## 4 Discussion  
Protcolytic and lipolytic microorganisms are constantly being researched for their possible application in several sectors, such as environmental remediation, detergent manufactory, biomedicine, the food industry and, recently, for food waste recovery and valorization. In this work, yeasts and bacteria, known for their proteolytic and lipolytic properties, were screened in vitro using common substrates, namely skim milk, Spirit Blue Agar and gelatin. Among the microorganisms tested, the best performing ones were _B. subtilis_ B5C, _B. amyloliguefaciens_ B5M and two strains of _Y. lipolytica_, YL2 and YL4. Regarding bacterial strains, the genus _Bacillus_ contains several species with documented proteolytic and lipolytic activities [33; 34]. For instance, _Bacillus subtilis_ and _B. amyloliguefaciens_, both Gram-positive bacteria with a generally recognized safe (GRAS) status, can break down many types of carbohydrates, proteins, and lipids due to their enzymes [35; 36]. Therefore, it is not surprising that the _Bacillus_ species performed well in the different assays (skim milk, gelatin and Spirit Blue Agar). _Y. lipolytica_ is one of the most extensively studied yeast species, after _S. cerevisiae_, which contributes to food ripening thanks to its proteolytic and lipolytic properties [37; 38; 39]. _Y. lipolytica_ possesses the \"qualified presumption of safety\" (QPS) and GRAS status for production purpose and it has been defined as \"novel food\" by the European Food Safety authority (EFSA) [40]. Due to its properties, this yeast has been proposed as a promising tool to recover and valorize food wastes and by-products [22]. Concerning proteolytic activity, strain YL4 possessed the strongest, followed by Bacilli and YL2. The values obtained were higher compared to those reported by [PERSON] et al. [14], but in line with other studies [41]. It is possible that the lower activity described by [PERSON] et al. [14] also depends on the very poor substrate (marine broth) applied to grow the strain tested. At the same time, higher proteolytic activities were reported when optimal conditions were selected [42]. The 4 proteolytic and lipolytic microorganisms were then tested in a fish waste solution as substrates. As already performed by [PERSON] et al. [43], samples were incubated for 72 h in order to observe differences in hydrolysis. All the tested strains grew in the fish waste solution with B5M, B5C, and YL4 increasing by around 2.2 log CFU/mL, while YL2 increased by 3.1 log CFU/mL. The pH increased during the first 48 h with _Y. lipolytica_, while it remained stable with the two species of _Bacillus_, as well as the control. After 72 h, instead, the pH decreased in all samples due to the growth of indigenous lactic acid bacteria present in the matrix that reached a final concentration of 5.8 log CFU/mL. Looking at the degrees of hydrolysis, the two strains of _Y. lipolytica_ were the best-performing ones. The values obtained are in line with the data reported by [PERSON] et al. [43] when brown crab processing side streams were fermented with _Pseudoalteromonas_ strains after 72 h. In our case, the two yeasts strains performed better than the two bacteria strains. The use of _Y. lipolytica_ to produce protein hydrolysates for fish waste has never been reported. In fact, most of the literature is focused on the production of single cell proteins, single cell oils, citric acid, polyols and enzymes from food waste [22], or single cell oils and lipid reduction from fish waste [6; 23]. Alternatively, protein hydrolysates have been used as substrates for _Y. lipolytica_ to produce oils [44; 45]. Only [PERSON] et al. [46] reported the use of _Y. lipolytica_ RO25 to obtain cricket powder hydrolysate richer in health-promoting molecules, while [PERSON] et al. [47] described the use of purified _Y. lipolytica_ proteases to reduce the antigenic response to bovine milk allergens. Regarding the functionality of the obtained hydrolysates, those of _Y.