{"title":"一种基于 PCR 的简单方法,用于检测木薯蚧 Phenacoccus manihoti(半翅目:伪球虫科)的主要寄生虫 Anagyrus lopezi(膜翅目: Encyrtidae)和 Prochiloneurus pulchellus(膜翅目: Encyrtidae)。","authors":"Shun-ichiro Takano, Ngoc Hung Nguyen, Thi Xuyen Le, Ah Nge Htwe, Keiji Takasu","doi":"10.1111/ens.12575","DOIUrl":null,"url":null,"abstract":"<p>Estimating parasitism rates in the field is essential for developing and evaluating biocontrol strategies using parasitoids. In this study, we developed a simple polymerase chain reaction (PCR)-based method for detecting parasitism of the cassava mealybug <i>Phenacoccus manihoti</i> Matile-Ferrero (Hemiptera: Pseudococcidae) by the primary parasitoid <i>Anagyrus lopezi</i> De Santis (Hymenoptera: Encyrtidae) and its hyperparasitoid <i>Prochiloneurus pulchellus</i> Silvestri (Hymenoptera: Encyrtidae). Primers were designed to amplify partial cytochrome <i>c</i> oxidase subunit I genes of each species, and their sensitivity was evaluated with mealybugs that had been parasitized by <i>A. lopezi</i> 0, 3, and 6 days earlier, and mummified mealybugs containing <i>A. lopezi</i> pupae that had been parasitized by <i>P. pulchellus</i> 0, 3, 6, 9, and 12 days earlier. The detection rate of parasitism by <i>A. lopezi</i> was 100% for all ages of <i>A. lopezi</i>. The detection rate of parasitism by <i>P. pulchellus</i> ranged from 94.1% to 100%, depending on its developmental stage. For <i>P. pulchellus</i>, template DNA was diluted 10 times before PCR because PCR with the original concentration showed low detection rates, presumably due to the presence of PCR inhibitors. Overall, our primers can be considered sufficiently sensitive to be used for detecting each species.</p>","PeriodicalId":11745,"journal":{"name":"Entomological Science","volume":"27 2","pages":""},"PeriodicalIF":0.7000,"publicationDate":"2024-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"A simple PCR-based method for detecting Anagyrus lopezi (Hymenoptera: Encyrtidae) and Prochiloneurus pulchellus (Hymenoptera: Encyrtidae), primary and hyper parasitoids of the cassava mealybug Phenacoccus manihoti (Hemiptera: Pseudococcidae)\",\"authors\":\"Shun-ichiro Takano, Ngoc Hung Nguyen, Thi Xuyen Le, Ah Nge Htwe, Keiji Takasu\",\"doi\":\"10.1111/ens.12575\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>Estimating parasitism rates in the field is essential for developing and evaluating biocontrol strategies using parasitoids. In this study, we developed a simple polymerase chain reaction (PCR)-based method for detecting parasitism of the cassava mealybug <i>Phenacoccus manihoti</i> Matile-Ferrero (Hemiptera: Pseudococcidae) by the primary parasitoid <i>Anagyrus lopezi</i> De Santis (Hymenoptera: Encyrtidae) and its hyperparasitoid <i>Prochiloneurus pulchellus</i> Silvestri (Hymenoptera: Encyrtidae). Primers were designed to amplify partial cytochrome <i>c</i> oxidase subunit I genes of each species, and their sensitivity was evaluated with mealybugs that had been parasitized by <i>A. lopezi</i> 0, 3, and 6 days earlier, and mummified mealybugs containing <i>A. lopezi</i> pupae that had been parasitized by <i>P. pulchellus</i> 0, 3, 6, 9, and 12 days earlier. The detection rate of parasitism by <i>A. lopezi</i> was 100% for all ages of <i>A. lopezi</i>. The detection rate of parasitism by <i>P. pulchellus</i> ranged from 94.1% to 100%, depending on its developmental stage. For <i>P. pulchellus</i>, template DNA was diluted 10 times before PCR because PCR with the original concentration showed low detection rates, presumably due to the presence of PCR inhibitors. Overall, our primers can be considered sufficiently sensitive to be used for detecting each species.