Engineered coumarin accumulation reduces mycotoxin-induced oxidative stress and disease susceptibility.

Coumarins can fight pathogens and are thus promising for crop protection. Their biosynthesis, however, has not yet been engineered in crops. We tailored the constitutive accumulation of coumarins in transgenic Nicotiana benthamiana, Glycine max and Arabidopsis thaliana plants, as well as in Nicotiana tabacum BY-2 suspension cells. We did so by overexpressing A. thaliana feruloyl-CoA 6-hydroxylase 1 (AtF6'H1), encoding the key enzyme of scopoletin biosynthesis. Besides scopoletin and its glucoside scopolin, esculin at low level was the only other coumarin detected in transgenic cells. Mechanical damage of scopolin-accumulating tissue led to a swift release of scopoletin, presumably from the scopolin pool. High scopolin levels in A. thaliana roots coincided with reduced susceptibility to the root-parasitic nematode Heterodera schachtii. In addition, transgenic soybean plants were more tolerant to the soil-borne pathogenic fungus Fusarium virguliforme. Because mycotoxin-induced accumulation of reactive oxygen species and cell death were reduced in the AtF6'H1-overexpressors, the weaker sensitivity to F. virguliforme may be caused by attenuated oxidative damage of coumarin-hyperaccumulating cells. Together, engineered coumarin accumulation is promising for enhanced disease resilience of crops.

Bacillus firmus I-1582 promotes plant growth and impairs infection and development of the cyst nematode Heterodera schachtii over two generations.

Plant-parasitic nematodes wreak havoc on crops by root parasitism worldwide. An approach to combat nematode root parasitism is the application of antagonistic microbes like the rhizobacterium Bacillus firmus I-1582 which is promoted as biological control agent. Although B. firmus is a known nematode antagonist in general, the underlying mechanisms about its interaction with nematodes and plants have not yet been elucidated. Therefore, we explored the influence of B. firmus I-1582 as well as its extracellular and secreted molecules on plant-nematode interaction utilizing the plant-pathogen system Arabidopsis thaliana-Heterodera schachtii. We demonstrated that B. firmus I-1582 is attracted by A. thaliana root exudates, particularly by those of young plants. The bacterium colonized the root and showed a strictly pH-dependent development and plant growth promotion effect. Our results revealed that root colonization by B. firmus I-1582 significantly protected A. thaliana from infestation by the beet cyst nematode whereas dead bacterial cells or the culture supernatant were not effective. The bacterium also negatively affected nematode reproduction as well as pathogenicity and development of next generation nematodes. The obtained results highlight B. firmus I-1582 as a promising biocontrol agent that is well suited as an element of integrated control management strategies in sustainable agriculture.

Sublethal fluazaindolizine doses inhibit development of the cyst nematode Heterodera schachtii during sedentary parasitism.

BACKGROUND: Fluazaindolizine is a new compound for the control of plant-parasitic nematodes (PPNs) with an unknown and novel mode-of-action. This compound is very effective against important PPNs. However, investigations elucidating the impact of sublethal fluazaindolizine doses on early nematode virulence and plant-nematode interaction parameters are lacking. RESULTS: The effect of direct exposure of Heterodera schachtii juveniles to 50 ppm fluazaindolizine was negligible. Infection assays revealed a 57% reduction in adult females at 1.25 ppm and a 46% reduction in offspring at 40 ppm when juveniles were soaked in the compound for 48 h and subsequently inoculated onto Arabidopsis thaliana. Pre-incubation of A. thaliana roots with fluazaindolizine was not effective against H. schachtii. Conversely, supplementing the plant growth medium with fluazaindolizine led to a significant reduction of adults (-35%), females (-75%) and female size at 1.25 ppm and nearly completely inhibited nematode parasitism at 5 ppm. The impact of fluazaindolizine on A. thaliana was dependent on plant age, compound concentration and duration of contact. Very low sublethal fluazaindolizine concentrations, 5 or 10 ppm, did not interfere with nematode mobility, host finding, penetration, and induction of the feeding site, but specifically inhibited sedentary nematode development inside the root in a concentration-dependent manner. CONCLUSION: Fluazaindolizine does not have direct toxicity against PPN infective juveniles, but has a clear effect on nematodes during sedentary development. The formation of females and the development of offspring are strongly reduced. It will be interesting to identify the underlying mechanism in the future. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Heterologous Production of ß-Caryophyllene and Evaluation of Its Activity against Plant Pathogenic Fungi.

