Proteome-Wide Response of Dormant Caryopses of the Weed, Avena fatua, After Colonization by a Seed-Decay Isolate of Fusarium avenaceum.

Promoting seed decay is an ecological approach to reducing weed persistence in the soil seedbank. Previous work demonstrated that Fusarium avenaceum F.a.1 decays dormant Avena fatua (wild oat) caryopses and induces several defense enzyme activities in vitro. The objectives of this study were to obtain a global perspective of proteins expressed after F.a.1-caryopsis colonization by conducting proteomic evaluations on (i) leachates, soluble extrinsic (seed-surface) proteins released upon washing caryopses in buffer and (ii) proteins extracted from whole caryopses; interactions with aluminum (Al) were also evaluated in the latter study because soil acidification and associated metal toxicity are growing problems. Of the 119 leachate proteins classified as defense/stress, 80 were induced or repressed. Defense/stress proteins were far more abundant in A. fatua (35%) than in F.a.1 (12%). Avena defense/stress proteins were also the most highly regulated category, with 30% induced and 35% repressed by F.a.1. Antifungal proteins represented 36% of Avena defense proteins and were the most highly regulated, with 36% induced and 37% repressed by F.a.1. These results implicate selective regulation of Avena defense proteins by F.a.1. Fusarium proteins were also highly abundant in the leachates, with 10% related to pathogenicity, 45% of which were associated with host cell wall degradation. In whole caryopsis extracts, fungal colonization generally resulted in induction of a similar set of Avena proteins in the presence and absence of Al. Results advance the hypothesis that seed decay pathogens elicit intricate and dynamic biochemical responses in dormant seeds.

Rhizoctonia Resistance Is Negatively Correlated to Early Root Growth Rate in Synthetic Hexaploid Wheat Derivatives.

Resistance to the soilborne fungal pathogen Rhizoctonia solani AG-8 is desirable in adapted wheat and barley but remains an elusive trait for prebreeders and breeders. In a previous study, we observed that emergence and root growth was faster in the Rhizoctonia-susceptible 'Scarlet' than in its resistant counterpart, 'Scarlet-Rz1'. The objective of the current study was to quantify early root growth rate and total root length in resistant and susceptible synthetic hexaploid wheat lines, including parental lines and 22 recombinant inbred lines derived crosses between parental lines. In Petri dish assays, the susceptible lines displayed a faster rate of root growth during the first 40 h of root emergence compared with resistant lines. This growth differential was observed in 14-day and 48-h greenhouse assays, in which the total root length of susceptible parental lines was significantly (P < 0.05) greater than that of resistant parental lines. However, the resistant lines sustained less root loss compared with susceptible lines when R. solani AG-8 was present in the soil. Early root growth rate and total root length were not correlated to freezing tolerance in a set of wheat cultivars selected for cold tolerance. The findings indicated that early root growth was negatively correlated to R. solani AG-8 damage in resistant synthetic wheat lines developed for the Pacific Northwest, United States, and suggested that the dynamics of root emergence affect resistance to this soilborne pathogen.

Potential Biocontrol Activities of Populus Endophytes against Several Plant Pathogens Using Different Inhibitory Mechanisms.

The plant microbiome can be used to bolster plant defense against abiotic and biotic stresses. Some strains of endophytes, the microorganisms within plants, can directly inhibit the growth of plant fungal pathogens. A previously isolated endophyte from wild Populus (poplar), WPB of the species Burkholderia vietnamiensis, had robust in vitro antifungal activity against pathogen strains that are highly virulent and of concern to Pacific Northwest agriculture: Rhizoctonia solani AG-8, Fusarium culmorum 70110023, and Gaemannomyces graminis var. tritici (Ggt) ARS-A1, as well as activity against the oomycete, Pythium ultimum 217. A direct screening method was developed for isolation of additional anti-fungal endophytes from wild poplar extracts. By challenging pathogens directly with dilute extracts, eleven isolates were found to be inhibitory to at least two plant pathogen strains and were therefore chosen for further characterization. Genomic analysis was conducted to determine if these endophyte strains harbored genes known to be involved in antimicrobial activities. The newly isolated Bacillus strains had gene clusters for production of bacillomycin, fengicyn, and bacillibactin, while the gene cluster for the synthesis of sessilin, viscosin and tolaasin were found in the Pseudomonas strains. The biosynthesis gene cluster for occidiofungin (ocf) was present in the Burkholderia vietnamiensis WPB genome, and an ocf deletion mutant lost inhibitory activity against 3 of the 4 pathogens. The new isolates lacked the gene cluster for occidiofungin implying they employ different modes of action. Other symbiotic traits including nitrogen fixation, phosphate solubilization, and the production of auxins and siderophores were investigated. Although it will be necessary to conduct in vivo tests of the candidates with pathogen-infected agricultural crops, the wild poplar tree microbiome may be a rich source of beneficial endophyte strains with potential for biocontrol applications against a variety of pathogens and utilizing varying modes of action.

