Showing papers in "Botanical Bulletin of Academia Sinica in 2004"

Studies on the production of some important secondary metabolites from medicinal plants by plant tissue cultures

Abstract: Plants are a tremendous source for the discovery of new products of medicinal value for drug development. Today several distinct chemicals derived from plants are important drugs currently used in one or more countries in the world. Many of the drugs sold today are simple synthetic modifications or copies of the naturally obtained substances. The evolving commercial importance of secondary metabolites has in recent years resulted in a great interest in secondary metabolism, particularly in the possibility of altering the production of bioactive plant metabolites by means of tissue culture technology. Plant cell culture technologies were introduced at the end of the 1960's as a possible tool for both studying and producing plant secondary metabolites. Different strategies, using an in vitro system, have been extensively studied to improve the production of plant chemicals. The focus of the present review is the application of tissue culture technology for the production of some important plant pharmaceuticals. Also, we describe the results of in vitro cultures and production of some important secondary metabolites obtained in our laboratory.

Copper-induced stress and antioxidative responses in roots of Brassica juncea L.

Abstract: Copper induced antioxidative reactions in the roots of Brassica juncea L. were investigated in both time-and concentration-dependent manners. The rapid uptake of Cu was observed immediately after the start of treatment. Application of Cu at 8µM caused 50 percent reduction in biomass of Cu-treated roots as compared with control. Cu-induced root growth inhibition paralleled the level of root oxidative damage. Treatment with Cu at 8µM induced a twofold increase in H2O2 content during the first 4 d, but it declined to the basal level thereafter. We also observed a twofold increase in superoxide dismutase activities with 8µM Cu during the first 2 d. The stimulation lasted for 4 d and then gradually declined. Activities of both ascorbate peroxidase and guaiacol peroxidase in roots were found to be low during the first 4 d after seedling exposure to 8µM Cu, but significantly increased after that, suggesting that increased enzyme activities would be responsible for the removal of H2O2. Catalase activities were always suppressed under Cu stress. Treatment of seedlings with 8µM Cu induced general decreases in both reduced ascorbate and dehydroascorbate. The reduced glutathione content decreased at early stages of Cu treatment. However, it was restored to the level of controls thereafter. In contrast, the oxidized glutathione contents showed a progressive increase during the time of Cu treatment. The total non-protein thiol content was shown to increase during the first several days, but it declined at later stages.

Growth Promotion and Induction of Systemic Resistance in Rice Cultivar Co-47 (Oryza sativa L.) by Methylobacterium spp.

Abstract: Pink-pigmented facultatively methylotrophic bacteria (PPFMs), persistent colonizers of plant leaf surfaces, belong to the genus Methrlobacterium and are mostly transmitted through seeds. Plant growth-promoting activity of methylotrophic bacteria and their effects on disease suppression were evaluated on rice under greenhouse conditions. Rice seeds were inoculated with Methylobacterium sp. strain PPFM-Os-07 and seed germination was evaluated in terms of morphometric measurements, seedling growth, rate of germination (R(subscript G)), and seedling vigour index (SVI). Another experiment was carried out to study the induction of pathogenesis-related proteins (PR-proteins) in rice plants that were inoculated with methylotrophic bacteria by seed imbibition or foliar spray. In the third experiment, sixty-day-old rice plants grown in pots were challenge inoculated with Rhizoctonia solani strain TNAU-01. Methylobacterium inoculation promoted seed germination and plant growth. Increased plant height, number of tillers, plant biomass, and grain yield were observed. The average yield increases for seed imbibition and phyllosphere spray were, respectively, 22.1% and 24.3% greater than control. The bacteria also significantly reduced the sheath blight incidence when applied as either bacterial culture through seed imbibition and or phyllosphere spray. The percent disease reduction recorded for seed imbibition alone and for combined applications of seed imbibition and phyllosphere spray were 17.8% and 23.5%. Rice plants sprayed with PPFM-Os-07 strain showed increased presence of PR-proteins and phenolic contents on day 1 after application. Maximum phenylalanine ammonia lyase (PAL) and peroxidase activity on day 4 and β-1,3-glucanase and chitinase activity on day 5 were recorded. The results suggest that Methylobacterium inoculation may alter rice susceptibility to R. solani. This work emphasizes the importance of evaluating induced systemic resistance while studying plant-associated growth promoting bacteria.

