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2010 Potomac Division Meeting Abstracts

Host range determination of Colletotrichum gloeosporioides f. sp. salsolae, a biological control agent of tumbleweed: From BLUPs to biomass loss
D. K. Berner (1), C. A. CAVIN (1)
(1) USDA, ARS, FDWSRU
Phytopathology 100:S205

Host range tests were conducted with Colletotrichum gloeosporioides f. sp. salsolae (CGS) in quarantine to determine whether the fungus is safe to release in N. America for biological control of tumbleweed (Salsola tragus L., Chenopodiaceae). Ninety two accessions were analyzed from 19 families and 10 tribes within the family Chenopodiaceae. These included 62 genera and 120 species. Disease reaction data were combined with a relationship matrix derived from internal transcribed spacer DNA sequences and analyzed with mixed model equations to produce Best Linear Unbiased Predictors (BLUPs) for each species. Twenty nine species from 7 closely-related Chenopodiaceae tribes had significant levels of disease severity as indicated by BLUPs. Most species in the genus Salsola, which are all introduced and weedy, were very susceptible and damaged by CGS. Of the 29 susceptible species, 10 native or commercially important species in N. America were identified as needing additional tests to determine the extent of any damage caused by disease. These additional tests were done by inoculating the non-target species of concern with CGS and weighing oven-dried shoots and roots of non-inoculated and inoculated plants one month after inoculation. The shoots and roots of each plant were scanned and the surface areas determined with image analysis software. The damage to the shoots and roots of each plant were standardized by dividing surface area by the corresponding dry weights to arrive at area per unit weight. Average differences in standardized plant damage between inoculated and controls for each plant species were combined with corresponding disease ratings and analyzed by principal component analysis. Results showed that most of the non-target species clustered as not-damaged while the target and several related weedy species were heavily damaged. Three non-target species were moderately damaged, but these species were either perennial or not ecologically sympatric with tumbleweed.

Extract of the brown seaweed Asophyllum nodosum and silicon reduce plant death due to Fusarium spp. of cucurbits
G. E. Brust (3), R. E. ROSS (1), J. Jayaraj (2)
(1) Acadian Seaplants LLC, Dartmouth, NS, Canada; (2) Department of Life Sciences, The University of the West Indies, St. Augustine, Trinidad; (3) University of Maryland, Upper Marlboro, MD, USA
Phytopathology 100:S205

Crop losses due to Fusarium spp. are important to cucurbit growers along with an increasing interest in natural ways to improve disease resistance. Extracts of the brown seaweed, Ascophyllum nodosum, and products containing silicon have both been shown to promote disease resistance in many crops. In a 2008 watermelon trial located in Upper Marlboro, MD, Fusarium solani symptoms were suppressed by extracts of A. nodosum. At the final rating, 30% of the watermelon plants were dead from this pathogen in the control plots vs. 10% in A. nodosum extract treatments. A second study was implemented in 2009 on Gladiator Pumpkins. Calcium silicate and A. nodosum extract were applied to pumpkins grown in a field known to have Fusarium spp. infected squash three years prior. At the final rating, 24.6% of the pumpkin plants were dead in the control plots vs. 19.2% in the silicon plots, 13.6% in the A. nodosum extract treatment, and just 6.1% in the plots with both calcium silicate and A. nodosum extract. These field trials were further supported by two greenhouse studies where applications A. nodosum extract applied to cucumber plants reduced incidence of Fusarium oxysporum and enhanced the activities of plant defense-related enzymes including chitinase, beta-1,3-glucanase, peroxidase, polyphenol oxidase, phenylalanine ammonia lyase and lipoxigenase as well as elevated levels of total phenols compared to the control. The jasmonic acid pathway has been found to be very important in plant defense responses elicited by A. nodosum. Pathogens that are inhibited by the jasmonic acid pathway are often necrotrophs such as Fusarium spp. A. nodosum extract may offer a valuable tool to improve the health and productivity of cucurbits.

Detection and distribution of Bean pod mottle virus in soybean and beetle vectors in eastern Virginia
M. E. CASSELL (1), T. P. Kuhar (2), P. B. Schultz (3), S. A. Tolin (1)
(1) Virginia Tech, Blacksburg, VA; (2) Virginia Tech-ESAREC, Painter, VA; (3) Virginia Tech-HRAREC, Virginia Beach, VA
Phytopathology 100:S205

