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Phagocytic uptake of Encephalitozoon cuniculi by nonprofessional phagocytes
S Couzinet1, E Cejas, J Schittny
1Institute for Medical Microbiology, University of Bern, Bern, Switzerland.
Abstract:
Encephalitozoon cuniculi is an obligate intracellular, spore-forming parasite belonging to the microsporidia that can cause disseminated infection in immunocompromised persons. E. cuniculi spores infect host cells by germination, i.e., by explosively everting the polar filament, through which the spore contents (sporoplasms) are subsequently injected into the cytoplasm. In addition, we observed intracellular, nongerminated spores in various nonprofessional phagocytes. In MRC5 cells, the number of internalized spores was approximately 10-fold higher than the number of injected sporoplasms. Compared to the rate of uptake by human monocyte-derived macrophages, internalization rates by A549 cells, MRC5 cells, and 293 cells were 0.6, 4.4, and 22.2%, respectively. The mechanism of uptake was studied in MRC5 cells. Killed spores were internalized at the same rate as live spores, indicating that nongerminated parasites do not actively participate in cell entry. Cytochalasin D inhibited uptake of spores by 95%, demonstrating an actin-dependent process. By electron and epifluorescence microscopy, intracellular spores were found in a tightly fitting membrane-bound compartment. The vacuole containing the spores was positive for the lysosomal membrane protein LAMP-1 and colocalized with the late endosomal-lysosomal content marker rhodamine dextran. Our results show that, in addition to the unique way in which microsporidia infect cells, E. cuniculi spores enter nonprofessional phagocytes by phagocytosis and traffic into a late endosomal-lysosomal compartment.
Insights
Encephalitozoon cuniculi spores infect host cells via phagocytosis, not just germination. These parasites are internalized by nonprofessional phagocytes and trafficked to a late endosomal-lysosomal compartment.
Area of Science:
- Cell biology
- Parasitology
- Infectious diseases
Background:
- Encephalitozoon cuniculi is an obligate intracellular parasite causing disseminated infections in immunocompromised individuals.
- Microsporidia infect host cells through germination, injecting sporoplasms via a polar filament.
- Nongerminated spores are also found within host cells, suggesting alternative entry mechanisms.
Purpose of the Study:
- To investigate the cellular entry mechanisms of Encephalitozoon cuniculi spores into nonprofessional phagocytes.
- To elucidate the intracellular trafficking pathway of E. cuniculi spores after cell entry.
Main Methods:
- Comparative analysis of spore uptake rates in different cell lines (A549, MRC5, 293, macrophages).
- Assessment of live vs. killed spore internalization and the effect of Cytochalasin D to determine the entry mechanism.
- Electron and epifluorescence microscopy to visualize intracellular spores and their compartment.
Main Results:
- Nonprofessional phagocytes internalized E. cuniculi spores via phagocytosis, with uptake rates varying by cell type.
- Spore internalization was an actin-dependent process, independent of parasite viability.
- Intracellular spores were enclosed in a membrane-bound vacuole positive for LAMP-1 and colocalized with late endosomal/lysosomal markers.
Conclusions:
- E. cuniculi spores utilize phagocytosis for entry into nonprofessional phagocytes, in addition to the known germination pathway.
- Internalized spores are directed to a late endosomal-lysosomal compartment within the host cell.