Protective immunity againstH uman infection with Leishmania chagasi, the protozoan causing South American visceral leishmaniasis, causes diverse sequelae ranging from subclinical infection to progressive fatal disease (1). Subclinical infection results in the development of cellular immune responses detected by a positive delayed-type hypersensitivity skin test. These immune responses are presumed to protect against subsequent disease (2). One would assume that "immunization" by inducing a subclinical infection with parasites that are rendered avirulent by virtue of attenuation or knockout of a conditionally essential gene might mimic this naturally acquired protective immunity. The purpose of this study was to determine whether subclinical infection with virulent or avirulent strains of L. chagasi also induces protective immunity in a susceptible mouse model.Murine models of infection with an agent of cutaneous leishmaniasis, Leishmania major, have provided important information on the development of protective vs permissive immune responses toward this pathogen. Expansion of a Th1 subset of CD4 ϩ lymphocytes that secretes IFN-␥ and IL-2 is associated with resistance to infection (3-5). IL-12, likely secreted by APCs, enhances expansion of Th1 cells through a pathway involving NK cells (6 -8). Susceptible mice instead expand CD4 ϩ lymphocytes belonging to the Th2 subset that secretes IL-4, IL-10, and IL-13 (3-5). Early IL-4 secretion by an as yet undefined population of cells plays a role in differentiation of T cells toward the Th2 phenotype and suppression of Th1 subset development (9, 10). Several laboratories have shown that, in contrast to murine L. major infection, permissive murine Leishmania donovani or L. chagasi infection results from failure to expand an Ag-specific Th1 subset of CD4 ϩ cells in the absence of a detectable Th2 response (11,12).Genetically susceptible strains of mice can mount a protective immune response against L. major infection after a variety of experimental manipulations that deplete the animal of disease-exacerbating CD4 ϩ cells. Protective immunity can also be induced by immunization of mice with parasite Ags combined with IL-12 (13), by cure of prior active infection (14), or by immunization with Leishmania Ags in the form of recombinant proteins (15)(16)(17)(18)(19) or expressed in DNA vaccines (20,21). In addition, expression of Leishmania Ags in recombinant organisms, such as bacillus Calmette-Guérin, Salmonella, and vaccinia virus, can lead to partial protection against infectious challenge (22)(23)(24). Although the use of attenuated Leishmania for immunization runs the risk of inoculating infectious organisms into the host, a novel means of immunization uses Leishmania that are rendered avirulent by deletion of genes encoding enzymes essential for the metabolism of the parasite (e.g., dihydrofolate-thymidylate synthase gene locus (DHFR-TS) 3 ). Such "genetically attenuated" parasites were

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The genomic sequences of 20 Leishmania infantum isolates collected in northeastern Brazil were compared with each other and with the available genomic sequences of 29 L. infantum/donovani isolates from Nepal and Turkey. The Brazilian isolates were obtained in the early 1990s or since 2009 from patients with visceral or non-ulcerating cutaneous leishmaniasis, asymptomatic humans, or dogs with visceral leishmaniasis. Two isolates were from the blood and bone marrow of the same visceral leishmaniasis patient. All 20 genomic sequences display 99.95% identity with each other and slightly less identity with a reference L. infantum genome from a Spanish isolate. Despite the high identity, analysis of individual differences among the 32 million base pair genomes showed sufficient variation to allow the isolates to be clustered based on the primary sequence. A major source of variation detected was in chromosome somy, with only four of the 36 chromosomes being predominantly disomic in all 49 isolates examined. In contrast, chromosome 31 was predominantly tetrasomic/pentasomic, consistent with its regions of synteny on two different disomic chromosomes of Trypanosoma brucei. In the Brazilian isolates, evidence for recombination was detected in 27 of the 36 chromosomes. Clustering analyses suggested two populations, in which two of the five older isolates from the 1990s clustered with a majority of recent isolates. Overall the analyses do not suggest individual sequence variants account for differences in clinical outcome or adaptation to different hosts. For the first known time, DNA of isolates from asymptomatic subjects were sequenced. Of interest, these displayed lower diversity than isolates from symptomatic subjects, an observation that deserves further investigation with additional isolates from asymptomatic subjects.