Antioxidant and antiproliferative activities of sweet potato (Ipomoea batatas (L.) Lam 'Tainong 57') constituents

Abstract: The aim of this study is to examine possible antioxidant and antiproliferative activities of the different extracts from sweet potato (Ipomoea batatas (L.) Lam 'Tainong 57') organs. DPPH staining, total phenolic com- pounds and flavonoid content, DPPH radical, reducing power method, FTC method, and cell proliferation were all employed. In the DPPH staining, ethanol extract of vein had the highest radical-scavenging activity when it was diluted to 6.25 mg dry matter/mL. Among all the extracts, the highest amount of total phenolic and flavonoid com- pounds was found in the ethanol extract of vein. In the DPPH colorimetric method, it was found that ethanol extract of leaf had the highest radical-scavenging activity, followed by water extract of vein. In the reducing power activity assay, it was found that the water extract of leaf had the highest reducing power activity, followed by ethanol extract of vein. Like phenolic compounds, the highest FTC activity was found in the ethanol extract of vein. The antipro- liferative activities aof sweet potato were studied in vitro using human lymphoma NB4 cells, and the following re- sults were found: water extract of vein had the highest antiproliferative activity with an EC 50 of 449.6 ± 27.73 µg/mL, followed by water extract of storage root, water extract of leaf, ethanol extract of storage root, and ethanol extract of leaf. Although the ethanol extract of vein showed strong antioxidant activity, it had no antiproliferative activity under the experimental conditions tested.

Effects of three allelopathic phenolics on chlorophyll accumulation of rice (Oryza sativa) seedlings: II. Stimulation of consumption-orientation

Abstract: The effects of three allelopathic phenolics, o-hydroxyphenyl acetic, ferulic and p-coumaric acids, on the chlorophyllase activity of rice leaf (Oryza sativa cv TN67) were investigated Ten-day-old green seedlings of rice were cultured in greenhouse for 16 d in Kimura's culture solution, which was changed every 4 days, with or without 50, 100 or 200 ppm of the phenolic compounds Just before changing the culture solution, leaves were harvested to determine their chlorophyll (Chl) and chlorophyllide (Chlide) contents, and their chlorophyllase a and b activities While the Chl and Chlide contents decreased and increased, respectively, causing the molar ratio of Chlide/Chl to increase, as the phenolic concentrations increased; the chlorophyllase a and b activities drastically increased This suggests that the consumption-orientation of Chl was significantly stimulated by the exogenously applied phenolics The order of inhibition of growth of the rice seedlings is: ferulic acid＞p-coumaric acid＞o-hydroxyphenylacetic acid The order of inhibition effect on Chl accumulation is: p-coumaric acid＞o-hydroxyphenylacetic＞ferulic acid The order of stimulation effect on chlorophyllase a activity is: o-hydroxyphenylacetic acid＞ferulic acid＞p-coumaric acid The order of promotion effect on chlorophyllase b is: ferulic acid＞o-hydroxyphenylacetic acid＞p-coumaric acid The different responses of chlorophyllase a and b activities to the same concentrations of allelochemical phenolics suggest that they may be two different enzymes It is apparent that the three phenolics may enhance the activities of enzymes, such as chlorophyllase and Mg-dechelatase, responsible for the Chl degradative pathway A combination of the present and the preceding data strongly suggest that the three allelopathic phenolics may comprehensively affect the biosynthetic and degradative pathways of Chl

Antrodia cinnamomea reconsidered and A. salmonea sp. nov. on Cunninghamia konishii in Taiwan

Abstract: Antrodia cinnamomea is reinstated as the correct name for a basidiomycete consistently associated with Cinnamomum kanehirai. Antrodia salmonea nov. sp., causing a brown heart rot of Cunninghamia konishii in Taiwan, is described and illustrated. Antrodia cinnamomea and A. salmonea are morphologically similar but can be separated by pore surface color of basidiomata and host preferences. Moreover, pairings between monokaryons of A. cinnamomea and those of A. salmonea demonstrate that these two fungi have different mating systems.

Analysis of toxic and osmotic effects of sodium chloride on leaf growth and economic yield of sugarcane

Abstract: Sugarcane (Saccharum officinarum L.) an important sugar crop, shows high sensitivity to salinity at various growth stages. This study was conducted to determine the comparative response of a salt-tolerant (CP-4333) and a salt-sensitive (CP-71-3002) sugarcane clone to the toxic or osmotic effects of sodium chloride at two growth stages. Clones indicated significant differences in terms of reductions in dry weight and area of leaves under salinity at the grand growth stage. Leaf dry weight was more affected than leaf area, resulting in reduced specific leaf weight. The tolerant clone produced supplementary tillers in greater number that appeared to offload the ion excess. Clones indicated significant difference with regard to increase in leaf Na+ and Cl- and decrease in K+, but no difference in K+:Na+ ratio. Na+ and Cl- were negatively correlated while K+ was positively correlated with leaf growth parameters, indicating an adverse effect of Na+ and Cl- and importance of K+ to salt tolerance. The tolerant clone displayed higher water content, water, and turgor potentials of leaf than sensitive clone, but the osmotic potential did not significantly change. Soluble sugars of tolerant clone indicated a ~2 fold increase over control, indicating their osmo-protective role. Free proline accumulation was more specific to the sensitive clone and was correlated with Na+ and Cl- showing its synthesis due to ion-toxicity. At maturity, increased salinity reduced the millable cane yield, extractable juice and juice-brix percentage, but increased juice osmolality. All these parameters were negatively correlated with EC, Na+, and Cl- and displayed their specific effect on the sugar levels in the internodes. In summary, ion-toxicity was the main determinant of salt tolerance at the grand growth stage while the osmotic component of NaCl mainly appeared to affect the transport of sucrose to stalks, followed by stimulated sucrolytic activity in the internodes, resulting in reduced final cane yield.

