Characterization of root-endophytic actinobacteria from cactus (Opuntia ficus-indica) for plant growth promoting traits.

The present study is the first report of isolation and characterization of endophytic actinobacteria from cactus (Opuntia ficus-indica). A total of 179 morphologically distinct endophytic actinobacterial isolates were purified from the roots of two different genetic accessions of cactus. All these isolates were screened for their plant growth promotion traits, namely, growth on N-free medium, P-solubilization, siderophore production, ACC deaminase activity and auxin production. A majority of the endophytic actinobacterial isolates (85%) exhibited their potential for plant growth promotion under in vitro conditions. Ten among the isolates were selected based on their multi-PGP traits and were identified as Streptomyces sp. following the 16S rRNA gene sequencing and phylogenetic analysis. Plant growth promotion potential of these selected endophytic Streptomyces was studied in wheat seedlings. All these selected isolates significantly enhanced the growth parameters such as seedling length and rootlets number compared to the uninoculated control. The wheat seeds inoculated with Streptomyces tuirus VL-70-IX exhibited maximum number of rootlets (6.33) compared to uninoculated control (3.67). The inoculation of endophytic actinobacteria Streptomyces levis VL-70-XII caused maximum seedling length (20.53 cm) and root length (8.26 cm), while the inoculation of S. radiopugnans HV-VIII resulted in highest shoot length (12.33 cm). These endophytic actinobacteria isolated from the roots of cactus accessions showed potential PGP traits. This work lays foundation for characterization and selection of endophytic actinobacteria from the under-exploited, drought tolerant species such as cactus with potential cross-compatibility for the improvement of plant growth of field crops especially under abiotic stress conditions.

Co-cultivation Approach to Decipher the Influence of Nitrogen-Fixing Cyanobacterium on Growth and N Uptake in Rice Crop.

The present study was performed to evaluate the efficacy of selected potential nitrogen-fixing cyanobacterial strain (Anabaena sp.), isolated from rhizospheric soil of rice plants on growth, pigments, N uptake, root architecture, and image-based phenotypic traits of rice crop using co-cultivation approach under controlled sand culture conditions. We studied the beneficial interaction of cyanobacterium to rice using sensor image-based Phenomics approach as well as conventional methods. Co-cultivation experiment revealed that inoculation with Anabaena sp. significantly improved plant growth, chlorophyll, leaf area, % nitrogen, and protein of rice by ~ 70%, ~ 22%, ~ 60%, and ~ 25% under 100% nitrogen input in comparison with un-inoculated control. Further, comparative evaluation revealed superior performance of Anabaena sp. at 100% and 75% N followed by 50% N input improving below-ground parameters as well as phenotypic traits as compared to control treatment. Hence, inoculation performed better with inorganic nitrogen input for overall growth of rice crop. Therefore, cyanobacterial strain can be used as an efficient bio-inoculant for sustainable rice production under integrated nutrient management.

Functional and phylogenetic diversity of cultivable rhizobacterial endophytes of sorghum [Sorghum bicolor (L.) Moench].

A diverse group of bacteria colonize the exo- and endo-rhizospheres of sorghum and play a critical role in its tolerance to drought and other abiotic stresses. Two hundred and eighty endophytic bacteria were isolated from the surface-sterilized roots of four sorghum cultivars that were grown on three soil types at three different phenological stages of growth. The isolates were subjected to in vitro screening for their plant growth promoting traits. Out of 280 isolates, 70 could produce Indole 3-Acetic Acid (IAA), 28 showed N-fixation, 28 could solubilize phosphate, 24 had ACC deaminase activity and 13 isolates were able to produce siderophores. Functional diversity grouping of the isolates indicated one isolate having five PGP traits and two isolates having four PGP traits; two and 29 isolates having three and two PGP traits, respectively. Among the thirty-four isolates that possessed multiple PGP traits, 19 and 17 isolates were able to produce significant quantities of IAA in the presence and absence of L-tryptophan, an inducer. Eight isolates possessed high levels of ACC deaminase activity. PCR-RFLP of the 16Sr RNA gene revealed a distinct clustering and considerable genetic diversity among these functionally characterized isolates. The 16S rRNA gene based identification of the isolates of single and multiple PGP traits revealed phylogenetic dominance of Firmicutes; Acinetobacter, Bacillus, Enterobacter, Geobacillus, Lysinibacillus, Microbacterium, Ochrobactrum, Paenibacillus and Pseudomonas were the major genera present in the endo-rhizosphere of sorghum. Results of this study are constructive in selection of effective rhizobacterial endophytes or consortia for drought stress alleviation in sorghum.

Optimizing plant growth, nutrient uptake, and yield of onion through the application of phosphorus solubilizing bacteria and endophytic fungi.

