Proteomic and Lipidomic Analysis of Mycobacteriophages Zalkecks and PotatoSplit
Ever since the invention of antibiotics nearly a century ago,the threat of antibiotic resistance has been gradually increasing. As antibiotics are continually prescribed, the rate at which bacteria are becoming resistant to antibiotics is increasing as well. It is projected that antibiotic resistance is one of the largest threats to overall world health, and bacteriophage therapy is one of the leading strategies to combat it. Bacteriophages are viruses that infect and kill specific host bacteria andcan potentially be utilized to kill desired bacteria causing infections that are resistant to antibiotics.
The purpose of this research project is to learn more about the bacteriophage-host interaction through mass spectrometry and bioinformatic tools. This is done through the analysis of proteins and lipids that are produced when the bacteriophage infects the host bacteria. The growth curve of a Passage One From Frozen (P1FF) and a Passage Two From Frozen (P2FF) sample of Mycobacterium smegmatiswas calculated to determine to optimum time for bacteriophage infection. Twobacteriophages were chosen, PotatoSplit and Zalkecks, the Mycobacterium smegmatis samples were infected, samples collected, and mass spectrometry performed. A large portion of this research project is based on the analysis of the proteins and lipids that are produced during each bacteriophage’s infection. Proteomic and lipidomic strategies can be implemented to understand more about the bacteriophage-host interaction and discover any proteins and lipids that are produced at varying timepoints throughout the inoculation process. Bioinformatic tools can then be used to understand the potential functions of each protein or lipid and potential functions or applications of the bacteriophage in general, including the pathogenicity of each bacteriophage.
Determined from proteomic and lipidomic analysis, a list of all proteins and lipids found within each phage infected sample was made. An important trend discovered is that more phage proteins were expressed at later times during the phage infection –Hour 7 and Hour 10, whereas more bacterial proteins were expressed initially –Hour 0 and Hour 3. A case study to investigate the usage of different intensity types produced from mass spectrometry was completed. Overall, it was determined that both the number of phageproteins and bacterial proteins can differ depending on if LFQ or iBAQ intensity type data was used. Correlation between proteins and lipid ontology classes was performed and shows whether groups of lipids are upregulated or downregulated at 14each time point. Understanding the function of lipid ontology groups and the type of regulation provides insight into how the phage or bacteria are potentially using the lipids produced. Some of the main findings include lipids that are involved in bacterial defense mechanisms/energy usage increase over time. Some correlation trends were not consistent across the different bacteriophages, which can be contributed to the different phage life cycles and therefore different phage-host interactions. Further investigation should be performed to determine the specific biological function of proteins and lipids to confidently make claims about potential applications for each phage. Also, further investigation should be performed to understand if the differences in results between bacteriophage PotatoSplit and Zalkecks are due to the varying life cycles.
- Master of Science
- Agricultural and Biological Engineering
- West Lafayette