Plant quality and predation risk mediated by plant ontogeny: consequences for herbivores and plants

Bottom‐up and top‐down impacts on herbivores can be influenced by plant productivity, structural complexity, vigor and size. Although these traits are likely to vary with plant development, the influence of plant ontogeny on the relative importance of plant quality (i.e. bottom‐up forces) and predation risk (i.e. top‐down forces) has been the focus of little previous investigation. We evaluated the role of plant ontogeny for the relative importance of bottom‐up and top‐down forces on insect herbivore abundance, species richness, and species diversity attacking the tropical tree Casearia nitida. We also quantified the cascading effects on herbivory, growth and reproduction of this plant species. Plant quality traits (nitrogen and phenolic compounds) were assessed in saplings and reproductive trees. Bottom‐up forces were manipulated by fertilizing plants from both ontogenetic stages. Top‐down forces were manipulated by excluding insectivorous birds from saplings and reproductive trees. Plant ontogeny influenced foliage quality in terms of total phenolics, which were in greater concentration in reproductive trees than in saplings; however, it did not influence bottom‐up forces as modified by fertilization. Bird exclusion increased herbivore density with the same magnitude on both stages. Ontogeny influenced species diversity, which was greater in reproductive trees than in saplings, and also influenced treatment impacts on species richness and diversity. Although top‐down forces increased herbivory equally on plants of each ontogenetic stage, the two stages showed different overcompensation responses to increased damage: caged saplings produced greater leaf biomass than non‐caged saplings, whereas caged trees increased in height proportionally more than non‐caged trees. In sum, plant ontogeny influenced the impact of bird predation on herbivore density, species richness, and species diversity, and the growth variables affected by increased damage in caged plants. We suggest that plant ontogeny can contribute to some extent to the influence of plant quality and the third trophic level on herbivores in this system.     

Modifications in dispersal and oviposition of Bt-resistant and Bt-susceptible Colorado potato beetles as a result of exposure to Bacillus thuringiensis subsp. tenebrionis Cry3A toxin

Laboratory strains of Colorado potato beetle, Leptinotarsa decemlineata (Say), physiologically resistant and susceptible to Bacillus thuringiensis (Berliner) subsp. tenebrionis Cry3A toxin were reared to adults on caged potato plants. Influence of three different diets (transgenic potatoes, regular potatoes, and regular potatoes followed by the transgenic potatoes) on beetle mortality, fecundity, and flight behavior were tested under laboratory conditions. A computer-linked flight mill system was used to quantify beetle flight, and dissections were performed to determine the level of flight muscle development. Susceptible beetles continuously fed on transgenic foliage suffered heavy mortality, did not develop flight muscles, and did not produce any eggs. Resistant beetles continuously fed on transgenic foliage were capable of flight and reproduction; however, it took them longer to initiate flight behavior, and their fecundity was lower than fecundity of other treatments. In both strains, detrimental effects became significantly less severe when the beetles were allowed to feed on regular foliage prior to toxin ingestion. In the resistant strain, ingestion of Cry3A toxin significantly increased flight activity, indicating that physiological resistance was probably reinforced by the behavioral escape from toxic environments. No such response was observed for susceptible beetles. When fed on regular foliage, resistant Colorado potato beetles engaged in significantly fewer flights than susceptible beetles. Behavioral differences between resistant and susceptible beetles observed in the present study are likely to affect gene flow between transgenic crops and adjacent refugia, and should be taken in consideration when designing resistance management plans for transgenic potato crops.     

Localized outbreak of a willow leaf beetle: plant vigor or natural enemies?