Bean pod mottle virus (BPMV) (genus Comovirus; family Comoviridae) is a reemerging disease of soybeans. Vectored by the Bean leaf beetle (Cerotoma trifurcata), this virus has become prevalent on the Eastern Shore of Virginia. In tissue blot immunoassays (TBIA) of soybean sentinel plots for the Legume IPM-PIPE in 2007, BPMV was detected at a high incidence at the Eastern Shore station, but not at the Tidewater station. In 2008, beetles collected at the Eastern Shore station were 80% positive for BPMV by TBIA, but all non-soybean legumes tested were TBIA-negative. In a systemic survey in 2009, BPMV was detected by TBIA in 16 of 42 soybean fields from the southern tip of the Eastern Shore to southern Maryland. Up to 100% of the beetles collected from 24 of 38 Virginia fields were positive for BPMV by ELISA of individual beetles. Infectious virus was recovered from beetle extracts prepared for ELISA. In 2009, an outbreak of BPMV was also detected in two counties in the Northern Neck of Virginia. The primary inoculum of BPMV remains unknown. Sampling is being conducted on the Eastern Shore to locate plants that might serve as an early season source of BPMV for acquisition by overwintering or first generation beetles.

Impact of mowing and fertility practices on weed species and brown patch dynamics in rhizomatous tall fescue
M. A. CUTULLE (3), D. McCall (2), B. Horvath (1), J. Derr (3)
(1) University of Tennessee, Knoxville, TN; (2) Virginia Tech, Blacksburg, VA; (3) Virginia Tech, Virginia Beach, VA
Phytopathology 100:S205

Tall Fescue (Festuca arundinacea) is a commonly utilized turfgrass in the temperate and transition zone areas of the United States. It establishes quickly, requires moderate amounts of nitrogen, and is resistant to most diseases. However, during hot humid summers, tall fescue is under stress and is susceptible to Rhizoctonia solani infection. The resulting disease, referred to as brown patch, causes turf thinning, leading to encroachment from weeds such as bermudagrass (Cynadon dactylon). Cultural practices such as fertility and mowing height may impact bermudagrass encroachment and brown patch disease in tall fescue. Improved brown patch control may result in lower weed infestations. Two mowing heights (5 and 10 cm), three levels of fertility (49, 171, and 220 kg of nitrogen annually per hectare), and preemerge herbicide application (ronstar or no herbicide applied in 2009 only) were evaluated in an established stand of ‘RTF’ tall fescue. Three plugs of common bermudagrass were planted in each plot in May 2008. Data collected monthly included weed composition and density, bermudagrass diameter, brown patch severity, and turf quality. The experiment was repeated in May of 2009. Mowing height had a significant effect on bermudagrass in year one and year two. A higher mowing height resulted in less bermudagrass encroachment. Fertility did not have an effect on bermudagrass diameter. In July and August, southern crabgrass (Digitaria ciliaris) density was much greater in the 5 cm mowing height plots. Tall fescue cover was significantly reduced in the 5 cm mowing treatment due to weed competition but was acceptable at the 10 cm height. Higher fertility resulted in increased brown patch severity. However, these plots recovered quickly when weather was cooler and dryer. The same trends were observed in year two, though incidence of brown patch was greater in year two due to the increased precipitation.

A new phytoplasma lineage is associated with diseased juniper (Juniperus occidentalis)
R. E. DAVIS (1), E. L. Dally (1), Y. Zhao (1), I.-M. Lee (1), R. Jomantiene (2), A. J. Detweiler (3), M. L. Putnam (4)
(1) USDA-Agricultural Research Service, Beltsville, MD, USA; (2) Nature Research Center, Vilnius LT-08406, Lithuania; (3) Oregon State University Deschutes County Extension, Redmond, OR, USA; (4) Oregon State University Plant Clinic, Corvallis, OR, USA
Phytopathology 100:S206

Phytoplasmas are wall-less, prokaryotic plant pathogens that are spread from plant-to-plant by insects and are the cause of diseases in a wide range of plant species that include angiosperms and gymnosperms. Worldwide, work is underway to determine the possible association of phytoplasmas with plant diseases of unsolved cause, in order to devise disease control and quarantine measures based on knowledge of the pathogens involved. The present work focused on a disease (juniper witches’ broom, JunWB) of Juniperus occidentalis, a native tree indigenous to parts of western USA. Amplification of ribosomal RNA gene sequences (rDNA) in polymerase chain reactions (PCRs) primed by phytoplasma-universal primers indicated that a phytoplasma was associated with the disease. Nucleotide sequences of the rDNA were analyzed using a computer-based interface, iPhyClassifier, to obtain virtual RFLP patterns of 16S rDNA; the results indicated that JunWB phytoplasma represented a new lineage in the pigeon pea witches’ broom phytoplasma group (16SrIX). The findings expand the known biodiversity of phytoplasmas infecting conifers and raise the question of whether J. occidentalis, previously undescribed as a phytoplasma host, could play a role in the spread of phytoplasmal disease potentially damaging to forest and/or landscape conifers in North America.