Antioxidant enzyme activities are upregulated in response to cadmium in sensitive, but not in tolerant, rice (Oryza sativa L.) seedlings

Abstract: Changes in H 2 O 2 and malondialdehyde (MDA) contents and antioxidant enzyme activities in Cd-treated rice (Oryza sativa L.) seedlings of two cultivars were investigated. On treatment with CdCl 2 , increases in H 2 O 2 and MDA contents and antioxidant enzyme activities (speroxide dismutase (SOD), ascorbate peroxidase, glutathione reductase, catalase, and peroxidase (POX)) were observed in the leaves of Cd-sensitive cultivar (cv. Taichung Native 1, TN1) but not in Cd-tolerant cultivar (cv. Tainung 67, TNG67). The increased content of MDA and activities of SOD and POX preceded the occurrence of toxicity in CdCl 2 -treated TN1 leaves. Pretreatment with abscisic acid (ABA) enhanced Cd tolerance and reduced Cd-induced increase in the content of MDA and increase in the activities of SOD and POX in TN1 leaves. Exogenous application of ABA biosynthesis inhibitor, fluridone, decreased Cd tolerance, increased the content of MDA, and increased the activities of SOD and POX in Cd-treated TNG67 leaves. Furthermore, fluridone's effects on toxicity, the content of MDA, and the activities of SOD and POX in Cd-treated TNG67 leaves were reversed by the application of ABA. In conclusion, the oxidative stress is differently expressed in TN1 and TNG67 rice seedlings in response to CdCl 2 . Results also suggest that CdCl 2 causes an oxidative stress and CdCl 2 -induced toxicity is mediated through oxidative stress in TN1 leaves.

Thidiazuron induced high frequency shoot organogenesis in callus from Kigelia pinnata L.

Abstract: An efficient regeneration system was developed for Kigelia pinnata L., a multipurpose tree belonging to the family Bignoniaceae. The nodal segments were cultured in vitro, and the optimum concentrations of plant growth regulators for callus induction were determined. The friable organogenic calli were derived from the basal cut end of the nodal segments. The highest yield of morphogenic callus (100%) was observed when nodal segments were cul- tured on Murashige and Skoog (MS) medium supplemented with 3 µM 2,4 dichlorophenoxyacetic acid (2, 4-D). The morphogenic callus maintained high regeneration during the first four subcultures in the callus induction medium. The maximum shoots (28/culture) were regenerated at the highest frequency of 100% when 3 µM thidiazuron (N-phenyl N' 1,2,3-thidiazol-5-yl urea) (TDZ) and 0.5 µM naphthaleneacetic acid (NAA) were added to MS medium. The emergence of multiple shoots from the calli was histologically documented. The regenerated shoots showed maxi- mum rooting on ½ MS medium containing 4 µM indole-3- butyric acid (IBA). The effect of vesicular arbuscular mycorrhizae (VAM) association in averting the transplantation shock was tested and proved to be highly beneficial, giving a 100% survival rate after 60 d of transplantation. This efficient plant regeneration system provides a founda- tion for generating transgenic plants of this multipurpose tree.

Purification and Characterization of Carboxymethyl Cellulase from Sinorhizobium Fredii

Abstract: Carboxymethyl cellulase (CMCase) is the key enzyme used by bacteria to decompose the plant root hair-wall during symbiosis. In this study, CMCase activities were routinely assayed using carboxymethyl cellulose (CMC) as a substrate. Coupled with the catalytic reaction of BCA (2,2'-bicinchoninic acid) solution, the color developed at 570 nm was measured to determine the enzyme activity. Sinorhizobium fredii CCRC 15769 was utilized to produce CMCase for this study. The optimum growth conditions of the cultural medium were studied in order to obtain sufficient cells for further CMCase purification, and further, for large-scale fermentation production. BIII medium containing 0.3% myo-inositol proved to be a suitable carbon source for bacterial cell growth and specific activity of CMCase. A 22.4 g wet weight of cells can be harvested from 3.5 liters of fermented medium through continuous centrifugation. The cells were suspended in 50 mM potassium phosphate-citric acid (PCA) buffer at pH 5.2 and disintegrated through ultra-sonication to obtain crude extract. Then, certain precipitates were collected at 40-60% ammonium sulfate saturation via ammonium sulfate fractionation of the crude extract. The supernatant obtained following centrifugation was loaded onto a DEAE Sepharose anion-exchange column, and the active fractions were collected and dialyzed against 10 mM Tris-HCl buffered at pH 7.4. For further purification, the dialyzed fraction was loaded onto a Phenyl-Sepharose column. The active fractions were dialyzed and then assayed by SDS-PAGE activity stain to confirm that it contained CMCase activity at the fraction of 94 kDa. The characterizations of CMCase were demonstrated as follows: the optimal temperature and pH were 35°C and 7.0, respectively. The purification fold was 9.08, and the recovery yield was 26.4%, and the specific activity was 3.822 U mg-1.

