Phosphoethanolamine-modified lipid A enables antimicrobial resistance and atypical signaling properties in the marine pathogen P. damselae subsp. damselae.

Lipid A, the immunostimulatory component of lipopolysaccharide (LPS), plays a multitude of roles in host-pathogen interactions. It functions as both a key antigen recognized by innate immune receptors and an essential structural element of the gram-negative bacterial outer membrane. Consequently, gram-negative bacteria exposed to stressful environments can modify their lipid A structure to preserve membrane integrity, evade immune cell recognition, and avoid disruption by antimicrobial agents. In this study, we isolated a strain (SF21) from a nurse shark stool, which was identified as the gram-negative marine pathogen Photobacterium damselae subsp. damselae via genomic sequencing. Mass spectrometric analysis revealed that its lipid A is extensively modified with phosphoethanolamine, which enabled inherent resistance to the cationic antimicrobial peptide colistin (polymyxin E). Additionally, growth under cold conditions (10°C) led to the replacement of the normally saturated C14 acyl chain in SF21 lipid A with an unsaturated C16:1 acyl chain. Stimulation of NF-κB reporter cells with purified SF21 LPS elicited stronger signaling through human TLR4/MD-2 compared to murine TLR4/MD-2, challenging the paradigm that hexa-acylated LPS is more stimulatory through the murine ortholog of this receptor complex. Altogether, this study characterizes a gram-negative bacterium whose ability to grow in various environments and be detected by the TLR4/MD-2 complex appears linked to its capacity to modify its lipid A. Gram-negative bacteria can remodel their membrane lipids to accommodate the various stresses that they encounter in the environment. One membrane lipid, called lipid A, is a key structural element of the outer leaflet of their outer membrane. While it provides structural integrity, it also serves as a molecular pattern that our innate immune system utilizes to recognize bacteria and mount an immune response. In this study, we show that a marine pathogen, present in the stool of nurse sharks, possesses lipid A that is modified with phosphoethanolamine. This lipid A modification enabled resistance to antimicrobials, promoted growth at temperatures beyond normal shark body temperature, and induced an altered immune response that differs from prototypical LPS. From this study, we conclude that phosphoethanolamine-modified lipid A improves bacterial fitness, enabling better survival across the various environments that a bacterium may encounter.