Search Results (36)

Sclerotinia sclerotiorum is a globally widespread and vast destructive plant pathogenic fungus that causes significant yield losses in crops. Due to the lack of effective resistant germplasm resources, the control of diseases caused by S. sclerotiorum largely relies on chemical fungicides. However, excessive use of these chemicals not only causes environmental concerns but also leads to the increased development of resistance in S. sclerotiorum. In contrast, trans-kingdom sRNA silencing-based technologies, such as host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS), offer novel, effective, and environmentally friendly methods for the management of S. sclerotiorum infection. This review summarizes recent advances in the identification of S. sclerotiorum pathogenic genes, target gene selection, categories, and application of trans-kingdom RNA interference (RNAi) technologies targeting this pathogen. Although some challenges, including off-target effects and the efficiency of external sRNA uptake, exist, recent findings have proposed solutions for further improvement. Combined with the latest developments in CRISPR/Cas gene editing and other technologies, trans-kingdom RNAi has significant potential to become a crucial tool in the control of sclerotinia stem rot (SSR), mitigating the impact of S. sclerotiorum on crop production. Full article

In planta RNAi or host-induced gene silencing (HIGS) has undergone significant advancements that have rendered it efficient and stable at the transgenerational level in plants for regulating host genes and targeting genes of insect pests and plant pathogens. Similarly, topical RNAi or spray-induced gene silencing (SIGS) has garnered considerable attention as an environmentally sustainable, selective, and alternative approach to chemical control of insect pests and plant pathogens. Several biotechnology companies and startups have focused their efforts on RNAi-based solutions for topical application in agriculture. Nevertheless, further technological advancements are required to enhance the efficacy of topical RNAi in agriculture, including improved dsRNA delivery systems, better target gene selection, and addressing biosafety regulatory issues. Herein, this review discusses key advances and bottlenecks in RNAi, and summarizes successful applications of these RNAi-based technologies in agriculture focusing on in planta and topical RNAi to control insect pests and plant pathogens. Furthermore, this review delves into the patenting landscape, biosafety considerations, risk evaluations, and the current regulatory status of RNAi in Latin America. Finally, it explores the contributions of RNAi to plant science, food production, and fostering a more sustainable form of agriculture. Full article

This review highlights the emerging role of cross-kingdom RNA interference in plant–microbe interactions, particularly the transfer of sRNAs from microbes to plants and vice versa, emphasizing the importance of this mechanism in both mutualistic and pathogenic contexts. As plants adapted to terrestrial life, they formed symbiotic relationships with microbes, essential for nutrient uptake and defense. Emerging evidence underscores sRNAs, including small interfering RNAs (siRNAs) and microRNAs (miRNAs), as critical regulators of gene expression and immune responses in plant–microbe interactions. In mutualistic symbioses, such as mycorrhizal fungi and nitrogen-fixing bacteria associations, sRNAs are hypothesized to regulate nutrient exchange and symbiotic stability. In pathogenic scenarios, microbes utilize sRNAs to undermine plant defenses, while plants employ strategies like host-induced gene silencing (HIGS) to counteract these threats. We further explore the emerging role of extracellular vesicles (EVs) in sRNA transport, which is critical for facilitating interspecies communication in both pathogenic and mutualistic contexts. Although the potential of ckRNAi in mutualistic interactions is promising, the review highlights the need for further experimental validation to establish its true significance in these relationships. By synthesizing current research, this review highlights the intricate molecular dialogues mediated by sRNAs in plant–microbe interactions and identifies critical gaps, proposing future research directions aimed at harnessing these mechanisms for agricultural advancements. Full article

Maize is one of the major crops that are susceptible to Aspergillus flavus infection and subsequent aflatoxin contamination, which poses a serious health threat to humans and domestic animals. Here, an RNA interference (RNAi) approach called Host-Induced Gene Silencing (HIGS) was employed to suppress the O-methyl transferase gene (omtA, also called aflP), a key gene involved in aflatoxin biosynthesis. An RNAi vector carrying part of the omtA gene was introduced into the B104 maize line. Among the six transformation events that were positive for containing the omtA transgene, OmtA-6 and OmtA-10 were self-pollinated from T1 to T4, and OmtA-7 and OmtA-12 to the T6 generation. These four lines showed at least an 81.3% reduction in aflatoxin accumulation at the T3 generation under laboratory conditions. When screened under field conditions with artificial inoculation, OmtA-7 at T5 and T6 generations and OmtA-10 at T4 generation showed a reduction in aflatoxin contamination between 60% and 91% (p < 0.02 to p < 0.002). In order to develop commercial maize lines with enhanced aflatoxin resistance, the omtA transgene in OmtA-7 was introduced into three elite inbred lines through crossing, and the resulting crosses also exhibited significantly lower aflatoxin accumulation compared to crosses with non-transgenic controls (p < 0.04). In addition, high levels of omtA-specific small RNAs were only detected in the transgenic kernel and leaf tissues. These results demonstrate that suppression of omtA through HIGS can enhance maize resistance to aflatoxin contamination, and this resistance can be transferred to elite backgrounds, providing a viable and practical approach to reduce aflatoxin contamination in maize. Full article

