Sinensis Plants Research Articles

Agrobacterium-mediated genetic transformation of Miscanthus sinensis

Miscanthus species are tall perennial rhizomatous grasses with C4 photosynthesis originating from East Asia, and they are considered as important bioenergy crops for biomass production. In this study, Agrobacterium-mediated transformation system for M. sinensis was developed using embryogenic calli derived from mature seeds. In order to establish a stable system, optimum conditions to obtain highly regenerable and transformation-competent embryogenic calli were investigated, and embryogenic calli were efficiently induced with callus induction medium containing 3 mg L−1 2,4-dichlorophenoxyacetic acid and 25 mM l-proline, at pH 5.7 with an induction temperature of 28 °C. In addition, the embryogenic callus induction and regeneration potentials were compared between seven M. sinensis germplasms collected from several sites in Korea, which revealed that the germplasm SNU-M-045 had superior embryogenic callus induction and regeneration potentials. With this germplasm, the genetic transformation of M. sinensis was performed using Agrobacterium tumefaciens EHA105 carrying pCAMBIA1300 with a green fluorescence protein gene as a reporter. After putative transgenic plants were obtained, the genomic integration of transgenes was confirmed by genomic PCR, transgene expression was validated by Northern blot analysis, and the number of transgene integration was confirmed by DNA gel blot analysis. Furthermore, the Agrobacterium-mediated transformation of M. sinensis was also performed with pCAMBIA3301 which contains an herbicide resistance gene (BAR), and we obtained transgenic M. sinensis plants whose herbicide resistance was confirmed by spraying with BASTA®. Therefore, we have established a stable Agrobacterium-mediated transformation system for M. sinensis, and also successfully produced herbicide-resistant Miscanthus plants by introducing BAR gene via the established method.

Hydrogen peroxide induces vessel occlusions and stimulates sesquiterpenes accumulation in stems of Aquilaria sinensis

Agarwood is highly valuable resinous and fragrant heartwood, produced principally from tropical tree species in the genus Aquilaria, which is used widely in countries of the Middle East, Southeast Asia and Japan. Generally, healthy trees will not produce agarwood, but wounding of the tree initiates the production of agarwood. In this study, the pruning of actively growing saplings of Aquilaria sinensis resulted in hydrogen peroxide (H2O2) burst, which was followed by formation of vessel occlusions and sesquiterpene biosynthesis in the pruned stems. Treatment of the pruned stems with scavenger of H2O2 (ascorbate, AsA) greatly reduced the amount of H2O2 released, the number of vessel occlusions, and the amount of sesquiterpenes produced. In addition, exogenous H2O2 also induced A. sinensis plants to form vessel occlusions and produce sesquiterpenes as pruning treatment. The results indicated that H2O2 may be an important post-wounding signal in A. sinensis that leads to the induction of vessel occlusions formation and sesquiterpene biosynthesis, and thus H2O2 might play a vital role in agarwood formation in pruned stems of A. sinensis.

Diversity in ploidy levels and nuclear DNA amounts in Korean Miscanthus species

Because of its high biomass productivity and nutrient-use efficiency, Miscanthus spp. have emerged as promising bioenergy crops. Polyploidism in Miscanthus species is important because it allows non-native countries to cultivate sterile types, such as triploid M. × giganteus, in order to prevent ecosystem disturbance. Although M. sacchariflorus and M. sinensis are considered to be native to Korea, China, and Japan, accurate information describing the ploidy levels of these species has not yet been well established. To evaluate rough ploidy levels, 215 accessions of Miscanthus species were estimated by relative fluorescence intensities of DAPI-stained nuclei. After evaluation of rough ploidy levels, 20 plants were randomly selected by species and ploidy levels, and ploidy levels were examined by counting chromosomes, estimating propidium iodide-stained DNA content with flow cytometry, and measuring sizes of stomata and caryopsis. Among the 20 plants examined, 3 were diploid and 6 were tetraploid M. sacchariflorus (4.56 ± 0.01 and 8.90 ± 0.14 pg/2C nuclear DNA content, respectively), while 6 were diploid and 3 were triploid M. sinensis (5.40 ± 0.18 and 8.29 ± 0.33 pg/2C nuclear DNA content, respectively). One plant was a putative triploid M. × giganteus having 7.31 pg/2C nuclear DNA content, which was similar to an M. × giganteus plant from Illinois, USA having 7.23 pg/2C nuclear DNA content. We confirmed the occurrence of diploid and tetraploid M. sacchariflorus and M. sinensis plants as well as a putative triploid M. × giganteus native to Korea. Thus, measurement of the size of stomata and caryopsis allowed prediction of ploidy levels in the field and laboratory.

