关键词：
citrus melanose;Diaporthe citri;RNA-Seq;sweet orange;reactive oxygen species
摘要：
Introduction: Reactive oxygen species (ROS) generation is a common disease defense mechanism in plants. However, it is unclear whether Citrus host activates defense response against Diaporthe citri causing citrus melanose disease by producing ROS, and the underlying molecular mechanisms are unknown. Methods: DAB staining and RNA-Seq technology were used to compare the active oxygen burst and differential gene expression, respectively, in uninfected and infected Citrus sinensis leaves at different time points during D. citri infection in vivo. The functions of CsRBOH (a significant DEG) were confirmed in N. benthamiana through the Agrobacterium-mediated transient expression system. Results: DAB staining indicated that C. sinensis initiated defense against D. citri infection within 24 h by generating ROS. Illumina sequencing revealed 25,557 expressed genes of C. sinensis. The most upregulated DEGs (n = 1,570) were identified 72 h after fungal inoculation (sample denoted as CD72). In the CD72 vs. Cs (samples at 0 h after fungal inoculation) comparison, the KEGG pathway category with the highest number of genes (n = 62) and most significant enrichment was Protein processing in endoplasmic reticulum, followed by Glutathione metabolism and MAPK signaling pathway-plant. GO analysis revealed that the DEGs of CD72 vs. Cs related to active oxygen burst and chitin recognition were significantly grouped into the regulation of biological processes and molecular functions, with GO terms including response to ROS, response to fungus, and oxidoreductase activity. Remarkably, CsRBOH was significantly enriched in the GO and KEGG analyses, and its expression pattern in qRT-PCR and DAB staining results were consistent. Among the 63 ROS-related DEGs, HSP genes and genes associated with the peroxidase family were highly significant as revealed by protein-protein interaction networks. Furthermore, ROS accumulation, cell death, and upregulation of defense-related genes were observed in N. benthamiana leaves with CsRBOH expressed through the Agrobacterium-mediated transient expression system. Conclusion: Our findings suggested that C. sinensis activates CsRBOH and ROS-related genes, leading to ROS accumulation to resist the invasion by D. citri. This study laid the foundation for future research on molecular mechanisms and breeding of C. sinensis cultivars resistant to citrus melanose.
摘要：
Citrus melanose, caused by Diaporthe citri, is one of the most prevalent and important fungal diseases of citrus crops globally. However, the overuse of chemical fungicides for disease control has an adverse impact on citrus production. In contrast, biological control agents (BCAs) are environmentally friendly and have become essential tools for plant disease control. In the present study, Bacillus subtilis M23, a strain isolated from healthy citrus leaves, significantly suppressed the mycelial growth of D. citri, the causal agent of citrus melanose, in vitro. The fermentation broth of B. subtilis M23 also exhibited good antifungal activity against D. citri on citrus plants in greenhouse and field experiments. Lipopeptides (LPs) produced by M23 exhibited excellent antifungal activity against D. citri, with a 50 % effective concentration (EC50) of approximately 1.00 and 0.28 mu g/mL for inhibition of mycelial growth and conidial germination, respectively. LP biosynthesis genes were identified in the M23 genome using a PCR assay. Notably, LP extracts significantly reduced the formation of pycnidia and ATP biosynthesis in D. citri. Comparative transcriptome analysis revealed significant differences in the expression of genes associated with fungal vacuole, oxidoreductase activity and endopeptidase activity in D. citri treated with B. subtilis M23. Both RNA-seq and RT-qPCR analysis provided similar results on the expression of selected genes. Collectively, our data provided convincing data supporting the biocontrol potential of B. subtilis M23 for the management of citrus melanose.
摘要：
The Med1 transcriptional coactivator is a crucial component of the Mediator middle complex, which regulates the expression of specific genes involved in cell development, differentiation, reproduction, and homeostasis. The Med1 LxxLL motif, a five-amino-acid peptide sequence, is essential for Med1-mediated gene expression. Our previous study revealed that the disruption of the Med1 subunit leads to a significant increase in fumonisin B1 (FB1) production in the maize pathogen Fusarium verticillioides. However, our understanding of how Med1 regulates FB1 biosynthesis in F. verticillioides, particularly through the Med1 LxxLL motifs, remains limited. To characterize the role of LxxLL motifs, we generated a series of Med1 LxxLL deletion and amino acid substitution mutants. These mutants exhibited impaired mycelial growth and conidia germination while demonstrating enhanced conidia production and virulence. Similar to the Med1 deletion mutant, Med1 LxxLL motif mutants also exhibited increased FB1 biosynthesis in F. verticillioides. Proteomic profiling revealed that the Med1 LxxLL motif regulated the biosynthesis of several key substances that affected FB1 production, including starch and carotenoid. Subsequent studies demonstrated that the production of amylopectin, which is strongly linked to FB1 biosynthesis, was significantly increased in Med1 LxxLL motif mutants. In addition, the disruption of carotenoid metabolic genes decreased carotenoid content, thus stimulating FB1 biosynthesis in F. verticillioides. Taken together, our results provide valuable insights into how the Med1 LxxLL motif regulates FB1 biosynthesis in the mycotoxigenic fungus F. verticillioides.
