Abstract:

Objective To establish a loop-mediated isothermal amplification(LAMP) assay for rapid detection of Echinococcus granulosus cytochrome c oxidase subunit 2(cox2) gene in dog feces. Methods Four primers were designed based on the cox2 gene of Echinococcus granulosus and were used for the LAMP assay. LAMP assay and conventional PCR were performed to detect E. granulosus, Cysticercus tenuicollis, Moniezia expansa, Thysaniezia ovilla, Avitellina centripunctata, flagella nematode, Fasciola hepatica, E. multilocularis and Haemonchus contortus, and the specificity of LAMP was evaluated. The sensitivity of LAMP was compared with that of conventional PCR using gradient dilutions of recombinant plasmid carrying cox2 fragment of E. granulosus. In addition, LAMP, conventional PCR and ELISA were performed to detect E. granulosus in 50 dog fecal samples. The positive rate of E. granulosus was calculated and the efficacy of LAMP was evaluated. Results The LAMP assay only produced specific bands for E. granulosus among various species. The sensitivity of LAMP for E. granulosus was 16.09 ag/μl, which was 1 000 times higher than that of conventional PCR(16.9 fg/μl). Regarding the fecal samples, the three methods revealed same positive rate of 8.0%(4/50). Conclusion The LAMP assay is a convenient method to detect E. granulosus based on the cox2 gene with a high specificity and sensitivity, revealing a suitable technique for rapid detection of E. granulosus in dogs.

Key words: Echinococcus granulosus, Cytochrome c oxidase subunit 2, LAMP