An important area in the cell biology of intracellular parasitism is the customization of parasitophorous vacuoles (PVs) by prokaryotic or eukaryotic intracellular microorganisms. We were curious to compare PV biogenesis in primary mouse bone marrow-derived macrophages exposed to carefully prepared amastigotes of either Leishmania major or L. amazonensis. While tight-fitting PVs are housing one or two L. major amastigotes, giant PVs are housing many L. amazonensis amastigotes. In this study, using multidimensional imaging of live cells, we compare and characterize the PV biogenesis/remodeling of macrophages i) hosting amastigotes of either L. major or L. amazonensis and ii) loaded with Lysotracker, a lysosomotropic fluorescent probe. Three dynamic features of Leishmania amastigote-hosting PVs are documented: they range from i) entry of Lysotracker transients within tight-fitting, fission-prone L. major amastigote-housing PVs; ii) the decrease in the number of macrophage acidic vesicles during the L. major PV fission or L. amazonensis PV enlargement; to iii) the L. amazonensis PV remodeling after homotypic fusion. The high content information of multidimensional images allowed the updating of our understanding of the Leishmania species-specific differences in PV biogenesis/remodeling and could be useful for the study of other intracellular microorganisms.
Leishmania parasites lodge in host cells within phagolysosome-like structures called parasitophorous vacuoles (PVs). Depending on the species, amastigote forms can be individually hosted within small, tight-fitting PVs or grouped within loose, spacious PVs. Using multidimensional live cell imaging, we examined the biogenesis of the two PV phenotypes in macrophages exposed to L. major (a representative of the tight PV phenotype) or L. amazonensis (an example of the loose PV phenotype) amastigotes. L. major PVs undergo fission as parasites divide; we demonstrate that in the course of fission there are transients of the lysosomotropic fluorescent probe Lysotracker. In contrast, during the course of amastigote population size expansion, L. amazonensis PVs do accumulate Lysotracker while increasing in diameter and volume. The large PVs fuse together, and the products of fusion undergo size and shape remodeling. The biogenesis/remodeling of the two types of Leishmania PVs is accompanied by a reduction in the number of macrophage acidic vesicles. The present imaging study adds new morphometric information to the cell biology of Leishmania amastigote intracellular parasitism.