Extracellular Matrix of Aspergillus Terreus HLP5 Biofilm Regulates the Stress Tolerance Mechanism Against Pb2+-Phenanthrene Co-Pollutants

Abstract

Filamentous fungi develop robust biofilm embedded in extracellular matrix (ECM), contributing to their adaptability and resilience against toxic pollutants. The present study explores the biofilm-forming filamentous fungus Aspergillus terreus HLP5 isolated from Bhitarkanika mangroves for its ability to tolerate lead (Pb2+)- phenanthrene co-pollutants. The variation in biofilm formation and interaction of ECM of A. terreus HLP5 with Pb2+-phenanthrene ranging from 25:25 ppm to 500:500 ppm concentration was investigated. Crystal-violet quantification indicated a shift in biofilm maturation at 48 h in control to 72 h in the presence of co-pollutants. The absorbance maximum (Am) was significantly reduced beyond 25:25 ppm concentration (P < 0.0001; twoway ANOVA), whereas the maximum growth rate (Kmax ) remained relatively constant. Confocal micrographs of the biofilm maturation revealed a total biofilm biomass of 21±2.5 μm³/μm² and a maximum thickness of 51.90±2.6 μm in the control condition, with a significant increase in total biofilm biomass and maximum thickness in the presence of co-pollutants (P < 0.0001; two-way ANOVA). SEM analysis depicted mycelial thickening and dense ECM deposition up to 200:200 ppm of co-pollutant concentration. Polysaccharide and protein levels significantly increased with co-pollutant concentration, supporting biofilm growth (P < 0.0001; two-way ANOVA). During ATR-FTIR analysis, the peak stretching and occurrence of additional peaks in the amide I and II regions indicated co-pollutant interaction with the protein functional groups. Fluorescence spectroscopy revealed the quenching of tryptophan-like protein with co-pollutant interaction. The present study suggests the significant role of fungal biofilm-ECM in mitigating the impact of co-pollutants and establishing fungal survival mechanisms.