Effect of shade treatment on theanine biosynthesis in Camellia sinensis seedlings

Theanine synthetase (TS) is a key enzyme involved in theanine biosynthesis. In our recent study, it has been revealed that theanine biosynthesis derived from nitrogen metabolism in tea (Camellia sinensis) plants can be influenced by shading treatment. The expression patterns of CsTS protein in the roots and shoots of tea seedlings were examined by western blot using a self-prepared polyclonal antibody with high specificity and sensitivity. The effect of long-term shade treatment on the levels of theanine synthesis was also investigated using roots and shoots of tea seedlings. Levels of theanine and total free amino acids gradually increased in shoots, reaching their maximum after 22 days of treatment (DOT). The immunoblotting analysis suggested that CsTS protein levels increased gradually up to 22 DOT and expression remained at a high level, except after 1 DOT where levels were low in both roots and shoots. The increased theanine concentration we observed in the shading treatment may be due to increased nitrogen assimilation and reduced theanine catabolism under shade conditions.

Diurnal changes in photosynthesis and antioxidants of Angelica sinensis as influenced by cropping systems

Diurnal dynamics of photosynthetic character of Angelica sinensis, as well as effect of continuous cropping on leaf photosynthetic character, antioxidant enzyme activity and growth of A. sinensis were investigated under field condition. The results showed that the diurnal net photosynthetic rate of A. sinensis in sunny day exhibited a double-peak pattern, and the peaks occurred at 9:45 and 16:45 h, respectively. There was a significant midday depression with A. sinensis, which was caused principally by stomatal factors such as stomatal conductance. The results also showed that net photosynthetic rate (P N), transpiration rate (E), stomatal conductance (g s), intercellular CO2 concentration (C i), and chlorophyll content (Chl) of A. sinensis plants under continuous cropping were significantly lower than those of the control. The activity of total superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT), and growth parameters of A. sinensis plants were significantly decreased under continuous cropping condition. This study provides evidence of continuous cropping obstacle effect on photosynthesis, antioxidant enzyme activity, and growth parameters of A. sinensis in a field experiment, which partly explained the yield reduction of A. sinensis in the field, when it was cultivated continuously on the same soil.

Citrus psorosis and Mirafiori lettuce big-vein ophiovirus coat proteins localize to the cytoplasm and self interact in vivo

Citrus psorosis (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV) belong to the family Ophioviridae, plant viruses with filamentous nucleocapsids and segmented genomes of negative polarity, causing the worldwide distributed citrus psorosis and lettuce big-vein diseases, respectively. To gain insight into the replication cycle of these viruses, the subcellular localization of the viral coat proteins (CP) was studied. Immunoblot analysis of fractionated extracts derived from natural and experimental infected hosts indicated that the CP of CPsV occurs in the soluble cytoplasmic fraction. The cytoplasmic localization of this protein was confirmed by confocal microscopy of fluorescent protein (FP)-tagged CP following its expression in either CPsV-infected and healthy Citrus sinensis plants or in Nicotiana benthamiana plants. The same localization was observed for FP-tagged CP of MiLBVV. The CPs of CPsV and MiLBBV can undergo homologous and heterologous interactions as revealed by fluorescent lifetime imaging microscopy and co-immunoprecipitation analysis. A putative leucine zipper motif that is conserved among ophiovirus CP sequences may account for these interactions.