Gas Exchange Characteristics of the Invasive Species Mikania Micrantha and its Indigenous Congener M. Cordata (Asteraceae) in South China

Abstract: In South China, the photosynthetic characteristics of an invasive exotic weed Mikania micrantha H.B. Kunth and its indigenous close congener Mikania cordata (Burm. f.) B.L. Robinson were studied. Mikania micrantha exhibited a greater biochemical capacity for photosynthesis than M. cordata, as the former had higher V(subscript cmax) and J(subscript max), which lead to its higher maximum assimilation rate A(subscript max) values. Similar light compensation point, apparent quantum yield (α), and dark respiration (R(subscript d)), suggested a similar shade-tolerance. Higher A(subscript max), light saturating point, and CO2 saturating point give M. micrantha greater potential to acclimate to brighter environments and a larger pool of available carbon. These differences in photosynthetic characteristics could be a basis for their niche partitioning. The leaf traits of the native and invasive Mikania species were as follows; higher construction cost (CC), lower specific leaf area (SLA), as well as lower mass-based leaf N and photosynthetic nitrogen-use efficiency (PNUE) for the native species, but M. micrantha had lower area-based leaf N. The higher water use efficiency (WUE) and δ^13C observed in M.micrantha was probably due to its high photosynthetic capacity and not to reduced stomatal conductance, explaining the absence of any trade-off between WUE and PNUE.

Effect of exogenous indole butyric acid on root formation and peroxidase and indole-3-acetic acid oxidase activities and phenolic contents in date Palm offshoots

Abstract: Exogenous indole butyric acid had a significant positive effect on the rooting response of date Palm (Phoenix dactylifera L.) offshoots. IBA-treated offshoots rooted earlier and with a much higher frequency while untreated offshoots rooted poorly, later, and tended to produce fewer roots. The effect of exogenous application of auxin was also reflected in the metabolic changes in offshoots during the rooting process. Pox activity showed a minimum on the sixth day and a maximum on the twelfth day after the root-inducing treatment. Changes of IAA oxidase activity showed a parallel time curve while phenolic contents and auxin protector levels showed an inverse trend. These parameters changed slightly one week later in the untreated offshoots.

Effect of nitrogen on root and shoot relations and gas exchange in winter wheat

Abstract: The seedling growth of two drought-resistant wheat varieties was studied under solution culture in a plant growth chamber. The results showed that the shoot dry weight and leaf gas exchange parameters increased with the increase of nitrogen supply, but decreased when nitrogen supply reached a certain level. The optimum nitrogen concentrations for shoot dry weight and gas exchange were different among the varieties. The root growth was negatively correlated with the increase of nitrogen supply. The distribution of root length in different layers was similar for the two varieties. The root length was the longest at the layer of 5-15 cm, the shortest below 15 cm, and in between at the layer of 0-5 cm. The water use efficiency (WUE) decreased with increasing ratio of root to shoot (R/S), while leaf photosynthetic rate tended to increase initially and then decrease. The increase in R/S was unfavorable to increase WUE, and the appropriate R/S for leaf photosynthetic rate was about 0.5.

The Role of Diversity and Functional Traits of Species in Community Invasibility

Abstract: The invasion of exotic species into assemblages of native plants is a pervasive and widespread phenomenon. Many theoretical and observational studies suggest that diverse communities are more resistant to invasion by exotic species than less diverse ones. However, experimental results do not always support such a relationship. Therefore, the hypothesis of diversity-community invasibility is still a focus of controversy in the field of invasion ecology. In this study, we established and manipulated communities with different species diversity and different species functional groups (16 species belong to C3, C4, forbs and legumes, respectively) to test Elton's hypothesis and other relevant hypotheses by studying the process of invasion. Alligator weed (Alternanthera philoxeroides) was chosen as the invader. We found that the correlation between the decrement of extractable soil nitrogen and biomass of alligator weed was not significant, and that species diversity, independent of functional groups diversity, did not show a significant correlation with invasibility. However, the communities with higher functional groups diversity significantly reduced the biomass of alligator weed by decreasing its resource opportunity. Functional traits of species also influenced the success of the invasion. Alternanthera sessilis, in the same morphological and functional group as alligator weed, was significantly resistant to alligator weed invasion. Because community invasibility is influenced by many factors and interactions among them, the pattern and mechanisms of community invasibility are likely to be far subtler than we found in this study. More careful manipulated experiments coupled with theoretical modeling studies are essential steps to a more profound understanding of community invasibility.

