Regulation of virulence mechanisms in plant-pathogenic Streptomyces.

Streptomyces have a uniquely complex developmental life cycle that involves the coordination of morphological differentiation with the production of numerous bioactive specialized metabolites. The majority of Streptomyces spp. are soil-dwelling saprophytes, while plant pathogenicity is a rare attribute among members of this genus. Phytopathogenic Streptomyces are responsible for economically important diseases such as common scab, which affects potato and other root crops. Following the acquisition of genes encoding virulence factors, Streptomyces pathogens are expected to have specifically adapted their regulatory pathways to enable transition from a primarily saprophytic to a pathogenic lifestyle. Investigations of the regulation of pathogenesis have primarily focused on Streptomyces scabiei and the principal pathogenicity determinant thaxtomin A. The coordination of growth and thaxtomin A production in this species is controlled in a hierarchical manner by cluster-situated regulators, pleiotropic regulators, signalling and plant-derived molecules, and nutrients. Although the majority of phytopathogenic Streptomyces produce thaxtomins, many also produce additional virulence factors, and there are scab-causing pathogens that do not produce thaxtomins. The development of effective control strategies for common scab and other Streptomyces plant diseases requires a more in-depth understanding of the genetic and environmental factors that modulate the plant pathogenic lifestyle of these organisms.

The barrier to radial oxygen loss protects roots against hydrogen sulphide intrusion and its toxic effect.

The root barrier to radial O2 loss (ROL) is a key root trait preventing O2 loss from roots to anoxic soils, thereby enabling root growth into anoxic, flooded soils. We hypothesized that the ROL barrier can also prevent intrusion of hydrogen sulphide (H2 S), a potent phytotoxin in flooded soils. Using H2 S- and O2 -sensitive microsensors, we measured the apparent permeance to H2 S of rice roots, tested whether restricted H2 S intrusion reduced its adverse effects on root respiration, and whether H2 S could induce the formation of a ROL barrier. The ROL barrier reduced apparent permeance to H2 S by almost 99%, greatly restricting H2 S intrusion. The ROL barrier acted as a shield towards H2 S; O2 consumption in roots with a ROL barrier remained unaffected at high H2 S concentration (500 µM), compared to a 67% decline in roots without a barrier. Importantly, low H2 S concentrations induced the formation of a ROL barrier. In conclusion, the ROL barrier plays a key role in protecting against H2 S intrusion, and H2 S can act as an environmental signalling molecule for the induction of the barrier. This study demonstrates the multiple functions of the suberized/lignified outer part of the rice root beyond that of restricting ROL.

Status of Phytotoxins Isolated from Necrotrophic Fungi Causing Diseases on Grain Legumes.

Fungal phytotoxins can be defined as secondary metabolites toxic to host plants and are believed to be involved in the symptoms developed of a number of plant diseases by targeting host cellular machineries or interfering with host immune responses. As any crop, legumes can be affected by a number of fungal diseases, causing severe yield losses worldwide. In this review, we report and discuss the isolation, chemical, and biological characterization of fungal phytotoxins produced by the most important necrotrophic fungi involved in legume diseases. Their possible role in plant-pathogen interaction and structure-toxicity relationship studies have also been reported and discussed. Moreover, multidisciplinary studies on other prominent biological activity conducted on reviewed phytotoxins are described. Finally, we explore the challenges in the identification of new fungal metabolites and their possible applications in future experiments.

(±)-3-Deoxyradicinin Induces Stomata Opening and Chloroplast Oxidative Stress in Tomato (Solanum lycopersicum L.).

