L-Tryptophan-dependent biosynthesis of indole-3-acetic acid (IAA) improves plant growth and colonization of maize by <Emphasis Type="BoldItalic">Burkholderia phytofirmans</Emphasis> PsJN

Burkholderia phytofirmans PsJN is a well-known plant growth-promoting bacterium that establishes rhizospheric and endophytic colonization in different plants. PsJN inoculation promotes growth of different horticultural crops. L-Tryptophan (L-TRP) application may further improve its effectiveness, due to substrate (L-TRP)-dependent inoculum (PsJN)-derived auxins in the rhizosphere. In the present study, the substrate (L-TRP)-dependent response of PsJN inoculation to maize growth and auxin biosynthesis was evaluated under pot conditions. In vitro auxin biosynthesis by PsJN was determined in the absence and presence of L-TRP, a physiological precursor of auxins. Surface-disinfected seeds were treated with peat-based inoculum and L-TRP solutions (10^?4 and 10^?5 M). Results revealed that L-TRP application and PsJN inoculation, when applied separately, significantly increased the growth parameters of maize compared to untreated control. However, PsJN inoculation supplemented with L-TRP (10^?5 M) gave the most promising results and significantly increased plant height, photosynthesis, chlorophyll content, root biomass and shoot biomass up to 18, 16, 45, 62 and 55 %, respectively, compared to the uninoculated control. Similarly, higher values of N, P and IAA content were observed with precursor (L-TRP)–inoculum (PsJN) interaction. The inoculant strain efficiently colonized maize seedlings and was recovered from the rhizosphere, root and shoot of plants. The results imply that substrate (L-TRP)-derived IAA biosynthesis in the rhizosphere by PsJN inoculation could be a useful approach for improving the growth, photosynthesis and nutrient content of maize plants.     

Comparative analysis of defence responses induced by the endophytic plant growth-promoting rhizobacterium Burkholderia phytofirmans strain PsJN and the non-host bacterium Pseudomonas syringae pv. pisi in grapevine cell suspensions

Plant growth-promoting rhizobacteria (PGPR) are beneficial microorganisms that colonize the rhizosphere of many plant species and confer beneficial effects, such as an increase in plant growth. PGPR are also well known as inducers of systemic resistance to pathogens in plants. However, the molecular mechanisms involved locally after direct perception of these bacteria by plant cells still remain largely unknown. Burkholderia phytofirmans strain PsJN is an endophytic PGPR that colonizes grapevine and protects the plant against the grey mould disease caused by Botrytis cinerea. This report focuses on local defence events induced by B. phytofirmans PsJN after perception by the grapevine cells. It is demonstrated that, after addition to cell suspension cultures, the bacteria were tightly attaching to plant cells in a way similar to the grapevine non-host bacteria Pseudomonas syringae pv. pisi. B. phytofirmans PsJN perception led to a transient and monophasic extracellular alkalinization but no accumulation of reactive oxygen species or cell death were detected. By contrast, challenge with P. syringae pv. pisi induced a sustained and biphasic extracellular alkalinization, a two phases oxidative burst, and a HR-like response. Perception of the PGPR also led to the production of salicylic acid (SA) and the expression of a battery of defence genes that was, however, weaker in intensity compared with defence gene expression triggered by the non-host bacteria. Some defence genes up-regulated after B. phytofirmans PsJN challenge are specifically induced by exogenous treatment with SA or jasmonic acid, suggesting that both signalling pathways are activated by the PGPR in grapevine.     

Endophytic Colonization of Vitis vinifera L. by Plant Growth-Promoting Bacterium Burkholderia sp. Strain PsJN

