Alternaria is an important fungus to study due to their different life style from saprophytes to endophytes and a very successful fungal pathogen that causes diseases to a number of economically important crops. Alternaria species have been well-characterized for the production of different host-specific toxins (HSTs) and non-host specific toxins (nHSTs) which depend upon their physiological and morphological stages. The pathogenicity of Alternaria species depends on host susceptibility or resistance as well as quantitative production of HSTs and nHSTs. These toxins are chemically low molecular weight secondary metabolites (SMs). The effects of toxins are mainly on different parts of cells like mitochondria, chloroplast, plasma membrane, Golgi complex, nucleus, etc. Alternaria species produce several nHSTs such as brefeldin A, tenuazonic acid, tentoxin, and zinniol. HSTs that act in very low concentrations affect only certain plant varieties or genotype and play a role in determining the host range of specificity of plant pathogens. The commonly known HSTs are AAL-, AK-, AM-, AF-, ACR-, and ACT-toxins which are named by their host specificity and these toxins are classified into different family groups. The HSTs are differentiated on the basis of bio-statistical and other molecular analyses. All these toxins have different mode of action, biochemical reactions and signaling mechanisms to cause diseases. Different species of Alternaria produced toxins which reveal its biochemical and genetic effects on itself as well as on its host cells tissues. The genes responsible for the production of HSTs are found on the conditionally dispensable chromosomes (CDCs) which have been well characterized. Different bio-statistical methods like basic local alignment search tool (BLAST) data analysis used for the annotation of gene prediction, pathogenicity-related genes may provide surprising knowledge in present and future.

In the present study, we have evaluated the comparative biochemical defense response generated against Alternaria alternata and its purified toxins viz. alternariol (AOH), alternariol monomethyl ether (AME), and tenuazonic acid (TeA). The necrotic lesions developed due to treatment with toxins were almost similar as those produced by the pathogen, indicating the crucial role of these toxins in plant pathogenesis. An oxidative burst reaction characterized by the rapid and transient production of a large amount of reactive oxygen species (ROS) occurs following the pathogen infection/toxin exposure. The maximum concentration of hydrogen peroxide (H2O2) produced was reported in the pathogen infected samples (22.2-fold) at 24 h post inoculation followed by TeA (18.2-fold), AOH (15.9-fold), and AME (14.1-fold) in treated tissues. 3,3′- Diaminobenzidine staining predicted the possible sites of H2O2 accumulation while the extent of cell death was measured by Evans blue dye. The extent of lipid peroxidation and malondialdehyde (MDA) content was higher (15.8-fold) at 48 h in the sample of inoculated leaves of the pathogen when compared to control. The cellular damages were observed as increased MDA content and reduced chlorophyll. The activities of antioxidative defense enzymes increased in both the pathogen infected as well as toxin treated samples. Superoxide dismutase (SOD) activity was 5.9-fold higher at 24 h post inoculation in leaves followed by TeA (5.0-fold), AOH (4.1-fold) and AME (2.3-fold) treated leaves than control. Catalase (CAT) activity was found to be increased upto 48 h post inoculation and maximum in the pathogen challenged samples followed by other toxins. The native PAGE results showed the variations in the intensities of isozyme (SOD and CAT) bands in the pathogen infected and toxin treated samples. Ascorbate peroxidase (APx) and glutathione reductase (GR) activities followed the similar trend to scavenge the excess H2O2. The reduction in CAT activities after 48 h post inoculation demonstrate that the biochemical defense programming shown by the host against the pathogen is not well efficient resulting in the compatible host-pathogen interaction. The elicitor (toxins) induced biochemical changes depends on the potential toxic effects (extent of ROS accumulation, amount of H2O2 produced). Thus, a fine tuning occurs for the defense related antioxidative enzymes against detoxification of key ROS molecules and effectively regulated in tomato plant against the pathogen infected/toxin treated oxidative stress. The study well demonstrates the acute pathological effects of A. alternata in tomato over its phytotoxic metabolites.

