<Emphasis Type="Italic">In vitro</Emphasis> morphogenic response of leaf sheath of <Emphasis Type="Italic">Phyllostachys bambusoides</Emphasis>

Nodal segments from secondary branches of saplings of Phyllostachys bambusoides were inoculated in MS medium to assess the in vitro morphogenic response of leaf sheath through the induction to callogenesis by Picloram(4-amino-3,5,6-trichloropicolinic acid) at different concentrations of carbohydrate under the same conditions with presence or absence of luminosity.In our experiment,secondary explants were kept in MS medium containing 8.0 mg·L-1 of Picloram for the callus formation.Calluses were transferred in MS medium supplemented with sucrose,fructose and glucose(control,2%,4% and 6%).Results show that Picloram induced the callogenesis in leaf sheath.The secondary embryogenesis was formed in yellow-globular callus.The sucrose as carbohydrate source in the absence of light was more efficient to induce rhizogenesis.Glucose was more efficiency in the presence of light.Callogenic induction and further embryogenesis evidenced the competence and determination of leaf sheath cells.     

In vitro clonal propagation of <Emphasis Type="Italic">Balanites aegyptiaca</Emphasis> (L.) Del

In vitro propagation technique of Balanites aegyptiaca, a multipurpose woody tree was studied. Nodal segments including axillary bud from mature tree were used as an explant and their morphogenetic potential was tested on MS media with various concentrations (2.5–15.0 μM) of 6-benzyladenine (BA), Kinetin, and Thidiazuron alone or in combination with different concentrations (0.5–2.5 μM) of α-naphthalene acetic acid (NAA). Nodal segments showed axillary bud proliferation in almost all media tried. MS medium containing 12.5 μM BA alone was effective for inducing multiple shoots (5.0 ± 0.22) with an average shoot length (3.7 ± 0.26 cm) in 67% of cultures. A better shoot differentiation and elongation was achieved in a combined treatment of BA (12.5 μM) and NAA (1.0 μM). Half strength MS medium supplemented with Indole-3-butyric acid (IBA) gave the best result for rooting. The maximum frequency of root formation (68%), number of roots (5.3 ± 0.32) and root length (4.1 ± 0.38 cm) was obtained on half strength MS medium containing 1.0 μM IBA. The regenerated plantlets were potted and acclimatized successfully in a growth chamber and then moved to the greenhouse.     

Spatial pattern of beech (<Emphasis Type="Italic">Fagus sylvatica</Emphasis> L.) and oak (<Emphasis Type="Italic">Quercus pubescens</Emphasis> Mill.) seedlings in natural pine (<Emphasis Type="Italic">Pinus sylvestris</Emphasis> L.) woodlands

Spatial pattern of recruitment is an important factor influencing population dynamics of plant communities. The spatial pattern is determined by seed dispersal and by the spatial variability of germination and initial survival. In the process of forest expansion following farmland abandonment, mid- and late-successional species are often dispersed in pioneer forests by birds. This could result in an aggregated spatial pattern of seeds that could influence the dynamics of these species in mixed pioneer forests. In the sub-Mediterranean area, mid- and late-successional species such as Quercus pubescens (downy oak) and Fagus sylvatica (European beech) are expected to replace pioneer Pinus species. Using a point sampling method we demonstrated that beech and oak seedlings (height <2 m) have a clumped distribution in the understorey of pine. This could result from an aggregated dispersal by jays (dispersal effect) or from preferential recruitment in particular habitats (habitat effect). To test these hypotheses we proposed a statistical analysis of spatial patterns of regeneration of beech and oak. Ground cover variables (i.e. cover by rock outcrops, herbs, box shrubs, mosses or pine) did not differ significantly around seedlings as compared with random sample plots. Likewise, clumped seedlings had growth similar to isolated seedlings, thus refuting the hypothesis of preferential location in the most favourable microsites. Aggregated dispersal seems to be involved in the process of regeneration. Since beech and oak seedlings have contrasting ecological demands, we discuss the implication of this pattern for the replacement dynamics of pine by these species.     

