Cellulase and xylanase activity in relation to biofilm formation by two intertidal filamentous fungi in a novel polymethylmethacrylate conico-cylindrical flask

Abstract

A polymethylmethacrylate (PMMA) conico-cylindrical flask (CCF) with an inner arrangement consisting of eight equidistantly spaced rectangular strips mounted radially on a circular disk to provide additional surface area for fungal attachment was employed for production of cellulase by Chaetomium crispatum and xylanase by Gliocladium viride. The design allowed comparison of production between CCFs with hydrophobic surface (PMMA-CCF), hydrophilic glass surface (GS-CCF) and 500-ml Erlenmeyer flask (EF). Compared with the EF, endo-β-1,4-glucanase and FPase (filter paper degradation) activities increased from 0.044 to 0.156 and from 0.008 to 0.021 IU/ml, respectively, in the PMMA-CCF, while growth of C. crispatum was higher by at most 1.38-fold compared with the other vessels. Xylanase production in the EF was at most 5.08-fold higher and growth of G. viride was at most 1.52-fold higher compared with the other vessels. Temporal pattern of biofilm development based on two-channel fluorescence detection of extracellular polymeric substances (EPSs) and whole cells in a confocal laser scanning microscope demonstrated increase by 100% in biovolume, 25% in thickness and 62.5% both in substratum coverage and total spreading of C. crispatum biofilm in PMMA-CCF over 6 days. Biovolume of G. viride biofilm in GS-CCF increased by 150% over 4 days while that in PMMA-CCF enhanced by 200% over 2 days. Biofilm thickness in PMMA-CCF was 44% higher compared with GS-CCF and increased by 175% over 2 days. Substratum coverage was 38% higher in GS-CCF compared with PMMA-CCF. Thus, reactor surface area and property, shear forces and biofilm formation influenced enzyme production.

Enhanced Biotransformation of Fluoranthene by Intertidally Derived Cunninghamella elegans under Biofilm-Based and Niche-Mimicking Conditions

“…The surfaces of the inner arrangement of the CCFs were roughened, and roughness value (R a ) was determined with a profilometer. For sterilization, the CCF components were immersed in 2% (vol/vol) benzalkonium chloride for 5 h and then surface sterilized (7)(8)(9).…”

“…Biofilm architecture was determined by confocal laser scanning microscopy (CLSM) using slides as described previously (8,9). Two sets of three flasks each were utilized during cultivation on PMMA as well as glass surfaces.…”

“…Two sets of three flasks each were utilized during cultivation on PMMA as well as glass surfaces. Slides were aseptically taken out (two each) 1 day after addition of FA, i.e., the 4th, 5th, and 6th days of cultivation, and washed twice with phosphate-buffered saline (PBS) buffer (8,9). EPS staining with fluorescein isothiocyanate-concanavalin A (FITC-ConA; Sigma, USA) was done as explained elsewhere (8,9).…”

“…Slides were aseptically taken out (two each) 1 day after addition of FA, i.e., the 4th, 5th, and 6th days of cultivation, and washed twice with phosphate-buffered saline (PBS) buffer (8,9). EPS staining with fluorescein isothiocyanate-concanavalin A (FITC-ConA; Sigma, USA) was done as explained elsewhere (8,9). Thereafter, slides were incubated with SYTO 64 (Molecular Probes, USA) as elucidated previously (8,9).…”

“…Marine surfaceassociated microorganisms may require conditions that resemble their native environment in order to produce the maximum amount of bioactives; for example, studies have shown an increased production of antimicrobial compounds when the surface-associated bacteria were grown in vitro to form surface-attached biofilms (5,6). In our previous studies, we showed that melanin production by estuarine bacterial isolate Shewanella colwelliana; endo-␤-1,4-glucanase activity of another estuarine fungal isolate, Chaetomium crispatum; and riboflavin production by Candida famata, also an estuarine fungal isolate, were dependent upon biofilm formation (7)(8)(9).…”