Water retting process with hemp pre-treatment: effect on the enzymatic activities and microbial populations dynamic.

Proper retting process of hemp stems, in which efficient separation of cellulose fiber from the rest of the stem is promoted by indigenous microorganisms able to degrade pectin, is essential for fiber production and quality. This research aimed to investigate the effect of a pre-treatment dew retting in field of hemp stalks on the pectinolytic enzymatic activity and microbiota dynamic during lab-scale water retting process. A strong increase in the pectinase activity as well as in the aerobic and anaerobic pectinolytic concentration was observed from 14 to 21 days, especially using hemp stalks that were not subjected to a pre-retting treatment on field (WRF0 0.690 ± 0.05 U/mL). Results revealed that the microbial diversity significantly varied over time during the water retting and the development of microbiota characterizing the water retting of hemp stalks of different biosystems used in this study was affected by pre-treatment conditions in the field and water retting process and by an interaction between the two methods. Although at the beginning of the experiment a high biodiversity was recorded in all biosystems, the water retting led to a selection of microbial populations in function of the time of pre-treatment in field, especially in bacterial populations. The use of hemp stems did not subject to a field pre-treatment seems to help the development of a homogeneous and specific pectinolytic microbiota with a higher enzymatic activity in respect to samples exposed to uncontrolled environmental conditions for 10, 20, or 30 days before the water retting process. KEY POINTS: • Microbial diversity significantly varied over time during water retting. • Water retting microbiota was affected by dew pre-treatment in the field. • Retting of no pretreated hemp allows the development of specific microbiota with high enzymatic activity.

Straw Incorporation with Exogenous Degrading Bacteria (ZJW-6): An Integrated Greener Approach to Enhance Straw Degradation and Improve Rice Growth.

Rice straw is an agricultural waste, the disposal of which through open burning is an emerging challenge for ecology. Green manufacturing using straw returning provides a more avant-garde technique that is not only an effective management measure to improve soil fertility in agricultural ecosystems but also nurtures environmental stewardship by reducing waste and the carbon footprint. However, fresh straw that is returned to the field cannot be quickly decomposed, and screening microorganisms with the capacity to degrade straw and understanding their mechanism of action is an efficient approach to solve such problems. This study aimed to reveal the potential mechanism of influence exerted by exogenous degradative bacteria (ZJW-6) on the degradation of straw, growth of plants, and soil bacterial community during the process of returning rice straw to the soil. The inoculation with ZJW-6 enhanced the driving force of cellulose degradation. The acceleration of the rate of decomposition of straw releases nutrients that are easily absorbed by rice (Oryza sativa L.), providing favorable conditions for its growth and promoting its growth and development; prolongs the photosynthetic functioning period of leaves; and lays the material foundation for high yields of rice. ZJW-6 not only directly participates in cellulose degradation as degrading bacteria but also induces positive interactions between bacteria and fungi and enriches the microbial taxa that were related to straw degradation, enhancing the rate of rice straw degradation. Taken together, ZJW-6 has important biological potential and should be further studied, which will provide new insights and strategies for the appropriate treatment of rice straw. In the future, this degrading bacteria may provide a better opportunity to manage straw in an ecofriendly manner.

Cordyceps cateniannulata: An endophyte of coffee, a parasite of coffee leaf rust and a pathogen of coffee pests.

Here, we report on a Cordyceps species entering into a multi-trophic, multi-kingdom association. Cordyceps cateniannulata, isolated from the stem of wild Coffea arabica in Ethiopia, is shown to function as an endophyte, a mycoparasite and an entomopathogen. A detailed polyphasic taxonomic study, including a multilocus phylogenetic analysis, confirmed its identity. An emended description of C. cateniannulata is provided herein. Previously, this species was known as a pathogen of various insect hosts in both the Old and New World. The endophytic status of C. cateniannulata was confirmed by re-isolating it from inoculated coffee plants. Inoculation studies have further shown that C. cateniannulata is a mycoparasite of Hemileia vastatrix, as well as an entomopathogen of major coffee pests; infecting and killing Hypothenemus hampei and Leucoptera coffeella. This is the first record of C. cateniannulata from Africa, as well as an endophyte and a mycoparasite. The implications for its use as a biocontrol agent are discussed.