Introduction: The application of mineral fertilizers deteriorates soil properties and affects crop yield and nutritional properties. However, plant growth-promoting microorganisms (PGPM- Serendipita indica, phosphorus solubilizing bacteria (PSB), and vesicular arbuscular mycorrhizae (VAM)) have great potential to reduce fertilizers and improve soil fertility, crop yield, and nutrient uptake and mitigate the environmental effect of mineral fertilizers. Material and methods: Hence, a field experiment was conducted involving nine treatments to evaluate the effects of PGPM along with 50% or 100% of the recommended dose of fertilizers on plant growth, soil fertility, nutrient uptake, and onion productivity. Results and discussion: Results indicated that 100% RDF combined with S. indica or PSB led to improved plant growth, and higher nutrient concentrations in both leaves and bulbs of onions compared to RDF alone. Moreover, the application of 100% RDF with S. indica increased total dry matter yield by 11.5% and 7.6% in the 2018-2019 and 2019-2020 seasons, respectively, compared to 100% RDF alone. This treatment also resulted in the highest nutrient uptake, with N uptake increasing by 6.9%-29.9%, P by 13.7%-21.7%, K by 20.0%-23.7%, and S by 18.1%-23.4%. Additionally, the combination of 100% RDF with S. indica inoculation led to a notable increase in bulb yield, with increments of 16.2% and 13.9% observed in 2018-2019 and 2019-2020, respectively, compared to 100% RDF alone. Similarly, the application of 100% RDF along with PSB inoculation resulted in an increase in bulb yield by 7.2% and 9.4% in the respective years. However, VAM did not exhibit satisfactory performance or improvements in the onion crop. Conclusion: Overall, the study suggests that combining 100% RDF with S. indica or PSB can enhance onion productivity and nutrient use efficiency. The present study may open a new avenue of PGPM application in enhancing onion yield and improving the bulb quality as well as soil health. However, field trials across different regions and soil types are necessary to validate these findings for practical adoption by farmers.

Community-forming traits play role in effective colonization of plant-growth-promoting bacteria and improved plant growth.

Community-forming traits (CFts) play an important role in the effective colonization of plant-growth-promoting bacterial communities that influence host plants positively by modulating their adaptive functions. In this study, by considering plant-growth-promoting traits (PGPts) and community-forming traits (CFts), three communities were constructed, viz., SM1 (PGPts), SM2 (CFts), and SM3 (PGPts+CFts). Each category isolates were picked up on the basis of their catabolic diversity of different carbon sources. Results revealed a distinctive pattern in the colonization of the communities possessed with CF traits. It was observed that the community with CFts colonized inside the plant in groups or in large aggregations, whereas the community with only PGPts colonized as separate individual and small colonies inside the plant root and leaf. The effect of SM3 in the microcosm experiment was more significant than the uninoculated control by 22.12%, 27.19%, and 9.11% improvement in germination percentage, chlorophyll content, and plant biomass, respectively. The significant difference shown by the microbial community SM3 clearly demonstrates the integrated effect of CFts and PGPts on effective colonization vis-à-vis positive influence on the host plant. Further detailed characterization of the interaction will take this technology ahead in sustainable agriculture.

Use of Phenomics for Differentiation of Mungbean (Vigna radiata L. Wilczek) Genotypes Varying in Growth Rates Per Unit of Water.

In the human diet, particularly for most of the vegetarian population, mungbean (Vigna radiata L. Wilczek) is an inexpensive and environmentally friendly source of protein. Being a short-duration crop, mungbean fits well into different cropping systems dominated by staple food crops such as rice and wheat. Hence, knowing the growth and production pattern of this important legume under various soil moisture conditions gains paramount significance. Toward that end, 24 elite mungbean genotypes were grown with and without water stress for 25 days in a controlled environment. Top view and side view (two) images of all genotypes captured by a high-resolution camera installed in the high-throughput phenomics were analyzed to extract the pertinent parameters associated with plant features. We tested eight different multivariate models employing machine learning algorithms to predict fresh biomass from different features extracted from the images of diverse genotypes in the presence and absence of soil moisture stress. Based on the mean absolute error (MAE), root mean square error (RMSE), and R squared (R 2) values, which are used to assess the precision of a model, the partial least square (PLS) method among the eight models was selected for the prediction of biomass. The predicted biomass was used to compute the plant growth rates and water-use indices, which were found to be highly promising surrogate traits as they could differentiate the response of genotypes to soil moisture stress more effectively. To the best of our knowledge, this is perhaps the first report stating the use of a phenomics method as a promising tool for assessing growth rates and also the productive use of water in mungbean crop.

Microbial Community and Function-Based Synthetic Bioinoculants: A Perspective for Sustainable Agriculture.

