3′Nucleotidase/nuclease is required for Leishmania infantum clinical isolate susceptibility to miltefosine

Juliana Carniellie, Anuja Dave, Audrey Romano, Sarah Forrester, Pedro Faria, Monti-Rocha, Renata, Carlos Costa, R Dietze, Ian Graham, Jeremy C. Mottram

Research output: Contribution to journalArticlepeer-review

5 Citations (Scopus)
14 Downloads (Pure)

Abstract

Summary
Background Miltefosine treatment failure in visceral leishmaniasis in Brazil has been associated with deletion of the
miltefosine susceptibility locus (MSL) in Leishmania infantum. The MSL comprises four genes, 3′
-nucleotidase/nucleases (NUC1 and NUC2); helicase-like protein (HLP); and 3,2-trans-enoyl-CoA isomerase (TEI).
Methods In this study CRISPR-Cas9 was used to either epitope tag or delete NUC1, NUC2, HLP and TEI, to
investigate their role in miltefosine resistance mechanisms. Additionally, miltefosine transporter genes and
miltefosine-mediated reactive oxygen species homeostasis were assessed in 26 L. infantum clinical isolates. A
comparative lipidomic analysis was also performed to investigate the molecular basis of miltefosine resistance.
Findings Deletion of both NUC1, NUC2 from the MSL was associated with a significant decrease in miltefosine
susceptibility, which was restored after re-expression. Metabolomic analysis of parasites lacking the MSL or NUC1
and NUC2 identified an increase in the parasite lipid content, including ergosterol; these lipids may contribute to
miltefosine resistance by binding the drug in the membrane. Parasites lacking the MSL are more resistant to lipid
metabolism perturbation caused by miltefosine and NUC1 and NUC2 are involved in this pathway. Additionally,
L. infantum parasites lacking the MSL isolated from patients who relapsed after miltefosine treatment were found
to modulate nitric oxide accumulation in host macrophages.
Interpretation Altogether, these data indicate that multifactorial mechanisms are involved in natural resistance to
miltefosine in L. infantum and that the absence of the 3’nucleotidase/nuclease genes NUC1 and NUC2 contributes to
the phenotype.
Original languageEnglish
Article number104378
JournalEBioMedicine
Volume86
DOIs
Publication statusPublished - 2022

Keywords

  • Miltefosine resistance
  • Visceral leishmaniasis
  • CRISPR-Cas9;
  • 3’nucleotidase/nuclease
  • Prognostic marker

Fingerprint

Dive into the research topics of '3′Nucleotidase/nuclease is required for Leishmania infantum clinical isolate susceptibility to miltefosine'. Together they form a unique fingerprint.

Cite this