Fungal Endophytes: Microfactories of Novel Bioactive Compounds with Therapeutic Interventions; A Comprehensive Review on the Biotechnological Developments in the Field of Fungal Endophytic Biology over the Last Decade.

The seminal discovery of paclitaxel from endophytic fungus Taxomyces andreanae was a milestone in recognizing the immense potential of endophytic fungi as prolific producers of bioactive secondary metabolites of use in medicine, agriculture, and food industries. Following the discovery of paclitaxel, the research community has intensified efforts to harness endophytic fungi as putative producers of lead molecules with anticancer, anti-inflammatory, antimicrobial, antioxidant, cardio-protective, and immunomodulatory properties. Endophytic fungi have been a valuable source of bioactive compounds over the last three decades. Compounds such as taxol, podophyllotoxin, huperzine, camptothecin, and resveratrol have been effectively isolated and characterized after extraction from endophytic fungi. These findings have expanded the applications of endophytic fungi in medicine and related fields. In the present review, we systematically compile and analyze several important compounds derived from endophytic fungi, encompassing the period from 2011 to 2022. Our systematic approach focuses on elucidating the origins of endophytic fungi, exploring the structural diversity and biological activities exhibited by these compounds, and giving special emphasis to the pharmacological activities and mechanism of action of certain compounds. We highlight the tremendous potential of endophytic fungi as alternate sources of bioactive metabolites, with implications for combating major global diseases. This underscores the significant role that fungi can play in the discovery and development of novel therapeutic agents that address the challenges posed by prevalent diseases worldwide.

Efficient production and characterization of melanin from Thermothelomyces hinnuleus SP1, isolated from the coal mines of Chhattisgarh, India.

In the present study, fungi were isolated and screened from barren land in south-eastern Coalfields limited (SECL) in Chhattisgarh, India. Out of 14 isolated fungi, only three fungal isolates exhibited pigmentation in screening studies. The isolated fungal strain SP1 exhibited the highest pigmentation, which was further utilized for in vivo production, purification, and characterization of melanin pigment. The physical and chemical properties of the fungal pigment showed insolubility in organic solvents and water, solubility in alkali, precipitation in acid, and decolorization with oxidizing agents. The physiochemical characterization and analytical studies of the extracted pigment using ultraviolet-visible spectroscopy and Fourier transform infrared (FTIR) confirmed it as a melanin pigment. The melanin-producing fungus SP1 was identified as Thermothelomyces hinnuleus based on 18S-rRNA sequence analysis. Furthermore, to enhance melanin production, a response surface methodology (RSM) was employed, specifically utilizing the central composite design (CCD). This approach focused on selecting efficient growth as well as progressive yield parameters such as optimal temperature (34.4°C), pH (5.0), and trace element concentration (56.24 mg). By implementing the suggested optimal conditions, the production rate of melanin increased by 62%, resulting in a yield of 28.3 mg/100 mL, which is comparatively higher than the actual yield (17.48 ± 2.19 mg/100 mL). Thus, T. hinnuleus SP1 holds great promise as a newly isolated fungal strain that could be used for the industrial production of melanin.

Characterization of Colletotrichum Isolates from Strawberry and Other Hosts with Reference to Cross-Inoculation Potential.

Colletotrichum is an important phytopathogenic fungus that causes anthracnose disease in diverse agronomically important tropical food crops. Accurate pathogen identification is critical for early diagnosis and efficient management of anthracnose. ITS is not a reliable marker for this fungal genus due to its failure to phylogenetically resolve cryptic species. In this study, 36 Colletotrichum isolates belonging to the Acutatum, Boninense and Gloeosporioides species complexes were characterized using multigene phylogenetic analyses, morphology and pathogenicity assays. Additionally, the cross-inoculation potential of a representative subset of isolates was evaluated revealing that cross-infection potential is possible among the isolates belonging to the same species complex.

Symbiosis and pathogenicity of Geosmithia and Talaromyces spp. associated with the cypress bark beetles Phloeosinus spp. and their parasitoids.

Fungi associated with cypress bark beetles are practically unknown in the Eastern Mediterranean. Our study focused on the fungi associated with the body parts and galleries of two indigenous cypress bark beetles, Phloeosinus armatus and P. bicolor, sampled from Cupressus sempervirens trees in different regions in Israel. Arbitrarily primed PCR, performed on genomic DNA of 302 isolates, clustered the fungal population into five distinct groups. Multilocus phylogeny, split-network analyses and morphological characterization identified the isolates as Geosmithia omnicola, Geosmithia langdonii, Geosmithia sp. 708b, Geosmithia cupressina sp. nov. CBS147103 and Talaromyces cupressi sp. nov. CBS147104. Of these fungal isolates, G. cupressina and T. cupressi are newly described, and their morphological features and phylogenetic designations are presented. Inoculation of intact cypress saplings in an outdoor net-house revealed that only the representative isolate T. cupressi sp. nov. CBS147104 causes 100% disease incidence, whereas Geosmithia spp. isolates are not pathogenic. A number of these fungi were isolated from parasitoids that emerged from branch and stem sections colonized by P. armatus. This study suggests a long and stable association between Phloeosinus and Geosmithia species, and a possible role for additional associated fungal species as pathogens or endophytes of C. sempervirens trees in Israel.

Production, Purification and Characterisation of a Potential Fibrinolytic Protease from Endophytic Xylaria curta by Solid Substrate Fermentation.