Radicinin is a phytotoxic dihydropyranopyran-4,5-dione isolated from the culture filtrates of Cochliobolus australiensis, a phytopathogenic fungus of the invasive weed buffelgrass (Cenchrus ciliaris). Radicinin proved to have interesting potential as a natural herbicide. Being interested in elucidating the mechanism of action and considering radicinin is produced in small quantities by C. australiensis, we opted to use (±)-3-deoxyradicinin, a synthetic analogue of radicinin that is available in larger quantities and shows radicinin-like phytotoxic activities. To obtain information about subcellular targets and mechanism(s) of action of the toxin, the study was carried out by using tomato (Solanum lycopersicum L.), which, apart from its economic relevance, has become a model plant species for physiological and molecular studies. Results of biochemical assays showed that (±)-3-deoxyradicinin administration to leaves induced chlorosis, ion leakage, hydrogen peroxide production, and membrane lipid peroxidation. Remarkably, the compound determined the uncontrolled opening of stomata, which, in turn, resulted in plant wilting. Confocal microscopy analysis of protoplasts treated with (±)-3-deoxyradicinin ascertained that the toxin targeted chloroplasts, eliciting an overproduction of reactive singlet oxygen species. This oxidative stress status was related by qRT-PCR experiments to the activation of transcription of genes of a chloroplast-specific pathway of programmed cell death.

Value of dehydrated food waste fertiliser products in increasing soil health and crop productivity.

Dehydration of food waste is a technique in which food waste is dewatered to form a low moisture product. This research characterised the physicochemical properties of different dehydrated food waste products and examined their value in improving physical, biological, and chemical properties of soils. Dehydrated food waste products were slightly acidic (4.7-5.1) with high levels of electrical conductivity (EC) (4.83-7.64 mS cm-1). The products were composed of complex carbohydrates, polysaccharides, alcohols, phenols, carboxylic acid, lipids, and fats and contained high levels of total and available nutrients. Dehydrated food wastes slightly impacted the soil pH; however, they significantly increased soil EC, which may cause soil salinity when applied repeatedly. The food waste products also increased macro-nutrients (N, P, and K) for plants across different soil types. Carbon and nutrients in dehydrated food waste increased microbial activity, measured by basal respiration. Delayed germination and reduced plant growth of corn (Zea mays) and wheat (Triticum aestivum) plants were observed at high application rates of dehydrated food waste. This may have resulted from a combination of phytotoxins, anoxic conditions, salinity as well as the water-repellent nature of dehydrated food waste. However, release of nutrients increased nutrient uptake and plant biomass in corn and wheat plants at low levels of food waste application. The dehydrated food waste products may require composting prior to soil application or incorporation into soil for a long duration prior to planting. These processes will overcome the limitations of phytotoxins, anoxic conditions, salinity, and water repellence. Further work is required to optimise the levels of dehydrated food waste application to improve soil health and crop productivity.

Potentiators of Disease During Barley Infection by Pyrenophora teres f. teres in a Susceptible Interaction.

Pyrenophora teres f. teres is a necrotrophic fungal pathogen and causal agent of net form net blotch (NFNB), a significant disease of barley. RNA-seq data encompassing asymptomatic and subsequent necrotrophic phases of the pathogen was obtained for P. teres f. teres isolate W1-1 in NFNB-sensitive cultivar Baudin. Host genes notably regulated during infection included concerted induction of over half the repertoire of disease resistance genes, together with genes involved in oxidation-reduction processes, characteristic of a hypersensitive response. Several systemic acquired resistance response genes were suppressed and there was a complete absence of defense-related thionin gene expression. In P. teres f. teres, genes involved in hydrolase activities and cell-wall catabolic processes were induced during infection, while nitrate assimilation and response to oxidative stress processes were suppressed. Timecourse data allowed a number of predicted P. teres f. teres effector genes with differing expression profiles to be identified that may underlie barley sensitivity to NFNB. Candidate genes involved in the host-pathogen interaction provide a basis for functional characterization and control strategies based on fungicide or mutation targets, which will facilitate further research aimed at controlling NFNB disease.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Disentangling Mechanisms in Natural Toxin Sorption to Soil Organic Carbon.