Cotton is often threatened by Verticillium wilt caused by V. dahliae. Understanding the molecular mechanism of V. dahlia–cotton interaction is important for the prevention of this disease. To analyze the transcriptome profiles in V. dahliae and cotton simultaneously, the strongly pathogenic strain Vd592 was inoculated into cotton, and the infected cotton roots at 36 h and 3 d post infection were subjected to dual RNA-seq analysis. For the V. dahliae, transcriptomic analysis identified 317 differentially expressed genes (DEGs) encoding classical secreted proteins, which were up-regulated at least at one time point during infection. The 317 DEGs included 126 carbohydrate-active enzyme (CAZyme) and 108 small cysteine-rich protein genes. A pectinesterase gene (VDAG_01782) belonging to CAZyme, designated as VdPE1, was selected for functional validation. VdPE1 silencing by HIGS (host-induced gene silencing) resulted in reduced disease symptoms and the increased resistance of cotton to V. dahliae. For the cotton, transcriptomic analysis found that many DEGs involved in well-known disease resistance pathways (flavonoid biosynthesis, plant hormone signaling, and plant–pathogen interaction) as well as PTI (pattern-triggered immunity) and ETI (effector-triggered immunity) processes were significantly down-regulated in infected cotton roots. The dual RNA-seq data thus potentially connected the genes encoding secreted proteins to the pathogenicity of V. dahliae, and the genes were involved in some disease resistance pathways and PTI and ETI processes for the susceptibility of cotton to V. dahliae. These findings are helpful in the further characterization of candidate genes and breeding resistant cotton varieties via genetic engineering. Full article

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), stands out as one of the most devastating epidemics impacting wheat production worldwide. Resistant wheat varieties had swiftly been overcome due to the emergence of new virulent Pst strains. Effectors secreted by Pst interfere with plant immunity, and verification of their biological function is extremely important for controlling wheat stripe rust. In this study, we identified an effector, Pst-18220, from Puccinia striiformis f. sp. tritici (Pst), which was induced during the early infection stage of Pst. Silencing the expression of Pst-18220 through virus-mediated host-induced gene silencing (HIGS) resulted in a decreased number of rust pustules. In Nicotiana benthamiana, it significantly suppressed cell death induced by Pseudomonas syringae pv. tomato (Pto) DC3000. In Arabidopsis, plants with stable overexpression of Pst-18220 showed increased susceptibility to Pto DC3000, accompanied by a decrease in the expression level of pattern-triggered immunity (PTI)/effector-triggered immunity (ETI)-related genes, namely, AtPCRK1, AtPCRK2, and AtBIK1. These results emphasize the significant role of the Pst candidate effector, Pst-18220, in rust pathogenicity and the suppression of plant defense mechanisms. This broadens our understanding of effectors without any known motif. Full article

Wheat leaf rust fungus is an obligate parasitic fungus that can absorb nutrients from its host plant through haustoria and secrete effector proteins into host cells. The effector proteins are crucial factors for pathogenesis as well as targets for host disease resistance protein recognition. Exploring the role of effector proteins in the pathogenic process of Puccinia triticina Eriks. (Pt) is of great significance for unraveling its pathogenic mechanisms. We previously found that a cysteine-rich effector protein, Pt1641, is highly expressed during the interaction between wheat and Pt, but its specific role in pathogenesis remains unclear. Therefore, this study employed techniques such as heterologous expression, qRT-PCR analysis, and host-induced gene silencing (HIGS) to investigate the role of Pt1641 in the pathogenic process of Pt. The results indicate that Pt1641 is an effector protein with a secretory function and can inhibit BAX-induced programmed cell death in Nicotiana benthamiana. qRT-PCR analyses showed that expression levels of Pt1641 were different during the interaction between the high-virulence strain THTT and low-virulence strains FGD and Thatcher, respectively. The highest expression level in the low-virulence strain FGD was four times that of the high-virulence strain THTT. The overexpression of Pt1641 in wheat near-isogenic line TcLr1 induced callose deposition and H₂O₂ production on TcLr1. After silencing Pt1641 in the Pt low-virulence strain FGD on wheat near-isogenic line TcLr1, the pathogenic phenotype of Pt physiological race FGD on TcLr1 changed from “;” to “3”, indicating that Pt1641 plays a non-toxic function in the pathogenicity of FGD to TcLr1. This study helps to reveal the pathogenic mechanism of wheat leaf rust and provides important guidance for the mining and application of Pt avirulent genes. Full article

