Key message

Host mediated silencing of COM1 gene of Colletotrichum gloeosporioides disables appressorial differentiation and effectively prevents the development of Anthracnose disease in chilli and tomato.

Abstract

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.

中文翻译：

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宿主诱导的炭疽菌分生孢子形态 1 基因 (CgCOM1) 的沉默赋予辣椒和番茄对炭疽病的抵抗力。

关键信息

宿主介导的胶状炭疽菌COM1基因沉默使附着胞分化失灵，有效防止辣椒和番茄炭疽病的发展。

抽象的

炭疽病是由子囊菌类真菌炭疽菌引起的，它是全球辣椒和番茄严重减产的原因。传统上，有害杀虫剂用于控制炭疽病，效果有限。在这项研究中，我们评估了宿主诱导基因沉默 (HIGS) 方法靶向与真菌分生孢子和附着胞形成有关的 胶孢炭疽菌 COM1 ( CgCOM1 ) 发育基因以限制辣椒和番茄果实中的真菌感染的潜力。对于这项研究，我们开发了稳定的辣椒和番茄转基因品系，利用农杆菌表达CgCOM1 -RNAi 构建体介导的转化。通过分子和基因表达分析表征转基因植物。茎环 RT-PCR 证实了转基因辣椒和番茄 RNAi 系中特异性CgCOM1 siRNA 的产生。对叶子和果实的真菌攻击试验表明，与野生型和空载体对照植物相比，转基因品系对引起炭疽病的C. gloeosporioides具有抗性。转基因系中的 RT-qPCR 分析显示C. gloeosporioides感染组织中CgCOM1转录物的丰度极低，表明病原体中CgCOM1基因表达几乎完全沉默。Cg的显微镜检查- 受攻击的辣椒叶 - CgCOM1 i 系显示出高度抑制的分生孢子萌发、胚管发育、附着胞形成和C. gloeosporioides的菌丝体生长，导致植物组织的感染减少。这些结果证明了 HIGS 在沉默基本真菌发育基因的表达以抑制病原真菌的生长方面的高效使用，从而提供了一种高度精确的方法来阻止疾病的传播。