Host cell wall composition and localized microenvironment implicated in resistance to basal stem degradation by lettuce drop (Sclerotinia minor).

BACKGROUND: Sclerotinia spp. are generalist fungal pathogens, infecting over 700 plant hosts worldwide, including major crops. While host resistance is the most sustainable and cost-effective method for disease management, complete resistance to Sclerotinia diseases is rare. We recently identified soft basal stem as a potential susceptibility factor to Sclerotinia minor infection in lettuce (Lactuca sativa) under greenhouse conditions. RESULTS: Analysis of stem and root cell wall composition in five L. sativa and one L. serriola accessions with varying growth habits and S. minor resistance levels revealed strong association between hemicellulose constituents, lignin polymers, disease phenotypes, and basal stem mechanical strength. Accessions resistant to basal stem degradation consistently exhibited higher levels of syringyl, guaiacyl, and xylose, but lower levels of fucose in stems. These findings suggest that stem cell wall polymers recalcitrant to breakdown by lignocellulolytic enzymes may contribute to stem strength-mediated resistance against S. minor. CONCLUSIONS: The lignin content, particularly guaiacyl and syringyl, along with xylose could potentially serve as biomarkers for identifying more resistant lettuce accessions and breeding lines. Basal stem degradation by S. minor was influenced by localized microenvironment conditions around the stem base of the plants.

Effects of arbuscular mycorrhizal fungi in the rhizosphere of two olive (Olea europaea) varieties Arbequina and Barnea under water deficit conditions.

One strategy to improve olive (Olea europaea ) tree drought tolerance is through the symbiosis of arbuscular mycorrhizal fungi (AMF), which helps alleviate water deficit through a combination of morphophysiological effects. Cuttings of olive varieties Arbequina (A) and Barnea (B) were grown with (+AMF) or without (-AMF) inoculum in the olive grove rhizosphere soil. One year after establishment, pots were exposed to four different water regimes: (1) control (100% of crop evapotranspiration); (2) short-period drought (20days); (3) long-period drought (25days); and (4) rewatering (R). To evaluate the influence of AMF on tolerance to water stress, stem water potential, stomatal conductance and the biomarkers for water deficit malondialdehyde, proline, soluble sugars, phenols, and flavonoids were evaluated at the end of the irrigation regimes. Stem water potential showed higher values in A(+) and B(+) in all water conditions, and the opposite was true for stomatal conductance. For proline and soluble sugars, the stem water potential trend is repeated with some exceptions. AMF inoculum spore communities from A(+ and -) and B(+ and -) were characterised at the morphospecies level in terms of richness and abundance. Certain morphospecies were identified as potential drought indicators. These results highlight that the benefits of symbiotic relationships between olive and native AMF can help to mitigate the effects of abiotic stress in soils affected by drought.

Agave Wilt Susceptibility by Reduction of Free Hexoses in Root Tissue of Agave tequilana Weber var. azul Commercial Plants in the Fructan Accumulation Process.

Agave tequilana stems store fructan polymers, the main carbon source for tequila production. This crop takes six or more years for industrial maturity. In conducive conditions, agave wilt disease increases the incidence of dead plants after the fourth year. Plant susceptibility induced for limited photosynthates for defense is recognized in many crops and is known as "sink-induced loss of resistance". To establish whether A. tequilana is more prone to agave wilt as it ages, because the reduction of water-soluble carbohydrates in roots, as a consequence of greater assembly of highly polymerized fructans, were quantified roots sucrose, fructose, and glucose, as well as fructans in stems of agave plants of different ages. The damage induced by inoculation with Fusarium solani or F. oxysporum in the roots or xylem bundles, respectively, was recorded. As the agave plant accumulated fructans in the stem as the main sink, the amount of these hexoses diminished in the roots of older plants, and root rot severity increased when plants were inoculated with F. solani, as evidence of more susceptibility. This knowledge could help to structure disease management that reduces the dispersion of agave wilt, dead plants, and economic losses at the end of agave's long crop cycle.

