Team:US AFRL CarrollHS/Demonstrate
Destroy Mechanism Testing
To determine the efficacy of our destroy mechanism, we decided to test the ability of cinnamaldehyde and chitinase to inhibit the growth of Yarrowia lipolytica. We used Yarrowia lipolytica for our testing because it was one of the fungal isolates from fuel tanks that Dr. Wendy Goodson provided us. We obtained the cinnamaldehyde and chitinase we used from Sigma. We used a chitinase from Streptomyces griseus for the testing. One of the chitinases we designed our microbe to produce also originated from Streptomyces griseus, which should act very similarly to the chitinase we tested with, even though they are not exactly the same chitinase. For our experiment, we plated dilute fungi culture on TSA plates, let them dry, and then pipetted spots of cinnamaldehyde, chitinase, and a combination of both onto the plates (the full protocol for our testing can be found on our Experiments page). After letting them incubate at 27°C for 1-2 days, we examined the plates for zones of clearing. Plates from one of our tests are shown below.
Figure 1. Chitinase test plate. The concentration of chitinase for the top row is 10 mg/ml, 1 mg/ml for the middle row, and 0.1 mg/ml for the bottom.
Figure 2. Cinnamaldehyde test plate. The concentration of cinnamaldehyde for the top row is 0.66 mg/ml, 0.33 mg/ml for the middle row, and 0.16 mg/ml for the bottom.
Figure 3. Combined test plate. The concentration of chitinase and cinnamaldehyde respectively for the top row is 10 mg/ml and 0.66 mg/ml, 10 mg/ml and 0.33 mg/ml for the second row, 1 mg/ml and 0.66 mg/ml mg/ml for the third row, and 1 mg/ml and 0.33 mg/ml for the bottom row.
To determine the efficacy of our destroy mechanism, we decided to test the ability of cinnamaldehyde and chitinase to inhibit the growth of Yarrowia lipolytica. We used Yarrowia lipolytica for our testing because it was one of the fungal isolates from fuel tanks that Dr. Wendy Goodson provided us. For our experiment, we plated dilute fungi culture on TSA plates, let them dry, and then pipetted spots of cinnamaldehyde, chitinase, and a combination of both onto the plates (the full protocol for our testing can be found on our Experiments page). After letting them incubate at 27°C for 1-2 days, we examined the plates for zones of clearing. Plates from one of our tests are shown below.