Exploration of Mangrove Endophytes as Novel Sources of Tannase Producing Fungi

Abstract

Tannase, a highly adaptive biocatalyst, plays a pivotal role in diverse bioconversion reactions in nature. This enzyme exhibits numerous applications across various industrial sectors, including food, pharmaceuticals, chemicals, and beverages. This study aimed to screen and characterize fungal endophytes isolated from mangrove plants for their enzyme tannase-producing ability. Eighty-five filamentous endophytic fungi were isolated from different mangrove samples and subsequently identified. These fungal strains were initially screened using the tannic acid agar plate method. Out of the screened strains, 13 fungal isolates demonstrated tannase production ability. The quantitative estimation of extracellular tannase was performed using the submerged fermentation technique. Among the studied endophytes, eight isolates, namely LV_084 (21.21 IU/mL), LV_074 (15.41 IU/mL), LV_078 (6.98 IU/mL), LV_038 (6.97 IU/mL), LV_077 (6.32 IU/mL), LV_016 and LV_066 (6.37 IU/mL), and LV_060 (6.18 IU/mL) exhibited excellent tannase activity. Among these isolates, LV_084 Phyllosticta capitalensis and LV_074 Aspergillus chevalieri showed the highest enzyme-producing ability. These isolates were authenticated using ITS rDNA sequencing, followed by BLAST search and phylogenetic analysis. Furthermore, the physical and chemical conditions for the maximum enzyme production were optimized. This is the first report of enzyme tannase production by Phyllosticta capitalensis and Aspergillus chevalieri.

Keywords: endophytic fungi; mangrove; screening; tannase.

Figure 9

Aspergillus chevalieri: (a) growth on TAA showing a dark zone around the colony; (b) colony on PDA verse; (c) PDA reverse; (d) a conidiophore with a vesicle; (e) a vesicle with sterigmata; (f) conidia; (g) ascomata; and (h) ascospores.

Figure 10

Phylogram of Aspergillus resulting from the maximum likelihood (RAxML) tree using ITS sequences. Confidence values for ML ≥50% (UFboot2/RAxML) are included near the nodes. The specimens described in this study are highlighted in bold. Aspergillus xerophilus NRRL6131, Aspergillus xerophilus NRRL6132, and Aspergillus osmophilus IRAN2090C were used as an outgroup.

Figure 1

The map represents the mangrove ecosystem in different states of India . The green zone in the box represents Mumbai coastal region. Pink coloured spots represent sample collection sites in the Godrej Mangrove Forest at Vikhroli.

Figure 2

Diversity of fungal endophytes in A. marina, A. officinalis, Acanthus ilicifolius, S. maritima, D. indica, D. trifoliata, R. mucronate, and Salvadora persica.

Figure 3

Fungi showing tannic acid degradation leading to the formation of a hydrolysis zone after 72 h: Penicillium sp. (LV_001); Chaetomium sp. (LV_002); Chaetomium sp. (LV_004); Aspergillus sp. (LV_010); Aspergillus sp. (LV_016); Curvularia brachyspora (LV_019); Alternaria tenuissima (LV_022); Chaetomium sp. (LV_038); Corynespora cassiicola (LV_042); Cladosporium limoniforme (LV_047); Curvularia lunata (LV_053); and control.

Figure 4

Quantitative estimation of the tannase enzyme activity of fungal isolates under submerged fermentation.

Figure 5

Optimization of the incubation period (48 h, 72 h, 96 h, and 120 h) for the maximum tannase activity of LV_074 and LV_084.

Figure 6

Optimization of the optimum temperature for the maximum tannase activity of LV_074 and LV_084.

Figure 7

Optimization of the pH level of the growth medium for the maximum tannase activity in LV_074 and LV_084.

Figure 8

Optimization of the incubation period for the maximum biomass production of isolates LV_074 and LV_084.

Figure 11

Phyllosticta capitalensis Henn.: (a) growth on tannic acid agar showing a dark zone around the colony; (b) colony on PDA verse; (c) PDA reverse; (d) mycelium; (e) an ascus; (f,g) asci and ascospores.

Figure 12

Phylogram of Phyllosticta resulting from the maximum likelihood (RAxML) tree using ITS sequences. Confidence values for ML ≥50% (UFboot2/RAxML) are included near the nodes. The specimens described in this study are highlighted in bold. B. obtusa (CMW 8232) and B. stevensii (CBS112553) were used as an outgroup.