Characterisation of volatile and non-volatile metabolites in etiolated leaves of tea (Camellia sinensis) plants in the dark

Aroma is an essential factor affecting the quality of tea (Camellia sinensis) products. While changes of volatile compounds during tea manufacturing have been intensively studied, the effect of environmental factors on volatile contents of fresh tea leaves has received less attention. We found that C. sinensis var. Yabukita kept in darkness by shading treatment for 3weeks developed etiolated leaves with significantly increased levels of volatiles, especially volatile phenylpropanoids/benzenoids (VPBs). Upstream metabolites of VPBs, in particular shikimic acid, prephenic acid, and phenylpyruvic acid, showed lower levels in dark treated than in control leaves, whereas the contents of most amino acids including l-phenylalanine, a key precursor of VPBs, were significantly enhanced. In addition, analysis by ultra performance liquid chromatography-time of flight mass spectrometry, capillary electrophoresis–time of flight mass spectrometry, high performance liquid chromatography, and gas chromatography–mass spectrometry indicated that volatile and non-volatile metabolite profiles differed significantly between dark treated and untreated leaves.

Pyridine metabolism in tea plants: salvage, conjugate formation and catabolism

Pyridine compounds, including nicotinic acid and nicotinamide, are key metabolites of both the salvage pathway for NAD and the biosynthesis of related secondary compounds. We examined the in situ metabolic fate of [carbonyl-(14)C]nicotinamide, [2-(14)C]nicotinic acid and [carboxyl-(14)C]nicotinic acid riboside in tissue segments of tea (Camellia sinensis) plants, and determined the activity of enzymes involved in pyridine metabolism in protein extracts from young tea leaves. Exogenously supplied (14)C-labelled nicotinamide was readily converted to nicotinic acid, and some nicotinic acid was salvaged to nicotinic acid mononucleotide and then utilized for the synthesis of NAD and NADP. The nicotinic acid riboside salvage pathway discovered recently in mungbean cotyledons is also operative in tea leaves. Nicotinic acid was converted to nicotinic acid N-glucoside, but not to trigonelline (N-methylnicotinic acid), in any part of tea seedlings. Active catabolism of nicotinic acid was observed in tea leaves. The fate of [2-(14)C]nicotinic acid indicates that glutaric acid is a major catabolite of nicotinic acid; it was further metabolised, and carbon atoms were finally released as CO(2). The catabolic pathway observed in tea leaves appears to start with the nicotinic acid N-glucoside formation; this pathway differs from catabolic pathways observed in microorganisms. Profiles of pyridine metabolism in tea plants are discussed.

Herbivore-Induced Volatiles from Tea (Camellia sinensis) Plants and Their Involvement in Intraplant Communication and Changes in Endogenous Nonvolatile Metabolites

As a defense response to attacks by herbivores such as the smaller tea tortrix ( Adoxophyes honmai Yasuda), tea ( Camellia sinensis ) leaves emit numerous volatiles such as (Z)-3-hexen-1-ol, linalool, α-farnesene, benzyl nitrile, indole, nerolidol, and ocimenes in higher concentration. Attack of Kanzawa spider mites ( Tetranychus kanzawai Kishida), another major pest insect of tea crops, induced the emission of α-farnesene and ocimenes from tea leaves. The exogenous application of jasmonic acid to tea leaves induced a volatile blend that was similar, although not identical, to that induced by the smaller tea tortrix. Most of these herbivore-induced plant volatiles (HIPV) were not stored in the tea leaves but emitted after the herbivore attack. Both the adaxial and abaxial epidermal layers of tea leaves emitted blends of similar composition. Furthermore, HIPV such as α-farnesene were emitted mostly from damaged but not from undamaged leaf regions. A principal component analysis of metabolites (m/z 70-1000) in undamaged tea leaves exposed or not to HIPV suggests that external signaling via HIPV may lead to more drastic changes in the metabolite spectrum of tea leaves than internal signaling via vascular connections, although total catechin contents were slightly but not significantly increased in the external signaling via HIPV.

