*2.2. Screening of Strains with Antitumor Activity*

All the isolated were cultured on ISP medium 2 (yeast extract-malt extract agar) and incubated at 28 ◦C for 7 days. The spores of the strains were transferred into 250 mL Erlenmeyer flasks containing 30 mL of the production broth containing maltodextrin 4%, lactose 4%, yeast extract 0.5%, and MOPS 2%, at pH 7.2–7.4. on a rotary shaker at 250 r.p.m at 28 ◦C. After seven days, the production broth was extracted with an equal volume of methanol for approximately 24 h. After filtration, the filtrate substances were evaporated under reduced pressure at 50 ◦C to yield the crude extract and then dissolved in DMSO (dimethyl sulfoxide) at concentrations of 20 μg/mL and 100 μg/mL. The HCT-116 (human colorectal carcinoma) cell lines were maintained in Dulbecco's modified Eagle's medium supplemented with 10% (*w*/*v*) fetal bovine serum in a humidified incubator at 37 ◦C of 5% CO2 incubator. The antitumor activities of extracts with two concentrations were investigated by the SRB (Sulforhodamine B) colorimetric method. Briefly, treated cells were harvested and seeded at a density of 5 <sup>×</sup> 104 cells/well into a sterile flat bottom 96-well plate for 24 h, the cells were treated with different concentrations of the extracts for 48 h and growth inhibition was measured by determining the optical density at 510 nm, and the assay was performed basing on an established method [51].
