Plant disease suppression and growth promotion by a fluorescent Pseudomonas strain

An antibiotic- and siderophore-producing Pseudomonas strain isolated from virgin soils (with forest trees) displayed in vitro antibiosis against many plant pathogenic fungi. The presence of iron had no effect on this in vitro antibiosis. Seed bacterization improved germination, shoot height, root length, fresh and dry mass, enhanced yield and chlorophyll content of leaves in the five test crop plants under field conditions. Seed bacterization also reduced the number of infected brinjal plants grown in soil infested with Rhizoctonia solani. The strain produced a yellowish green siderophore in the standard succinate medium and both siderophore and a yellow viscous antibiotic compound in King's B medium. The results confirmed that the plant growth promotion was due to siderophore production whereas the disease suppression was due to the antibiotic substance.     

Testicular disorders induced by plant growth regulators: cellular protection with proanthocyanidins grape seeds extract

The present study aims to investigate the adverse effects of plant growth regulators : gibberellic acid (GA₃) and indoleacetic acid (IAA) on testicular functions in rats, and extends to investigate the possible protective role of grape seed extract, proanthocyanidin (PAC). Male rats were divided into six groups; control group, PAC, GA₃, IAA, GA₃ + PAC and IAA + PAC groups. The data showed that GA₃ and IAA caused significant increase in total lipids, total cholesterol, triglycerides, phospholipids and low-density-lipoprotein cholesterol in the serum, concomitant with a significant decrease in high-density-lipoprotein cholesterol, total protein, and testosterone levels. In addition, there was significant decrease in the activity of alkaline phosphatase, acid phosphatase, and gamma-glutamyl transferase. A significant decrease was detected also in epididymyal fructose along with a significant reduction in sperm count. Testicular lipid peroxidation product and hydrogen peroxide (H₂O₂) levels were significantly increased. Meanwhile, the total antioxidant capacity, glutathione, sulphahydryl group content, as well as superoxide dismutase, catalase, and glucose-6-phosphate dehydrogenase activity were significantly decreased. Moreover, there were a number of histopathological testicular changes including Leydig’s cell degeneration, reduction in seminiferous tubule and necrotic symptoms and sperm degeneration in both GA₃- and IAA-treated rats. However, an obvious recovery of all the above biochemical and histological testicular disorders was detected when PAC seed extract was supplemented to rats administered with GA₃ or IAA indicating its protective effect. Therefore it was concluded that supplementation with PAC had ameliorative effects on those adverse effects of the mentioned plant growth regulators through its natural antioxidant properties.     

Plant protection and growth stimulation by microorganisms: biotechnological applications of Bacilli in agriculture

The increasing demand for a steady, healthy food supply requires an efficient control of the major pests and plant diseases. Current management practices are based largely on the application of synthetic pesticides. The excessive use of agrochemicals has caused serious environmental and health problems. Therefore, there is a growing demand for new and safer methods to replace or at least supplement the existing control strategies. Biological control, that is, the use of natural antagonists to combat pests or plant diseases has emerged as a promising alternative to chemical pesticides. The Bacilli offer a number of advantages for their application in agricultural biotechnology. Several Bacillus-based products have been marketed as microbial pesticides, fungicides or fertilisers. Bacillus-based biopesticides are widely used in conventional agriculture, by contrast, implementation of Bacillus-based biofungicides and biofertilizers is still a pending issue.     

Crop protection by seed coating

Providence of sufficient and healthy food for increasing human population clears the importance of notice to increasing crop production in company with environmental loss reduction. Growth and yield of every plant with sexual reproduction, depends on germination & emergence of sown seeds. Seed is a small alive plant that its biological function is protection and nutrition of embryo. Biological, chemical and physiological characteristics of seed, affect on plant performance & its resistance to undesirable environmental conditions, and even on its total yield. So attention to seed and try to increase its performance is so important. One of the factors that cause reduction in germination percentage and seedling establishment, is seed disease. It's possible to control these diseases by treating the seed before planting it. Coating the seed with pesticides, is one of the ways to gain this goal. Seed coating is a technique in which several material as fertilizers, nutritional elements, moisture attractive or repulsive agents, plant growth regulators, rhizobium inocolum, chemical & pesticide etc, add to seed by adhesive agents and cause to increase seed performance and germination. Seed coating, leads to increase benefits in seed industry, because seeds can use all of their genetic vigor. This technique is used for seeds of many garden plants, valuable crops (such as corn, sunflower, canola, alfalfa,...) and some of the grasses. In this technique that was first used in coating cereal seeds in 1930, a thin and permeable layer of pesticide is stuck on seed surface and prevent damage of seedborn pathogens. This layer is melted or splited after absorption of moisture and suitable temperature by seed, and let the radical to exit the seed. In this approach materials are used accurately with seed, evaporation & leakage of pesticide and also adverse effects of some pesticides on seeds are diminished, and these factors cause to increase the accuracy and performance of pesticide, decrease their consumption, environmental pollution and costs. This technique in new and there is a few information about it. So after searching and studying about this technique this paper is written to introduce it and its applications in crop protection.     