Evolution of Apocarpy in Alismatidae Using Phylogenetic Evidence from Chloroplast rbcL Gene Sequence Data

Abstract: The apocarpous groups in the monocotyledons are mainly concentrated in the subclass Alismatidae. The molecular phylogeny of Alismatidae based on analysis of chloroplast rbcL gene sequence data serves as a framework with which to evaluate character evolution with respect to the derivation of apocarpy in the group. 20 of the 27 genera in the subclass that display apocarpy have been included in our study. Our analysis indicates that apocarpy is polyphyletic within the subclass Alismatidae. Two independent origins of apocarpy in Alismatidae are explored in this study. Three separate origins of a single carpel and two separate origins of syncarpy in the subclass are also proposed. Basally connate carpel condition was the ancestral character in Alismatidae and evolved in two directions. It is possible for the unicarpellate condition to have been directly derived by reduction from syncarpy, and it could also be that the unicarpellate state has been derived from apocarpy by reduction in carpel number. The present results indicate that a progression has occurred in the evolution of carpels in Alismatidae from basally connate carpels through syncarpy or apocarpy to a single carpel.

Statistical approaches on discriminating spatial variation of species diversity

Abstract: This study applied statistical approaches to the discrimination of spatial variations between sites and between forest types in the upper area of the Liukuei Experimental Forest of Taiwan Forestry Research institute, Taiwan The main purpose was to compare the effectiveness of various statistical approaches and then present the best strategy for discriminating the spatial variations of species diversity The two methods used were (1) univariate methods by diversity measures, Shannon t-test, and (2) multivariate methods by cluster analysis, ordination by non-metric multi-dimensional scaling, and principal component analysis The results by univariate methods indicate that diversity differences exist between sites and between forest types Meanwhile, the natural forest has more diversity than the plantation, and the hardwood plantation has more diversity than the conifer plantation The differences between forest types are very significant at the 100 significance level according to the Shannon t-test The results indicate that univariate methods by diversity measures are a flexible way to reduce the complexity of ”species by sites” matrices into a single coefficient The results of using multivariate methods indicate that cluster analysis and ordination by non-metric multi-dimensional scaling and principal component analysis are useful techniques for discriminating spatial variations However, ordination by non-metric multi-dimensional scaling discriminates better than principal component analysis in addition, ordination by non-metric multi-dimensional scaling is a more informative summary than cluster analysis, and the combination of both the analyses is more effective than either alone for the mutual consistency of representations it is concluded that the most powerful tools for discriminating the spatial variations of species diversity are in the multivariate category Among multivariate methods, ordination by non-metric multidimensional scaling is preferable, and its superimposition with cluster analysis is recommended in order to obtain more information regarding the relationship between sites and between forest types

Phylogenetic Relationships of Monascus Species Inferred from the ITS and the Partial β-tubulin Gene

Abstract: ITS and partial β-tubulin genes of 17 ATCC reference strains of Monaccus species were PCR amplified and sequenced. Monascus pilosus and M. ruber could not be differentiated with these sequences, suggesting a synonymy. In maximum parsimony analyses on both data sets, M. ruber, M. pilosus, M. purpureus, and M. sanguineus were placed into the same clade. ITS sequence alignment revealed a number of gaps in ITS1 and ITS2 of M. pallens, M. lunisporas, and M. eremophilus compared to M. purpureus, M. ruber, and M. pilosus. Accordingly, analyses with the ITS sequences placed these species into clades, incongruent with the analyses using the partial β-tubulin genes and the previous results with the partial large subunit rRNA genes. The phylogenetic relationship derived from the partial β-tubulin genes was similar to those postulated by the 5'-partial LSU rRNA genes. This finding strongly suggests that evolutionary or phylogenetic classification with ITS sequence information should be performed with caution. in the phylogenetic trees with the ITS sequences, M. lunisporas was distantly associated with Aspergillus ustus; M. pallens was placed in a clade that shares a common node with A. versicolor; and M. eremophilus was placed on a branch separate from the M. purpureus, M. ruber and M. pilosus group while M. pallens and M. lunisporas were placed into the related clades sharing a common node in the tree derived from the partial β-tubulin gene. Each of the phylogenetic analyses with the partial β-tubulin genes, the ITS, or the 5-end of the LSU rRNA, as previously carried out, placed M. eremophilus into a different lineage. Molecular analyses with these molecular targets generated three different topologies for M. eremophilus, indicating a unique and unpredictable genetic combination for this species. It might reflect extreme environmental stress on this species and subsequent genetic changes.

Effects of Cryptococcus laurentii (Kufferath) Skinner in combination with sodium bicarbonate on biocontrol of postharvest green mold decay of citrus fruit

Abstract: The potential of using Cryptococcus laurentii alone or in combination with sodium bicarbonate solution for control of Penicillium digitatum (green mold) on oranges was investigated. Agar disks of C. laurentii NYDA cultures placed on PDA plates seeded with pathogen did not inhibit the growth of P. digitatum. Spore germination of P. digitatum in PDB was significantly controlled in the presence of living C. laurentii cell suspensions. Cryptococcus laurentii significantly controlled green mold on oranges after challenge with 5 × 104 spores/ml of P. digitatum. The higher the concentrations of the antagonist, the lower the disease incidence regardless of whether the fruit was stored at 20°C for 5 days or 2°C for 50 days. The efficacy of C. laurentii for control of green mold was improved when combined with sodium bicarbonate. The combination of sodium bicarbonate and C. laurentii could be an alternative to fungicides for control of postharvest green mold disease on citrus fruits.

