Leaf Disc Assay Research Articles

Photosynthetic Responses to Salt Stress in Two Rice (Oryza sativa L.) Varieties

Assessing salt tolerance in plants under field conditions is a challenging task. The objective of this research was to assess the effectiveness of different methods (leaf disc assay and pot experiment) for evaluating salt tolerance in rice. Using two varieties with different salt tolerance, Changmaogu (CM) and 9311, under three NaCl levels (0, 0.3%, and 1.0%), we evaluated the photosynthetic performance in terms of chlorophyll content in leaf disc assays, as well as the photosynthetic rate (Pn), chlorophyll content, linear electron flow (LEF), and non-photochemical quenching (NPQ), in a semi-controlled pot experiment. In the leaf disc assay, CM showed a smaller decrease in chlorophyll content compared to 9311, especially under 1.0% salinity. Simultaneously, in the pot experiment, the CM variety employed flexible photosynthetic strategies, actively decreasing LEF and Pn after 5 days of salt stress (day 5) and then increasing photosynthetic capacity (chlorophyll content, LEF, and Pn) on day 10. Notably, the total chlorophyll content for the CM variety under 1.0% salinity was significantly higher than in the control, showing a 25.0% increase. Additionally, CM demonstrated NPQt sensitivity under 0.3% salinity, requiring an LEF of 150 to achieve an NPQt value of 3.0, compared to an LEF of 180 in the control. These results suggest that a simple leaf disc assay may not fully capture the adaptive mechanisms of rice plants under salinity stress. Therefore, we advocate for the use of more comprehensive methods, such as outdoor pot or field experiments, to gain a deeper understanding and more accurate evaluation of salt tolerance in rice.

Foliar Resistance to a Natural Population of Plasmopara viticola in Some Grapevine Varieties Grown in Türkiye

ABSTRACTIt is critical to use a population of the pathogen that contains different strains to determine the resistance of grapevine varieties to the downy mildew pathogen Plasmopara viticola in a specific region. This study tested six local grape varieties (Çavuş, Kalecik Karası, Müşküle, Papaz Karası, Sultana and Yapıncak), eight Turkish registered varieties (Barış, Bozbey, Güzgülü, Özer Karası, Reçel Üzümü, Tekirdağ Çekirdeksizi, Trakya İlkeren and Yalova İncisi) and seven varieties of foreign origin (Cabernet Sauvignon, Cardinal, Chardonnay, Cinsault, Isabella, Semillon and Gamay) using detached leaf and leaf disc assays with pathogen populations collected from vineyards. There was a significant correlation for disease severity between the two assay methods. No sporulation was observed on the varieties Çavuş and Isabella in both assay methods. The variety Yapıncak had very low disease severity (2.00%) in the leaf disc assay, and the pathogen did not sporulate on this variety in the detached leaf assay. Cluster analysis was performed to determine the resistance level of the varieties against the pathogen, as different disease severities were observed in some varieties depending on the test method used, and the 21 varieties were classified into four clusters. Barış, Bozbey, Cardinal, Çavuş, Isabella, Semillon and Yapıncak in Cluster I exhibited the lowest disease severity mean (2.88%), while those in Cluster 3, including Cabernet Sauvignon, Güzgülü, Papaz Karası and Yalova Incisi, had the highest disease severity mean (56.75%). These varieties in Clusters 1 and 3 were considered highly resistant and highly susceptible, respectively. Leaf hair density, as reported in official variety descriptions, did not associate with reduced disease severity in the two assays. This study assessed the response of grapevine varieties to the P. viticola population in the region for the first time. The highly resistant and resistant varieties identified will provide new material for breeders and contribute to the improvement of organic viticulture in the area.