The possible roles of plant quality (vigor) and natural enemies in the development of a localized out-break of the leaf beetle Galerucella lineola (Coleoptera: Chrysomelidae) in a stand of Salix cinerea were investigated. Caged and uncaged larvae on six bushes in the outbreak area were compared with caged and uncaged larvae on six bushes in an adjacent nonoutbreak area in terms of performance. In 1997, when the studies were performed, the natural density of the insect (beetles plus eggs) was six times higher in the outbreak area compared with the nonoutbreak area. Even though the vigor (measured as shoot length) of bushes in the outbreak was 72% higher than that of bushes in the nonoutbreak area, we found no difference between areas in the performance (survival, developmental time, pupal weight) of caged larvae or in the willingness of caged females to lay eggs. Among larvae exposed to natural enemies, the disappearance rate was significantly higher in the nonoutbreak area. The density of generalist predators was significantly higher in the nonoutbreak than in the outbreak area. We conclude that differences in plant quality, despite the observed difference in plant vigor, could not explain the observed difference in beetle density between areas. Lower predation pressure in the outbreak area could, however, not be excluded as a possible reason for the higher density of leaf beetles in this area. Received: October 18, 1999 / Accepted: February 4, 2000     

Plant regeneration and stable transformation in the floricultural plant Cleome spinosa, a C3 plant closely related to the C4 plant C. gynandra

Cleome spinosa is widely used as a garden ornamental in many countries. Here we determined the optimal conditions for plant regeneration from different tissue explants grown in vitro. Induction medium containing MS salts, MS vitamins, 3% sucrose, 1 mg l?1 BA, 200 mg l?1 timentin, and 0.8% agar was sufficient for shoot regeneration of all the tissue explants examined, including leaf, hypocotyl, and cotyledon. Subsequently, an Agrobacterium tumefaciens-mediated method was developed to transform the vector pCHS, which carries the transgenes Petunia chalcone synthase (chs) and selection marker neomycin phosphotransferase II (nptII), into C. spinosa. From a total of 368 cotyledon explants, 13 putative transgenic lines were regenerated from selection medium supplemented with 50 mg l?1 kanamycin and 200 mg l?1 timentin, and transferred to the greenhouse. Genomic PCR and Southern blot analyses revealed that the nptII transgene was present in all 13 transgenic plants. Similarly, when the Petunia chs transgene was used as a probe in Southern blot analysis, single or multiple hybridization bands were detected in 12 out of the 13 transgenic plants. In addition, T? progeny assay from selected transformants showed that the nptII transgene can be transmitted in a Mendelian manner from transgenic parents into their progeny. This is the first report of stable transformation of the C? dicotyledon C. spinosa, which will facilitate functional comparison of cell-type specific genes with counterpart C? dicotyledon C. gynandra using transgenic approaches.     

Effects of social conditions on adult and subadult female rat-like hamsters (<Emphasis Type="Italic">Cricetulus triton</Emphasis>)

As a solitary species, rat-like hamsters (Cricetulus triton) still live in family groups before they become mature and leave their families for a solitary life. This study aimed to investigate by a laboratory experiment if housing conditions have a different effect on physiological aspects of immature and mature females. We found that paired caged adult females became significantly heavier than their original weights; whereas the singly caged did not show significant change in their body weight. Although the subadults body weights increased significantly compared to their initial weights in both paired or singly caged groups, significant changes in body weight did not occur between the two groups. Although spleen and adrenal gland sizes were not significantly different between the two adult groups, the cortisol levels were significantly elevated by paired caging. In subadults, the adrenal size of the singly caged group was larger than that in the paired caged group despite there being no significant difference in cortisol level. Flank glands became significantly larger in paired caged adults than in singly caged adults, and there were no significant differences in subadults between the two groups. Additionally, ovaries and uteri of the paired caged adult females were comparatively lighter than those of the singly caged group; in contrast, ovaries and uteri of the paired caged group were larger than those of the singly caged group in subadults, although progesterone and estradiol levels did not show significant differences between the two adult groups. These different changes in physiological traits caused by housing conditions indicated that paired caging depressed adults and facilitated subadults; isolation facilitated adults and depressed subadults.     

