Alive Pathogenic and Saprophytic Leptospires Enter and Exit Human and Mouse Macrophages With No Intracellular Replication

Abstract

Leptospira interrogans are pathogenic bacteria responsible for leptospirosis, a zoonosis impacting 1 million people per year worldwide. Leptospires can infect all vertebrates, but not all hosts develop similar symptoms. Human and cattle may suffer from mild to acute illnesses and are therefore considered as sensitive to leptospirosis. In contrast, mice and rats remain asymptomatic upon infection, although they get chronically colonized in their kidneys. Upon infection, leptospires are stealth pathogens that partially escape the recognition by the host innate immune system. Although leptospires are mainly extracellular bacteria, it was suggested that they could also replicate within macrophages. However, contradictory data in the current literature led us to reevaluate these findings. Using a gentamicin–protection assay coupled to high-content (HC) microscopy, we observed that leptospires were internalized in vivo upon peritoneal infection of C57BL/6J mice. Additionally, three different serotypes of pathogenic L. interrogans and the saprophytic L. biflexa actively infected both human (PMA differentiated) THP1 and mouse RAW264.7 macrophage cell lines. Next, we assessed the intracellular fate of leptospires using bioluminescent strains, and we observed a drastic reduction in the leptospiral intracellular load between 3 h and 6 h post-infection, suggesting that leptospires do not replicate within these cells. Surprisingly, the classical macrophage microbicidal mechanisms (phagocytosis, autophagy, TLR–mediated ROS, and RNS production) were not responsible for the observed decrease. Finally, we demonstrated that the reduction in the intracellular load was associated with an increase of the bacteria in the supernatant, suggesting that leptospires exit both human and murine macrophages. Overall, our study reevaluated the intracellular fate of leptospires and favors an active entrance followed by a rapid exit, suggesting that leptospires do not have an intracellular lifestyle in macrophages.

Evolutionary insights into the emergence of virulentLeptospiraspirochetes

“…Genome evolution is a key determinant in host adaptation, but other mechanisms should also be considered. Previous studies had demonstrated that genes controlling adhesion, uptake and intracellular survival within macrophages and neutralization of the complement system [36, 37] are involved in Leptospira pathogenicity. However, we have shown here that most of these genes involved in virulence and host adaptation are also present within the P1- group ( Fig.…”

Differential modulation of innate immune response by lipopolysaccharide of Leptospira

“…Leptospires were labelled with CFSE following the published protocol with minor modifications [55]. Briefly, Leptospira was washed twice in DPBS to remove the proteins in the EMJH media and diluted in 1 ml of DPBS containing 5 µM CFSE and then incubated for 30 min at room temperature, protected from light.…”

Use of an Integrated Multi-Omics Approach To Identify Molecular Mechanisms and Critical Factors Involved in the Pathogenesis of Leptospira

“…Although pathogenic Leptospira is not considered a typical intracellular pathogen, several studies have shown that it can attach to, invade, and induce the apoptosis of mammalian macrophages to escape an innate response during the early stage of infection ( 11 , 12 ). A previous study has demonstrated the differential survivability of L. interrogans within murine and human macrophages; however, a recent study has shown that the bacteria exit both types of macrophages without intracellular replication ( 13 , 14 ). The uptake of L. interrogans by phagocytes was also demonstrated by the zebrafish embryo model, which suggests phagocytosis as a critical defense during early stages of infection ( 15 ).…”