Graduation Year

2013

Document Type

Dissertation

Degree

Ph.D.

Degree Granting Department

Public Health

Major Professor

Dennis E. Kyle

Keywords

Drug Resistance, Leishmania, Molecular Mechanism

Abstract

Abstract

Visceral leishmaniasis is a devastating neglected parasitic disease caused by infection with Leishmania donovani. It life cycle has two stages with promastigote (insect stage) and amastigote (animal stage) morphologies. Miltefosine is currently the only commercially available oral drug available to treat leishmaniasis and recent evidence suggests clinical resistance has emerged. Due to the importance of this drug and the scarcity of new drugs in the pipeline, work has been done on understanding the mechanism(s) of miltefosine, yet the mechanism of action for resistance is still not known.

In previous studies investigators generated miltefosine resistance on the insect vector stage (promastigotes), yet there is an important gap in understanding miltefosine resistance in the human infectious stage (amastigotes) of the disease. Before we could accomplish this goal a L. donovani cell line was converted into a stable, continuously cultured axenic amastigote cell line and characterized by disease burden generated in vitro and in vivo.

The axenic line of L. donovani (MHOM/SD/75/1246/130) retains characteristics of amastigotes infecting macrophages and proliferated better than metacyclic promastigotes in macrophages in vitro and hamsters. The axenic amastigote line was used to induce miltefosine resistance by using discontinuous stepwise increasing drug pressure. Stable high-grade resistance was established (62-fold) and characterized in vitro and in vivo. Lastly, the fitness of miltefosine resistant versus susceptible parasite was established for the first time. This work adds the first efforts characterizing and understanding the complex problem of miltefosine resistance in amastigotes.

Download

Included in

Parasitology Commons

Share

COinS