Characterisation of antimony-resistant Leishmania donovani isolates: biochemical and biophysical studies and interaction with host cells

Recent clinical isolates of Leishmania donovani from the hyperendemic zone of Bihar were characterised in vitro in terms of their sensitivity towards sodium stibogluconate (SSG) in a macrophage (MPhi) culture system. The resulting half maximal effective concentration (EC(50)) values were compared with those of known sensitive isolates. Fifteen of the isolates showed decreased sensitivity towards SSG with an average EC(50) of 25.7+/-4.5mug/ml pentavalent antimony (SbV) (defined as antimony resistant (Sb-R)), whereas nine showed considerable sensitivity with an average EC(50) of 4.6+/-1.7mug/ml (defined as antimony sensitive (Sb-S)). Out of those nine, seven were recent clinical isolates and the remaining two were known sensitive isolates. Compared with the Sb-S, Sb-R isolates showed enhanced expression of thiol metabolising enzymes in varying degrees coupled with increased intracellular non-protein thiol content, decreased fluorescence anisotropy (inversely proportional with membrane fluidity) and over-expression of the terminal glycoconjugates (N-acetyl-d-galactosaminyl residue). MPhis infected with Sb-R but not with Sb-S showed up-regulation of the ATP Binding Cassette transporter multidrug resistance protein 1 (MRP1) and permeability glycoprotein (P-gp), while the supernatant contained abundant IL-10. The above results reinforce the notion that Sb-R parasites have undergone a number of biochemical and biophysical changes as part of their adaptation to ensure their survival in the host.