Comparative Appraisal of Leaf Proteomic and Mass Spectrometry Analyses During Fusarium Wilt Infection in Resistance and Susceptible Genotypes of Castor (Ricinus communis L.).

The resistant and susceptible genotypes of castor were utilized for leaf proteomic study during Fusarium wilt infection. The histopathological study was observed under SEM and it confirmed that the infection of Fusarium oxysporum f. sp. ricini was higher in the root of susceptible JI-35, while incompatible interaction is observed in resistant SKI-215 genotype. The acidic and neutral proteins were maximally up-expressed with 2 to 171 kDa in treated resistant and 2 to 150 kDa in treated susceptible interactions. In resistant genotype, the leaf proteins were recognized with 3.0- and 5.8-fold higher at infection stage and post-infection stage, respectively, as compared to susceptible genotype. The highly up expressions of leaf acidic (4.76 pI) and basic (8.77 pI) proteins were found with 224.94- and 61.68-fold change, respectively during the post-infection stage in treated resistance compared to its control. The protein spots at 4.76 pI and 8.77 pI were characterized with nanoLC-MS Triple TOF and were recognized as signalling molecules small GTP binding protein (23 kDa) and actin (8 kDa), respectively, on the basis of mass spectrometry and peptide sequences. However, basic and neutral proteins were up regulated as 30.11- and 20.30-fold changes in treated susceptible compared to its control. These proteins were identified as HSP90 (10 kDa) and LEA (27 kDa) proteins. The 148 kDa protein is recognized as histidine kinase in incompatible resistant interaction compared to compatible susceptible (serine threonine protein kinase, 65 kDa) as common acidic protein at 3.80 pI during infection stage. Some acidic proteins were maximally up-regulated in the leaf of resistant castor genotype and played a significant role in defense response.

Biosynthesis and characterization of extracellular metabolites-based nanoparticles to control the whitefly.

The Beauveria spp. were isolated from soil and insect cadavers and confirmed as Beauveria bassiana by molecular identification using a specific primer. The bioefficacy of 14 B. bassiana against whiteflies indicated the highest percent mortality in JAU2, followed by JAU1. The LC50 and LC90 values were found to be 0.043 × 105 and 0.05 × 1014 conidia.ml-1, respectively, in JAU2. Extracellular metabolites of B.bassiana are derived and used for the green synthesis of silver nanoparticles (AgNPs). The synthesized green AgNPs were characterized for size (24.8 nm), shape (scanning electron microscopy), stability (200 mV zeta), and purity (energy-dispersive X-ray spectroscopy, 3 keV). A total of 63 extracellular metabolites were identified using LC-MS/QTOF in potent JAU2 with recognition of alcohols, phenols, carboxylic acids, amines, alkynes, and amides as functional groups. The functional groups of green AgNPs were also confirmed in Fourier transforms infrared spectroscopy (FTIR) with the specific spectra in the electromagnetic spectrum. The relationship between identified metabolites of antagonist and the FTIR spectrum of the functional group indicated the involvement of extracellular novel compounds, viz., homoisocitrate, aconitine, phodexin A, capillone, solanocapsine, and anethole in the synthesis of green AgNPs. The efficacy of green AgNPs on whiteflies suggested that corrected percent mortality was observed at 60 µg Ag.ml-1 at 120 h, which corresponds to the LC50 value (66.42 µg Ag.ml-1). Results were interpreted to show that green AgNPs synthesized from extracellular metabolites of B.bassiana JAU2 gave better insecticidal activity at LC50 as compared to live antagonist JAU2.

Characterization and bio-efficacy of entomopathogenic Beauveria associated with cuticle-degrading enzymes to restrain sucking pest Bemisia tabaci.

The entomopathogenic Beauveria spp. were acquired from insect cadavers and soil rhizosphere of cotton, groundnut, and castor. Among Beauveria, five spp. derived from infected insects, eight Beauveria found from soil, and one strain of Beauveria bassiana collected from MTCC 9544. Beauveria were characterized for morphology and cuticle-degrading enzyme activity associated with virulence against Bemisia tabaci. The colony morphology, conidial arrangement, size, and shape confirmed all isolates as Beauveria. The chitinase (EC 3.2.1.14) and lipase (EC 3.1.1.3) activities were observed the highest in Beauveria JAU2, while higher protease (EC 3.4.21.4) activity found in JAU4 followed by JAU2 at 240 h. The bio-efficacy of Beauveria (1 × 107 conidia.ml-1) illustrated that potent JAU2 was examined with the highest % mortality and corrected mortality of B. tabaci at 144 h followed by JAU1. The LC90 and LC50were determined from potent (JAU1 and JAU2) and weak (JAU6), and it was found the lowest in JAU2. The most potent Beauveria JAU2, isolated from insect cadaver (Harmivora armigera), was illustrated higher virulence than other isolates. The Beauveria JAU2 were recognized as Beauveria bassiana based on the shape of conidia and size (2.00 to 2.09 µm dia) as examined in SEM. Study insight into recognition of potent Beauveria bassiana JAU2 was linked with cuticle-degrading enzyme activity for insecticidal action. The JAU2 isolate established the most positive correlation (P0.01: 0.864) between chitinase activity and corrected mortality of insect.

Green synthesis and characterization of nanosilver derived from extracellular metabolites of potent Bacillus subtilis for antifungal and eco-friendly action against phytopathogen.