Evaluation of genetic variation and host resistance to wheat stem rust pathogen (Puccinia graminis f. sp. tritici) in bread wheat (Triticum aestivum L.) varieties grown in Türkiye.

Wheat stem rust, which is caused by Puccinia graminis f. sp. tritici (Pgt), is a highly destructive disease that affects wheat crops on a global scale. In this study, the reactions of 150 bread wheat varieties were evaluated for natural Pgt infection at the adult-plant stage in the 2019-2020 and 2020-2021 growing seasons, and they were analyzed using specific molecular markers to detect stem rust resistance genes (Sr22, Sr24, Sr25, Sr26, Sr31, Sr38, Sr50, and Sr57). Based on phenotypic data, the majority of the varieties (62%) were resistant or moderately resistant to natural Pgt infection. According to molecular results, it was identified that Sr57 was present in 103 varieties, Sr50 in nine varieties, Sr25 in six varieties, and Sr22, Sr31, and Sr38 in one variety each. Additionally, their combinations Sr25 + Sr50, Sr31 + Sr57, Sr38 + Sr50, and Sr38 + Sr57 were detected in these varieties. On the other hand, Sr24 and Sr26 were not identified. In addition, many varieties had low stem rust scores, including a large minority that lacked Sr57. These varieties must have useful resistance to stem rust and could be the basis for selecting greater, possibly durable resistance.

Metataxonomic analysis of endophytic bacteria of blackberry (Rubus ulmifolius Schott) across tissues and environmental conditions.

(1) Background: Endophytic bacteria represent an important component of plant wellness. They have been widely studied for their involvement in plant development and enhancement of stress tolerance. In this work, the endophytic communities of roots, stems, and leaves of blackberry (Rubus ulmifolius Schott) were studied in three different niches: natural, riverside, and human-impacted niches. (2) Results: The microbiome composition revealed that Sphingomonadaceae was the most abundant family in all samples, accounting for 9.4-45.8%. In contrast, other families seem to be linked to a specific tissue or niche. Families Microbacteriaceae and Hymenobacteraceae increased their presence in stem and leaf samples, while Burkholderiaceae abundance was important in riverside samples. Alpha and beta diversity analyses showed that root samples were the most diverse, and they gathered together in the same cluster, apart from the rest of the samples. (3) Conclusions: The analysis of the microbiome of R. ulmifolius plants revealed that the composition was essentially the same in different niches; the differences were primarily influenced by plant tissue factors with a core genome dominated by Sphingomonadaceae. Additionally, it was observed that R. ulmifolius can select its own microbiome, and this remains constant in all tissues evaluated regardless the niche of sampling.

Differential response of bacteria and fungi to drought on the decomposition of Sarcocornia fruticosa woody stems in a saline stream.

Inland saline ecosystems suffer multiple stresses (e.g., high radiation, salinity, water scarcity) that may compromise essential ecosystem functions such as organic matter decomposition. Here, we investigated the effects of drought on microbial colonization and decomposition of Sarcocornia fruticosa woody stems across different habitats in a saline watershed: on the dry floodplain, submerged in the stream channel and at the shoreline (first submerged, then emerged). Unexpectedly, weight loss was not enhanced in the submerged stems, while decomposition process differed between habitats. On the floodplain, it was dominated by fungi and high cellulolytic activity; in submerged conditions, a diverse community of bacteria and high ligninolytic activity dominated; and, on the shoreline, enzyme activities were like submerged conditions, but with a fungal community similar to the dry conditions. Results indicate distinct degradation paths being driven by different stress factors: strong water scarcity and photodegradation in dry conditions, and high salinity and reduced oxygen in wet conditions. This suggests that fungi are more resistant to drought, and bacteria to salinity. Overall, in saline watersheds, variations in multiple stress factors exert distinct environmental filters on bacteria and fungi and their role in the decomposition of plant material, affecting carbon cycling and microbial interactions.

Spatial distribution and community composition of endophytic fungi within Mussaenda pubescens stems.