Natural toxins are multifunctional, often ionizable organic compounds increasingly detected in the environment. Surprisingly little is known about their interactions with soil organic carbon, although sorption largely controls transport, bioavailability, and dissipation. For a set of 117 natural toxins from 36 compound classes the pH-dependent organic carbon-water distribution coefficient (Doc) was quantified using a soil column chromatography approach under changing conditions with regards to pH, ionic strength, and the major inorganic cation in solution. Natural toxins could be assigned to groups with either hydrophobic partitioning or specific interactions (complexation reactions, cation exchange) as dominating sorption mechanisms. The complex interplay of interactions in the sorption of natural toxins was equally influenced by sorbate, sorbent, and solution specific characteristics. High variability in sorption was particularly observed in the presence of Ca2+ resulting in Doc being enhanced by a factor of 10 when the pH was increased from 4.5 to 6. Sorbates following this trend contain either functional groups able to form ternary complexes via Ca2+ or aromatic moieties adjacent to protonated N presumably stabilizing cation exchange reactions. Although sorption was often stronger than predicted, investigated natural toxins were highly mobile under all considered conditions.

Exposure and Transport of Alkaloids and Phytoestrogens from Soybeans to Agricultural Soils and Streams in the Midwestern United States.

Phytotoxins are naturally produced toxins with potencies similar/higher than many anthropogenic micropollutants. Nevertheless, little is known regarding their environmental fate and off-field transport to streams. To fill this research gap, a network of six basins in the Midwestern United States with substantial soybean production was selected for the study. Stream water (n = 110), soybean plant tissues (n = 8), and soil samples (n = 16) were analyzed for 12 phytotoxins (5 alkaloids and 7 phytoestrogens) and 2 widely used herbicides (atrazine and metolachlor). Overall, at least 1 phytotoxin was detected in 82% of the samples, with as many as 11 phytotoxins detected in a single sample (median = 5), with a concentration range from below detection to 37 and 68 ng/L for alkaloids and phytoestrogens, respectively. In contrast, the herbicides were ubiquitously detected at substantially higher concentrations (atrazine: 99% and metolachlor: 83%; the concentrations range from below detection to 150 and 410 ng/L, respectively). There was an apparent seasonal pattern for phytotoxins, where occurrence prior to and during harvest season (September to November) and during the snow melt season (March) was higher than that in December-January. Runoff events increased phytotoxin and herbicide concentrations compared to those in base-flow conditions. Phytotoxin plant concentrations were orders of magnitude higher compared to those measured in soil and streams. These results demonstrate the potential exposure of aquatic and terrestrial organisms to soybean-derived phytotoxins.

Phytotoxin of rice aggregate sheath spot pathogen Rhizoctonia oryzae-sativae and its biological activities.

Rice aggregate sheath spot disease occurs in many countries and causes serious yield losses. In China, the disease-causing fungus Rhizoctonia oryzae-sativae was reported in 1985, and since then, it has rarely been reported in major rice-growing areas after almost 30 years. Compared with Rhizoctonia solani, R. oryzae-sativae has a significantly different physiological morphology and growth status, although both fungi affect rice leaves in very similar ways. The optimum temperature for the suitable growth of R. oryzae-sativae is 31 °C, which is consistent with previous reports. We extracted phytotoxins from R. oryzae-sativae and analyzed its biological activity via the detached leaf and radicle inhibition methods. Rhizoctonia solani and R. oryzae-sativae exhibit differences in terms of pathogenicity and toxin activity, which indicates that these fungi may produce different toxin components. Based on gas chromatography-mass spectrometry data, esters, phenols, and other components were present in the crude toxin extract of R. oryzae-sativae. Our research provides a new method for studying the phytotoxins of R. oryzae-sativae. However, further studies are needed to elucidate the pathogenic mechanisms responsible for aggregate sheath spot disease in rice.

Identification and Quantification of a Phytotoxic Metabolite from Alternaria dauci.

Alternaria dauci is the causal agent of Alternaria leaf blight (ALB) in carrot (Daucus carota) crops around the world. However, to date, A. dauci has received limited attention in its production of phytotoxic metabolites. In this investigation, the bioassay-guided isolation of the extract from liquid cultures of A. dauci resulted in the isolation of two metabolites identified as α-acetylorcinol (1) and p-hydroxybenzoic acid (2), based on their spectroscopic data and results from chemical correlation reactions. Testing of both metabolites in different assays showed an important phytotoxic activity for p-hydroxybenzoic acid (2) when tested in the leaf-spot assay on parsley (Petroselinum crispum), in the leaf infiltration assay on tobacco (Nicotiana alata) and marigold (Tagetes erecta), and in the immersion assay on parsley and parsnip (Pastinaca sativa) leaves. Quantification of the two metabolites in the crude extract of A. dauci kept at different times showed that p-hydroxybenzoic acid (2) is one of the first metabolites to be synthesized by the pathogen, suggesting that this salicylic acid derivative could play an important role in the pathogenicity of the fungus.

