Tongmin Sa

Enhancement of growth and salt tolerance of red pepper seedlings (Capsicum annuum L.) by regulating stress ethylene synthesis with halotolerant bacteria containing 1-aminocyclopropane-1-carboxylic acid deaminase activity

Three 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase-producing halotolerant bacteria were isolated from West Coast soil of Yellow Sea, Incheon, South Korea and evaluated for their efficiency in improving red pepper plant growth under salt stress. The strains RS16, RS656 and RS111 were identified by 16S rRNA gene sequencing as Brevibacterium iodinum, Bacillus licheniformis and Zhihengliuela alba, respectively. Two hour exposure of 100, 150 and 200 mM NaCl stress on 8 day old red pepper seedlings caused 44, 64 and 74% increase ethylene production, while at 150 mM NaCl stress, inoculation of B. licheniformis RS656, Z. alba RS111, and Br. iodinum RS16 reduces ethylene production by 44, 53 and 57%, respectively. Similarly, 3 week old red pepper plants were subjected to salt stress for two weeks and approximately ∼50% reduction in growth recorded at 150 mM NaCl stress compared to negative control whereas bacteria inoculation significantly increase the growth compared to positive control. Salt stress also caused 1.3-fold reduction in the root/shoot dry weight ratio compared to the absence of salt while bacteria inoculation retained the biomass allocation similar to control plants. The salt tolerance index (ratio of biomass of salt stressed to non-stressed plant) was also significantly increased in inoculated plants compared to non-inoculated. Increase nutrient uptakes under salt stress by red pepper further evident that bacteria inoculation ameliorates salt stress effect. In summary, this study indicates that the use of ACC deaminase-producing halotolerant bacteria mitigates the salt stress by reducing salt stress-induced ethylene production on growth of red pepper plants.

Ecological occurrence of Gluconacetobacter diazotrophicus and nitrogen-fixing Acetobacteraceae members: their possible role in plant growth promotion.

Gluconacetobacter diazotrophicus has a long-standing history of bacterial-plant interrelationship as a symbiotic endophyte capable of fixing atmospheric nitrogen. In low nitrogen fertilized sugarcane fields it plays a significant role and its occurrence was realised in most of the sugarcane growing countries. In this mini review, the association of G. diazotrophicus with sugarcane, other crop plants and with various hosts is discussed. The factors affecting survival in the rhizosphere and the putative soil mode of transmission are emphasized. In addition, other N2-fixing Acetobacteraceae members, including Gluconacetobacter azotocaptans, Gluconacetobacter johannae and Swaminathania salitolerans, occurring in coffee, corn and rice plants are also covered. Lastly, the plant-growth-promoting traits identified in this group of bacteria, including N2 fixation, phytohormone synthesis, P and Zn solubilization and biocontrol, are analysed.

Growth promotion and induction of systemic resistance in rice cultivar Co-47 (Oryza sativa L.) by Methylobacterium spp.

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

Bacillus methylotrophicus sp. nov., a methanol-utilizing, plant-growth-promoting bacterium isolated from rice rhizosphere soil.

A Gram-positive bacterium, designated strain CBMB205(T), was isolated from the rhizosphere soil of traditionally cultivated, field-grown rice. Cells were strictly aerobic, motile, rod-shaped and formed endospores. The best growth was achieved at 30°C and pH 7.0 in ammonium mineral salts (AMS) medium containing 600 mM methanol. A comparative 16S rRNA gene sequence-based phylogenetic analysis placed strain CBMB205(T) in a clade with the species Bacillus amyloliquefaciens, Bacillus vallismortis, Bacillus subtilis, Bacillus atrophaeus, Bacillus mojavensis and Bacillus licheniformis and revealed pairwise similarities ranging from 98.2 to 99.2 %. DNA-DNA hybridization experiments revealed a low level (<36 %) of DNA-DNA relatedness between strain CBMB205(T) and its closest relatives. The major components of the fatty acid profile were C₁₅:₀ anteiso, C₁₅:₀ iso, C₁₆:₀ iso and C₁₇:₀ anteiso. The diagnostic diamino acid of the cell wall was meso-diaminopimelic acid. The G+C content of the genomic DNA was 45.0 mol%. The lipids present in strain CBMB205(T) were diphosphatidylglycerol, phosphatidylglycerol, a minor amount of phosphatidylcholine and two unknown phospholipids. The predominant respiratory quinone was MK-7. Studies of DNA-DNA relatedness, morphological, physiological and chemotaxonomic analyses and phylogenetic data based on 16S rRNA gene sequencing enabled strain CBMB205(T) to be described as representing a novel species of the genus Bacillus, for which the name Bacillus methylotrophicus sp. nov. is proposed. The type strain is CBMB205(T) (=KACC 13105(T)=NCCB 100236(T)).

