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We have identified a novel 75 kbp large extrachromosomal DNA (LED) which is stably maintained during developmental conversion of Trypanosoma cruzi. It has a covalently closed circular conformation and is not derived from the kinetoplast network. In all T. cruzi strains analysed, LED contains 18S rRNA and spliced leader (sl) sequences. LED from the T. cruzi Y strain contains a minimum of 15 copies of the sl repeat arrayed in a head-to-tail configuration and 50 copies of a 196 bp repeat. LED is also present in Trypanosoma dionisii (subgenus Schizotrypanosoma) and in other members of the family Trypanosomatidae. LED from different T. cruzi strains and from other members of the Trypanosomatidae differ in their content of large ribosomal subunit rRNA sequences and the 196 bp repeat. The presence of LED in four evolutionarily distant trypanosomatid species suggests that it plays an important role in the biology of these parasites. [ABSTRACT FROM AUTHOR]