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Even at low concentrations in environmental waters, some viruses are highly infective, making them a threat to human health. They are the leading cause of waterborne enteric diseases. In agriculture, plant viruses in irrigation and runoff water threat the crops. The low concentrations pose a challenge to early contamination detection. Thus, concentrating the virus particles into a small volume may be mandatory to achieve reliable detection in molecular techniques. This paper reviews the organic monoliths developments and their applications to concentrate virus particles from waters (waste, surface, tap, sea, and irrigation waters). Free‐radical polymerization and polyaddition reactions are the most common strategies to prepare the monoliths currently used for virus concentration. Here, the routes for preparing and functionalizing both methacrylate and epoxy‐based monoliths will be shortly described, following a revision of their retention mechanisms and applications in the concentration of enteric and plant viruses in several kinds of waters. [ABSTRACT FROM AUTHOR]

Background: The tricarboxylic acid (TCA) cycle is crucial for energy supply in animal, plant, and microbial cells. It is not only the main pathway of carbohydrate catabolism but also the final pathway of lipid and protein catabolism. Some TCA genes have been found to play important roles in the growth and development of tomato and potato, but no comprehensive study of TCA cycle genes in Solanaceae crops has been reported. Results: In this study, we analyzed TCA cycle genes in four important Solanaceae vegetable crops (potato (Solanum tuberosum), tomato (Solanum lycopersicum), eggplant (Solanum melongena), and pepper (Capsicum annuum)) based on comparative genomics. The four Solanaceae crops had a total of 180 TCA cycle genes: 43 in potato, 44 in tomato, 40 in eggplant, and 53 in pepper. Phylogenetic analysis, collinearity analysis, and tissue expression patterns revealed the conservation of and differences in TCA cycle genes between the four Solanaceae crops and found that there were unique subgroup members in Solanaceae crops that were independent of Arabidopsis genes. The expression analysis of potato TCA cycle genes showed that (1) they were widely expressed in various tissues, and some transcripts like Soltu.DM.01G003320.1(SCoAL) and Soltu.DM.04G021520.1 (SDH) mainly accumulate in vegetative organs, and some transcripts such as Soltu.DM.12G005620.3 (SDH) and Soltu.DM.02G007400.4 (MDH) are preferentially expressed in reproductive organs; (2) several transcripts can be significantly induced by hormones, such as Soltu.DM.08G023870.2 (IDH) and Soltu.DM.06G029290.1 (SDH) under ABA treatment, and Soltu.DM.07G021850.2 (CSY) and Soltu.DM.09G026740.1 (MDH) under BAP treatment, and Soltu.DM.02G000940.1 (IDH) and Soltu.DM.01G031350.4 (MDH) under GA treatment; (3) Soltu.DM.11G024650.1 (SDH) can be upregulated by the three disease resistance inducers including Phytophthora infestans, acibenzolar-S-methyl (BTH), and DL-β-amino-n-butyric acid (BABA); and (4) the levels of Soltu.DM.01G045790.1 (MDH), Soltu.DM.01G028520.3 (CSY), and Soltu.DM.12G028700.1 (CSY) can be activated by both NaCl and mannitol. The subcellular localization results of three potato citrate synthases showed that Soltu.DM.01G028520.3 was localized in mitochondria, while Soltu.DM.12G028700.1 and Soltu.DM.07G021850.1 were localized in the cytoplasm. Conclusions: This study provides a scientific foundation for the comprehensive understanding and functional studies of TCA cycle genes in Solanaceae crops and reveals their potential roles in potato growth, development, and stress response. [ABSTRACT FROM AUTHOR]

A new set of loop-mediated isothermal amplification assays was developed for detection of phytoplasmas associated with plant diseases and optimised for rapid laboratory detection. This set of assays was designed based on the leucyl tRNA synthetase gene expressed in most of the phytoplasmas and showing a good level of sequence divergence among strains from different 16Sr groups. This makes it a good target for developing discrete assays that can detect specific phytoplasma 16Sr groups. In this study, unique sets of LAMP assays were designed and validated for seven phytoplasma ribosomal groups (16SrI, 16SrII, 16SrIII, 16SrV, 16SrVI, 16SrX, and 16SrXII). These assays are specific, rapid and as sensitive as the nested PCR assays and the results could be obtained in less than 30 min in real-time LAMP assays. [ABSTRACT FROM AUTHOR]

Plants develop a plethora of defense strategies during their acclimation and interactions with various environmental stresses. Secondary metabolites play a pivotal role in the processes during stress acclimation, therefore deciphering their relevant responses exchange the interpretation of the underlying molecular mechanisms that may contribute to improved adaptability and efficacy. In the current study, tomato plants were exposed to short-term cold stress (5 °C for 16 h) or inoculated (20 d) with either Cucumber Mosaic Virus (CMV) or Potato Virus Y (PVY). Responses were recorded via the assessments of leaf total phenolic (TP) content, total flavonoid (TF) levels, and phenylalanine ammonia-lyase (PAL) enzyme activity. The transcription of the gene families regulating the core phenylpropanoid biosynthetic pathway (PBP) at an early (PAL, cinnamic acid 4-hydroxylase, 4-coumarate-CoA ligase) or late (chalcone synthase and flavonol synthase) stage was also evaluated. The results showed that cold stress stimulated an increase in TP and TF contents, while PAL enzyme activity was also elevated compared to viral infection. Besides genes transcription of the enzymes involved in the core PBP was mostly induced by cold stress, whereas transcription of the genes regulating flavonoid biosynthesis was mainly triggered by viral infection. In conclusion, abiotic and biotic stressors induced differential regulation of the core PBP and flavonoid biosynthetic metabolism. Taking the above into consideration, our results highlight the complexity of tomato responses to diverse stimuli allowing for better elucidation of stress tolerance mechanisms at this crop. [ABSTRACT FROM AUTHOR]