The potent antagonist Bacillus isolated from the soil rhizosphere elucidated the highest antagonism against the phytopathogen Fusarium oxysporum f. sp. cumini and was identified as Bacillus subtilis strain JSD-RSCu-8D based on molecular recognition by 16S rRNA sequencing (NCBI Accession No. KT894724). Live Bacillus may not work as effectively against phytopathogen under unfavorable environmental conditions like temperature, humidity, or other abiotic stresses. The extracellular metabolites, obtained from culturing potent B. subtilis, were exploited for the creation of green nanosilver for proficient actions in a changing climate. The synthesized green nanosilver was illustrated for shape (spherical with 65.21 ± 3.71 nm under SEM), size (70.9 nm in PSA), purity (2.69 keV peak corresponded to the binding energy of silver under EDAX), and stability (44.2 mV as ZETA). The formation of green Ag-NPs from extracellular metabolites was confirmed by a comparative appraisal of the electromagnetic peak of the metabolite's functional groups, silver nitrate, and green nanoparticles in Fourier transform infrared spectroscopy. The novel mode of action of pathogen mycelium degradation was elucidated by the minimum inhibitory concentration (MIC) of green nanosilver as 40 µg Ag ml-1 to diminish F. oxysporum (SEM morphology). The green nanosilver at 2 DAI renowned the leakage of sugars from mycelia of the cell membrane and defeated the activity of respiratory chain dehydrogenases, followed by lipid peroxidation and the highest leakage of proteins at 3 DAI on MIC. The in-vivo study might allow for novel insight to utilize green nanosilver at MIC (40 µg Ag ml-1) as an eco-friendly and fungicide alternate way for antifungal action to demolish Fusarium wilt infection under harsh conditions.

Characterization and bioefficacy of green nanosilver particles derived from fungicide-tolerant Tricho-fusant for efficient biocontrol of stem rot (Sclerotium rolfsii Sacc.) in groundnut (Arachis hypogaea L.).

An efficient and eco-friendly bioefficacy of potent Tricho-fusant (Fu21) and its green nanosilver formulation against stem rot (Sclerotium rolfsii) in groundnut was established. Fu21 demonstrated higher in-vitro growth inhibition of pathogen with better fungicide tolerance than the parental strains. The green nanosilver particles were synthesized from the extracellular metabolites of Fu21 and characterized for shape (spherical, 59.34 nm in scanning electron microscope), purity (3.00 KeV, energy dispersive X-ray analysis), size (54.3 nm in particle size analyzer), and stability (53.7 mv, zeta). The field efficacy study exhibited that the seedling emergence was high in seeds treated with green nanosilver (minimum inhibitory concentration-[MIC] 20 µg Ag/ml), and a low disease severity index of stem rot during the crop growth was followed by the live antagonist (Fu21) in addition to seed treatment with a fungicide mix under pathogen infestation. The seed quality analysis of harvested pods revealed a high oil content with balanced fatty acid composition (3.10 oleic/linoleic acid ratio) in green nanosilver treatment under pathogen infestation. The residual analysis suggested that green nanosilver applied at the MIC level as seed treatment yielded similar effects as the control for silver residue in the harvested groundnut seeds. The green nanosilver at MIC has a high pod-yield under S. rolfsii infestation, demonstrating green chemistry and sustainability of the nanoproduct.

Metabolomics of extracellular compounds and parasitic enzymes of Beauveria bassiana associated with biological control of whiteflies (Bemisia tabaci).

The Beauveria spp. were isolated from soil and insect cadavers of crop rhizosphere and characterized for parasitic enzyme activity and virulence against whiteflies (Bemisia tabaci). The colony morphology and molecular identification using ITS specific marker were carried out and confirmed entomopathogenic fungi as Beauveria bassiana. The bioefficacy of B. bassiana against whiteflies demonstrated highest corrected mortality and lowest LC50 in isolate B. bassiana JAU2 (SEM morphology) followed by JAU1 on 6th days. Parasitic enzymes chitinase and lipase were determined highest in JAU2 and protease activity examined higher in isolate JAU4 followed by JAU2 isolate on 6th days after inoculation. Comparative extracellular metabolomics carried out from potent (JAU1 and JAU2), moderate (JAU4 and JAU14) and weak (JAU6) B. bassiana isolates in normal suborder dextrose agar with yeast extrect (SDAY) and chitin induced media. Results illustrated that total 105 metabolites identified common for all five B. bassiana isolates differing in virulence. However, the color intensity of the metabolites changes in heat map showing differential concentration of that extracellular compound compared to other isolates. The volcano plot analysis illustrated 58 compounds significanlty diverse between potent JAU1 and JAU2 under two different culture conditions of which 34 compounds recognized up regulated in most potent JAU2 under chitin induced media. Out of 34 metabolites, ten compounds viz., fumaricine, resazurin, N-methyldioctylamine, penaresidun B, tetralin, squamocin B, oligomycin C, pubesenolide, epirbuterol and gentamicin C1a were recognized significantly upregulated in most potent JAU2 and reported for antimicrobial, nematicidal, larvicidalor insecticidal activities. The mass spectra and fragment structure were elucidated under LCMS-QTOF for some novel and unique compounds recognized in most potent B. bassiana JAU2, involved in parasitic activity against whiteflies.