simultaneous detection of chlamydia pneulvloniae and mycoplasma pneumoniae by pcr

both c. pneumoniae and m. pneumoniae are common causes of respiratory tract infection. at present, both are still diagnosed in the laboratory retrospectively by serology. this is despite many publications which indicate that pcr, which is not retrospective, is extremely good at detecting these organisms. we thought that a single pcr test which could detect both organisms simultaneously in a routine diagnostic laboratory would be more economic than using two separate pcr tests. chlamydia pcr was developed and optimized to detect c. pneumoniae using primer cpna and cpnb which targets the momp gene. this test was very sensitive and could detect 10 organisms. to detect m. pneumoniae, the tu!pcr reported by luneberg6 was selected. after optimization of the duplex test, it was found that the sensitivity of the test for mycoplasma pcr was 100 times less than the sensitivity of the single tests, due to the inhibitory role of c. pneumoniae primer cpnb. reducing the concentration of this primer helped but we decided to redesign it instead. the final form of the duplex has sufficient sensitivity, detecting 10 copies of each organism. the new primer cpnb2 was a great improvement. the test was then developed to detect the product by hybridization rather than analysis with agarose gel electrophoresis.