The present investigation highlights the optimal conditions for production of a non-toxic, bi-functional fibrinolytic enzyme xylarinase produced by endophytic fungus Xylaria curta by solid substrate fermentation using rice chaff medium. The purified enzyme is a monomeric protein with a molecular mass of ∼33 kDa. The enzyme exhibits cleavage of Aα and Bß chains of fibrin(ogen) and has no effect on γ chain. The optimal fibrinolytic activity of the enzyme was observed at 35 °C and pH 8. The fibrinolytic activity was enhanced in the presence of Ca2+, whereas it was completely inhibited in the presence of Fe2+ and Zn2+ ions and inhibitors like EDTA and EGTA suggesting it to be a metalloprotease. The K m and V max of the enzyme for azocasein were 326 µM and 0.13 µM min-1. The N-terminal sequence of the enzyme (SNGPLPGGVVWAG) was same when compared to xylarinase isolated from culture broth of X. curta. Thus, xylarinase could be exploited as a potent clot busting enzyme which could be produced on large scale using solid substrate fermentation.

Mycoherbicidal Potential of Phaeoacremonium italicum, A New Pathogen of Eichhornia crassipes Infesting Harike Wetland, India.

Mycoherbicides are exclusive biotechnology products which offer a non-chemical solution to control noxious weeds on the land as well as aquatic in systems, viz a viz saving environment from hazardous impact of synthetic chemicals. The present paper highlights the mycobiota associated with Eichhornia crassipes infesting Harike wetland area of Punjab and evaluation of their pathogenic potential for futuristic application as a mycoherbicide. Of the 20 isolates tested by leaf detached assay and whole plant bioassays, only one isolate (#8 BJSSL) caused 100% damage to E. crassipes. Further, the culture filtrate of this isolate also exhibited a similar damage to the leaves in an in vitro detached leaf assay. The potential isolate was identified as Phaeoacremonium italicum using classical and modern molecular methods. This is the first report of P. italicum as a pathogen of E. crassipes and of its potential use as a biological control agent for the management of water hyacinth.

Xylarinase: a novel clot busting enzyme from an endophytic fungus Xylaria curta.

Xylarinase is a bi-functional fibrinolytic metalloprotease isolated from the culture filtrate of endophytic fungus Xylaria curta which is monomeric with a molecular mass of ∼33.76 kDa. The enzyme displayed both plasmin and tissue plasminogen activator like activity under in vitro conditions. It hydrolyses Aα and Bß chains of the fibrinogen. Optimal fibrinolytic activity of xylarinase is observed at 35 °C, pH 8. Ca(2+) stimulated the fibrinolytic activity of xylarinase while Fe(2+) and Zn(2+) inhibited suggesting it to be a metalloprotease. The Km and Vmax values of xylarinase were 240.9 µM and 1.10 U/ml for fibrinogen and 246 µM and 1.22 U/ml for fibrin, respectively. Xylarinase was found to prolong the activated partial thromboplastin time and prothrombin time. The N-terminal sequence of xylarinase (SNGPLPGGVVWAG) did not show any homology with previously known fibrinolytic enzymes. Thus xylarinase is a novel fibrinolytic metalloprotease which could be possibly used as a new clot busting enzyme.

Potential fibrinolytic activity of an endophytic Lasiodiplodia pseudotheobromae species.

Cost-effective, fibrinolytic agents possessing least or no antigenic properties are much in demand for their prospective use as clot busters in thrombolytic therapy. The present communication explores the potential of 22 endophytic Botryosphaeria species isolated from stem of Aegle marmelos for their fibrinolytic potential. Only nine fungal isolates exhibited proteolytic activity out of which only four possessed plasminogen-independent fibrinolytic activity. The endophytic fungal isolate #1088 AMSTITYEL showed maximum in vitro proteolytic and fibrinolytic activity with a halo formation of 153.86 and 113.04 mm2, respectively. The partially purified protein exhibited a fibrinolytic activity equivalent to 6.51 U/ml of plasmin. SDS-PAGE of the dialysed fraction of #1088 AMSTITYEL resolved into six bands ranging from 26 to 80 kDa. Fibrin zymogram exhibited that a single band of molecular size ~80 KDa possessing the fibrinolytic activity. Furthermore, the bioactive isolate #1088 AMSTITYEL was identified as Lasiodiplodia pseudotheobromae using classical and molecular taxonomic tools.

Muscodor kashayum sp. nov. - a new volatile anti-microbial producing endophytic fungus.

Muscodor kashayum (MycoBank no.: MB 803800; GenBank no.: KC481680) is a newly described endophytic fungus of a medicinal plant Aegle marmelos (Bael tree), growing in the tropical conserved rainforest in the Western Ghats of India. Muscodor kashayum possesses distinct morphological, molecular and physiological features from the earlier reported Muscodor species. The fungus forms characteristic rings of the ropy mycelium on potato dextrose agar medium. This sterile fungus is characterised by the presence of a pungent smell which is attributable to a blend of more than 23 volatile organic constituents, predominantly 3-cyclohexen-1-ol,1-(1,5-dimethyl-4-hexenyl)-4-methyl; 1,6-dioxacyclododecane-7,12-dione; 2,6-bis(1,1-dimethylethyl)-4-(1-oxopropyl) phenol; 2,4-di-tert-butylthiophenol and 4-octadecylmorpholine. In the in vitro anti-microbial assay using M. kashayum, growth of 75% of test fungi/yeasts and 72% of the test bacteria were completely inhibited. Therefore, M. Kashayum holds potential for future application to be used as a myco-fumigation agent.