Plants are often attacked by multiple antagonists and traits of the attacking organisms and their order of arrival onto hosts may affect plant defences. However, few studies have assessed how multiple antagonists, and varying attack order, affect plant defence or nutrition. To address this, we assessed defensive and nutritional responses of Pisum sativum plants after attack by a vector herbivore (Acrythosiphon pisum), a nonvector herbivore (Sitona lineatus), and a pathogen (Pea enation mosaic virus, PEMV). We show viruliferous A. pisum induced several antipathogen plant defence signals, but these defences were inhibited by S. lineatus feeding on peas infected with PEMV. In contrast, S. lineatus feeding induced antiherbivore defence signals, and these plant defences were enhanced by PEMV. Sitona lineatus also increased abundance of plant amino acids, but only when they attacked after viruliferous A. pisum. Our results suggest that diverse communities of biotic antagonists alter defence and nutritional traits of plants through complex pathways that depend on the identity of attackers and their order of arrival onto hosts. Moreover, we show interactions among a group of biotic stressors can vary along a spectrum from antagonism to enhancement/synergism based on the identity and order of attackers, and these interactions are mediated by a multitude of phytohormone pathways. [ABSTRACT FROM AUTHOR]

Bois noir (BN) grapevine disease is associated with 'Candidatus Phytoplasma solani' (CaPsol), a pathogen with a complex ecology including multiple insect vectors and plant hosts. A key point to improve the effectiveness of BN control strategies consists in determining the epidemiological role of ground‐cover weeds. The present study employed a multidisciplinary approach, based on the application of spatial (spatial analysis by distance indices) and molecular (stamp gene typing) analyses, to identify weeds with a potential role in BN epidemiology in northern Italy. Generated data showed that in addition to Convolvulus arvensis, one of the main CaPsol inoculum sources, Chenopodium album, Polygonum aviculare and Trifolium repens were found associated with BN epidemiology. CaPsol molecular typing highlighted that the strains prevalent in symptomatic grapevines were characterised by stamp sequence variants St19, St11 (nettle‐related) and St5 (bindweed‐related). The latter was prevalent also in Hyalesthes obsoletus and weeds, suggesting their main association with bindweed‐related epidemiology. On the other hand, nettle‐related CaPsol strains were occasionally found in H. obsoletus and weeds. Considering that H. obsoletus‐mediated transmission of CaPsol occurs mainly with young instars, further investigations will confirm if, in addition to bindweed and nettle, weeds associated with BN epidemiology in Franciacorta can represent larval developmental hosts and, consequently, act as CaPsol reservoirs for transmission to grapevine. Moreover, other studies are needed to clarify the relationship between such weeds and CaPsol alternative vectors. [ABSTRACT FROM AUTHOR]

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High‐throughput droplet‐based digital PCR (ddPCR) is a refinement of the conventional polymerase chain reaction (PCR)‎ methods. In ddPCR, DNA/RNA is encapsulated stochastically inside the microdroplets as reaction chambers. A small percentage of the reaction chamber contains one or fewer copies of the DNA or RNA. After PCR amplification, concentrations are determined based on the proportion of nonfluorescent partitions through the Poisson distribution. Some of the main features of ddPCR include high sensitivity and specificity, absolute quantification without a standard curve, high reproducibility, good tolerance to PCR inhibitor, and high efficacy compared to conventional molecular methods. These advantages make ddPCR a valuable addition to the virologist's toolbox. The following review outlines the recent technological advances in ddPCR methods and their applications in viral identification. [ABSTRACT FROM AUTHOR]

In order to survive in nature, different pathogens follow different procedures to manipulate their host plants for the pathogen favor. Plant viruses are not an exception of this rule. They are often found to alter the host plant traits in the way that affects the community of organisms in the host plant as well as the vectoring insects. It has been indicated that virus-infected plants are more preferable than virus-free plants with respect to the growth rates, longevity and reproduction of the vector. Viruses use several strategies in order to reprogram their host's cell to make it more conducive to replication and spread. Consequently, phytohormone signaling pathway in virus-infected plants can be disrupted either directly or indirectly. In plants, there are hormone pathways contribute to all aspects of plant physiology. Sometimes, virus infection can be advantageous to the infected host by providing the plant with tolerance to biotic and abiotic stresses. This article summarizes some aspects where the virus found to reprogram the host's cell to make it more conducive to virus' cycle of life. It also provides an important basic knowledge about how biotic and abiotic stress affects the interaction among virus, vector and the host plant; this knowledge could open the gate to understand the effect of multi-stress effect on the host plant in future studies through recognizing the necessity for plants to have an integrated system of defense against different threats. [ABSTRACT FROM AUTHOR]

Viruses cause a big fraction of economically important diseases in major crops, including tomato. In the past decade (2011–2020), many emerging or re-emerging tomato-infecting viruses were reported worldwide. In this period, 45 novel viral species were identified in tomato, 14 of which were discovered using high-throughput sequencing (HTS). In this review, we first discuss the role of HTS in these discoveries and its general impact on tomato virome research. We observed that the rate of tomato virus discovery is accelerating in the past few years due to the use of HTS. However, the extent of the post-discovery characterization of viruses is lagging behind and is greater for economically devastating viruses, such as the recently emerged tomato brown rugose fruit virus. Moreover, many known viruses still cause significant economic damages to tomato production. The review of databases and literature revealed at least 312 virus, satellite virus, or viroid species (in 22 families and 39 genera) associated with tomato, which is likely the highest number recorded for any plant. Among those, here, we summarize the current knowledge on the biology, global distribution, and epidemiology of the most important species. Increasing knowledge on tomato virome and employment of HTS to also study viromes of surrounding wild plants and environmental samples are bringing new insights into the understanding of epidemiology and ecology of tomato-infecting viruses and can, in the future, facilitate virus disease forecasting and prevention of virus disease outbreaks in tomato. [ABSTRACT FROM AUTHOR]

Plant pathogens, including viruses, negatively impact global crop production. Plants have evolved complex immune responses to pathogens. These responses are often controlled by nucleotide-binding leucine-rich repeat proteins (NLRs), which recognize intracellular, pathogen-derived proteins. Genetic resistance to plant viruses is often phenotypically characterized by programmed cell death at or near the infection site; a reaction termed the hypersensitive response. Although visualization of the hypersensitive response is often used as a hallmark of resistance, the molecular mechanisms leading to the hypersensitive response and associated cell death vary. Plants with extreme resistance to viruses rarely exhibit symptoms and have little to no detectable virus replication or spread beyond the infection site. Both extreme resistance and the hypersensitive response can be activated by the same NLR genes. In many cases, genes that normally provide an extreme resistance phenotype can be stimulated to cause a hypersensitive response by experimentally increasing cellular levels of pathogen-derived elicitor protein(s). The molecular mechanisms of extreme resistance and its relationship to the hypersensitive response are largely uncharacterized. Studies on potato and soybean cultivars that are resistant to strains of Potato virus Y (PVY), Potato virus X (PVX), and Soybean mosaic virus (SMV) indicate that abscisic acid (ABA)-mediated signaling and NLR nuclear translocation are important for the extreme resistance response. Recent research also indicates that some of the same proteins are involved in both extreme resistance and the hypersensitive response. Herein, we review and synthesize published studies on extreme resistance in potato and soybean, and describe studies in additional species, including model plant species, to highlight future research avenues that may bridge the gaps in our knowledge of plant antiviral defense mechanisms. [ABSTRACT FROM AUTHOR]

