# Unveiling Biological Activities of Marine Fungi: The Effect of Sea Salt
## 4 Discussion
Chitinases can be used as antifungal agents and biopesticides [61]. They are also useful in the manufacture of pharmaceutical products, as tools in cell engineering, and in medical diagnosis [62]. Some marine fungi capable of synthesizing chitinases are already known, such as species belonging to _Aspergillus_, _Rhizopus_, _Penicillium_, _Beuveria_[9], and _Clonostachys rosea_ isolated from seagrass [63]. Another relevant group of enzymes for the lignin degradative process with bleaching applications in the wood, paper, and cellulose industries are lacases [52], which are afamily of multicopper oxidases distributed across bacteria, fungi, and plants [64]. These enzymes are constitutively secreted and are stable in the extracellular environment contributing to their attractiveness for biotechnological applications [65]. Several marine fungal strains isolated from seagrass, algae, and decaying wood have been reported to produce lacases, such as _Penicillium_ spp., _Trichoderma_ spp., _Lulworthia_ spp., _Z. maritima_, _Aspergillus scleroorum_, _Cladosporium cladosporoides_, and _Muvor racemosus_[52; 66; 67]. In this study, only _T. aestariumum_ and _V. ambiguum_ expressed extracellular lacases. Proteases also have a relevant role in the biotechnology industry, including the pharmaceutical industry. As reviewed earlier [68], proteases, namely collagenases, have important applications in the health industry, such as the use of collagenases in the enzymatic debridement of wounds and of other injuries where the removal of devitalized tissue is necessary. Some studies have explored the influence of salt in the enzymatic activity of marine fungi. For example, [PERSON] et al. [69] evaluated different enzymatic activities of halotolerant and halophilic fungi and [PERSON] et al. [70] reported the presence of more than 400 lignocellulosic enzymes from the mangrove fungus _Pestalotiopsis_ sp. The authors concluded that the presence of salt modifies lignocellulosic enzyme composition, with an increase in the secretion of xylanases and cellulases and a decrease in the production of oxidases, as we also observed in this study. Additionally, [PERSON] et al. [71] and [PERSON] et al. [72] achieved similar results in lignin enzymes when fungi were grown in saline conditions. In this study, sea salt induced the alteration of the enzymatic profiles detected. Eight strains were selected for a deeper characterization of their biotechnological potential. These strains belong to genera widely known to produce active extracellular metabolites with antibacterial activity, such as _Penicillium_, _Aspergillus_, _Trichoderma_, and _Cladosporium_[73], or even _Emericellopsis_[74]. In the present work, 80% of the 25 strains tested in the preliminary screening were able to inhibit the growth of at least one bacterium, mainly Gram-positive bacteria, during the primary screening. However, only four strains (_A. affinis_, _E. cladophrae_, _Pen. Lusitanum_, and _T. aestariumum_) of the eight fungi selected for the fermentation assay retained their antibacterial activity. Similar observations were reported in other studies, in which fungi showed activity in solid media but not when subjected to fermentation. This apparent loss of activity may be due to the type of medium, culture conditions, or to the extraction process [75; 76; 77]. In general, crude extracts of extracellular media showed higher activity than mycelia extracts. _Aspergillus affinis_ led to the largest inhibition halos, inhibiting the growth of the three-Gran negative bacteria, including the multiresistant strains _K. pneumoniae_ and _P. aeruginosa_. These two multiresistant strains were isolated from the Lis River (Central Portugal) [37]. These authors highlighted that the presence of antibiotic-resistant bacteria and resistance genes in this particular environment is a major public health risk.