Neutrophil Extracellular Traps are Involved in the Innate Immune Response to Infection with Leptospira

NETosis is a process by which neutrophils extrude their DNA together with bactericidal proteins that trap and/or kill pathogens. In the present study, we evaluated the ability of Leptospira spp. to induce NETosis using human ex vivo and murine in vivo models. Microscopy and fluorometric studies showed that incubation of human neutrophils with Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 (LIC) resulted in the release of DNA extracellular traps (NETs). The bacteria number, pathogenicity and viability were relevant factors for induction of NETs, but bacteria motility was not. Entrapment of LIC in the NETs resulted in LIC death; however, pathogenic but not saprophytic Leptospira sp. exerted nuclease activity and degraded DNA. Mice infected with LIC showed circulating NETs after 2 days post-infection (dpi). Depletion of neutrophils with mAb1A8 significantly reduced the amount of intravascular NETs in LIC-infected mice, increasing bacteremia at 3 dpi. Although there was a low bacterial burden, scarce neutrophils and an absence of inflammation in the early stages of infection in the kidney and liver, at the beginning of the leptospiruric phase, the bacterial burden was significantly higher in kidneys of neutrophil-depleted-mice compared to non-depleted and infected mice. Surprisingly, interstitial nephritis was of similar intensity in both groups of infected mice. Taken together, these data suggest that LIC triggers NETs, and that the intravascular formation of these DNA traps appears to be critical not only to prevent early leptospiral dissemination but also to preclude further bacterial burden.Author Summary: Neutrophils extracellular traps (NETs) are a relatively novel pathogen-killing mechanism for extracellular microbes independent of phagocytic uptake and degranulation. Although it was originally proposed that NETs are formed exclusively in tissues at sites of infection, NETs have also been found within blood vessels where they ensnare microbes in circulation during sepsis. Leptospirosis is a global zoonosis that has become prevalent in slum areas, where its diagnosis and treatment may be overlooked. Here, for the first time, we characterized NETs formation as a result of Leptospira spp. stimulus. We also showed that mice lacking neutrophils exhibit early reduced levels of circulating NETs, but higher bacteremia, a fact that was later associated with higher bacterial burden in kidney. The present results demonstrate that Leptospira spp. can trigger NET formation and that this innate immune mechanism can play a role in the pathogenesis of leptospirosis.