Pathogenic effects of <Emphasis Type="Italic">Xenorhabdus nematophilus</Emphasis> and <Emphasis Type="Italic">Photorhabdus luminescens</Emphasis> (Enterobacteriaceae) against pupae of the Diamondback Moth, <Emphasis Type="Italic">Plutella xylostella</Emphasis> (L.)

The relationship between Xenorhabdus nematophilus and Photorhabdus luminescens, the bacterial symbionts of the entomopathogenic nematodes Steinernema carpocapsae and Heterorhabditis bacteriophora, against the diamondback moth, P. xylostella (L.), pupae showed the pathogenic capability of P. luminescens to be over that of X. nematophilus. They gave 60 and 40?% mortality with LC₅₀ values of 5?×?10⁴ and 5.5?×?10⁵ cells/ml, respectively.The number of bacterial cells influences the rate of killing the pupae of P. xylostella and a direct correlation between mortality of the pupae, percentage of deformed adults and the spraying dose with significant differences were observed. An inverse correlation found between the no. of eggs laid/female, percentage of hatching, adult survival and the spraying dose. These bacterial effects have been attributed to different toxic bacterial enzymes which damage the haemocytes and inhibit activation of the humoral immune system of the insect.     

Influence of cytokinins,basal media and pH on adventitious shoot regeneration from excised root cultures of <Emphasis Type="Italic">Albizia lebbeck</Emphasis>

A highly reproducible and efficient in vitro shoot regeneration system was developed in a potential medicinal plant, Albizia lebbeck using root explants. Root explants from 15 day-old-aseptic seedlings were cultured on Murashige and Skoog (MS) medium supplemented with different concentrations (0.5, 2.5, 5.0, 7.5 and 10.0 μM) of 6-Benzyladenine (BA), Kinetin (Kn), 2-Isopentenyl adenine (2-iP) singly as well as in combination with α-Naphthalene acetic acid (NAA) (0.1, 0.5, 1.0, 1.5 and 2.0 μM). The highest rate of shoot multiplication (16.0 ± 1.87 for the average shoot number and 5.16 ± 0.38 cm for shoot length) was achieved on MS medium supplemented with 7.5 μM BA and 0.5 μM NAA. The effects of medium type, medium strength, pH and subculture on shoot induction and proliferation were also tested. An average of 21.6±2.87 shoots per explants could be obtained following this protocol. Rooting was achieved on microshoots using half strength MS medium with 2.0 μM Indole-3-butyric acid (IBA) after four weeks of culture. The in vitro raised healthy plantlets were successfully established in earthen pots containing garden soil and grown in greenhouse with >80% survival rate.     

Osmoregulators in <Emphasis Type="Italic">Hymenaea courbaril</Emphasis> and <Emphasis Type="Italic">Hymenaea stigonocarpa</Emphasis> under water stress and rehydration

The objective of this work was to evaluate the effect of different water deficiency and rehydration levels on the concentrations of osmoregulators in two plant species (Hymenaea courbaril and H. Stigonocarpa) in the Amazon. We adopted a 2 × 5 × 5 factorial system, referring to 2 species (H. courbaril and H. stigonocarpa) and 5 stages of hydration and rehydration. The five hydration and rehydration stages were established in: (1) Control treatment E0; (2) Plants with 13 days of stress after incubation—E13; (3) Plants with 26 days of stress E26; (4) The plants that were established after 26 days after incubation and rehydrated for two days (RD2); (5) rehydrated for two days (RD4). The plants that were established after 26 days after incubation and rehydrated for four days. The experiment totaled fifty young plants with five replicates. Biochemical measurements were performed at the beginning of the experiment (E0) at 13 (E13) and 26 (E26) days after the water stress, in which the plants were rehydrated, repeating the analyses after two (RD2) and four (RD4) days. Both species increased the sucrose concentration by 18%, with a decrease of 52% in starch content. The RD4 time presented the highest mean starch concentration (0.19 mmol g^?1 of the residue for H. courbaril and 0.27 mmol g^?1 of residue for H. stigonocarpa). Increased proline concentrations were recorded for controls until RD2 for both species. For glycine betaine, the highest increases in treatments E26 and RD2 were observed for the H. courbaril species. Our rehydration period was not sufficient for total recovery of pre-stress concentrations of all studied solutes.     

