Authors

Discussion

INTRODUCTION: Visceral leishmaniasis (VL) is transmitted by sand flies and varies from an asymptomatic to a life-threatening condition with limited treatment options. The Leishmania-host interaction is shaped by an evolutionary arms race involving inflammatory host responses and parasite immunomodulatory molecules. The macrophage migration inhibitory factor (MIF) is a key immune regulator produced by both the mammalian host (hMIF) and the parasite (pMIF). Upon infection, hMIF induces an anti-parasitic, proinflammatory state, whereas in cutaneous leishmaniasis, L. major mif (LmMIF) supports parasite survival and persistence in macrophages. Although LmMIF has been extensively studied, research regarding the role of hMIF and its parasite orthologs in VL is lacking. To address this knowledge gap, a search for orthologs in L. infantum identified two genes, Linf259 (LINF_330025900) and Linf260 (LINF_330026000). Overexpression and deletion of these genes were used to elucidate their individual roles.

METHODOLOGY: Single overexpressors (OE) were generated through gene cloning and introduction into the chromosomal ssu locus, whereas single and double knockout (KO) lines were obtained through CRISPR-Cas9 gene editing. All transgenic lines were characterized in vitro by assessing (i) their growth and metacyclogenesis rates, and (ii) their ability to infect, differentiate into amastigotes and multiply in primary peritoneal macrophages.

RESULTS: No distinct differences could be observed in promastigote proliferation, metacyclogenesis, and parasite survival in culture. Deletion of LinfMIF caused profound suppression of macrophage infection and in situ proliferation.