As the incidence of fungicide resistance in plant pathogens continues to increase, control of diseases and the management of resistance would be greatly aided by rapid diagnostic methods. Quantitative allele-specific PCR (ASqPCR) is an ideal technique for the in-field analysis of fungicide resistance as it can quantify the frequency of mutations in fungicide targets. We have applied this technique to the fungal pathogen Blumeria graminis f. sp. tritici (Bgt), the causal agent of wheat powdery mildew. In Australia, strobilurin-resistant Bgt was first discovered in 2016. Molecular analysis revealed a nucleotide transversion in the cytochrome b (cytb) gene in the cytochrome bc1 enzyme complex, resulting in a substitution of alanine for glycine at position 143 (G143A). We have developed an in-field ASqPCR assay that can quantify both the resistant (A143) and sensitive (G143) cytb alleles down to 1.67% in host and Bgt DNA mixtures, within 90 min of sample collection. The in situ analysis of samples collected during a survey in Tasmania revealed A143 frequencies ranging between 9–100%. Validation of the analysis with a newly developed laboratory based digital PCR assay found no significant differences between the two methods. We have successfully developed an in-field quantification method, for a strobilurin-resistant allele, by pairing the ASqPCR assay on a lightweight qPCR instrument with a quick DNA extraction method. The deployment of these type of methodologies in the field can contribute to the effective in-season management of fungicide resistance. [ABSTRACT FROM AUTHOR]

The leaf curl disease of Jatropha caused by geminiviruses results in heavy economic losses. In the present study, we report the identification of a new strain of a Jatropha leaf curl Gujarat virus (JLCuGV), which encodes six ORFs with each one having RNA silencing suppressor activity. Therefore, three artificial microRNAs (amiRNAs; C1/C4, C2/C3 and V1/V2) were designed employing overlapping regions, each targeting two ORFs of JLCuGV genomic DNA and transformed in tobacco. The C1/C4 and C2/C3 amiRNA transgenics were resistant while V1/V2 amiRNA transgenics were tolerant against JLCuGV. The relative level of amiRNA inversely related to viral load indicating a correlation with disease resistance. The assessment of photosynthetic parameters suggests that the transgenics perform significantly better in response to JLCuGV infiltration as compared to wild type (WT). The metabolite contents were not altered remarkably in amiRNA transgenics, but sugar metabolism and tricarboxylic acid (TCA) cycle showed noticeable changes in WT on virus infiltration. The overall higher methylation and demethylation observed in amiRNA transgenics correlated with decreased JLCuGV accumulation. This study demonstrates that amiRNA transgenics showed enhanced resistance to JLCuGV while efficiently maintaining normalcy in their photosynthesis and metabolic pathways as well as homeostasis in the methylation patterns. [ABSTRACT FROM AUTHOR]

Plant–virus interactions are greatly influenced by environmental factors such as temperatures. In virus‐infected plants, enhanced temperature is frequently associated with more severe symptoms and higher virus content. However, the mechanisms involved in such regulatory effects remain largely uncharacterized. To provide more insight into the mechanisms whereby temperature regulates plant–virus interactions, we analysed changes in the proteome of potato cv. Chicago plants infected with potato virus Y (PVY) at normal (22 °C) and elevated temperature (28 °C), which is known to significantly increase plant susceptibility to the virus. One of the most intriguing findings is that the main enzymes of the methionine cycle (MTC) were down‐regulated at the higher but not at normal temperatures. With good agreement, we found that higher temperature conditions triggered consistent and concerted changes in the level of MTC metabolites, suggesting that the enhanced susceptibility of potato plants to PVY at 28 °C may at least be partially orchestrated by the down‐regulation of MTC enzymes and concomitant cycle perturbation. In line with this, foliar treatment of these plants with methionine restored accumulation of MTC metabolites and subverted the susceptibility to PVY at elevated temperature. These data are discussed in the context of the major function of the MTC in transmethylation processes. [ABSTRACT FROM AUTHOR]

SUMMARY: Whereas the activation of resistance (R) proteins has been intensively studied, the downstream signaling mechanisms leading to the restriction of the pathogen remain mostly unknown. We studied the immunity network response conditioned by the potato Ny‐1 gene against potato virus Y. We analyzed the processes in the cell death zone and surrounding tissue on the biochemical and gene expression levels in order to reveal the spatiotemporal regulation of the immune response. We show that the transcriptional response in the cell death zone and surrounding tissue is dependent on salicylic acid (SA). For some genes the spatiotemporal regulation is completely lost in the SA‐deficient line, whereas other genes show a different response, indicating multiple connections between hormonal signaling modules. The induction of NADPH oxidase RBOHD expression occurs specifically on the lesion border during the resistance response. In plants with silenced RBOHD, the functionality of the resistance response is perturbed and the spread of the virus is not arrested at the site of infection. RBOHD is required for the spatial accumulation of SA, and conversely RBOHD is under the transcriptional regulation of SA. Using spatially resolved RNA‐seq, we also identified spatial regulation of an UDP‐glucosyltransferase, another component in feedback activation of SA biosynthesis, thus deciphering a novel aspect of resistance signaling. Significance Statement: To progress towards sustainable resistance in the field, it is important to understand the mechanisms of immune signaling, and therefore we studied the regulation of the resistance response of potato against potato virus Y on biochemical, ultrastructural and gene expression levels. We show that SA synthesis and NADPH oxidase RBOHD are interconnected through a regulatory loop that is required for the efficient spatial regulation of the resistance response against potato virus Y in potato. [ABSTRACT FROM AUTHOR]