Direct and admixture toxicity of diatomaceous earth and monoterpenoids against the storage pests <Emphasis Type="Italic">Callosobruchus</Emphasis><Emphasis Type="Italic">maculatus</Emphasis> (F.) and <Emphasis Type="Italic">Sitophilus oryzae</Emphasis> (L.)

We investigated the effects of two commercial diatomaceous earth based insecticides (DE), Protect-It^® and SilicoSec^®, the nano-structured silica product AL06, developed by the section for Urban Plant Ecophysiology at Humboldt University Berlin, and the monoterpenoids, eugenol, and cinnamaldehyde on two stored product pests, Callosobruchus maculatus and Sitophilus oryzae. Protect-It^® was more effective than SilicoSec^® against C. maculatus while the reverse was true for S. oryzae. Generally C. maculatus was more sensitive towards DE and silica treatment than S. oryzae. Mortality rate of both pest species increased when DE’s were applied to food commodities previously treated with a monoterpenoid. In admixture experiments, the toxicity of SilicoSec^® + cinnamaldehyde (LD₅₀ = 42.73 ppm), SilicoSec^® + eugenol (LD₅₀ = 24.30 ppm), and Protect-It^® + eugenol (LD₅₀ = 2.60 ppm) was increased over DE alone against S. oryzae. Both substances showed a synergistic effect considering their co-toxicity coefficient relative to the LD₅₀-value. In contrast, we could not find any synergistic effects in experiments with C. maculatus. Here only Protect-It^® + cinnamaldehyde (LD₅₀ = 20.84 ppm) showed an additive effect while all other combinations of monoterpenoid and DE indicated antagonistic effects. In addition to contact insecticidal effects both monoterpenoids showed a strong fumigant action. The presented results indicate that the natural product DE has great potential to replace synthetic pesticides commonly used in stored product pest management. Efficacy of DE can be improved by adding certain monoterpenoids against certain insect pests.     

Successful <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of <Emphasis Type="Italic">Populus tomentosa</Emphasis> with apple <Emphasis Type="Italic">SPDS</Emphasis> gene

The problem of salinized soils has become one of the most serious constraints to agricultural and forest productivity. With the purpose of enhancing salt stress tolerance of Populus tomentosa, we transformed this tree species with spermidine synthase （SPDS） genes derived from an apple by an Agrobacterium-mediated method. Four transgenic clones were confu＇med by PCR and Southern blot analysis. As well, the expression of introduced SPDS genes was analyzed by real-time quantitative PCR.     

Species proportions by area in mixtures of Scots pine (<Emphasis Type="Italic">Pinus sylvestris</Emphasis> L.) and European beech (<Emphasis Type="Italic">Fagus sylvatica</Emphasis> L.)

Scots pine (Pinus sylvestris L.) and European beech (Fagus sylvatica L.) dominate many of the European forest stands. Also, mixtures of European beech and Scots pine more or less occur over all European countries, but have been scarcely investigated. The area occupied by each species is of high relevance, especially for growth evaluation and comparison of different species in mixed and monospecific stands. Thus, we studied different methods to describe species proportions and their definition as proportion by area. 25 triplets consisting of mixed and monospecific stands were established across Europe ranging from Lithuania to Spain in northern to southern direction and from Bulgaria to Belgium in eastern to western direction. On stand level, the conclusive method for estimating the species proportion as a fraction of the stand area relates the observed density (tree number or basal area) to its potential. This stand-level estimation makes use of the potential from comparable neighboring monospecific stands or from maximum density lines derived from other data, e.g. forest inventories or permanent observations plots. At tree level, the fraction of the stand area occupied by a species can be derived from the proportions of their crown projection area or of their leaf area. The estimates of the potentials obtained from neighboring monospecific stands, especially in older stands, were poorer than those from the maximum density line depending on the Martonne aridity index. Therefore, the stand-level method in combination with the Martonne aridity index for potential densities can be highly recommended. The species’ proportions estimated with this method are best approximated by the proportions of the species’ leaf areas. In forest practice, the most commonly applied method is an ocular estimation of the proportions by crown projection area. Even though the proportions of pine were calculated here by measuring crown projection areas in the field, we found this method to underestimate the proportion by 25% compared to the stand-level approach.     