The effect of natural herbicide from Fusarium equiseti crude extract on the aquatic weed water hyacinth (Eichornia crassipes (Mart.) Solms)

In this study, Fusarium equiseti was isolated from the weed plant Tridax procumbens in an agricultural field and a crude extract produced with 75% ethanol for use as active ingredient material in natural herbicides. The herbicidal effect of F. equiseti extract was tested on water hyacinth (Eichornia crassipes), an invasive aquatic weed, by leaf disk assay at concentrations of 0.05%, 0.1%, and 0.2% w/v crude extract. Dose-dependent visual toxicity symptoms were evident after three days, namely chlorosis, yellow leaves surrounded by dark brown edges. Photosynthetic pigments (chlorophyll a, b, and carotenoids) and membrane integrity (as electrolyte leakage and malondialdehyde content) were evaluated following the leaf disk test. 3 days after treatment, photosynthetic pigment contents showed dose-dependent decreases, while both measures of membrane integrity showed dose-dependent increases with increasing extract concentration. In addition, a cytogenetic assay was conducted on Allium cepa L. root, in which mitotic index reduction and depigmentation were evident as early as 24 h after herbicide application. Finally, anatomical analysis of treated E. crassipes leaves revealed degradation or damage of the ground tissue. All told, our results support the F. equiseti crude-based natural herbicide cloud as a sustainable alternative in agriculture.

Pathogenicity of Lasiodiplodia theobromae isolated from cocoa dieback disease in South Sulawesi, Indonesia

AbstractDieback disease is one of the constraints to cocoa production in Indonesia. A newly identified dieback symptom has been observed in cocoa farms in East Luwu since 2015 and later observed in Luwu, Pinrang, Enrekang and Soppeng Regency, South Sulawesi. The potential causal agent identity was determined using morphological characteristics, DNA sequences, and phylogenetic analyses. A total of four fungal isolates out of seven isolates that were recovered from diseased stems morphologically resembled Lasiodiplodia species. ITS and EF1α sequencing confirmed that three of the Lasiodiplodia isolates were Lasiodiplodia theobromae, and one isolate was Lasiodiplodia pseudotheobromae. The aggresiveness of all fungal isolates and control was further determined by a leaf disk assay. Experiments showed that L. theobromae isolate CAS0321 was the most aggressive isolate in a leaf disk assay, while other Lasiodiplodia isolates, Fusarium isolates and Diaporthe isolate, were less aggressive. L. theobromae isolate CAS0321 was further used for a pathogenicity test on cocoa seedlings. Two months after inoculation, L. theobromae isolate CAS0321 caused elongated streaks on seedling stems with dark brown to black streaking of vascular tissues similar to that observed in natural infections. This is the first report of a virulent isolate of L. theobromae associated with cocoa dieback in Sulawesi.

Pesticidal activity of extra cellular metabolites extracted from the green muscardian fungi Metarhizium anisopliae

AbstractThe present study was undertaken to screen the pesticidal activity of extra cellular metabolites derived from entomopathogenic fungus Metarhizium anisopliae against Spodoptera litura and Aphis crassivora an important aphid pest under laboratory and pot culture condition. Fungal strain used in the study was isolated from the soil by serial dilution technique. Isolated fungal strain was mass multiplied in a liquid medium sabouraud maltose yeast extract broth under optimum condition (28°C). Culture filtrate and the collected culture derived from the medium was used for the source of metabolites. The culture filtrate was extracted with ethyl acetate, acetone and the concentrated extract was screened for pesticidal activity against Spodoptera litura and Aphis crassivora adopting leaf disc and pot culture assay respectively. Extracted metabolites showed pesticidal activity against both insect pests. However, notable effect was recorded in ethyl acetate derived metabolites. The ethyl acetate extract treatment recorded 91.2% mortality at a concentration of 100 μg against Spodoptera litura and 100% mortality at a concentration of 100 μg against Aphis crassivora. Phytotoxicity was determined by measuring the germination index (GI) of groundnut seeds by exposing it to the crude metabolites extracted in ethyl acetate and acetone. The results showed that the germination index was high on ethyl acetate extract which was recorded to be 98.0% and germination index of acetone extract was found to be 90.0%. Production of insecticidal metabolites from M. anisopliae under laboratory condition proposed the possible utilization of metabolites as an effective, safe pesticidal agent against economic important insect pests. No impact on the seedling emergence revealed non phytotoxic effect of the metabolites.