Endophytic fungi, pivotal in facilitating plant co-evolution, significantly enhance plant growth, stress resistance, and environmental adaptability. Despite their importance, the spatial distribution of stem endophytic fungi (SEF) within host plants remains poorly characterized. Here, we employed high-throughput sequencing to conduct a comparative analysis of SEF communities in Mussaenda pubescens on a regional scale. Our findings reveal that whole-SEF communities were overwhelmingly dominated by members of the phylum Ascomycota, accounting for 85.9 %, followed by Basidiomycota at 13.9 %, and that alpha diversity within the whole-SEF community of M. pubescens remains relatively consistent across sampling sites. However, significant variation was observed within conditionally abundant taxa (CAT), conditionally rare or abundant taxa (CRAT), and conditionally rare taxa (CRT). Climatic factors emerged as the primary influence on SEF community distribution, followed by spatial distance and stem chemical properties. Neutral community modeling results suggested that both stochastic and deterministic processes play a role in shaping whole-SEF communities, with deterministic processes having a stronger influence on CRT subcommunities. Furthermore, the CRT co-occurrence network exhibited a more complex structure, characterized by higher values of network betweenness and degree relative to CAT and CRAT subcommunities. These findings enhance our understanding of community assembly and ecological interactions between stem fungal endophytes, presenting opportunities for harnessing fungal resources for the benefit of humanity.

Unsaturated fatty acid, Nonacosenoic acid isolated from an endophyte Chaetomium nigricolor inhabiting the stem of Catharanthus roseus and its bioactivity.

The endophytic fungus Chaetomium nigricolor culture filtrate's hexane extract was used to identify a cytotoxic very long-chain fatty acid. Based on multiple spectroscopic investigations, the structure of the compound was predicted to be an unsaturated fatty acid, Nonacosenoic acid (NA). Using the MTT assay, the compound's cytotoxic potential was evaluated against MCF-7, A-431, U-251, and HEK-293 T cells. The compound was moderately cytotoxic to breast carcinoma cell line, MCF-7 cells and negligibly cytotoxic to non-cancerous cell line HEK-293 T cells. The compound exhibited mild cytotoxic activity against A-431 and U-251 cells. The compound also induced ROS generation and mitochondrial depolarization in MCF-7 cells when assessed via the NBT and JC-1 assays, respectively. This is the first report on the production of nonacosenoic acid from the endophytic fungus Chaetomium nigricolor and the assessment of its bioactivity.

Resurgence of wheat stem rust infections in western Europe: causes and how to curtail them.

Ten years ago, (black) stem rust - the most damaging of wheat (Triticum aestivum) rusts - re-emerged in western Europe. Disease incidences have since increased in scale and frequency. Here, we investigated the likely underlying causes and used those to propose urgently needed mitigating actions. We report that the first large-scale UK outbreak of the wheat stem rust fungus, Puccinia graminis f. sp. tritici (Pgt), in 2022 may have been caused by timely arrival of airborne urediniospores from southwest Europe. The drive towards later-maturing wheat varieties in the UK may be exacerbating Pgt incidences, which could have disastrous consequences. Indeed, infection assays showed that two UK Pgt isolates from 2022 could infect over 96% of current UK wheat varieties. We determined that the temperature response data in current disease risk simulation models are outdated. Analysis of germination rates for three current UK Pgt isolates showed substantial variation in temperature response functions, suggesting that the accuracy of disease risk simulations would be substantially enhanced by incorporating data from prevailing Pgt isolates. As Pgt incidences continue to accelerate in western Europe, we advocate for urgent action to curtail Pgt losses and help safeguard future wheat production across the region.

Identification of Candidate Avirulence and Virulence Genes Corresponding to Stem Rust (Puccinia graminis f. sp. tritici) Resistance Genes in Wheat.