Influence of umbrella pine (<Emphasis Type="Italic">Pinus</Emphasis><Emphasis Type="Italic">pinea</Emphasis> L.) stand type and tree characteristics on cone production

Most umbrella pine (Pinus pinea L.) stands are managed as agroforestry systems, whose main production is fruit, due to the edible and highly nutritious kernels, and are frequently associated to natural or seeded pastures and grazing. The stands have low density, in order to enhance crown growth and fruit production. Nevertheless, cone production, both with regard to number and weight, varies greatly between stands, trees and years. In this study were selected three agroforestry systems, representative of umbrella pine stands whose main production is fruit, and one stand representative of the timber production system, where fruit is the secondary production. It was evaluated the variability in cone production as a function of the tree’s diameter at breast height and crown diameter and the individual tree’s competition status. The results indicate that stands managed in agroforestry systems with lower competition and individuals with larger diameter at breast height and crown diameter tend to produce more and heavier cones per tree. The first two principal components of the principal component analysis explain 84 % of the variance in cone production, trees’ dimensions and competition index. Tree competition status has a negative impact on production per tree.     

Effect of different pre-sowing seed treatments on the germination of <Emphasis Type="Italic">Leucaena leucocephala</Emphasis> (Lam.) and <Emphasis Type="Italic">Acacia farnesiana</Emphasis> (L.)

Leucaena leucocephala and Acacia farnesiana are tree species used for several agricultural purposes in the Mediterranean region. The seeds of these species exhibit dormancy, causing delayed germination. Two experiments were conducted to investigate the effect of pre-sowing treatments (scarification, hot water, or soaking) on seed germination of L. leucocephala and A. farnesiana. In one experiment, seeds were exposed to three pre-sowing treatments: control, sandpaper scarification, or soaking in 70°C water for 4, 8, 12, 16, 20, or 24 min. In another experiment, seeds were soaked in 70°C water for 20 min, and then soaked in water at room temperature for an additional 24, 48, or 72 h or blade scarified. In general, soaking the seeds of the two species in hot water was more effective in breaking seed dormancy than scarification. Sandpaper scarification was not effective for either species. Blade scarification increased A. farnesiana seed germination to 56%, indicating that seed dormancy was mainly a consequence of hardseededness. L. leucocephala seeds collected from Jordan University of Science and Technology (JUST) site and soaked in 70°C water for 20 min and then soaked for 24, 48, or 72 h had germination rates above 97%. Our results suggest that blade scarification of A. farnesiana seeds and soaking of L. leucocephala seeds in 70°C water for 20 min are effective treatments to break seed dormancy and enhance seed germination of these vital species.     

Insecticidal activity of synthetic antioxidants,natural phytochemicals,and essential oils against an <Emphasis Type="Italic">Aspergillus</Emphasis> section <Emphasis Type="Italic">Flavi</Emphasis> vector (<Emphasis Type="Italic">Oryzaephilus surinamensis</Emphasis> L.) in microcosm

The purpose of this study is to investigate the insecticidal activity of two benzoic acids 2(3)-tert-butyl-4 hydroxyanisole (BHA) and 2,6-di(tert-butyl)-p-cresol (BHT); two phenolic acids 3-phenyl-2-propenoic acid (CA) and trans-4-hydroxy-3-methoxycinnamic acid (FA); and two essential oils of Eugenia caryophyllata (clove tree) and Thymus vulgaris (thyme) against Oryzaephilus surinamensis (L.) vector carrier of aflatoxigenic fungi on stored peanut. The weight loss of peanut, susceptibility of insects, Aspergillus isolation frequency from insects and peanut, and aflatoxin B₁ analyses from peanut were determined. BHA, BHT, and BHA/BHT mixture were highly effective against O. surinamensis, these chemical agents give 100% mortality at the doses assayed. Essential oil of thyme at 2,000 and 3,000 ppm were highly effective against O. surinamensis, these concentrations gave 100% mortality. No Aspergillus section Flavi contamination was observed in dead insects collected from peanut treated with BHA/BHT mixture. No differences were observed in the fresh weight of pods peanut among treatments with and without chemical agents. All samples of treated peanut pods showed complete inhibition of this toxin after 120 days of storage. Our results indicate that these substances could be evaluated further for the control of pest vectors of aflatoxigenic fungi of stored peanut.     

