ELSEVIER Molecular and Biochemical Parasitology 69 (1995) 197-209 zyxwvutsrqponmlkjihgfedcbaZYXWV MOLECULAR i&EMICfiL PARASITOLOGY Genotypic polymorphisms in experimental metastatic dermal leishmaniasis Raquel S. Pacheco a, *, Julia Elvira Martinez b, Liliana Valderrama b, Hooman Momen a, Nancy G. Saravia b zyxwvutsrqponmlkjihgfedcbaZYXWVUTS a Department zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA of Biochemistry and Molecular Biology, Institute Oswald0 Crux, FIOCRUZ, Au. Brasil4365, Manguinhos 21045. Rio de Janeiro, Brazil b Centro International de Inuestigaciones Medicas, CIDEIM, Cali, Colombia Received 6 July 1994; revised 14 November 1994; accepted 22 November 1994 zyxwvutsrqponmlkjihgfedcbaZY Abstract Molecular karyotype and kDNA restriction analyses were utilized to examine the genetic heterogeneity and plasticity of the Leishmaniu (Viunniu) guyanensis strain WHI/BR/78/M5313, composed of metastatic and non-metastatic populations. Cloning revealed that the strain was constituted by multiple closely related populations that were distinguishable by restriction fragment polymorphisms in kDNA. Size polymorphisms in molecular karyotype were not detected. Passage of clones in hamsters and recovery of parasites from cutaneous metastatic lesions yielded evidence of further genetic heterogeneity among some of the progeny populations. Overall, six kDNA minicircle restriction patterns or schizodemes were observed among clones, subclones and progeny. Although the possibility that population heterogeneity was not resolved by cloning cannot be ruled out, subcloning and kDNA restriction analysis to determine whether the putative clones consisted of homogeneous populations showed the schizodeme of subclones of 3 out of 4 clones to be identical to the clone of origin, while a subclone of the fourth had a co-efficient of similarity of 0.95. Metastasis did not segregate with a particular schizodeme: all six restriction profiles were represented among populations isolated from metastatic lesions and some clones with the same restriction profile did not produce metastatic lesions. The strain from which the clones, subclones and progeny were derived had a kDNA restriction pattern identical to the most prevalent schizodeme (38%) among these subpopulations. This finding together with the reappearance of the repertoire of schizodemes found among clones in the populations recovered from metastatic lesions in hamsters inoculated with a single clone, suggest that sequence polymor- phisms in kDNA can emerge during infection. Keywords: Minicircle; Kinetoplast DNA; Schizodeme; Karyotype; Metastasis; Leishmania 1. Introduction Secondary manifestations of Leishmania infec- Abbreviations: kDNA, kinetoplast DNA; TAPE, transverse alternating field electrophoresis; RFLP, restriction fragment length polymorphism tion are among the most destructive and challenging disease presentations to manage, both clinically and epidemiologically [l-3]. In the Americas, metastasis * Corresponding author. and recurrent cutaneous and mucosal leishmaniasis 0166-6851/95/$09.50 0 1995 Elsevier Science B.V. All rights reserved SSDI 0166-6851(94)00212-6