Comparison of Aspergillus Flavus and Aspergillus Oryzae by Amplified Fragment Length Polymorphism

Abstract: The aflatoxin producer Aspergillus flavus and the koji mold Aspergillus oryzae are morphologically similar species that belong to the Aspergillus section Flavi. We used amplified fragment length polymorphism (AFLP) to differentiate these two species. In this study, we tested thirteen A. flavus, nine A. oryzae, and three A. flavus var. columnaris strains. DNA fragment profiles amplified with each of three selective primer pairs displayed similar patterns for the various A. flavus strains. Different patterns were observed with these primer pairs for the A. oryzae strains. We combined these data to increase the grouping of the various strains within each species and to distinguish between A. flavus and A. oryzae. By unweighted pair group method with arithmetic average (UPGMA) analysis, the AFLP data obtained from the three selective primer pairs, EcoRI-AC/MseI-CAT, EcoRI-TA/MseI-CAT, and EcoRI-TA/MseI-CTT, differentiated A. flavus from A. oryzae successfully. Three strains of A. flavus received from ATCC grouped outside of the other A. flavus strains we examined. The morphology of these isolates and our results indicated those strains were originally misidentified and they should be classified as A. parasiticus. We found that the AFLP technique is a reliable and reproducible tool with the ability to differentiate A. flavus from A. oryzae and should be generally useful in distinguishing between closely related species or strains

Effects of Partial Ringing and Heating of Trunk on Shoot Growth and Fruit Quality of Peach Trees

Abstract: Small-sized peach (Prunus persica [L.] Batsch.) trees for commercial fruit production have not been available due to the lack of suitable dwarfing rootstocks that have a wide range of compatibility among cultivars. Effects of partial girdling and trunk heating on the growth, yield, and fruit quality of peach were studied. A 4 cm wide partial ring of bark was removed at a height of 25 cm from the ground leaving a connecting strip of 5 mm. Furthermore, the dc-barked spot was heated at 150℃ for fifteen minutes using an electric heater. The partial girdling was effective in reducing shoot growth and improving fruit quality. The total soluble solids content of fruit in treatment trees was higher than that of control. The heating had little additive effects over partial ringing.

Comparisons of allozyme variation of narrow endemic and widespread species of Far East Euphorbia (Euphorbiaceae)

Abstract: Genetic variation from 19 populations representing four species of Euphorbia was compared using isozyme markers. Euphorbia fauriei and E. garanbiensis are rare endemic species in Korea and Taiwan, respectively while E. pekinensis and E. jolkinii are widespread in Korea and Taiwan. A low level of genetic variation and high genetic differentiation among populations was found in E. garanbiensis, which has a restricted distribution, small population sizes, gravity-dispersed seeds, and is possibly a self-pollinator. Alternatively, E. garanbiensis may have recently evolved from a continental progenitor or may be a relictual lineage from the continental flora. The high genetic diversity found in E. fauriei is unusual and may be explained by its recent origin from a widespread continental progenitor, E. pekinensis, An alternative explanation is that E. fauriei survived in refugia during the last glaciation in the Korean peninsula. The idea of progenitor-derivative species pairs between E. pekinensis and E. fauriei is supported by the high genetic identity values between the two species and the putative derivative E. fauriei having no unique allele and lacking some rare alleles present in the putative progenitor E. pekinensis.

Asymbiotic and Symbiotic Seed Germination of Anoectochilus Formosanus and Haemaria Discolor and Their F1 Hybrids

Abstract: Two orchid species Anoectochilus formosanus Hayata and Haemaria discolor var. dawsoniana, which have different flowering times in nature, were induced to flowering synchronously by controlling the culture temperature and light duration in the phytotrons. They were crossed, and the F1 hybrid seeds were collected and named as H. discolor × A. formosanus (HA) and A. formosanus × H. discolor (AH). It was found that fifty days after the asymbiotic culture in MS medium, the germination rates of all the four kinds of seed were higher than 70%. For the symbiotic seed germination on oatmeal agar (OMA) medium, only the R02, as compared to the R01 and R04 isolates of Rhizoctonia spp., had a more than 80% germination rate. SEM observation of various seed germination stages for the non-mycorrhizal and the mycorrhizal protocorms of A. formosanus showed that 70 days after seed sowing, the inoculated seeds developed to a more advanced stage than the non-inoculated control. Asymbiotic and symbiotic germination occurred after the uptake of water, and the seed coat was ruptured by the enlargement of embryo. Afterward the papilla, protocorm and apical meristem appeared. In the symbiotic germinated embryos, the fungal hyphae penetrated the protocorm and formed pelotons, which was the tolypophagy type of infection. No hypha infection was found in the asymbiotic germinated embryos.

