Comparative Fitness of a Parent Leishmania donovani Clinical Isolate and Its Experimentally Derived Paromomycin-Resistant Strain

Abstract

Paromomycin has recently been introduced for the treatment of visceral leishmaniasis and emergence of drug resistance can only be appropriately judged upon its long term routine use in the field. Understanding alterations in parasite behavior linked to paromomycin-resistance may be essential to assess the propensity for emergence and spread of resistant strains. A standardized and integrated laboratory approach was adopted to define and assess parasite fitness of both promastigotes and amastigotes using an experimentally induced paromomycin-resistant Leishmania donovani strain and its paromomycin-susceptible parent wild-type clinical isolate. Primary focus was placed on parasite growth and virulence, two major components of parasite fitness. The combination of in vitro and in vivo approaches enabled detailed comparison of wild-type and resistant strains for which no differences could be demonstrated with regard to promastigote growth, metacyclogenesis, in vitro infectivity, multiplication in primary peritoneal mouse macrophages and infectivity for Balb/c mice upon infection with 2 x 107 metacyclic promastigotes. Monitoring of in vitro intracellular amastigote multiplication revealed a consistent decrease in parasite burden over time for both wild-type and resistant parasites, an observation that was subsequently also confirmed in a larger set of L. donovani clinical isolates. Though the impact of these findings should be further explored, the study results suggest that the epidemiological implications of acquired paromomycin-resistance may remain minimal other than the loss of one of the last remaining drugs effective against visceral leishmaniasis.

Evolution of resistance to single and combined floral phytochemicals by a bumble bee parasite

“…paromycin resistance were found in L. donovani (Hendrickx et al, 2015); in L. infantum, miltefosine resistance was costly, but paromycin resistance resulted in increased growth and enhanced tolerance to stress (Hendrickx et al, 2016). Resistance to pro-oxidant antimonial drugs can actually improve L. donovani infectivity and establishment in hosts, presumably because the superior antioxidant defences of resistant lines allow them to tolerate host immune responses and the stress of initial establishment (Vanaerschot et al, 2011).…”

Phenotypic adaptations of Leishmania donovani to recurrent miltefosine exposure and impact on sand fly infection

“…Although a Sbresistance background may alter the interaction with several drugs [30], it does not impact on the experimental selection for MIL resistance [31], hence allowing the use of both strains without challenging our stated research aims. The intrinsically low susceptibility of BPK282/0-cl4 towards paromomycin, a feature that has been associated with increased infectivity, could possibly explain its relatively more easy adaptation to the hamster model [22].…”

“…Promastigote growth curves were made [22] to compare the in vitro growth of the WT parent and MIL-exposed derived strains. After passage through fine needles (21 G × 1½″, 0.8 × 40 mm and 25 G × 5/8″, 0.5 × 16 mm) to break clustering, the promastigotes were diluted in PBS and counted by flow cytometry.…”

“…The flagellum and cell body lengths of at least 50 parasites per slide were determined using the Axiovision ® software and the percentage of metacyclics present in each sample was calculated. Parasites were considered metacyclic when the flagellum/cell body ratio exceeded 2 [22].…”

Transcriptome profiling identifies genes/pathways associated with experimental resistance to paromomycin in Leishmania donovani

“…PMM is an effective treatment option for antileishmanial therapy and emergence of resistance towards this drug can only be appropriately judged upon its regular use in the field. However, understanding the alterations in the parasite behaviour associated with PMM resistance is essential to assess the propensity for emergence and spread of resistant strains ( Hendrickx et al, 2015 ). To understand the mechanism of resistance towards PMM, drug resistant parasite lines were selected in vitro and mRNA expression profiling was compared between PMM-S and PMM-R parasites.…”