Differential Protein Expression Analysis of Two Sugarcane Varieties in Response to Diazotrophic Plant Growth-Promoting Endophyte Enterobacter roggenkampii ED5.

Plant endophytic bacteria have many vital roles in plant growth promotion (PGP), such as nitrogen (N) fixation and resistance to biotic and abiotic stresses. In this study, the seedlings of sugarcane varieties B8 (requires a low concentration of nitrogen for growth) and GT11 (requires a high concentration of nitrogen for growth) were inoculated with endophytic diazotroph Enterobacter roggenkampii ED5, which exhibits multiple PGP traits, isolated from sugarcane roots. The results showed that the inoculation with E. roggenkampii ED5 promoted the growth of plant significantly in both sugarcane varieties. 15N detection at 60 days post-inoculation proved that the inoculation with strain ED5 increased the total nitrogen concentration in the leaf and root than control in both sugarcane varieties, which was higher in B8. Biochemical parameters and phytohormones in leaf were analyzed at 30 and 60 days after the inoculation. The results showed that the inoculation with E. roggenkampii ED5 improved the activities of superoxide dismutase (SOD), catalase (CAT), NADH-glutamate dehydrogenase (NADH-GDH), glutamine synthetase (GS), and endo-ß-1,4-glucanase, and the contents of proline and indole acetic acid (IAA) in leaf, and it was generally more significant in B8 than in GT11. Tandem Mass Tags (TMT) labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were used to perform comparative proteomic analysis in the sugarcane leaves at 30 days after inoculation with strain ED5. A total of 27,508 proteins were detected, and 378 differentially expressed proteins (DEPs) were found in the treated sugarcane variety B8 (BE) as compared to control (BC), of which 244 were upregulated and 134 were downregulated. In contrast, a total of 177 DEPs were identified in the treated sugarcane variety GT11 (GE) as compared to control (GC), of which 103 were upregulated and 74 were downregulated. The DEPs were associated with nitrogen metabolism, photosynthesis, starch, sucrose metabolism, response to oxidative stress, hydrolase activity, oxidative phosphorylation, glutathione metabolism, phenylpropanoid metabolic process, and response to stresses in Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) database. To the best of our knowledge, this is the first proteomic approach to investigate the molecular basis of the interaction between N-fixing endophytic strain E. roggenkampii ED5 and sugarcane.

Whole Genome Analysis of Sugarcane Root-Associated Endophyte Pseudomonas aeruginosa B18-A Plant Growth-Promoting Bacterium With Antagonistic Potential Against Sporisorium scitamineum.

Sugarcane smut is a significant fungal disease that causes a major loss in sugar yield and quality. In this study, we isolated an endophytic strain B18 from a sugarcane root, which showed plant growth-promotion, hydrolytic enzyme production, antifungal activity against sugarcane pathogens (Sporisorium scitamineum, Ceratocystis paradoxa, Fusarium verticillioides), and the presence of nifH, acdS, and antibiotic genes (hcn, prn, and phCA) under in vitro conditions. BIOLOG(R) phenotypic profiling of B18 established its ability to use various carbon and nitrogen sources and tolerate a range of pH and osmotic and temperature stresses. Whole-genome analysis of B18, identified as Pseudomonas aeruginosa, showed that it consists of a single circular chromosome of 6,490,014 bp with 66.33% GC content. Genome annotation has identified 5,919 protein-coding genes, and 65 tRNA, and 12 rRNA genes. The P. aeruginosa B18 genome encodes genes related to ethylene, nitrogen (nifU, norBCDERQ, gltBDPS, and aatJMPQ), and phosphate (pstABCS and phoBDHRU) metabolism and produce indole-3-acetic acid and siderophores. This also includes genes encoding hydrolases and oxidoreductases, those associated with biocontrol mechanisms (hcnABC, phzA_B, phzDEFGMS, and pchA), colonization (minCDE and lysC), and biofilm formation (efp, hfq, flgBCDEFGHI, and motAB), and those associated with metabolism of secondary metabolites. Collectively, these results suggest a role for P. aeruginosa B18 in plant growth enhancement and biocontrol mechanisms. The P. aeruginosa B18 strain was found to be an efficient colonizer in sugarcane; it can improve growth through modulation of plant hormone production and enhanced host-plant resistance to smut pathogen S. scitamineum in a smut-susceptible sugarcane variety (Yacheng71-374). These biocontrol and plant growth promotion properties of P. aeruginosa B18 area are discussed in this report.

Diversity of nitrogen-fixing rhizobacteria associated with sugarcane: a comprehensive study of plant-microbe interactions for growth enhancement in Saccharum spp.

