Team:Penn/CdTolerance

Introduction

In order to appropriately determine the effectiveness of using AMB-1 as a viable bioremediation alternative to E.Coli, we performed tests to measure the natural tolerance of both strands to the pollutant cadmium. The experiment allowed us to identify the threshold concentration of normal growth of bacteria. We used the tests to ensure that the AMB-1 strain could withstand the levels of cadmium present in the environment currently. We were able to measure the growth curves of three different strains of bacteria: NEB Turbo, MG1665 and W3110. The growth curve of W3110 is shown here. To see how the other two strains of bacteria react under cadmium challenge, visit the supplementary materials page. . Additionally, our procedure for creating the growth curves is outlined in . From the data, we can conclude that Magnetospirillum magneticum AMB-1 maintains normal growth under 10 uM concentration of cadmium and showed no growth in media with above 100 uM of cadmium ions. E.Coli showed unaffected growth in media with less than 500uM of cadmium ions and showed zero growth with 5000uM of cadmium ions. Although E. coli seemed to be inherently more tolerant towards cadmium than AMB-1, the data does show that AMB-1 can tolerate cadmium concentrations usually toxic to ecosystems. This shows that the strain could potentially still be a viable bioremediation tool.

Purpose

To investigate the effects of increasing cadmium concentrations on the growth of different strains of E. coli and AMB-1. To identify the threshold concentration of normal growth of bacteria

Experiments Protocol

1. Inoculate from glycerol stock in 2mL E-MSGM in culture tube, and a fourth tube of 2ml E-MSGM only
2. Grow culture in 30 C until saturation
3. Check if the media only tube has any growth
4. Dilute bacterial culture 1:100 into 50mL falcon tube (add 400 uL culture to 39.6 mL media)
5. check OD600 of the culture every 24 hours until it reaches approximately OD600 0.1.
6. Prepare 18 2mL eppendorf tubes and add cadmium to corresponding concentration
7. Grow the spiked culture in 30C incubator and measure OD600 every two hours right after addition of cadmium and every 24 hours afterwards.

Conclusion

From the data, we can conclude that Magnetospirillum magneticum AMB-1 maintains normal growth under 10 uM of cadmium ions concentration and showed no growth in media with above 100 uM of cadmium ions. All three strains of E.coli showed unaffected growth in media with less than500uM of cadmium ions and showed zero growth with 5000uM of cadmium ions. AMB-1 (grown in MSGM) is more sensitive to cadmium ions in media than E.coli (grown in LB media).

We recommend future studies into the effect of cadmium on these strains of bacteria under different growth media and determination of the threshold concentration for normal growth.