Plant growth-promoting traits of culturable seed microbiome of citrus species from Purvanchal Himalaya.

Despite Northeastern India being "Treasure House of Citrus Genetic Wealth," genetic erosion of citrus diversity poses severe concern with a corresponding loss in seed microbial diversity. The seed microbiome of citrus species unique to the Purvanchal Himalaya is seldom explored for their use in sustainable orchard management. Isolation and characterization of culturable seed microbiomes of eight citrus species, namely, Citrus reticulata Blanco, C. grandis (L.) Osbeck, C. latipes Tanaka, C. megaloxycarpa Lushaigton, C. jambhiri Lush, C. sinensis (L.) Osbeck, C. macroptera Montr, and C. indica Tanaka collected from NE India were carried out. The isolates were then screened for an array of plant growth-promoting (PGP) traits [indole acetic acid (IAA) production, N2 fixation, phosphate and zinc complex dissolution, siderophores, and Hydrogen Cyanide (HCN) production]. The pure culture isolates of seed microbiomes were capable of dissolving insoluble Ca3(PO4)2 (1.31-4.84 µg Pi ml-1 h-1), Zn3(PO4)2 (2.44-3.16 µg Pi ml-1 h-1), AlPO4 (1.74-3.61 µg Pi ml-1 h-1), and FePO4 (1.54-4.61µg Pi ml-1 h-1), mineralized phytate (12.17-18.00 µg Pi ml-1 h-1) and produced IAA-like substances (4.8-187.29 µg ml-1 h-1). A few isolates of the seed microbiome were also able to fix nitrogen, secrete siderophore-like compounds and HCN, and dissolve ZnSO4 and ZnO. The 16S ribosomal Ribonucleic Acid (rRNA)-based taxonomic findings revealed that Bacillus was the most dominant genus among the isolates across citrus species. Isolates CG2-1, CME6-1, CME6-4, CME6-5, CME6-9, CJ7-1, CMA10-1, CI11-3, and CI11-4 were identified as promising bioinoculants for development of microbial consortium having multifaceted PGP traits for nutritional benefits of nitrogen, phosphorus and zinc, and IAA hormonal benefits to citrus crops for better fitness in acid soils.

Rapid and precise detection of cryptic tea pathogen Exobasidium vexans: RealAmp validation of LAMP approach.

This work embodies the development of a real time loop mediated isothermal amplification (RealAmp) assay for the rapid detection of the cryptic tea phytopathogen, Exobasidium vexans, the causal organism of blister blight disease. Due to the widespread popularity of tea as a beverage and the associated agro-economy, the rapid detection and management of the fast-spreading blister blight disease have been a longstanding necessity. Loop-mediated isothermal amplification (LAMP) primers were designed targeting the E. vexans ITS rDNA region and the reaction temperature was optimized at 62 °C with a 60 min reaction time. Amplification of the E. vexans isolates in the initial LAMP reactions was confirmed by both agarose gel electrophoresis and SYBR Green I dye based colour change visualization. The specificity of the LAMP primers for E. vexans was validated by negative testing of seven different phytopathogenic test fungi using LAMP and RealAmp assay. The positive findings in RealAmp assay for E. vexans strain were corroborated via detecting fluorescence signals in real-time. Further, the LAMP assays performed with gDNA isolated from infected tea leaves revealed positive amplification for the presence of E. vexans. The results demonstrate that this rapid and precise RealAmp assay has the potential to be applied for field-based detection of E. vexans in real-time.

Bipartite molecular approach for species delimitation and resolving cryptic speciation of Exobasidium vexans within the Exobasidium genus.

Exobasidium vexans, a basidiomycete pathogen, is the causal organism of blister blight disease in tea. The molecular identification of the pathogen remains a challenge due to the limited availability of genomic data in sequence repositories and cryptic speciation within its genus Exobasidium. In this study, the nuclear internal transcribed spacer rDNA region (ITS) based DNA barcode was developed for E. vexans, to address the problem of molecular identification within the background of cryptic speciation. The isolation of E. vexans strain was confirmed through morphological studies followed by molecular identification utilizing the developed ITS barcode. Phylogenetic analysis based on Maximum Parsimony (MP), Maximum Likelihood (ML) and Bayesian Inference (BI) confirmed the molecular identification of the pathogen as E. vexans strain. Further, BI analysis using BEAST mediated the estimation of the divergence time and evolutionary relationship of E. vexans within genus Exobasidium. The speciation process followed the Yule diversification model wherein the genus Exobasidium is approximated to have diverged in the Paleozoic era. The study thus sheds light on the molecular barcode-based species delimitation and evolutionary relationship of E. vexans within its genus Exobasidium.