Thirty years of plant transformation technology development

Technology development is seminal to many aspects of basic and applied plant transgenic science. Through the development and commercialization of genetically modified crops, the evolution of plant transgenic technologies is also relevant to society as a whole. In this study, literature statistics were used to uncover trends in the development of these technologies. Publication volume and impact (citation) over the past 30 years were analysed with respect to economic zones, countries, species and DNA delivery method. This revealed that, following a dramatic expansion in the 1980s, publications focusing on the development of transgenic technology have been slowing down worldwide since the early mid-1990s, except in a few leading Asian countries. The implications of these trends on the future of plant transgenic science as a whole are discussed.     

A recently acquired host plant provides an oligophagous insect herbivore with enemy-free space

Enemy-free space (EFS) is a potentially important factor affecting host plant use by phytophagous insects. Yet only a few field studies have demonstrated that natural enemy activity is the sole mechanism underlying use of novel host plants by herbivorous insects. This may be due to the fact that in earlier studies, both herbivores and natural enemies had the opportunity to adapt to the new host plant. Here we studied the possibility that EFS underlies the recently recorded increase in Phthorimaea operculella densities on tomato plants in a few areas within its geographical range. Through field experiments in Ethiopia, we show that all three conditions proposed by Berdegue et al. to demonstrate EFS are fulfilled. First, a significantly higher proportion of larvae survive on caged than on exposed potato plants, showing that natural enemies are an important mortality factor on the original host, potato. Second, larval survival was significantly higher on exposed tomato than potato plants, implying greater protection for the herbivore from its natural enemies on tomato than on potato plants. Thus tomato plants provide P. operculella with an EFS. Finally, larval survival was significantly higher on caged potato than on caged tomato plants at the preblossom stage, indicating that, in the absence of natural enemies, there is a fitness cost when larvae feed on the sub-optimal tomato plants. Fulfillment of this third condition points to the importance of natural enemy activity relative to that of other unidentified factors, such as food quality and competition. An intensive field survey provides further support for this conclusion.     

The influence of caging conditions and hormone treatments on fighting in male and female hamsters

After gonadectomy, more individually caged female hamsters fought prior to the initiation of hormone treatments than did group-caged females. Daily injections of testosterone propionate (TP), estradiol benzoate (EB), or progesterone (Prog) had no influence on the number of individually caged females that fought. However, TP and EB were effective in increasing the number of group-caged females that fought. In contrast to females, both individually and group-caged males fought infrequently after castration. Daily injections of TP, EB, or Prog were effective in increasing the number of individually caged males that fought, while only TP and EB were effective in group caged males. Prog failed to increase the number of group-caged hamsters of either sex that fought.     

基因工程植物的安全性问题

转基因植物的研究进展很迅速,但基因工程植物是否安全一直争论不休,主要表现在转基因食品的安全性及生态安全性问题上.转基因食品的安全性涉及这些食品的过敏性、毒性以及抗生素标记基因的安全性几个方面.转基因植物的生态安全性包括基因漂流、是否能诱发昆虫产生Bt抗性和对生物多样性的影响等.本文针对这些问题,对转基因植物潜在危害以及国际上现有的评价作简要综述.     

Design and synthesis of photolabile caged cytokinin

Cytokinins are phytohormones that regulate diverse developmental processes throughout the life of a plant. trans-Zeatin, kinetin, benzyladenine and dihydrozeatin are adenine-type cytokinins that are perceived by the AHK cytokinin receptors. Endogenous cytokinin levels are critical for regulating plant development. To manipulate intracellular cytokinin levels, caged cytokinins were designed on the basis of the crystal structure of the AHK4 cytokinin receptor. The caged cytokinin was photolyzed to release the cytokinin molecule inside the cells and induce cytokinin-responsive gene expression. The uncaging of intracellular caged cytokinins demonstrated that cytokinin-induced root growth inhibition can be manipulated with photo-irradiation. This caged cytokinin system could be a powerful tool for cytokinin biology.     

