Crop-Friendly Bacteria Tapped To Battle Fungal Marauders

Soil-dwelling bacteria that depend on wheat and barley roots for their “room and board” could soon make good on their debt. Researchers are investigating the microbes’ potential to biologically control root-rot fungi that cause crop yield losses of 10-30 percent annually in the U.S. Pacific Northwest and other parts of the world. The bacteria are members of the genus Pseudomonas and include 11 strains that stymie the growth of Pythium and Rhizoctonia fungi, which are responsible for dampingoff and root-rot diseases of wheat and barley. The pathogens thrive in cool, moist soils and can reach especially high levels in crop fields where conservation tillage is practiced to save on fuel costs, avoid soil erosion, and attain other ecological and environmental benefits. “They’re most problematic to seedlings of spring crops that are 4 to 6 weeks old,” notes Pat Okubara, a geneticist in the Agricultural Research Service’s Root Diseases and Biological Control Research Unit in Pullman, Washington. “Fungicides are not very effective, and there are no resistant wheat or barley varieties yet,” she adds. Rotating wheat with nonhost crops is difficult too, because of the fungi’s extensive plant-host range. Over the past year, Okubara and university colleagues have evaluated the biocontrol potential of 26 Pseudomonas strains. From those, they chose 11 for further study based on 3 important characteristics: rapid colonization of and persistence on roots, high antifungal activity, and reduction of plant disease symptoms

Detection and Quantification of Pratylenchus thornei in DNA Extracted from Soil Using Real-Time PCR

The root-lesion nematode Pratylenchus (hornet is one of the most important pests restricting productivity of wheat in the Pacific Northwest (PNW). It is laborious and difficult to use microscopy to count and identify the nematodes in soils. A SYBR Green I-based real-time polymerase chain reaction (PCR) assay was developed to detect and quantify this species from DNA extracts of soil. A primer set, designed from the internal transcribed spacer region (ITSI) of rDNA, was highly specific to P. thornei and did not amplify DNA from 27 isolates of other Pratylenchus spp., other nematodes, and six fungal species present in PNW wheat fields. A standard curve relating threshold cycle and log values of nematode number was generated from artificially infested soils. The standard curve was supported by a high correlation between the numbers of P thornei added to soil and the numbers quantified using real-time PCR. Examination of 15 PNW dryland field soils and 20 greenhouse samples revealed significant positive correlations between the numbers determined by real-time PCR and by the Whitehead tray and microscopic method. Real-time PCR is a rapid, sensitive alternative to time-consuming nematode extractions, microscopic identification, and counting of P thornei from field and greenhouse soils.Keywords: root disease, Pacific Northwest, Potato cyst nematode, Root lesion nematodes, Quantitative PCR, Meloidogyne incognita, in silico analysis, Genus pratylenchus, Polymerase chain reaction, Fragment length polymorphism, Wheat, IdentificationKeywords: root disease, Pacific Northwest, Potato cyst nematode, Root lesion nematodes, Quantitative PCR, Meloidogyne incognita, in silico analysis, Genus pratylenchus, Polymerase chain reaction, Fragment length polymorphism, Wheat, Identificatio

Species-Specific PCR Assays for Differentiating Heterodera filipjevi and H. avenae

Heterodera avenae and H. filipjevi are economically important cyst nematodes that restrict production of cereal crops in the Pacific Northwest United States and elsewhere in the world. Identification of these two species is critical for recommending and implementing effective management practices. Primers were designed from the internal transcribed spacer (ITS) regions of H. avenae and H. filipjevi ribosomal DNA. The primers were highly specific when examined on target isolates but did not amplify DNA from nontarget Heterodera, Globodera, Meloidogyne, Pratylenchus, and other nematode species tested. Polymerase chain reaction (PCR) and amplification conditions were established, and H. avenae and H. filipjevi were clearly distinguished by PCR fragments of 242 and 170 bp, respectively. Robust PCR amplification was achieved with DNA extracted from a single egg or second-stage juvenile (J2) using a laboratory-made worm lysis buffer, and DNA from 0.5 egg or J2 using a commercial kit. The PCR assays were successfully employed for differentiation of H. filipjevi and H. avenae populations collected from eight locations in three Pacific Northwest states. This is the first report of a species-specific ITS PCR assay to detect and identify H. filipjevi. The assays for both species will enhance diagnosis of cereal cyst nematode species in infested fields

