Posted online September 30, 2005
An early warning system for detection of aerially transported plant pathogens: Puccinia graminis as a test case. C. W. BARNES (1), L. J. Szabo (1,3), J. J. Johnson (1), V. C. Bowersox (2), S. V. Krupa (3), D. A. Gay (2), and K. S. Harlin (2). (1) CDL, USDA-ARS, St. Paul, MN; (2) NADP, Illinois State Water Survey, Champaign, IL; (3) University of Minnesota, St. Paul, MN.
Atmospheric transport and deposition in rain is a common dispersal mechanism of certain rust fungi. Trap nurseries of susceptible plant material are labor intensive and reflective of previous infection events. An early warning system is needed. Puccinia graminis was used as a model to test whether real-time PCR can detect rust spores in rain samples. Weekly precipitation samples from 19 National Atmospheric Deposition Program (NADP) sites in the “Puccinia Pathway” of the Great Plains were used. The assay utilizes primers specific for rust fungi and a P. graminis specific TaqMan probe nested within a general primer pair within the ITS1 region of the rDNA. Lower limits of the assay are roughly 1-10 spores per sample based on spiking experiments. P. graminis spores were detected by June 1, 2004 in Minnesota and North Dakota, roughly one month prior to the first field observations in Minnesota. Frequency of detection increased through June, was weekly in July and August, and decreased by September. This study demonstrated that use of a PCR assay to detect spores in rain samples collected at geographically dispersed NADP sites can serve as a powerful early warning system for the movement of P. graminis. This methodology may be useful for monitoring the movement of other plant pathogens dispersed in the atmosphere.
Bionomics and management of Heterodera glycines in Michigan: With special reference to HG types. G. F. BIRD, G. F. Warner, A. Tenney, C. Bates, J. Davenport, and R. Gore. Dept. of Entomology, Michigan State Univ., E. Lansing, MI 48824.
Heterodera glycines (soybean cyst nematode, SCN) was first detected in MI in 1987. It is a key pest and known to exist in 33 counties. The MI Soybean Promotion Committee sponsors a soil sampling program for SCN diagnostics and grower education. Two to five-year rotation systems are use in MI. A four-year rotation is required to decrease SCN populations below the pathogenicity threshold. Use of resistant varieties derived PI54842, PI 88788 or PI437654 is common. Yield response is excellent to fair and SCN population reduction is excellent to poor. There is an increase in the availability of varieties derived from multiple sources of resistance, eg. PI88788 × PI437654 or PI88788 × PI54842. HG Types 0, 2, 5.7 and 2.5.7 have been identified. Under field conditions, HG Type classification and strength is impacted by continuous exposure to a source of resistance. Cultivars of poor hosts are being evaluated as SCN trap crops. Most SCN infested sites have additional soil quality (SQ) challenges. SQ restoration protocols are being evaluated for impacts on SCN population dynamics, host-parasite relationships and soybean productivity.
Combining disease control tactics and the utility of registered fungicides when applied according to disease predictors for managing late blight of celery. R. S. BOUNDS and M. K. Hausbeck. Dept. Plant Pathology, Michigan State University, East Lansing, MI 48824.
Fungicides are relied on as the primary tool to control late blight of celery, caused by Septoria apiicola, which can be applied when the threat of disease is minimal. Field trials were conducted to evaluate disease thresholds for delaying the initial fungicide application and to test disease predictors on ‘Dutchess’ celery in 2003 and 2004. Fungicide programs initiated three weeks after the common management practice and reapplied according to the S. apiicola and TOM-CAST disease predictors required 3 to 6 fewer applications relative to the standard weekly schedule. Azoxystrobin, propiconazole, and chlorothalonil were applied in various alternation programs every seven days or according to the S. apiicola and TOM-CAST disease predictors. All fungicide programs and application intervals were equally effective in limiting disease. Coupling preventive applications with disease predictors and the use of various fungicide alternation programs applied according to disease predictors were effective in managing late blight while reducing fungicide use.