Laser-induced fluorescence imaging method to monitor citrus greening disease

The potential use of laser-induced fluorescence imaging to monitor citrus greening disease in sweet orange ( Citrus sinensis) plants was investigated. For this purpose, an experiment using healthy plants as the control and plants of the same variety inoculated with Candidatus Liberibacter asiaticus (CLas) was performed to verify the fluorescence response through 8 months after bacterial inoculation. A system using laser-induced fluorescence at 473 nm of wavelength excitation was configured. Color descriptors from the collected fluorescence images were determined using the average values for red (R); green (G); blue (B); hue (H); saturation (S); value (V, or Intensity); relative red (rR); relative green (rG); relative blue (rB) and luminosity (L). The data were analyzed using a paired Student’s t-test by means of comparison between diseased and healthy plants. The results demonstrate a potential method to identify citrus greening at an early stage of the disease (first month), prior to the onset of visual symptoms at confidence level of 95%.

Brassinolide alleviates salt stress and increases antioxidant activity of cowpea plants (Vigna sinensis)

Soil salinity is one of the most severe factors limiting growth and physiological response in Vigna sinensis plants. Plant salt stress tolerance requires the activation of complex metabolic activities including antioxidative pathways, especially reactive oxygen species and scavenging systems within the cells which can contribute to continued growth under water stress. The present investigation was carried out to study the role of brassinolide in enhancing tolerance of cowpea plants to salt stress (NaCl). Treatment with 0.05ppm brassinolide as foliar spray mitigated salt stress by inducing enzyme activities responsible for antioxidation, e.g., superoxide dismutase, peroxidase, polyphenol oxidase, and detoxification as well as by elevating contents of ascorbic acid, tocopherol, and glutathione. On the other hand, total soluble proteins decreased with increasing NaCl concentrations in comparison with control plants. However, lipid peroxidation increased with increasing concentrations of NaCl. In addition to, the high concentrations of NaCl (100 and 150mM) decreased total phenol of cowpea plants as being compared with control plants. SDS-PAGE of protein revealed that NaCl treatments alone or in combination with 0.05ppm brassinolide were associated with the disappearance of some bands or appearance of unique ones in cowpea plants. Electrophoretic studies of α-esterase, β-esterase, polyphenol oxidase, peroxidase, acid phosphatase, and superoxide dismutase isoenzymes showed wide variations in their intensities and densities among all treatments.

First Report of Basal Stem Rot and Foliar Blight Caused by Pythium sylvaticum on Miscanthus sinensis in Illinois.