The ecology of plant viruses began to be explored at the end of the 19th century. Since then, major advances have revealed mechanisms of virus-host-vector interactions in various environments. These advances have been accelerated by new technlogies for virus detection and characterization, most recently including high throughput sequencing (HTS). HTS allows investigators, for the first time, to characterize all or nearly all viruses in a sample without a priori information about which viruses might be present. This powerful approach has spurred new investigation of the viral metagenome (virome). The rich virome datasets accumulated illuminate important ecological phenomena such as virus spread among host reservoirs (wild and domestic), effects of ecosystem simplification caused by human activities (and agriculture) on the biodiversity and the emergence of new viruses in crops. To be effective, however, HTS-based virome studies must successfully navigate challenges and pitfalls at each procedural step, from plant sampling to library preparation and bioinformatic analyses. This review summarizes major advances in plant virus ecology associated with technological developments, and then presents important considerations and best practices for HTS use in virome studies. [ABSTRACT FROM AUTHOR]

Viral diseases and their damage causing significant loss to economically important crops have increased by several folds during the last decade. All the conventional approaches are not able to eradicate the viral infection. Therefore, there is a need to look for efficient and eco-friendly viral disease-preventive measures. The genomic material of the majority of deleterious viruses of higher plants is RNA. One of the possible measures to control viruses is the use of ribonucleases (RNases), which can cleave RNA in the viral genome. Based on this, we investigated the RNase activity of endophytic Bacillus spp., which can enrich in 103–105 colony-forming units per gram of wet mass of aboveground part of potato plants. A high level of RNase activity was observed in the culture medium of Bacillus thuringiensis B-6066, Bacillus sp. STL-7, Bacillus sp. TS2, and Bacillus subtilis 26D. B. thuringiensis B-5351 had low RNase activity but high ability to colonize internal plant tissues, Bacillus sp. STL-7 with high RNase activity have relatively low number of cells in internal tissues of plants. B. thuringiensis B-6066, B. subtilis 26D, and Bacillus sp. TS stimulate RNase activity in potato plants for a long time after application. Strains with high ability to colonize internal plant tissues combined with high RNase activity reduced severity of viral diseases symptoms on plants and reduced the incidence of potato viruses M, S, and Y. It is worth noting that Bacillus spp. under investigation reduced the number of Leptinotarsa decemlineata Say. egg clusters and larvae on treated plants and showed antifeedant activity. This results in increase of potato productivity mainly in the fraction of major tubers. B. subtilis 26D and Bacillus sp. TS2 combining endophytic lifestyle, RNase, and antifeedant activity may become the basis for the development of biocontrol agents for plant protection. [ABSTRACT FROM AUTHOR]

The emerald ash borer (EAB), Agrilus planipennis (Fairmaire), is the most destructive invasive insect species of ash (Fraxinus spp.) in North America. An accurate method for early detection of this noxious insect pest is indispensable to providing adequate warning of A. planipennis infestation. A loop-mediated isothermal amplification (LAMP) assay (EAB-LAMP) was developed based on mitochondrial cytochrome c oxidase subunit I (COI) gene. The EAB-LAMP required only 30 min at 65°C to amplify A. planipennis DNA from specimens collected from geographically distinct locations. There was no cross-reactivity with other Agrilus and insect species. The developed EAB-LAMP differentially detected traces of A. planipennis genome (COI) within frass from various Fraxinus species. EAB-LAMP was also able to distinguish among A. planipennis DNA and other Agrilus species and nontarget insect species in trap captures. By detecting A. planipennis DNA in two additional trap captures (in situ), the EAB-LAMP was more sensitive and reliable than visual inspection. We tested the quantitative nature of the assay by evaluating pooled trap samples and demonstrated that the EAB-LAMP was capable of functioning optimally using a pool size of at least five individual trap samples. This potentially circumvents the need to perform large-scale individual analysis for processing trap samples. Considering its performance, specificity, sensitivity, and repeatability, the developed EAB-LAMP could be a valuable tool to support strategy and operation of large-scale surveillance for A. planipennis and could profitably be used in routine monitoring programs for effective management of A. planipennis. [ABSTRACT FROM AUTHOR]

Colorado potato beetle (CPB) is an agricultural pest of solanaceous crops, notorious for its rapid resistance development to chemical pesticides. Foliar spraying of dsRNA formulations is a promising innovative technology providing highly specific and environmentally acceptable option for CPB management. We designed dsRNA to silence CPB mesh gene (dsMESH) and performed laboratory feeding trials to assess impacts on beetle survival and development. We compared the effectiveness of in vivo and in vitro produced dsRNA in a series of laboratory experiments. We additionally performed a field trial in which the efficacy of dsRNA sprayed onto potato foliage was compared to a spinosad-based insecticide. We showed that dsMESH ingestion consistently and significantly impaired larval growth and decreased larval survival in laboratory feeding experiments. In vivo produced dsRNA performed similarly as in vitro synthesized dsRNA in laboratory settings. In the field trial, dsMESH was as effective in controlling CPB larvae as a commercial spinosad insecticide, its activity was however slower. We discuss limitations and benefits of a potential dsMESH-based CPB management strategy and list some important RNAi based CPB research topics, which will have to be addressed in future. [ABSTRACT FROM AUTHOR]

Background Coronavirus disease 2019 (COVID-19) has become a public health emergency. The widely used reverse transcription–polymerase chain reaction (RT-PCR) method has limitations for clinical diagnosis and treatment. Methods A total of 323 samples from 76 COVID-19–confirmed patients were analyzed by droplet digital PCR (ddPCR) and RT-PCR based 2 target genes (ORF1ab and N). Nasal swabs, throat swabs, sputum, blood, and urine were collected. Clinical and imaging data were obtained for clinical staging. Results In 95 samples that tested positive by both methods, the cycle threshold (Ct) of RT-PCR was highly correlated with the copy number of ddPCR (ORF1ab gene, R 2 = 0.83; N gene, R 2 = 0.87). Four (4/161) negative and 41 (41/67) single-gene positive samples tested by RT-PCR were positive according to ddPCR with viral loads ranging from 11.1 to 123.2 copies/test. The viral load of respiratory samples was then compared and the average viral load in sputum (17 429 ± 6920 copies/test) was found to be significantly higher than in throat swabs (2552 ± 1965 copies/test, P  < .001) and nasal swabs (651 ± 501 copies/test, P  < .001). Furthermore, the viral loads in the early and progressive stages were significantly higher than that in the recovery stage (46 800 ± 17 272 vs 1252 ± 1027, P  < .001) analyzed by sputum samples. Conclusions Quantitative monitoring of viral load in lower respiratory tract samples helps to evaluate disease progression, especially in cases of low viral load. [ABSTRACT FROM AUTHOR]