BACKGROUND: Nitrogen is an essential element for sugarcane growth and development and is generally applied in the form of urea often much more than at recommended rates, causing serious soil degradation, particularly soil acidification, as well as groundwater and air pollution. In spite of the importance of nitrogen for plant growth, fewer reports are available to understand the application and biological role of N2 fixing bacteria to improve N2 nutrition in the sugarcane plant. RESULTS: In this study, a total of 350 different bacterial strains were isolated from rhizospheric soil samples of the sugarcane plants. Out of these, 22 isolates were selected based on plant growth promotion traits, biocontrol, and nitrogenase activity. The presence and activity of the nifH gene and the ability of nitrogen-fixation proved that all 22 selected strains have the ability to fix nitrogen. These strains were used to perform 16S rRNA and rpoB genes for their identification. The resulted amplicons were sequenced and phylogenetic analysis was constructed. Among the screened strains for nitrogen fixation, CY5 (Bacillus megaterium) and CA1 (Bacillus mycoides) were the most prominent. These two strains were examined for functional diversity using Biolog phenotyping, which confirmed the consumption of diverse carbon and nitrogen sources and tolerance to low pH and osmotic stress. The inoculated bacterial strains colonized the sugarcane rhizosphere successfully and were mostly located in root and leaf. The expression of the nifH gene in both sugarcane varieties (GT11 and GXB9) inoculated with CY5 and CA1 was confirmed. The gene expression studies showed enhanced expression of genes of various enzymes such as catalase, phenylalanine-ammonia-lyase, superoxide dismutase, chitinase and glucanase in bacterial-inoculated sugarcane plants. CONCLUSION: The results showed that a substantial number of Bacillus isolates have N-fixation and biocontrol property against two sugarcane pathogens Sporisorium scitamineum and Ceratocystis paradoxa. The increased activity of genes controlling free radical metabolism may at least in part accounts for the increased tolerance to pathogens. Nitrogen-fixation was confirmed in sugarcane inoculated with B. megaterium and B. mycoides strains using N-balance and 15N2 isotope dilution in different plant parts of sugarcane. This is the first report of Bacillus mycoides as a nitrogen-fixing rhizobacterium in sugarcane.

Diazotrophic Bacteria Pantoea dispersa and Enterobacter asburiae Promote Sugarcane Growth by Inducing Nitrogen Uptake and Defense-Related Gene Expression.

Sugarcane is a major crop in tropical and subtropical regions of the world. In China, the application of large amounts of nitrogen (N) fertilizer to boost sugarcane yield is commonplace, but it causes substantial environmental damages, particularly soil, and water pollution. Certain rhizosphere microbes are known to be beneficial for sugarcane production, but much of the sugarcane rhizosphere microflora remains unknown. We have isolated several sugarcane rhizosphere bacteria, and 27 of them were examined for N-fixation, plant growth promotion, and antifungal activity. 16S rRNA gene sequencing was used to identify these strains. Among the isolates, several strains were found to have a relatively high activity of nitrogenase and ACC deaminase, the enzyme that reduces ethylene production in plants. These strains were found to possess nifH and acdS genes associated with N-fixation and ethylene production, respectively. Two of these strains, Pantoea dispersa-AA7 and Enterobacter asburiae-BY4 showed maximum plant growth promotion (PGP) and nitrogenase activity, and thus they were selected for detailed analysis. The results show that they colonize different sugarcane tissues, use various growth substrates (carbon and nitrogen), and tolerate various stress conditions (pH and osmotic stress). The positive effect of AA7 and BY4 strains on nifH and stress-related gene (SuCAT, SuSOD, SuPAL, SuCHI, and SuGLU) expression and the induction of defense-related processes in two sugarcane varieties, GT11 and GXB9, showed their potential for stress amelioration and PGP. Both bacterial strains increased several sugarcane physiological parameters. i.e., plant height, shoot weight, root weight, leaf area, chlorophyll content, and photosynthesis, in plants grown under greenhouse conditions. The ability of rhizobacteria on N-fixing in sugarcane was also confirmed by a 15N isotope-dilution study, and the estimate indicates a contribution of 21-35% of plant nitrogen by rhizobacterial biological N fixation (BNF). This is the first report of sugarcane growth promotion by N-fixing rhizobacteria P. dispersa and E. asburiae strains. Both strains could be used as biofertilizer for sugarcane to minimize nitrogen fertilizer use and better disease management.

Chitinolytic activity of cold tolerant antagonistic species of streptomyces isolated from glacial sites of Indian himalaya.

Seventy-eight isolates of actinomycetes were isolated from the soil samples collected from alpine zones of Pindari glacier region in Indian Himalaya. Following a plate based rapid screening using two test fungi, five efficient isolates (nos. HA1, HA2, HA6, HA40, and HA142) were selected for further characterization with special reference to their antagonistic properties. Based on phenotypic and genotypic characters, the isolates were identified up to species level. All the isolates belonged to the genus Streptomyces. The isolate nos. HA1 and HA2 were S. sampsonii and HA6, HA40 and HA142 were S. griseobrunneus, S. aurantiacus, and S. griseoluteus, respectively. The isolates showed strong antifungal properties against phytopathogenic test fungi in plate assays. All the isolates hydrolyzed glycol-chitin as a substrate in denaturing conditions showing variable amount of different isoforms.