Chitinase genes, as a class of cell wall hydrolases, are essential for the development and pathogenesis of Fusarium oxysporum f.sp. vasinfectum (F. ox) in cotton, but related research focused on chitinase genes are limited. This study explored two island cotton root secretions from the highly resistant cultivar Xinhai 41 and sensitive cultivar Xinhai 14 to investigate their interaction with F. ox by a weighted correlation network analysis (WGCNA). As a result, two modules that related to the fungal pathogenicity emerged. Additionally, a total of twenty-five chitinase genes were identified. Finally, host-induced gene silencing (HIGS) of FoChi20 was conducted, and the cotton plants showed noticeably milder disease with a significantly lower disease index than the control. This study illuminated that chitinase genes play crucial roles in the pathogenicity of cotton wilt fungi, and the FoChi20 gene could participate in the pathogenesis of F. ox and host–pathogen interactions, which establishes a theoretical framework for disease control in Sea Island cotton. Full article

Recent advancements in molecular biology have revolutionized plant disease diagnosis and management. This review focuses on disease diagnosis through serological techniques, isothermal amplification methods, CRISPR-based approaches, and management strategies using RNA-based methods. Exploring high-throughput sequencing and RNA interference (RNAi) technologies like host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS), this review delves into their potential. Despite the precision offered by RNAi in pest and pathogen management, challenges such as off-target effects and efficient dsRNA delivery persist. This review discusses the significance of these strategies in preventing aphid-mediated plant virus transmission, emphasizing the crucial role of meticulous dsRNA design for effective viral RNA targeting while minimizing harm to plant RNA. Despite acknowledged challenges, including off-target effects and delivery issues, this review underscores the transformative potential of RNA-based strategies in agriculture. Envisaging reduced pesticide dependency and enhanced productivity, these strategies stand as key players in the future of sustainable agriculture. Full article

Puccinia striiformis f. sp. tritici is an obligate biotrophic fungus that causes destructive stripe rust disease in wheat. During infection, Pst secretes virulence effectors via a specific infection structure—the haustorium—inside host cells to disturb host immunity and promote fungal colonization and expansion. Hence, the identification and functional analyses of Pst effectors are of great significance in deciphering the Pst pathogenicity mechanism. Here, we identified one candidate Pst effector Pst_9302 that could suppress Bax-triggered cell death in Nicotiana benthamiana. qRT-PCR analyses showed that the transcript levels of Pst_9302 were highly increased during the early infection stages of Pst. The transient expression of Pst_9302 in wheat via the type-three secretion system (T3SS) significantly inhibited the callose deposition induced by Pseudomonas syringae EtHAn. During wheat–Pst interaction, Pst_9302 overexpression suppressed reactive oxygen species (ROS) accumulation and cell death caused by the avirulent Pst race CYR23. The host-induced gene silencing (HIGS) of Pst_9302 resulted in decreased Pst pathogenicity with reduced infection area. The results suggest that Pst_9302 plays a virulence role in suppressing plant immunity and promoting Pst pathogenicity. Moreover, wheat voltage-dependent anion channel 1 protein (TaVDAC1) was identified as candidate Pst_9302-interacting proteins by yeast two-hybrid (Y2H) screening. Pull-down assays using the His-Pst_9302 and GST-TaVDAC1 protein verified their interactions. These results suggest that Pst_9302 may modulate wheat TaVDAC1 to regulate plant immunity. Full article

Sclerotinia sclerotiorum is a devastating fungal pathogen that causes severe crop losses worldwide. It is of vital importance to understand its pathogenic mechanism for disease control. Through a forward genetic screen combined with next-generation sequencing, a putative protein kinase, SsCak1, was found to be involved in the growth and pathogenicity of S. sclerotiorum. Knockout and complementation experiments confirmed that deletions in SsCak1 caused defects in mycelium and sclerotia development, as well as appressoria formation and host penetration, leading to complete loss of virulence. These findings suggest that SsCak1 is essential for the growth, development, and pathogenicity of S. sclerotiorum. Therefore, SsCak1 could serve as a potential target for the control of S. sclerotiorum infection through host-induced gene silencing (HIGS), which could increase crop resistance to the pathogen. Full article

