Plant-parasitic Nematodes Associated with Grasses Grown for Seed in the Willamette Valley of Oregon.

Plant-parasitic nematodes (PPN) are an understudied pathogen group in the Oregon cool-season grass seed cropping system. In this survey, the PPN associated with annual ryegrass, bentgrass, fine fescue, orchardgrass, perennial ryegrass, and tall fescue were determined. Thirty-seven fields were sampled in the 2022 or 2023 growing season by collecting 10 soil cores in each of six 100-m transects for nematode extraction and visual identification. PerMANOVA testing indicated significant differences in PPN community composition across grass host and sampling time. Pratylenchus and Meloidogyne were the most commonly encountered nematodes, with maximum population densities of 1,984 and 2,496 nematodes/100 g soil, respectively. Sequencing of the COX1 gene region indicated the presence of P. crenatus, P. fallax, P. neglectus, P. penetrans, and P. thornei, with some of these species being detected for the first time on these grass hosts. The only Meloidogyne sp. found in these grasses was M. nassi, based upon sequencing of the ITS gene region. This first-of-its-kind survey indicates the need for further assessment of the impact of these PPNs on yield and stand longevity in cool-season grass seed fields in Oregon.

Floral Colonization Dynamics and Specificity of Aureobasidium pullulans Strains Used to Suppress Fire Blight of Pome Fruit.

Aureobasidium pullulans is used as a biocontrol agent for fire blight protection in organic apple and pear production. We assessed colonization of pome flowers by A. pullulans in orchards located near Corvallis, OR and Wenatchee, WA. Blossom Protect, a mix of A. pullulans strains CF10 and CF40, and its citrate-based companion, Buffer Protect, were sprayed at 70% bloom. Later in bloom, the population size of putative A. pullulans on flowers was estimated by dilution plating; plate scrapings of putative A. pullulans were then sampled and subjected to a PCR analysis. Sequenced PCR amplicons of the internal transcribed spacer region and the elongase gene confirmed the presence of A. pullulans, whereas a multiplex PCR with primers specific to CF10 and CF40 was used to determine the presence of the introduced strains. At Corvallis, a wet spring environment, A. pullulans, was recovered from most (>90%) Bartlett pear and Golden Delicious apple flowers sampled from experimental trees, regardless of whether the trees were treated with Blossom Protect. Nevertheless, population size estimates of A. pullulans on the flowers were correlated with the number of times Blossom Protect was sprayed on the trees. At Wenatchee, an arid spring environment, A. pullulans was detected on most flowers from trees treated with Blossom Protect, but only on a minority of flowers from nontreated controls. In both locations, the combined incidence of strains CF10 and CF40 on flowers averaged 89% on Blossom Protect-treated trees, but only 27% on adjacent, nontreated trees. During subsequent trials, the efficacy of Blossom Protect for fire blight control was compared with alternative yeast isolates, with each applied with Buffer Protect; local isolates of A. pullulans and Cryptococcus neoformans and a postharvest biocontrol strain of Cystofilobasidium infirmominiatum were used All yeast strains suppressed fire blight to a degree; however, in each of four trials, the level of suppression was highest with Blossom Protect, and it was significantly superior (P ≤ 0.05) to other yeast isolates in two of the trials. Because A. pullulans strains CF10 and CF40 were detected primarily on flowers on trees treated with Blossom Protect, and because they were detected much less frequently on nearby nontreated tress, we recommend treating every tree row with Blossom Protect at least once for organic fire blight suppression.

First Report of Fusarium solani on Utah Sweetvetch in the United States.

