Zhang Huan, Tong Yan, Liu Huifang, Guo Lin, Jin Wenyi, Li Xuan, Meng Qian, Yu Xupeng, Fang Fenfen, Qin Qilian, Yang Miaomiao
State Key Laboratory of Integrated Management of Pest Insects and Rodents, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.
University of Chinese Academy of Sciences, Beijing, China.
Heliyon. 2023 Apr 7;9(4):e15170. doi: 10.1016/j.heliyon.2023.e15170. eCollection 2023 Apr.
Plutella xylostella granulovirus (PlxyGV) biopesticide is an effective tool to control the long-term damage of Plutella xylostella (Linnaeus) to cruciferous vegetables. In China, PlxyGV can be produced on a large scale using host insects, and its products have been registered in 2008. In experiments and biopesticide production, the routine enumeration method of PlxyGV virus particles is to use the Petroff-Hausser counting chamber in dark field microscope. However, the accuracy and repeatability of granulovirus (GV) counting are affected due to the small particle size of GV occlusion bodies (OBs), the limitations of optical microscope, the judgment of different operators, host impurities, the addition of biological products. This limits the convenience of its production, product quality, trading and field application. Here we use PlxyGV as an example, the method based on Real-time fluorescence quantitative PCR (qPCR) was optimized from two aspects of sample treatment and specific primers design, which improved the repeatability and accuracy of absolute quantitative OBs of GV. This study provides basic information for accurate quantitative PlxyGV by qPCR method.
小菜蛾颗粒体病毒(PlxyGV)生物农药是控制小菜蛾(Linnaeus)对十字花科蔬菜长期危害的有效工具。在中国,PlxyGV可利用宿主昆虫大规模生产,其产品已于2008年注册。在实验和生物农药生产中,PlxyGV病毒粒子的常规计数方法是在暗视野显微镜下使用血细胞计数板。然而,由于颗粒体病毒(GV)包涵体(OBs)粒径小、光学显微镜的局限性、不同操作人员的判断、宿主杂质、生物制品添加等因素,影响了GV计数的准确性和重复性。这限制了其生产便利性、产品质量、贸易及田间应用。在此,我们以PlxyGV为例,从样品处理和特异性引物设计两方面对基于实时荧光定量PCR(qPCR)的方法进行优化,提高了GV绝对定量OBs的重复性和准确性。本研究为采用qPCR方法准确定量PlxyGV提供了基础信息。