Antioxidant Activities of Methanolic and hot-water Extracts from Leaves of Three Cultivars of Mai-Men-Dong (Liriope spicata L.)

Abstract: 1,1-dipheny-2-picrylhydrazyl (DPPH) scavenging activities of the 80% methanolic leaf extracts of three cultivars (small leaf, SL; big leaf, BL; thin leaf, TL) of Mai-Men-Dong (Liriope spicata L.) are analyzed by spectrophotometry. The concentrations required for 50% inhibition (IC50) of DPPH radicals were 81.08, 96.97, and 53.78μg/mL, respectively. The methanolic extracts were further partitioned into three n-hexane-, ethylacetate-, and water-soluble fractions, among which the ethylacetate-soluble fraction exhibited the highest DPPH scavenging activity. The IC50 of ethylacetate-soluble fractions of SL, BL, and TL for DPPH radical scavenging activity were 41.55, 24.55, and 53.33μg/mL, respectively. Each Mai-Men-Dong powder (1g) was deposited in a tea bag and then dipped in hot water (100℃, 100mL) for 3 min with triplicate samples. These hot-water extracts were then freeze-dried for an anti-DPPH radical capacity test, which found a positive correlation with the phenolic contents of each hot water extract. The IC50 of hot water extracts of SL, BL, and TL for DPPH radical scavenging activities were 378.97, 171.12, and 95.84mg/mL, respectively. All three hot water extracts can effectively scavenge hydroxyl radical using electron spin resonance (ESR) spectrometry. The IC50 against hydroxyl radical were 80.8, 69.7, and 116μg/mL, respectively, for the SL, BL, and TL cultivars.

Effects of Cryptococcus laurentii (Kufferath) Skinner on biocontrol of postharvest decay of arbutus berries

Abstract: Cryptococcus laurentii was tested as a biocontrol agent for reducing natural decay of arbutus berries caused by Penicillium citrinum and Verticicladiella abielina in semi-commercial postharvest trials. Three different preparations of C. laurentii were compared for antagonistic efficiency. Washed C. laurentii cells provided better protection against decay than yeast cultured in broth without washing while the culture supernatant free of yeast cells provided no protection. The protection provided by the washed yeast cells was dose-dependent. Cryptococcus laurentii was also effective in controlling decay at low temperature (4°C). The efficacy of C. laurentii was enhanced by the addition of 2% CaCl2. Agar disks of C. laurentii NYDA cultures placed on PDA plates seeded with pathogens did not inhibit the growth of P. citrinum or V. abielina. Spore germination of the pathogens in potato dextrose broth was strongly inhibited in the presence of active cell suspensions.

Begonia coptidifolia (Begoniaceae), a new species from China

Abstract: Begonia coptidifolia H. G. Ye, F. G. Wang, Y. S. Ye & C.-I Peng, a new species from Guangdong province, China, is here described and illustrated. Its chromosome number (2n=22) and karyotype are reported. Begonia coptidifolia belongs to sect. Platycentrum and somewhat resembles B. hemsleyana Hook. f., from which it differs by the aerial stems seen only at anthesis (vs. all year round), palmately trisected leaves with pinnately parted lobules (vs. leaves palmately compound with 7-9 petiolulate leaflets) and petioles grooved on adaxial surface (vs. terete). The new species also resembles Begonia pedatifida H. Lev., which, however, has much stouter rhizomes (2-6 cm vs. 0.3-0.5 cm thick), leaves plmatifid with triangular lobules, and terete petioles. Begonia coptidifolia is rare, known only from a small ravine in Ehuangzhang Nature Reserve, Yangchun, Guangdong Province.

The occurrence of extrafloral nectaries in Hong Kong plants

Abstract: This is a study of the extrafloral nectarines in Hong Kong plants. Five major types can be discerned: button-shaped, cup-shaped, stalk-shaped, pit-shaped, and pore-shaped. Euphorbiaceae is the largest family with extrafloral nectaries which are always visible structures, attracting ants. SEM micrographs of extrafloral nectarines are included.