Stem rust, caused by the biotrophic fungal pathogen Puccinia graminis f. sp. tritici (Pgt), is an important disease of wheat. However, the majority of Pgt virulence/avirulence loci and underlying genes remain uncharacterized due to the constraints of developing bi-parental populations with this obligate biotroph. Genome-wide association studies (GWAS) using a sexual Pgt population mainly collected from the Pacific Northwestern United States were used to identify candidate virulence/avirulence effector genes corresponding to the six wheat Sr genes: Sr5, Sr21, Sr8a, Sr17, Sr9a, and Sr9d. The Pgt isolates were genotyped using whole-genome shotgun sequencing that identified approximately 1.2 million single nucleotide polymorphisms (SNPs) and were phenotyped at the seedling stage on six Sr gene differential lines. Association mapping analyses identified 17 Pgt loci associated with virulence or avirulence phenotypes on six Pgt resistance genes. Among these loci, 16 interacted with a specific Sr gene, indicating Sr-gene specific interactions. However, one avirulence locus interacted with two separate Sr genes (Sr9a and Sr17), suggesting two distinct Sr genes identifying a single avirulence effector. A total of 24 unique effector gene candidates were identified, and haplotype analysis suggests that within this population, AvrSr5, AvrSr21, AvrSr8a, AvrSr17, and AvrSr9a are dominant avirulence genes, while avrSr9d is a dominant virulence gene. The putative effector genes will be fundamental for future effector gene cloning efforts, allowing for further understanding of rust effector biology and the mechanisms underlying virulence evolution in Pgt with respect to race-specific R-genes. [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.

Proteo-metabolomic Dissection of Extracellular Matrix Reveals Alterations in Cell Wall Integrity and Calcium Signaling Governs Wall-Associated Susceptibility during Stem Rot Disease in Jute.

The plant surveillance system confers specificity to disease and immune states by activating distinct molecular pathways linked to cellular functionality. The extracellular matrix (ECM), a preformed passive barrier, is dynamically remodeled at sites of interaction with pathogenic microbes. Stem rot, caused by Macrophomina phaseolina, adversely affects fiber production in jute. However, how wall related susceptibility affects the ECM proteome and metabolome remains undetermined in bast fiber crops. Here, stem rot responsive quantitative temporal ECM proteome and metabolome were developed in jute upon M. phaseolina infection. Morpho-histological examination revealed that leaf shredding was accompanied by reactive oxygen species production in patho-stressed jute. Electron microscopy showed disease progression and ECM architecture remodeling due to necrosis in the later phase of fungal attack. Using isobaric tags for relative and absolute quantitative proteomics and liquid chromatography-tandem mass spectrometry, we identified 415 disease-responsive proteins involved in wall integrity, acidification, proteostasis, hydration, and redox homeostasis. The disease-related correlation network identified functional hubs centered on α-galactosidase, pectinesterase, and thaumatin. Gas chromatography-mass spectrometry analysis pointed toward enrichment of disease-responsive metabolites associated with the glutathione pathway, TCA cycle, and cutin, suberin, and wax metabolism. Data demonstrated that wall-degrading enzymes, structural carbohydrates, and calcium signaling govern rot responsive wall-susceptibility. Proteomics data were deposited in Pride (PXD046937; PXD046939).

Microbial communities of Schisandra sphenanthera Rehd. et Wils. and the correlations between microbial community and the active secondary metabolites.

Background: Schisandra sphenanthera Rehd. et Wils. is a plant used in traditional Chinese medicine (TCM). However, great differences exist in the content of active secondary metabolites in various parts of S. sphenanthera. Do microorganisms critically influence the accumulation of active components in different parts of S. sphenanthera? Methods: In this study, 16S/ITS amplicon sequencing analysis was applied to unravel microbial communities in rhizospheric soil and different parts of wild S. sphenanthera. At the same time, the active secondary metabolites in different parts were detected, and the correlation between the secondary metabolites and microorganisms was analyzed. Results: The major components identified in the essential oils were sesquiterpene and oxygenated sesquiterpenes. The contents of essential oil components in fruit were much higher than that in stem and leaf, and the dominant essential oil components were different in these parts. The dominant components of the three parts were γ-muurolene, δ-cadinol, and trans farnesol (stem); α-cadinol and neoisolongifolene-8-ol (leaf); isosapathulenol, α-santalol, cedrenol, and longiverbenone (fruit). The microbial amplicon sequences were taxonomically grouped into eight (bacteria) and seven (fungi) different phyla. Community diversity and composition analyses showed that different parts of S. sphenanthera had similar and unique microbial communities, and functional prediction analysis showed that the main functions of microorganisms were related to metabolism. Moreover, the accumulation of secondary metabolites in S. sphenanthera was closely related to the microbial community composition, especially bacteria. In endophytic bacteria, Staphylococcus and Hypomicrobium had negative effects on five secondary metabolites, among which γ-muurolene and trans farnesol were the dominant components in the stem. That is, the dominant components in stems were greatly affected by microorganisms. Our results provided a new opportunity to further understand the effects of microorganisms on the active secondary metabolites and provided a basis for further research on the sustainable utilization of S. sphenanthera.