Potato (Solanum tuberosum L.) is the worldwide most important nongrain crop after wheat, rice, and maize. The autotetraploidy of the modern commercial potato makes breeding of new resistant and high-yielding cultivars challenging due to complicated and time-consuming identification and selection processes of desired crop features. On the other hand, plant protection of existing cultivars using conventional synthetic pesticides is increasingly restricted due to safety issues for both consumers and the environment. Chitosan is known to display antimicrobial activity against a broad range of plant pathogens and shows the ability to trigger resistance in plants by elicitation of defense responses. As chitosan is a renewable, biodegradable and nontoxic compound, it is considered as a promising next-generation plant-protecting agent. However, the molecular and cellular modes of action of chitosan treatment are not yet understood. In this study, transcriptional changes in chitosan-treated potato leaves were investigated via RNA sequencing. Leaves treated with a well-defined chitosan polymer at low concentration were harvested 2 and 5 h after treatment and their expression profile was compared against water-treated control plants. We observed 32 differentially expressed genes (fold change ≥ 1; p-value ≤ 0.05) 2 h after treatment and 83 differentially expressed genes 5 h after treatment. Enrichment analysis mainly revealed gene modulation associated with electron transfer chains in chloroplasts and mitochondria, accompanied by the upregulation of only a very limited number of genes directly related to defense. As chitosan positively influences plant growth, yield, and resistance, we conclude that activation of electron transfer might result in the crosstalk of different organelles via redox signals to activate immune responses in preparation for pathogen attack, concomitantly resulting in a generally improved metabolic state, fostering plant growth and development. This conclusion is supported by the rapid and transient production of reactive oxygen species in a typical oxidative burst in the potato leaves upon chitosan treatment. This study furthers our knowledge on the mode of action of chitosan as a plant-protecting agent, as a prerequisite for improving its ability to replace or reduce the use of less environmentally friendly agro-chemicals. [ABSTRACT FROM AUTHOR]

A number of isothermal DNA amplification technologies claim to be ideal for point-of-need (PON) applications as they enable reactions to be performed using a single-temperature heat source (e.g. water bath). Thus, we examined several isothermal amplification methods focusing on simplicity, cost, sensitivity and reproducibility to identify the most suitable method(s) for low resource PON applications. A number of methods were found unsuitable as they either involved multiple temperature incubations, were relatively expensive or required relatively large amounts target DNA for amplification. Among the methods examined, loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) were found to be the most suitable for PON applications as they are both single step methods that provide highly sensitive and reproducible amplifications. The speed of LAMP reactions was greatly enhanced, up to 76%, with the addition of loop primers while the presence of swarm primers and the sequestration of free magnesium ions with nucleotides also enhanced the amplification speed. In contrast, we were unable to enhance RPA's performance from the original published literature. While both RPA and LAMP have some drawbacks, either isothermal technology can reliably be used for on-site diagnostics with minimal equipment. [ABSTRACT FROM AUTHOR]

Tomato brown rugose fruit virus (ToBRFV) is a member of Tobamovirus infecting tomato and pepper. Within North America, both the United States and Mexico consider ToBRFV to be a regulated pest. In Canada, the presence of ToBRFV has been reported, but an efficient diagnostic system has not yet been established. Here, we describe the development and assessment of a loop-mediated isothermal amplification (LAMP)-based assay to detect ToBRFV. The LAMP test was efficient and robust, and results could be obtained within 35 min with an available RNA sample. Amplification was possible when either water bath or oven were used to maintain the temperature at isothermal conditions (65°C), and results could be read by visual observation of colour change. Detection limit of the LAMP was eight target RNA molecules. Under the experimental conditions tested, LAMP was as sensitive as qPCR and 100 times more sensitive than the currently used RT-PCR. We recommend this sensitive, efficient LAMP protocol to be used for routine lab testing of ToBRFV. [ABSTRACT FROM AUTHOR]

BACKGROUND Plant viral infections induce changes in metabolic components in the host plant, with potential effects on compositional, organoleptic and storability features of agricultural products. Identification of modulated metabolites may provide clues concerning pathways implementing responses in plant–pathogen interactions. A time course study of metabolic fingerprinting of onion yellow dwarf virus (OYDV)‐infected versus healthy 'Rossa di Tropea' onion bulbs was performed using proton high‐resolution magic angle spinning nuclear magnetic resonance (1H HR‐MAS NMR) and ultra‐performance liquid chromatography (UPLC), providing an overview of the metabolic state of the bulb in response to OYDV infection during storage. RESULTS: Metabolites accumulated/depleted upon infection were identified, belonging to flavonoid, saccharide, amino acid and organic acid classes. A decrease in quercetin glucosides content and antioxidant activity was observed in infected bulbs; some amino acids (Arg, Asn, Phe, Val) accumulated, while others were depleted (Leu); for some metabolites, a bimodal time‐course was observed during storage (Glc, Lys). Virus interference on metabolic pathways, and the effects of the metabolic shift on edible product storability, organoleptic and nutritional quality were discussed. CONCLUSIONS: OYDV infection induces a metabolic shift in 'Rossa di Tropea' onion during bulb storage, involving several pathways and affecting storability and organoleptic and nutritional quality of bulbs at marketable stage. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Investigation of diseases caused by phytoplasmas, a group of cell-wall-less gram-positive bacteria has received significant attention in plant pathology. Grapevine is a host of two, genetically distinct phytoplasmas: Line Flavescence dorée (FD) phytoplasma associated to 'flavescence dorée' and 'Candidatus Phytoplasma solani' responsible for 'bois noir' (BN) disease. In the current study, we focused on BN diseased grapevines (Vitis vinifera L. cv. 'Kékfrankos'), measured their photosynthetic performance and leaf hydrogen peroxide (H2O2) concentration. The latter is generally considered as a key molecule in the process of 'recovery' which is a spontaneous and unpredictable long-term remission of disease symptoms. This phenomenon also occurred during the time of our experiment. Infection resulted in reduced gas exchange performance and maximum quantum efficiency of PSII with an increased regulated non-photochemical quenching of PSII and H2O2 concentration. Changes in gas exchange seem to be a systemic response, while reduced photochemistry is a local response to 'Ca. P. solani' infection. H2O2 accumulation in BN phytoplasma infected plants, unlike in FD disease, was found to be a typical response to the appearance of a biotic stressor. [ABSTRACT FROM AUTHOR]

