Uncovering bacteriophage–LPS interactions with Gram-negative model membranes

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a Gram-negative enterobacteria of public health concern. The lipopolysaccharide (LPS) leaflet of the Gram-negative outer membrane (OM) has a defensive role and modulates the interactions between the bacterial cell and the microenvironment. Due to the increasing number of antimicrobial-resistant bacteria, interest in alternative treatments, such as phage therapy, has grown. Thus, understanding the mechanisms of phage–host interaction become increasingly important. The bacteriophages Lederbergvirus P22 (P22) and Nonanavirus nv9NA (9NA) infect S. Typhimurium after binding and depolymerizing the O-antigen portion of LPS¹. In vitro studies have shown that a complete LPS structure (lipid A, core, and O-antigen) is essential for P22 and 9NA’s DNA release^2,3. However, the dynamics of the early steps that precede infection, in which phages locate, recognize, and cleave the LPS O-antigen before DNA entry, remain largely unresolved. To address this, we developed LPS-enriched supported lipid bilayers (LPS-SLBs) as model membranes that mimic the S. Typhimurium OM, showcasing their usability as versatile biomimetic platforms for tracking microbial virus interactions. In addition, we investigated phage–surface interactions by tracking single 9NA particles on distinct LPS-SLB surfaces using total internal reflection fluorescence (TIRF) microscopy. This allowed us to monitor phage diffusion on the model membrane surface and characterize the dynamic events leading to the start of the infection. Our results provide mechanistic insight into how the OM composition of Gram-negative hosts guides the efficiency of bacteriophage infection.

References

1. Broeker, N. K. & Barbirz, S. Mol. Microbiol. 105, 353–357 (2017).
2. Andres, D. et al. Mol. Microbiol. 83, 1244–1253 (2012).
3. Stephan, M. S. et al. Preprint at https://doi.org/10.1101/2024.08.19.608551 (2024).