Establishment of a Rapid Detection Method for Yeast-like Symbionts in Brown Planthopper Based on Droplet Digital PCR Technology.

The brown planthopper Nilaparvata lugens (Stål) (BPH) is a typical monophagous sucking rice pest. Over the course of their evolution, BPH and its symbionts have established an interdependent and mutually beneficial relationship, with the symbionts being important to the growth, development, reproduction, and variation in virulence of BPH. Yeast-like symbionts (YLS), harbored in the abdomen fat body cells of BPH, are vital to the growth and reproduction of the host. In recent research, the symbionts in BPH have mainly been detected using blood cell counting, PCR, real-time quantitative PCR, and other methods. These methods are vulnerable to external interference, cumbersome, time consuming and laborious. Droplet digital PCR (ddPCR) does not need a standard curve, can achieve absolute quantification, does not rely on Cq values, and is more useful for analyzing copy number variation, gene mutations, and relative gene expression. A rapid detection method for the YLS of BPH based on ddPCR was established and optimized in this study. The results showed that the method's limits of detection for the two species of YLS (Ascomycetes symbionts and Pichia guilliermondii) were 1.3 copies/µL and 1.2 copies/µL, respectively. The coefficient of variation of the sample repetition was less than 5%; therefore, the ddPCR method established in this study had good sensitivity, specificity, and repeatability. It can be used to detect the YLS of BPH rapidly and accurately.

Effects of Secondary Metabolites of Rice on Brown Planthopper and Its Symbionts.

The brown planthopper Nilaparvata lugens (Stål) (BPH) is a main rice pest in China and many other Asian countries. In the control of BPH, the application of insect-resistant rice has proven to be quite effective. Secondary metabolites are essential weapons in plants' defense against phytophagous insects. Studies have found that differences in the content of secondary metabolites play a crucial role in determining whether rice exhibits resistance or susceptibility to BPH. Simultaneously, symbionts are essential to the BPH. Nevertheless, there is limited research on the impact of secondary metabolites on the symbionts within BPH. Therefore, investigating the influence of secondary metabolites on both BPH and their symbionts is significant for the control of BPH. In this experiment, newly emerged female adults of BPH were fed artificial diets containing 10 different secondary metabolites. The results indicated that methyl jasmonate had inhibitory effects on the survival rate, weight gain, and reproductive capacity of BPH. Using qPCR methods, it was discovered that the number of symbiotic fungi (Ascomycetes symbionts) within BPH significantly decreased under methyl jasmonate stress. In conclusion, this experiment has preliminarily revealed the inhibitory effects of methyl jasmonate on BPH and its symbionts, demonstrating its potential for controlling BPH.

Detection of Yeast-like Symbionts in Brown Planthopper Reared on Different Resistant Rice Varieties Combining DGGE and Absolute Quantitative Real-Time PCR.

The brown planthopper (BPH), Nilaparvata lugens, is a serious pest of rice throughout Asia. Yeast-like symbionts (YLS) are endosymbionts closely linked with the development of BPH and the adapted mechanism of BPH virulence to resistant plants. In this study, we used semi-quantitative DGGE and absolute quantitative real-time PCR (qPCR) to quantify the number of the three YLS strains (Ascomycetes symbionts, Pichia-like symbionts, and Candida-like symbionts) that typically infect BPH in the nymphal stages and in newly emerged female adults. The quantities of each of the three YLS assessed increased in tandem with the developing nymphal instar stages, peaking at the fourth instar stage, and then declined significantly at the fifth instar stage. However, the amount of YLS present recovered sharply within the emerging adult females. Additionally, we estimated the quantities of YLS for up to eight generations after their inoculation onto resistant cultivars (Mudgo, ASD7, and RH) to reassociate the dynamics of YLS with the fitness of BPH. The minimum number of each YLS was detected in the second generation and gradually increased from the third generation with regard to resistant rice varieties. In addition, the Ascomycetes symbionts of YLS were found to be the most abundant of the three YLS strains tested for all of the development stages of BPH.