Chitin contents of different organisms Fungi 5-60% Worms 20-38% Squid 3-20% Scorpions 30% Spiders 38% Cockroaches 35% Silkworms 44% Crabs 70% Fungal source No seasonal variation Demineralization is not required during isolation Homogenous preparation Free of heavy metals as nickel, copper Low molecular weight and low viscosity High degree of deacetylation High polydispersity Useful to make tissue engineering scaffolds Biocompatible Have good mechanical, water absorption and lysozyme degradation properties Not heavily melanised Fungi are second largest group of organisms on earth Estimated number - 15,00,000 Known species - 82,000 Zygomycetes are 1,056 (fungi containing more chitosan in the cell wall) Non-zygomycetous plant and insect-pathogenic fungi have also chitosan in the cell wall for self defence from host chitinases

Three types of fungal sources Use of waste mycelium from Mycotech industries Fermentation of fungus for chitin/chitosan production Fermentation of fungus for the production of chitin/chitosan and value addition

Mycotech industries producing waste biomass Brewing and baking Sacharomyces cerevisiae, Saccharomyces carlbergensis Mushroom cultivation Agaricus bisporus, Auricularia sp., Flammulina velutipes, Lentinus edodes, Pleurotus sp., Volvariella volvacea Antibiotics and other pharmaceuticals (Penicillin, cephalosporin, cyclosporin, ergot alkaloids, griseofulvin mevalonin) Aspergillus terreus , Cephalosporium acremonium,, Claviceps pupurea, Penicillium chrysogenum, Penicillium griseofulvum, Tolypocladium inflatum and others Organic acids (Citric acid, itaconic acid, lactic acid) Aspergillus niger, Aspergillus terreus, Mucor rouxii , Rhizopus oryzae Enzymes (Amylase, cellulase, glucanase, glucoamylase, glucose oxidase, hemicellulase, laccase, lipase, pectinase, protease, rennet, tannase, xylanase) Aspergillus niger, Aspergillus oryzae, Endothia parasitica , Mucor miehei, Penicillium emersonii , Penicillium funiculosum, Pyricularia oryzae, Rhizopus oryzae, Trichoderma reesei, Trichoderma viride to name a few From citric acid production >80,000 tons of waste Aspergillus niger biomass per year is generated In Australian wineries, > 800 tons yeast biomass is generated per year Penicillin production > 80 % of the total yield of antibiotics. 1.2 million tons of bio waste, mainly Penicillium mycelial biomass per year Mushroom waste- mainly stalks and mushrooms of irregular dimensions and shape. Approximately 50,000 metric tons of waste material per year ‎

Fungi studied for chitosan production Chitosan % DDA g/kg biomass Zygomycetes 29-110 75-94 Absidia coerulea 110 90 Absidia glauca 59 76 Benjaminiella poitrasii 51 94 Cunninghamella bertholletiae 55 88 Gongrenella butleri 40-60 87 Mucour rouxii 60-77 83 Rhizopus arrhizus 29 86 Rhizopus oryzae 58 89

Agricultural industry Use of fungi and fungal products for the control of insect pests and pathogens ( NCL work) Use of conidia as mycoinsecticide After solid state fermentation 70 g conidia and 5 g mycelial biomass are produced; Metarhizium anisopliae , chitosan, 18-23 g/kg biomass; %DDA ,82 M. anisopliae M34412: 67 g conidia / kg rice (4.4 x 1010 conidia/g)

Commercial chitosan 1M NaOH treatment CDA treatment Another M. anisopliae strain produces extracellular CDA in submerged fermentation Deacetylation of chitosan from marine sources using M. anisopliae CDA Fungal biomass can be used for chitosan extraction CDA treatment 1M NaOH treatment Commercial chitosan DDA-75% DDA-80.8% DDA-83.3% 1H-NMR spectra Use of extracellular CDE/ME complex for biocontrol After submerged fermentation 100 g enzyme complex and 100 g biomass Myrothecium verrucaria , chitosan, 27 g/kg biomass; %DDA, 87

Advanced strain improvement method: Metabolic Engineering Utilize metabolic engineering technologies to develop fungal strains with improved production capability for lactic acid by Rhizopus (Chitosan- enriched aqueous medium) Metabolic engineering of the morphology of Aspergillus oryzae by altering chitin synthesis Mycelial morphology affects the rheological properties of the cultivation medium Metabolic engineering to produce higher biomass with higher chitosan contents

Epilouge Finally priority is important- Quantity in tons,kgs, or gram Quality industrial/food /pharma grade Ease of isolation protocol Eco-friendly approach Cost