Synergistic effects of inoculating arbuscular mycorrhizal fungi and Methylobacterium oryzae strains on growth and nutrient uptake of red pepper (Capsicum annuum L.)

A greenhouse experiment was conducted to examine the effects of inoculation with two Methylobacterium oryzae strains (CBMB20 and CBMB110) and a consortium of three arbuscular mycorrhizal (AM) fungi on the growth of red pepper (Capsicum annum L.). Inoculation of red pepper plants with the M. oryzae strains resulted in a significant increase in root length and root fresh weight compared to untreated control plants. The combined inoculation of M. oryzae strains and AM fungi significantly increased various plant growth parameters and chlorophyll content compared to uninoculated controls. Mycorrhizal colonisation and the number of AM fungal spores were higher in co-inoculation treatments. In addition, the combined inoculation of M. oryzae strains and AM fungi resulted in significantly higher nitrogen (N) accumulation in the roots and shoots of red pepper plants compared to uninoculated controls. The combined inoculation of M. oryzae strain CBMB110 and AM fungi increased the phosphorus (P) content by 23.3% compared to untreated controls. The micronutrient content of the red pepper plants also increased in most of the inoculation treatments. A perfect mutualism among CBMB100-AMF was found which was attributed to the improved macro- and micronutrient uptake along with higher chlorophyll content in red pepper. Further research on in-depth understanding of the co-operative microbial interactions will facilitate the successful application of Methylobacterium-AM fungi products in biotechnology.

Isolation and characterization of plant growth promoting endophytic bacterial isolates from root nodule of Lespedeza sp.

Thirty-nine endophytic bacterial strains were isolated from the nodule of Lespedeza sp. grown in two different locations of South Korea. All strains were checked for their plant growth promoting (PGP) abilities under in vitro conditions. Most of the isolates showed multiple PGP activity, i.e., indole acetic acid production, ACC deaminase activity, siderophore production, and phosphate solubilization. The strains were identified by using 16S rRNA gene sequence analysis as belonging to Alphaproteobacteria, Betaproteobacteria, Actinobacteria, and Firmicutes phylum with nine different genera Arthrobacter, Bacillus, Bradyrhizobium, Burkholderia, Dyella, Methylobacterium, Microbacterium, Rhizobium, and Staphylococcus. Gene nodA amplification showed positive results only for strains from Bradyrhizobium and Rhizobium genera. The strains from Bradyrhizobium and Rhizobium genera enhanced plant growth, nodulation, and acetylene reduction activity when inoculated on Vigna unguiculata L. (cowpea), whereas other strains did not induce nodule formation but enhanced plant growth. Herbaceous legume Lespedeza sp. formed root nodules with diverse bacterial group, and probably, these bacteria can be used for stimulating plant growth.

Effect of co-inoculation of methylotrophic Methylobacterium oryzae with Azospirillum brasilense and Burkholderia pyrrocinia on the growth and nutrient uptake of tomato, red pepper and rice