Interactions among the plant microbiome and its host are dynamic, both spatially and temporally, leading to beneficial or pathogenic relationships in the rhizosphere, phyllosphere, and endosphere. These interactions range from cellular to molecular and genomic levels, exemplified by many complementing and coevolutionary relationships. The host plants acquire many metabolic and developmental traits such as alteration in their exudation pattern, acquisition of systemic tolerance, and coordination of signaling metabolites to interact with the microbial partners including bacteria, fungi, archaea, protists, and viruses. The microbiome responds by gaining or losing its traits to various molecular signals from the host plants and the environment. Such adaptive traits in the host and microbial partners make way for their coexistence, living together on, around, or inside the plants. The beneficial plant microbiome interactions have been exploited using traditional culturable approaches by isolating microbes with target functions, clearly contributing toward the host plants' growth, fitness, and stress resilience. The new knowledge gained on the unculturable members of the plant microbiome using metagenome research has clearly indicated the predominance of particular phyla/genera with presumptive functions. Practically, the culturable approach gives beneficial microbes in hand for direct use, whereas the unculturable approach gives the perfect theoretical information about the taxonomy and metabolic potential of well-colonized major microbial groups associated with the plants. To capitalize on such beneficial, endemic, and functionally diverse microbiome, the strategic approach of concomitant use of culture-dependent and culture-independent techniques would help in designing novel "biologicals" for various crops. The designed biologicals (or bioinoculants) should ensure the community's persistence due to their genomic and functional abilities. Here, we discuss the current paradigm on plant-microbiome-induced adaptive functions for the host and the strategies for synthesizing novel bioinoculants based on functions or phylum predominance of microbial communities using culturable and unculturable approaches. The effective crop-specific inclusive microbial community bioinoculants may lead to reduction in the cost of cultivation and improvement in soil and plant health for sustainable agriculture.

Multi-trait PGP rhizobacterial endophytes alleviate drought stress in a senescent genotype of sorghum [Sorghum bicolor (L.) Moench].

Root-tissue colonizing bacteria demonstrated with multiple PGP traits from sorghum plants were identified as Ochrobactrum sp. EB-165, Microbacterium sp. EB-65, Enterobacter sp. EB-14 and Enterobacter cloacae strain EB-48 on the basis of 16S rRNA gene sequencing. Here, the in vivo experiments using ½-MS media and ½-MS media + 15% PEG 8000 (for inducing drought stress) indicated stress tolerance imparting ability of these rhizobacterial endophytes in a non-stay green and senescent genotype (R-16) of sorghum. In the experiment with sterile soilrite mix base, seed bacterization with these isolates showed improved plant growth specifically the roots, in terms of root length (~ 44.2 to 50.8% over controls), root dry weight (~ 91.3 to 99.8% over controls) and root surface area (~ 1 to 1.5 fold over controls) under drought stress. Rhizobacterial endophytes were successful, not only in providing better cellular osmotic adjustment in leaves (≥ 1-fold increase in proline accumulation over controls), but favorable physiological responses like Relative Water Content (RWC) and cell Membrane Stability Index (MSI) in the inoculated plants during the drought stress induction. Up-regulation of drought responsive genes like sbP5CS2 and sbP5CS1 was observed in these endophytes-treated plants as compared to untreated control and Escherichia coli DH5α (negative control)-treated plants. Interestingly, the stress imparting traits of rhizobacterial endophytes, including up-regulation of specific genes, were observed during sorghum seedling growth only under drought stresses. The results of this study lead to the conclusion that the potential endophytic rhizobacterial interactions can contribute to plant growth promotion as well as induced stress tolerance in sorghum.

Whole genome shotgun sequence of Bacillus paralicheniformis strain KMS 80, a rhizobacterial endophyte isolated from rice (Oryza sativa L.).

Bacillus paralicheniformis strain KMS 80 (MTCC No. 12704) is an isolate from the root tissues of rice (Oryza sativa L.) that displays biological nitrogen fixation and plant growth promoting abilities. Here, we report the complete genome sequence of this strain, which contains 4,566,040 bp, 4424 protein-coding genes, 8692 promoter sequences, 67 tRNAs, 20 rRNA genes with six copies of 5S rRNAs along with a single copy of 16S-23S rRNA and genome average GC-content of 45.50%. Twenty one genes involved in nitrogen metabolism pathway and two main transcriptional factor genes, glnR and tnrA responsible for regulation of nitrogen fixation in Bacillus sp. were predicted from the whole genome of strain KMS 80. Analysis of the ~ 4.57 Mb genome sequence will give support to understand the genetic determinants of host range, endophytic colonization behaviour as well as to enhance endophytic nitrogen fixation and other plant beneficial role of B. paralicheniformis in rice.

