*4.4. Extracellular Hydrolytic Enzymes*

The enzymatic activity of the studied *Actinomycetes* isolates was screened by carrying out an assay of extracellular hydrolytic enzymes on KB media supplemented with the below specific substrates for each enzyme: chitin azure (1%) or chitin from crab shells (1%) for chitinase [48]; skim milk (1%) for protease [48]; and lichenan (0.2%) for glucanase [49]. In addition, soluble starch (1%), pectin (0.5%), and polygalacturonic acid (1%) were used for amylase, pectinase, and polygalacturanase, respectively [50,51]. All plates were incubated at 30 ◦C for 96 h and then flooded with specific staining solutions as follows: Congo red (0.03%) for chitinase and glucanase; lugol solution for amylase; CTAB: hexadecyltrimethylammonium bromide (2%) for pectinase and ruthenium red (0.1%) for polygalacturanase. The enzymatic activity was taken as evidence of the appearance of hydrolysis clear zones around the colonies, and their diameters were measured in millimeters.