Fine root biomass and morphology of <Emphasis Type="Italic">Pinus densiflora</Emphasis> under competitive stress by <Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis>

The fine root (diameter ≤2.0 mm) biomass and morphology of Japanese red pines (Pinus densiflora) grown under different aboveground conditions (i.e., high and low competitive environments) were examined in a pine–cypress mixed forest. All P. densiflora subject trees were about 40 years old, and the aboveground condition (i.e., size) of red pines appeared to be influenced by the surrounding Japanese cypress (Chamaecyparis obtusa). Smaller P. densiflora exhibited lower fine root biomasses, shorter root lengths, and lower root tip densities, but longer specific root lengths and higher specific root tip densities relative to larger pines. These results suggest that P. densiflora may adjust the morphological traits of fine roots to the different conditions in biomass allocation to fine roots of individuals with different aboveground growth.     

Effectiveness of Neem (<Emphasis Type="Italic">Azadirachta indica</Emphasis>) insecticides against Brassica pod midge (<Emphasis Type="Italic">Dasineura brassicae</Emphasis> Winn<Emphasis Type="Italic">.</Emphasis>)

The efficiency of the botanical insecticide (BI) NeemAzal T/S (containing 1% Azadirachtin A), on the basis of azadirachtin applied in a dose of 20 g a.i. ha⁻¹ against Brassica pod midge (Dasineura brassicae), has been monitored for 4 years. The biological efficiency of BI was compared with the efficiency of some synthetic insecticides. It was ascertained that BI was very efficient in decreasing the number of damaged oilseed rape pods (ranging from 56.5 to 85.9% compared to untreated plants) and its efficiency was comparable with synthetic insecticides based on Chloronicotinyl (Thiacloprid) and Neonicotinoid (Acetamiprid). BI’s efficiency was, in some years, even significantly higher compared to pyrethroid (λ-cyhalothrin). The high biological efficiency of azadirachtin that we ascertained was significantly reflected in increasing potential crop yields. The yield increase of azadirachtin ranged between 9.3 and 19.4% compared to the control sample. Azadirachtin showed the highest yield for the whole time of experimentation, and in some years the yield increase was significantly higher compared to some synthetic agents.     

Insecticidal properties of <Emphasis Type="Italic">Thymus persicus</Emphasis> essential oil against <Emphasis Type="Italic">Tribolium castaneum</Emphasis> and <Emphasis Type="Italic">Sitophilus oryzae</Emphasis>

Red flour beetle Tribolium castaneum (Herbst) and rice weevil Sitophilus oryzae (L.) are considered to be the major insect pests in storage. Essential oils from aromatic plants are recognized as proper alternatives to fumigants. Thymus persicus (Ronniger ex Rech. f.) is one of these plants that have medicinal properties and is indigenous to Iran. The essential oil was obtained from aerial parts of the plant and analyzed by GC and GC–MS. Carvacrol (44.69%) and thymol (11.05%) were the major constituents of the oil extracted. In this experiment, fumigant toxicity of the essential oil was studied against T. castaneum, S. oryzae at 27 ± 1°C and 60 ± 5% RH in dark condition. The adult insects were exposed to the concentrations of 51.9, 111.1, 207.4 and 370.4 μl/l air to estimate median lethal time (LT₅₀) values. The fumigant toxicity was increased in response to increased essential oil concentrations. The LT₅₀ values at the lowest and the highest concentrations tested were ranged from 28.09 to 13.47 h for T. castaneum, and 3.86 to 2.30 h for S. oryzae. It was found that S. oryzae adults were much more susceptible to the oil than T. castaneum. After 24 h of exposure, the LC₅₀ values (95% fiducial limit) for T. castaneum and S. oryzae were estimated to be 236.9 (186.27–292.81) and 3.34 (2.62–4.28) μl/l air, respectively. These results suggest that T. persicus essential oil merits further study as potential fumigant for the management of these stored-product insects.     