Modelling metabolic fluxes of tomato stems reveals that nitrogen shapes central metabolism for defence against Botrytis cinerea.

Among plant pathogens, the necrotrophic fungus Botrytis cinerea is one of the most prevalent, leading to severe crop damage. Studies related to its colonization of different plant species have reported variable host metabolic responses to infection. In tomato, high N availability leads to decreased susceptibility. Metabolic flux analysis can be used as an integrated method to better understand which metabolic adaptations lead to effective host defence and resistance. Here, we investigated the metabolic response of tomato infected by B. cinerea in symptomless stem tissues proximal to the lesions for 7 d post-inoculation, using a reconstructed metabolic model constrained by a large and consistent metabolic dataset acquired under four different N supplies. An overall comparison of 48 flux solution vectors of Botrytis- and mock-inoculated plants showed that fluxes were higher in Botrytis-inoculated plants, and the difference increased with a reduction in available N, accompanying an unexpected increase in radial growth. Despite higher fluxes, such as those involved in cell wall synthesis and other pathways, fluxes related to glycolysis, the tricarboxylic acid cycle, and amino acid and protein synthesis were limited under very low N, which might explain the enhanced susceptibility. Limiting starch synthesis and enhancing fluxes towards redox and specialized metabolism also contributed to defence independent of N supply.

Effective Control of Sclerotinia Stem Rot in Canola Plants Through Application of Exogenous Hairpin RNA of Multiple Sclerotinia sclerotiorum Genes.

Sclerotinia stem rot is a globally destructive plant disease caused by Sclerotinia sclerotiorum. Current management of Sclerotinia stem rot primarily relies on chemical fungicides and crop rotation, raising environmental concerns. In this study, we developed an eco-friendly RNA bio-fungicide targeting S. sclerotiorum. Six S. sclerotiorum genes were selected for double-stranded RNA (dsRNA) synthesis. Four genes, a chitin-binding domain, mitogen-activated protein kinase, oxaloacetate acetylhydrolase, and abhydrolase-3, were combined to express hairpin RNA in Escherichia coli HT115. The effect of application of total RNA extracted from E. coli HT115 expressing hairpin RNA on disease progressive and necrosis lesions was evaluated. Gene expression analysis using real-time PCR showed silencing of the target genes using 5 ng/µl of dsRNA in a fungal liquid culture. A detached leaf assay and greenhouse application of dsRNA on canola stem and leaves showed variation in the reduction of necrosis symptoms by dsRNA of different genes, with abhydrolase-3 being the most effective. The dsRNA from a combination of four genes reduced disease severity significantly (P = 0.01). Plants sprayed with hairpin RNA from four genes had lesions that were almost 30% smaller than those of plants treated with abhydrolase-3 alone, in lab and greenhouse assays. The results of this study highlight the potential of RNA interference to manage diseases caused by S. sclerotiorum; however, additional research is necessary to optimize its efficacy.

Fusarium amaranthi sp. nov. from Amaranth Is an Emergent Species Closely Related to F. circinatum.

