A non-canonical activation of the host's ESCRT machinery is required for the scission of parasitophorous vacuoles and the replication of Leishmania donovani.

Leishmania donovani (Ld) is the causative agent of visceral leishmaniasis, which results in death if not treated. In mammalian cells, Ld live in vacuolar compartments called Leishmania parasitophorous vacuoles (LdLPVs) that enigmatically divide following parasite replication. We evaluated the role of the endosomal sorting complex required for transport (ESCRT) machinery in the scission of LdLPVs. We found that ESCRT components are constitutively recruited to LdLPVs. We propose that this recruitment depends on the expression of PI(3,4)P2 on LdLPVs. The knockdown (KD) of upstream components of the ESCRT machinery revealed that ALIX, but not TSG101 or VPS28, led to a significant reduction in the parasite burden in infected cultures. Interestingly, LdLPVs in ALIXKDs were more distended and harbored more than 2 parasites. Incorporation of BrdU into Leishmania in THP-1 macrophages revealed that parasite replication was inhibited in ALIXKD due to defective LdLPV scission. These findings establish that non-canonical activation of the ESCRT machinery is required for Leishmania to replicate within macrophages.