Positive and Negative Regulation of the Virulence-Associated Coronafacoyl Phytotoxin in the Potato Common Scab Pathogen Streptomyces scabies.

The potato common scab pathogen Streptomyces scabies produces N-coronafacoyl-l-isoleucine (CFA-Ile), which is a member of the coronafacoyl family of phytotoxins that are synthesized by multiple plant pathogenic bacteria. The CFA-Ile biosynthetic gene cluster contains a regulatory gene, cfaR, which directly controls the expression of the phytotoxin structural genes. In addition, a gene designated orf1 encodes a predicted ThiF family protein and is cotranscribed with cfaR, suggesting that it also plays a role in the regulation of CFA-Ile production. In this study, we demonstrated that CfaR is an essential activator of coronafacoyl phytotoxin production, while ORF1 is dispensable for phytotoxin production and may function as a helper protein for CfaR. We also showed that CFA-Ile inhibits the ability of CfaR to bind to the promoter region driving expression of the phytotoxin biosynthetic genes and that elevated CFA-Ile production by overexpression of both cfaR and orf1 in S. scabies increases the severity of disease symptoms induced by the pathogen during colonization of potato tuber tissue. Overall, our study reveals novel insights into the regulatory mechanisms controlling CFA-Ile production in S. scabies and it provides further evidence that CFA-Ile is an important virulence factor for this organism.

Comparative genomics and pathogenicity potential of members of the Pseudomonas syringae species complex on Prunus spp.

BACKGROUND: Diseases on Prunus spp. have been associated with a large number of phylogenetically different pathovars and species within the P. syringae species complex. Despite their economic significance, there is a severe lack of genomic information of these pathogens. The high phylogenetic diversity observed within strains causing disease on Prunus spp. in nature, raised the question whether other strains or species within the P. syringae species complex were potentially pathogenic on Prunus spp. RESULTS: To gain insight into the genomic potential of adaptation and virulence in Prunus spp., a total of twelve de novo whole genome sequences of P. syringae pathovars and species found in association with diseases on cherry (sweet, sour and ornamental-cherry) and peach were sequenced. Strains sequenced in this study covered three phylogroups and four clades. These strains were screened in vitro for pathogenicity on Prunus spp. together with additional genome sequenced strains thus covering nine out of thirteen of the currently defined P. syringae phylogroups. Pathogenicity tests revealed that most of the strains caused symptoms in vitro and no obvious link was found between presence of known virulence factors and the observed pathogenicity pattern based on comparative genomics. Non-pathogenic strains were displaying a two to three times higher generation time when grown in rich medium. CONCLUSION: In this study, the first set of complete genomes of cherry associated P. syringae strains as well as the draft genome of the quarantine peach pathogen P. syringae pv. persicae were generated. The obtained genomic data were matched with phenotypic data in order to determine factors related to pathogenicity to Prunus spp. Results of this study suggest that the inability to cause disease on Prunus spp. in vitro is not the result of host specialization but rather linked to metabolic impairments of individual strains.

Virulence mechanisms of plant-pathogenic Streptomyces species: an updated review.

Gram-positive Actinobacteria from the genus Streptomyces are best known for their morphological complexity and for their ability to produce numerous bioactive specialized metabolites with useful applications in human and veterinary medicine and in agriculture. In contrast, the ability to infect living plant tissues and to cause diseases of root and tuber crops such as potato common scab (CS) is a rare attribute among members of this genus. Research on the virulence mechanisms of plant-pathogenic Streptomyces spp. has revealed the importance of the thaxtomin phytotoxins as key pathogenicity determinants produced by several species. In addition, other phytotoxic specialized metabolites may contribute to the development or severity of disease caused by Streptomyces spp., along with the production of phytohormones and secreted proteins. A thorough understanding of the molecular mechanisms of plant pathogenicity will enable the development of better management procedures for controlling CS and other plant diseases caused by the Streptomyces.

