Blog #4 Research Continues!

These past few months have been filled with many lab trials, beginning initially with the analysis of the mid-logarithmic M. bovis-BCG cultures supplemented with glutathione. Each time, these trials showed inconsistent optical density readings and only one general trend. The cells both those in liquid broth media, conditioned with and without glutathione, would lyse. Dr. Kelly and I are now currently in the process of ordering (and eagerly awaiting!) new M.bovis-BCG for that particular set of experiments.However, more importantly  to my research are the latent infection cultures. In replication of my lab’s previous work with the mycobacteria and glutathione, I have manipulated the protocol to my needs, specifically tripling the culture size (and thus 3-fold more bacteria) in order to have a sufficient RNA yield. Data analysis of these samples recently revealed very similar results in accordance with the growth trend on days 4 and 5, however were not the best possible results I believe we are able to obtain. 

Initially, this appeared to be a huge obstacle. We theorized ways to compensate for the physical limitations of upscaling the culture size, which resulted in the inadequate volume displacement (due to the glass beads addition to achieve anaerobic conditions), preventing bacterial contact. Although strikingly, this observation then led my lab to a new hypothesis on the effects of reactivation tuberculosis at the cellular level, illustrating the scientific method at its best. From these results, we suggest that the mechanism of resuscitation from dormancy in the previous cultures was dependent on quorum sensing, or cell to cell communication. Little is currently known about how tuberculosis may function in reactivation, however it is possible that glutathione is, or highly resembles, a small peptide that which mycobacteria may recognize/import as a means of determining cell density, promoting virulence, and resuscitating from long-established dormant cultures. I spent many hours with Dr. Kelly methodically reviewing results from our lab and discussing possible outcomes. I am truly grateful for this experience in biological research, and not only for the kinesthetic practices of techniques and handling of microbial systems, but especially for the mentorship that has progressed my analytical thinking in the methods and developments of biological research.

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