Plant and fungal identity determines pathogen protection of plant roots by arbuscular mycorrhizas

1.  A major benefit of the mycorrhizal symbiosis is that it can protect plants from below-ground enemies, such as pathogens. Previous studies have indicated that plant identity (particularly plants that differ in root system architecture) or fungal identity (fungi from different families within the Glomeromycota) can determine the degree of protection from infection by pathogens. Here, we test the combined effects of plant and fungal identity to assess if there is a strong interaction between these two factors.
2.  We paired one of two plants ( Setaria glauca , a plant with a finely branched root system and Allium cepa , which has a simple root system) with one of six different fungal species from two families within the Glomeromycota. We assessed the degree to which plant identity, fungal identity and their interaction determined infection by Fusarium oxysporum , a common plant pathogen.
3.  Our results show that the interaction between plant and fungal identity can be an important determinant of root infection by the pathogen. Infection by Fusarium was less severe in Allium (simple root system) or when Setaria (complex root system) was associated with a fungus from the family Glomeraceae. We also detected significant plant growth responses to the treatments; the fine-rooted Setaria benefited more from associating with a member of the family Glomeraceae, while Allium benefited more from associating with a member of the family Gigasporaceae.
4.   Synthesis . This study supports previous claims that plants with complex root systems are more susceptible to infection by pathogens, and that the arbuscular mycorrhizal symbiosis can reduce infection in such plants – provided that the plant is colonized by a mycorrhizal fungus that can offer protection, such as the isolates of Glomus used here.     

Effects of actinobacteria on plant disease suppression and growth promotion

Biological control and plant growth promotion by plant beneficial microbes has been viewed as an alternative to the use of chemical pesticides and fertilizers. Bacteria and fungi that are naturally associated with plants and have a beneficial effect on plant growth by the alleviation of biotic and abiotic stresses were isolated and developed into biocontrol (BCA) and plant growth-promoting agents (PGPA). Actinobacteria are a group of important plant-associated spore-forming bacteria, which have been studied for their biocontrol, plant growth promotion, and interaction with plants. This review summarizes the effects of actinobacteria as BCA, PGPA, and its beneficial associations with plants.     

Status and future of disease protection and grape berry quality alteration by micro‐organisms in viticulture

Grapevine is one of the most widely grown fruit crops in the world. At present, however, there is much concern regarding chemical pollution in viticulture due to the application of chemical fungicides and fertilizers. One viticultural practice to resolve this issue is the application of micro‐organisms to grapevine as a substitute for chemicals. Some micro‐organisms act as an enhancer of grape berry quality as well as a suppresser of disease in grapevine through their antagonistic ability and/or systemic resistance inducing ability. Herein, we review current and prospective applications of micro‐organisms in viticulture.

Significance and Impact of the Study

In this review, we evaluate the applicability of micro‐organisms in viticulture. Micro‐organisms can improve grape berry quality through grapevine disease protection and grape berry quality alteration. Because the use of micro‐organisms to protect grapevine from plant diseases is safer than the use of chemical fungicides, the use of biofungicides in viticulture is expected to be enhanced by the increasing consumer concern towards chemical fungicides. Micro‐organisms also modify plant secondary metabolites for use as flavours, pharmaceuticals and food additives. Studies of micro‐organisms that promote polyphenol, anthocyanin and aroma compound biosynthesis are in progress with an eye to improving grape berry quality.     

