The obligate intracellular pathogen Simkania negevensis (Sne) resides within a Simkania Containing Vacuole (SnCV), closely interacting with the host endoplasmic reticulum and mitochondria [1]. It is dependent on host metabolites, like sphingolipids, due to limited metabolic capacity. To unravel the role of sphingolipids in both Sne infection and its life cycle, our aim was to investigate the involvement of mitochondrial sphingolipids in Sne infection and examine the trafficking of ceramide among host cell organelles, plasma membrane, and SnCV.

To check the mitochondrial status, Mitochondria-ceramide transport protein expression (Mito-CERT), altering CERT to target mitochondria, was performed. Despite cell survival, this modification had no effect on Sne infection. Subsequently, we screened various inhibitors of sphingolipid metabolism and employed knockout cell lines to identify key factors for Sne survival. We showed that, unlike Chlamydia trachomatis, Sne does not depend on de novo ceramide synthesis or CERT. However, the synthesis of sphingomyelin from ceramide by sphingomyelin synthases 1/-2 is necessary for successful infection. Acid sphingomyelinase and acid ceramidase are crucial as well, indicating the role of lysosomes and the salvage pathway. Sphingolipidome analyses in two cell lines showed a significant change in the ratio between sphingomyelin and ceramide in favor of ceramide, as well as the increase of sphingosine-1-phosphate in infected cells.

Currently, our focus is on whether Sne, similar to Chlamydia, serves as a platform for synthesizing sphingomyelin or ceramide within the host cell. Additionally, we aim to explore the role of lysosomal sphingolipid and cholesterol trafficking, as well as cholesterol modification, in Sne infection.

1.Kunz TC, Kozjak-Pavlovic V. Diverce Facets of Sphingolipid Involvement in Bacterial Infections, Front Cell Dev Biol. 2019; 7:203. doi: 10.3389/fcell.2019.00203

2.Derré I, Swiss R, Agaisse H. The Lipid Transfer Protein CERT Interacts with the Chlamydia Inclusion Protein IncD and Participates to ER-Chlamydia Inclusion Membrane Contact Sites, PLoS Pathog. 2011; 7(6): e1002092 doi:10.1371/journal.ppat.1002092