Optimisation and comparison of three PCR procedures for molecular identification of Taenia solium

Abstract

Abstract The aim of the study was to optimise selected PCR methods for identification of T. solium, and to compare their effectiveness and usefulness. The investigation concerned three PCR methods described earlier: PCR I (specific to oncospherespecific protein Tso31 gene), PCR II (specific to large subunit rRNA gene), and PCR III (cytochrome c oxidases ubunit 1 gene). Each of them needed optimisation in connection with some changes in the procedures. Among the examined procedures, PCR I was found to be the most useful, requiring the least corrections during optimisation - only a higher concentration of polymerase was necessary. Testing an optimised PCR II method showed strong unspecific reactions with E. granulosus and T. saginata. This method was not considered diagnostically useful in distinguishing T. solium. PCR III method yielded products only when annealing temperature was lowered by 2 C. Under such conditions, there were no unspecific reactions with three others Taenidae parasites; however, annealing at a temperature only 1oC lower generated a distinct unspecific PCR product from T. saginata DNA. Therefore, this method was of limited usefulness. Comparison of the effectiveness of the two selected methods (PCR I and III) in detection of T. solium in successive DNA dilutions showed a large difference between them: in the same DNA sample, PCR I showed positive results in a sample diluted 1:3200, while PCR III failed at dilutions greater than 1:50. The results showed that among the three different methods used in the investigations, the most specific and effective for identification of T. solium was PCR I.