Fungal glucose oxidase (GOD) is widely employed in the different sectors of food industries for use in baking products, dry egg powder, beverages, and gluconic acid production. GOD also has several other novel applications in chemical, pharmaceutical, textile, and other biotechnological industries. The electrochemical suitability of GOD catalyzed reactions has enabled its successful use in bioelectronic devices, particularly biofuel cells, and biosensors. Other crucial aspects of GOD such as improved feeding efficiency in response to GOD supplemental diet, roles in antimicrobial activities, and enhancing pathogen defense response, thereby providing induced resistance in plants have also been reported. Moreover, the medical science, another emerging branch where GOD was recently reported to induce several apoptosis characteristics as well as cellular senescence by downregulating Klotho gene expression. These widespread applications of GOD have led to increased demand for more extensive research to improve its production, characterization, and enhanced stability to enable long term usages. Currently, GOD is mainly produced and purified from Aspergillus niger and Penicillium species, but the yield is relatively low and the purification process is troublesome. It is practical to build an excellent GOD-producing strain. Therefore, the present review describes innovative methods of enhancing fungal GOD production by using genetic and non-genetic approaches in-depth along with purification techniques. The review also highlights current research progress in the cost effective production of GOD, including key advances, potential applications and limitations. Therefore, there is an extensive need to commercialize these processes by developing and optimizing novel strategies for cost effective GOD production.

Plant defense against their pathogens can be induced by a complex network of different inducers. The present study investigates the synergistic effect of Trichoderma harzianum, exogenous salicylic acid (SA) and methyl jasmonate (MeJA) over the response and regulation of the antioxidant defense mechanisms and lipid peroxidation in tomato plants against Fusarium wilt disease. In the present work, tomato plants were infected by Fusarium oxysporum f. sp. lycopersici 3 days after inoculated with T. harzianum and/or sprayed daily for 3 days with chemical inducers (SA and MeJA). Plants were analysed at 0, 24, 48, 72 and 96 h after inoculation with Fusarium oxysporum f. sp. lycopersici. Infection of tomato plants by pathogen led to strong reduction in the dry weight of roots and shoots with the enhanced concentration of H2O2 and varying degree of lipid peroxidation. Concurrently, exogenous SA, when applied with pathogen greatly enhanced H2O2 content as well as activities of antioxidant enzymes except catalase (CAT) and ascorbate peroxidase (APx). The pathogen challenged plants pretreated with T. harzianum and MeJA together exhibited less lipid peroxidation and as well as the elevated level of ascorbic acid and enhanced activities of antioxidant enzymes. All applied treatments protected tomato seedlings against Fusarium wilt disease but the percentage of protection was found higher in plants pretreated with the combination of T. harzianum and chemical inducers.

Vascular wilt of tomato caused by Fusarium oxysporum f.sp. lycopersici (FOL) is one of the most devastating diseases, that delimits the tomato production worldwide. Fungal short-chain dehydrogenases/reductases (SDRs) are NADP(H) dependent oxidoreductases, having shared motifs and common functional mechanism, have been demonstrated as biochemical targets for commercial fungicides. The 1,3,6,8 tetra hydroxynaphthalene reductase (T4HNR) protein, a member of SDRs family, catalyzes the naphthol reduction reaction in fungal melanin biosynthesis. We retrieved an orthologous member of T4HNR, (complexed with NADP(H) and pyroquilon from Magnaporthe grisea) in the FOL (namely; FOXG_04696) based on homology search, percent identity and sequence similarity (93% query cover; 49% identity). The hypothetical protein FOXG_04696 (T4HNR like) had conserved T-G-X-X-X-G-X-G motif (cofactor binding site) at N-terminus, similar to M. grisea (1JA9) and Y-X-X-X-K motif, as a part of the active site, bearing homologies with two fungal keto reductases T4HNR (M. grisea) and 17-β-hydroxysteroid dehydrogenase from Curvularia lunata (teleomorph: Cochliobolus lunatus PDB ID: 3IS3). The catalytic tetrad of T4HNR was replaced with ASN115, SER141, TYR154, and LYS158 in the FOXG_04696. The structural alignment and superposition of FOXG_04696 over the template proteins (3IS3 and 1JA9) revealed minimum RMSD deviations of the C alpha atomic coordinates, and therefore, had structural conservation. The best protein model (FOXG_04696) was docked with 37 fungicides, to evaluate their binding affinities. The Glide XP and YASARA docked complexes showed discrepancies in results, for scoring and ranking the binding affinities of fungicides. The docked complexes were further refined and rescored from their docked poses through 50 ns long MD simulations, and binding free energies (ΔGbind) calculations, using MM/GBSA analysis, revealed Oxathiapiprolin and Famoxadone as better fungicides among the selected one. However, Famoxadone had better interaction of the docked residues, with best protein ligand contacts, minimum RMSD (high accuracy of the docking pose) and RMSF (structural integrity and conformational flexibility of docking) at the specified docking site. The Famoxadone was found to be acceptable based on in silico toxicity and in vitro growth inhibition assessment. We conclude that the FOXG_04696, could be employed as a novel candidate protein, for structure-based design, and screening of target fungicides against the FOL pathogen.