Enhancing arsenic sequestration on ameliorated waste molasses nanoadsorbents using response surface methodology and machine-learning frameworks.

The development of a novel nanobiosorbent derived from waste molasses for the adsorptive removal of arsenic (As) has been attempted in this study. Waste molasses were chemically ameliorated through a solvothermal route for the incorporation of iron oxide, thereby producing iron oxide incorporated carbonaceous nanomaterial (IOCN). Synthesis of IOCN was confirmed through transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and atomic emission spectroscopy (AES) analysis. The surface area and porous behavior of IOCN were elucidated by Brunauer-Emmett-Teller (BET) assessments. The experimental conditions for adsorption were first modeled using response surface methodology (RSM) based on the central composite design (CCD), considering the parameters: adsorbate dosage, adsorbent dosage, pH, and contact time. RSM optimizations were improved upon using a three-layer feed-forward multilayer perceptron (MLP) based Artificial Neural Network (ANN) model. Optimization through ANN model resulted in the increase of the maximal As adsorption efficiency to ~ 96% for IOCN. The IOCN isotherm plots show the best fit for the Sips isotherm, and the reaction kinetics follows the pseudo-second-order model, indicating the chemisorption mechanism for As adsorption. Evidence for direct coordination of As to the surface of adsorbents was further confirmed by FTIR spectroscopic studies before and after As adsorption. The high adsorption efficiencies and the low-cost facile synthesis of the IOCN nanosorbent from agro-industrial waste indicate their potential for commercial applications.

Optimizing In vitro Culture Conditions for the Biotrophic Fungi Exobasidium vexans Through Response Surface Methodology.

The blister blight disease caused by the fungus, Exobasidium vexans has serious implications on the quality of tea production. The disease however, has been poorly studied and hence there is very limited information on the pathogen and as such the pathogenesis of blister blight infection. One of the major roadblocks in understanding E. vexans is the obligate and biotrophic nature of the fungus which limits the establishment and maintenance of in vitro cultures. To address this issue, a Central Composite Design based Response Surface Methodology (RSM) was adopted to study the modification of three fungal culture media viz. czapek dox, potato dextrose, and v8 juice, and the effect of altered media composition on growth conditions and media compositions were assessed. The response parameter for the RSM experiments was the mycelial biomass produced under different culture conditions. The uni and bi-parametric interactions among the experimental variables provided the basis for the statistically optimized conditions for maximal fungal growth. The study thus presents the recommended modifications of existing media that can lead to the successful establishment and maintenance of E. vexans in vitro cultures.

Glycans as Modulators of Plant Defense Against Filamentous Pathogens.

Plants and microbes utilize glycoconjugates as structural entities, energy reserves for cellular processes, and components of cellular recognition or binding events. The structural heterogeneity of carbohydrates in such systems is a result of the ability of the carbohydrate biosynthetic enzymes to reorient sugar monomers in a variety of forms, generating highly complex, linear, branched, or hierarchical structures. During the interaction between plants and their microbial pathogens, the microbial cell surface glycans, cell wall derived glycans, and glycoproteins stimulate the signaling cascades of plant immune responses, through a series of specific or broad spectrum recognition events. The microbial glycan-induced plant immune responses and the downstream modifications observed in host-plant glycan structures that combat the microbial attack have garnered immense interest among scientists in recent times. This has been enabled by technological advancements in the field of glycobiology, making it possible to study the ongoing co-evolution of the microbial and the corresponding host glycan structures, in greater detail. The new glycan analogs emerging in this evolutionary arms race brings about a fresh perspective to our understanding of plant-pathogen interactions. This review discusses the role of diverse classes of glycans and their derivatives including simple sugars, oligosaccharides, glycoproteins, and glycolipids in relation to the activation of classical Pattern-Triggered Immunity (PTI) and Effector-Triggered Immunity (ETI) defense responses in plants. While primarily encompassing the biological roles of glycans in modulating plant defense responses, this review categorizes glycans based on their structure, thereby enabling parallels to be drawn to other areas of glycobiology. Further, we examine how these molecules are currently being used to develop new bio-active molecules, potent as priming agents to stimulate plant defense response and as templates for designing environmentally friendly foliar sprays for plant protection.