Genomic and metabolomic polymorphism among experimentally selected paromomycin-resistant Leishmania donovani strains

Shaw, C. D. and Imamura, H. and Downing, T. and Blackburn, G. and Westrop, G. D. and Cotton, J. A. and Berriman, M. and Sanders, M. and Rijal, S. and Coombs, G. H. and Dujardin, J. C. and Carter, K. C. (2019) Genomic and metabolomic polymorphism among experimentally selected paromomycin-resistant Leishmania donovani strains. Antimicrobial Agents and Chemotherapy. ISSN 0066-4804 (In Press)

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    Abstract

    Understanding the mechanism(s) underpinning drug resistance could lead to novel treatments to reverse the increased tolerance of a pathogen. In this study paromomycin (PMM) resistance (PMM-R) was induced in three Nepalese clinical strains of L. donovani, with different inherent susceptibility to antimony drugs (Sb), by step-wise exposure of promastigotes to PMM. Exposure to PMM resulted in the production of mixed populations of parasites even though a single cloned population was used at the start of selection. PMM IC50 values for PMM-R parasites varied between 104-481 µM at the promastigotes stage and 32-195 µM at the intracellular amastigotes stage. PMM resistance was associated with increased resistance to nitric oxide at the amastigote but not the promastigote stage (p < 0.05). This effect was most marked in the Sb-R PMM-R clone, where PMM-R resistance was associated with a significant upregulation in glutathione compared to its WT (p < 0.05) although there was no change in trypanothione (detected in its oxidised form). Interestingly, PMM-R strains showed an increase in either the keto acid derivative of isoleucine (Sb-I PMM-R) or the 2-hydroxy acids derived from arginine and tyrosine (Sb-S PMM-R and Sb-R PMM-R). These results are consistent with the recent finding that upregulation of the branch-chain amino acid aminotransferase and the D-lactate dehydrogenase are linked to PMM-R. In addition, we found that PMM-R was associated with a significant increase in aneuploidy during PMM selection in all the strains, which could allow rapid selection of genetic changes that confer a survival advantage.