Structural determination of Rickettsia lipid A without chemical extraction confirms shorter acyl chains in later-evolving Spotted Fever Group pathogens.

Deforestation, urbanization, and homelessness lead to spikes in Rickettsioses. Vector-borne human pathogens of Transitional Group (TRG), Typhus Group (TG), and Spotted Fever Group (SFG) rickettsiae differ by clinical manifestations, immunopathology, genome composition, and morphology. We previously showed that lipid A (or endotoxin), the membrane anchor of Gram-negative bacterial lipopolysaccharide (LPS), structurally differs in (later-evolving SFG) relative to (basal SFG), (TG), and (TRG). As lipid A structure influences recognition potential in vertebrate LPS sensors, further assessment of lipid A structural heterogeneity is needed. Here, we sidestepped the difficulty of lipid A chemical extraction by utilizing FLAT , a new procedure for generating lipid A structures directly from host cell-purified bacteria. These data confirm later-evolving SFG pathogens synthesize structurally distinct lipid A. Our findings impact interpreting immune responses to different pathogens and utilizing lipid A adjuvant or anti-inflammatory properties in vaccinology.