Native yeast and non-yeast fungal communities of Cabernet Sauvignon berries from two Washington State vineyards, and persistence in spontaneous fermentation.

To address a knowledge gap about the grape berry mycobiome from Washington State vineyards, next-generation sequencing of the internal transcribed spacer region (ITS1) was used to identify native yeast and fungal species on berries of cultivar 'Cabernet Sauvignon' from two vineyards at veraison and harvest in 2015 and 2016. Four hundred fifty-six different yeast amplicon sequence variants (ASV), representing 184 distinct taxa, and 2467 non-yeast fungal ASV (791 distinct taxa) were identified in this study. A set of 50 recurrent yeast taxa, including Phaeococcomyces, Vishniacozyma and Metschnikowia, were found at both locations and sampling years. These yeast species were monitored from the vineyard into laboratory-scale spontaneous fermentations. Taxa assignable to Metschnikowia and Saccharomyces persisted during fermentation, whereas Curvibasidium, which also has possible impact on biocontrol and wine quality, did not. Sulfite generally reduced yeast diversity and richness, but its effect on the abundance of specific yeasts during fermentation was negligible. Among the 106 recurring non-yeast fungal taxa, Alternaria, Cladosporium and Ulocladium were especially abundant in the vineyard. Vineyard location was the primary factor that accounted for the variation among both communities, followed by year and berry developmental stage. The Washington mycobiomes were compared to those from other parts of the world. Sixteen recurrent yeast species appeared to be unique to Washington State vineyards. This subset also contained a higher proportion of species associated with cold and extreme environments, relative to other localities. Certain yeast and non-yeast fungal species known to suppress diseases or modify wine sensory properties were present in Washington vineyards, and likely have consequences to vineyard health and wine quality.

Purinoceptor P2K1/DORN1 Enhances Plant Resistance Against a Soilborne Fungal Pathogen, Rhizoctonia solani.

The purinoceptor P2K1/DORN1 recognizes extracellular ATP, a damage-associated molecular pattern (DAMP) released upon cellular disruption by wounding and necrosis, which in turn, boost plant innate immunity. P2K1 is known to confer plant resistance to foliar biotrophic, hemi-biotrophic, and necrotrophic pathogens. However, until now, no information was available on its function in defense against root pathogens. In this report, we describe the contribution of P2K1 to resistance in Arabidopsis against Rhizoctonia solani, a broad host range, necrotrophic soilborne fungal pathogen. In pot assays, the Arabidopsis P2K1 overexpression line OxP2K1 showed longer root length and a greater rosette surface area than wild type in the presence of the pathogen. In contrast, the knockout mutant dorn1-3 and the double mutant rbohd/f, defective in two subunits of the respiratory burst complex NADPH oxidase, exhibited significant reductions in shoot and root lengths and rosette surface area compared to wild type when the pathogen was present. Expression of PR1, PDF1.2, and JAZ5 in the roots was reduced in dorn1-3 and rbohd/f and elevated in OxP2K1 relative to wild type, indicating that the salicylate and jasmonate defense signaling pathways functioned in resistance. These results indicated that a DAMP-mediated defense system confers basal resistance against an important root necrotrophic fungal pathogen.

Real-time PCR quantification of Rhizoctonia solani AG-3 from soil samples.

