Detecting Fasciola hepatica and Fasciola gigantica microRNAs with loop-mediated isothermal amplification (LAMP).

Abstract

Fascioliasis is a parasitic infection typically caused by two common parasites of class Trematodo, genus Fasciola, namely Fasciola hepatica and Fasciola gigantica. The widespread of these species in water and food makes fascioliasis become a global zoonotic disease that affects 2.4 million people in more than 75 countries worldwide. Typically, F. hepatica and F. gigantica can be recognized by parasitological techniques to detect Fasciola spp. eggs, immunological techniques to detect worm-specific antibodies, or by molecular techniques such as PCR to detect parasitic genomic DNA. Recently, miRNAs have been raised as a key regulator and potential diagnostic biomarkers of diseases, including parasitic infection. An isothermal PCR called loop-mediated isothermal amplification (LAMP) is rapid, sensitive, and its amplification process is so extensive that making LAMP well-suited for field diagnostics. LAMP reactions for miRNA detection have been introduced and were able to detect the target miRNA amounts in the wide range of 1.0amol to 1.0pmol, exhibiting high selectivity to differentiate one-base between miRNA sequences. Here, we introduced a modified LAMP to detect a species-specific miRNA of F. hepatica and F. gigantica. Our method did not demand an initial heating step and the reactions had a high sensitivity that greater than 1000 times in comparison to that reported in previous studies. Most importantly, the technique could perform well with parasitic miRNA presenting in bovine serum samples without sophisticated equipment required. These results create a promising technique basis for some novel and simple device to diagnose fascioliasis and other parasitic infection diseases at point-of-care.