Terpenoids constitute one of the largest and most diverse groups within the class of secondary metabolites, comprising over 80,000 compounds. They not only exhibit important functions in plant physiology but also have commercial potential in the biotechnological, pharmaceutical, and agricultural sectors due to their promising properties, including various bioactivities against pathogens, inflammations, and cancer. In this work, we therefore aimed to implement the plant sesquiterpenoid pathway leading to ß-caryophyllene in the heterologous host Rhodobacter capsulatus and achieved a maximum production of 139 ± 31 mg L-1 culture. As this sesquiterpene offers various beneficial anti-phytopathogenic activities, we evaluated the bioactivity of ß-caryophyllene and its oxygenated derivative ß-caryophyllene oxide against different phytopathogenic fungi. Here, both compounds significantly inhibited the growth of Sclerotinia sclerotiorum and Fusarium oxysporum by up to 40%, while growth of Alternaria brassicicola was only slightly affected, and Phoma lingam and Rhizoctonia solani were unaffected. At the same time, the compounds showed a promising low inhibitory profile for a variety of plant growth-promoting bacteria at suitable compound concentrations. Our observations thus give a first indication that ß-caryophyllene and ß-caryophyllene oxide are promising natural agents, which might be applicable for the management of certain plant pathogenic fungi in agricultural crop production.

The Plant Sesquiterpene Nootkatone Efficiently Reduces Heterodera schachtii Parasitism by Activating Plant Defense.

Plant parasitic nematodes, including the beet cyst nematode Heterodera schachtii, constitute a devastating problem for crops worldwide. The limited availability of sustainable management options illustrates the need for new eco-friendly control means. Plant metabolites represent an invaluable source of active compounds for the discovery of such novel antagonistic agents. Here, we evaluated the impact of eight plant terpenoids on the H. schachtii parasitism of Arabidopsis thaliana. None of the metabolites affected the plant development (5 or 10 ppm). Nootkatone decreased the number of adult nematodes on A. thaliana to 50%, with the female nematodes being smaller compared to the control. In contrast, three other terpenoids increased the parasitism and/or female size. We discovered that nootkatone considerably decreased the number of nematodes that penetrated A. thaliana roots, but neither affected the nematode viability or attraction to plant roots, nor triggered the production of plant reactive oxygen species or changed the plant's sesquiterpene profile. However, we demonstrated that nootkatone led to a significant upregulation of defense-related genes involved in salicylic and jasmonic acid pathways. Our results indicate that nootkatone is a promising candidate to be developed into a novel plant protection agent acting as a stimulator of plant immunity against parasitic nematodes.

Insights into the Mechanism of Action of Highly Diluted Biologics.

The therapeutic use of Abs in cancer, autoimmunity, transplantation, and other fields is among the major biopharmaceutical advances of the 20th century. Broader use of Ab-based drugs is constrained because of their high production costs and frequent side effects. One promising approach to overcome these limitations is the use of highly diluted Abs, which are produced by gradual reduction of an Ab concentration to an extremely low level. This technology was used to create a group of drugs for the treatment of various diseases, depending on the specificity of the used Abs. Highly diluted Abs to IFN-γ (hd-anti-IFN-γ) have been demonstrated to be efficacious against influenza and other respiratory infections in a variety of preclinical and clinical studies. In the current study, we provide evidence for a possible mechanism of action of hd-anti-IFN-γ. Using high-resolution solution nuclear magnetic resonance spectroscopy, we show that the drug induced conformational changes in the IFN-γ molecule. Chemical shift changes occurred in the amino acids located primarily at the dimer interface and at the C-terminal region of IFN-γ. These molecular changes could be crucial for the function of the protein, as evidenced by an observed hd-anti-IFN-γ-induced increase in the specific binding of IFN-γ to its receptor in U937 cells, enhanced induced production of IFN-γ in human PBMC culture, and increased survival of influenza A-infected mice.