Effects of exogenous indole-3-acetic acid on transcriptional reprogramming of hormone signaling and metabolism genes in potato purple top phytoplasma-infected tomato plants
Y. DING (1), W. Wei (1), W. Wu (1), Y. Jiang (1), R. E. Davis (1), Y. Zhao (1)
(1) Molecular Plant Pathology Laboratory, ARS-USDA, Beltsville, MD
Phytopathology 100:S206

Phytoplasmas are plant pathogenic bacteria that lack a cell wall. Plants infected by phytoplasmas exhibit various symptoms indicative of disrupted hormonal balance. Observations that exogenous application of auxins on aster yellows phytoplasma-infected periwinkle plants could induce symptom remission or even phytoplasma elimination further point to crucial roles of plant hormones in phytoplasma pathogenesis. The present study was designed to gain an insight into expression profiles of plant hormone signaling and metabolism genes in healthy vs phytoplasma-infected plants, and to examine whether exogenously applied indole-3-acetic acid (IAA, a naturally occurring auxin) would modify the expression patterns of these genes. Columbia Basin potato purple top (PPT) phytoplasma (a member of subgroup 16SrVI-A) and its alternate host Rutgers tomato were used as a model pathogen-host pair. Our preliminary data revealed that, following graft inoculation of plants with PPT phytoplasma, expression patterns of a putative IAA biosynthesis gene and an F-box protein-encoding gene responsible for IAA signaling were altered. Exogenously applied IAA was able to reverse the course, bringing expression of the two genes to the levels comparable to those in healthy and mock-inoculated tomato plants. The findings provide a clue to understanding mechanisms of phytoplasma pathogenesis and exogenous auxin-induced phytoplasmal disease remission.

The science and art of photography for art and science
J. D. EISENBACK (1)
(1) Department of Plant Pathology, Physiology, and Weed Science, Virginia Tech, Blacksburg, VA
Phytopathology 100:S206

Several art historians have noted that paintings suddenly improved in detail and realistic representation sometime around 1420 AD. This remarkable enhancement of drawings and paintings was credited to use of a convex mirror to project a scene onto a canvas. As the technology of glass production improved, conventional lenses were made for projecting images onto a canvas inside a large dark room, called a camera obscura. In the early 1800s light sensitive paper replaced the canvas and pigments, and the photograph was born. The camera obscura was downsized to the more mobile camera, a small box with a lens and a holder for light sensitive film. Cameras and photography became incorporated into nearly every facet of human activity, including scientific documentation. The discovery of the photovoltaic effect by Albert Einstein initiated the development of digital cameras in the late 1900s. Digital photography allows images to be readily manipulated in several ways, including high dynamic range (HDR) photography, multiple focus photography, and the production of megapixel mosaic photographs that can be utilized for art and science.

Effect of aging on survival and heat tolerance of anhydrobiotic seed-gall nematode, Anguina agrostis
J. D. EISENBACK (1), C. W. Roane (1), W. Ma (1)
(1) Department of Plant Pathology, Physiology, and Weed Science, Virginia Tech, Blacksburg, VA
Phytopathology 100:S206

A seed gall nematode, Anguina agrostis, was found parasitizing redtop creeping bentgrass (Agrostis stolonifera), that over winters in an anhydrobiotic state. Infested grass seed was collected on Aug. 24, 2003 and Aug. 16, 2009 from a naturally infested site on Butt Mountain Lookout, near the fire watchtower, in Giles County, Virginia. Samples that were stored in an open plastic bag in a laboratory cabinet for more than 5 years were compared to freshly collected specimens. Seed galls were soaked in tap water for 24 hr. to evaluate the survival of juvenile nematodes. Additional seed galls were exposed to high temperatures in a glass test tube immersed in hot water for 5, 10, 15, and 30 min. at 80, 90, and 100°C each. The nematodes were freed from the gall with sharply pointed forceps after the gall was placed into water for 24 hr. All treatments were replicated 6 times. Fresh galls contained an average of 630 nematodes, of which 82% were alive; five-year-old galls had 694 nematodes, of which 72% survived. The survival of nematodes in galls that were heat-treated at 80°C for 30 min. was 90% in fresh galls and reduced to 68% in 5 year old galls. Fresh galls exposed to 90°C for 30 min. survival was 68% and only 10% in five year old galls. All of the individuals were killed in five year old galls that were treated at 100°C for 15 min.; however, there was 35% survival in fresh galls. These experiments demonstrate that Anguina agrostis has remarkable heat tolerance that gradually declines as it ages.