Rice acclimation to soil flooding: Low concentrations of organic acids can trigger a barrier to radial oxygen loss in roots.

Waterlogged soils contain monocarboxylic acids produced by anaerobic microorganisms. These "organic acids" can accumulate to phytotoxic levels and promote development of a barrier to radial O2 loss (ROL) in roots of some wetland species. Environmental cues triggering root ROL barrier induction, a feature that together with tissue gas-filled porosity facilitates internal aeration, are important to elucidate for knowledge of plant stress physiology. We tested the hypothesis that comparatively low, non-toxic, concentrations of acetic, propionic, butyric, and/or hexanoic acids might induce root ROL barrier formation in rice. Each organic acid, individually, triggered the ROL barrier in roots but with no effect (acetic or butyric acids) or with only slight effects (propionic or hexanoic acids) on root extension. Transcripts of four genes related to suberin biosynthesis were increased by some of the organic acid treatments. Respiration in root tissues was not, or moderately, inhibited. Beyond a narrow concentration range, however, respiration declined exponentially and the order (least to greatest) for EC50 (effective concentration for 50% inhibition) was butyric, propionic, acetic, then hexanoic acid. An understanding of the environmental cue for root ROL barrier induction should enhance future work to elucidate the molecular regulation of this root trait contributing to plant flooding tolerance.

NMR-based metabolomics and bioassays to study phytotoxic extracts and putative phytotoxins from Mediterranean plant species.

INTRODUCTION: Mediterranean plants are characterised by a high content of bioactive secondary metabolites that play important roles in plant-plant interactions as plant growth regulators and could be useful for the development of new eco-friendly herbicides. OBJECTIVE: An NMR-based metabolomics approach was reported to seek selective phytotoxic plant extracts and putative plant-derived active molecules. METHODS: Plant extracts derived from five Mediterranean donor species (Pistacia lentiscus, Bellis sylvestris, Phleum subulatum, Petrohrhagia saxifraga and Melilotus neapolitana) were used to treat the hydroponic cultures of three receiving plants (Triticum durum, Triticum ovatum and Avena fatua). Morphological analyses of the treated receiving plants were carried out. NMR-based metabolomics was applied both to characterise the donor plant extracts and to study the effects of the treatments on the receiving plants. RESULTS: This study allowed the identification of Melilotus neapolitana and Bellis sylvestris as phytotoxic plant and good candidates for further studies. Specifically, the NMR-based metabolomics investigation showed that these species affect a specific set of metabolites (such as sugars, amino and organic acids) and therefore metabolic pathways [i.e. tricarboxylic acid (TCA) cycle, amino acid metabolism, etc.] that are crucial for the plant growth and development. Moreover, it was possible to identify the metabolite(s) probably responsible for the phytotoxicity of the active extracts. CONCLUSION: The NMR-based metabolomics approach employed in this study led to the identification of two phytotoxic plant extracts and their putative active principles. These new insights will be of paramount importance in the future to find plant derived molecules endowed with phytotoxic activities.

Synthesis and Herbicidal Activity Against Buffelgrass (Cenchrus ciliaris) of (±)-3-deoxyradicinin.

A novel synthetic strategy for obtainment of (±)-3-deoxyradicinin (2) is reported. This synthetic methodology is more efficient than those previously reported in the literature and also shows higher versatility towards the introduction of different side-chains at both C-7 and C-2. The obtained compound (±)-2 shows phytotoxicity against the grass-weed buffelgrass comparable to that of the natural phytotoxin radicinin (1). Therefore, (±)-2 can constitute a more practical synthetic alternative to 1 as bioherbicide for buffelgrass control.

Discovery of Three New Phytotoxins from the Fungus Aspergillus nidulans by Pathway Inactivation.

