DNA Fingerprinting of the Human Intestinal Parasite Giardia intestinalis with Hypervariable Minisatellite Sequences

Abstract

Individual isolates of the Giardia duodenalis group of protozoan intestinal parasites were identified by DNA fingerprinting with hypervariable minisatellite sequences. A morphologically identical parasite is found in some forty different animal species. Although the species name intestinalis is reserved for the human isolates, electrophoretic karyotyping suggests that most duodenalis isolates fall into the same species grouping. Distinction based upon morphology, restriction endonuclease cleavage of genomic DNA or isoenzyme analysis has not been adequate to identify individual strains. The successful use of hypervariable sequences in the identification of individual human genomes encouraged us to examine the use of these same sequences for the possible identification of parasite isolates. We initially use as a fingerprinting probe the genome of the bacteriophage M13, which has repeated sequences recognising homologous hypervariable sequences in the human genome. The M13 probe recognises a weakly homologous set of hypervariable sequences in Giardia. The number of informative bands is comparable to those seen in mammals, since the lower molecular weight bands are also useful. There is considerable divergence in the sequences of individual Giardia minisatellites. Some cloned Giardia hypervariable sequences are more homologous to M13 than they are to each other. Similar results were observed with the hypervariable repeat sequences 3' to the human alpha-globin gene when they were used as a probe to distinguish Giardia isolates. The poly(dA-dC).poly(dG-dT) probe which recognises frequent TG tracts in a number of organisms also detects a few variable bands amidst a hybridisation background in the Giardia genome. Thus Giardia isolates which could not be distinguished by restriction endonuclease cleavage, antibody typing or isoenzyme analysis have been identified by DNA fingerprinting procedures. Detailed analysis of strain movement, resurgence, variation, host range and drug resistance is now possible. Similar families of sequences may be widespread in lower eukaryotes and useful for generating individual specific fingerprints. A procedure for detecting individual parasites is also presented. Since Giardia is regarded as the most ancient eukaryote before the occurrence of symbiosis with purple non-sulphur bacteria to generate mitochondria, the identification of hypervariable sequences in the Giardia genome should also aid in understanding the mechanism of generation and evolution of these sequences.