Development of new gene diagnosis by detecting NTPase gene of Toxoplasma gondii.

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We tried to amplify a gene fragment (about 800 base pair) in NTPase, which is present in tachyzoite cells causing acute toxoplasmosis, gene by PCR and RT-PCR using DNA and RNA extracted from tissues of animals infected with Toxoplasma gondii and suspected human patient for toxoplasmosis as the tempelates. The results indicated that it was rigidly possible to detect the gene and mRNA of NTPase from the tissues of animal infected with T.gondii. Then, we tried to detect the gene for diagnosis of a suspected human patient as congenital toxoplasmosis. We succeeded to detect the mRNA but not to detect the DNA suggesting that RT-PCR was more sensitive than simple PCR.Anyway, we succeeded to diagnose congenital toxoplasmosis within a short period. Preliminal experiment indicated that only one tachyzoite cell was detected by RT-PCR and the sensitivity of RT-PCR was 10 times higher than that of PCR.The reason why RT-PCR is more sinsitive than PCR is that tachyzoite cells contain lot of mRNA and copy number of NTPase is abundant. In the present study, we have succeeded to detect NTPase mRNA by RT-PCR indicating that RT-PCR is useful for diagnosis of acute toxoplasmosis in general. Comparing with previous diagnostic methods for acute toxoplasmosis, which took long time to complete, this RT-PCR and PCR methods have a merit being practicable to diagnose acute toxoplasosis at once.