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SummaryLeishmania parasites and dendritic cell interactions (DCs) play an essential role in initiating and directing T cell responses and influence disease evolution. These interactions may vary depending on Leishmania species and strains. To evaluate the correlation between Leishmania major (Lm) virulence and in-vitro human DC response, we compared the ability of high (HV) and low virulent (LV) Lm clones to invade, modulate cytokine production and interfere with differentiation of DCs. Clones derived from HV and LV (HVDlmpdi and LVDlmpdi), and deleted for the gene coding for a Lm protein disulphide isomerase (LmPDI), probably involved in parasite natural pathogenicity, were also used. Unlike LV, which fails to invade DCs in half the donors, HV promastigotes were associated with a significant increase of the infected cells percentage and parasite burden. A significant decrease of both parameters was observed in HVDlmpdi-infected DCs, compared to wild-type cells. Whatever Lm virulence, DC differentiation was accompanied by a significant decrease in CD1a expression. Lm clones decreased interleukin (IL)-12p70 production similarly during lipopolysaccharide (LPS)-induced maturation of DCs. LPS stimulation was associated with a weak increase in tumour necrosis factor (TNF)-a and IL-10 productions in HV-, HVDlmpdi-and LVDlmpdi-infected DCs. These results indicate that there is a significant variability in the capacity of Lm clones to infect human DCs which depends upon their virulence, probably involving LmPDI protein. However, independently of their virulence, Lm clones were able to down-regulate CD1a expression during DC differentiation and IL-12p70 production during DC maturation, which may favour their survival.

Trying to identify virulence genes of wild Leishmania (L.) major parasites, the species responsible for zoonotic cutaneous leishmaniasis, we compared, using differential display technique, gene expression in two L. major isolates obtained from human lesions and characterized by their contrasting pathogenicity in the BALB/c mouse model. The analysis was performed on amastigotes derived from BALB/c mice lesions. A total of 13 different clones were identified, but the use of reverse transcription and real-time polymerase chain reaction technique did not allow us to confirm any of these clones as differentially expressed. However, the fact that we used the amastigote stage of the parasite led us the identification of amastigote-specific genes, essentially (8 among 13). They are overexpressed, two to seven times, in amastigotes relative to promastigotes. Sequence analysis revealed that two of them namely LPG3 and the ATP dependent RNA helicase correspond to previously described amastigote-specific genes. The others correspond to genes involved in important biological process. Their better characterization could help the development of new drugs targeting the processes in which these molecules are involved.

Cellular and humoral immune responses of healed cutaneous leishmaniasis and Mediterranean visceral leishmaniasis patients were evaluated against results for Leishmania major virulence proteins L. major protein disulfide isomerase (LmPDI) and mitogen-activated protein kinase kinase (MAPKK). Only MAPKK induces significant peripheral blood mononuclear cell proliferation with gamma interferon production as well as antibody responses. Thus, MAPKK may be of interest in Leishmania vaccination and serodiagnosis.The leishmaniases are diseases caused by vector-borne pathogens that represent a major public health problem affecting the lives of millions of people worldwide (http://www.who .int/whr/en). Depending on the parasite species and on the immunological response of the human host, leishmaniasis ranges from an asymptomatic infection to a self-limiting cutaneous lesion(s) or a fatal visceral form.No anti-Leishmania vaccine is available at the moment. Different studies showed that development of Th1-and Leishmania-specific cytotoxic immune responses correlate with healing of patients with cutaneous leishmaniasis (CL) (3,12). An intense effort is being made to identify antigens that could induce an immune state similar to that developed by individuals who recover from symptomatic infection and are resistant to a subsequent natural challenge. Such antigens may contribute to the development of an anti-Leishmania vaccine.Diagnostic tools targeting leishmaniasis are available. However, parasite detection is invasive and poorly sensitive. Serological diagnosis using various techniques based on detection of Leishmania-specific antibodies that are developed during acute disease are often more sensitive, less time-consuming, and more user friendly (4,11,15).In an attempt to identify new antigens to be used as vaccines or for serodiagnosis, we focused on Leishmania virulence factors. Indeed, several studies described these factors as potentially immunogenic in humans, mice, and, more recently, dogs (6,8,9,13). Here, cellular and humoral immune responses of healed CL (hCL) and Mediterranean visceral leishmaniasis (MVL) patients were evaluated against results for Leishmania major protein disulfide isomerase (LmPDI) and mitogen-activated protein kinase kinase (MAPKK), which we and others previously described as potential virulence factors (2, 10).We produced LmPDI (55 kDa) and MAPKK (40 kDa) in the prokaryotic expression pET system (Novagen, Gibbstown, NJ) and then purified them by affinity chromatography (Fig. 1). Lymphoproliferative responses (Fig. 2) and gamma interferon (IFN-␥) and interleukin-10 (IL-10) production (Fig. 3) induced by recombinant LmPDI and MAPKK (10 g/ml) were characterized in vitro using peripheral blood mononuclear cells (PBMC) from two groups. The first group consisted of 18 hCL patients (age range, 17 to 42 years; mean Ϯ standard deviation [SD], 28 Ϯ 10.9 years) living in an area of L. major infection endemicity (the governorate of Sidi Bouzid, Tunisia). Diagnosis of leishmaniasis was based on the presence o...