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Using Bacterial Cytological Profiling to Study the Interactions of Bacteria and the Defense Systems of Multicellular Eukaryotes

  • Author(s): Kalla, Gayathri
  • Advisor(s): Pogliano, Kit
  • et al.
No data is associated with this publication.
Abstract

Multicellular eukaryotes host diverse microbiomes that play important roles in host defense and health. However, little is known about the molecular interactions between the microbiota and the host, and if antimicrobial molecules produced by either the host or commensal bacteria differentially affect beneficial and pathogenic species. This research focuses on antibacterial defense molecules in two settings: plants and human skin. Secondary plant defense metabolites have been noted for their antimicrobial bioactivity in vitro, but their exact mechanisms of action (MOAs) against individual pathogens and if they differentially affect commensal species are unknown. Human skin commensal species Staphylococcus felis has been shown to produce antimicrobial peptides that inhibit the growth of drug-resistant Staphylococcus aureus (MRSA), but the MOA of these bacterial peptides is unknown. To determine the MOAs in each of these contexts, I used Bacterial Cytological Profiling (BCP), a fluorescent microscopy technique to rapidly determine the MOA of a compound in vivo against a bacterial species. BCP was optimized in two plant-associated strains, Agrobacterium tumefaciens and Pseudomonas fluorescens, and databases of cytological profiles were established against compounds with a variety of MOAs. These databases provide references for studying novel plant defense metabolites, setting BCP up as a powerful in vivo assay for future plant research. The differences in BCP phenotypes of Agrobacterium tumefaciens in comparison to the more common distributed growth bacteria may also help us learn more about polar growth organisms. BCP was also used to determine the MOA of the S. felis extract in inhibiting MRSA growth. I observed BCP profiles of condensed chromosomes, suggesting inhibition of translation, and staining of DNA with dyes that can only enter the cell when the cell membrane is compromised, suggesting permeabilization of the membrane. There may be one compound with multiple activities, or multiple compounds with different MOAs. Understanding the MOA of compounds like these is important to developing new alternatives to antibiotics that increasing numbers of infectious bacteria have built up resistance to. This newfound knowledge in each microbiome-host context, as well as the further development of BCP in new species, can enable us to better harness these anti-pathogen properties of hosts and commensal bacterial species for future infection treatment and disease prevention methods.

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This item is under embargo until January 14, 2023.