Antibiotic Resistance in Tuberculosis
Determining the Utility of Cell Envelope Labeling as a Readout for Mycobacterial Growth
Tuberculosis presents a growing danger to the world where strains are becoming resistant to some or all antibiotics used to normally treat the sickness. The causative agent of tuberculosis, Mycobacterium tuberculosis, is difficult to treat because of its complex cellular envelope. Using Mycobacterium smegmatis as a model organism, we hope to shine light on the complexity of the cell wall using several fluorescent D-amino acid labels attached to specific chemical probes that allow us to label the three distinct parts of the cell wall. We believe that incorporation of the probes either intracellular or extracellular happens when the cell is still dividing, therefore, a cell that isn’t dividing due to antibiotic treatment should have less fluorescence. The fluorescent tags are attached using a Copper Alkyne Azide Cycloaddition (CuAAC) reaction, also known as a Copper Click reaction. To get an understanding of how antibiotics effect the incorporation of the probes and fluorescence of the labels, microscopy and flow cytometry were used to allow us to visualize and quantify the results. Analysis of the results show that there was a difference in fluorescence when cells were treated with antibiotics. Cells often lost the polarity of their fluorescence, resulting in fluorescence across the cell when it is usually concentrated at the poles. Several different antibiotics and fluorescent labels were used to determine which is most successful at labeling the cell as a readout for growth.