Sensitive and rapid detection of Clonorchis sinensis infection in fish by loop-mediated isothermal amplification (LAMP)

Abstract

The fish-borne clonorchiasis caused by the oriental liver fluke Clonorchis sinensis is endemic in a number of countries with over 35 million people being infected globally. Rapid and accurate detection of C. sinensis in its intermediate host fish is important for the control and prevention of clonorchiasis in areas where the disease is endemic. In the present study, we established a loop-mediated isothermal amplification (LAMP) approach for the sensitive and rapid detection of C. sinensis metacercariae in fish. The specificity and sensitivity of primers designed from the C. sinensis cathepsins B3 gene were evaluated, and specific amplification products were obtained with C. sinensis, while no amplification products were detected with DNA of related trematodes, demonstrating the specificity of the assay. The LAMP assay was proved to be 100 times more sensitive than a conventional polymerase chain reaction for detection of C. sinensis. The established LAMP assay provides a useful tool for the rapid and sensitive detection of C. sinensis in fish, which has important implications for the effective control of human clonorchiasis.

Rapid Detection and Differentiation ofClonorchis sinensisandOpisthorchis viverriniUsing Real-Time PCR and High Resolution Melting Analysis

“…All parasite materials were preserved in 70% ethanol and kept at −20°C until the extraction of genomic DNA. Metacercariae of C. sinensis were collected from the fish muscles essentially according to a previously reported method [9]. Fecal samples were obtained from the residents in Guangdong Province, China.…”

“…A number of methods have been developed to detect these two species, such as direct microscopy, ELISA [7], conventional PCR [8], and loop-mediated isothermal amplification (LAMP) [9]. There are, however, many drawbacks of these methods.…”

Sensitive and rapid detection of Paragonimus westermani infection in humans and animals by loop-mediated isothermal amplification (LAMP)

“…Recently, this method has also been developed successfully for the diagnosis of parasitic infections such as malaria, trypanosomiasis, theileriosis, babesiosis, schistosomiasis, and clonorchiasis (Kuboki et al 2003;Poon et al 2006;Alhassan et al 2007;Iseki et al 2007;Thekisoe et al 2007;Xu et al 2009;Cai et al 2010).…”

Sequences and gene organization of the mitochondrial genomes of the liver flukes Opisthorchis viverrini and Clonorchis sinensis (Trematoda)

“…O. viverrini infection is confined to the liver bile duct, and gallbladder, and chronic infection is associated with many severe diseases such as cholangitis, cholecystitis, cholelithiasis, and hepatomegaly (Boonjaraspinyo et al 2010;Kiatsopit et al 2011). C. sinensis is endemic in many countries, including China, Korea, and North Vietnam (Lun et al 2005;Chen et al 2011), about 35 million people were estimated to be infected with C. sinensis globally, and approximately 15 million of these were in China (Lun et al 2005;Cai et al 2010). C. sinensis infection is often associated with many human diseases such as biliary calculi, cholecystitis, liver cirrhosis, and even cholangiocarcinoma (Pak et al 2009).…”