Involvement of Peptidoglycan Receptor Proteins in Mediating the Growth-Promoting Effects of Bacillus pumilus TUAT1 in Arabidopsis thaliana.

Bacillus pumilus TUAT1 acts as plant growth-promoting rhizobacteria for various plants like rice and Arabidopsis. Under stress conditions, B. pumilus TUAT1 forms spores with a thick peptidoglycan (PGN) cell wall. Previous research showed that spores were significantly more effective than vegetative cells in enhancing plant growth. In Arabidopsis, lysin motif proteins, LYM1, LYM3 and CERK1, are required for recognizing bacterial PGNs to mediate immunity. Here, we examined the involvement of PGN receptor proteins in the plant growth promotion (PGP) effects of B. pumilus TUAT1 using Arabidopsis mutants defective in PGN receptors. Root growth of wild-type (WT), cerk1-1, lym1-1 and lym1-2 mutant plants was significantly increased by TUAT1 inoculation, but this was not the case for lym3-1 and lym3-2 mutant plants. RNA-seq analysis revealed that the expression of a number of defense-related genes was upregulated in lym3 mutant plants. These results suggested that B. pumilus TUAT1 may act to reduce the defense response, which is dependent on a functional LYM3. The expression of the defense-responsive gene, WRKY29, was significantly induced by the elicitor flg-22, in both WT and lym3 mutant plants, while this induction was significantly reduced by treatment with B. pumilus TUAT1 and PGNs in WT, but not in lym3 mutant plants. These findings suggest that the PGNs of B. pumilus TUAT1 may be recognized by the LYM3 receptor protein, suppressing the defense response, which results in plant growth promotion in a trade-off between defense and growth.

Effect of Bacterial Extracellular Polymeric Substances from Enterobacter spp. on Rice Growth under Abiotic Stress and Transcriptomic Analysis.

Plant biostimulants have received attention as sustainable alternatives to chemical fertilizers. Extracellular polymeric substances (EPSs), among the compounds secreted by plant growth-promoting rhizobacteria (PGPRs), are assumed to alleviate abiotic stress. This study aims to investigate the effect of purified EPSs on rice under abiotic stress and analyze their mechanisms. A pot experiment was conducted to elucidate the effects of inoculating EPSs purified from PGPRs that increase biofilm production in the presence of sugar on rice growth in heat-stress conditions. Since all EPSs showed improvement in SPAD after the stress, Enterobacter ludwigii, which was not characterized as showing higher PGP bioactivities such as phytohormone production, nitrogen fixation, and phosphorus solubilization, was selected for further analysis. RNA extracted from the embryos of germinating seeds at 24 h post-treatment with EPSs or water was used for transcriptome analysis. The RNA-seq analysis revealed 215 differentially expressed genes (DEGs) identified in rice seeds, including 139 up-regulated and 76 down-regulated genes. A gene ontology (GO) enrichment analysis showed that the enriched GO terms are mainly associated with the ROS scavenging processes, detoxification pathways, and response to oxidative stress. For example, the expression of the gene encoding OsAAO5, which is known to function in detoxifying oxidative stress, was two times increased by EPS treatment. Moreover, EPS application improved SPAD and dry weights of shoot and root by 90%, 14%, and 27%, respectively, under drought stress and increased SPAD by 59% under salt stress. It indicates that bacterial EPSs improved plant growth under abiotic stresses. Based on our results, we consider that EPSs purified from Enterobacter ludwigii can be used to develop biostimulants for rice.

Screening of NIAS World Rice Core Collection for Seeds with Long Longevity as Useful Potential Breeding Materials Focusing on the Stability of Embryonic RNAs.

Seed longevity is a crucial trait for the seed industry and genetic resource preservation. To develop excellent cultivars with extended seed lifespans, it is important to understand the mechanism of keeping seed germinability long term and to find useful genetic resources as prospective breeding materials. This study was conducted to identify the best cultivars with a high and stable seed longevity trait in the germplasm of rice (Oryza sativa L.) and to analyze the correlation between seed longevity and embryonic RNA integrity. Seeds from 69 cultivars of the world rice core collection selected by the NIAS in Japan were harvested in different years and subjected to long-term storage or controlled deterioration treatment (CDT). The long-term storage (4 °C, RH under 35%, 10 years) was performed on seeds harvested in 2010 and 2013. The seeds harvested in 2016 and 2019 were used for CDT (36 °C, RH of 80%, 40 days). Seed longevity and embryonic RNA integrity were estimated by a decrease in the germination percentage and RNA integrity number (RIN) after long-term storage or CDT. The RIN value was obtained by the electrophoresis of the total RNA extracted from the seed embryos. Seeds of "Vandaran (indica)", "Tupa 729 (japonica)", and "Badari Dhan (indica)" consistently showed higher seed longevity and embryonic RNA integrity both under long-term storage and CDT conditions regardless of the harvest year. A strong correlation (R2 = 0.93) was observed between the germination percentages and RIN values of the seeds after the long-term storage or CDT among nine cultivars selected based on differences in their seed longevity. The study findings revealed the relationship between rice seed longevity and embryo RNA stability and suggested potential breeding materials including both japonica and indica cultivars for improving rice seed longevity.