Amaranth (Amaranthus spp. L) is not native to South Korea but is cultivated in small scales for ornamental purposes as well as leafy vegetables and pseudo cereals. In this study, a new species within the genus Fusarium was isolated from amaranth, showing stem rot symptoms from a farmer field in Hwaseong, South Korea. The disease is characterized by dark-brown spots with black borders, leading to withering. Phylogenetic analysis-based concatenated sequences of translation elongation factor 1-alpha (TEF1), beta-tubulin (tub2), calmodulin (cmdA), RNA polymerase largest subunit (RPB1), and RNA polymerase II second largest subunit (RPB2) genes revealed that the obtained isolates formed a distinct clad within the Fusarium fujikuroi species complex and is closely related to F. circinatum. Cultural and morphological characteristics and pathogenicity on healthy amaranth plants (stem and leaves) were examined. The isolates readily differentiated from F. circinatum based on one- to five-septate macroconidia and the absence of sterile hyphae. Based on molecular and morphological characteristics, this fungus is demonstrated to be a new species and is described here as F. amaranthi, the causal agent of stem rot of amaranth in South Korea.

Effective, Consistent, and Rapid Noncontact Application Methods for Seedling Basal Stem Infection by Sclerotinia sclerotiorum.

Sclerotinia sclerotiorum (Lib.) de Bary, an economically devastating soilborne fungal pathogen known to cause disease across a wide range of plants, produces long-term inoculum called sclerotia that can germinate either carpogenically by ascospores infecting aboveground plant parts or myceliogenically to infect stem base and roots. Typically, for research purposes, S. sclerotiorum diseases are initiated by direct contact methods, using S. sclerotiorum mycelium agar plugs wrapped around the stem or sclerotia placed directly beneath root mass. However, reproducible noncontact methods leading to basal stem infection are not currently available. Therefore, the objective of this study was to develop effective noncontact protocols that consistently generate basal plant stem infection from S. sclerotiorum in the soil. Using three host plant species (canola, lupin, and lettuce), we determined two methods that reliably produced basal stem infection. The first method, where mycelial agar plugs were positioned just below the soil surface at a distance of 5 mm from each seedling, led to 100% infection in all plants. The second method used pathogen-infested soil by mixing the soil with dry inoculum in the form of a powder prepared from mycelium-colonized organic substrates. Four substrates consistently produced 100% seedling infection at 4 days after inoculation (DAI): wheat bran, wheat grain, red rice, and hulled millet. In contrast, chia, canary, sesame, and ryegrass seed substrates resulted in less than 50% seedling infection at 10 DAI, and infection levels did not progress further. The two soil inoculation methods outlined in this study will enhance future research on the progression of S. sclerotiorum diseases, with the potential to screen disease-resistant host genotypes to basal S. sclerotiorum infection and, in particular, to test the effectiveness of soil applications of fungicides or biocontrol agents against S. sclerotiorum basal infection.

Effect of Fungicide and Timing of Application on Management of Phoma Black Stem of Cultivated Sunflowers in the United States.

Phoma black stem (PBS), caused by Phoma macdonaldii Boerema (teleomorph Leptosphaeria lindquistii Frezzi), is the most common stem disease of sunflower (Helianthus annuus L.) in the northern Great Plains region of the United States. However, the impact of PBS on sunflower yield in the United States is unclear, and a near complete absence of information on the impact of fungicides on disease management exists. The objectives of this study were to determine the impact of PBS on sunflower yield, the efficacy of available fungicides, the optimal fungicide application timing, and the economic viability of fungicides as a management tool. Fungicide timing efficacy was evaluated by applying single and/or sequential applications of pyraclostrobin fungicide at three sunflower growth stages in 10 field trials between 2017 and 2019. Efficacy of 10 fungicides from the Fungicide Resistance Action Committee (FRAC) groups 3, 7, and 11 were evaluated in four field trials between 2018 and 2019. The impact of treatments on PBS were evaluated by determination of incidence, severity, maximum lesion height, disease severity index (DSI), and harvested yield. Nine of the 10 fungicides evaluated and all fungicide timings that included an early bud application resulted in disease reductions when compared with the nontreated controls. The DSI was negatively correlated to sunflower yield in high-yield environments (P = 0.0004; R2 = 0.3425) but not in low- or moderate-yield environments. Although FRAC 7 fungicides were generally most efficacious, the sufficient efficacy and lower cost of FRAC 11 fungicides make them more economically viable in high-yielding environments at current market conditions.