**2.4 Sterilization of glassware and nutrients**

All tools used in microorganism experiments were sterilized to avoid any contamination of microbial solutions. The same procedure was carried out for their nutrients. All glassware and nutrients were sterilized for 20 min at 120°C. For the development of the microorganism, the experimental process of cultivation in nutrient solutions is described as follows: the reculture of the microorganism takes place after recovery from ampoules stored at −80 <sup>ο</sup> C. The nutrient medium is vaccinated with a small amount of the ampoule of the microorganism. The procedure takes place under aseptic conditions, conditions achieved by the use of a reducing flame in order to avoid contamination of the sample. Vaccination of the microorganism is done with a sterile Pasteur pipette, where its nose has been sterilized in flame. The nutrient solution with the microorganism is incubated in a special stirring chamber at 37°C, at approximately 100 rpm for 24 hours in order to optimally develop the microorganism. For the development of the microorganism used each

time, the experimental process of growing it in a Petri dish is as follows: ampoules of the microorganism are inoculated in a Petri dish forming zeta as shown in the following image. The procedure takes place under aseptic conditions using a reducing flame to avoid any complications of the sample. The microorganism is vaccinated in the dish with sterile Pasteur pipette, where its nose has been sterilized as above. The vaccinated dish is then placed in an incubation oven at 37°C for about 24 hours.
