Development of a rapid assay to detect the jellyfish Cyanea nozakii using a loop-mediated isothermal amplification method

Abstract

Blooms of the harmful jellyfish Cyanea nozakii, which are a severe nuisance to fisheries and tourisms, frequently occur in the northern East China Sea, Yellow Sea, and Bohai Sea. To provide early warning of this species, a simple and effective molecular method for identifying C. nozakii was developed using the loop-mediated isothermal amplification method (LAMP). The LAMP assay is highly specific and uses a set of four primers that target six different regions on the mitochondrial cytochrome c oxidase subunit I (COI) gene of C. nozakii. The amplification conditions, including the dNTP and betaine concentrations, the inner primer to outer primer concentration ratio, reaction time and temperature, were optimized. The LAMP assay amplified DNA extracted from tissue samples of C. nozakii but did not amplify DNA from other common scyphozoans and hydrozoans collected in the same region. In addition, the LAMP assay was more sensitive than conventional PCR. Therefore, the established LAMP assay is a sensitive, specific, fast, and easily performed method for detection of C. nozakii at different stages in their life cycle.

An isothermal DNA amplification method for detection of Onchocerca volvulus infection in skin biopsies

“…Since every cell contains hundreds of mitochondrial DNA copies, it makes mitochondrial encoded sequences ideal targets for molecular detection requiring high sensitivity, as is the case for O. volvulus detection in skin biopsies [41]. For this reason, the cox 1 gene has already been used as a target for other molecular assays to detect the presence of the jellyfish Cyanea nozakii in seawater samples or the presence of the olive tree pest Bactrocera oleae in the guts of arthropods [42, 43]. It will be of interest for future work to study whether a correlation exists between number of mf as determined by microscopy and time to LAMP.…”

Evaluation of the complete nuclear rDNA unit sequence of the jellyfish Cyanea nozakii Kishinouye (Scyphozoa: Semaeostomeae) for molecular discrimination

“…The BLAST search of the 18S region showed that the C. nozakii in the present study was 100% identical to C. nozakii (KT445894) collected from Chinese coastal waters. As noted previously, C. nozakii is widely distributed around the coasts of China, Korea, and Japan (Dong et al 2008 ; Hong and Lin 2010 ; Liu et al 2016 ). Therefore, 18S and 28S rDNA can be utilized to easily detect and monitor this harmful organism in the marine environment.…”

“…The jellyfish Cyanea nozakii Kishinouye 1891 belongs to the family Cyaneidae and is commonly known as the ghost jellyfish. The species is widely distributed around the coasts of China, Korea, and Japan (Dong et al 2008 ; Hong and Lin 2010 ; Liu et al 2016 ). It can be industrially used as a source of collagen for immune-enhancing activity (Tang et al 2013 ; Zhang et al 2014 ).…”

“…For example, within Semaeostomeae, morphology has inconsistently resolved relationships among families (Cyaneidae, Pelagiidae, and Ulmaridae), within family Pelagiidae, and within genera such as Aurelia and Cyanea (Mayer 1910 ; Kramp 1961 ; Dawson 2003 ). In particular, the genus Cyanea contains 18 species (WoRMS 2021 ) that are morphologically similar; specifically, the larval and juvenile stages of C. nozakii vary in size and shape; hence their morphological identification is difficult and time-consuming (Dong et al 2008 ; Liu et al 2016 ). For example, the planulae of C. nozakii varies from slipper-shaped to irregularly oval, measuring 90-180 mm in length and 60-95 mm in width, and the benthic scyphistomae varies in size and shape (Dong et al 2006 ).…”

“…In recent years, molecular investigations have reduced systematic instability by resolving inconsistencies of some morphological hypotheses at various nodes throughout the scyphozoan tree of life (Collins et al 2006 ; Bayha et al 2010 ; Chae et al 2018 ). In addition, it offers more accurate identifications of cryptic taxa (Dawson 2003 ; Liu et al 2016 ; Jang et al 2020 ). Among the molecular methods, sequencing of ribosomal DNA is quite effective in exploring discrimination, phylogenies, and elucidating evolutionary inferences of jellyfishes (Ki et al 2008 ; Chae et al 2018 ).…”