Rhizoctonia solani anastomosis group 3 (AG-3) causes several diseases of potato, including black scurf and stem canker, affecting potato production in the Skagit Valley, Washington, and around the world. Primers for a SYBR-Green II-based real-time polymerase chain reaction (qPCR) assay were designed from sequences of the nuclear internal transcribed spacer (ITS) regions of fungal isolates of potato and onion from the Pacific Northwest, USA. The primers preferentially amplified R. solani AG-3 DNA, compared to DNA from R. solani AG-4, AG-5 and AG-8. In silico analysis of primer-template duplex stability indicated that the assay also will detect R. solani AG-3 isolates from pea and onion in Washington State and from diverse crop species around the world, but not R. solani AG-9 and AG-2-1. The assay was used to quantify R. solani AG-3 populations in pathogen-infested field soils after temporary flooding rotation, a practice found to be effective for reducing Sclerotinia sclerotiorum and R. solani AG-3 in potatoes in growth chamber studies. Population densities of the pathogen were not significantly reduced in saturated (flooded) soils relative to fallow. However, the qPCR approach was more sensitive and quantitative than the toothpick baiting method for diagnosis of these soil samples. Accurate detection and quantification of R. solani AG-3 in soil will facilitate the development of integrated management plans for Rhizoctonia diseases of potato.

Chronic Sublethal Aluminum Exposure and Avena fatua Caryopsis Colonization Influence Gene Expression of Fusarium avenaceum F.a.1.

Fusarium avenaceum F.a.1 is a novel strain of a fungal plant pathogen capable of preferentially decaying wild oat (Avena fatua) caryopses compared with those of wheat (Triticum aestivum). Understanding the molecular mechanisms governing weed seed-pathogen interactions is crucial to developing novel weed seed suppression technologies. Additionally, wild oat often competes with wheat in regions undergoing soil acidification, which leads to increases in soluble concentrations of many metals, including aluminum (Al). There is a dearth of information regarding the gene expression responses of Fusarium species to Al toxicity, or how metal toxicity might influence caryopsis colonization. To address this, a transcriptomic approach was used to investigate molecular responses of F.a.1 during wild oat caryopsis colonization in the presence and absence of chronic, sublethal concentrations of Al (400 µM). Caryopsis colonization was associated with induction of genes related to virulence, development, iron metabolism, oxidoreduction, stress, and detoxification, along with repression of genes associated with development, transport, cell-wall turnover, and virulence. Caryopsis colonization during Al exposure resulted in the induction of genes associated with virulence, detoxification, stress, iron metabolism, oxidoreduction, and cell wall turnover, along with repression of genes associated with cell wall metabolism, virulence, development, detoxification, stress, and transcriptional regulation. Aluminum exposure in the absence of caryopses was associated with induction of genes involved in siderophore biosynthesis, secretion, uptake, and utilization, along with several other iron metabolism-related and organic acid metabolism-related genes. The siderophore-related responses associated with Al toxicity occurred concurrently with differential regulation of genes indicating disruption of iron homeostasis. These findings suggest Al toxicity is attenuated by siderophore metabolism in F.a.1. In summary, both caryopsis colonization and Al toxicity uniquely influence transcriptomic responses of F.a.1.

Real-time PCR assays for the quantification of native yeast DNA in grape berry and fermentation extracts.

Native yeasts comprise part of the microbial community in grape vineyards and play roles in alcoholic fermentation and wine quality. Monitoring populations of native yeast in vineyards, during fermentation and after bottling will provide viticulturalists and oenologists with information needed to help control spoilage and to enhance desirable wine properties. This is especially crucial for low-intervention winemaking, in which fermentation is driven by native rather than starter microbes. In this study, we report real-time polymerase chain reaction (qPCR) assays for rapid quantification of seven grape yeast species or species combinations that occur in vineyards of Washington State and throughout the world. The assays targeted Candida californica, Curvibasidium pallidicorallinum, Metschnikowia spp., Meyerozyma caribbica/Me. guilliermondii, and Saccharomyces cerevisiae/S. bayanus. We also developed assays for the spoilage yeast Brettanomyces bruxellensis, and the yeast-like fungus Aureobasidium pullulans. Primers were designed for sequences in the internal transcribed spacer (ITS) and large ribosome subunit (LSU) gene. Known populations of yeast cells, added to fermentation extract, were significantly correlated to amounts of purified DNA in picograms (pg) for most of the yeasts; exceptions were A. pullulans and Cu. pallidicorallinum. The utility of the Metschnikowia, Meyerozyma and Saccharomyces assays was further validated by good correlations (R2 = 0.75-0.83) between the number of target sequences and pg of DNA from qPCR for selected vineyard and fermentation samples. Overall, the assays will aid in species identification and monitoring of specific yeasts from cultures, vineyards and fermentation samples. Topics: Food Microbiology, Microbiological Method.