Host range determination of Synchytrium solstitiale: Issues as a candidate for biological control of yellow starthistle
F. M. ESKANDARI (1), W. L. Bruckart (1), T. L. Widmer (1)
(1) USDA, ARS, FDWSRU, Ft. Detrick, MD, USA
Phytopathology 100:S206

Synchytrium solstitiale, a chytrid recently evaluated for biological control of yellow starthistle (YST, Centaurea solstitialis), was found to cause infections on seedlings of commercially-important safflower (Carthamus tinctorius) and two North American natives, Centaurea americana and C. rothrockii (10%, 10%, and 25% incidence, respectively) when cotyledons were exposed to zoospores released in water. This compared to >90% incidence for inoculated YST seedlings in that same study. The object of the present study was to confirm susceptibility of safflower to S. solstitiale. Surface-sterilized leaves of YST (as susceptible control), safflower, Russian knapweed (Rhaponticum [Acroptilon] repens), and common crupina (Crupina vulgaris), were floated on sterile water in large (15 cm) glass Petri dishes containing zoospores. Five pieces of surface-sterilized galled leaf tissue were placed at four locations around the perimeter and at the center of each dish to give uniform distribution of zoospores in each test. Leaves of each test species were paired with YST leaves in each dish, thus providing uniform inoculum for each test species and YST within a dish. At least three dishes were set up for tests. Results confirm that both YST and safflower are susceptible to S. solstitiale and that Russian knapweed and common crupina are not. Data from both studies suggest differential susceptibility to S. solstitiale occurs among plants within the Asteraceae. Before proposal is made to introduce S. solstitiale for biological control of YST in the U.S., additional data will be needed concerning potential risk associated with such action, particularly relating to safflower and safflower culture in California.

Phytophthora pini Leonian, a valid and distinct species
M. GALLEGLY (2), C. Hong (1), P. Richardson (1), P. Kong (1)
(1) Virginia Tech, Virginia Beach, VA; (2) West Virginia University, Morgantown, WV
Phytopathology 100:S207

Leonian described Phytophthora pini in 1925. His one culture, isolated from roots of red pine growing in Minnesota, was essentially ignored as a species until 1956 when Waterhouse included it in her compilation of original descriptions. Later, in 1963, Waterhouse considered P. pini to be invalid because it was morphologically identical to P. citricola described by Sawada in 1927. We secured the ex-type and ex-authentic cultures of P. citricola, P. pini, and Chester’s P. cactorum var. applanata and compared them morphologically and molecularly with each other and with isolates of P. plurivora Jung and Burgess (same as Gallegly and Hong’s P. citricola II), P. citricola I and P. citricola III. The results show that P. pini is identical to P. citricola I in all molecular and morphological characters and different from P. plurivora, the ex-type P. citricola, and P. citricola III. Incidentally, the ex-type culture of P. cactorum var. applanata is identical to isolates of P. plurivora. Thus, Phytophthora pini is resurrected to distinct species status.

Characterization of Pythium species causing Pythium blight on snap beans in the eastern US
L. HARRISON (1), S. L. Rideout (1)
(1) Virginia Tech, Painter, VA, USA
Phytopathology 100:S207

The Eastern Shore of Virginia (ESV) is an important snap bean (Phaseolus vulgaris L.) growing region, but profitable yields are threatened by Pythium blight, one of the most severe snap bean diseases in the US. Although this disease is well documented, the species of Pythium causing this disease have not been well characterized. This information is important for determining management strategies. Isolates were collected to establish the causal agent(s) of Pythium blight on snap beans. Because of the pathogen’s wide host range and distribution, isolates were recovered from different hosts, including other legumes, cucurbits and solanaceous crops, and from multiple snap bean-growing areas in the ESV, Georgia, and New Jersey. Isolates were collected from soil by baiting and from plant tissue showing water-soaking and/or white, cottony growth. For each isolate, pathogenicity on snaps beans was verified, and each isolate was characterized by morphology and sequence analysis of the rDNA-internal transcribed spacer (ITS) regions. All ESV isolates were identified as Pythium aphanidermatum, except for one Pythium myriotylum and one Pythium ultimum isolate. Both P. aphanidermatum and P. ultimum were recovered from New Jersey crops. P. aphanidermatum was also isolated from symptomatic plants in Georgia; however, multiple Georgia isolates had 99-100% ITS sequence similarity with Pythium deliense Meurs accessions in GenBank. These isolates also had P. deliense-characteristic morphology, producing oospores measuring 16.5 µm diameter and similar, but less inflated, sporangia than P. aphanidermatum. Putative P. deliense isolates will be further characterized by sequence analysis of the cytochrome oxidase II gene. P. deliense has not yet been reported on common bean. This research verifies that multiple Pythium spp. are responsible for Pythium blight symptoms on snap beans.