Assimilates mobilization, stable canopy temperature and expression of expansin stabilizes grain weight in wheat cultivar LOK-1 under different soil moisture conditions.

BACKGROUND: Grain yield of wheat is primarily determined by both grain number and grain weight, which often influence each other in response to environmental stimuli. Some of the genotypes are capable of maintaining high single grain weight (SGW) across the environments. Understanding mechanisms and factors associated with the superiority of such genotypes over others is necessary to enhance productivity of wheat. RESULTS: Experiments were conducted to elucidate the physiological basis of high SGW of LOK-1, a wheat cultivar grown in dry and hot environments in the central and peninsular zones of India. SGW of LOK-1 was least affected by removal of spikelets indicating little competition between the grains within the spike for assimilates. Reduction in SGW due to defoliation was less and the contribution of stem reserves to the grain development was high in LOK-1 relative to other cultivars. It seems that high level of expression of genes such as expansin (TaExpA6) contributes to the high SGW of LOK-1. CONCLUSIONS: Source was not a limiting factor for grain growth of LOK-1 in contrast to other cultivars, whereas sink appeared to be a limiting factor in recently released/identified cultivars. Differences in the amounts of water soluble stem carbohydrate reserves translocated to grain could be one of the factors contributing to higher grain weight in LOK-1. High level expression of TaExpA6, one of the genes contributing to the elongation of endosperm, seems to be crucial for grain growth in wheat.

Draft Genome Sequence of Pseudomonas stutzeri Strain KMS 55, an Endophytic Diazotroph Isolated from Rice Roots.

Pseudomonas stutzeri strain KMS 55 (MTCC 12703) is an isolate from the root tissues of rice (Oryza sativa L.) that displays a high biological nitrogen fixation ability. Here, we report the complete genome sequence of this strain, which contains 4,637,820 bp, 4,289 protein-coding genes, 5,006 promoter sequences, 62 tRNAs, a single copy of 5S-16S-23S rRNA, and a genome average GC content of 51.18%. Analysis of the ~4.64-Mb genome sequence will give support to increased understanding of the genetic determinants of host range, endophytic colonization behavior, endophytic nitrogen fixation, and other plant-beneficial roles of Pseudomonas stutzeri.

Comparative conventional and phenomics approaches to assess symbiotic effectiveness of Bradyrhizobia strains in soybean (Glycine max L. Merrill) to drought.

Symbiotic effectiveness of rhizobitoxine (Rtx)-producing strains of Bradyrhizobium spp. in soybean (cultivar NRC-37/Ahilya-4) under limited soil moisture conditions was evaluated using phenomics tools such as infrared(IR) thermal and visible imaging. Red, green and blue (RGB) colour pixels were standardized to analyse a total of 1017 IR thermal and 692 visible images. Plants inoculated with the Rtx-producing strains B. elkanii USDA-61 and USDA-94 and successive inoculation by B. diazoefficiens USDA-110 resulted in cooler canopy temperatures and increased canopy greenness. The results of the image analysis of plants inoculated with Rtx-producing strains were correlated with effective nodulation, improved photosynthesis, plant nitrogen status and yield parameters. Principal component analysis (PCA) revealed the reliability of the phenomics approach over conventional destructive approaches in assessing the symbiotic effectiveness of Bradyrhizobium strains in soybean plants under watered (87.41-89.96%) and water-stressed (90.54-94.21%) conditions. Multivariate cluster analysis (MCA) revealed two distinct clusters denoting effective (Rtx) and ineffective (non-Rtx) Bradyrhizobium inoculation treatments in soybean. Furthermore, correlation analysis showed that this phenotyping approach is a dependable alternative for screening drought tolerant genotypes or drought resilience symbiosis. This is the first report on the application of non-invasive phenomics techniques, particularly RGB-based image analysis, in assessing plant-microbe symbiotic interactions to impart abiotic stress tolerance.

Isolation and characterization of ACC deaminase gene from two plant growth-promoting rhizobacteria.

Lowering of plant ethylene by deamination of its immediate precursor 1-aminocyclopropane-1-carboxylate (ACC) is a key trait found in many rhizobacteria. We isolated and screened bacteria from the rhizosphere of wheat for their ACC-degrading ability. The ACC deaminase gene (acdS) isolated from two bacterial isolates through PCR amplification was cloned and sequenced. Nucleotide sequence alignment of these genes with previously reported genes of Pseudomonas sp. strain ACP and Enterobacter cloacae strain UW4 showed variation in their sequences. In the phylogenetic analysis, distinctness of these two genes was observed as a separate cluster. 16S rDNA sequencing of two isolates identified them to be Achromobacter sp. and Pseudomonas stutzeri.