Real-time PCR quantification of Fusarium avenaceum in soil and seeds.

The pathogenic fungus Fusarium avenaceum infects a broad range of plant hosts across the globe. While primarily soilborne, F. avenaceum can colonize all plant tissues, including buds, seeds, fruits, stems, crowns, and roots, resulting in significant crop yield reductions and economic losses for growers. In addition to its impact on crop productivity, F. avenaceum produces toxic metabolites that can be transferred to humans and livestock through consumption of infected foods. The ability of F. avenaceum to cause seed decay may be utilized to deplete the weed seedbank in soil, an important integrated weed management strategy. We developed a SYBR Green I-based real-time polymerase chain reaction (qPCR) assay to efficiently detect and quantify F. avenaceum in soil, wild oat (Avena fatua L.) seed caryopses, and wild oat seed hulls. The primer pair was designed from the translation elongation factor 1-alpha (TEF1) gene. In silico and wet lab testing were done to assess the ability of the primers to bind TEF1 sequences from Fusarium spp. and common soil fungi. The findings indicated that the primers were specific to F. avenaceum, and also recognized GenBank TEF1 accessions annotated as F. arthrosporioides, which has been listed as a foliar pathogen of wheat in Oregon, and conspecific with F. avenaceum. Standard curves of F. avenaceum DNA diluted with soil, caryopsis, or hull extracts indicated primer amplification efficiency was not significantly affected by PCR inhibitors. This real-time PCR assay effectively assesses the presence and abundance of F. avenaceum and its close relative F. arthrosporioides, if present, in soil and seed tissues. The assay can be used for endpoint PCR as well.

Corrigendum: An In vitro Study of Bio-Control and Plant Growth Promotion Potential of Salicaceae Endophytes.

[This corrects the article DOI: 10.3389/fmicb.2017.00386.].

Biological Control of Botrytis cinerea: Interactions with Native Vineyard Yeasts from Washington State.

Native yeasts are of increasing interest to researchers, grape growers, and vintners because of their potential for biocontrol activity and their contributions to the aroma, flavor, and mouthfeel qualities of wines. To assess biocontrol activity, we tested 11 yeasts from Washington vineyards, representing isolates of Candida saitoana, Curvibasidium pallidicorallinum, Metschnikowia chrysoperlae, M. pulcherrima, Meyerozyma guilliermondii, Saccharomyces cerevisiae, and Wickerhamomyces anomalus, for ability to colonize Thompson Seedless grape berries, inhibit the growth of Botrytis cinerea in vitro, and suppress disease symptoms on isolated berries. The yeast-like fungus Aureobasidium pullulans was also included based on its known biocontrol activity against B. cinerea in studies on apple and grape. All yeast strains multiplied rapidly in grape berries and reached densities of over log 6 cells per wound as early as 2 days after inoculation with 200 cells. One of the Botrytis isolates used in this study was much less virulent than the others and was provisionally identified as B. prunorum based on multilocus sequence analysis. Suppression of the growth of B. cinerea isolates 111bb, 207a, 207cb, and 407cb occurred on berries treated with A. pullulans P01A006, Metschnikowia chrysoperlae P34A004 and P40A002, M. pulcherrima P01A016 and P01C004, Meyerozyma guilliermondii P34D003, and S. cerevisiae HNN11516. Inhibition of Botrytis isolates by the yeast strains was more common on berries than in vitro, suggesting the possibility that niche competition was a more likely biocontrol mechanism than antibiosis in planta. Metabolic profiling of yeast strains and B. cinerea isolates using Biolog YT plates revealed seven distinct metabolic groups. Furthermore, the yeast strains showed partial to complete tolerance to the commonly used fungicides fluopyram, triflumizole, metrafenone, pyraclostrobin, and boscalid. Implications of these findings for field deployment of native Washington yeasts as biocontrol agents against B. cinerea are discussed.

An In vitro Study of Bio-Control and Plant Growth Promotion Potential of Salicaceae Endophytes.