作者：
Yuan, S. Q.;Wang, Y. C.;Lei, L.;Hong, J. Y.;Yi, T. Y.;...
期刊：
PLANT DISEASE,2022年106(7):1996 ISSN：0191-2917
通讯作者：
T. Y. Yi<&wdkj&>Y. Y. Hong
作者机构：
[Yuan, S. Q.; Wang, Y. C.; Lei, L.; Yi, T. Y.; Hong, Y. Y.] Hunan Agr Univ, Coll Plant Protect, Hunan Prov Key Lab Biol & Control Plant Pests, Changsha 410128, Peoples R China.;[Hong, J. Y.] Hunan Agr Univ, Orient Sci & Technol Coll, Changsha 410128, Peoples R China.
通讯机构：
[T. Y. Yi; Y. Y. Hong] H;Hunan Provincial Key Laboratory for Biology and Control of Plant Pests, College of Plant Protection, Hunan Agricultural University, Changsha, 410128, China
摘要：
Carbamoyl phosphate synthetase is involved in arginine biosynthesis in many organisms. In this study, we investigate the biological function of Cpa1, a small subunit of carbamoyl phosphate synthetase of Colletotrichum gloeosporioides. The deletion of the CPA1 gene affected vegetative growth, arginine biosynthesis, and fungal pathogenicity. Genetic complementation with native CPA1 fully recovered all these defective phenotypes. We observed that Cpa1-RFP fusion protein is localized at the mitochondria, which is consistent with Cpa2, a large subunit of carbamoyl phosphate synthetase. We identified the proteins that interact with Cpa1 by using the two-hybrid screen approach, and we showed that Dut1 interacts with Cpa1 but without Cpa2 in vivo. Dut1 is dispensable for hyphal growth, appressorial formation, and fungal pathogenicity. Interestingly, the Dut1-Cpa1 complex is localized at the mitochondria. Further studies showed that Dut1 regulates Cpa1-Cpa2 interaction in response to arginine. In summary, our studies provide new insights into how Cpa1 interacts with its partner proteins to mediate arginine synthesis. (C) 2020 British Mycological Society. Published by Elsevier Ltd. All rights reserved.
作者机构：
[Lei, Ling] Hunan Agricultural University, 12575, Hunan Provincial Key Laboratory for Biology and Control of Plant Pests, College of Plant Protection, Hunan Agricultural University, Changsha, Hunan province, China,, Changsha, Hunan, China;[Lei, Ling] leiling19970805@163.com;[Hong, Yan-Yun] Hunan Agricultural University, 12575, College of Plant Protection & Hunan Provincial Key Laboratory for Biology and Control of Plant Diseases and Plant Pests, Nongda Road 1, Furong District, Changsha, Hunan, China, 410128;[Hong, Yan-Yun] hongyanyun@126.com;[Yi, Tu-Yong] Hunan Agricultural University, 12575, College of Plant Protection & Hunan Provincial Key Laboratory for Biology and Control of Plant Diseases and Plant Pests, No. 1 Nongda Road, Furong Distrcit, Changsha, Hunan, China, 410128
通讯机构：
[T. Y. Yi] H;Hunan Key Laboratory of Biology and Prevention and Control of Plant Diseases and Insect Pests, College of Plant Protection, Hunan Agricultural University, Changsha 410128, China
摘要：
Sarcandra glabra, belonging to the family Chloranthaceae, is a Chinese medicinal plant. The whole dry plant can be used as a medicine; it is rich in bioactive phytochemicals that possess anti-bacterial, anti-inflammatory, anti-oxidant, and anti-tumor properties (Xie et al. 2020). The current market price of S. glabra is around US$5/kg, and the annual demand is 3 500 000~4 000 000 kg in China (Pan et al. 2007). To meet consumer demand for safe and high-quality herbal products, the artificial cultivation of S. glabra has been vigorously promoted. In 2020, it was observed that a plant disease affected S. glabra growth in Hunan province. The disease symptoms included constriction at the base of the stem, with decay and a white mycelium covering. The plants finally died with a disease incidence ranging from 15% to 20%. Using our previously published methods (Yi et al. 2019), one fungal isolate was isolated from the cultured symptomatic stem tissue on potato dextrose agar (PDA) medium and was named as Kb. The isolate was subsequently transferred into 70% glycerol for preservation. The Kb colony varied in color from white to light yellow. The septate hyphae grew rapidly on PDA medium, at approximately 25 mm/day, at 28 °C. On the fifth day, rhizomorphs were formed at the edge and on the center of the PDA plate. On the sixth day, sclerotia developed into a rapeseed shape (d = 1.2~2.3 mm) with a smooth surface, and with white, yellow, or chestnut