TERI School of Advanced Studies, 10 Institutional Area, New Delhi, 110070, India.
The Energy and Resources Institute, Lodi Road, New Delhi, 110003, India.
Plant Mol Biol. 2020 Nov;104(4-5):381-395. doi: 10.1007/s11103-020-01046-3. Epub 2020 Aug 16.
Host mediated silencing of COM1 gene of Colletotrichum gloeosporioides disables appressorial differentiation and effectively prevents the development of Anthracnose disease in chilli and tomato. Anthracnose disease is caused by the ascomycetes fungal species Colletotrichum, which is responsible for heavy yield losses in chilli and tomato worldwide. Conventionally, harmful pesticides are used to contain anthracnose disease with limited success. In this study, we assessed the potential of Host-Induced Gene Silencing (HIGS) approach to target the Colletotrichum gloeosporioides COM1 (CgCOM1) developmental gene involved in the fungal conidial and appressorium formation, to restrict fungal infection in chilli and tomato fruits. For this study, we have developed stable transgenic lines of chilli and tomato expressing CgCOM1-RNAi construct employing Agrobacterium-mediated transformation. Transgenic plants were characterized by molecular and gene expression analyses. Production of specific CgCOM1 siRNA in transgenic chilli and tomato RNAi lines was confirmed by stem-loop RT-PCR. Fungal challenge assays on leaves and fruits showed that the transgenic lines were resistant to anthracnose disease-causing C. gloeosporioides in comparison to wild type and empty-vector control plants. RT-qPCR analyses in transgenic lines revealed extremely low abundance of CgCOM1 transcripts in the C. gloeosporioides infected tissues, indicating near complete silencing of CgCOM1 gene expression in the pathogen. Microscopic examination of the Cg-challenged leaves of chilli-CgCOM1i lines revealed highly suppressed conidial germination, germ tube development, appressoria formation and mycelial growth of C. gloeosporioides, resulting in reduced infection of plant tissues. These results demonstrated highly efficient use of HIGS in silencing the expression of essential fungal developmental genes to inhibit the growth of pathogenic fungi, thus providing a highly precise approach to arrest the spread of disease.
宿主介导的 Colletotrichum gloeosporioides COM1 基因沉默可抑制附着胞分化，有效防治辣椒和番茄炭疽病。炭疽病由子囊菌真菌 Colletotrichum 引起，是全球辣椒和番茄减产的主要原因。传统上，使用有害农药来控制炭疽病，但效果有限。在这项研究中，我们评估了宿主诱导基因沉默（HIGS）方法的潜力，以靶向真菌分生孢子和附着胞形成所涉及的 Colletotrichum gloeosporioides COM1（CgCOM1）发育基因，从而限制辣椒和番茄果实中的真菌感染。为此，我们利用农杆菌介导的转化，开发了表达 CgCOM1-RNAi 构建体的辣椒和番茄稳定转基因系。通过分子和基因表达分析对转基因植物进行了表征。通过茎环 RT-PCR 证实了转基因辣椒和番茄 RNAi 系中产生了特异性的 CgCOM1 siRNA。叶片和果实上的真菌挑战试验表明，与野生型和空载体对照植物相比，转基因系对炭疽病病原菌 C. gloeosporioides 具有抗性。在转基因系中进行的 RT-qPCR 分析显示，感染组织中 CgCOM1 转录本的丰度极低，表明 CgCOM1 基因在病原体中的表达几乎完全沉默。对 Cg 挑战的辣椒叶片进行的显微检查表明，Cg-challenged 叶片的附着胞形成和菌丝生长受到高度抑制，导致病原菌对植物组织的感染减少。这些结果表明，HIGS 可高效用于沉默关键真菌发育基因的表达，从而抑制致病真菌的生长，从而为阻止疾病传播提供了一种高度精确的方法。
