Abstract
Protozoan parasites of the genus Leishmania alternate between flagellated, elongated extracellular promastigotes found in insect vectors, and round-shaped amastigotes enclosed in phagolysosome-like Parasitophorous Vacuoles (PVs) of infected mammalian host cells. Leishmania amazonensis amastigotes occupy large PVs which may contain many parasites; in contrast, single amastigotes of Leishmania major lodge in small, tight PVs, which undergo fission as parasites divide. To determine if PVs of these Leishmania species can fuse with each other, mouse macrophages in culture were infected with non-fluorescent L. amazonensis amastigotes and, 48 h later, superinfected with fluorescent L. major amastigotes or promastigotes. Fusion was investigated by time-lapse image acquisition of living cells and inferred from the colocalization of parasites of the two species in the same PVs. Survival, multiplication and differentiation of parasites that did or did not share the same vacuoles were also investigated. Fusion of PVs containing L. amazonensis and L. major amastigotes was not found. However, PVs containing L. major promastigotes did fuse with pre-established L. amazonensis PVs. In these chimeric vacuoles, L. major promastigotes remained motile and multiplied, but did not differentiate into amastigotes. In contrast, in doubly infected cells, within their own, unfused PVs metacyclic-enriched L. major promastigotes, but not log phase promastigotes - which were destroyed - differentiated into proliferating amastigotes. The results indicate that PVs, presumably customized by L. major amastigotes or promastigotes, differ in their ability to fuse with L. amazonensis PVs. Additionally, a species-specific PV was required for L. major destruction or differentiation – a requirement for which mechanisms remain unknown. The observations reported in this paper should be useful in further studies of the interactions between PVs to different species of Leishmania parasites, and of the mechanisms involved in the recognition and fusion of PVs.
Highlights
In a classic review of intracellular parasitism, James Moulder proposed that microbial parasites customize the morphology, composition and function of parasitophorous vacuoles (PVs) in which they are sheltered [1]
In phagocytes infected with two different Leishmania species, would the two parasites be found in the same or in separate vacuoles? Of the species chosen, Leishmania amazonensis develops within large vacuoles which shelter many parasites; in contrast, Leishmania major lodges in small PVs containing one or two parasites
Whereas L. major amastigotes remained within their individual vacuoles, L. major promastigotes were delivered to L. amazonensis vacuoles, in which they survived and multiplied but were unable to differentiate into amastigotes
Summary
In a classic review of intracellular parasitism, James Moulder proposed that microbial parasites customize the morphology, composition and function of parasitophorous vacuoles (PVs) in which they are sheltered [1]. In some instances it was possible to generate chimeric vacuoles in cells coinfected with different pathogens [2,3,4,5]. In the present studies macrophages were coinfected with two species of Leishmania parasites normally lodged in PVs that differ in their biogenesis, morphology and parasite occupancy. Elongated, proliferating, extracellular procyclic promastigote forms colonize the midgut of sandfly vectors. These forms, which can be grown axenically, differentiate into infective, stationary phase metacyclics promastigotes that can be released into the dermis of mammalian hosts in the course of the insect bloodmeal. Macrophages and other mammalian cells internalize infective promastigotes within PVs, in which parasites differentiate into the smaller, internally flagellated, oval-shaped amastigote forms. Amastigotes divide intracellularly and spread the infection in the mammal host [6]
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