CRISPR-Cas9 high-throughput screening to study drug resistance in .

Parasites of the genus pose a global health threat with limited treatment options. New drugs are urgently needed, and genomic screens have the potential to accelerate target discovery, mode of action, and resistance mechanisms against these new drugs. We describe here our effort in developing a genome-wide CRISPR-Cas9 screen in , an organism lacking a functional nonhomologous end joining system that must rely on microhomology-mediated end joining, single-strand annealing, or homologous recombination for repairing Cas9-induced double-stranded DNA breaks.

Increased copy number of the target gene squalene monooxygenase as the main resistance mechanism to terbinafine in Leishmania infantum.

We use here two genomic screens in an attempt to understand the mode of action and resistance mechanism of terbinafine, an antifungal contemplated as a potential drug against the parasite Leishmania. One screen consisted in in vitro drug evolution where 5 independent mutants were selected step-by-step for terbinafine resistance. Sequencing of the genome of the 5 mutants revealed no single nucleotide polymorphisms related to the resistance phenotype.

Mutations in an Aquaglyceroporin as a Proven Marker of Antimony Clinical Resistance in the Parasite Leishmania donovani.

Background: Antimonial drugs have long been the mainstay to treat visceral leishmaniasis. Their use has been discontinued in the Indian subcontinent because of drug resistance, but they are still clinically useful elsewhere. The goal of this study was to find markers of antimony resistance in Leishmania donovani clinical isolates and validate experimentally their role in resistance.