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Detection of mutant P2 adenosine transporter (TbAT1) gene in Trypanosoma brucei gambiense isolates from northwest Uganda using allele‐specific polymerase chain reaction
Author(s) -
Nerima Barbara,
Matovu Enock,
Lubega George W.,
Enyaru John C. K.
Publication year - 2007
Publication title -
tropical medicine and international health
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.056
H-Index - 114
eISSN - 1365-3156
pISSN - 1360-2276
DOI - 10.1111/j.1365-3156.2007.01918.x
Subject(s) - biology , trypanosoma brucei , restriction fragment length polymorphism , polymerase chain reaction , variants of pcr , microbiology and biotechnology , mutant , allele , gene , virology , genetics
Summary Objective To assess the application of allele‐specific PCR (AS‐PCR) as a fast, cheap and reliable method for detecting mutant TbAT1 associated with melarsoprol relapse in Trypanosoma brucei gambiense isolates from northwest Uganda. Methods A total of 105 trypanosome isolates were analysed using Sfa N1 restriction fragment length polymorphism (RFLP) and AS‐PCR, the former used as the gold standard. Sensitivity, specificity, positive and negative predictive values of AS‐PCR as well as agreement between the tests were determined. Results Eleven trypanosome isolates had mutant TbAT1 while 94 exhibited the wild‐type TbAT1 genes. There was a highly significant agreement between Sfa N1 RFLP and AS‐PCR with kappa and intra‐class correlation values of 1.0. The sensitivity and specificity of AS‐PCR were both 100%, while the positive and negative predictive values were found to be equal to 1.0. Cost and time analyses were performed and AS‐PCR was 4.3 times cheaper than Sfa N1 RFLP, in addition to the less time required for its execution. Conclusion AS‐PCR should be the test of choice for screening for mutant TbAT1 in the ever‐increasing numbers of field trypanosome isolates.