Patterns of colonization of Vitis vinifera L. cv. Chardonnay plantlets by a plant growth-promoting bacterium, Burkholderia sp. strain PsJN, were studied under gnotobiotic conditions. Wild-type strain PsJN and genetically engineered derivatives of this strain tagged with gfp (PsJN::gfp2x) or gusA (PsJN::gusA11) genes were used to enumerate and visualize tissue colonization. The rhizospheres of 4- to 5-week-old plantlets with five developed leaves were inoculated with bacterial suspensions. Epiphytic and endophytic colonization patterns were then monitored by dilution plating assays and microscopic observation of organ sections. Bacteria were chronologically detected first on root surfaces, then in root internal tissues, and finally in the fifth internode and the tissues of the fifth leaf. Analysis of the PsJN colonization patterns showed that this strain colonizes grapevine root surfaces, as well as cell walls and the whole surface of some rhizodermal cells. Cells were also abundant at lateral root emergence sites and root tips. Furthermore, cell wall-degrading endoglucanase and endopolygalacturonase secreted by PsJN explained how the bacterium gains entry into root internal tissues. Host defense reactions were observed in the exodermis and in several cortical cell layers. Bacteria were not observed on stem and leaf surfaces but were found in xylem vessels of the fifth internode and the fifth leaf of plantlets. Moreover, bacteria were more abundant in the fifth leaf than in the fifth internode and were found in substomatal chambers. Thus, it seems that Burkholderia sp. strain PsJN induces a local host defense reaction and systemically spreads to aerial parts through the transpiration stream.     

Endophytic colonization of Vitis vinifera L. by plant growth-promoting bacterium Burkholderia sp. strain PsJN

Patterns of colonization of Vitis vinifera L. cv. Chardonnay plantlets by a plant growth-promoting bacterium, Burkholderia sp. strain PsJN, were studied under gnotobiotic conditions. Wild-type strain PsJN and genetically engineered derivatives of this strain tagged with gfp (PsJN::gfp2x) or gusA (PsJN::gusA11) genes were used to enumerate and visualize tissue colonization. The rhizospheres of 4- to 5-week-old plantlets with five developed leaves were inoculated with bacterial suspensions. Epiphytic and endophytic colonization patterns were then monitored by dilution plating assays and microscopic observation of organ sections. Bacteria were chronologically detected first on root surfaces, then in root internal tissues, and finally in the fifth internode and the tissues of the fifth leaf. Analysis of the PsJN colonization patterns showed that this strain colonizes grapevine root surfaces, as well as cell walls and the whole surface of some rhizodermal cells. Cells were also abundant at lateral root emergence sites and root tips. Furthermore, cell wall-degrading endoglucanase and endopolygalacturonase secreted by PsJN explained how the bacterium gains entry into root internal tissues. Host defense reactions were observed in the exodermis and in several cortical cell layers. Bacteria were not observed on stem and leaf surfaces but were found in xylem vessels of the fifth internode and the fifth leaf of plantlets. Moreover, bacteria were more abundant in the fifth leaf than in the fifth internode and were found in substomatal chambers. Thus, it seems that Burkholderia sp. strain PsJN induces a local host defense reaction and systemically spreads to aerial parts through the transpiration stream.     

Endophytic colonization of Vitis vinifera L. by Burkholderia phytofirmans strain PsJN: from the rhizosphere to inflorescence tissues

The colonization pattern of Vitis vinifera L. by Burkholderia phytofirmans strain PsJN was determined using grapevine fruiting cuttings with emphasis on putative inflorescence colonization under nonsterile conditions. Two-week-old rooted plants harbouring flower bud initials, grown in nonsterile soil, were inoculated with PsJN:gfp2x. Plant colonization was subsequently monitored at various times after inoculation with plate counts and epifluorescence and/or confocal microscopy. Strain PsJN was chronologically detected on the root surfaces, in the endorhiza, inside grape inflorescence stalks, not inside preflower buds and flowers but rather as an endophyte inside young berries. Data demonstrated low endophytic populations of strain PsJN in inflorescence organs, i.e. grape stalks and immature berries with inconsistency among plants for bacterial colonization of inflorescences. Nevertheless, endophytic colonization of inflorescences by strain PsJN was substantial for some plants. Microscopic analysis revealed PsJN as a thriving endophyte in inflorescence organs after the colonization process. Strain PsJN was visualized colonizing the root surface, entering the endorhiza and spreading to grape inflorescence stalks, pedicels and then to immature berries through xylem vessels. In parallel to these observations, a natural microbial communities was also detected on and inside plants, demonstrating the colonization of grapevine by strain PsJN in the presence of other microorganisms.     