Influence of potassium-solubilizing bacteria on the growth and radiocesium phyto-transfer of Brassica rapa L. var. perviridis grown in contaminated Fukushima soils.

The supply of K, being the chemical analog of Cs, affects the phytotransfer of radiocesium such as 137Cs from contaminated soils and its accumulation in plant tissues. Since K and Cs have high affinity to the same clay particle surfaces, the presence of potassium-solubilizing bacteria (KSB) could increase the availability of not only K+ in the rhizosphere but also of radiocesium. In this study, we obtained five KSB isolates with the highest solubilization capacities from soybean rhizosphere on modified Aleksandrov medium containing sericite as K source. Based on biochemical and 16S rRNA gene sequence analysis, we identified the bacteria as Bacillus aryabhattai MG774424, Pseudomonas umsongensis MG774425, P. frederiksbergensis MG774426, Burkholderia sabiae MG774427, and P. mandelii MG774428. We evaluated the KSB isolates based on plant growth promotion and 137Cs accumulation in komatsuna (Brassica rapa L. var. Perviridis) grown in three soils collected from Miyanoiri, Takanishi, and Ota contaminated by 137Cs from the Fukushima accident. Inoculation with KSB showed beneficial effects on plant growth and increased the overall plant biomass production (~40%). On the average, KSB inoculation resulted in the removal of 0.07 ± 0.04% of 137Cs from the soil, more than twice the control. But similar to the effect of KSB inoculation on komatsuna biomass production, different KSBs performed variably and exhibited site-specific responses independent of their K-solubilizing capacities, with higher 137Cs phyto-transfer in roots than in shoots. In terms of root transfer factor (TF), values were highest in komatsuna plants grown in Miyanoiri and Ota soils inoculated with P. frederiksbergensis and Burkholderia sabiae, while they were highest in Takanishi soils inoculated with Bacillus aryabhattai and P. umsongensis. These TF values were also much higher than previously reported values for komatsuna grown in 137Cs-contaminated Fukushima soils inoculated with other rhizobacteria. Thus, KSB inoculation significantly enhance not only the growth of komatsuna but 137Cs uptake.

Isolation and Screening of Indigenous Plant Growth-promoting Rhizobacteria from Different Rice Cultivars in Afghanistan Soils.

To develop biofertilizers for rice in Afghanistan, 98 plant growth-promoting rhizobacteria were isolated from rice plants and their morphological and physiological characteristics, such as indole-3-acetic acid production, acetylene reduction, phosphate and potassium solubilization, and siderophore production, were evaluated. The genetic diversity of these bacteria was also analyzed based on 16S rRNA gene sequences. Of 98 bacteria, 89.7% produced IAA, 54.0% exhibited nitrogenase activity, and 40% showed phosphate solubilization and siderophore production. Some isolates assigned to Pseudomonas (brassicacearum, chengduensis, plecoglossicida, resinovorans, and straminea) formed a relationship with rice, and P. resinovorans and P. straminea showed nitrogen fixation. Rhizobium borbori and R. rosettiformans showed a relationship with rice plants and nitrogen fixation. Among the isolates examined, AF134 and AF137 belonging to Enterobacter ludwigii and P. putida produced large amounts of IAA (92.3 µg mL-1) and exhibited high nitrogenase activity (647.4 nmol C2H4 h-1), respectively. In the plant growth test, more than 70% of the inoculated isolates showed significantly increased root and shoot dry weights. Highly diverse bacterial isolates showing promising rice growth-promoting traits were obtained from Afghanistan alkaline soils.

Genetic Characterization of Soybean Rhizobia Isolated from Different Ecological Zones in North-Eastern Afghanistan.

Seventy rhizobial isolates were obtained from the root nodules of two soybean (Glycine max) cultivars: Japanese cultivar Enrei and USA cultivar Stine3300, which were inoculated with different soil samples from Afghanistan. In order to study the genetic properties of the isolates, the DNA sequences of the 16S rRNA gene and symbiotic genes (nodD1 and nifD) were elucidated. Furthermore, the isolates were inoculated into the roots of two soybean cultivars, and root nodule numbers and nitrogen fixation abilities were subsequently evaluated in order to assess symbiotic performance. Based on 16S rRNA gene sequences, the Afghanistan isolates obtained from soybean root nodules were classified into two genera, Bradyrhizobium and Ensifer. Bradyrhizobium isolates accounted for 54.3% (38) of the isolates, and these isolates had a close relationship with Bradyrhizobium liaoningense and B. yuanmingense. Five out of the 38 Bradyrhizobium isolates showed a novel lineage for B. liaoningense and B. yuanmingense. Thirty-two out of the 70 isolates were identified as Ensifer fredii. An Ensifer isolate had identical nodD1 and nifD sequences to those in B. yuanmingense. This result indicated that the horizontal gene transfer of symbiotic genes occurred from Bradyrhizobium to Ensifer in Afghanistan soil. The symbiotic performance of the 14 tested isolates from the root nodules of the two soybean cultivars indicated that Bradyrhizobium isolates exhibited stronger acetylene reduction activities than Ensifer isolates. This is the first study to genetically characterize soybean-nodulating rhizobia in Afghanistan soil.