Open AccessRapid and Visual Detection of Meloidogyne hapla Using Recombinase Polymerase Amplification Combined with a Lateral Flow Dipstick Assay
Author(s) -
Zhiqiang Song,
Xiai Yang,
Xiaowei Zhang,
Mingbao Luan,
Bing Guo,
Chunjie Liu,
Jiangpeng Pan,
Shuqi Mei
Publication year - 2021
Publication title -
plant disease
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.663
H-Index - 108
eISSN - 1943-7692
pISSN - 0191-2917
DOI - 10.1094/pdis-06-20-1345-re
Subject(s) - recombinase polymerase amplification , biology , dipstick , amplicon , polymerase chain reaction , microbiology and biotechnology , virology , gene , genetics , anatomy , urinary system
The northern root-knot nematode, Meloidogyne hapla, is a biotrophic parasite that infects many crops and causes severe economic losses worldwide. Rapid and accurate detection of M. hapla is crucial for disease forecasting and control. We developed a recombinase polymerase amplification combined with a lateral flow dipstick (RPA-LFD) assay for rapid detection of M. hapla. The primers and probe were designed based on the effector gene 16D10 sequence and were highly specific to M. hapla. The RPA reaction was performed at a wide range of temperatures from 25 to 45°C within 5 to 25 min, and the amplicon was visualized directly on the LFD within 5 min. The detection limits of the RPA-LFD assay were 10 −3 females and 10 −2 second-stage juveniles/0.5 g of soil, which was 10 times more sensitive than the conventional PCR assay. In addition, the RPA-LFD assay can detect M. hapla from infested plant roots and soil samples, and the entire detection process can be completed within 1.5 h. These results indicate that the RPA-LFD assay is a simple, rapid, specific, sensitive, and visual method that can be used for rapid detection of M. hapla in the field and in resource-limited conditions.