Ammonia cometabolism and product inhibition vary considerably among species of methanotrophic bacteria

The structural gene for 1-aminocyclopropane-1-carboxylate (ACC) deaminase ( acdS ) from the endophytic plant growth-promoting bacterium Burkholderia phytofirmans PsJN was isolated and used to construct a mutant strain B. phytofirmans YS2 ( B. phytofirmans PsJN/Δ acdS ), in which an internal segment of the acdS gene was deleted. The mutant YS2 lost ACC deaminase activity as well as the ability to promote the elongation of the roots of canola seedlings. Concomitant with the creation of this deletion mutant, a number of physiological changes were observed in the bacterium, including an increase in indole acetic acid synthesis, a decrease in the production of siderophores and an increase in the cellular level of the stationary-phase σ factor, RpoS. Introduction of the wild-type acdS gene into the mutant YS2 to construct strain B. phytofirmans YS3 ( B. phytofirmans YS2/pRK-AcdS) restored both ACC deaminase activity and plant growth-promotion activity in strain YS3. However, the complemented mutant still showed the above-mentioned physiological changes.     

Enhancement of chilling resistance of inoculated grapevine plantlets with a plant growth-promoting rhizobacterium, Burkholderia phytofirmans strain PsJN

In vitro inoculation of Vitis vinifera L. cv. Chardonnay explants with a plant growth-promoting rhizobacterium, Burkholderia phytofirmans strain PsJN, increased grapevine growth and physiological activity at a low temperature. There was a relationship between endophytic bacterial colonization of the grapevine plantlets and their growth at both ambient (26 degrees C) and low (4 degrees C) temperatures and their sensitivities to chilling. The major benefits of bacterization were observed on root growth (11.8- and 10.7-fold increases at 26 degrees C and 4 degrees C, respectively) and plantlet biomass (6- and 2.2-fold increases at 26 degrees C and 4 degrees C, respectively). The inoculation with PsJN also significantly improved plantlet cold tolerance compared to that of the nonbacterized control. In nonchilled plantlets, bacterization enhanced CO(2) fixation and O(2) evolution 1.3 and 2.2 times, respectively. The nonbacterized controls were more sensitive to exposure to low temperatures than were the bacterized plantlets, as indicated by several measured parameters. Moreover, relative to the noninoculated controls, bacterized plantlets had significantly increased levels of starch, proline, and phenolics. These increases correlated with the enhancement of cold tolerance of the grapevine plantlets. In summary, B. phytofirmans strain PsJN inoculation stimulates grapevine growth and improves its ability to withstand cold stress.     

Involvement of quinolinate phosphoribosyl transferase in promotion of potato growth by a Burkholderia strain

Burkholderia sp. strain PsJN stimulates root growth of potato explants compared to uninoculated controls under gnotobiotic conditions. In order to determine the mechanism by which this growth stimulation occurs, we used Tn5 mutagenesis to produce a mutant, H41, which exhibited no growth-promoting activity but was able to colonize potato plants as well as the wild-type strain. The gene associated with the loss of growth promotion in H41 was shown to exhibit 65% identity at the amino acid level to the nadC gene encoding quinolinate phosphoribosyltransferase (QAPRTase) in Ralstonia solanacearum. Complementation of H41 with QAPRTase restored growth promotion of potato explants by this mutant. Expression of the gene identified in Escherichia coli yielded a protein with QAPRTase activities that catalyzed the de novo formation of nicotinic acid mononucleotide (NaMN). Two other genes involved in the same enzymatic pathway, nadA and nadB, were physically linked to nadC. The nadA gene was cotranscribed with nadC as an operon in wild-type strain PsJN, while the nadB gene was located downstream of the nadA-nadC operon. Growth promotion by H41 was fully restored by addition of NaMN to the tissue culture medium. These data suggested that QAPRTase may play a role in the signal pathway for promotion of plant growth by PsJN.     

Enhancement of Chilling Resistance of Inoculated Grapevine Plantlets with a Plant Growth-Promoting Rhizobacterium, Burkholderia phytofirmans Strain PsJN

In vitro inoculation of Vitis vinifera L. cv. Chardonnay explants with a plant growth-promoting rhizobacterium, Burkholderia phytofirmans strain PsJN, increased grapevine growth and physiological activity at a low temperature. There was a relationship between endophytic bacterial colonization of the grapevine plantlets and their growth at both ambient (26°C) and low (4°C) temperatures and their sensitivities to chilling. The major benefits of bacterization were observed on root growth (11.8- and 10.7-fold increases at 26°C and 4°C, respectively) and plantlet biomass (6- and 2.2-fold increases at 26°C and 4°C, respectively). The inoculation with PsJN also significantly improved plantlet cold tolerance compared to that of the nonbacterized control. In nonchilled plantlets, bacterization enhanced CO₂ fixation and O₂ evolution 1.3 and 2.2 times, respectively. The nonbacterized controls were more sensitive to exposure to low temperatures than were the bacterized plantlets, as indicated by several measured parameters. Moreover, relative to the noninoculated controls, bacterized plantlets had significantly increased levels of starch, proline, and phenolics. These increases correlated with the enhancement of cold tolerance of the grapevine plantlets. In summary, B. phytofirmans strain PsJN inoculation stimulates grapevine growth and improves its ability to withstand cold stress.     

