Live-cell imaging and ultrastructure expansion microscopy reveal the dynamic intracellular life cycle of Encephalitozoon intestinalis

Microsporidia are a unique group of fungal pathogens that infect various hosts. Encephalitozoon intestinalis, a human-infecting species, possesses the smallest known eukaryotic genome at 2.3 megabases. As an obligate intracellular parasite, E. intestinalis relies on its host for survival and replication. Understanding E. intestinalis cell biology has been challenging due to genetic limitations and the parasite's small size, which falls within the diffraction limit of traditional microscopy.

To overcome these obstacles, researchers have developed a live-cell imaging technique called silhouette microscopy. This method does not require genetic manipulation of microsporidia and allows for the study of E. intestinalis replication in host cells, from entry to exit. Through this technique, scientists have been able to observe the timing of cell division, the kinetics of the parasite life cycle, and the dynamics of the parasitophorous vacuole, the compartment within which E. intestinalis resides.

The parasitophorous vacuole is observed to frequently undergo fission and fusion, with the ultimate rupture necessary for parasite release from the host cell. To further enhance the resolution of E. intestinalis cell biology, ultrastructure expansion microscopy (U-ExM) was employed on infected cells. U-ExM revealed the organization of the E. intestinalis cytoskeleton and the development of a specialized invasion organelle, the polar tube.

By combining live-cell imaging and U-ExM, researchers have established a platform to investigate the E. intestinalis life cycle and better understand its dynamics and cell biology within host cells.