植物转基因技术的诞生和发展

在过去的几十年里,转基因技术经历了从诞生、成长到大规模商业化应用等阶段,在农业生产中表现出巨大优势和潜力。本文简单介绍转基因技术的诞生、发展历程以及产业化情况。     

Agrobacterium-mediated transformation and plant regeneration of Brassica oleracea var. capitata

An efficient protocol for Agrobacterium tumefaciens-mediated transformation of four commercial cultivars of Brassica oleracea var. capitata is described. A strain of A. tumefaciens LBA4404 with the neomycin phosphotransferase gene (nptII) and a CaMV 35S-peroxidase gene cassette were used for co-cultivation. Preliminary selection of regenerated transgenic plants was performed on kanamycin-containing medium. The frequency of transgenic plants was calculated on the basis of GUS (β-glucuronidase) activity detected by the histochemical X-gluc test. Tissue-specific GUS expression driven by the peroxidase gene promoter in transgenic plants was analysed by GUS staining. The transformation rates of the commercial cultivars of B. oleracea was higher than in previous reports. Southern blot analysis revealed that integration of marker genes occurred in single and multiple loci in the genome. All transgenic plants grew normally after a brief vernalization period and showed stable inheritance of the marker gene. The present study demonstrates that morphologically normal, fertile transgenic plants of B. oleracea can be obtained. Received: 24 August 1999 / Revision received: 23 November 1999 / Accepted: 3 December     

Molecular tailoring of farnesylation for plant drought tolerance and yield protection

Protecting crop yield under drought stress is a major challenge for modern agriculture. One biotechnological target for improving plant drought tolerance is the genetic manipulation of the stress response to the hormone abscisic acid (ABA). Previous genetic studies have implicated the involvement of the beta-subunit of Arabidopsis farnesyltransferase (ERA1) in the regulation of ABA sensing and drought tolerance. Here we show that molecular manipulation of protein farnesylation in Arabidopsis, through downregulation of either the alpha- or beta-subunit of farnesyltransferase enhances the plant's response to ABA and drought tolerance. To test the effectiveness of tailoring farnesylation in a crop plant, transgenic Brassica napus carrying an ERA1 antisense construct driven by a drought-inducible rd29A promoter was examined. In comparison with the non-transgenic control, transgenic canola showed enhanced ABA sensitivity, as well as significant reduction in stomatal conductance and water transpiration under drought stress conditions. The antisense downregulation of canola farnesyltransferase for drought tolerance is a conditional and reversible process, which depends on the amount of available water in the soil. Furthermore, transgenic plants were more resistant to water deficit-induced seed abortion during flowering. Results from three consecutive years of field trial studies suggest that with adequate water, transgenic canola plants produced the same amount of seed as the parental control. However, under moderate drought stress conditions at flowering, the seed yields of transgenic canola were significantly higher than the control. Using protein farnesyltransferase as an effective target, these results represent a successful demonstration of engineered drought tolerance and yield protection in a crop plant under laboratory and field conditions.     

''Caged cytoskeletons'': a rapid method for the isolation of microtubule-associated proteins from synchronized plant suspension cells

In the cytoskeleton method for isolating microtubule-associated proteins MAP65, DcKRP120-1 and DcKRP120-2, carrot cells are first converted to protoplasts but this method cannot be used to isolate mitotic MAPs as mitotic synchrony is eroded during lengthy cellulase treatment. Anti-microtubule cycle blocks would also be unsuitable. We report here a method for overcoming these problems. Cellulase degradation of tobacco BY-2 cells for only several minutes allows extraction of detergent-soluble proteins, leaving synchronized "caged cytoskeletons" for depolymerization and enabling affinity purification of MAPs on neurotubules. This rapid and simple method should be of general utility: it can be bulked up, avoids anti-microtubule blocks, and is applicable to other cell suspensions. The effectiveness of the caged cytoskeleton method is demonstrated by comparing known MAPs (the 65 kDa structural MAPs and the kinesin-related protein, TKRP125) in synchronized cells taken at the mitotic peak with those in unsynchronized cells.     