Phenotypic Plasticity of Mosla Chinensis and M. Scabra (Labiatae) Response to Soil Water Status

Abstract: The growth and architectural plasticity of Mosla chinensis Maxim. in response to soil water status were compared with the congeneric plant, Mosla scabra (Thunb.) C. Y. Wu et H. W. Li. Two-week-old seedlings were exposed to five levels of soil water for a 6-week period. The results indicated that: an individual’s total mass, root mass, apical height, basal diameter, accumulative branch length and branch fresh weight/dry weight ratio (FW(subscript B) /DW(subscript B)) of both species had high plasticity in response to soil water content (p＜0.05), and the plasticity of these traits in M. scabra is mostly higher than in M. chinensis. The leaf mass ratio (LMR), specific leaf area (SLA), root mass ration (RMR), and root/shoot ratio (R/S) of both species had low plasticity. Furthermore, leaf mass, branch mass, branch mass ratio (BMR). and branch length ratio (BLR) had high plasticity (P＜0.05) in M. chinensis but not in M. scabra (P＞0.05) while branch number exhibited contrary trends. In response to soil water, M. scabra adjusted the traits of total mass and size, in terms of a bigger PI, more than M. chinensis while M. chinensis only adjusted partial branch and root traits, such as BMR, BLR, FW(subscript B)/DW(subscript B), RMR and R/S, more than M. scabra. The optimum water niches (OWN) of both M. chinensis and M. scabra are from 40% soil water holding capacity (WHC) to constant saturation, but M. chinensis is only found in relatively dry environments while M. scabra is distributed from dry to wet environments in the field, so the actual water niche (AWN) was separated from the OWN in M. chinensis, but not in M. scabra. Mosla chinensis grew slower and remained smaller than M. scabra and other neighbor species in the field, and it therefore had no competitive superiority in the community. Mosla scabra was very competitive because of its higher yield and taller growth.

Heat shock proteins of thermophilic and thermotolerant fungi from Taiwan

Abstract: Fifteen species of thermophilic and four species of thermotolerant fungi were isolated from soil samples collected at various localities in Taiwan. Fourteen species and three strains responded to three h of thermal stress at elevated temperature from 30℃ to 50℃ by synthesizing 30 heat shock proteins (HSPs) with molecular weights ranging between 20-150 kDa. Heat shock treatments at 40℃, resulted in the synthesis of 22 HSPs with molecular weights of 30-150 kDa. Nine of the seventeen fungal species produced a 46 kDa HSPs, seven species a 52 kDa HSP, and five species a 94 kDa HSP. Four of the seventeen species produced two kinds of HSPs with molecular weights of 87 kDa and 40 kDa. Ten fungal strains responded to an elevated temperature of 50℃ and synthesized 12 HSPs of molecular weights ranging from 20 to 92 kDa. Six species produced a 35 kDa HSP, and four species produced HSPs with molecular weights of 30 kDa, 28 kDa and 22 kDa. When subjected to heat shock treatments at 40℃, thermophilic and thermotolerant fungi synthesized mostly high and medium molecular weight HSPs while at 50℃, they synthesized mostly low molecular weight HSPs.

Purification and characterization of isoforms of β-N- acetylhexosaminidase from mungbean seedlings

Abstract: Three isoforms of β-N-acetylhexosaminidase (β-NAHA), named β-NAHAs Ⅰ, Ⅱ and Ⅲ, were isolated from six-day-old etiolated mungbean (Vigna radiata) seedlings β-NAHA Ⅰ was purified to apparent homogeneity by a procedure involving Con A-Sepharose chromatography, chromatofocusing, and gel filtration β-NAHAs Ⅱ and Ⅲ were highly purified β-NAHAs Ⅰ, Ⅱ and Ⅲ had molecular masses of 135, 127 and 110kDa, respectively β-NAHA Ⅰ was dissociated into a single 67kDa protein band Ⅱ was dissociated into two protein bands corresponding to 60 and 48kDa, and Ⅲ was dissociated into a single 48kDa protein band in SDS-polyacrylamide gel electrophoresis The results suggest that isoforms Ⅰ and Ⅲ are homodimeric enzymes, each comprising two identical subunits with molecular masses of 67kDa and 48kDa, respectively, while isoform Ⅱ is a heterodimeric enzyme, comprising two non-identical subunits with molecular masses of 60kDa and 48kDa All the enzymes were active against paranitrophenyl-β-N-acetylglucosaminide (PNP-β-N-acetylglucosaminide) and PNP-β-N-galactosaminide The enzymes were inhibited by 5,5'-dithiobis (2-nitrobenzoic acid)(DTNB), Ag(superscript +), Hg(superscript 2+), and N,N'-diacetylchitobiose Km values for isoforms Ⅰ, Ⅱ and Ⅲ were 067mM, 104mM and 176mM, respectively, using PNP-β-N-acetylglucosaminide as a substrate These three isoforms had acidic p1 values (Ⅰ, 63; Ⅱ, 61; and Ⅲ, 59) Their optimal pH in the reaction towards PNP-β-N-acetylglucosaminide was 54, 47 and 57, and optimal temperatures were 65℃, 65℃ and 50℃ for isoforms Ⅰ, Ⅱ and Ⅲ, respectively

Astrosphaeriella from Taiwan, Including Two New Species

Abstract: A modified generic concept is adopted in the study of ascomycetes referred to Astrosphaeriella from Taiwan Seven species, including two new species, are described and illustrated A key to the Taiwan species is also provided Those species with striate ascospores are found to constitute a distinct group within Astrosphaeriella and can readily be distinguished from the other species in Astrosphaeriella