Our duty to conserve global natural ecosystems is increasingly in conflict with our need to feed an expanding population. The use of conventional pesticides not only damages the environment and vulnerable biodiversity but can also still fail to prevent crop losses of 20–40% due to pests and pathogens. There is a growing call for more ecologically sustainable pathogen control measures. RNA-based biopesticides offer an eco-friendly alternative to the use of conventional fungicides for crop protection. The genetic modification (GM) of crops remains controversial in many countries, though expression of transgenes inducing pathogen-specific RNA interference (RNAi) has been proven effective against many agronomically important fungal pathogens. The topical application of pathogen-specific RNAi-inducing sprays is a more responsive, GM-free approach to conventional RNAi transgene-based crop protection. The specific targeting of essential pathogen genes, the development of RNAi-nanoparticle carrier spray formulations, and the possible structural modifications to the RNA molecules themselves are crucial to the success of this novel technology. Here, we outline the current understanding of gene silencing pathways in plants and fungi and summarize the pioneering and recent work exploring RNA-based biopesticides for crop protection against fungal pathogens, with a focus on spray-induced gene silencing (SIGS). Further, we discuss factors that could affect the success of RNA-based control strategies, including RNA uptake, stability, amplification, and movement within and between the plant host and pathogen, as well as the cost and design of RNA pesticides. Full article

Wheat stripe rust, caused by the obligate biotrophic fungus Puccinia striiformis f. sp. tritici (Pst), seriously affects wheat production. Here, we report the complete genome sequence and biological characterization of a new mitovirus from P. striiformis strain GS-1, which was designated as “Puccinia striiformis mitovirus 2” (PsMV2). Genome sequence analysis showed that PsMV2 is 2658 nt in length with an AU-rich of 52.3% and comprises a single ORF of 2348 nt encoding an RNA-dependent RNA polymerase (RdRp). Phylogenetic analysis indicated that PsMV2 is a new member of the genus Unuamitovirus within the family Mitoviridae. In addition, PsMV2 multiplied highly during Pst infection and it suppresses programmed cell death (PCD) triggered by Bax. Silencing of PsMV2 in Pst by barley stripe mosaic virus (BSMV)-mediated Host Induced Gene Silencing (HIGS) reduced fungal growth and decreased pathogenicity of Pst. These results indicate PsMV2 promotes host pathogenicity in Pst. Interestingly, PsMV2 was detected among a wide range of field isolates of Pst and may have coevolved with Pst in earlier times. Taken together, our results characterized a novel mitovirus PsMV2 in wheat stripe rust fungus, which promotes the virulence of its fungal host and wide distribution in Pst which may offer new strategies for disease control. Full article

Aflatoxins are immunosuppressive and carcinogenic secondary metabolites, produced by the filamentous ascomycete Aspergillus flavus, that are hazardous to animal and human health. In this study, we show that multiplexed host-induced gene silencing (HIGS) of Aspergillus flavus genes essential for fungal sporulation and aflatoxin production (nsdC, veA, aflR, and aflM) confers enhanced resistance to Aspergillus infection and aflatoxin contamination in groundnut (<20 ppb). Comparative proteomic analysis of contrasting groundnut genotypes (WT and near-isogenic HIGS lines) supported a better understanding of the molecular processes underlying the induced resistance and identified several groundnut metabolites that might play a significant role in resistance to Aspergillus infection and aflatoxin contamination. Fungal differentiation and pathogenicity proteins, including calmodulin, transcriptional activator-HacA, kynurenine 3-monooxygenase 2, VeA, VelC, and several aflatoxin pathway biosynthetic enzymes, were downregulated in Aspergillus infecting the HIGS lines. Additionally, in the resistant HIGS lines, a number of host resistance proteins associated with fatty acid metabolism were strongly induced, including phosphatidylinositol phosphate kinase, lysophosphatidic acyltransferase-5, palmitoyl-monogalactosyldiacylglycerol Δ-7 desaturase, ceramide kinase-related protein, sphingolipid Δ-8 desaturase, and phospholipase-D. Combined, this knowledge can be used for groundnut pre-breeding and breeding programs to provide a safe and secure food supply. Full article

Chitin is the main component of fungal cell walls, which can be recognized by pattern recognition receptors (PRRs) as pathogen-associated molecular patterns (PAMP). Chitinase in filamentous fungi has been reported to degrade immunogenic chitin oligomers, thereby preventing chitin-induced immune activation. In this study, we identified the chitinase families in 10 fungal genomes. A total of 131 chitinase genes were identified. Among the chitinase families, 16 chitinase genes from Puccinia striiformis f. sp. tritici (Pst) were identified, and the expression of PstChia1 was the highest during Pst infection. Further studies indicated that PstChia1 is highly induced during the early stages of the interaction of wheat and Pst and has chitinase enzyme activity. The silencing of PstChia1 revealed that PstChia1 limited the growth and reduced the virulence of Pst. The expression level of TaPR1 and TaPR2 was induced in PstChia1 knockdown plants, suggesting that PstChia1 is involved in regulating wheat resistance to Pst. Our data suggest that PstChia1 contributes to pathogenicity by interfering with plant immunity and regulating the growth of Pst. Full article