Does a Vicia villosa cover crop induce general suppression of Fusarium wilt of watermelon?
J. HIMMELSTEIN (2), K. Everts (2), J. Maul (1)
(1) Sustainable Agriculture Systems Laboratory, USDA-ARS, Beltsville, MD, USA; (2) University of Maryland, College Park, MD, USA
Phytopathology 100:S207

Current triploid watermelon cultivars have little resistance to Fusarium wilt, caused by Fusarium oxysporum f. sp. niveum (FON) and yield losses are increasing in the eastern U.S. A Vicia villosa (hairy vetch) green manure, suppressed watermelon Fusarium wilt in previous trials, but the mechanism of this suppression is unknown. The objective of this experiment was to determine if the V. villosa cover crop suppresses Fusarium wilt via general suppression by looking at the rate of soil respiration and microbial activity in the presence of three cover crops at two locations in Maryland. Fall planted V. villosa, Trifolium incarnatum (crimson clover), and Secale cereale (rye) were grown and incorporated as green manures into the soil prior to planting the watermelon. FON was applied three days after transplanting at the Salisbury location and five days after transplanting in Beltsville. No visible wilt symptoms were observed at either location. Hairy vetch and crimson clover caused a significant increase in total fruit yield at Beltsville. The respiration data revealed that microbial activity increased significantly directly after cover crop incorporation, then dropped down to pre incorporation levels for 3 months. As the watermelon matured the respiration rate increased once again. Crimson clover and hairy vetch plots had similar respiration rates and we are testing for correlations between cover crop induced changes in soil respiration and disease suppressiveness.

Multianalyte immunohistochemical investigation of relative hormone levels in potato purple top phytoplasma-infected tomato plants
Y. JIANG (1), W. Wei (1), Y. Ding (1), W. Wu (1), R. E. Davis (1), R. W. Hammond (1), Y. Zhao (1)
(1) Molecular Plant Pathology Laboratory, ARS-USDA, Beltsville, MD
Phytopathology 100:S207

Phytoplasmas are small, cell wall-less bacteria responsible for numerous diseases in agriculturally and environmentally important plant species worldwide. Phytoplasma infections of plants induce symptoms including excessive shoot proliferation, witches’-broom growths, general stunting, rapid senescence (yellowing), and abnormal floral development (virescence and phyllody). These symptoms indicate that hormonal balance may be disrupted in affected plants. The lack of plant hormone biosynthesis genes in all completely-sequenced phytoplasma genomes implies that the presumed hormonal imbalance in phytoplasma-infected plants may be caused either by changes in endogenous hormone levels or by alterations in sensitivity to hormones. The present study was aimed at understanding the mechanism underlying phytoplasma-induced host hormonal imbalance. Tissue sections prepared from Columbia Basin potato purple top (PPT) phytoplasma-infected and healthy tomato plants were subjected to comparative immunohistochemical analyses using antibodies against auxin (IAA), cytokinins (6-BA, trans-zeatin riboside, and cis-zeatin riboside), abscisic acid (ABA), and gibberellic acid (GA3). The results revealed notable changes in levels of plant hormones in PPT-infected vs healthy plants. Findings from the study will aid understanding of the roles of plant hormones in phytoplasma pathogenesis and disease symptom expression.

Phytophthora phaseoli; destroyer of lima bean production
S. G. KUNJETI (1), N. M. Donofrio (1), A. G. Marsh (1), B. C. Meyers (1), T. A. Evans (1)
(1) University of Delaware
Phytopathology 100:S207

Phytophthora phaseoli Thaxt., an oomycete plant pathogen and close relative to Phytophthora infestans (Blair et al. 2008), causes downy mildew of lima bean (Phaseolus lunatus L.) during cool and humid weather conditions. Since its first report in 1889, the vegetable processing industry of the humid eastern US has been negatively affected by this disease. Since 1889 P. phaseoli has evolved six races A,B,C,D,E and currently prevalent race F in lima bean fields. Developing lima bean cultivars with durable resistance to this pathogen is a more environmentally friendly and cost-efficient method of disease management than pesticide application. To develop such a cultivar, it is necessary to understand the underlying mechanism of how the pathogen breaks down the plant’s defenses. To date, nothing is known about the molecular interactions that occur during this plant-pathogen interaction. Towards a better understanding of these mechanisms, we used next generation sequencing technology (Illumina) to compare global gene expression of plate-grown and plant-grown P. phaseoli. Our computational analysis of the transcripts showed that most of the effector genes that were over-expressed in P. infestans while infecting potato leaf tissue were also over-expressed in P. phaseoli while infecting lima bean hypocotyls. Some of the well-characterized effectors like INF1, and INF4 were confirmed by performing RT-PCR using plate-grown, plant-grown mycelium and lima bean pods infected with P. phaseoli as template. Effector genes that were expressed in P. phaseoli when infecting lima bean pods were consistent with the genes expressed when infecting lima bean hypocotyls. Our results suggest that like in P. infestans infection of potato, P. phaseoli requires the same effector genes for the infection of lima bean pods and hypocotyls.