Miscanthus sinensis Anderss., a perennial grass, is native to eastern Asia. It has been widely grown as an ornamental in temperate regions of the world, including the United States, and recently has become an important component of public and private sector bioenergy feedstock Miscanthus selection programs. In August 2008, stem rot and blight was observed on M. sinensis plants in two irregular patches, ~2 to 2.5 × 1 to 1.5 m each in a trial plot that was preceded by corn, at the University of Illinois Energy Farm near Urbana, IL. At the time of the observation, most plants were dead and the wilted tillers had black, soft rotted basal stems. A few plants were stunted and the crowns of the tillers had black-to-brown soft rot. Some tillers' leaves were dead and others had turned light brown. Sample tissue fragments were surface disinfested in 0.5% NaOCl and plated on 1% water agar (WA). After 3 days of incubation in the dark at 23°C, colonies were transferred to corn meal agar (CMA), potato dextrose agar (PDA), or 10% V8 juice agar and incubated at 23°C under continuous white light for up to 2 weeks. Morphological characteristics of the isolates correspond to those originally described for Pythium sylvaticum W.A. Campb. & J.W. Hendrix (1). The mycelia grew and covered the 10-cm-diameter plates within 5 days. On PDA, the culture was a creamy white mycelial mat of coenocytic hyphae. The isolates produced only globose, terminal or intercalary hyphal swellings ranging from 28 to 48 μm in diameter, but no oogonia were produced on any of the three growth media. No zoospores were produced when agar blocks bearing mycelium were flooded with distilled water or 1% soil water. Sequence analysis was performed with the internal transcribed spacer (ITS) region of the rDNA amplified with primer pair ITS1/ITS4 (3) and the mitochondrially encoded cytochrome c oxydase subunit II (cox II) gene using primers FM58/FM66 (2). The resulting 871-bp ITS nucleotide sequence (Accession No. HM991706) was identical among all three isolates analyzed and 99% identical (100% coverage) to ITS sequences of multiple isolates of P. sylvaticum in GenBank. Likewise, the 544-bp cox II sequence (Accession No. HQ454429) was 99% identical (97% coverage) to cox II sequences of multiple isolates of P. sylvaticum. Six pots of M. sinensis seedlings were inoculated by placing two CMA plugs of a 2-week-old culture of isolate F71 at the crown. The control pots were mock inoculated with sterile CMA plugs. The plants were incubated at ~90% relative humidity (RH) and 25°C day and 22°C night for 3 days, and thereafter left on the greenhouse bench at ~65% RH with alternating 9 h of darkness and 15 h of light. Three weeks after inoculation, two of the inoculated seedlings wilted, others were stunted with leaves wilting from the tip downwards and the stems rotting from the crown upward. A thick mat of mycelia was seen on the rotted basal stems. No symptoms were observed in the control. P. sylvaticum was reisolated from both the rotted basal stems and the wilted foliage. To our knowledge, this is the first report of P. sylvaticum on M. sinensis. Infestation of farm soils with P. sylvaticum could limit M. sinensis biomass production significantly by limiting seedling establishment.

Establishment of a Regeneration System by Callus Induction from Explants of Miscanthus sinensis

Immature inflorescence sections of greenhouse-grown Miscanthus sinensis plants and collected seeds were cultivated on Murashige and Skoog medium containing benzyladeninepurine (BAP) and 2,4-dichlorophenoxyacetic acid or BAP and 1-naphthaleneacetic acid (NAA). From immature inflorescence sections, the rate of callus induction depended on the collection area of the cultivar. The percentage shoot regeneration through embryogenic callus was increased and a new shoot-forming type was formed after inclusion of BAP during shoot regeneration. A higher percentage of shoot regeneration was achieved from immature inflorescence sections cultured with 5 mg L-1 BAP and 0.24 mg L-1 NAA. Plants from callus induction were regenerated from embryogenic callus, but regeneration capacity differed depending on cultivar collection area.

Synergistic effect of kinetin and spermine on some physiological aspects of seawater stressed Vigna sinensis plants

The effect of kinetin, spermine and their combination on growth vigor, photosynthetic pigments, some metabolites, some enzymes, polyamines and productivity of salt-stressed Vigna sinensis plants was investigated. Salt stress reduced all evaluated growth criteria and yield components of used plants. Chlorophyll (CHL (a, b, carotenoids, carbohydrates, protein, spermidine and spermine level as well as and amylase activity were also decreased in response to salinity. On the other hand, proline, K +, Na + and putrescine concentration, and peroxidase activity were increased in the salt-stressed plants. Exogenous application of kinetin and spermine mitigated the deleterious effects of salinity stress on growth and yield of the used plants. Conversely, the combined treatment of kinetin and spermine induced additional reduction in growth and yield of the stressed plants, and the effect appeared to be constitutive. The protective effect of kinetin and spermine on V. sinensis plants appeared mainly due to the enhancement effect of these growth regulators on chlorophylls and protein content and polyamines titer.