Microbial communities in the endosphere of Salicaceae plants, poplar (Populus trichocarpa) and willow (Salix sitchensis), have been demonstrated to be important for plant growth promotion, protection from biotic and abiotic stresses, and degradation of toxic compounds. Our study aimed to investigate bio-control activities of Salicaceae endophytes against various soil borne plant pathogens including Rhizoctonia solani AG-8, Fusarium culmorum, Gaeumannomyces graminis var. tritici, and Pythium ultimum. Additionally, different plant growth promoting traits such as biological nitrogen fixation (BNF), indole-3-acetic acid (IAA) biosynthesis, phosphate solubilization, and siderophore production were assessed in all bio-control positive strains. Burkholderia, Rahnella, Pseudomonas, and Curtobacterium were major endophyte genera that showed bio-control activities in the in-vitro assays. The bio-control activities of Burkholderia strains were stronger across all tested plant pathogens as compared to other stains. Genomes of sequenced Burkholderia strains WP40 and WP42 were surveyed to identify the putative genes involved in the bio-control activities. The ocf and hcnABC gene clusters responsible for biosynthesis of the anti-fungal metabolites, occidiofungin and hydrogen cyanide, are present in the genomes of WP40 and WP42. Nearly all endophyte strains showing the bio-control activities produced IAA, solubilized tricalcium phosphate, and synthesized siderophores in the culture medium. Moreover, some strains reduced acetylene into ethylene in the acetylene reduction assay, a common assay used for BNF. Salicaceae endophytes could be useful for bio-control of various plant pathogens, and plant growth promotion possibly through the mechanisms of BNF, IAA production, and nutrient acquisition.

Defense Enzyme Responses in Dormant Wild Oat and Wheat Caryopses Challenged with a Seed Decay Pathogen.

Seeds have well-established passive physical and chemical defense mechanisms that protect their food reserves from decay-inducing organisms and herbivores. However, there are few studies evaluating potential biochemical defenses of dormant seeds against pathogens. Caryopsis decay by the pathogenic Fusarium avenaceum strain F.a.1 was relatively rapid in wild oat (Avena fatua L.) isoline "M73," with >50% decay after 8 days with almost no decay in wheat (Triticum aestivum L.) var. RL4137. Thus, this fungal strain has potential for selective decay of wild oat relative to wheat. To study defense enzyme activities, wild oat and wheat caryopses were incubated with F.a.1 for 2-3 days. Whole caryopses were incubated in assay reagents to measure extrinsic defense enzyme activities. Polyphenol oxidase, exochitinase, and peroxidase were induced in whole caryopses, but oxalate oxidase was reduced, in response to F.a.1 in both species. To evaluate whether defense enzyme activities were released from the caryopsis surface, caryopses were washed with buffer and enzyme activity was measured in the leachate. Significant activities of polyphenol oxidase, exochitinase, and peroxidase, but not oxalate oxidase, were leached from caryopses. Defense enzyme responses were qualitatively similar in the wild oat and wheat genotypes evaluated. Although the absolute enzyme activities were generally greater in whole caryopses than in leachates, the relative degree of induction of polyphenol oxidase, exochitinase, and peroxidase by F.a.1 was greater in caryopsis leachates, indicating that a disproportionate quantity of the induced activity was released into the environment from the caryopsis surface, consistent with their assumed role in defense. It is unlikely that the specific defense enzymes studied here play a key role in the differential susceptibility to decay by F.a.1 in these two genotypes since defense enzyme activities were greater in the more susceptible wild oat, compared to wheat. Results are consistent with the hypotheses that (1) dormant seeds are capable of mounting complex responses to pathogens, (2) a diversity of defense enzymes are involved in responses in multiple plant species, and (3) it is possible to identify fungi capable of selective decay of weed seeds without damaging crop seeds, a concept that may be applicable to weed management in the field. While earlier work on seed defenses demonstrated the presence of passive defenses, this work shows that dormant seeds are also quite responsive and capable of activating and releasing defense enzymes in response to a pathogen.

SYBR(®) Green-based real-time quantitative reverse-transcription PCR for detection and discrimination of grapevine viruses.

