Isolating Group A Streptococcus Infected Phagosomes

The number of Group A Streptococcus (GAS) infections has dramatically risen in the last several decades and is an increasing public health concern worldwide. Normally, the body’s innate immune system responds to pathogens by phagocytosing and degrading the bacteria within the phagolysosome by a number of different molecular processes, including the oxidative burst response and lysosomal acidification. However, it has been shown by our lab and others that GAS has a number of different mechanisms to subvert immune defenses, including evasion of digestion in the phagolysosome. Because it is unusual for pathogens to reside in phagolysosomes, we want to develop a proteomic profile of GAS‐infected phagosomes. Because isolation of phagosomes is technically difficult, we set out to develop a method to isolate and purify GAS‐infected phagosomes using magnetic particles. We coupled magnetic particles to live GAS and demonstrated that we can effectively isolate viable coupled bacteria. Following infection of THP‐1 macrophages, we employ a magnetic separation protocol that yields a high concentration of GAS‐infected phagosomes largely absent of other organellar contamination. With these magnetically‐isolated GAS phagolysosomes, we aim to conduct a proteomics survey using mass spectrometry to characterize GAS phagosome modification to elucidate its mechanisms of survival. Our work will help identify novel therapeutic targets and strategies to treat GAS infections to enhance normal immune processes aimed at combating GAS infections, and thus limiting the use of antibiotics. Support or Funding Information American Heart Association (17GRNT3410851), Occidental College Undergraduate Research Center