Efficacy of fungicide seed treatments on soybean in North Dakota. C. A. BRADLEY (1), C. D. Chesrown (1), and T. C. Helms (2). (1) Dept. Plant Pathology, North Dakota State University; (2) Dept. Plant Sciences, North Dakota State University, Fargo, ND 58105.
Seed and seedling diseases can cause stand and yield reductions to soybeans planted in cool and wet North Dakota spring seasons. Field trials were conducted in 2003 and 2004 at several locations throughout North Dakota to evaluate the efficacy of two chemical, Warden RTA (mefenoxam + fludioxonil) and SoyGard (metalaxyl + azoxystrobin), and one biological, Yield Shield (Bacillus pumilus), fungicide seed treatments compared to an untreated control. In 2003, conditions were favorable for quick seed germination and emergence, and few differences among the treatments were observed. Conversely, in 2004, cool and wet soil conditions were present after planting, which slowed down seed germination and emergence, and differences among treatments were observed at some locations. In 2004, plots treated with Warden RTA, SoyGard, or Yield Shield had greater stand than the untreated control at 3 of 8 locations, 4 of 8 locations, and 3 of 8 locations, respectively. Plots treated with Warden RTA, SoyGard, or Yield Shield in 2004 had greater yield than the untreated control at 3 of 8 locations.
Monitoring surface irrigation water for Phytophthora capsici. A. J. GEVENS (1), K. H. Lamour (2), and M. K. Hausbeck (1). (1) Plant Pathology, Michigan State University, East Lansing, MI 48824; (2) Entomology and Plant Pathology, University of Tennessee, Knoxville, TN 37996.
Phytophthora capsici infects a range of solanaceous and cucurbit hosts worldwide. Green pears and cucumbers were used to bait the pathogen from Michigan irrigation sources during the growing seasons of 2002 to 2004. Baits were collected every 3 to 7 days, lesions excised, and incubated for 3 days on amended water agar. Isolates (186) were screened for sensitivity to mefenoxam and characterized for compatibility type (CT). Selected isolates (58) were compared using amplified fragment length polymorphism (AFLP) analysis. River isolates (146) were 45% insensitive, 29% intermediately and 26% fully sensitive to mefenoxam. Pond isolates (19) were primarily sensitive to mefenoxam. A run-off ditch in 2004 yielded 62% intermediately sensitive and 29% insensitive isolates. Approximately 60% of isolates from all sources were of the A1 CT. AFLP analysis suggests a lack of clonal lineage persisting over time or in specific geographical locations. While P. capsici was present in monitored sources, the pathogen did not overwinter in water.
Resistance to Sclerotinia sclerotiorum in PI391589A. E. HELLIWELL (1), T. Smith (1), S. A. Berry (1), S. G. Gordon (1), S. K. St. Martin (2), and A. E. Dorrance (1). (1) Dept of Plant Pathology, The Ohio State University, Wooster, OH 44691; (2) Dept of Horticulture and Crop Science, The Ohio State University, Columbus, OH 43210.
A total of 28 QTLs have been identified in soybean cultivars that contribute to resistance to S. sclerotiorum, the causal agent of Sclerotinia stem rot in soybean. Recently, a number of PIs, including PI391589B, were identified that have high levels of resistance to S. sclerotiorum. PI391589A was not evaluated for resistance; A and B designations have been assigned to PIs where off types occur. For these PIs the primary question is: are there additional resistance QTLs that may be unique from those that were identified previously? A cross of Kottman(2) × 391589A was made and increased to the BC1F4:6 generation. Resistance to S. sclerotiorum was evaluated with the cotyledon assay. Genotypic analysis was completed with 53 SSR markers covering 18 linkage groups. Preliminary data analysis identified two QTLs on MLGs A1 (Satt050) and G (Satt191) contributing resistance to S. sclerotiorum.
A PCR-based method to distinguish Eutypa lata and Eutypella vitis from grapevine. S. A. JORDAN, M. Catal, S. C. Butterworth, and A. M. C. Schilder. Dept. Plant Pathology, Michigan State University, East Lansing, MI 48824.