A SYBR(®) Green-based real-time quantitative reverse transcription PCR (qRT-PCR) assay in combination with melt-curve analysis (MCA) was optimized for the detection of nine grapevine viruses. The detection limits for simplex qRT-PCR for all nine grapevine viruses were estimated to be in the range of 214-1112 copies of the virus genome. Amplicons with melting temperatures (Tm) separated by at least 2°C in the MCA could differentiate two viruses in the same reaction. Therefore, eight of the nine viruses could be co-diagnosed in five different combinations of duplex assays. Of 305 grape leaf samples from the field or greenhouse, 162 were positive for at least one of the nine grapevine viruses using the duplex qRT-PCR assays. In contrast, only 127 samples were positive using endpoint RT-PCR and PCR assays, indicating the enhanced sensitivity of duplex real-time PCR. In addition, the duplex qRT-PCR assays were be used to detect Grapevine leafroll associated virus 3 (GLRaV-3) in its vector, the grape mealybug (Pseudococcus maritimus Ehrhorn), and Grapevine red blotch-associated virus (GRBaV) in Virginia creeper leafhopper (Erythroneura ziczac Walsh). The simplex and duplex real-time PCR assays developed in this study can be used to examine transmission of co-occruing viruses by insect vectors as well as for rapid and sensitive detection of viruses in infected grapevines.

Rapid Quantitative Assessment of Rhizoctonia Resistance in Roots of Selected Wheat and Barley Genotypes.

Rhizoctonia solani AG8, causal agent of Rhizoctonia root rot and bare patch in dryland cereal production systems of the Pacific Northwest United States and Australia, reduces yields in a wide range of crops. Disease is not consistently controlled by available management practices, so genetic resistance would be a desirable resource for growers. In this report, we describe three rapid and low-cost assays for R. solani AG8 resistance in wheat and barley, with the view of facilitating screens for genetic resistance in these hosts. The first assay uses 50-ml conical centrifuge tubes containing soil infested with R. solani AG8 on a substrate of ground oats. The second assay uses roots of 3-day-old seedlings directly coated with infested ground oats, followed by incubation in plastic dishes. The third assay, suitable for barley, uses whole infested oat kernels in 50-ml tubes. Symptoms are quantified on the bases of root fresh weight and total root length at 7 and 3 days for the tube and coating assays, respectively. Each of the assays show the same disease differential between susceptible and partially resistant wheat genotypes. The assays can be conducted in the laboratory, growth chamber, or greenhouse.

Genome Sequences of Three Phytopathogenic Species of the Magnaporthaceae Family of Fungi.

Magnaporthaceae is a family of ascomycetes that includes three fungi of great economic importance: Magnaporthe oryzae, Gaeumannomyces graminis var. tritici, and Magnaporthe poae. These three fungi cause widespread disease and loss in cereal and grass crops, including rice blast disease (M. oryzae), take-all disease in wheat and other grasses (G. graminis), and summer patch disease in turf grasses (M. poae). Here, we present the finished genome sequence for M. oryzae and draft sequences for M. poae and G. graminis var. tritici. We used multiple technologies to sequence and annotate the genomes of M. oryzae, M. poae, and G. graminis var. tritici. The M. oryzae genome is now finished to seven chromosomes whereas M. poae and G. graminis var. tritici are sequenced to 40.0× and 25.0× coverage respectively. Gene models were developed by the use of multiple computational techniques and further supported by RNAseq data. In addition, we performed preliminary analysis of genome architecture and repetitive element DNA.

Use of pressure cycling technology for cell lysis and recovery of bacterial and fungal communities from soil.

Selection of cell lysis methodology is critical to microbial community analyses due to the inability of any single extraction technology to recover the absolute genetic structure from environmental samples. Numerous methodologies are currently applied to interrogate soil communities, each with its own inherent bias. Here we compared the efficacy and bias of three physical cell lysis methods in conjunction with the PowerLyzer PowerSoil DNA Isolation Kit (MO BIO) for direct DNA extraction from soil: bead-beating, vortex disruption, and hydrostatic pressure cycling technology (PCT). PCT lysis, which is relatively new to soil DNA extraction, was optimized for soils of two different textures prior to comparison with traditional bead-beating and vortex disruption lysis. All cell lysis methods successfully recovered DNA. Although the two traditional mechanical lysis methods yielded greater genomic, bacterial, and fungal DNA per gram soil than the PCT method, the latter resulted in a greater number of unique terminal restriction fragments by terminal RFLP (T-RFLP) analysis. These findings indicate the importance of diversity and quantity measures when assessing DNA extraction bias, as soil DNA retrieved by PCT lysis represented populations not found using traditional mechanical lysis methods.

