Pathogens as sensors for measuring immune defence efficiency in the on-going infection
Parasitology and Biology of Tropical Infectious Disease Pathogens
Final Report Abstract
To fight intracellular pathogens, the immune system has evolved powerful defence mechanisms, such as T cell responses, which promote the induction of reactive oxygen and nitrogen species within phagocytes. These antimicrobial molecules enable phagocytic cells to control pathogens that they have taken up. It has however remained unclear how such intracellular defence mechanisms contain the infection in vivo: is killing of the pathogen the only way of achieving control over the pathogen, or does the inhibition of pathogen growth critically contribute as well? The main difficulty to investigate this question has been the lack of tools to determine how exactly the immune response influences the pathogen in order to contain it. However, telling apart such different modes of action of pathogen control is of fundamental interest for understanding infectious diseases and for improving antimicrobial therapy, as antibiotic treatment success can vary dramatically depending on pathogen metabolic activity and growth rate. Furthermore, quantitative differences in pathogen killing can influence the activation of inflammatory signaling: Dying pathogens release different molecules than alive microbes, and are thus perceived differnently by the immune system. In the present project, we have established and characterized genetically encoded reporter systems to measure the viability and growth rate of the unicellular parasite L. major in the ongoing infection of the skin. This has enabled us to determine how the immune response impacts on the parasite over the course of an infection, which is marked by an acute phase with severe skin pathology, and a late asymptomatic phase with low pathogen numbers in the tissue. Specifically, we have set up a new pH-based approach to measure L. major viability based on the fact that, at the site of infection, the parasite resides within acidified compartments of infected cells. Consequently, a loss of parasite membrane integrity is expected to decrease the parasite’s pH. Thus, the reporter system enabled us to monitor the loss of parasite membrane integrity in real time. We could show that this loss in membrane integrity preceded parasite death by several hours, thus, dying parasite could be readily identified in vivo. While parasite death increased with the onset of the immune response, surprisingly, nearly no parasites were killed at late stages of the infection, a phase at which L. major had been widely controlled and was only present at low numbers. How can such low numbers of parasites be maintained without overt killing activity by the immune system? In order to answer this question, we employed a different reporter system that allows to measure the speed by which L. major grows in the tissue. Unexpectedly, we observed that parasite proliferation is highest at the peak of the pathology, that is, when the largest number of activated immune cells is present at the infection site. In contrast, in the late phase of the infection, when no overt killing of parasites by the immune system was observed, L. major growth was strongly reduced. Thus, a small but steady population of persistent parasites seems to be maintained at the site of infection, which is known to play an important role for the immune system to defend the organism against further infections by L. major. Using the parasite growth reporter system, we were also able to distinguish cells which harboured L. major that had a high growth rate from cells infected by slow-growing parasites. Therefore, we sought to identify infected cell populations which were permissive or non-permissive for high L. major growth. We identified cells that were expressing the surface marker CD11c to be especially overrepresented among cells infected with fast-growing L. major. Interestingly, this cell population had been described earlier to be important for the induction of an efficient immune response. Therefore, CD11c-expressing phagocytes at the site of leishmanial infection seem to fill a dual role, on the one hand representing a niche for fast parasite growth, but on the other hand being important for the induction of an efficient immune response.
Publications
- De novo phosphorylation and conformational opening of the tyrosine kinase Lck act in concert to initiate T cell receptor signaling. Sci Signal. 2017 Jan 17;10(462)
Philipsen L, Reddycherla AV, Hartig R, Gumz J, Kästle M, Kritikos A, Poltorak MP, Prokazov Y, Turbin E, Weber A, Zuschratter W, Schraven B, Simeoni L, Müller AJ
(See online at https://doi.org/10.1126/scisignal.aaf4736) - Frontline Science: Leishmania mexicana amastigotes can replicate within neutrophils. J Leukoc Biol. 2017 Nov;102(5):1187-1198
Hurrell BP, Beaumann M, Heyde S, Regli IB, Müller AJ, Tacchini-Cottier F
(See online at https://doi.org/10.1189/jlb.4HI0417-158R) - CD11c-expressing Ly6C+CCR2+ monocytes constitute a reservoir for efficient Leishmania proliferation and cell-to-cell transmission. PLoS Pathog. 2018 14:e1007374
Heyde S, Philipsen L, Formaglio P, Fu Y, Baars I, Höbbel G, Kleinholz CL, Seiß EA, Stettin J, Gintschel P, Dudeck A, Bousso P, Schraven B, Müller AJ
(See online at https://doi.org/10.1371/journal.ppat.1007374)