Inhibition of flowering and growth in Pharbitis nil by the growth retardant Ancymidol

Flower formation and growth of the short day plant Pharbitisnil, strain "Violet", were inhibited when the growth retardantAncymidol was applied prior to an inductive dark period viacotyledons or roots. Inhibition of flower formation by Ancymidolcould be completely reversed by an application of gibberellinA₃ (GA₃) to the plumule before the inductive dark period. Adose of 0.01 µg GA₃/plant was almost sufficient to restoreflowering, but about a hundred times more GA₃ was required torestore the internode length to that of control. Ancimidol greatlyreduced the endogenous gibberellin content. (Received July 18, 1980; )     

Inhibition of flowering and growth in Pharbitis nil by the growth retardant Ancymidol

Flower formation and growth of the short day plant Pharbitisnil, strain "Violet", were inhibited when the growth retardantAncymidol was applied prior to an inductive dark period viacotyledons or roots. Inhibition of flower formation by Ancymidolcould be completely reversed by an application of gibberellinA₃ (GA₃) to the plumule before the inductive dark period. Adose of 0.01 µg GA₃/plant was almost sufficient to restoreflowering, but about a hundred times more GA₃ was required torestore the internode length to that of control. Ancimidol greatlyreduced the endogenous gibberellin content. (Received July 18, 1980; )     

Endophytic bacteria enhancing growth and disease resistance of potato (Solanum tuberosum L.)

Priming plants by non-pathogenic bacteria allows the host to save energy and to reduce time needed for development of defense reaction during a pathogen attack. However, information on the role of endophytes in plant defense is limited. Here, the ability of endophytic bacteria to promote growth and resistance of potato plants towards infection by the necrotroph Pectobacterium atrosepticum was studied. A Pseudomonas sp. strain was selected due to antagonism towards bacterial pathogens and a Methylobacterium sp. strain because of efficient plant colonization. The aim of this study was to find if there is any correlation between plant growth promotion and induction of resistance by endophytes of potato, as well as to study the putative mechanisms of endophytes interacting with the plant during resistance induction. Both tested strains promoted growth of potato shoots but only the Pseudomonas sp. increased potato resistance towards the soft rot disease. Induction of disease resistance by the Methylobacterium sp. was inversely proportional to the size of bacterial population used for inoculation. The plant antioxidant system was moderately activated during the induction of resistance by the biocontrol strains. qPCR data on expression of marker genes of induced systemic resistance and acquired systemic resistance in endophyte-infected Arabidopsis plants showed activation of both salicylic acid and jasmonate/ethylene-dependent pathways after challenge inoculation with the pathogen. We suggest that some endophytes have the potential to activate both basal and inducible plant defense systems, whereas the growth promotion by biocontrol strains may not correlate with induction of disease resistance.     

The effects of genetic and environmental factors on disease expression (stroma formation) and plant growth in Brachypodium sylvaticum infected by Epichloë sylvatica

Fungi in the genus Epichloë (Clavicipitaceae, Ascomycota) are endophytic and often mutualistic symbionts of many grasses in temperate areas. Species with a sexual cycle suppress host flowering and seed formation, whereas asexual fungi remain asymptomatic and transmit vertically by seed. Thus, the mode of reproduction may determine whether the symbiosis is mutualistic or parasitic. The level of sexual reproduction (disease expression) varies among different endophytes and on different grass hosts, but factors responsible for this variation, and evolutionary mechanisms leading to one or the other life strategy are not understood. As experimental system, we chose Brachypodium sylvaticum in which the endophyte E. sylvatica can express both reproduction modes. A field experiment was done in plots of a free air carbon dioxide enrichment (FACE) facility. We investigated the effects of three environmental factors (elevated CO2 concentration, shading and fertilisation) and one genetic factor (plant and fungal genotype combination), on plant growth and disease expression. Variation in plant growth was mainly dependant on the genotype and was increased by fertilisation. Elevated CO2 and shading slightly stimulated plant growth, but only in fertilised plants. Disease expression was overwhelmingly dependent on the genotype, hence genetic factors. Fertilisation slightly stimulated disease expression in some genotypes, while the effect of elevated CO2 was negligible, and by interaction with fertilisation inconsistent in the two years. Horizontal transmissions during the experiment, presumably mediated by ascospores, confounded the original infection status of the plants. Contagious infections occurred more frequently in the shade, and in endophyte‐free host plants. The latter suggests that pre‐existing infections render host plants less susceptible to superinfection by choke forming strains. Although our results clearly indicate that disease expression of E. sylvatica has a genetic basis, it is still unclear whether selection on the plants or the fungi is driving the evolution of this association.     

