Evaluation of Trade-Offs in Silver-Resistant E. coli

Student Classification

Jordan Miller, 4th-Year, Biology, Chemistry & Psychology Leilani Whyte, 2nd-Year, Biology


Dr. Misty Thomas, Biology, North Carolina Agricultural and Technical State University

Document Type


Publication Date

Spring 2023


Escherichia coli use two-component response systems (TCRS) to sense, adapt and respond to their environment. To acclimate, E. coli use these TCRSs to activate pathways that allow them to maintain homeostasis. When cultured in stressful environments, bacteria gain adaptive mutations to improve fitness at the cost of losing fitness in other areas. Previous experimental evolution studies produced several adaptive mutations (T14P, R15L, T17P and N279H) in cusS, a sensor protein involved in silver ion efflux. These mutations were recombinantly inserted in the chromosomal DNA of wildtype-E. coli, producing E. coli with each mutation. We have now shown that each of the mutations cause constitutive activation of CusS and we hypothesize that this gain-of-function (GoF) mutation comes at a fitness cost of a slower growth rate. To test this we conducted a competition assay with the wildtype and mutant E. coli using green and red fluorescent proteins (GFP, RFP) to differentiate between them. Growth was measured using a Varioskan plate reader; however, we saw a discrepancy between the growth curves and the number of colonies grown on the plates. From there, it was determined a new excitation and emission wavelength was needed to measure GFP fluorescence. Excitation and emission scans identified new wavelengths at 509 and 407 nm, respectively. Furthermore, when completing competition assays we identified that the serial dilutions for plating required optimization. We concluded the wildtype E. coli has been out-competing the mutant E. coli which supports our hypothesis that the GoF mutations have a growth rate fitness cost.

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