Influence of beneficial microorganisms during in vivo acclimatization of in vitro-derived tea (Camellia sinensis) plants

The effects of native isolates of Pseudomonas fluorescens, Azospirillum brasilense, and Trichoderma harzianum on rooting and acclimatization of in vitro-grown shoots and plantlets of tea were evaluated. In vitro bacterization of P. fluorescens failed to establish, while both T. harzianum and A. brasilense retarded shoot growth, eventually overtaking shoot cultures in in vitro rooting. Acclimatization of rooted plantlets in soil amended with bioinoculants, either individually or in various combinations, promoted plantlet survival. Moreover, efficiency of nutrient uptake of plantlets was higher in the presence of microorganisms. Root rot or wilting of tissue culture-derived plants was not observed in bioinoculant-treated plants, as they possessed relatively higher activities of defense enzymes, including peroxidase and phenylalanine ammonia lyase.

First Report of Wisteria vein mosaic virus on Wisteria sinensis in New Zealand.

Wisteria vein mosaic virus (WVMV) is a member of the Potyvirus genus. The virus has been reported in Wisteria spp. in Australia, China, the United States, and a number of European countries (2). In 2006, several W. sinensis plants with mottling and mosaic symptoms were observed in a commercial plant nursery in Whenuapai, north of Auckland, New Zealand. These plants had been propagated from a nursery in the New Plymouth area of New Zealand. Sap from the symptomatic Wisteria plants was examined with an electron microscope and elongated and flexuous potyvirus-like particles approximately 750 nm long were observed. RNA was extracted from the symptomatic plants with a Qiagen RNeasy Plant Mini Kit (Doncaster, Australia). The RNA was initially tested using general potyvirus primers, PV1/SP6 (4) and U335 (3), with the cycling conditions of 94°C for 5 min followed by 40 cycles of 94°C for 45 s, 50°C for 45 s, 72°C for 90 s, and a final extension of 72°C for 7 min. The polymerase chain reaction (PCR) product (695 bp) was directly sequenced (GenBank Accession No. EU580146) and a BLAST search in GenBank showed 98% nucleotide identity with WVMV (GenBank Accession no. AF484549). The RNA was then tested using WVMV-specific primers, WVMVF1 and WVMVR1, and the published cycling conditions (2). PCR amplicons of 701 bp were obtained. PCR products were directly sequenced (GenBank Accession No. EU308592), and a BLAST search in GenBank showed 98% nucleotide identity with published sequences of WVMV (GenBank Accession Nos. AF484549 and AY656816). In addition, RNA was extracted from the original isolate of WVMV that was reported in the Netherlands (1; supplied by R. van der Vlugt, Plant Research International) and the RNA was amplified using the WVMV-specific primer pair. The sequence obtained from PCR amplicons of the type isolate (GenBank Accession No. EU308593) showed a 98% nucleotide identity with the New Zealand WVMV isolate and with published sequences of WVMV (as shown above). From the symptomatology, particle morphology, and nucleotide sequences, it is concluded that WVMV is present in New Zealand. The distribution of the virus in New Zealand is not known, but the affected plants at the New Plymouth nursery may have been imported into New Zealand as many as 30 years ago. Although WVMV infection can reduce the quality of commercial plants, the disease is not economically significant in New Zealand.

First Report of Dodder Transmission of 'Candidatus Liberibacter asiaticus' to Tomato (Lycopersicon esculentum).