Colonisation of barley roots by endophytic Fusarium equiseti and Pochonia chlamydosporia: Effects on plant growth and disease

Colonisation of plant roots by endophytic fungi may confer benefits to the host such as protection against abiotic or biotic stresses or plant growth promotion. The exploitation of these properties is of great relevance at an applied level, either to increase yields of agricultural crops or in reforestation activities. Fusarium equiseti is a naturally occurring endophyte in vegetation under stress in Mediterranean ecosystems. Pochonia chlamydosporia is a nematode egg-parasitic fungus with a worldwide distribution. Both fungi have the capacity to colonise roots of non-host plants endophytically and to protect them against phytopathogenic fungi under laboratory conditions. The aim of this study was to evaluate the root population dynamics of these fungi under non-axenic practical conditions. Both fungal species were inoculated into barley roots. Their presence in roots and effects on plant growth and incidence of disease caused by the pathogen Gaeumannomyces graminis var. tritici were monitored periodically. Both fungi colonised barley roots endophytically over the duration of the experiment and competed with other existing fungal root colonisers. Furthermore, colonisation of roots by P. chlamydosporia promoted plant growth. Although a clear suppressive effect on disease could not be detected, F. equiseti isolates reduced the mean root lesion length caused by the pathogen. Results of this work suggest that both F. equiseti and P. chlamydosporia are long-term root endophytes that confer beneficial effects to the host plant.     

Water and temperature relations of softrot bacteria: growth and disease development

The effect of water availability and the temperature of the growth substrate on growth and disease development of softrot bacteria were studied using artificial media and plant material. Water availability was measured as the osmotic potential of a solution (ψ_osm) and was assessed for solutions of PEG4000 and KNO₃ as artificial osmotica and for plant tissue of chicory heads. Growth of softrot bacteria was found at water potentials from ψ= -0.12 MPa to ψ= -8.0 MPa but the lag phase of the growth curve increased with decreasing water potential. The relative growth rates of the three softrot pathogens showed a sigmoidal relationship with water potential, the relative growth rates decreasing rapidly at water potentials lower than ψ= -1.5 MPa. The water potential of harvested chicory heads decreased with storage time of the harvested crop but was still within the growth limits for softrot bacteria. In relation to temperature, the relative growth rate of Erwinia carotovora subsp. carotovora (Ecc) was highest at 10°C, of Erwinia carotovora subsp. atroseptica (Eca) at 15°C and of Pseudomonas marginalis (Pm) at 5°C. Chicory heads of two cultivars, Rumba and Salsa, inoculated with Ecc, had a significantly higher disease severity at 30°C (0.72 for Rumba and 0.47 for Salsa) than at lower or higher temperatures. In conclusion, temperature and water availability during forcing of chicory were not factors limiting populations of softrot bacteria. Possibilities for crop protection thus only avail during chicory root storage. During storage a high death rate combined with a low growth rate of the softrot bacteria may result in a decrease of bacterial populations below the minimum densities needed for infection during the forcing of chicory heads.     

The root growth of wheat plants,the water conservation and fertility status of sandy soils influenced by plant growth promoting rhizobacteria

Plant growth promoting rhizobacteria were isolated and characterized from sandy soils in Pakistan. The role of the rhizobacteria, in association with plant growth regulators, was studied on the roots of wheat grown under water stressed conditions. The plant growth promoting rhizobacteria were characterized on the basis of colony morphology, biochemical traits and identified on the basis of 16S-rRNA gene sequencing which identified the selected isolates Planomicrobium chinense, Bacillus cereus and Pseudomonas fluorescens. Antibacterial and antifungal activities were determined. The fresh cultures (24 h old) of isolates were used to soak the seeds for 2–3 h prior to sowing. The growth regulators salicylic acid and putrescine were applied to the plant as foliar spray at three leaf stage. The plant growth promoting rhizobacteria produced exopolysaccharides that formed soil aggregation around roots of the plants and significantly enhanced water holding capacity of sandy soil. The relative water content (80%) of leaves and root fresh (80%) and dry weight (68%) were higher in plant growth promoting rhizobacteria inoculated plants. The nutrient content of rhizosphere soil of treated plants was also enhanced (Ca 35%, K 34%, Mg 52% and Na 42%) over stressed controls. Integrative use of effective plant growth promoting rhizobacteria in combination with salicylic acid appears to be an effective eco-friendly approach to increase drought tolerance in wheat plants to combat desertification.     