Antisense repression of sucrose phosphate synthase in transgenic muskmelon alters plant growth and fruit development

To unravel the roles of sucrose phosphate synthase (SPS) in muskmelon (Cucumis melo L.), we reduced its activity in transgenic muskmelon plants by an antisense approach. For this purpose, an 830 bp cDNA fragment of muskmelon sucrose phosphate synthase was expressed in antisense orientation behind the 35S promoter of the cauliflower mosaic virus. The phenotype of the antisense plants clearly differed from that of control plants. The transgenic plant leaves were markedly smaller, and the plant height and stem diameter were obviously shorter and thinner. Transmission electron microscope observation revealed that the membrane degradation of chloroplast happened in transgenic leaves and the numbers of grana and grana lamella in the chloroplast were significantly less, suggesting that the slow growth and weaker phenotype of transgenic plants may be due to the damage of the chloroplast ultrastructure, which in turn results in the decrease of the net photosynthetic rate. The sucrose concentration and levels of sucrose phosphate synthase decreased in transgenic mature fruit, and the fruit size was smaller than the control fruit. Together, our results suggest that sucrose phosphate synthase may play an important role in regulating the muskmelon plant growth and fruit development.     

Cytokinin-induced activity of antioxidant enzymes in transgenic Pssu-ipt tobacco during plant ontogeny

Cytokinin (CK) content and activities of several antioxidant enzymes were examined during plant ontogeny with the aim to elucidate their role in delayed senescence of transgenic Pssu-ipt tobacco. Control Nicotiana tabacum L. (cv. Petit Havana SR1) and transgenic tobacco with the ipt gene under the control of the promoter of small subunit of Rubisco (Pssu-ipt) were both grown either as grafts on control rootstocks or as rooted plants. Both control plant types showed a decline in total content of CKs with proceeding plant senescence. Contrary to this both transgenic plant types exhibited at least ten times higher content of CKs than controls and a significant increase of CK contents throughout the ontogeny with maximal values in the later stages of plant development. Significantly higher portion of O-glucosides was found in both transgenic plant types compared to control ones. In transgenic plants, zeatin and zeatin riboside were predominant type of CKs. Generally, Pssu-ipt tobacco exhibited elevated activities of antioxidant enzymes compared to control tobacco particularly in the later stages of plant development. While in control tobacco activity of glutathione reductase (GR) and superoxide dismutase (SOD) showed increasing activity up to the onset of flowering and then gradually decreased, in both transgenic types GR increased and SOD activity showed only small change throughout the plant ontogeny. Ascorbate peroxidase (APOD) was stimulated in both transgenic types. The manifold enhancement of syringaldazine and guaiacol peroxidase activities was observed in transgenic grafts throughout plant ontogeny in contrast to control and transgenic rooted plants, where the increase was found only in the late stages. Electron microscopic examination showed higher number of crystallic cores in peroxisomes and abnormal interactions among organelles in transgenic tobacco in comparison with control plant. The overproduction of cytokinins resulted in the stimulation of activities of AOE throughout the plant ontogeny of transgenic Pssu-ipt tobacco.     

Cytokinin vectors mediate marker-free and backbone-free plant transformation

Conventional Agrobacterium-mediated transformation methods rely on complex and genotype-specific tissue culture media for selection, proliferation, and regeneration of genetically modified cells. Resulting transgenic plants may not only contain selectable marker genes but also carry fragments of the vector backbone. Here, we describe a new method for the production of transgenic plants that lack such foreign DNA. This method employs vectors containing the bacterial isopentenyltransferase (ipt) gene as backbone integration marker. Agrobacterium strains carrying the resulting ipt gene-containing "cytokinin" vectors were used to infect explants of various Solanaceous plant species as well as canola (Brassica napus). Upon transfer to hormone-free media, 1.8% to 9.9% of the infected explants produced shoots that contained a marker-free T-DNA while lacking the backbone integration marker. These frequencies often equal or exceed those for backbone-free conventional transformation.     

