Leishmania are evolutionarily ancient protozoans (Kinetoplastidae) and important human pathogens that cause a spectrum of diseases ranging from the asymptomatic to the lethal. The Leishmania genome is relatively small [Ϸ34 megabases (Mb)], lacks substantial repetitive DNA, and is distributed among 36 chromosomes pairs ranging in size from 0.3 Mb to 2.5 Mb, making it a useful candidate for complete genome sequence determination. We report here the nucleotide sequence of the smallest chromosome, chr1. The sequence of chr1 has a 257-kilobase region that is densely packed with 79 protein-coding genes. This region is f lanked by telomeric and subtelomeric repetitive elements that vary in number and content among the chr1 homologs, resulting in an Ϸ27.5-kilobase size difference. Strikingly, the first 29 genes are all encoded on one DNA strand, whereas the remaining 50 genes are encoded on the opposite strand. Based on the gene density of chr1, we predict a total of Ϸ9,800 genes in Leishmania, of which 40% may encode unknown proteins.The Kinetoplastidae are flagellated protozoans found in terrestrial and aquatic environments that cause diseases in organisms ranging from plants to vertebrates. These diseases result in widespread human suffering and death, as well as considerable economic loss from infection of livestock, wildlife, and crops. In addition, kinetoplastids have been particularly valuable for the study of fundamental molecular and cellular phenomena, such as RNA editing (1), mRNA transsplicing (2), glycosylphosphatidylinositol-anchoring of proteins (3), antigenic variation (4), and telomere organization (5). The early evolutionary divergence of these organisms makes comparison of their sequences with those of other eukaryotes, as well as prokaryotes, useful for the identification of ancient conserved motifs, and their protein sequences may be a useful source of diversity for protein engineering.The numerous human-infective Leishmania spp. cause a spectrum of diseases with pathologies ranging from the asymptomatic to the lethal, and there are correlations between species and disease type and severity (6). The Leishmania haploid genome content is Ϸ34 megabases (Mb; ref. 7), consisting of 36 chromosomes ranging in size from 0.3 Mb to 2.5 Mb (8). It contains Ϸ30% repeated sequence (9), half of which is a series of telomeric hexamer repeats, whereas the remainder comprises other simple sequence repeats, transposons, as well as tandem and dispersed gene families such as rRNA, spliced-leader, tubulin, and gp63. The Leishmania molecular karyotype is conserved between Leishmania strains and species (10) with most genes syntenic among species (8). There are modest chromosome size polymorphisms between strains and larger size polymorphisms between species. Thus, this organism is an ideal candidate for a genome-sequencing project to elucidate its full genetic complement. The Leishmania Genome Network, established with the support of the World Health Organization, initiated a coordinated effort to map and sequenc...