Under the field conditions crop plants interact with diverse microorganisms. These include beneficial (symbiotic) and phytopathogenic microorganisms, which jointly affect growth and productivity of the plants. In last decades, production of potato (Solanum tuberosum L.) suffers from increased incidence of potato virus Y (PVY), which is one of most important potato pests. Arbuscular mycorrhizal fungi (AMF) are common symbionts of potato, however the impact of mycorrhizal symbiosis on the progression of PVY-induced disease is scarcely known. Therefore, in the present study we investigated the effect of joint PVY infection and mycorrhizal colonization by Rhizophagus irregularis on growth traits of the host potato plant (cv. Pirol). The tested PVY isolate belonged to N-Wilga strain group, which is considered to be predominant in Europe and many other parts of the world. The viral particles were concentrated in the leaves, but decreased the root growth. Furthermore, the infection with PVY evoked prolonged oxidative stress reflected by increased level of endogenous H2O2. AMF alleviated oxidative stress in PVY-infected host plants by a substantial decrease in the level of shoot- and root-derived H2O2, but still caused asymptomatic growth depression. It was assumed that mycorrhizal symbiosis of potato might mask infection by PVY in field observations. [ABSTRACT FROM AUTHOR]

Copyright of Canadian Journal of Plant Pathology is the property of Taylor & Francis Ltd and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The genome content of adeno-associated virus (AAV) vectors is critical to the safety and potency of AAV-based gene therapy products. Empty capsids are considered a product-related impurity and a critical quality attribute (CQA) of the drug product, thus requiring characterization throughout the production process to demonstrate they are controlled to acceptable levels in the final drug product. Anion exchange chromatography has been used to achieve separation between empty and full capsids, but requires method development and gradient optimization for different serotypes and formulations. Here, we describe an alternative approach to quantitation that does not rely on achieving separation between empty and full capsids, but instead uses the well-established relationship between absorbance at UV A 260 /A 280 and relation to DNA/protein content, in combination with anion-exchange chromatography to allow one to calculate the relative proportion of empty and full capsids in AAV samples from a single peak. We call this approach ACUVRA: Anion-exchange Chromatography UV-Ratio Analysis, and show the applicability of the method through a case study with recombinant AAV2 (rAAV2) process intermediates and drug substance. Method qualification and GMP validation in a quality control (QC) laboratory results show that ACUVRA is a fit-for-purpose method for process development support and characterization, while also being a QC-friendly option for GMP release testing at all stages of clinical development. Graphical abstract., (© 2022. The Author(s).)

Maize Iranian mosaic virus (MIMV) infects several gramineous plants and is an economically important nucleorhabdovirus in Iran. Maize responds to MIMV infection at the transcriptional level. Alternative splicing (AS) is a mechanism that generates multiple mRNAs from a single pre-mRNA, often encoding protein isoforms with functional differences. We carried out genome-wide analysis of AS responses to MIMV in maize seedlings and identified genes involved in this molecular response. The AS events we investigated included skipped exons, alternative 3′ splice site, alternative 5′ splice site, mutually exclusive exons, and retained introns. In total 10,881 maize genes showed AS, of which 601 genes were involved in response to MIMV-infection and 186 were found only in uninfected maize. AS was identified in some of the genes that are involved in disease resistance or pathogenicity pathways. We demonstrated that in MIMV-infects maize, host genes that are involved in symptom development, virus multiplication, resistance to pathogens and host-pathogen interaction are affected by AS mechanism. Gene network analysis showed that ten genes represent the hubs for the protein network in maize and that they are involved in response to pathogen attack and include 26S proteasome, 14–3-3-like protein A, Rop family, mitogen-activated protein kinase, ubiquitin and serine/threonine-protein kinases. In conclusion, we showed that AS occurs as a transcriptional regulatory mechanism in maize response to MIMV infection and we identified genes that have the key roles in pathogenicity pathways that were differentially spliced in infected seedlings. [ABSTRACT FROM AUTHOR]

Citrus stubborn disease (CSD) is caused by a Gram-positive bacterium, Spiroplasma citri, and is an endemic, but manageable, citrus disease. CSD-affected trees are low yielding but a key problem is that its symptoms are similar to and easily mistaken for Huanglongbing (HLB), a devastating citrus disease controlled by quarantine and eradication. Therefore, a rapid and simple test for S. citri is needed to readily distinguish CSD from HLB. To this end, a Loop-Mediated Isothermal Amplification technique (LAMP) was developed to detect S. citri, targeting the spiralin gene. The protocol was optimized for crude plant extracts from infected trees to allow on-site field testing. The LAMP assay showed high specificity to S. citri and detected DNA to a level of 100 fg/μl with no inhibition by crude plant extracts. Although the LAMP assay was 9 times less sensitive than qPCR with purified DNA templates, it performed well in field validations using a portable BioRanger device with citrus crude extracts. The LAMP assay showed detection efficiency and percentage yes/no calls similar to those obtained by real time PCR conducted with DNA extracted and purified from the same sample. The LAMP procedure allows growers, pest control or diagnostic services to rapidly test for S. citri in the field without a laboratory or DNA purification. [ABSTRACT FROM AUTHOR]

While one of the biggest problems we are facing today is water scarcity, enormous quantities of water are still being used in irrigation. If contaminated, this water can act as an effective pathway for the spread of disease-causing agents, like viruses. Here, we present a novel, environmentally friendly method known as cold atmospheric plasma for inactivation of viruses in water used in closed irrigation systems. We measured the plasma-mediated viral RNA degradation as well as the plasma-induced loss of viral infectivity using potato virus Y as a model virus due to its confirmed water transmissibility and economic as well as biological importance. We showed that only 1 min of plasma treatment is sufficient for successful inactivation of viruses in water samples with either high or low organic background. The plasma-mediated inactivation was efficient even at markedly higher virus concentrations than those expected in irrigation waters. Obtained results point to reactive oxygen species as the main mode of viral inactivation. Our laboratory-scale experiments confirm for the first time that plasma has an excellent potential as the eukaryotic virus inactivation tool for water sources and could thus provide a cost-effective solution for irrigation mediated plant virus transmission. The outstanding inactivation efficiency demonstrated by plasma treatments in water samples offers further expansions of its application to other water sources such as reused wastewater or contaminated drinking waters, as well as other plant, animal, and human waterborne viruses, ultimately leading to the prevention of water scarcity and numerous human, animal, and plant infections worldwide. [ABSTRACT FROM AUTHOR]