Exploring host plant resistance in chilli (Capsicum spp) against Southeast Asian thrips (Thripsparvispinus Karny)

Southeast Asian thrips (Thrips parvispinus Karny), poses a significant threat to chilli crop production causing severe economic losses. They feed heavily on leaves, flowers and young fruits causing severe damage to the crop. Exploring host plant resistance is the best ecofriendly management strategy to combat this pest. Large-scale field screening of 481 germplasm accessions, followed by further screening of selected accessions under polyhouse conditions led to identification of fourteen accessions that showed resistance against Southeast Asian thrips. Among the screened accessions, IIHR-B-HP-79 showed the highest level of Southeast Asian thrips resistance with significantly low leaf and fruit damage scores of 0.28 and 0.13, respectively. Further, screening of selected fourteen field resistant accessions was performed through whole plant bio-assay where in four accessions i.e., IIHR-B-HP-79 (C. frutescens), IIHR 4550, IIHR 4501 & IIHR 4410 (C. chinense) showed good level of resistance. Multi-choice leaf disc assay, demonstrated that IIHR-B-HP-79 exhibited a high level of resistance with the lowest percent of scraping damage during both the vegetative (10.98) and flowering stages (15.31). The identified resistant accession IIHR-B-HP-79 (C. frutescens), has the potential to be a valuable genetic resource as donor in breeding for Southeast Asian thrips resistance.

Efficacy of ergosterol peroxide obtained from the endophytic fungus Acrophialophora jodhpurensis against Rhizoctonia solani.

To investigate antifungal activity of the extract and major metabolite of the endophytic fungus Acrophialophora jodhpurensis (belonging to Chaetomiaceae) against crown and root rot caused by Rhizoctonia solani (teleomorph: Thanatephorus cucumeris), as an important pathogen of tomato. The endophytic fungus A. jodhpurensis, has high inhibitory effect against R. solani AG4-HG II in vitro and in vivo. The media conditions were optimized for production of the endophyte's metabolites. The highest amounts of secondary metabolites were produced at pH 7, 30°C temperature, and in the presence of 0.5% glucose, 0.033% sodium nitrate, and 1 gl-1 asparagine as the best carbon, nitrogen, and amino acid sources, respectively. The mycelia were extracted by methanol and the obtained extract was submitted to various chromatography techniques. Phytochemical analysis via thin-layer chromatography (TLC) and nuclear magnetic resonance (NMR) spectroscopy showed that ergosterol peroxide was the major component in the extract of this endophyte. Antifungal activities of the methanolic extract and ergosterol peroxide in the culture media were studied against R. solani. Minimum inhibitory concentrations of the extract and ergosterol peroxide against the pathogen were 600 and 150µg ml-1, respectively. Ergosterol peroxide revealed destructive effects on the pathogen structures in microscopic analyses and induced sclerotia production. Histochemical analyses revealed that it induced apoptosis in the mycelia of R. solani via superoxide production and cell death. Application of ergosterol peroxide in the leaf disc assay reduced the disease severity in tomato leaves. Antifungal metabolites produced by A. jodhpurensis, such as ergosterol peroxide, are capable of controlling destructive Rhizoctonia diseases on tomato.

Newly Isolated Trichoderma spp. Show Multifaceted Biocontrol Strategies to Inhibit Potato Late Blight Causal Agent Phytophthora infestans both In Vitro and In Planta

Potato growers worldwide have been at war for more than 150 years with an enemy whose lifecycle, genome size and architecture, infection rate, and economic impacts are the epitome of a plant pathogen. Phytophthora infestans is an oomycete that causes the notorious late blight infection in potato and tomato fields. This study explored the benefits of the multitalented plant symbiotic fungi Trichoderma spp. and their metabolites as potential biopesticides against P. infestans. Eleven strains of Trichoderma spp. were obtained from soil and tree barks and were identified using DNA sequence analysis of three molecular markers. The antagonistic potential of the strains against P. infestans was first evaluated in vitro. In dual-culture assays, P. infestans growth was significantly inhibited (53 to 95%) by different Trichoderma spp. through direct mycoparasitism, competition for space and nutrients, or antibiosis. The cell-free filtrates (CFFs) of different Trichoderma strains were obtained and characterized for anti- Phytophthora activities as well as biochemical stability. The obtained results indicated that Trichoderma CFFs were chemically stable and strongly decreased P. infestans’ mycelial growth and zoospore motility and viability. Similarly, in leaf-disk assays, Trichoderma CFFs showed significant protection against P. infestans infection. Ultraperformance liquid chromatography analysis revealed the presence of harzianic acid, iso-harzianic acid, and 6-pentyl-2H-pyran-2-one as major compounds in different Trichoderma CFFs. Furthermore, selected Trichoderma strains significantly protected potato plants against soil-mediated late blight infection. Finally, Trichoderma spp. showed high compatibility with a copper-based fungicide, especially at lower concentrations, suggesting that both protective agents could be combined in integrated pest management programs. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Development of a soybean leaf disc assay for determining oral insecticidal activity in the lepidopteran agricultural pest Helicoverpa armigera

