Detection of CD4+CD45RO+ T lymphocytes producing IL-4 in response to antigens on Plasmodium falciparum erythrocytes: an in vitro correlate of protective immunity induced with attenuated Plasmodium falciparum sporozoites.

Malaria is caused by Plasmodium spp. and is one of the major infectious diseases leading to morbidity and mortality in tropical areas of the world. The model of protective immunity induced by immunization with radiation-attenuated Plasmodia sporozoites (SPZ) has become the framework for the elucidation of protective immune mechanisms and the prototype for a promising vaccine strategy. We have previously reported that although considered stage specific based on antibody and CD8+ cytolytic T lymphocyte responses directed against preerythrocytic stage antigens, in particular, the circumsporozoite protein and sporozoite surface protein 2, protective immunity induced in humans by attenuated Plasmodium falciparum SPZ may also involve CD4+ T cell responding to antigens present on parasitized red blood cells (pRBC). In this study we examined the functional role of pRBC responding CD4+ T cells by comparing in vitro pRBC-stimulated responses of CD4+ T cells from persons during preimmunity to irradiated SPZ, during induction of protection, and infection induced with SPZ. The results reported herein corroborate previously published observations that antigens associated with pRBC induce proliferative CD4+ lymphocytes responses in subjects exposed to malaria parasite-derived antigens and not malaria-naive persons; however, now we demonstrate that pRBC-proliferative CD4+ T cells did not coincide with protective immunity. Similarly, pRBC-induced IFN-gamma levels did not distinguish malaria protected from susceptible persons, although IFN-gamma was observed only in lymphocyte cultures from malaria parasite-exposed volunteers and not in lymphocyte cultures from malaria-naive persons. In contrast, we noted an increase in the IL-4-producing CD4+ T cells that also exhibited the memory phenotype, CD45RO, and an upregulated expression of CD25 in cultures from malaria protected persons as compared to malaria naive persons and subjects who became parasitemic. Hence, these observations suggest that the induction of memory CD4+ T cell subset distinguished by the expression of CD45RO and CD25 and production of IL-4 coincides with protective immune responses generated by immunization with attenuated SPZ.

Rapid detection of Leishmania amastigotes in fluid aspirates and biopsies of human tissues.

When using a genus-specific monoclonal antibody (83-J3D2) as the primary reagent in an indirect immunofluorescent antibody assay (IFA), intracellular amastigotes of Leishmania were easily identified in 9 of 9 biopsies and in 11 of 12 needle aspirates taken from human lesions. In contrast, only 5 of the biopsies and 4 of the aspirates yielded promastigotes upon culture in vitro. Similarly, all but 2 of the aspirates and one-half of the biopsies were reported as negative for parasites when stained with Wright's and hematoxylin-eosin, respectively. Serum antibody titers, ranging from 1:8 to 1:128, corroborated the results of the amastigote detection assays when histopathology and isolation were negative. These findings support the practicality of using the genus-specific monoclonal IFA in those field situations where it becomes necessary to differentiate leishmaniasis from other skin infections.