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Research articles

ScienceAsia 45 (2019): 268-274 |doi: 10.2306/scienceasia1513-1874.2019.45.268

Rapid and sensitive detection of Nosema bombycis using loop-mediated isothermal amplification and colorimetric nanogold

Weijiang Daia, Jingru Qia, Hongli Chena, Zhilin Zhanga, Ruisha Shanga, Yiling Zhanga,b,c, Shunming Tanga,b,c, Zhongyuan Shena,b,c,*

ABSTRACT:     Salt-induced self-aggregation of gold nanoparticles (AuNP) carrying the ssDNA probes can be prevented specifically by complementary DNA. Loop-mediated isothermal amplification (LAMP) can amplify DNA rapidly. In the present study, we established a LAMP-AuNP method for detection of the microsporidium Nosema bombycis by combining the two techniques. The DNA templates of microsporidia were extracted by FTA card. The amplified product can be visually detected via hybridization at 63 °C for 5 min with a ssDNA-labelled nanogold probe, followed by salt-induced AuNP aggregation. The LAMP-AuNP assay needs approximately 65 min. This method can detect 10 spores/ml and is specific for N. bombycis. When it was used to detect pebrine disease of the silkworm eggs, the maximum sample size for silkworm eggs was 700, in which only one egg was infected with N. bombycis. The new LAMP-AuNP is a simple, sensitive and specific detection assay for N. bombycis.

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a Jiangsu University of Science and Technology, Zhenjiang 212018, Jiangsu Province, China
b Sericultural Research Institute, Chinese Academy of Agricultural Sciences, Zhenjiang 212018, Jiangsu Province, China
c Key Laboratory of Genetic Improvement of Silkworm and Mulberry of Agricultural Ministry, Zhenjiang 212018, Jiangsu Province, China

* Corresponding author, E-mail: szysri@163.com

Received 24 Feb 2019, Accepted 30 Jun 2019