The promoter from SlREO, a highly-expressed, root-specific Solanum lycopersicum gene, directs expression to cortex of mature roots

Root-specific promoters are valuable tools for targeting transgene expression, but many of those already described have limitations to their general applicability. We present the expression characteristics of SlREO, a novel gene isolated from tomato (Solanum lycopersicum L.). This gene was highly expressed in roots but had a very low level of expression in aerial plant organs. A 2.4-kb region representing the SlREO promoter sequence was cloned upstream of the uidA GUS reporter gene and shown to direct expression in the root cortex. In mature, glasshouse-grown plants this strict root specificity was maintained. Furthermore, promoter activity was unaffected by dehydration or wounding stress but was somewhat suppressed by exposure to NaCl, salicylic acid and jasmonic acid. The predicted protein sequence of SlREO contains a domain found in enzymes of the 2-oxoglutarate and Fe(II)-dependent dioxygenase superfamily. The novel SlREO promoter has properties ideal for applications requiring strong and specific gene expression in the bulk of tomato root tissue growing in soil, and is also likely to be useful in other Solanaceous crop

Developing a Real-Time PCR Assay for Detection and Quantification of Pratylenchus neglectus in Soil

Pratylenchus neglectus is one of the most widespread and economically important nematodes that invades plant roots and restricts wheat productivity in the Pacific Northwest. It is challenging to quantify P neglectus using microscopic methods for studies that require large-scale sampling, such as assessment of rotation crops, wheat cultivars, and other management practices. A real-time quantitative polymerase chain reaction (qPCR) assay was developed to detect and quantify P. neglectus from DNA extracts of soil. The primers, designed from the internal transcribed spacer region of rDNA, showed high specificity with a single melt curve peak to DNA from eight isolates of P. neglectus but did not amplify DNA from 28 isolates of other plant-parasitic and non-plant-parasitic nematodes: A standard curve (R-2 = 0.96; P < 0.001) was generated by amplifying DNA extracted from soil to which nematodes were added. The soil standard curve was validated using sterilized soil inoculated with lower numbers of P. neglectus. A significant positive relationship (R-2 = 0.66; P < 0.001) was observed for nematode numbers quantified from 15 field soils using qPCR and the Whitehead tray and microscopic method but the qPCR generally tended to provide higher estimates. Real-time PCR potentially provides a useful platform for efficient detection and quantification of P. neglectus directly from field soils.Keywords: Verticillium dahliae, Fragment length polymorphism, Root lesion nematodes, Potato cyst nematode, Northwest United States, Pacific Northwest, Polymerase chain reaction, Genus pratylenchus, Quantitative detection, DN

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.]

Antibody-mediated Prevention of Fusarium Mycotoxins in the Field

Fusarium mycotoxins directly accumulated in grains during the infection of wheat and other cereal crops by Fusarium head blight (FHB) pathogens are detrimental to humans and domesticated animals. Prevention of the mycotoxins via the development of FHB-resistant varieties has been a challenge due to the scarcity of natural resistance against FHB pathogens. Various antibodies specific to Fusarium fungi and mycotoxins are widely used in immunoassays and antibody-mediated resistance in planta against Fusarium pathogens has been demonstrated. Antibodies fused to antifungal proteins have been shown to confer a very significantly enhanced Fusarium resistance in transgenic plants. Thus, antibody fusions hold great promise as an effective tool for the prevention of mycotoxin contaminations in cereal grains. This review highlights the utilization of protective antibodies derived from phage display to increase endogenous resistance of wheat to FHB pathogens and consequently to reduce mycotoxins in field. The role played by Fusarium-specific antibody in the resistance is also discussed