Molecular and genetic aspects of controlling the soilborne necrotrophic pathogens Rhizoctonia and Pythium.

The soilborne necrotrophic pathogens Rhizoctonia and Pythium infect a wide range of crops in the US and worldwide. These pathogens pose challenges to growers because the diseases they cause are not adequately controlled by fungicides, rotation or, for many hosts, natural genetic resistance. Although a combination of management practices are likely to be required for control of Rhizoctonia and Pythium, genetic resistance remains a key missing component. This review discusses the recent deployment of introduced genes and genome-based information for control of Rhizoctonia, with emphasis on three pathosystems: Rhizoctonia solani AG8 and wheat, R. solani AG1-IA and rice, and R. solani AG3 or AG4 and potato. Molecular mechanisms underlying disease suppression will be addressed, if appropriate. Although less is known about genes and factors suppressive to Pythium, pathogen genomics and biological control studies are providing useful leads to effectors and antifungal factors. Prospects for resistance to Rhizoctonia and Pythium spp. will continue to improve with growing knowledge of pathogenicity strategies, host defense gene action relative to the pathogen infection process, and the role of environmental factors on pathogen-host interactions.

Polyphenol oxidase as a biochemical seed defense mechanism.

Seed dormancy and resistance to decay are fundamental survival strategies, which allow a population of seeds to germinate over long periods of time. Seeds have physical, chemical, and biological defense mechanisms that protect their food reserves from decay-inducing organisms and herbivores. Here, we hypothesize that seeds also possess enzyme-based biochemical defenses, based on induction of the plant defense enzyme, polyphenol oxidase (PPO), when wild oat (Avena fatua L.) caryopses and seeds were challenged with seed-decaying Fusarium fungi. These studies suggest that dormant seeds are capable of mounting a defense response to pathogens. The pathogen-induced PPO activity from wild oat was attributed to a soluble isoform of the enzyme that appeared to result, at least in part, from proteolytic activation of a latent PPO isoform. PPO activity was also induced in wild oat hulls (lemma and palea), non-living tissues that cover and protect the caryopsis. These results are consistent with the hypothesis that seeds possess inducible enzyme-based biochemical defenses arrayed on the exterior of seeds and these defenses represent a fundamental mechanism of seed survival and longevity in the soil. Enzyme-based biochemical defenses may have broader implications since they may apply to other defense enzymes as well as to a diversity of plant species and ecosystems.

Agroecological factors correlated to soil DNA concentrations of Rhizoctonia in dryland wheat production zones of Washington state, USA.

The necrotrophic soilborne fungal pathogens Rhizoctonia solani AG8 and R. oryzae are principal causal agents of Rhizoctonia root rot and bare patch of wheat in dryland cropping systems of the Pacific Northwest. A 3-year survey of 33 parcels at 11 growers' sites and 60 trial plots at 12 Washington State University cereal variety test locations was undertaken to understand the distribution of these pathogens. Pathogen DNA concentrations in soils, quantified using real-time polymerase chain reaction, were correlated with precipitation, temperature maxima and minima, and soil texture factors in a pathogen-specific manner. Specifically, R. solani AG8 DNA concentration was negatively correlated with precipitation and not correlated with temperature minima, whereas R. oryzae concentration was correlated with temperature minima but not with precipitation. However, both pathogens were more abundant in soils with higher sand and lower clay content. Principal component analysis also indicated that unique groups of meteorological and soil factors were associated with each pathogen. Furthermore, tillage did not affect R. oryzae but affected R. solani AG8 at P = 0.06. Lower soil concentrations of R. solani AG8 but not R. oryzae occurred when the previously planted crop was a broadleaf (P < 0.05). Our findings showed that R. solani AG8 concentrations were consistent with the general distribution of bare patch symptoms, based on field observations and surveys of other pathogens, but was present at many sites in which bare patch symptoms were not evident. Management of Rhizoctonia root rot and bare patch should account for the likelihood that each pathogen is affected by a unique group of agroecological variables.