Fig. 1. Live attenuated LmCen^−/− parasites do not cause any pathology in hamster model and do not cause any infection in sand flies.

a Lesion size was monitored every week in hamsters injected with 10⁶-total stationary phase either LmWT or LmCen^−/− parasites by intradermal (ID) injection. Ear lesion diameters were measured at indicated days post inoculation. Results (SD) are representative cumulative effect of two (3 d, 15 d, and 28 d) to three (49 d) independent experiments, (p values were determined by Mann–Whitney two-tailed test). b Photographs of representative ears of LmWT and LmCen^−/− immunized hamsters at 49 days post inoculation. Red arrow indicates the lesion development. c, d Parasite load in the ear (c) and draining lymph node (dLN) (d) of LmWT and LmCen^−/− immunized hamsters were determined by serial dilution assay at 3, 15, and 28 days (n = 6/group of hamsters) and 49 days (n = 15/group of hamsters) post inoculation. Results (Mean ± SD) represent cumulative effect of two (15 d) to three (49 d) independent experiments (p values were determined by Mann–Whitney two-tailed test). e Schematic representation of xenodiagnoses to determine infectiousness of immunized hamsters for sand flies. f Photographs of representative ears of LmWT and LmCen^−/− infected hamsters for xenodiagnoses at 2- and 8- weeks post inoculation. Red arrows indicate the lesion development. g After exposed with infected hamsters (n = 6/group), blood-fed flies were isolated, and parasite positive flies were identified after dissection of flies isolated from both LmWT and LmCen^−/− infected groups at 4 days post-blood feeding and 8 days post-blood feeding. Results (Mean ± SD) are representative one experiment. h Schematic representation of immune-suppression by DXM treatment of LmCen^−/− immunized hamsters. i Photographs of representative ears of LmWT, LmCen^−/−, and LmCen^−/− + DXM treated hamsters. Red arrow indicates the lesion development. j Ear lesion diameters were measured after 4 weeks of DXM treatment (total 15 weeks post parasite infection) in LmWT (n = 6) and LmCen^−/− (n = 12) and LmCen^−/− + DXM (n = 12) treated hamsters. Results (Mean ± SD) represent cumulative effect of two independent experiments, 1 ear, total 6–12 hamsters per group (p values were determined by Mann–Whitney two-tailed test). k, l Parasite load in the inoculated ear (k) and dLN (l) of each group of hamsters (LmWT, n = 6; LmCen^−/−, n = 12; and LmCen^−/− + DXM, n = 12) were determined by limiting dilution assay. Results (Mean ± SD) represent cumulative effect of two independent experiments (p values were determined by Mann–Whitney two-tailed test). BF Blood Fed, DXM Dexamethasone.