brown coloring. Morphologically, Kb was similar to Sclerotium rolfsii (Sun et al. 2020). Vigorously growing aerial hyphae were selected for molecular identification. The internal transcribed spacers (ITS) were amplified using the primer pairs ITS1/ITS4 (Glass et al. 1995). BLAST searches against Genbank indicated that Kb's ITS sequence shared 97% similarity with that of Athelia rolfsii (MN696630.1). Based on morphological and molecular characteristics, Kb was identified as A. rolfsii. The sequence was deposited in GenBank (MW288292). Pathogenicity tests were carried out using the following procedures. Three healthy S. glabra seedlings were inoculated at the stem base with a PDA plug (5 mm in diameter) covered with 5-day-old fungal mycelium cultured at 28 °C, while the remaining three seedlings were inoculated with distilled water only, as the control. Plants were incubated in a greenhouse at 28 °C. At 7 days post inoculation, the inoculated sites infected with the putative pathogen displayed identical constrictions as previously observed in the field. In contrast, the controls remained symptomless. The pathogen was reisolated from these infected seedlings, and its culture showed the same morphological and molecular traits as the original isolates. No pathogens were isolated from the control plants. Pathogenicity tests were repeated three times. Koch's postulates were fulfilled. Although S. rolfsii has been previously reported to cause Southern Blight on mung bean crops in China (Sun et al. 2020), this is the first report on A. rolfsii causing similar symptoms of Southern Blight on S. glabra in Hunan Province, China. Identification of the pathogens causing each disease is important for the development of effective disease management strategies and for extensive artificial cultivation.
作者机构：
[洪艳云; 易图永; 张玮] College of Plant Protection, Hunan Agricultural University / Hunan Provincial Key Laboratory for Biology and Control of Plant Pests, Changsha, 410128, China;[刘登望; 李林; 张博文] Institute of Dryland Crops of Hunan Agricultural University, Changsha, 410128, China
通讯机构：
[Yi, T.-Y.] C;[Li, L.] I;College of Plant Protection, China;Institute of Dryland Crops of Hunan Agricultural UniversityChina
摘要：
<jats:title>Abstract</jats:title><jats:sec>
<jats:title>Background</jats:title>
<jats:p>Peanut (<jats:italic>Arachis hypogaea</jats:italic> L.) is an important oil and economic crop. Calcium modulates plants in response to abiotic stresses and improves plant resistance to pathogens. Enrichment of beneficial microorganisms in the rhizosphere is associated with plant disease resistance and soil development. The purpose of this study was to analyze the differences in peanut rhizosphere microbial community structure between the calcium treatment and the control during two growth stages and to explain why calcium application could improve the resistance of peanuts to soil-borne pathogens.</jats:p>
</jats:sec><jats:sec>
<jats:title>Results</jats:title>
<jats:p>The 16S rDNA amplicon sequencing of rhizosphere microbiome showed that calcium application significantly enriched <jats:italic>Serratia marcescens</jats:italic> and other three dominant strains at the seedling stage. At the pod filling stage, ten dominant stains such as <jats:italic>Sphingomonas changbaiensis</jats:italic> and <jats:italic>Novosphingobium panipatense</jats:italic> were enriched by calcium. <jats:italic>Serratia marcescens</jats:italic> aseptic fermentation filtrate was mixed with PDA medium and inoculated with the main soil-borne pathogens in the seedling stage, which could inhibit the growth of <jats:italic>Fusarium solani</jats:italic> and <jats:italic>Aspergillus flavus</jats:italic>. The aseptic fermentation filtrate of <jats:italic>Novosphingobium panipatense</jats:italic> was mixed with PDA medium and inoculated with the main soil-borne pathogens in the pod filling stage, which could inhibit the growth of <jats:italic>Sclerotium rolfsii</jats:italic> and <jats:italic>Leptosphaerulina arachidicola</jats:italic>.</jats:p>
</jats:sec><jats:sec>
<jats:title>Conclusions</jats:title>
<jats:p>Calcium application increases the resistance of peanuts to soil-borne pathogens by enriching them with specific dominant bacteria.</jats:p>
</jats:sec>