Enhancement of in vitro growth and resistance to gray mould of Vitis vinifera co-cultured with plant growth-promoting rhizobacteria

The potential of a plant growth-promoting rhizobacterium, Pseudomonas sp. (strain PsJN), to stimulate the growth and enhancement of the resistance of grapevine (Vitis vinifera L.) transplants to gray mould caused by Botrytis cinerea has been investigated. In vitro inoculation of grapevine plantlets induced a significant plant growth promotion which made them more hardy and vigorous when compared to non-inoculated plantlets. This ability increased upon transplanting. When grown together with B. cinerea, the causal agent of gray mould, significant differences of aggressiveness were observed between the inoculated and non-inoculated plants. The presence of bacteria was accompanied by an induction of plant resistance to the pathogen. The beneficial effect from this plant-microbe association is being postulated.     

Involvement of Quinolinate Phosphoribosyl Transferase in Promotion of Potato Growth by a Burkholderia Strain

Burkholderia sp. strain PsJN stimulates root growth of potato explants compared to uninoculated controls under gnotobiotic conditions. In order to determine the mechanism by which this growth stimulation occurs, we used Tn5 mutagenesis to produce a mutant, H41, which exhibited no growth-promoting activity but was able to colonize potato plants as well as the wild-type strain. The gene associated with the loss of growth promotion in H41 was shown to exhibit 65% identity at the amino acid level to the nadC gene encoding quinolinate phosphoribosyltransferase (QAPRTase) in Ralstonia solanacearum. Complementation of H41 with QAPRTase restored growth promotion of potato explants by this mutant. Expression of the gene identified in Escherichia coli yielded a protein with QAPRTase activities that catalyzed the de novo formation of nicotinic acid mononucleotide (NaMN). Two other genes involved in the same enzymatic pathway, nadA and nadB, were physically linked to nadC. The nadA gene was cotranscribed with nadC as an operon in wild-type strain PsJN, while the nadB gene was located downstream of the nadA-nadC operon. Growth promotion by H41 was fully restored by addition of NaMN to the tissue culture medium. These data suggested that QAPRTase may play a role in the signal pathway for promotion of plant growth by PsJN.     

Combined use of Alkane-Degrading and Plant Growth-Promoting Bacteria Enhanced Phytoremediation of Diesel Contaminated soil

Inoculation of plants with pollutant-degrading and plant growth-promoting microorganisms is a simple strategy to enhance phytoremediation activity. The objective of this study was to determine the effect of inoculation of different bacterial strains, possessing alkane-degradation and 1-amino-cyclopropane-1-carboxylic acid (ACC) deaminase activity, on plant growth and phytoremediation activity. Carpet grass (Axonopus affinis) was planted in soil spiked with diesel (1% w/w) for 90 days and inoculated with different bacterial strains, Pseudomonas sp. ITRH25, Pantoea sp. BTRH79 and Burkholderia sp. PsJN, individually and in combination. Generally, bacterial application increased total numbers of culturable hydrocarbon-degrading bacteria in the rhizosphere of carpet grass, plant biomass production, hydrocarbon degradation and reduced genotoxicity. Bacterial strains possessing different beneficial traits affect plant growth and phytoremediation activity in different ways. Maximum bacterial population, plant biomass production and hydrocarbon degradation were achieved when carpet grass was inoculated with a consortium of three strains. Enhanced plant biomass production and hydrocarbon degradation were associated with increased numbers of culturable hydrocarbon-degrading bacteria in the rhizosphere of carpet grass. The present study revealed that the combined use of different bacterial strains, exhibiting different beneficial traits, is a highly effective strategy to improve plant growth and phytoremediation activity.     

