Genome and secretome analysis of jute endophyte Grammothele lineata strain SDL-CO-2015-1: Insights into its lignocellulolytic structure and secondary metabolite profile.

Grammothele lineata strain SDL-CO-2015-1, jute (Corchorus olitorius) endophyte has been reported to produce anti-cancer drug paclitaxel in culture condition. Here we investigated the genome using different bioinformatic tools to find its association with the production of commercially important compounds including taxol. Carbohydrate-active enzymes, proteases, and secretory proteins were annotated revealing a complex endophytic relationship with its plant host. The presences of a diverse range of CAZymes including numerous lignocellulolytic enzymes support its potentiality in biomass degradation. Genome annotation led to the identification of 28 clusters for secondary metabolite biosynthesis. Several biosynthesis gene clusters were identified for terpene biosynthesis from antiSMASH analysis but none could be specifically pinned to taxol synthesis. This study will direct us to understand the genomic organization of endophytic basidiomycetes with a potential for producing numerous commercially important enzymes and secondary metabolites taking G. lineata as a model.

Population diversity of bacterial endophytes from jute (Corchorus olitorius) and evaluation of their potential role as bioinoculants.

Endophytes are bacterial or fungal organisms associated with plants in an obligate or facultative manner. In order to maintain a stable symbiosis, many of the endophytes produce compounds that promote plant growth and help them adapt better to the environment. This study was conducted to explore the potential of jute bacterial endophytes for their growth promotion ability in direct and indirect ways. A total of 27 different bacterial species were identified from different varieties of a jute plant (Corchorus olitorius) and different parts of the plant (leaf, root, seed, and seedling) based on 16S rRNA gene sequence. Two of the isolates showed ACC deaminase activity with Staphylococcus pasteuri strain MBL_B3 and Ralstonia solanacearum strain MBL_B6 producing 18.1 and 8.08 µM mg-1 h-1 α-ketobutyrate respectively while eighteen had the ACC deaminase gene (acdS). Fourteen were positive for siderophore activity while Kocuria sp. strain MBL_B19 (133.36 µg/ml) and Bacillus sp. strain MBL_B17 (124.72 µg/ml) showed high IAA production ability. Seven bacterial strains were able to fix nitrogen with only one testing positive for nifH gene. Five isolates exhibited phosphorus utilization ability with Bacillus sp. strain MBL_B17 producing 218.47 µg P/ml. Three bacteria were able to inhibit the growth of a phytopathogen, Macrophomina phaseolina and among them Bacillus subtilis strain MBL_B4 was found to be the most effective, having 82% and 53% of relative inhibition ratio (RIR) and percent growth inhibition (PGI) values respectively. Nine bacteria were tested for their in vivo growth promotion ability and most of these isolates increased seed germination potential and vigour index significantly. Bacillus subtilis strain MBL_B13 showed 26.8% more vigour index than the control in which no bacterial inoculum was used. All inoculants were found to increase the dry weight of jute seedlings in comparison to the control plants and the most increase in fresh weight was found for Staphylococcus saprophyticus strain MBL_B9. Staphylococcus pasteuri strain MBL_B3 exhibited diverse in vitro growth promotion activity and significant growth promoting effect in in vivo pot experiments. These bacterial strains with plant growth enhancing abilities have the potential to be used as bioinoculants.

Draft Genome Sequence of Grammothele lineata SDL-CO-2015-1, a Jute Endophyte with a Potential for Paclitaxel Biosynthesis.

Grammothele lineata strain SDL-CO-2015-1, a basidiomycete fungus, was identified as an endophyte from a jute species, Corchorus olitorius var. 2015, and found to produce paclitaxel, a diterpenic polyoxygenated pseudoalkaloid with antitumor activity. Here, we report the draft genome sequence (42.8 Mb with 9,395 genes) of this strain.

An endophytic Basidiomycete, Grammothele lineata, isolated from Corchorus olitorius, produces paclitaxel that shows cytotoxicity.

Grammothele lineata, an endophyte isolated in our laboratory from jute (Corchorus olitorius acc. 2015) was found to be a substantial paclitaxel producer. Taxol and its related compounds, produced by this endophyte were extracted by growing the fungus in simple nutrient media (potato dextrose broth, PDB). Taxol was identified and characterized by different analytical techniques (TLC, HPLC, FTIR, LC-ESI-MS/MS) following its extraction by ethyl acetate. In PDB media, this fungus was found to produce 382.2 µgL-1 of taxol which is about 7.6 x103 fold higher than the first reported endophytic fungi, Taxomyces andreanae. The extracted taxol exhibited cytotoxic activity in an in vitro culture of HeLa cancer cell line. The fungal extract also exhibited antifungal and antibacterial activities against different pathogenic strains. This is the first report of a jute endophytic fungus harboring the capacity to produce taxol and also the first reported taxol producing species that belongs to the Basidiomycota phylum, so far unknown to be a taxol producer. These findings suggest that the fungal endophyte, Grammothele lineata can be an excellent source of taxol and can also serve as a potential species for chemical and genetic engineering to enhance further the production of taxol.

Modification of Monolignol Biosynthetic Pathway in Jute: Different Gene, Different Consequence.

Lignin, a cross-linked macromolecule of hydrophobic aromatic structure, provides additional rigidity to a plant cell wall. Although it is an integral part of the plant cell, presence of lignin considerably reduces the quality of the fiber of fiber-yielding plants. Decreasing lignin in such plants holds significant commercial and environmental potential. This study aimed at reducing the lignin content in jute-a fiber crop, by introducing hpRNA-based vectors for downregulation of two monolignoid biosynthetic genes- cinnamate 4-hydroxylase (C4H) and caffeic acid O-methyltransferase (COMT). Transgenic generations, analyzed through Southern, RT-PCR and northern assays showed downregulation of the selected genes. Transgenic lines exhibited reduced level of gene expression with ~ 16-25% reduction in acid insoluble lignin for the whole stem and ~13-14% reduction in fiber lignin content compared to the control lines. Among the two transgenic plant types one exhibited an increase in cellulose content and concomitant improvement of glucose release. Composition of the lignin building blocks was found to alter and this alteration resulted in a pattern, different from other plants where the same genes were manipulated. It is expected that successful COMT-hpRNA and C4H-hpRNA transgenesis in jute will have far-reaching commercial implications leading to product diversification and value addition.

High-Throughput Sequencing Reveals Diverse Sets of Conserved, Nonconserved, and Species-Specific miRNAs in Jute.

MicroRNAs play a pivotal role in regulating a broad range of biological processes, acting by cleaving mRNAs or by translational repression. A group of plant microRNAs are evolutionarily conserved; however, others are expressed in a species-specific manner. Jute is an agroeconomically important fibre crop; nonetheless, no practical information is available for microRNAs in jute to date. In this study, Illumina sequencing revealed a total of 227 known microRNAs and 17 potential novel microRNA candidates in jute, of which 164 belong to 23 conserved families and the remaining 63 belong to 58 nonconserved families. Among a total of 81 identified microRNA families, 116 potential target genes were predicted for 39 families and 11 targets were predicted for 4 among the 17 identified novel microRNAs. For understanding better the functions of microRNAs, target genes were analyzed by Gene Ontology and their pathways illustrated by KEGG pathway analyses. The presence of microRNAs identified in jute was validated by stem-loop RT-PCR followed by end point PCR and qPCR for randomly selected 20 known and novel microRNAs. This study exhaustively identifies microRNAs and their target genes in jute which will ultimately pave the way for understanding their role in this crop and other crops.