatment, purple-stem accession (E092) showed a lower degree of inhibition or decrease than green-stem one (E092-1). With the exception of malondialdehyde level, both proline content and antioxidant enzymes were upregulated to a greater extent in E092 following exposure to salinity condition. Meanwhile, significant enhancements of anthocyanin accumulation and total protein synthesis were detected in E092 after salt treatment, but not in E092-1. These results demonstrated that E092 favor better accumulation of anthocyanins under salinity condition, which contribute to salt tolerance by adjusting physiological functions and osmotic balance, and better maintenance of high turf quality. Hence, genetic phenotype can be utilized as a key indicator in E. ophiuroides breeding for salt-tolerance.

Transgenic Centipedegrass (Eremochloa ophiuroides [Munro] Hack.) Overexpressing S-Adenosylmethionine Decarboxylase (SAMDC) Gene for Improved Cold Tolerance Through Involvement of H2O2 and NO Signaling.

Centipedegrass (Eremochloa ophiuroides [Munro] Hack.) is an important warm-season turfgrass species. Transgenic centipedgrass plants overexpressing S-adenosylmethionine decarboxylase from bermudagrass (CdSAMDC1) that was induced in response to cold were generated in this study. Higher levels of CdSAMDC1 transcript and sperimidine (Spd) and spermin (Spm) concentrations and enhanced freezing and chilling tolerance were observed in transgenic plants as compared with the wild type (WT). Transgenic plants had higher levels of polyamine oxidase (PAO) activity and H2O2 than WT, which were blocked by pretreatment with methylglyoxal bis (guanylhydrazone) or MGBG, inhibitor of SAMDC, indicating that the increased PAO and H2O2 were a result of expression of CdSAMDC1. In addition, transgenic plants had higher levels of nitrate reductase (NR) activity and nitric oxide (NO) concentration. The increased NR activity were blocked by pretreatment with MGBG and ascorbic acid (AsA), scavenger of H2O2, while the increased NO level was blocked by MGBG, AsA, and inhibitors of NR, indicating that the enhanced NR-derived NO was dependent upon H2O2, as a result of expression CdSAMDC1. Elevated superoxide dismutase (SOD) and catalase (CAT) activities were observed in transgenic plants than in WT, which were blocked by pretreatment with MGBG, AsA, inhibitors of NR and scavenger of NO, indicating that the increased activities of SOD and CAT depends on expression of CdSAMDC1, H2O2, and NR-derived NO. Our results suggest that the elevated cold tolerance was associated with PAO catalyzed production of H2O2, which in turn led to NR-derived NO production and induced antioxidant enzyme activities in transgenic plants.

Differential Responses of Polyamines and Antioxidants to Drought in a Centipedegrass Mutant in Comparison to Its Wild Type Plants.

Centipedegrass (Eremochloa ophiuroides [Munro] Hack.) is an important warm-season turfgrass species with low turf maintenance requirements. However, our knowledge on physiological adaptation of centipedegrass to drought stress is limited. Physiological responses to drought in a gamma-ray-induced mutant 22-1 as compared with two wild type (WT) lines were analyzed for understanding of drought tolerance mechanism of centipedegrass. The mutant showed an elevated drought tolerance with higher levels of relative water content, net photosynthetic rate (A) and stomatal conductance (gs) and lower levels of ion leakage and malondialdehyde (MDA) under drought stress as compared with WT plants. A showed significant correlation with gs and MDA. Higher levels of antioxidant enzymes activities, non-enzyme antioxidants, and polyamines including putrescine (Put), spermidine (Spd), and spermine (Spm) were maintained in 22-1 than in WT plants. Superoxide dismutase (SOD), catalase (CAT), ascorbate-peroxidase (APX), and glutathione reductase (GR) activities and ascorbic acid (AsA) content were significantly correlated with both Put and Spd levels, and reduced glutathione level was correlated with Put during drought stress. Exogenous application of Put, Spd, and Spm increased drought tolerance and activities of SOD, CAT, APX, and GR in WT plants. The results suggest that higher levels of polyamines and antioxidant defense system are associated with the elevated drought tolerance in 22-1, which may improve protection on photosynthesis against drought induced oxidative damage.

Preference of cattle grazing conterminous monocultures of centipedegrass (Eremochloa ophiuroides) and bahiagrass (Paspalum notatum) with contrasting regrowth durations.

This study conducted four experiments to assess preference of cattle between centipedegrass (Eremochloa ophiuroides; CG) and bahiagrass (Paspalum notatum; BG) under manipulated vegetation conditions. In each experiment, three Japanese Black cows (Bos taurus) were individually allowed to graze four plots of conterminous monocultures of CG and BG (30 min for each 10 × 10 m plot). Vegetation of the plots was the combinations of two patch sizes (coarse and fine; 5 × 10 m and 5 × 5 m, respectively) × two relative availabilities of the grasses (CGtall /BGshort and CGshort /BGtall ). Tall and short patches were created by varying regrowth durations after cutting, and almost always offered a clear CG-BG contrast in availability (height and mass) and quality (digestibility and protein) with the trade-off relationship. Cows always preferred or equally selected CG to BG, with the preference being affected by the relative availability of grasses in three of the four experiments but not by the patch size at all. Test of intake rate maximization on both short-term and daily bases indicated that cows chose between the grasses to simultaneously enhance daily potential intake of dry matter, digestible energy and protein. The results strengthen the value of CG as a forage.