Drought stress amelioration in wheat through inoculation with Burkholderia phytofirmans strain PsJN

Plant growth promoting endophytic bacteria Burkholderia phytofirmans PsJN was used to investigate the potential to ameliorate the effects of drought stress on growth, physiology and yield of wheat (Triticum aestivum L.) under natural field conditions. Inoculated and uninoculated (control) seeds of wheat cultivar Sahar 2006 was sown in the field. The plants were exposed to drought stress at different stages of growth (tillering stage and flowering stage) by skipping the respective irrigation. The results showed that drought stress adversely affected the physiological, biochemical and growth parameters of wheat seedlings. It decreased the CO₂ assimilation, stomatal conductance, relative water content, transpiration rate and chlorophyll contents in wheat. Inoculation of wheat with PsJN significantly diluted the adverse effects of drought on relative water contents and CO₂ assimilation rate thus improving the photosynthetic rate, water use efficiency and chlorophyll content over the uninoculated control. Grain yield was also decreased when plants were exposed to drought stress at the tillering and flowering stage, but inoculation resulted in better grain yield (up to 21 and 18 % higher, respectively) than the respective uninoculated control. Similarly, inoculation improved the ionic balance, antioxidant levels, and also increased the nitrogen, phosphorus, potassium and protein concentration in the grains of wheat. The results suggested that B. phytofirmans strain PsJN could be effectively used to improve the growth, physiology and quality of wheat under drought conditions.     

Treatment of potato tubers with a growth promotingPseudomonas sp.: Plant growth responses and bacterium distribution in the rhizosphere

Rifampicin-nalidixic acid resistant mutants of a plant growth promotingPseudomonas sp., strain PsJN, were evaluated for their ability to stimulate in vitro growth of potato. Two mutant strains, MFE (a consistent growth promoter), and IIM15 (an inconsistent growth promoter), were selected for root colonization study. Root colonization of potato plants was consistently greater with MFE than with IIM15. The population density of indigenous bacteria on the root surface of potato plants inoculated with strain MFE was significantly lower as compared to non-bacterized controls and to the plants bacterized with strain IIM15. Soil sterilization did not affect plant growth in any of the treatments. Bacterization of seed tubers with strain MFE stimulated plant emergence and root development in the field, during the first two weeks after planting. Bacterized plants also formed stolons and tubers earlier and had increased yields of commercial size tubers (55 mm) as compared to non-bacterized controls. Root colonization by strain MFE was positively correlated with plant growth stimulation.     

Burkholderia phytofirmans PsJN acclimates grapevine to cold by modulating carbohydrate metabolism

Low temperatures damage many temperate crops, including grapevine, which, when exposed to chilling, can be affected by symptoms ranging from reduced yield up to complete infertility. We have previously demonstrated that Burkholderia phytofirmans PsJN, a plant growth-promoting rhizobacteria (PGPR) that colonizes grapevine, is able to reduce chilling-induced damage. We hypothesized that the induced tolerance may be explained at least partly by the impact of bacteria on grapevine photosynthesis or carbohydrate metabolism during cold acclimation. To investigate this hypothesis, we monitored herein the fluctuations of photosynthesis parameters (net photosynthesis [P(n)], intercellular CO(2) concentration, stomatal conductances, ΦPSII, and total chlorophyll concentration), starch, soluble sugars (glucose, fructose, saccharose, mannose, raffinose, and maltose), and their precursors during 5 days of chilling exposure (4°C) on grapevine plantlets. Bacterization affects photosynthesis in a non-stomatal dependent pattern and reduced long-term impact of chilling on P(n). Furthermore, all studied carbohydrates known to be involved in cold stress tolerance accumulate in non-chilled bacterized plantlets, although some of them remained more concentrated in the latter after chilling exposure. Overall, our results suggest that modification of carbohydrate metabolism in bacterized grapevine plantlets may be one of the major effects by which this PGPR reduces chilling-induced damage.     

香港西贡牛尾海邻近岛屿野生植物资源的初步研究

作者通过野外调查,报道了香港西贡牛尾海邻近岛屿的植被概况和物种多样性。结果表明:该地区共有维管束植物108科254属345种。作者将其植物资源按用途分为10类:药用植物、观赏植物、材用植物、纤维植物、油脂植物、饲用植物、有毒植物、食用植物、鞣料植物和芳香植物,并论述了各类资源植物的主要种类。并对合理开发、利用和保护该地区植物资源提出了建议。