Abstract
In the present work, loop-mediated isothermal amplification (LAMP) and hyperbranched rolling circle amplification (HRCA) methods were developed to detect and distinguish different lethal Amanita species. Specific LAMP primers and HRCA padlock probes for species-specific identification and a set of universal LAMP primers for lethal Amanita species were designed and tested. The results indicated that the LAMP-based assay was able to discriminate introclade lethal Amanita species but was not able to discriminate intraclade species perfectly, while the HRCA-based assay could discriminate whether introclade or intraclade species. The universal LAMP primers were positive for 10 lethal species of Amanita section Phalloideae and negative for 16 species of Amanita outside section Phalloideae. The detection limits of LMAP and HRCA were 10 and 1 pg of genomic DNA per reaction, respectively. In conclusion, the two methods could be rapid, specific, sensitive and low-cost tools for the identification of lethal Amanita species.
Highlights
Mushroom poisoning is the main cause of mortality in food poisoning incidents in China
The containing substances of various peptide toxins were another critical characteristics of lethal amanitas and the peptide toxins in Amanita can be divided into three major groups, including amatoxins, phallotoxins and virotoxins, which are bicyclic octapeptides, bicyclic heptapeptides and Isothermal Amplification Detection of Amanita monocyclic heptapeptides, respectively (Wieland, 1986)
Some lethal Amanita species are similar to the edible species of section Caesareae Singer, for example, A. chepangiana vs. A. exitialis and A. hemibapha vs. A. subjunquillea, this is the main reason for mistaken collection and ingestion
Summary
Mushroom poisoning is the main cause of mortality in food poisoning incidents in China. More than 90% of fatal mushroom poisoning cases were caused by mistaken ingestion of lethal amanitas in Europe, North America and East Asia (Enjalbert et al, 2002; Chen et al, 2014). There have been approximately 50 lethal Amanita species reported worldwide (Cai et al, 2016). These lethal Amanita species have four common morphologic characteristics as bases distinguished from other taxa of Amanita, including a non-appendiculate pileus, the persistent presence of an annulus, a bulbous stipe base with a limate volva and amyloid basidiospores (Cai et al, 2014). Some lethal Amanita species are similar to the edible species of section Caesareae Singer, for example, A. chepangiana (edible) vs. A. exitialis (lethal) and A. hemibapha (edible) vs. A. subjunquillea (lethal), this is the main reason for mistaken collection and ingestion
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