Feeding damage by Bagrada hilaris (Hemiptera: Pentatomidae) and impact on growth and chlorophyll content of Brassicaceous plant species

Bagrada hilaris Burmeister is an invasive species native to the old world and is currently threatening commercial vegetable production in the southwestern USA. A series of no-choice experiments were conducted to investigate multiple plant responses in six Brassica crops to feeding by B. hilaris. Varying numbers of adults were caged onto cotyledon, 2-true leaf, and 4-true leaf-stage plants of broccoli, green cabbage, red cabbage, cauliflower, kale, and radish for a 48-h infestation period. Feeding damage on leaf surfaces, total leaf area, and relative chlorophyll content on plants of each crop were measured before and after the 48-h infestation period. In addition, dry weights and total leaf area for the 4-leaf-stage plants were measured at 21 days post-infestation to estimate the residual impacts on older plants. In all crops tested, feeding damage increased with greater numbers of B. hilaris adults caged on cotyledon and 2-leaf-stage plants. Significantly more feeding damage occurred on the upper (younger) two leaves than on the lower (older) two leaves of the 4-true leaf plants for all host plants suggesting that B. hilaris feeds preferentially on newer leaf tissue. Significant reductions in leaf area, relative chlorophyll content, and dry weight in all crops indicated negative impacts on plant growth by B. hilaris. Moreover, cotyledon and 2-leaf plants were more severely impacted by B. hilaris-induced injury than the 4-leaf plants, and kale appeared to be less sensitive to B. hilaris feeding than the other five Brassicaceous hosts.     

Development of caged non-hydrolyzable phosphoamino acids and application to photo-control of binding affinity of phosphopeptide mimetic to phosphopeptide-recognizing protein

The design and synthesis of caged non-hydrolyzable phospho-serine, -threonine, and -tyrosine derivatives that generate parent non-hydrolyzable phosphoamino acids, containing a difluoromethylene unit instead of the oxygen of a phosphoester, after UV-irradiation are described. The caged non-hydrolyzable amino acids were incorporated into peptides by standard Fmoc solid-phase peptide synthesis, and the obtained peptides were successfully converted to the parent non-hydrolyzable phosphopeptides by UV-irradiation. Application of the caged non-hydrolyzable phosphoserine-containing peptide to photo-control the binding affinity of the peptide to 14-3-3β protein is also reported.     

Kinetics and mechanism of DNA repair. Preparation, purification and some properties of caged dideoxynucleoside triphosphates.

Caged dideoxyribosylthymine triphosphate, dideoxyadenosine triphosphate and arabinosylcytosine triphosphate were prepared in high yield by reaction with 1-(2-nitrophenyl)diazoethane at pH 4 and room temperature for 24 h. Synthesis of caged alpha-32P-labelled dideoxyadenosine triphosphate (approx. 5000 Ci/mmol) in 85% yield was achieved by a modification of the method used for the synthesis of the unlabelled compounds. ATP was shown to be an excellent buffer in the synthesis of alpha-32P-labelled material, and in caged form to be an effective carrier in h.p.l.c. purification. Preparative h.p.l.c. was used to achieve purification of unlabelled caged compounds to greater than 98% purity and 32P-labelled material to 97% purity. Photolysis of unlabelled and 32P-labelled caged compounds by using XeF-excimer laser irradiation at 351 nm was characterized by using difference spectrophotometry and h.p.l.c. analysis. The stability of caged dideoxyadenosine [a-32P]triphosphate in the presence of cultured mammalian cells was evaluated; the adenosine derivative is essentially stable for 1 h.