Durable and broad-spectrum disease protection measure against airborne phytopathogenic fungi by using the detachment action of gelatinolytic bacteria

We had previously obtained collagenolytic/gelatinolytic bacteria, which degrade the fungal extracellular matrix, to establish a novel biological control measure that inhibits germling adhesion of airborne phytopathogenic fungi on the host plant surface. By using barley-Magnaporthe oryzae pathosystem, Chryseobacterium sp. was most effective biocontrol agents as tested. The selected bacteria were evaluated for durable disease protection against M. oryzae on barley leaves by using chloramphenicol-resistant mutants. Chryseobacterium sp. from the soil was less likely to settle on leaf surfaces. Therefore, we tried to manipulate Chryseobacterium sp. to inhabit the leaf’s surface. The gelatin supplementation dramatically improved the settlement of gelatinolytic bacteria Chryseobacterium sp. from the soil, and the disease protection effect lasted for more than 2 weeks on barley. Moreover, exploitation of Chryseobacterium sp. for disease protection was extended against other airborne pathogens, Alternaria alternata Japanese pear pathotype on Japanese pear and Colletotrichum orbiculare on cucumber.     

Piriformospora indica enhances plant growth by transferring phosphate

Piriformospora indica is an endophytic fungus that colonized monocot as well as dicot. P. indica has been termed as plant probiotic because of its plant growth promoting activity and its role in enhancement of the tolerance of the host plants against abiotic and biotic stresses. In our recent study, we have characterized a high affinity phosphate transporter (PiPT) and by using RNAi approach, we have demonstrated the involvement of PiPT in P transfer to the host plant. When knockdown strains of PiPT-P. indica was colonized with the host plant, it resulted in the impaired growth of the host plants. Here we have analyzed and discussed whether the growth promoting activity of P. indica is its intrinsic property or it is dependent on P availability. Our data explain the correlation between the availability of P and growth-promoting activity of P. indica.Key words: Piriformospora indica, phosphate transport, plant growth promotionPhosphorous (P) is one of the most essential mineral nutrients for plant growth and development. In the soil P is present mainly in the form of sparingly soluble complexes that are not directly accessible to plants. Thus, it is the nutrient that limits crop production throughout the world.¹ Plants have therefore evolved a range of strategies to increase the availability of soil P, which include both morphological and biochemical changes at the soil-root interface. For example, increased root growth and branching, proliferation of root hairs, and release of root exudates can increase plant access to inorganic phosphate (Pi) from otherwise poorly available sources.^2,3 Plant root possess two distinct modes of phosphate uptake, direct uptake by its own transporters and indirect uptake through mycorrhizal associations. In plants several high affinity P transporters specifically associated with the uptake of Pi from soil solution. Expression of these transporters is induced in response to P deficiency and enables Pi to be effectively taken up against the large concentration gradient that occurs between the soil solution and internal plant tissues.⁴ However, in arbuscular mycorrhizal associations (indirect uptake), plants acquire Pi from the extensive network of fine extra radical hyphae of fungus, that extend beyond root depletion zones to mine new regions of the soil.⁵ In the case of arbuscular mycorrhizal fungi (AMF), including Glomus versiforme and G. intraradices, the regulation of phosphate transporters that are expressed, typically upregulated under P deficiency but their role in P transfer to the host plant have not been characterized.^5,6P. indica was reported to be involved in high salt tolerance, disease resistance and strong growth-promoting activities leading to enhancement of host plant yield.^7–9 Recently, we have shown the role of PiPT in the P transport to the host plant.¹⁰ Here we discuss the performance of P. indica (grown under P-rich and -deprived conditions and colonized with the host plant) and its involvement in the P transportation to, and the growth of the host plant.     