Pest insects pose a heavy burden on global agricultural industries with small molecule insecticides being predominantly used for their control. Unwanted side effects and resistance development plagues most small molecule insecticides such as the neonicotinoids, which have been reported to be harmful to honeybees. Bioinsecticides like Bacillus thuringiensis (Bt) toxins can be used as environmentally-friendly alternatives. Arachnid venoms comprise another promising source of bioinsecticides, containing a multitude of selective and potent insecticidal toxins. Unfortunately, no standardised insect models are currently available to assess the suitability of insecticidal agents under laboratory conditions. Thus, we aimed to develop a laboratory model that closely mimics field conditions by employing a leaf disk assay (LDA) for oral application of insecticidal agents in a bioassay tray format. Neonate larvae of the cotton bollworm (Helicoverpa armigera) were fed with soybean (Glycine max) leaves that were treated with different insecticidal agents. We observed dose-dependent insecticidal effects for Bt toxin and the neonicotinoid insecticide imidacloprid, with imidacloprid exhibiting a faster response. Furthermore, we identified several insecticidal arachnid venoms that were active when co-applied with sub-lethal doses of Bt toxin. We propose the H. armigera LDA as a suitable tool for assessing the insecticidal effects of insecticidal agents against lepidopterans.

Baseline Susceptibility to a Novel dsRNA-Based Insecticide across US Populations of Colorado Potato Beetle

The Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae) is an insect defoliator of the potato, Solanum tuberosum L. This species thrives in agricultural environments because of its flexible and complex life history, as well as its ability to evolve insecticide resistance. As a result, it has become a widely distributed agricultural pest. Ledprona (trade name Calantha) is a recently developed novel double-stranded RNA (dsRNA) insecticide that controls populations of Colorado potato beetle through RNA interference (RNAi). Previous studies have demonstrated the efficacy of ledprona through laboratory, greenhouse, and field studies. Colorado potato beetles from geographically distinct populations are known to vary in their response to insecticides, including experimental compounds based on RNAi. We tested the mortality and foliage consumption of beetles from different areas in the US treated with ledprona and found significant variation in both parameters. The beetles originating from New York were significantly less susceptible to ledprona in leaf disc assays compared to other populations. However, currently there is no evidence of reduced performance of ledprona against that population under field conditions, possibly because intoxicated beetles cannot withstand multiple stressors present in the field. The results of this study confirmed that the ledprona efficacy differs among geographically distinct populations, which may have implications for managing Colorado potato beetles.

Caterpillar Responses to Gustatory Stimuli in Potato Tuber Moths: Electrophysiological and Behavioral Insights.

This research investigates how fourth-instar larvae of the potato tuber moth, Phthorimaea operculella, respond to plant secondary metabolites (sucrose, glucose, nicotine, and tannic acid) both in terms of gustatory electrophysiology and feeding behavior. The objective is to establish a theoretical foundation for employing plant-derived compounds in potato tuber moth control. We employed single-sensillum recording techniques and dual-choice leaf disk assays to assess the gustatory electrophysiological responses and feeding preferences of these larvae towards the mentioned compounds. Sensory neurons responsive to sucrose, glucose, nicotine, and tannic acid were identified in the larvae's medial and lateral sensilla styloconica. Neuronal activity was influenced by stimulus type and concentration. Notably, the two types of sensilla styloconica displayed distinct response patterns for sucrose and glucose while they had similar firing patterns towards nicotine and tannic acid. Sucrose and glucose significantly promoted larval feeding, while nicotine and tannic acid had significant inhibitory effects. These findings demonstrate that the medial and lateral sensilla styloconica house sensory neurons sensitive to both feeding stimulants and inhibitors, albeit with differing response profiles and sensitivities. This study suggests that sucrose and glucose are promising candidates for feeding stimulants, while nicotine and tannic acid show potential as effective feeding inhibitors of P. operculella larvae.

