Content area
Full text
About the Authors:
Deepak Kumar Deep
Affiliations National Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delhi, India, Department of Biotechnology, Faculty of Science, Jamia Hamdard, New Delhi, India
Ruchi Singh
Affiliation: National Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delhi, India
Vasundhra Bhandari
Current address: School of Life Sciences, University of Hyderabad, Hyderabad, Telangana, India
Affiliation: National Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delhi, India
Aditya Verma
Affiliation: National Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delhi, India
Vanila Sharma
Affiliation: National Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delhi, India
Saima Wajid
Affiliation: Department of Biotechnology, Faculty of Science, Jamia Hamdard, New Delhi, India
Shyam Sundar
Affiliation: Institute of Medical Sciences, Banaras Hindu University, Varanasi, India
V. Ramesh
Affiliation: Dermatology Department, Safdarjung Hospital and Vardhman Mahavir Medical College (VMMC), New Delhi, India
Jean Claude Dujardin
Affiliation: Unit of Molecular Parasitology, Department of Parasitology, Institute of Tropical Medicine, Antwerp, Belgium
Poonam Salotra
* E-mail: [email protected]
Affiliation: National Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delhi, India
ORCID http://orcid.org/0000-0002-4057-4872Abstract
Background
Miltefosine (MIL) is an oral antileishmanial drug used for treatment of visceral leishmaniasis (VL) in the Indian subcontinent. Recent reports indicate a significant decline in its efficacy with a high rate of relapse in VL as well as post kala-azar dermal leishmaniasis (PKDL). We investigated the parasitic factors apparently involved in miltefosine unresponsiveness in clinical isolates of Leishmania donovani.
Methodology
L. donovani isolated from patients of VL and PKDL at pretreatment stage (LdPreTx, n = 9), patients that relapsed after MIL treatment (LdRelapse, n = 7) and parasites made experimentally resistant to MIL (LdM30) were included in this study. MIL uptake was estimated using liquid chromatography coupled mass spectrometry. Reactive oxygen species and intracellular thiol content were measured fluorometrically. Q-PCR was used to assess the differential expression of genes associated with MIL resistance.
Results
LdRelapse parasites exhibited higher IC50 both at promastigote level (7.92 ± 1.30 MM) and at intracellular amastigote level (11.35 ± 6.48 MM) when compared with LdPreTx parasites (3.27 ± 1.52 MM) and (3.85 ± 3.11 MM), respectively. The percent infectivity (72 hrs post infection) of LdRelapse parasites was significantly higher (80.71 ± 5.67%, P<0.001) in comparison to LdPreTx (60.44 ± 2.80%). MIL accumulation was significantly...