Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, College of Modern Science and Technology, China Jiliang University, Hangzhou 310018, China.
Int J Mol Sci. 2023 Jul 4;24(13):11071. doi: 10.3390/ijms241311071.
The brown planthopper (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 ( symbionts and ) 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.
褐飞虱（Stål）（BPH）是一种典型的单食性吸食水稻害虫。在进化过程中，BPH 及其共生菌建立了相互依存、互利共生的关系，共生菌对 BPH 的生长、发育、繁殖和毒力变异至关重要。酵母样共生菌（YLS）存在于 BPH 的腹部脂肪体细胞中，对宿主的生长和繁殖至关重要。在最近的研究中，主要使用血细胞计数、PCR、实时定量 PCR 等方法检测 BPH 中的共生菌。这些方法容易受到外部干扰，繁琐、耗时费力。微滴数字 PCR（ddPCR）不需要标准曲线，可以实现绝对定量，不依赖于 Cq 值，更有助于分析拷贝数变异、基因突变和相对基因表达。本研究建立并优化了基于 ddPCR 的 BPH YLS 快速检测方法。结果表明，该方法对两种 YLS（共生体和）的检测限分别为 1.3 拷贝/μL 和 1.2 拷贝/μL。样品重复的变异系数小于 5%；因此，本研究中建立的 ddPCR 方法具有良好的灵敏度、特异性和重复性。它可以用于快速准确地检测 BPH 的 YLS。
