Investigation of Proteomic and Lipidomic Mycobacteriophage Mass Spectrometry Data

Antibiotic resistance has been an increasing threat to humans since the inception of antibiotics. It can affect anyone, and the issue gets increasingly worse the more antibiotics are used over time. Bacteriophages are one potential way to fight back by harnessing their ability to infect and kill specific bacterial hosts, however, more needs to be studied about them before potential medical applications can be implemented.

The purpose of this research is to analyze the proteins and lipids being produced in the bacteriophage-host interaction to better understand their relationship through the use of mass spectrometry and bioinformatics tools. In this study, the bacterial growth curve of Mycobacterium smegmatis was measured to determine the time of ideal bacteriophage inoculation. Methods of protein and lipid extraction were then tested on M. smegmatis to determine the most effective protocols applicable to the bacterial host and thereby mycobacteriophages. With the ideal extraction protocols and time at which to inoculate the host, two phages were chosen, extraction was implemented, and mass spectrometry was performed on the proteins and lipids present. This exploratory study is based on the data analysis, showing what proteins and lipids are produced as a result of bacteriophage inoculation over time and what that illuminates about the bacteriophage- host interaction. Through the use of modern methods of untargeted proteomics and lipidomics, one has the capability to fill these gaps of what is being produced by the bacteriophage and host in this interaction and expand upon potential bacteriophage functions to provide a more comprehensive understanding of the pathogenesis of the infection.

Proteomic analysis determined that the acetone method of extraction was the most applicable to M. smegmatis and was used for further phage-treated samples and the method of proteomic data analysis tested on readily available mycobacteriophage data. Lipidomic analysis determined that the Bligh Dyer method of extraction was the most applicable to M. smegmatis and was used for further phage-treated samples. The lipids extracted from the phage-treated samples were categorized according to classification and showed functions relating to the cell membrane and energy utilization. Specific lipids from the phage-treated samples also indicated involvement in the abortive infection mechanisms of the phage-host interaction.