Utah sweetvetch (Hedysarum boreale Nutt.) is a native American perennial nitrogen fixing legume used mainly in rangeland reclamation, soil rejuvenation, and erosion control. In June 2011, a field of Utah sweetvetch grown for seeds in central Oregon had approximately 15% of the plants exhibiting chlorosis, defoliation, stunting, wilting, and/or death. Dissection of the crown of symptomatic plants revealed discolored pinkish brown vascular tissue. Symptomatic tissues from six random plants were surface sterilized, placed on acidified potato dextrose agar (PDA) medium, and cultured for 7 days at room temperature, which allowed six fungal isolates (SS1 through SS6) to be collected. On PDA, all six isolates had rapid, creamy white colored growth. Based on observations of 1-week-old isolates, microconidia were oval to kidney shaped, single celled, 8 to 10 × 2.5 to 4 µm, and formed at the tips of long unbranched monophialides. Macroconidia were three to four septate, cylindrical to slightly curved, with characteristic foot shaped basal cell and blunt apical cell, 37 to 49 × 4.4 to 5.3 µm. Chlaymydospores observed were 8.5 to 11 × 7.6 to 9 µm. Based on fungal references (1,2,3), the isolates were identified as Fusarium solani (Mart.) Sacc. Identification of the isolates at the molecular level was determined by amplification of the internal transcribed spacer (ITS) region using PCR and amplicon sequencing. Botrytis cinerea and F. graminearum cultures were used as controls for the extraction, amplification, and sequencing steps. Genomic DNA was extracted from mycelia using protocols of the MOBIO Ultraclean Soil DNA Isolation Kit (MO-BIO Laboratories Inc, Carlsbad, CA, USA). PCR was performed using ITS1/ITS4 primers and resulted in 563- to 573-bp amplicons, which were sequenced. Analysis of the ITS sequences (GenBank Accession Nos. JX524018 to JX524023) for the six fungal isolates using BLASTn revealed a 99% sequence identity with F. solani strains (AB470903, AB513851, AJ608989, EF152426, EU029589, and HM214456). Pathogenicity was confirmed on Utah sweetvetch plants in the greenhouse. Seeds of Utah sweetvetch were first plated on acidified PDA for germination; healthy seedlings were then selected and transplanted into pots with sterilized soil after 2 weeks of growth. The plants were kept in a greenhouse at Central Oregon Agricultural Research Center, Madras, Oregon. Ten 40-day-old healthy vetch plants were inoculated by drenching with a mixed conidial suspension (107 conidia/ml) of the six F. solani isolates. Ten plants drenched with sterile distilled water were included as controls. Symptoms of chlorosis and stunting similar to those in the commercial field were observed within 30 days of inoculation on 8 of 10 inoculated plants, while control plants were symptomless. Fungal isolates identical to F. solani were reisolated from the symptomatic plants. To our knowledge, this is the first report of F. solani on Utah sweetvetch plants. References: (1) C. Booth. The Genus Fusarium. CMI, Kew, Surrey, UK, 1971. (2) P. E. Nelson et al. Fusarium species: An illustrated manual for identification. The Pennsylvania State University Press, USA, 1983. (3) H. I. Nirenberg. A simplified method for identifying Fusarium spp. occurring on wheat. Can. J. Bot. 59:1599, 1980.

Implications of pathogenesis by Erwinia amylovora on rosaceous stigmas to biological control of fire blight.

As a prerequisite to infection of flowers, Erwinia amylovora grows epiphytically on stigmas, which provide a conducive habitat for bacterial growth. Stigmas also support growth of several other bacterial genera, which allows for biological control of fire blight; although, in practice, it is very difficult to exclude E. amylovora completely from this habitat. We investigated the dynamics of growth suppression of E. amylovora by comparing the ability of virulent and avirulent strains of E. amylovora to compete with each other on stigmas of pear, apple, and blackberry, and to compete with a co-inoculated mixture of effective bacterial antagonists. When strains were inoculated individually, virulent E. amylovora strain Ea153N attained the highest population size on stigmas, with population sizes that were approximately double those of an avirulent hrpL mutant of Ea153 or the bacterial antagonists. In competition experiments, growth of the avirulent derivative was suppressed by the antagonist mixture to a greater extent than the virulent strain. Unexpectedly, the virulent strain enhanced the population size of the antagonist mixture. Similarly, a small dose of virulent Ea153N added to inoculum of an avirulent hrpL mutant of Ea153 significantly increased the population size of the avirulent strain. A pathogenesis-gene reporter strain, Ea153 dspE::gfp, was applied to flowers and a subset of the population expressed the green fluorescent protein while growing epiphytically on stigmas of apple. These results are consistent with the hypothesis that virulent E. amylovora modifies the epiphytic habitat presented by the stigma through a pathogenesis-related process, which increases host resources available to itself and, coincidentally, to nonpathogenic competitors. Over nine orchard trials, avirulent Ea153 hrpL significantly suppressed the incidence of fire blight four times compared with six for the antagonist mixture. The degree of biological control achievable with an avirulent strain of E. amylovora likely is limited by its inability to utilize the stigmatic habitat to the same degree as a virulent strain.

Rates of Epiphytic Growth of Erwinia amylovora on Flowers Common in the Landscape.

We evaluated epiphytic growth of the fire blight bacterium, Erwinia amylovora, on flowers of plant species common to landscapes where pears and apples are grown. The plants were from genera regarded as important nectar and pollen sources for pollinating insects: Acer, Amelanchier, Brassica, Cytisus, Populus, Prunus, Rubus, Salix, Taraxacum, Trifolium, and Symphoricarpos. Floral bouquets were inoculated with E. amylovora and incubated in growth chambers at 15°C for 96 h. Regardless of their susceptibility to fire blight, all species from the rose family except Prunus domestica (European plum) supported epiphytic populations of E. amylovora that exceeded 1 × 106 CFU/flower with relative growth rates for the populations that averaged 7% per hour. Nonrosaceous plants were generally poor supporters of epiphytic growth of the fire blight pathogen with relative growth rates averaging <4% per hour. In two seasons of field inoculations, the rosaceous non-disease-host plants, Prunus avium (sweet cherry) and Rubus armeniacus (Himalayan blackberry), yielded mean population sizes of E. amylovora that exceeded 1 × 106 CFU/flower; in contrast, at 8 days after inoculation, mean population sizes of the pathogen were in the range of 5 × 103 to 5 × 104 CFU/flower on Cytisus scoparius (Scotch broom) and <1 × 102 CFU on Acer macrophylum (big leaf maple). Because vectors of E. amylovora, principally bees, visit many kinds of flowers in landscape areas between pear and apple orchards, flowers of rosaceous, non-disease-host species could serve as potential sites of inoculum increase during their periods of bloom.