</p>\",\"PeriodicalId\":11745,\"journal\":{\"name\":\"Entomological Science\",\"volume\":\"27 2\",\"pages\":\"\"},\"PeriodicalIF\":0.7000,\"publicationDate\":\"2024-05-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Entomological Science\",\"FirstCategoryId\":\"97\",\"ListUrlMain\":\"https://onlinelibrary.wiley.com/doi/10.1111/ens.12575\",\"RegionNum\":4,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"ENTOMOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Entomological Science","FirstCategoryId":"97","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1111/ens.12575","RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"ENTOMOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
估算田间寄生率对于制定和评估使用寄生虫的生物防治策略至关重要。在这项研究中,我们开发了一种基于聚合酶链式反应(PCR)的简单方法,用于检测主要寄生虫 Anagyrus lopezi De Santis(膜翅目:Encyrtidae)及其超寄生虫 Prochiloneurus pulchellus Silvestri(膜翅目:Encyrtidae)对木薯蚧 Phenacoccus manihoti Matile-Ferrero(半翅目:伪球虫科)的寄生情况。设计了引物来扩增每个物种的部分细胞色素 c 氧化酶亚单位 I 基因,并用 0、3 和 6 天前寄生过 A. lopezi 的蛤蚧以及 0、3、6、9 和 12 天前寄生过 P. pulchellus 的含有 A. lopezi 蛹的木乃伊蛤蚧来评估它们的敏感性。所有年龄段的 A. lopezi 寄生虫检出率均为 100%。P.pulchellus的寄生虫检出率为94.1%至100%,具体取决于其发育阶段。对于 P. pulchellus,在进行 PCR 之前将模板 DNA 稀释了 10 倍,因为使用原始浓度进行 PCR 的检出率较低,这可能是由于 PCR 抑制剂的存在。总体而言,我们的引物灵敏度较高,可用于检测每个物种。
A simple PCR-based method for detecting Anagyrus lopezi (Hymenoptera: Encyrtidae) and Prochiloneurus pulchellus (Hymenoptera: Encyrtidae), primary and hyper parasitoids of the cassava mealybug Phenacoccus manihoti (Hemiptera: Pseudococcidae)
Estimating parasitism rates in the field is essential for developing and evaluating biocontrol strategies using parasitoids. In this study, we developed a simple polymerase chain reaction (PCR)-based method for detecting parasitism of the cassava mealybug Phenacoccus manihoti Matile-Ferrero (Hemiptera: Pseudococcidae) by the primary parasitoid Anagyrus lopezi De Santis (Hymenoptera: Encyrtidae) and its hyperparasitoid Prochiloneurus pulchellus Silvestri (Hymenoptera: Encyrtidae). Primers were designed to amplify partial cytochrome c oxidase subunit I genes of each species, and their sensitivity was evaluated with mealybugs that had been parasitized by A. lopezi 0, 3, and 6 days earlier, and mummified mealybugs containing A. lopezi pupae that had been parasitized by P. pulchellus 0, 3, 6, 9, and 12 days earlier. The detection rate of parasitism by A. lopezi was 100% for all ages of A. lopezi. The detection rate of parasitism by P. pulchellus ranged from 94.1% to 100%, depending on its developmental stage. For P. pulchellus, template DNA was diluted 10 times before PCR because PCR with the original concentration showed low detection rates, presumably due to the presence of PCR inhibitors. Overall, our primers can be considered sufficiently sensitive to be used for detecting each species.
期刊介绍:
Entomological Science is the official English language journal of the Entomological Society of Japan. The Journal publishes original research papers and reviews from any entomological discipline or from directly allied field in ecology, behavioral biology, physiology, biochemistry, development, genetics, systematics, morphology, evolution and general entomology. Papers of applied entomology will be considered for publication if they significantly advance in the field of entomological science in the opinion of the Editors and Editorial Board.