MSACL 2018 US Abstract

Topic: Microbiology/Virology

Exploring the Hidden World of MRSA Pathogenesis

Jacob Wozniak (Presenter)
UCSD

Authorship: Jacob Wozniak (1), John Lapek (1), David Gonzalez (1)
(1) University of California San Diego

Short Abstract

Staphylococcus aureus remains the leading cause of skin and soft tissue infections in the US. During the course of S. aureus infection, the interplay between host and pathogen involves a complex crosstalk mediated by several protein factors. To date, many of the bacterial proteins remain beyond our limits of detection. Specifically, in the context of infection, high host protein levels can mask the signal of bacterial proteins. By using a targeted proteomics approach, we hope to overcome the limitations of identifying bacterial proteins that are typically lost or discarded when analyzing host-pathogen interactions. Implementation of this technique in a clinical setting would theoretically shorten time to positive diagnosis because bacterial proteins could be targeted directly from samples without the need for culture.

Long Abstract

Introduction

Staphylococcus aureus remains the leading cause of skin and soft tissue infections in the US. During the course of S. aureus infection, the interplay between host and pathogen involves a complex crosstalk mediated by several protein factors. To date, many of the bacterial proteins remain beyond our limits of detection. Specifically, in the context of infection, high host protein levels can mask the signal of bacterial proteins. This obscuration severely hampers the ability to directly investigate the proteins involved in infections at a systems scale. Therefore, new approaches must be undertaken to dig deeper into the hidden world of S. aureus pathogenesis. By using a targeted proteomics approach, we hope to overcome the limitations of identifying bacterial proteins that are typically lost or discarded when analyzing host-pathogen interactions. This innovative approach will provide novel biological insight into how S. aureus remains such a prevalent pathogen despite the fact that it has been the focus of intense investigations for decades. Additionally, we envision this technique can be readily applied to clinical samples as a diagnostic tool. Implementation in a clinical setting would theoretically shorten time to positive diagnosis because bacterial proteins could be targeted directly from samples without the need for culture.

Methods

Using a combination of Tandem-mass Tags (TMT) zero and 10plex reagents, we have targeted bacterial proteins in the background of the host proteins during an infection. Bacterial cell lysate will be labeled with TMTzero and the infection samples will be labeled with TMT10plex. Utilizing the mass offset of TMTzero, we will guide the mass spectrometer to target bacterial proteins that would typically not be selected for fragmentation and quantitation.

Results

Utilizing our targeted approach, we have reported the detection of bacterial peptides that were not identified in a conventional proteomics experiment.

Conclusions & Discussion

This proof of principle example shows our ability to detect low level bacterial proteins in the background of host cells. This technique reduces the analysis time of typical proteomics experiments and increases the sensitivity for detecting specific peptides. Therefore, we envision it to be an incredibly useful tool for rapid diagnosis of infections in a clinical setting.


References & Acknowledgements:


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