Formanilides from the culture broth of Perenniporia fraxinea.

A new formanilide dimer, fraxinin (1), and three known formanilides (2‒4) were isolated from the culture broth of Perenniporia fraxinea using silica gel and Sephadex LH-20 column chromatographies, medium-pressure liquid chromatography (MPLC), and preparative HPLC. The structures of these compounds were determined by spectroscopic methods, such as NMR and mass analysis, and by comparison of the spectra with previously reported data. The free radical scavenging activities of the isolated compounds were assessed using 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals. Compounds 1‒3 exhibited ABTS radical scavenging activity with IC50 values in the range of 57.2-250.2 μM. Compounds 2 and 4 marginally reduced disease incidence of powdery mildew with a control value of 42% at 1.0 mg ml-1 in cucumber leaf disk assay.

Evaluation of morpho-physiological and biochemical indicators of salinity stress in Argemone mexicana L., medicinal plant from Papaveraceae

Abstract Plant growth is significantly impacted by abiotic stress factors including soil salinity and plants do have different mechanism to nullify the effect of these stresses. The current study is to understand the various physio-biochemical characteristics of Mexican prickly poppy (Argemone mexicana L.), a medicinally valued species belongs to poppy family under different saline conditions (0 mM, 100 mM, 200 mM and 300 mM) to investigate the mechanisms underlying salinity tolerance. The results showed that salinity caused a negative impact on number of leaves, stomatal index and chlorophyll content in plants. Decreased chlorophyll content suggests a decline in photosynthetic rate. The concentrations of stress specific amino acid proline, soluble sugar sucrose, and secondary metabolite phenol were increased in plants treated with increasing concentrations of NaCl solution. Sucrose helps to protect the isolated chloroplast from damage caused by salinity stress. The degree of bleaching seen in the leaf disc assay was an indication of the damage caused by stress. These findings indicated that higher proline, sugar, and phenol content could be the key factors, which offer advantage to Argemone under saline conditions.

Peptide analogs of a Trichoderma peptaibol effectively control downy mildew in the vineyard.

Plasmopara viticola, the agent of grapevine downy mildew, causes enormous economic damage and its control is primarily based on synthetic fungicides. The European Union (EU) policies promote reducing reliance on synthetic plant protection products. Biocontrol agents (BCA) such as Trichoderma constitute a resource for the development of biopesticides. Trichoderma species produce secondary metabolites such as peptaibols, whose poor water solubility hampers their practical use as agrochemicals. To identify new bio-inspired molecules effective against P. viticola, some water-soluble peptide analogs of the peptaibol trichogin were synthesized. In grapevine leaf disk assays, various peptides at 50 µM completely prevented P. viticola infection after zoosporangia inoculation. Microscopic observations carried out with one of the most effective peptides showed that it causes membrane lysis and cytoplasm granulation of both zoosporangia and zoospores. Among the effective peptides, 4r was selected for a two-year field trial experiment. In the vineyard, the peptide administered at 100 µM (equivalent to 129.3 g/ha) overall reduced significantly disease incidence and severity on both leaves and bunches, allowing protection levels similar to those obtained with a cupric fungicide. In the second-year trial, reduced dosages were also tested, and results indicated that even by reducing the peptide concentration by 50 or 75%, a significant decrease in the disease level was obtained at the end of the trial. The peptide did not show any phytotoxic effect. Previously, peptide 4r had been demonstrated to be active against other fungal pathogens, including the grapevine fungus Botrytis cinerea. Thus, this peptide may be a candidate for broad-spectrum fungicide whose biological properties deserve further investigation.

An image-analysis based leaf disc assay for the rapid evaluation of genetic resistance to fire blight in apples

An image-analysis based leaf disc assay for the rapid evaluation of genetic resistance to fire blight in apples

Phenolic secondary metabolites from Acorus calamus (Acorales: Acoraceae) rhizomes: the feeding deterrents for Spodoptera litura (Lepidoptera: Noctuidae).

