Development of reverse transcription loop-mediated isothermal amplification (RT-LAMP) as a diagnostic tool of Toxoplasma gondii in pork

Abstract

A fast, sensitive and specific reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for the detection of Toxoplasma gondii (T. gondii) in pork was developed. In this study, we used a conserved sequence of 18s rRNA of Toxoplasma gondii to design primers for RT-LAMP test. The amplication was able to finish in 60 min under isothermal condition at 63°C by employing a set of six primers. The assay showed higher sensitivity than RT-PCR using T. gondii RNA as template. The RT-LAMP assay was also assessed for specificity and was found to precisely discriminate between positive and negative test samples. Furthermore, the assay correctly detected T. gondii from contaminated pork, and had the detect limit of 1 tachyzoite in 1g pork. This is the first report of a study which applied the RT-LAMP method to detect T. gondii from pork. As RT-LAMP requires very basic instruments and the results can be obtained by visual observation, this technique provides a simple and reliable tool for inspecting food which are T. gondii-contaminated.