Copyright of Journal of Cultivated Plants / Journal für Kulturpflanzen is the property of Verlag Eugen Ulmer and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Trichomes, also simply referred to as hairs, are fine outgrowths of epidermal cells in many organisms including plants and bacteria. Plant trichomes have long been known for their multiple beneficial roles, ranging from protection against insect herbivores and ultraviolet light to the reduction of transpiration. However, there is increasing evidence that the presence of trichomes may have detrimental consequences for plants. For example, plant pathogenic bacteria can enter hosts through the open bases or broken stalks of damaged trichomes. Similarly, trichomes are considered a preferred site for fungal infection, and in this regard, the colonization and penetration of trichomes by fungi and oomycetes have been visualized using light, fluorescence, and scanning electron microscopy in a variety of plants from grasses to shrubs and trees. In addition to parasitic interactions, trichomes also form a host site for endophytic relationships with fungi, thereby serving as an unusual fungal niche. The replication and presence of plant viruses in trichomes have also been confirmed after inoculation. In contrast, the well-known beneficial Azolla–Anabaena symbiosis is facilitated through epidermal trichomes of the seedless vascular plant Azolla. These observations indicate that plant trichomes are involved in multiple interactions in terms of providing microbial habitats and infection sites as well as functioning as protective structures. Trichome-related microbial parasitism and endophytism can, in many ways, be considered comparable to those associated with root hairs. [ABSTRACT FROM AUTHOR]

Phytoplasmas are bacterial plant pathogens that need a plant host and an insect vector for their spread and survival. In plants, the physiological responses that phytoplasmas trigger result in symptom development through effects on hormonal, nutritional, and stress signaling pathways, and the interactions between these. In this review, recent advances on the involvement of plant hormones together with their known and deduced roles in plants infected with phytoplasmas are discussed. Several studies have directly, or in many cases indirectly, addressed plant hormone systems in phytoplasma-infected plants. These have provided accumulating evidence that phytoplasmas extensively affect plant hormone pathways. Phytoplasmas thus, with disturbing complex plant hormone networks, suppress plant immunity and modify plant structure, while optimizing their nutrient acquisition and facilitating their colonization of the plants, and their dissemination among plants by their insect vectors. [ABSTRACT FROM AUTHOR]

Specific scope: This Standard describes a diagnostic protocol for the generic detection of phytoplasmas. This Standard should be used in conjunction with PM 7/76 Use of EPPO diagnostic protocols Specific approval and amendment: Approved in 2018–09. This diagnostic protocol was prepared on the basis of the IPPC diagnostic protocol adopted in 2016 (Appendix 12 to ISPM 27). Although this EPPO Diagnostic Standard differs in terms of format it is fully consistent with the content of the IPPC Standard. [ABSTRACT FROM AUTHOR]

Aphids have serious damage and well known of the most important pests, which may transmit different viruses on potato. Potato has immense significance as a food crop throughout the world. In the last research, the most important and different aspects of predominant potato varieties; Agria, Arinda, Fontane, Banba and Sante were studied during 2015–2016 in Karaj, Iran. This research has been conducted into several disparate categories: the effect of environmental conditions on the physiological trends, aphid characteristics and virus infection. The results showed that potato fields had an increasing trend to the end of season in the aspect of infestation by Aphis gossypii Glover and Myzus persicae Sulzer. The potato cultivars showed the same reaction in the lab and no host preference was observed between them against cotton aphid (A. gossypii). Finally, Arinda, and Agria varieties had the most photosynthesis activity, chlorophyll concentration, and leaf dry weight values and had the least electrical conductivity and susceptibility to some potato virus diseases. These results may help to provide better potato varieties by considering to the important physiological characteristic related to some destructive pests. [ABSTRACT FROM AUTHOR]

Accurate management practices in crop health and food safety are critical, especially regarding the detection of plant pathogens in the early stages of a disease. To date, specific, fast and sensitive technologies for point‐of‐care diagnosis and simple or grower‐friendly devices are very valuable, as no specialized staff are required for diagnosing a disease in the field. This is especially the case today, when factors such as climate change may cause the appearance of pathogens in areas where years ago they were unexpected. The aim of this research is to review some of the promising techniques that can be applied to in‐field molecular detection of plant pathogens and how these techniques can change the way farmers and pathologists are diagnosing plant diseases. Some of them, like loop‐mediated isothermal amplification and recombinase polymerase amplification, are already being successfully used for routine diagnosis. However, most technologies still need validation in the plant pathology field, where they have a promising future for in‐field diagnosis when combined with simple DNA extraction methods, reagent stabilization techniques and their integration into portable devices. [ABSTRACT FROM AUTHOR]

Micro(mi)RNAs play crucial roles in plant developmental processes and in defense responses to biotic and abiotic stresses. In the last years, many works on small RNAs in grapevine (Vitis spp.) were published, and several conserved and putative novel grapevine-specific miRNAs were identified. In order to reorganize the high quantity of available data, we produced “miRVIT,” the first database of all novel grapevine miRNA candidates characterized so far, and still not deposited in miRBase. To this aim, each miRNA accession was renamed, repositioned in the last version of the grapevine genome, and compared with all the novel and conserved miRNAs detected in grapevine. Conserved and novel miRNAs cataloged in miRVIT were then used for analyzing Vitis vinifera plants infected by Flavescence dorée (FD), one of the most severe phytoplasma diseases affecting grapevine. The analysis of small RNAs from healthy, recovered (plants showing spontaneous and stable remission of symptoms), and FD-infected “Barbera” grapevines showed that FD altered the expression profiles of several miRNAs, including those involved in cell development and photosynthesis, jasmonate signaling, and disease resistance response. The application of miRVIT in a biological context confirmed the effectiveness of the followed approach, especially for the identification of novel miRNA candidates in grapevine. miRVIT database is available at http://mirvit.ipsp.cnr.it. Highlights: The application of the newly produced database of grapevine novel miRNAs to the analysis of plants infected by Flavescence dorée reveals key roles of miRNAs in photosynthesis and jasmonate signaling. [ABSTRACT FROM AUTHOR]