含羞草叶子提取物对常见农作物种子萌发生长的抑制作用

观察了含羞草叶子提取物对孟加拉常见农作物种子萌发生长的抑制作用.实验在27-30℃的室温和24小时的光照条件下进行.结果表明：不同浓度的含羞草叶子提取物明显影响所选的植物种子根,茎和侧根的延长和生长.生物学测定表明：高浓度（50%-100%）的提取物有较强的抑制作用.而低浓度 （10%-25%） 的提取物的抑制作用较弱.研究还显示,提取物对植物根和侧根的抑制作用比对种子萌发和茎的生长抑制作用更显著.     

<Emphasis Type="Italic">Bursaphelenchus mucronatus</Emphasis> (Nematoda: Aphelenchoididae) vectored by <Emphasis Type="Italic">Monochamus </Emphasis><Emphasis Type="Italic">urussovi</Emphasis> (Coleoptera: Cerambycidae) in Hokkaido,Japan

Bursaphelenchus mucronatus Mamiya et Enda has been recovered for the first time from adults of the cerambycid beetle, Monochamus urussovi (Fischer), in Hokkaido, Japan. The nematode was also recovered from the inner bark of Picea jezoensis (Siebold et Zuccarini) Carrière and Abies sachalinensis (Fr Schmidt) Masters infested with M. urussovi larvae. PCR–RFLP analysis indicated that B. mucronatus in Hokkaido is the European type.     

In vitro regeneration of <Emphasis Type="Italic">Populus tomentosa</Emphasis> from petioles

A reliable in vitro regeneration procedure for Populus tomentosa is a prerequisite for its trait improvement through genetic transformation. We established a systematic protocol for indirect regeneration of P. tomentosa using in vitro petioles of Chinese poplar cultivar ‘fasta-3’. A high frequency of callus induction (>97 %) was obtained from isolated petioles cultured on the modified 1/2MS basal medium supplemented with 0.5 mg/L ZT and 1.0 mg/L NAA, and the tested calli were subsequently plated on 1/2MS basal medium supplemented with 0.25 mg/L BA, 0.25 mg/L ZT, 0.25 mg/L NAA, 0.01 mg/L TDZ, and 0.5 mg/L KT for efficient regeneration of shoots after being cultured for 6 weeks. The regenerated shoots were vigorously rooted on the tested media supplemented with 1.0 mg/L IBA and 0.5 mg/L NAA. These results can facilitate genetic transformation of P. tomentosa for trait improvements in future.     

Phenotypic plasticity blurs ecotypic divergence in the response of <Emphasis Type="Italic">Quercus coccifera</Emphasis> and <Emphasis Type="Italic">Pinus halepensis</Emphasis> to water stress

The Mediterranean evergreen woody plants Quercus coccifera and Pinus halepensis grow in a range of environments where selection by drought, heat and high irradiance can drive genetic and phenotypic differentiation of populations. However, the role of these stresses in filtering out maladaptive genotypes remains unknown. We hypothesize that this filtering is an important process for woody Mediterranean species due to their low phenotypic plasticity reported in previous studies. We have studied the response of saplings of Q. coccifera and P. halepensis, originating from two contrasting populations (a rock outcrop and a garrigue formation), to water stress. Isozyme characterization of genetic diversity was done to determine whether populations were genetically distinct. Water response analysis was based on water relations, gas exchange, chlorophyll a fluorescence, pigment content, antioxidant status and morphological and structural parameters. Ecotypic differentiation was found for both Q. coccifera and P. halepensis populations, with a higher population isozyme similarity and a higher frequency of dominance of a few genotypes at the rock outcrop in both the species. P. halepensis exhibited small but significant differences between populations for plastic responses to water, with lower phenotypic plasticity in saplings from the rock outcrop. Although it was not found in Q. coccifera, this pattern suggests that ecotypic differentiation rendering stress-tolerant ecotypes involves a decreased plasticity. Phenotypic plasticity was not high but it explained over 75% of the total variability among individual plants. Thus, and although evidence for ecotypic divergence was found in both the species, saplings were plastic enough to blur ecotypic differentiation. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.