Citrus huanglongbing (HLB) is one of the most devastating diseases of citrus worldwide. The disease is associated with three different species of fastidious α-proteobacteria, namely 'Candidatus Liberibacter asiaticus', Ca. L. americanus, and Ca. L. africanus (1). 'Ca. L. asiaticus' was first detected in South Florida in 2005 and has spread throughout the citrus-growing areas of Florida. 'Ca. L. asiaticus' is transmitted naturally by the Asian citrus psyllid, Diaphorina citri, and can also be transmitted by graft propagation and via various species of dodder (Cuscuta). HLB affects most if not all citrus and citrus relatives within the family Rutaceae (2), including the ornamental shrub Murraya paniculata (4). In addition, 'Ca. L. asiaticus' and 'Ca. L. americanus' can infect tobacco (Nicotiana xanthi) and periwinkle (Catharanthus roseus) (1,4). Here we report that 'Ca. L. asiaticus' can infect tomato (Lycopersicon esculentum) cvs. Manapal and FL47. Manapal and FL 47 plants grown from seed were placed adjacent to 'Ca. L. asiaticus'-infected sweet orange (Citrus sinensis) plants with dodder (Cuscuta pentagona) already well established on them. Young dodder shoots still connected to the citrus were draped over the tomato plants and subsequently also became attached to the tomato stems. After 1 month, the tomato plants were detached from the citrus and most of the dodder removed. One month later, these tomato plants started to show vein clearing and subsequently the mature leaves became thicker and leathery. Some leaves showed blotchy mottle symptoms and some fruits became lopsided in a manner similar to HLB symptom expression on citrus. PCR amplification of the 'Ca. L. asiaticus' 16S rDNA with primers OI1/OI2c and the β-operon with primers A2/J5 (1) revealed the presence of 'Ca. L. asiaticus' DNA. Sequence analysis confirmed that the sequences of the cloned amplicons were identical to those from the HLB-infected citrus source plant. Both conventional and quantitative real-time PCR (3) revealed a much lower abundance of 'Ca. L. asiaticus' DNA in tomato as compared with 'Ca. L. asiaticus'-infected citrus or periwinkle, indicating that 'Ca. L. asiaticus' bacteria multiplied at a lower titer in these tomato cultivars.

Evaluation of forest grazing as a management practice to utilize and control Miscanthus sinensis in a young tree plantation in southern Kyushu, Japan

Abstract Miscanthus sinensis, a tall cespitose warm‐season perennial grass, occurs widely in tree plantations across Japan as a major weed. A 4‐year grazing trial was conducted in a young Chamaecyparis obtusa (an evergreen conifer) plantation in south‐western Japan, in an effort to evaluate forest grazing as a management practice to utilize and control M. sinensis and to deepen a mechanistic understanding of the responses of this grass to herbivory. The plantation was continuously stocked by Japanese Black cattle during the grazing season from May–November (July–November in the first year) at rates of 0.38–0.63 animals ha−1. M. sinensis was always the major species selected by animals, with most of the plants (80–90%) in the paddocks being defoliated during the grazing season. Reflecting these, M. sinensis plants declined with time under grazing, mainly in shoot number per tuft and partly in leaf appearance rate and the weight of the youngest fully expanded leaf, leading to a drastic decrease in production of leaves (laminae) as a photosynthetic organ and a main feed component for animals. Animals did little fatal damage to C. obtusa trees. Performance of animals was acceptable for cows that had experienced forest grazing in preceding years, but needed improvement for inexperienced heifers and calves. The results demonstrate the potential value of cattle grazing in utilizing and controlling M. sinensis in young tree plantations, which enables multifunctional land use linking forest and animal industries. Further studies are necessary to explore the optimum grazing intensity that balances effective control and extended utilization of the grass.

Water relations and growth vigour ofVigna sinensisplants in relation to indole acetic acid, gibberellic acid or kinetin

The objective of this study was designed to investigate the effect of seed priming with different concentrations of growth bioregulators ( i.e. indole acetic acid, gibberellic acid or kinetin) on growth and water relations of Vigna sinensis plants throughout various stages of plant growth and development.In the majority of cases, indole acetic acid, gibberellic acid or kinetin at different concentrations stimulated the growth vigour of cowpea plants (root length, root fresh and dry weight, shoot length, shoot fresh and dry weights and leaf area production) all over the growth periods.Seed pretreatment with IAA or kinetin induced significant increases in total leaf conductance and transpiration rate that was accompanied by a decrease in RWC. On the other hand, GA 3 induced reverse situation in these parameters.

Lifestyles of Colletotrichum acutatum.

Lifestyles of Colletotrichum acutatum.