The present greenhouse study was undertaken to evaluate the effects of co-inoculating methylotrophic Methylobacterium oryzae CBMB20 along with nitrogen-fixing Azospirillum brasilense CW903 or a phosphate solubilizing bacterium Burkholderia pyrrocinia CBPB-HOD on the growth and nutrient uptake of tomato, red pepper and rice. Seed inoculation and soil/foliar application of the bacterial strains alone or under dual inoculation increased the plant growth in terms of shoot or root length and increased the nutrient uptake in the plants studied compared to uninoculated control plants. Co-inoculation of M. oryzae CBMB20 with A. brasilense CW903 or B. pyrrocinia CBPB-HOD improved the N and P concentration of plants, while the results varied among the plant species tested. Also, co-inoculation of the bacterial strains increased the activity of nitrogenase, urease and phosphatase enzymes in soil when compared to uninoculated control or individual inoculations. Though the inoculation effects were analyzed at an early stage of plant growth, the results conclusively suggest that M. oryzae being compatible with other microorganisms in the rhizosphere can potentially be used as individual inoculant or co-inoculated with other plant growth promoting bacteria to increase the production in sustainable agricultural systems.

Cultivable bacteria associated with larval gut of prothiofos-resistant, prothiofos-susceptible and field-caught populations of diamondback moth, Plutella xylostella and their potential for, antagonism towards entomopathogenic fungi and host insect nutrition

Aims:  To evaluate whether the gut bacteria of insecticide‐resistant, insecticide‐susceptible and field‐caught populations of the lepidopteran insect pest diamondback moth (DBM) –Plutella xylostella (L.) – are variable and their role in host protection and nutrition.

Colonization pattern of plant root and leaf surfaces visualized by use of green-fluorescent-marked strain of Methylobacterium suomiense and its persistence in rhizosphere

The localization of bacterial cell, pattern of colonization, and survival of Methylobacterium suomiense CBMB120 in the rhizosphere of rice and tomato plants were followed by confocal laser scanning, scanning electron microscopy, and selective plating. M. suomiense CBMB120 was tagged with green fluorescent protein (gfp), and inoculation was carried out through seed source. The results clearly showed that the gfp marker is stably inherited and is expressed in planta allowing for easy visualization of M. suomiense CBMB120. The colonization differed in rice and tomato—intercellular colonization of surface-sterilized root sections was visible in tomato but not in rice. In both rice and tomato, the cells were visible in the substomatal chambers of leaves. Furthermore, the strain was able to compete with the indigenous microorganisms and persist in the rhizosphere of tomato and rice, assessed through dilution plating on selective media. The detailed ultra-structural study on the rhizosphere colonization by Methylobacterium put forth conclusively that M. suomiense CBMB120 colonize the roots and leaf surfaces of the plants studied and is transmitted to the aerial plant parts from the seed source.

A meta-analysis of arbuscular mycorrhizal effects on plants grown under salt stress

Salt stress limits crop yield and sustainable agriculture in most arid and semiarid regions of the world. Arbuscular mycorrhizal fungi (AMF) are considered bio-ameliorators of soil salinity tolerance in plants. In evaluating AMF as significant predictors of mycorrhizal ecology, precise quantifiable changes in plant biomass and nutrient uptake under salt stress are crucial factors. Therefore, the objective of the present study was to analyze the magnitude of the effects of AMF inoculation on growth and nutrient uptake of plants under salt stress through meta-analyses. For this, data were compared in the context of mycorrhizal host plant species, plant family and functional group, herbaceous vs. woody plants, annual vs. perennial plants, and the level of salinity across 43 studies. Results indicate that, under saline conditions, AMF inoculation significantly increased total, shoot, and root biomass as well as phosphorous (P), nitrogen (N), and potassium (K) uptake. Activities of the antioxidant enzymes superoxide dismutase, catalase, peroxidase, and ascorbate peroxidase also increased significantly in mycorrhizal compared to nonmycorrhizal plants growing under salt stress. In addition, sodium (Na) uptake decreased significantly in mycorrhizal plants, while changes in proline accumulation were not significant. Across most subsets of the data analysis, identities of AMF (Glomus fasciculatum) and host plants (Acacia nilotica, herbs, woody and perennial) were found to be essential in understanding plant responses to salinity stress. For the analyzed dataset, it is concluded that under salt stress, mycorrhizal plants have extensive root traits and mycorrhizal morphological traits which help the uptake of more P and K, together with the enhanced production of antioxidant enzymes resulting in salt stress alleviation and increased plant biomass.