Clonal heterogeneity in populations of Leishmania major

Forty clones of Leishmania major were derived by direct plating from lesions and from cultures of recent isolates, followed by plating as well as by additional limit dilution in some. The parental strains originated from and represented three geographical areas in each of which a distinct electrophoretic type of the enzyme 6-phosphogluconate dehydrogenase (6PGD) is found. Each clone was characterized in terms of its virulence in Sabra mice, morphology, serotype, enzyme electrophoretic profile based on 11 enzymes, and kinetoplast DNA endonuclease restriction pattern. All the clones showed a similar virulence in the mice and similar polymorphism in fresh drops of living cultures and in Giemsa-stained preparations. Ten clones from the Jordan Valley were all serotype A1, 15 clones from the Arava were all A1Bz--confirming the mixed serological nature of parasites from this region, 11 of the 20 clones from the western Negev-eastern Sinai included a new A subserotype, and 9 were A1. Generally, the geographical separation of the 6PGD variants was upheld. Two of the Jordan Valley clones were similar to those of the western Negev. On two occasions, different enzyme types were isolated from the same individual, in one case from the same lesion. All clones from the western Negev were alike and similar to the type previously found in that region. The situation in the Arava was more complicated, since clones representative of all three regional types were found. In cases where the first two types were found, the clones and their parental strains were similar. However, one parental culture yielded three clones that resembled the reference strain for the Jordan Valley and two which resembled that of the western Negev. In one of these cases, two dissimilar clones originated from the same lesion. One aberrant clone from the western Negev was variant for nucleoside hydrolase. The kinetoplast DNA of each clone was distinct but relatively complicated, which made differentiation of these regional types difficult. However, clones from the same area displayed more similarity to each other than to those of other regions, implying a closer genetic relationship.