Leishmania (Viannia) braziliensis transfectants overexpressing the miniexon gene lose virulence in vivo.

The miniexon gene has a central role in the processing of polycistronic pre-mRNA of kinetoplastids. It is added to the 5' extremity of each mRNA, supplying the 5'-capped structure to the molecule. Previous studies in Leishmania (Leishmania) major showed that the overexpression of the miniexon array attenuates the virulence of the parasite in in vivo assays. The results presented here extend those findings to Viannia subgenus. Leishmania (Viannia) braziliensis was transfected with a cosmid harboring a tandem array of one hundred miniexon gene copies and then characterized by Northern blot analysis. The overexpression of the exogenous gene was confirmed and its effect on the virulence of L. (V.) braziliensis was investigated in hamsters. In BALB/c mice we could not detect parasites during the course of 15 weeks of infection. In addition, hamsters infected with transfectants overexpressing the miniexon gene exhibited only a minor footpad swelling of late onset and failed to develop progressive lesion, these attenuated parasites could be recovered from the inoculation site 1 year after infection. The persistence of parasites in the host indicates that a stable line overexpressing the miniexon may be tested as live vaccine against leishmaniasis.

Leishmania braziliensis isolates differing at the genome level display distinctive features in BALB/c mice.

Leishmania braziliensis is the species responsible for the majority of cases of human cutaneous leishmaniasis in Brazil. In the present study, L. braziliensis isolates from two different geographic areas in Brazil were studied by RAPD, using arbitrary primers. We also evaluated other biological features of these two isolates. We compared (a) the clinical features they initiate or not once delivered subcutaneously as stationary-phase promastigotes in the footpad of BALB/c mice; (b) the parasite load in both the footpad and the draining lymph node; (c) the cytokines present in the supernatant of cultures of the cell suspensions from the draining lymph nodes; and (d) the cell types present at the site of parasite delivery. The results show that the L. braziliensis strain from Ceará (H3227) is genotypically different from the L. braziliensis strain from Bahia (BA788). H3227-parasitized mice developed detectable lesions, whereas BA788-parasitized mice did not. Fifteen days post parasite inoculation there was an increase in the numbers of macrophages and lymphocytes in the footpads, whatever the parasite inoculum. Parasite load at the inoculation site--namely the footpad--did not differ significantly; in draining lymph nodes, however, it increased over the period under study. Early after parasite inoculation, the cells recovered from the draining lymph nodes of BA788-parasitized mice produced higher levels of IFN-gamma, a feature coupled to a higher number of NK cells. Later, after the parasite inoculation, there was an increased content of IL-12p70 and IL-10 in the supernatant of cells recovered from the lymph nodes of H3227-parasitized mice. This comparative analysis points out that L. braziliensis isolates differing in their genomic profiles do establish different parasitic processes in BALB/c mice.