Cattelan, N. | bioRxiv Science

bioRxiv2026

Bloodstream-associated Salmonella Typhimurium and Enteritidis iNTS pathovariants hyper-replicate in human macrophages

Invasive non-typhoidal Salmonella (iNTS) are a major cause of bloodstream infections in sub-Saharan Africa, yet the host-pathogen interaction mechanisms remain poorly understood. Here, we developed and optimised a human macrophage infection model based on PMA-differentiated THP-1 cells to investigate infection dynamics of clinically relevant Salmonella Typhimurium and Salmonella Enteritidis strains. We compared intracellular survival and replication of gastroenteritis-associated and bloodstream-associated pathovariants, including S. Typhimurium ST313 Lineage 2, the novel S. Typhimurium ST313 Lineage 3 and the understudied S. Enteritidis Central/Eastern African (CEAC) clades. Our results reveal that the CEAC S. Enteritidis and ST313 S. Typhimurium iNTS pathovariants hyper-replicate within host cells, compared to global epidemic isolates. The cellular model achieved robust pro-inflammatory polarisation of human macrophages, while revealing limitations in modelling macrophage plasticity. Overall, this work defines clear differences in intracellular behaviour of Salmonella pathovariants and establishes a robust experimental framework for future studies on invasive disease pathogenesis and therapeutic interventions that target iNTS bacteria.

microbiology↗

bioRxiv2020

Lipidation of a bacterial effector is critical for bacterial evasion of host-defense

The Rab32 antimicrobial pathway has been shown to restrict Salmonella Typhi, in mouse macrophages. The broad-host pathogen Salmonella Typhimurium however has evolved a strategy to evade the Rab32 antimicrobial pathway, via its effector protein GtgE. GtgE is a cysteine protease that specifically mediates the cleavage and inactivation of Rab32. Here we show that GtgE association and targeting to membranes is critical for its efficient proteolytic activity. The C-terminus of GtgE contains a CaaX motif, which can be post-translationally modified by the hosts prenylation machinery. Using a combination of confocal microscopy and subcellular fractionation we show that a cysteine in the CaaX motif is crucial for GtgE membrane targeting and, more importantly, GtgE localization to the Salmonella-containing vacuole. We also demonstrated that prenylation of CaaX is important for an effective and fast Rab32 cleavage, which in turn helps Salmonella to successfully survive in macrophages and establish an in vivo infection in mice. Our findings shed light on the importance of a host mediated post-translational modification that targets GtgE to the membranes where it can efficiently cleave and inactivate Rab32, leading to a better Salmonella survival in macrophages. Author summarySalmonella species includes a large group of bacteria that cause disease in different hosts. While some serovars are host generalists, others are restricted to humans. This is the case of Salmonella Typhi, responsible for Typhoid fever, a disease that affects millions globally. We have previously discovered an antimicrobial activity in macrophages that is controlled by Rab32. While the broad-host bacterium Salmonella Typhimurium effectively counteracts this mechanism through the delivery of two effectors, GtgE and SopD2, Salmonella Typhi does not express those effectors and cannot survive in mouse macrophages. In this article, we demonstrate how Salmonella Typhimurium exploits a host machinery to modify GtgE. We show that this host mediated modification is important for GtgE intracellular localization and effective Rab32 targeting, resulting in both a better intracellular survival and infection in vivo.

microbiology↗