Spodoptera litura Fabricius (Lepidoptera: Noctuidae) is one of the most destructive pests of various crops cultivated in Thailand. Spodoptera litura larvae, at early stages, attack the leaves and feed on every part of infested crops in later stages. Acorus calamus essential oil contains toxic asarones, which are generalistic cytotoxic compounds. However, the present study is the first attempt to look at safer metabolites from the rhizomes that could deter insect feeding. The objective was to use such compounds as safer residues on crops that would prevent the feeding of herbivorous lepidopterans. Accordingly, phenolic metabolites were isolated and evaluated to establish the feeding deterrence against polyphagous S. litura larvae. Methanol extract of A. calamus, chrysin, and 4-hydroxy acetophenone compounds were the most effective feeding deterrents with FD50 of 87.18, 10.33, and 70.77 µg/cm2, respectively, after 4 h of feeding on treated kale leaves in a no-choice leaf disc assay. Chrysin also reduced carboxylesterase activities (1.37-fold), whereas A. calamus methanol extract reduced glutathione-S-transferase activities (1.44-fold). Some larvae were also seen dead if they consumed the treated kale leaves. Feeding deterrent activity in the methanol extract of A. calamus was due to chrysin and 4-hydroxy acetophenone. The large-scale utilization of such compounds could help develop feeding deterrent strategies in the integrated pest management of lepidopterans.

Failure of methanol detoxification in pests confers broad spectrum insect resistance in PME overexpressing transgenic cotton

Methanol is noxious to insect pests, but most plants do not make enough of it to shield themselves from encroaching insects. Methanol emission is known to increase in the instance of herbivory. In the current study, we showed that Aspergillus niger pectin methylesterase over-expression increases methanol emission and confers resistance to polyphagous insect pests on transgenic cotton plants by impeding the possible methanol detoxification pathways. Transgenic plants emitted ∼11 fold higher methanol displaying insect mortality of 96% and 93% in Helicoverpa armigera and Spodoptera litura, respectively. The larvae were unable to survive and finish their life cycle and the surviving larvae exhibited severe growth retardation. Insects try to detoxify methanol via catalase, carboxylesterase and cytochrome P450 monooxygenase enzymes, amongst which cytochrome P450 plays a major role in oxidizing methanol to formaldehyde and formaldehyde to formic acid, which is broken down into carbon dioxide and water. In our study, catalase and esterase enzymes were found to be upregulated, but cytochrome P450 monooxygenase levels were not much affected. Leaf disc assays and In-planta bioassays also showed 50–60% population reduction in the sap sucking pests, such as Bemisia tabaci and Phenacoccus solenopsis. These findings imply that elevated methanol emissions confer resistance in plants against chewing and sap-sucking pests by tampering the methanol detoxification pathways. Such mechanism will be useful in imparting expansive resistance against pests in plants.

CRISPR/Cas9-Mediated Mutation in XSP10 and SlSAMT Genes Impart Genetic Tolerance to Fusarium Wilt Disease of Tomato (Solanum lycopersicum L.)

Fusarium wilt is a major devastating fungal disease of tomato (Solanum lycopersicum L.) caused by Fusarium oxysporum f. sp. lycopersici (Fol) which reduces the yield and production. Xylem sap protein 10 (XSP10) and Salicylic acid methyl transferase (SlSAMT) are two putative negative regulatory genes associated with Fusarium wilt of tomato. Fusarium wilt tolerance in tomato can be developed by targeting these susceptible (S) genes. Due to its efficiency, high target specificity, and versatility, CRISPR/Cas9 has emerged as one of the most promising techniques for knocking out disease susceptibility genes in a variety of model and agricultural plants to increase tolerance/resistance to various plant diseases in recent years. Though alternative methods, like RNAi, have been attempted to knock down these two S genes in order to confer resistance in tomato against Fusarium wilt, there has been no report of employing the CRISPR/Cas9 system for this specific intent. In this study, we provide a comprehensive downstream analysis of the two S genes via CRISPR/Cas9-mediated editing of single (XSP10 and SlSAMT individually) and dual-gene (XSP10 and SlSAMT simultaneously). Prior to directly advancing on to the generation of stable lines, the editing efficacy of the sgRNA-Cas9 complex was first validated using single cell (protoplast) transformation. In the transient leaf disc assay, the dual-gene editing showed strong phenotypic tolerance to Fusarium wilt disease with INDEL mutations than single-gene editing. In stable genetic transformation of tomato at the GE1 generation, dual-gene CRISPR transformants of XSP10 and SlSAMT primarily exhibited INDEL mutations than single-gene-edited lines. The dual-gene CRISPR-edited lines (CRELs) of XSP10 and SlSAMT at GE1 generation conferred a strong phenotypic tolerance to Fusarium wilt disease compared to single-gene-edited lines. Taken together, the reverse genetic studies in transient and stable lines of tomato revealed that, XSP10 and SlSAMT function together as negative regulators in conferring genetic tolerance to Fusarium wilt disease.

