Intracellular amastigote replication may not be required for successful in vitro selection of miltefosine resistance in Leishmania infantum.

Although miltefosine (MIL) has only recently been positioned as a first-line therapeutic option for visceral leishmaniasis, field reports note an increasing trend in treatment failures. Study of laboratory selected MIL-resistant strains is needed in the absence of confirmed resistant clinical isolates. In contrast to promastigotes, experimental in vitro selection of MIL-resistance on intracellular amastigotes has not yet been documented. This study reports for the first time the selection of MIL-resistance in Leishmania infantum LEM3323, a strain which clearly shows active intracellular replication. Starting from the hypothesis that active multiplication may be essential in the resistance selection process; several other L. infantum strains were evaluated. Although strain LEM5269 showed only marginally lower intracellular multiplication, selection for resistance failed, as was also the case for several other strains showing poor or no intracellular replication. These results suggest that intracellular multiplication may not be an absolute prerequisite for the outcome of experimental in vitro MIL-resistance selection in clinical field isolates.