Comparison of the detection of Xanthomonas campestris pv. campestris in Brassica seeds using agar plating and real-time PCR
A. MARQUES (2), S. Abramova (1), A. Ignatov (1), A. Sechler (3), N. Schaad (3)
(1) All-Russian Institute of Phytopathology; (2) Embrapa Recursos Genéticos e Biotecnologia; (3) FDWSRU, USDA-ARS
Phytopathology 100:S207

Black rot of Crucifers, caused by Xanthomonas campestris pv. campestris (Xcc), is a serious seedborne disease worldwide. Although several assays are available for detection of Xcc from seed, seeds remain the major source of inoculum. We compared the recovery of Xcc on three media, mFS, NSCAA, and mCS20ABN from spiked and naturally infested seed lots using the industry standard seed testing protocol. The highest recovery of Xcc was found to be equal in spiked seed extracts on mFS and NSCAA. Recovery of Xcc from naturally infested seed was found to be higher on mFS (100 cells/ml) than NSCAA (30 cells/ml). Media mCS20ABN (Chang et al., 1991) was found to be frequently overgrown by non-target bacteria making identification difficult. For PCR we adapted the hrpF PCR primers of Berg et al. (2005) for RT-PCR by designing a probe sequence between primers DLH120 and DLH125. The resulting primer and probe set was optimized, tested, and used for the confirmation of suspect Xcc colonies from agar media, and detection of Xcc from the seed soak. This adapted RT-PCR primer and probe set was found to react with Xcc and other Xanthomonas crucifer pathogens, including X. raphani, X. armoraciae, X. barbarae, and X. aberrans. The sensitivity of the primer and probe set was 500 cells per milliliter. Direct RT-PCR of the spiked seed soak was not reliable; detection was possible only from DNA extractions of the spiked seed soak.

Use of isoparaffin-based oil for controlling dollar spot and gray leaf spot in turfgrasses
D. S. MCCALL (1), A. E. Nichols (1), M. A. Cutulle (1)
(1) Virginia Tech, Blacksburg, VA, USA
Phytopathology 100:S208

Civitas fungicide is an isoparaffin-based oil, reported to trigger Induced Systemic Resistance (ISR) within plants. Little is known about how isoparaffin-based oils control turf diseases. Field trials were conducted on perennial ryegrass (Lolium perenne) and creeping bentgrass (Agrostis stolonifera) for control of gray leaf spot and dollar spot, respectively. Civitas (45.2 liters mineral oil per ha) plus Civitas Harmonizer (2.86 liters proprietary pigment dispersion per ha) was applied alone and in mixture with common fungicides for controlling each disease. Disease was assessed by visually estimating percentage of plots showing symptoms. The isoparaffin oil plus pigment has consistently reduced each disease tested when compared with untreated checks. Tank mixture of the oil plus pigment combination with reduced rates of thiophanate methyl (3336 Plus) controlled gray leaf spot as well as full rates of thiophanate-methyl. These results indicate that this isoparaffin oil plus pigment product suppressed disease during mild outbreaks of dollar spot and gray leaf spot, and can be mixed with reduced rates of several common fungicides during more severe outbreaks.

Clusters of defense-related genes in the genome of Arabidopsis thaliana
O. A. POSTNIKOVA (2), L. G. Nemchinov (2), A. M. Boutanaev (1)
(1) Institute of Basic Biological Problems, Russian Academy of Sciences, Pushchino, Moscow Region, Russia; (2) USDA/ARS, Plant Sciences Institute, Beltsville, MD, USA
Phytopathology 100:S208

Functional and physical clustering of unrelated genes known as operons is a characteristic of prokaryotic genomes. A concept and consequences of gene clusters in eukaryotic genomes are largely unexplored. In this work, we performed computer-generated analysis of the chromosomal distribution of genes associated with defense response in Arabidopsis thaliana. This analysis revealed numerous clustered genes whose co-regulation may be related to the defense responses. The genes were distributed among all chromosomes of A. thaliana. To support computer data, we arbitrarily selected two clusters and analyzed expression levels of their gene-members in Arabidopsis ecotypes Col-0 and C24 during infection with yellow strain of Cucumber mosaic virus (CMV(Y). Ecotype Col-0 is susceptible to CMV(Y), whereas C24 contains a dominant resistance gene RCY1. Our data showed that genes compiling two clusters were activated only in resistant ecotype C24. This indicated that co-regulation of neighboring, defense-related genes in the genome of Arabidopsis is strongly affected not only by their chromosomal location, but also by the basic mechanisms of genetic resistance to pathogens.

