000			07827nam a22002297a 4500
999			_c374096 _d374096
082			_a632.6 _bSHR/EN Ph.D
100			_aShree Naveena, P
245			_aEndophytic and growth promoting activities of entomopathogenic fungus Lecanicillium saksenae (Kushwaha) Kurihara and Sukarno in rice
260			_aVellayani _bDepartment of Agricultural Entomology, College of Agriculture _c2025
300			_a165p.
502			_aPh.D
520	3		_aThe study entitled “Endophytic and growth promoting activities of the entomopathogenic fungus, Lecanicilliumsaksenae (Kushwaha) Kurihara and Sukarno in rice” was carried out during 2020-2024 in the Department of Agricultural Entomology, College of Agriculture, Vellayani, Thiruvananthapuram. The objective of the study was to establish the entomopathogenic, endophytic and growth promoting attributes of L. saksenae in rice. The infection process of L. saksenae in the rice bug, investigated using Scanning Electron Microscopy (SEM), revealed five step infection processes comprising adhesion, germination, penetration, colonisation/invasion, and conidiation/dissemination at 24, 48, 72, 96 and 120 hours post inoculation (HPI), culminating in dissemination of spores facilitating horizontal transmission and complete mycosis evidenced by mummified appearance, by 144 HPI. Biochemical mechanisms involved in pathogenesis were deciphered using metabolomic analyses of the infected rice bug. Gas Chromatography-Mass Spectrometry (GC-MS) and High-Resolution ORBITRAP Liquid Chromatography Mass Spectrometry (HR-LC-MS) revealed an array of 19 insecticidal compounds, three immunosuppressors, three antimicrobial compounds, and 13 other metabolites related to insect metabolism, highlighting the complex biochemical arsenal employed by L. saksenae in insect pathogenesis. In addition to entomopathogenicity, L. saksenae exhibited significant plant growth-promoting traits. Plate assays and spectrophotometric observations, displayed elevated levels of phytohormones, notably gibberellic acid (GA3: 334.68 µg mL-1) and indole acetic acid (IAA: 30.00 µg mL-1). It also exhibited zinc and phosphate solubilization efficiency (1.15 SE and 1.70 SE), which are related to nutrient assimilation. The siderophore (2.23 AU) and ammonia production (15.42 µmol mL-1) levels which help in nutrient uptake were significant and higher than those in other entomopathogenic fungi. The study further examined endophytic association of L. saksenae upon seed inoculation, in rice through re-isolation, microscopy, and PCR. The fungus could be reisolated from roots, stems, and leaves of the inoculated plants up to 90 days after seed inoculation (DAI) demonstrating its systemic translocation within then plant. Maximum colonisation was noted in roots (72.86 per cent), followed by stem (64.29 per cent) and leaves (40.00 per cent) at 30 DAI, which tapered to 32.86, 17.14 and 11.4 per cent at 90 DAI. SEM observations confirmed internal colonisation of L. saksenae in the leaf stem and root tissues of inoculated rice, which was further established by PCR amplification, that unequivocally matched with the original Accession. MN545844 deposited by Rani et al. (2015). Comparative metabolomic analysis between inoculated and uninoculated plants revealed significant biochemical changes due to L. saksenae colonisation. Significant elevation of proteins, sugars, and phenolic compounds in the inoculated plants indicated a metabolic shift that favours enhanced growth and defence. Untargeted GC-MS analysis revealed predominance of carbohydrates at maximum tillering stage which indicates protection at cellular levels and promotion of plant growth. Untargeted ORBITRAP HR LC-MS detected the presence of 10 insecticidal/insectistatic, one nematicidal, and three antimicrobial compounds, suggesting that L. saksenae induced the defence mechanism in inoculated plants. LC-MS/MS revealed profound modulation of growth and defence related hormones at critical plant growth stages due to fungal inoculation. Inoculated plants showed increased growth related hormones such as indole-3-butyric acid (IBA), IAA, GAs, N6-benzylaminopurine (6-BAP), trans-zeatin (tZ), and trans-zeatin riboside (tZR) at the seedling stage, and IAA, IBA, GA3, GA4, GA7, and tZR at the maximum tillering stage. Defence hormones such as abscisic acid (ABA), salicylic acid (SA), jasmonic acid (JA), cis-jasmone (CJ), methyl jasmonate (MeJA), epibrassinolide (EBL), and 1 aminocyclopropane-1-carboxylic acid (ACC) were significantly higher in the inoculated plants at the seedling stage and SA, JA, CJ, MeJ, and ACC at the tillering stage, which are related to increased resistance to pest infestation. Inoculated seeds exhibited improved germination (6.38 per cent) and the seedlings exhibited increase in shoot and root length (30.30 per cent, 55.58 per cent), number of leaves and roots (35.29 per cent, 26.58 per cent) and the seedling vigor index (51.46 per cent), compared to the control. Pot culture experiments proved enhancedplant height (25.80 per cent), number of tillers (36.99 per cent), panicle count (30.00 per cent), grain number (24.20 per cent), grain yield (30.00 per cent), and straw yield (28.43 per cent) in the inoculated plants. In L. saksenae colonised, rice bug infested plants, there was a significant reduction in the number of eggs laid (29.40 per cent), number of feeding punctures (32.83 per cent) and grain damage (37.45 per cent) compared to control plants. Gene expression analysis revealed the upregulation of defence related genes, phenyl ammonia lyase (PAL) and lipoxygenase (LOX) in both inoculated and control plants, indicating it as a general stress response to herbivory. However, the pathogenicity related genes of the entomopathogens viz. subtilin like protease (PR1) and chitinase II (CHIT II) were not expressed in the colonised plants, as these genes are typically involved in breaching of insect cuticle. Headspace volatile analysis (GC-MS) of L. saksenae inoculated plants before rice bug infestation revealed the presence of 2-heptanone (natural enemy attractant and insect repellent), and 4-methylbenzaldehyde (oviposition deterrent). L. saksenae inoculated, rice bug infested plants, displayed defence related volatiles, such as S linalool (natural enemy attractant and insect repellent), methyl salicylate (natural enemy attractant), 9-octadecenamide (Z) (insecticidal), N-methyl-1-adamantaneacetamide (insecticidal and repellent). Additionally, there was upregulation of defence related enzymes, PAL, catalase (CAT), superoxide dismutase (SOD), polyphenol oxidase (PPO), peroxidase (POD), and ascorbate peroxide (APX) in inoculated plants. Temporal profiling revealed a dynamic activation pattern, where PPO, SOD, and APX peaked early at 48 hours after insect release (HAR), followed by a surge in PAL and POD at 72 HAR, and a delayed peak of CAT at 96 HAR. Importantly, L. saksenae inoculated plants displayed earlier and more intense upregulation of SOD and PAL, indicating a primed defence in plants during herbivore attack. These biochemical changes may underlie the observed reduction in pest performance and feeding efficiency. The above findings established the entomopathogenic, endophytic and growth promoting attributes of the indigenous isolate, L. saksenae in rice. By elucidating the tritrophic interactions involving the fungus, plant and insect, the study positions L. saksenae, as a "two-in-one" eco-friendly solution for plant health management in rice.
650			_a Agricultural Entomology
650			_aEntomopathogenic fungus
650			_aLecanicillium saksenae
650			_aKushwaha
650			_aSukarno
650			_aRice
700			_aReji Rani, O P (guide)
856			_uhttps://krishikosh.egranth.ac.in/handle/1/5810228094
942			_2ddc _cTH