Fungi are a source of novel phytotoxic compounds to be explored in the search for effective and environmentally safe herbicides. The genetic inactivation of the biosynthetic pathway of the new phytotoxin cichorine has led to the isolation of three novel phytotoxins from the fungus Aspergillus nidulans: 8-methoxycichorine (4), 8-epi-methoxycichorine (5), and N-(4'-carboxybutyl) cichorine (6). The structure of the new compounds was clearly determined by a combination of nuclear magnetic resonance (NMR) analysis and high-resolution electrospray ionization (HRESIMS). The phytotoxic bioassay was studied on leaves from Zea mays and Medicago polymorpha L. at the concentration of 5 × 10-3 M by using a moist chamber technique. Novel phytotoxins 8-methoxycichorine (4), 8-epi-methoxycichorine (5), and N-(4'-carboxybutyl) cichorine (6) exhibited a better phytotoxic effect than cichorine.

Radicinin, a Fungal Phytotoxin as a Target-Specific Bioherbicide for Invasive Buffelgrass (Cenchrus ciliaris) Control.

The fungal pathogens Cochliobolus australiensis and Pyricularia grisea have recently been isolated from diseased leaves of buffelgrass (Cenchrus ciliaris) in its North American range, and their ability to produce phytotoxic metabolites that could potentially be used as natural herbicides against this invasive weed was investigated. Fourteen secondary metabolites obtained from in vitro cultures of these two pathogens were tested by leaf puncture assay on the host plant at different concentrations. Radicinin and (10S, 11S)-epi-pyriculol proved to be the most promising compounds. Thus, their phytotoxic activity was also evaluated on non-host indigenous plants. Radicinin demonstrated high target-specific toxicity on buffelgrass, low toxicity to native plants, and no teratogenic, sub-lethal, or lethal effects on zebrafish (Brachydanio rerio) embryos. It is now under consideration for the development of a target-specific bioherbicide to be used against buffelgrass in natural systems where synthetic herbicides cause excessive damage to native plants.

Assignment Through Chiroptical Methods of The Absolute Configuration of Fungal Dihydropyranpyran-4-5-Diones Phytotoxins, Potential Herbicides for Buffelgrass (Cenchrus ciliaris) Biocontrol.

Radicinin and cochliotoxin (1 and 2) two phytotoxic pyranpyran-4,5-diones were isolated together with their close metabolites 3-epi-radicinin, radicinol, and its 3-epimer (3-5), from the culture filtrates of Cochliobolus australiensis, a fungus proposed as mycoherbcide for biocontrol of buffelgrass, a very noxious and dangerous weed. The absolute configuration of cochliotoxin was determined by chiroptical Optical Rotatory Dispersion (ORD), Electronic Circular Dichroism (ECD), and Vibrational Circular Dichroism (VCD)) and computational methods. The same methods were used to confirm that of radicinin, radicinol and their 3-epimers, previously determined with chemical, spectroscopic and ECD methods.

Niche partitioning of intertidal seagrasses: evidence of the influence of substrate temperature.

The influence of soil temperature on rhizome depths of four intertidal seagrass species was investigated in central Queensland, Australia. We postulated that certain intertidal seagrass species are soil temperature-sensitive and vertically stratify rhizome depths. Below-ground vertical stratification of intertidal seagrass rhizome depths was analysed based upon microclimate (soil temperature) and microhabitat (soil type). Soil temperature profiles exhibited heat transfer from surface layers to depth that varied by microhabitat, with vertical stratification of rhizome depths between species. Halodule uninervis rhizomes maintain a narrow median soil temperature envelope; compensating for high surface temperatures by occupying deeper, cooler soil substrates. Halophila decipiens, Halophila ovalis and Zostera muelleri rhizomes are shallow-rooted and exposed to fluctuating temperatures, with broader median temperature envelopes. Halodule uninervis appears to be a niche specialist, with the two Halophila species considered as generalist niche usage species. The implications of niche use based upon soil temperature profiles and rhizome rooting depths are discussed in the context of species' thermal tolerances and below-ground biomass O2 demand associated with respiration and maintenance of oxic microshields. This preliminary evidence suggests that soil temperature interaction with rhizome rooting depths may be a factor that influences the distribution of intertidal seagrasses.