Solid-state fermentation of tropical agro-industrial wastes using Pleurotus ostreatus for fungal protein production

Solid-state fermentation (SSF) of lignocellulosic agro-industrial wastes using white-rot fungi represents a scalable, low-energy strategy for converting underutilised tropical biomass into high-quality fungal protein. This study optimised six substrate formulations combining cocoa pod husk (CPH), hardwood sawdust, and composted vegetable waste for Pleurotus ostreatus NIHORT EM-1 (ITS rDNA authenticated; GenBank MN165852; 99.8% identity to CBS 244.31) under Nigerian tropical conditions. The strain is deposited at the National Horticultural Research Institute (NIHORT), Ibadan, Nigeria, and is publicly available for research purposes upon written request to the Director of Research, NIHORT, P.M.B. 5432, Idi-Ishin, Ibadan, Nigeria (email: info@nihort.gov.ng; Tel: +234-803-000-0001; online germplasm catalogue: www.nihort.gov.ng/germplasm ). Substrate carbon-to-nitrogen (C:N) ratio, mycelial colonisation kinetics, fresh-weight yield, proximate nutritional quality (protein digestibility-corrected amino acid score, PDCAAS), and contamination incidence were evaluated across a completely randomised design (n = 5 replicates per treatment; 30 bags total). The optimal formulation [T5: 58% sawdust, 25% CPH, 10% composted vegetable waste, 5% rice bran; C:N 31.2:1] achieved biological efficiency (BE) of 116.9%, protein yield of 359 g kg⁻¹ dry substrate (PDCAAS = 0.87; in vitro digestibility 89.2%), and full mycelial colonisation in 18.4 ± 1.0 days - improvements of 57%, 126%, and 21%, respectively, over the sawdust-only control. T5 fruiting bodies contained 30.7 ± 1.8% crude protein (dry weight), with eight of nine essential amino acids exceeding WHO/FAO (2007) adult reference requirements. ISO 14040-compliant life cycle assessment confirmed 98-99% reductions in global warming potential, water use, and land use relative to beef protein. Enterprise modelling demonstrated a return on investment of 321% per 100 kg production cycle (Monte Carlo-confirmed; n = 10,000 iterations). Multi-site pilot validation across three contrasting Nigerian agro-ecological zones yielded 95.7% mean yield retention (ANOVA: F(2,27) = 2.14, p = 0.136), confirming protocol transferability. These results establish CPH-based SSF as a globally competitive, low-carbon fungal protein platform with direct relevance to agro-waste valorisation and food security in tropical West Africa.