Plant growth promoting microorganisms as alternative to chemical protection from pathogens (review)

The review analyses data on physiological and biochemical influence of rhizospheric and endophytic microorganisms promoting plant growth (PGPR-plant growth promoting rhizobacteria) on induced resistance of plants and the possibility of its use in plant cultivation to protect crops from pathogens and phytophages. Resistance of plants provided by PGPR due to their endosymbiotic interrelationships is directly achieved because they produce peptide antibiotics and hydrolases ofchitin and glucan and also because plants form their own system of induced resistance, followed by changes in the balance of defensive proteins, phytohormones, and pro-/antioxidant status.     

Pea-derived vaccines demonstrate high immunogenicity and protection in rabbits against rabbit haemorrhagic disease virus

Vaccines against rabbit haemorrhagic disease virus (RHDV) are commercially produced in experimentally infected rabbits. A genetically engineered and manufactured version of the major structural protein of RHDV (VP60) is considered to be an alternative approach for vaccine production. Plants have the potential to become an excellent recombinant production system, but the low expression level and insufficient immunogenic potency of plant‐derived VP60 still hamper its practical use. In this study, we analysed the expression of a novel multimeric VP60‐based antigen in four different plant species, including Nicotiana tabacum L., Solanum tuberosum L., Brassica napus L. and Pisum sativum L. Significant differences were detected in the expression patterns of the novel fusion antigen cholera toxin B subunit (CTB)::VP60 (ctbvp60_SEKDEL) at the mRNA and protein levels. Pentameric CTB::VP60 molecules were only detected in N. tabacum and P. sativum, and displayed equal levels of CTB, at approximately 0.01% of total soluble protein (TSP), and traces of detectable VP60. However, strong enhancement of the CTB protein content via self‐fertilization was only observed in P. sativum, where it reached up to 0.7% of TSP. In rabbits, a strong decrease in the protective vaccine dose required from 48–400 µg potato‐derived VP60 [ Castanon, S., Marin, M.S., Martin‐Alonso, J.M., Boga, J.A., Casais, R., Humara, J.M., Ordas, R.J. and Parra, F. (1999) Immunization with potato plants expressing VP60 protein protects against rabbit hemorrhagic disease virus. J. Virol. 73 , 4452–4455; Castanon, S., Martin‐Alonso, J.M., Marin, M.S., Boga, J.A., Alonso, P., Parra, F. and Ordas, R.J. (2002) The effect of the promoter on expression of VP60 gene from rabbit hemorrhagic disease virus in potato plants. Plant Sci. 162 , 87–95] to 0.56–0.28 µg antigenic VP60 (measured with VP60 enzyme‐linked immunosorbent assay) of crude CTB::VP60 pea extracts was demonstrated. Rabbits immunized with pea‐derived CTB::VP60 showed anti‐VP60‐specific antibodies, similar to RikaVacc^®‐immunized rabbits, and survived RHDV challenge.     

Pinitol occurrence in soybean plants as affected by temperature and plant growth regulators

Unsuitable temperatures are frequently encountered by soybean(Glycine max L. Merr.) plants grown in the field. Certain polyolshave been reported to protect plants from high temperature orfrost damage. Controlled environment studies were conductedto investigate the effect of stressful temperature regimes onthe content of pinitol (3-O-methyl-D-chiro-inositol) in soybeanplants. Hydroponically-grown soybean plants were subjected tohigh (35/30 C) or low (15/10 C) day/night temperature stresses,and pinitol content in different plant parts was determinedusing high performance liquid chromatography (HPLC). A syntheticplant growth regulator, PGR-IV, was foliarly applied to theplants to evaluate its effect on pinitol content in differentplant components. Uniformly-labelled ¹⁴C-glucose was fed intothe leaves via the transpiration stream, and the effects ofhigh temperature and EXP-S1089, another synthetic plant growthregulator, on the incorporation of ¹⁴C-glucose into pinitolwas evaluated using HPLC separation and scintillation spectrometry.High-temperature stress significantly increased plant pinitolcontent and the incorporation of ¹⁴C-glucose into pinitol, butdecreased the content of sucrose, glucose and fructose. Underlow-temperature stress, there was hardly any change in pinitolcontent, but a drastic increase in soluble sugars. PGR-IV enhancedpinitol translocation from leaves to stems and roots, whileEXP-S1089 increased pinitol/sucrose ratio. Accumulation of pinitolmay be an adjustment mechanism of the plant to reduce high-temperaturedamage, but not low-temperature injuries. Key words: Pinitol, soybean, temperature, plant growth regulator