Isolates of Fusarium graminearum collected 40 to 300 meters above ground level cause Fusarium head blight in wheat and produce trichothecene mycotoxins
D. G. SCHMALE (1), T. Fetters (1), S. Ross (2), P. Tallapragada (2), B. Dingus (1)
(1) Department of Plant Pathology, Physiology, and Weed Science, Virginia Tech, Blacksburg, VA; (2) Department of Engineering Science and Mechanics, Virginia Tech, Blacksburg, VA
Phytopathology 100:S208

The genus Fusarium contains important plant and animal pathogens, some of which produce dangerous secondary metabolites (mycotoxins). Many fusaria use the atmosphere to travel from one habitat to another, yet their atmospheric transport is poorly understood. We used autonomous (self-controlling) unmanned aerial vehicles (UAVs) to collect fusaria tens to hundreds of meters above ground level (AGL) at Virginia Tech’s Kentland Farm in Blacksburg, VA. Eleven single-spored isolates of Fusarium graminearum collected with UAVs 40 to 300 meters AGL during fall, winter, spring, and summer months were able to cause Fusarium head blight on a susceptible cultivar of spring wheat and produce trichothecene mycotoxins. Nine of these isolates produced the mycotoxins deoxynivalenol (DON)/15-acetyl-DON, one isolate produced DON/3-acteyl-ADON, and one isolate produced nivalenol (NIV). To our knowledge, this is the first report of a NIV-producing isolate of F. graminearum in Virginia, and isolates producing DON/3-acetyl-DON are rare in populations of the fungus recovered from infected wheat plants in the eastern U.S. A new framework for understanding punctuated changes in the population structure of atmospheric fusaria based on the concepts of atmospheric transport barriers (ATBs) and Langrangian coherent structures (LCS) is being developed and tested at both local and regional scales. This work aims to transform our knowledge of the atmospheric transport of microorganisms and develop new paradigms that link field and atmospheric populations of toxigenic fusaria.

Detection of Phytophthora ramorum chlamydospores in soil by baiting and dilution plating
P. W. TOOLEY (1), M. M. Carras (1)
(1) USDA-ARS, Ft. Detrick, MD
Phytopathology 100:S208

Chlamydospores of P. ramorum produced by mixing 20 percent V8 juice broth cultures with sand and incubating over a 1 month period were used to infest field soil at densities ranging from 0.2 to 42 chlamydospores/cc soil. Chlamydospore recovery was determined by baiting with rhododendron leaf discs and dilution plating both when soil infestation was performed (time 0) and following 30 days storage at 4°C, as recommended in the soil and growing medium sampling protocol on the APHIS website (http://www.aphis.usda.gov/plant_health/plant_pest_info/pram/protocols.shtml). Baiting was slightly more sensitive than dilution plating at time 0, allowing detection of P. ramorum down to 0.2 chlamydospores/cc soil compared with 1 chlamydospore/cc for dilution plating. Following 30 days of infested soil storage at 4°C, P. ramorum was detected using both methods at significantly (P = 0.05) higher levels than at time 0. The results indicate that storage of P. ramorum-infested soil at 4°C for 30 days can enhance recovery of the pathogen.

Increasing atmospheric carbon dioxide amplifies Alternaria alternata sporulation and antigen production, but does not impact sporulation of Cladosporium phlei
J. WOLF (3), N. R. O’Neill (2), C. A. Rogers (4), M. M. Muilenberg (4), L. H. Ziska (1)
(1) USDA-ARS, Crop Systems and Global Change Lab, Beltsville, MD, USA; (2) USDA-ARS, Systematic Mycology and Microbiology Lab, Beltsville, MD, USA; (3) University of Maryland, College Park, MD, USA; (4) University of Massachusetts School of Public Health, Amherst, MA, USA
Phytopathology 100:S208