Grapevine fanleaf virus (GFLV) causes grapevine fanleaf degeneration, one of the oldest known viral diseases of grapevines. The virus has been found in all winegrowing regions around the world. In the seasons 2011–12 a comparison between field grown GFLV-infected and healthy grapevines was conducted for the cultivars Schioppettino in North-Eastern Italy and Refošk in South-Western Slovenia. Our research showed that GFLV infection caused a drop of the yield due to reduction of both cluster weight and berry weight. Besides the yield, the berry composition was also affected; in detail, anthocyanin concentration increased in both varieties but significantly only in the case of Schioppettino. Upregulation of the F3050H gene and downregulation of F30H gene in the berries of GFLV infected vines compared with the ones of healthy control vines resulted in modified proportions between di- and tri- hydroxylated or methylated derivatives of anthocyanins. The F3H1 gene was identified to be the most strongly regulated gene of the flavonoid biosynthetic pathway by GFLV infection, indicating its important role in increasing anthocyanin concentration in grapes of GFLV infected vines as compared with healthy controls. [ABSTRACT FROM AUTHOR]

RNA viruses have a great potential for high genetic variability and rapid evolution that is generated by mutation and recombination under selection pressure. This is also the case of Potato virus Y (PVY), which comprises a high diversity of different recombinant and non-recombinant strains. Consequently, it is hard to develop reverse transcription real-time quantitative PCR (RT-qPCR) with the same amplification efficiencies for all PVY strains which would enable their equilibrate quantification; this is specially needed in mixed infections and other studies of pathogenesis. To achieve this, we initially transferred the PVY universal RT-qPCR assay to a reverse transcription droplet digital PCR (RT-ddPCR) format. RT-ddPCR is an absolute quantification method, where a calibration curve is not needed, and it is less prone to inhibitors. The RT-ddPCR developed and validated in this study achieved a dynamic range of quantification over five orders of magnitude, and in terms of its sensitivity, it was comparable to, or even better than, RT-qPCR. RT-ddPCR showed lower measurement variability. We have shown that RT-ddPCR can be used as a reference tool for the evaluation of different RT-qPCR assays. In addition, it can be used for quantification of RNA based on in-house reference materials that can then be used as calibrators in diagnostic laboratories. [ABSTRACT FROM AUTHOR]

Virus‐induced systemic necrosis accounts for great yield losses in potato (Solanum tuberosum) cultivars susceptible to the necrotic strain of potato virus Y (PVYNTN). To extend understanding of systemic necrosis, the tissue of healthy and PVYNTN‐infected potato tubers (necrotic and non‐necrotic) during storage was profiled for host gene expression using microarrays while virus concentration was determined using RT‐qPCR. Comparison of non‐necrotic tissue types revealed minor transcriptional differences, while tuber necrosis was characterized by high virus concentration and pronounced expression of defence response‐associated genes. Creation of programmed cell death‐associated gene ontology enabled assessment of expression and regulation of relevant differentially expressed genes implicated in the execution of plant programmed cell death. Vacuolar cell death and unfolded protein response to viral infection were implicated in the necrotic tuber tissue by transcriptional induction of genes possessing caspase‐like activity and genes coding for endoplasmic reticulum chaperones and folding proteins. A meta‐analysis, including all relevant publicly available gene expression datasets differing in genotype, pathogen interaction type and potato organ, revealed common defence responses associated with cell wall fortification, oxidative and biotic stress. The response in necrotic tissues of susceptible tubers resembled the hypersensitive response of resistant primary infected plants, but with hormonal signalling as an important differentiating factor. This study will be useful for in‐depth analysis of programmed cell death regulation in plants by providing gene ontology. It specifically addresses the issue of systemic necrosis formation in PVYNTN‐infected potato tubers during storage. [ABSTRACT FROM AUTHOR]

Abstract: BACKGROUND: Rapid genetic on‐site identification methods at points of entry, such as seaports and airports, have the potential to become important tools to prevent the introduction and spread of economically harmful pest species that are unintentionally transported by the global trade of plant commodities. This paper reports the development and evaluation of a loop‐mediated isothermal amplification (LAMP)‐based identification system to prevent introduction of the three most frequently encountered regulated quarantine insect species groups at Swiss borders, Bemisia tabaci, Thrips palmi and several regulated fruit flies of the genera Bactrocera and Zeugodacus. RESULTS: The LAMP primers were designed to target a fragment of the mitochondrial cytochrome c oxidase subunit I gene and were generated based on publicly available DNA sequences. Laboratory evaluations analysing 282 insect specimens suspected to be quarantine organisms revealed an overall test efficiency of 99%. Additional on‐site evaluation at a point of entry using 37 specimens performed by plant health inspectors with minimal laboratory training resulted in an overall test efficiency of 95%. During both evaluation rounds, there were no false‐positives and the observed false‐negatives were attributable to human‐induced manipulation errors. To overcome the possibility of accidental introduction of pests as a result of rare false‐negative results, samples yielding negative results in the LAMP method were also subjected to DNA barcoding. CONCLUSION: Our LAMP assays reliably differentiated between the tested regulated and non‐regulated insect species within <1 h. Hence, LAMP assays represent suitable tools for rapid on‐site identification of harmful pests, which might facilitate an accelerated import control process for plant commodities. © 2018 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Abstract: Sustainable disease management depends on the ability to monitor the development of fungicide resistance in pathogen populations. A point mutation resulting in an alteration (F200Y) at codon 200 of the target protein β‐tubulin leads to a moderate level of resistance to carbendazim in Botrytis cinerea. Although traditional methods remain a cornerstone in detection of fungicide resistance, molecular methods that do not require the isolation of pathogens, can detect the presence of resistance alleles at low frequencies, and require less time and labour than traditional methods. In this study, we present an efficient, rapid, and highly specific method for detecting the moderately carbendazim‐resistant isolates in B. cinerea based on loop‐mediated isothermal amplification (LAMP). By using specific LAMP primers, we detected the resistance‐conferring mutation underlying β‐tubulin F200Y. The concentrations of LAMP components and LAMP parameters were optimised, resulting in reaction temperatures and times of 61–65°C and 45 min, respectively. The feasibility of the LAMP assay was verified by assaying the diseased samples with artificial inoculation in the different hosts. The LAMP assay developed in the current study was specific, stable, repeatable and sensitive, and was successfully applied for detection of moderately carbendazim‐resistant isolates of B. cinerea in plant samples. [ABSTRACT FROM AUTHOR]