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Published on: October 15, 2013
Opsonization Improves Dual Proteomic Analysis of Burkholderia cenocepacia Infections
Michael G Bacus1, Leila Jebeli1, Jessica M Lewis1
1Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne 3000, Australia.
Abstract:
Burkholderia cenocepacia is an opportunistic pathogen associated with severe cystic fibrosis (CF) lung infections, where macrophages serve as both a crucial reservoir and a key mediator of intense inflammation. Currently, there is a paucity of insight into how B. cenocepacia infection impacts both the host and the B. cenocepacia proteomes. A key limitation for understanding the proteomic changes during intracellular replication of B. cenocepacia is its low infectivity, which results in in vitro infection models dominated by uninfected cells. Using antibody-mediated opsonization, we show that improving the efficiency and uniformity of B. cenocepacia internalization enhances dual proteomic analysis and changes in both the host and internalized B. cenocepacia, which can be assessed within a single experimental framework. Opsonization enhances the detection of protein changes, including proinflammatory signaling and macrophage activation markers. Using this dual proteomic approach, we assessed the impact of the B. cenocepacia type 6 secretion system (T6SS) and the T6SS effector TecA at 3 and 24 h postinfection, demonstrating that the presence/activity of the T6SS or TecA does not significantly alter the proinflammatory response of THP-1 cells. Thus, this work demonstrates a simple means for enhancing proteomic analysis of B. cenocepacia infections, enabling dual proteomic studies.
Insights
Improving Burkholderia cenocepacia internalization via opsonization enhances dual proteomic analysis. This method reveals host and bacterial protein changes during infection, aiding cystic fibrosis research.
Area of Science:
- Microbiology
- Immunology
- Proteomics
Background:
- * Burkholderia cenocepacia is an opportunistic pathogen causing severe lung infections in cystic fibrosis (CF) patients.
- * Macrophages are central to B. cenocepacia infection, acting as reservoirs and driving inflammation.
- * Understanding host and bacterial proteomic changes during infection is limited by low in vitro infectivity.
Purpose of the Study:
- * To develop a method for enhanced dual proteomic analysis of B. cenocepacia-macrophage interactions.
- * To investigate the impact of B. cenocepacia infection on host and bacterial proteomes.
- * To assess the role of the type 6 secretion system (T6SS) and TecA in the host inflammatory response.
Main Methods:
- * Antibody-mediated opsonization to improve B. cenocepacia internalization efficiency.
- * Dual proteomic analysis of host (THP-1 cells) and intracellular B. cenocepacia.
- * Assessment of protein changes related to inflammation and macrophage activation.
- * Evaluation of T6SS and TecA effects at 3 and 24 hours postinfection.
Main Results:
- * Opsonization significantly enhanced the efficiency and uniformity of B. cenocepacia internalization.
- * Dual proteomic analysis successfully identified host and bacterial protein alterations.
- * Opsonization improved the detection of proinflammatory signaling and macrophage activation markers.
- * The B. cenocepacia T6SS and its effector TecA did not significantly alter the proinflammatory response in THP-1 cells.
Conclusions:
- * Antibody-mediated opsonization is a simple yet effective method to enhance dual proteomic studies of B. cenocepacia infections.
- * This approach facilitates a deeper understanding of host-pathogen interactions and proteomic dynamics during infection.
- * The study provides insights into the limited role of T6SS and TecA in modulating the early inflammatory response in this model.
