Leishmania braziliensis: characterisation of a complex specific subtelomeric repeat sequence and its use in the detection of parasites.
A 1.6-kb tandem repeat sequence had previously been identified in the subtelomeric region of mini- and megabase chromosomes from Leishmania braziliensis. Southern hybridisation was used to demonstrate that the repeat is complex specific. The sequence was characterised in strains representing four species of the L. braziliensis complex. This data allowed an assessment of the evolutionary relationship of the four species. PCR primers targeted to the repeat amplify only DNA from species of the L. braziliensis complex. Titration assays indicate that a minimum of 50 fg of parasite DNA can be detected by PCR alone. Southern hybridisation increases the limit of detection to 5 fg. Interspecies variation in the repeat sequence enabled restriction enzyme digestion of PCR products to distinguish individual species within the L. braziliensis complex.
Fu
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MRC Outstation of NIMR, Molteno Laboratories, Department of Pathology, University of Cambridge, CB2 1QP, U.K.
Perona-Wright
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Barker
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D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D017124:Conserved Sequence; D017931:DNA Primers / Q000737:chemistry; D016054:DNA, Protozoan / Q000737:chemistry*; D004275:DNA, Ribosomal / Q000737:chemistry; D007892:Leishmania braziliensis / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D016773:Leishmaniasis, Cutaneous / Q000175:diagnosis*; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D015183:Restriction Mapping; D016415:Sequence Alignment; D013045:Species Specificity; D020080:Tandem Repeat Sequences / Q000235:genetics*; D016615:Telomere / Q000235:genetics*
D017931:DNA Primers; D016054:DNA, Protozoan; D004275:DNA, Ribosomal