Sensitive detection of trypanosomes in tsetse flies by DNA amplification.
African trypanosome species were identified using the Polymerase Chain Reaction (PCR) by targeting repetitive DNA for amplification. Using oligonucleotide primers designed to anneal specifically to the satellite DNA monomer of each species/subgroup, we were able to accurately identify Trypanosoma simiae, three subgroups of T. congolense, T. brucei and T. vivax. The assay was sensitive and specific, detecting one trypanosome unequivocally and showing no reaction with non-target trypanosome DNA or a huge excess of host DNA. The assay was used to identify developmental stage trypanosomes in the tsetse fly. The use of radioisotopes was not necessary and mixed infections could be detected easily by incorporating more than one set of primers in a single reaction. The use of crude preparations of template made the process very rapid. The methodology should be suitable for large-scale epidemiological studies.
Masiga
D K
DK
Department of Pathology and Microbiology, School of Veterinary Science, University of Bristol, Langford, U.K.
Smyth
A J
AJ
Hayes
P
P
Bromidge
T J
TJ
Gibson
W C
WC
D000818:Animals; D001483:Base Sequence; D016054:DNA, Protozoan / Q000032:analysis* / Q000737:chemistry; D005784:Gene Amplification; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012091:Repetitive Sequences, Nucleic Acid; D012689:Sequence Homology, Nucleic Acid; D014345:Trypanosoma / Q000235:genetics / Q000302:isolation & purification*; D014370:Tsetse Flies / Q000469:parasitology*
D016054:DNA, Protozoan