In vitro Antimicrobial activity and Qualitative Enzymatic assays of Cladosporium pseudocladosporidiales, AVNK4, an endophytic fungus isolated from seeds of Nigella sativa

Endophytes are vital reservoirs of bioactive compounds with pharmaceutical and industrial significance. This study reports isolating and characterising a novel fungal endophyte, AVNK4, from surface-sterilised Nigella sativa seeds, identified as Cladosporium pseudocladosporidiales using morphological observations and ITS-rDNA sequencing. AVNK4 is the first reported isolate of Cladosporium pseudocladosporidiales from Nigella sativa seeds, showing both diverse lytic enzyme production and broad-spectrum antimicrobial activity against human and plant pathogens. Ethyl acetate extracts exhibited strong inhibition zones and low minimal inhibitory concentrations, indicating potent bioactivity. Enzymatic assays revealed substantial production of cellulase, amylase and protease, which enhance the endophyte’s bioactive potential. These hydrolytic enzymes enhance the endophyte’s bioactive potential by degrading cell walls and assisting in metabolite delivery, synergising with antimicrobial compounds. Furthermore, some enzymes are directly involved in secondary metabolite biosynthesis such as non-ribosomal peptide synthetases and polyketide synthases, which are responsible for the production of antibiotics and antifungals. The dual functionality of AVNK4, as a producer of both antimicrobial metabolites and industrially relevant enzymes, underscores its promise for biotechnological applications in pharmaceuticals, agriculture and enzyme-based industries as novel drugs and biocatalysts. This study lays the foundation for further metabolite purification and mechanistic investigations to support drug development and industrial enzyme deployment.