Spermidine biosynthesis by hypervirulent Francisella tularensis promotes fitness and salvages adenine

Francisella tularensis is a highly infectious Tier 1 select agent that causes tularemia, a potentially fatal disease. F. tularensis subspecies tularensis (type A), including subtypes A.I and A.II, and subspecies holarctica (type B) differ considerably in pathogenicity, with A.I strains being recognized as some of the most virulent bacterial pathogens known. We previously revealed that only the A.I clade can synthesize spermidine de novo. In this study, we show that deletion of spermidine synthase speE (ΔspeE) in prototype A.I strain SCHU S4 reduced spermidine levels relative to wild-type SCHU S4, type B live vaccine strain (LVS), and LVS ΔspeE. In conventional spermidine-containing chemically defined medium (cCDM), SCHU S4 and SCHU S4 ΔspeE grew substantially faster than LVS and LVS ΔspeE; however, SCHU S4 ΔspeE grew slower than SCHU S4. In modified cCDM without spermidine, SCHU S4 ΔspeE grew in a biphasic manner, which was alleviated by methylthioadenosine, adenine, or adenosine supplementation. Conversely, spermidine addition caused diauxic growth by LVS and LVS ΔspeE and was inhibitory above 0.1 mM, while SCHU S4 replication was substantially and similarly promoted by spermidine or spermine supplementation, with 0.02 mM spermidine being sufficient for maximum growth enhancement. These previously unknown traits demonstrate that de novo spermidine synthesis by A.I strains salvages adenine and promotes fitness, while exogenous spermidine and spermine further increase the rapid replication rate of hypervirulent A.I SCHU S4. These F. tularensis A.I attributes are likely an adaptation to an intracellular environment that contributes to persistence, while outcompeting an effective immune response during an infection.IMPORTANCEFrancisella tularensis is one of the most pathogenic bacteria known due to a low infectious dose, expansive host range, and high rate of mortality. This zoonotic pathogen poses a serious risk to public health and, therefore, is considered a potential bioterrorism agent. Our study revealed that hypervirulent strains of F. tularensis endogenously produce spermidine, which promotes fitness and salvages adenine, while exogenous spermidine or spermine further enhances the rapid replication rate of this intracellular pathogen. A better understanding of the inherent traits that allow F. tularensis to persist and outcompete an effective immune response is needed and may provide insight into preventing lethal infections by other intracellular pathogens.