Development of a loop-mediated isothermal amplification method for the rapid detection of Venturia carpophila on peach.

Abstract

Peach scab, caused by Venturia carpophila, can significantly reduce both the yield and quality of peach fruit. It is difficult to distinguish peach scab from other peach diseases such as black spot and bacterial spot. An efficient assay is needed to identify V. carpophila in order to develop scientific management strategies. A set of loop-mediated isothermal amplification (LAMP) primers was designed based on the internal transcribed spacer (rDNA-ITS) sequence to detect V. carpophila. Compared with the conventional polymerase chain reaction (PCR) method, the LAMP method not only exhibited higher sensitivity and specificity in the detection of V. carpophila, but also required simpler equipment and less operational time. The minimum detectable concentration of V. carpophila genomic DNA with the LAMP method was 56.6 fg μl-1 , which was 100 times lower than with the conventional PCR method. When eight fungal species including V. carpophila (23 isolates from 14 provinces) and one bacterial species were used with LAMP detection, only V. carpophila showed a color change, from brown to yellowish green, and ladder-like bands in electrophoresis, indicating successful amplification. Moreover, when crude DNAs of peach fruit samples were used in LAMP detection, amplification was observed only from diseased fruits, and not from healthy fruits and the negative control. The LAMP assay shows simplicity, rapidity, high sensitivity and specificity, and will be useful in distinguishing scab caused by Venturia carpophila from other diseases with similar symptoms. © 2020 Society of Chemical Industry.