Although the positive impact of elevated atmospheric carbon dioxide on pollen production has been established, impacts on fungal sporulation and antigen production have not been elucidated. This study examines the effects of rising atmospheric carbon dioxide on the quantity and quality of spores produced by fungi growing on timothy hay. Timothy grass (Phleum pratense) was grown at recent and projected future levels of carbon dioxide (300, 400, 500 and 600 µmol mol^–1). Leaves were used as substrate for the growth of Alternaria alternata and Cladosporium phlei. The abundance of spores produced by both fungi, as well as the size (microscopy) and antigenic protein content (ELISA) for A. alternata, were quantified. Timothy grass leaf dry weight and carbon-to-nitrogen ratio both increased at higher carbon dioxide levels. Leaf carbon-to-nitrogen ratio was positively correlated with the log of A. alternata spores produced per gram of leaf, but negatively correlated with antigenic protein content per spore. At the two highest levels of carbon dioxide, A. alternata produced nearly three-fold more spores and more than twice the total antigen per plant. C. phlei spore abundance increased with leaf carbon-to-nitrogen ratio, but overall spore numbers were much lower and per-plant production did not vary with carbon dioxide level. Elevated carbon dioxide often increases the biomass and carbon-to-nitrogen ratio of plant leaves. This study demonstrates that leaf changes induced by increasing carbon dioxide greatly enhance spore production by A. alternata, a ubiquitous allergenic fungus. This response may contribute to the increasing prevalence of allergies and asthma. Sporulation of C. phlei, a specialized pathogen of timothy grass, did not respond to increasing carbon dioxide in this study, suggesting that specialized and generalist fungal species may respond differently to rising atmospheric carbon dioxide. More study is needed to predict responses of different fungal groups to global changes.

Salicylic acid preconditioning increases tomato resistance to infection by potato purple top phytoplasma
W. WU (1), W. Wei (1), R. E. Davis (1), I. Lee (1), Y. Zhao (1)
(1) Molecular Plant Pathology Laboratory, ARS-USDA, Beltsville, MD
Phytopathology 100:S208

Columbia Basin potato purple top (PPT) phytoplasma is a newly discovered pathogen that causes serious diseases in potato and has the potential to affect other vegetable crops. Since the insect vector of PPT phytoplasma, the beet leafhopper, is a polyphagous species and has a wide geographic distribution, diseases associated with PPT phytoplasma infections may spread rapidly. The current study was aimed at investigating strategies to increase natural resistance of crops to PPT phytoplasma infections. The expression profiles of a set of defense/pathogenesis-related genes were examined in PPT phytoplasma-infected tomato plants. Results indicated that a delayed onset and a lack of sustained expression of a subset of defense-related genes may be key factors involved in PPT phytoplasmal disease development. Pretreatment of plants with SA prior to graft inoculation with PPT phytoplasma significantly altered the expression patterns of the same subset of genes and resulted in partial resistance of tomato to PPT infection. The findings shed new light on molecular mechanisms of phytoplasma pathogenesis and should aid in devising new strategies to mitigate phytoplasmal diseases.

Analysis of visual symptomatology in peach and plum inoculated with U.S. Plum pox virus isolates
A. Younkins (1), A. L. Stone (3), D. J. SHERMAN (3), W. L. Schneider (3), R. Scorza (2), V. D. Damsteegt (3)
(1) Dickinson College, Carlisle, PA, USA; (2) USDA-ARS-AFRS, Kearneysville, WV, USA; (3) USDA-ARS-FDWSRU, Frederick, MD, USA
Phytopathology 100:S209

Plum pox potyvirus (PPV) is an economically devastating potyvirus that affects Prunus species. Discovered in the United States in 1999, the Pennsylvania PPV isolates were primarily found in peaches (Prunus persica). When several of these original Pennsylvania isolates were inoculated onto plums (Prunus domestica), the isolates either did not transmit or showed few symptoms. This suggests that Pennsylvania PPV isolates were more adapted to peach as a host. An expanded experiment was designed using a greater number of Pennsylvania and New York PPV isolates to identify a U.S. PPV isolate with severe visual symptoms in plums, and to determine if symptom severity correlated with PPV titer. Two plum varieties (Bluebyrd and Stanley) were inoculated with fourteen PPV isolates from New York and Pennsylvania by aphid (Myzus persicae) or by grafting. Visual symptom severity was determined using a standardized symptom rating system. PPV titers were measured using Enzyme Linked Immunosorbent Assay (ELISA) and Real-time one step reverse transcription-PCR (RT-PCR). In contrast to PPV infection in peach, there was little correlation between average symptom rating and average ELISA titer or average Real-time RT-PCR Ct value in plum. One PPV isolate had been maintained for 10 years in both hosts: peach and plum. When this isolate was inoculated onto plum from both peach and plum sources, differences in titer and symptoms showed a possible host adaptation between PPV maintained in plum or in peach tissue.