Trypanosome identification in wild tsetse populations in Tanzania using generic primers to amplify the ribosomal RNA ITS-1 region.
Tsetse flies transmit many species of trypanosomes in Africa, some of which are human and livestock pathogens of major medical and socio-economic impact. Identification of trypanosomes is essential to assess the disease risk posed by particular tsetse populations. We have developed a single generic PCR test to replace the multiple species-specific PCR tests used previously to identify the trypanosome species carried by individual tsetse flies. In the generic PCR test, inter-species size variation in the PCR product of the internal transcribed spacer (ITS-1) region of the ribosomal RNA repeat region enables species identification. The test was applied to identify trypanosomes in midgut samples stored on FTA cards from wild-caught flies in two regions of Tanzania. Identifications were verified by sequencing the amplified ITS-1 region and/or species-specific PCR tests. The method facilitated the identification of large numbers of field samples quickly and accurately. Whereas species-specific tests are incapable of recognising previously unknown species, the generic test enabled a new species to be identified by the unique size of the amplified product. Thus, even without access to any isolate of this new species, we could collect data on its distribution, prevalence and co-occurrence with other trypanosomes. The combined molecular and ecological profiles should facilitate the isolation and full biological characterization of this species in the future.
Adams
E R
ER
School of Biological Sciences, University of Bristol, Bristol, UK.
Malele
I I
II
Msangi
A R
AR
Gibson
W C
WC
D000818:Animals; D017931:DNA Primers; D016054:DNA, Protozoan / Q000032:analysis; D021903:DNA, Ribosomal Spacer / Q000032:analysis*; D012335:RNA, Ribosomal / Q000235:genetics; D012680:Sensitivity and Specificity; D013636:Tanzania; D014345:Trypanosoma / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D014370:Tsetse Flies / Q000469:parasitology*
D017931:DNA Primers; D016054:DNA, Protozoan; D021903:DNA, Ribosomal Spacer; D012335:RNA, Ribosomal