Characterization of a Disease-Suppressive Isolate of Lysobacter enzymogenes with Broad Antagonistic Activity against Bacterial, Oomycetal and Fungal Pathogens in Different Crops.

Although synthetic pesticides play a major role in plant protection, their application needs to be reduced because of their negative impact on the environment. This applies also to copper preparations, which are used in organic farming. For this reason, alternatives with less impact on the environment are urgently needed. In this context, we evaluated eight isolates of the genus Lysobacter (mainly Lysobacter enzymogenes) for their activity against plant pathogens. In vitro, the investigated Lysobacter isolates showed broad antagonistic activity against several phytopathogenic fungi, oomycetes and bacteria. Enzyme assays revealed diverse activities for the tested isolates. The most promising L. enzymogenes isolate (LEC) was used for further detailed analyses of its efficacy and effective working concentrations. The experiments included in vitro spore and sporangia germination tests and leaf disc assays as well as ad planta growth chamber trials against Alternaria solani and Phytophthora infestans on tomato plants, Pseudoperonospora cubensis on cucumbers and Venturia inaequalis on young potted apple trees. When applied on leaves, dilutions of a culture suspension of LEC had a concentration-dependent, protective effect against the tested pathogens. In all pathosystems tested, the effective concentrations were in the range of 2.5-5% and similarly efficacious to common plant protection agents containing copper hydroxide, wettable sulphur or fenhexamid. Thus, the isolate of L. enzymogenes identified in this study exhibits a broad activity against common plant pathogens and is therefore a promising candidate for the development of a microbial biocontrol agent.

Evaluation of the Efficacy of Flupyradifurone against Bemisia tabaci on Cassava in Tanzania.

Simple SummaryCassava-colonizing Bemisia tabaci whitefly negatively impact cassava production in sub-Saharan Africa through transmission of viruses that cause cassava mosaic disease (CMD) and cassava brown streak disease (CBSD). This study evaluated the efficacy of a novel insecticide flupyradifurone against this whitefly pest through standard spraying and cutting dip methods. The results from the laboratory, screenhouse and field experiments consistently showed flupyradifurone is effective at reducing whiteflies compared to control treatments. A single cutting dip using this insecticide was highly effective at reducing whiteflies under field conditions and, therefore, should be considered as an alternative to insecticide spraying in cassava systems. Additionally, whiteflies were least abundant during the long rainy season (Masika) and on cassava variety Mkuranga1.A novel butenolide insecticide—flupyradifurone (Sivanto SL 200)—was evaluated for efficacy against cassava-colonizing Bemisia tabaci whitefly under laboratory, screenhouse and field conditions. LC50 values from leaf disc spray assays were comparable for both flupyradifurone (12.7 g a.i/100 L) and imidacloprid (12.6 g a.i/100 L). Both insecticides caused high levels of adult whitefly mortality in leaf disc and leaf dip assays when compared to untreated controls. In screenhouse-based trials, longer soaking (60 min) with flupyradifurone or imidacloprid was more effective than shorter soaking durations (15 or 30 min). In field spraying experiments, flupyradifurone significantly reduced whiteflies, and both insecticides demonstrated powerful knockdown effects on whitefly adult abundances over a period up to 24 h. Single cutting dip application of flupyradifurone reduced whitefly adult abundance by 2 to 6 times, and nymphs by 2 to 13 times. Lower whitefly abundances resulting from insecticide application reduced the incidence of CMD or CBSD. In addition, in field experiments, whiteflies were fewer during the long rainy season (Masika) and on cassava variety Mkuranga1. The findings from this study demonstrate that cutting dips with flupyradifurone could be incorporated as a management tactic against cassava whiteflies. This would ideally be combined in an IPM strategy with other cassava virus and virus vector management tactics including host-plant resistance, phytosanitation and the use of clean seed.