Opsonization Improves Dual Proteomic Analysis of <i>Burkholderia cenocepacia</i> Infections

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.