Two fungi were isolated from grapevines in Michigan vineyards with Eutypa dieback symptoms: Eutypa lata and Eutypella vitis. These fungi are difficult to distinguish morphologically but are genetically distinct as determined by sequencing of the ITS region. The ITS region of 25 E. lata and 15 E. vitis isolates was sequenced. The sequences were compared to each other and to existing sequences in GenBank. Eutypa lata sequences were more variable than those of E. vitis. PCR primers were designed for each species and evaluated against isolates of both fungi and 27 isolates of other fungi representing various pathogenic, saprophytic and endophytic genera on grapes and other small fruit crops. The primers were specific for their intended species. A multiplex PCR protocol was developed and successfully detected these fungi in wood samples from infected vines.
Genotypes of the soybean brown stem rot pathogen Phialophora gregata in major soybean production areas of North America. D. K. Malvick and E. Grunden. Dept. of Crop Sciences, U. of Illinois, Urbana, IL 61801.
Brown stem rot (BSR) of soybean is caused by Phialophora gregata (Pg). Genotype A of Pg causes stem and leaf symptoms, whereas genotype B primarily causes internal stem browning. Mature stem samples of BSR-resistant and susceptible soybean cvs. were collected arbitrarily in 2003 from plots in IL, IA, MN, MI, OH, WI, and Ontario. In 2004, samples were collected from plots and commercial fields in IL, IA, MN, MI, WI, IN, KS, ND, MO, NE, SD and Ontario. Samples were assayed by PCR with species-specific, genotype-discriminating primers to determine the distribution and frequency of Pg genotypes. In 2003, Pg was detected in all states and in 8.5% of total samples, with genotype A detected in 17% of positive samples, B in 75%, and A+B in 8%. In 2004, Pg was detected in all states except ND, NE, and MO and in 23% of total samples, with A detected in 30% of the positive samples, B in 57%, and A+B in 13%. In both years, A was the main genotype detected in susceptible cvs, whereas B predominated in the resistant cvs. Genotypes A and B of Pg are widespread, and genotype B infects resistant cultivars across the North Central USA and Ontario.
Fungicide treatment strategies for winter wheat in ND. M. McMullen and S. Meyer. Dept. Plant Pathology, North Dakota State University, Fargo, ND 58105.
Timings and rates of various registered fungicides were evaluated from 2001-2004 for maximum disease control and yield response of winter wheat cultivars grown in ND. Previous crop for each experiment was barley in 2000, spring wheat in 2001, and soybean in 2002 and 2003. Over the 4 years, a combination treatment of a half-rate, early season (tillering) fungicide application followed by a full rate, head emergence to flowering fungicide application resulted in the best disease control and yield; 75.5% reduction of fungal leaf spots, 85.9% reduction in leaf rust, 54.3% reduction in Fusarium head blight, and a 17.3% yield increase across cultivars. A single, flag leaf emergence treatment resulted in the least reduction in leaf and head diseases and only a 7.4% yield increase. A single, full rate late season (full heading to early flowering) application provided disease control equal to the early plus late combination treatment, with an average 11.1% yield increase. Of the cultivars tested, CDC Falcon generally had the highest leaf spot severities (up to 35.5%, untreated), while Wesley consistently had the highest Fusarium head blight severity index (up to 21.9%, untreated).
Growth of Phytophthora sojae on media amended with metalaxyl and mefenoxam. B. D. NELSON and I. Mallik. Dept. Plant Pathology, North Dakota State University, Fargo, ND 58105.
Phytophthora sojae is an important soilborne pathogen of soybean. Experiments were conducted over 4 to 7 weeks to examine growth of selected isolates on metalaxyl at various concentrations and with different sources (technical and commercial grade) of the compound. Eight isolates grew in the dark at 23°C after 2 to 3 weeks on V8 with metalaxyl at 5 µg a.i./ml. A distinct halo, consisting of a clearing to the medium, appeared around all colonies, but disappeared after 7 to 10 days. All 8 isolates had active hyphal growth for 5 weeks, then growth appeared to stop. After 9 weeks, transfers onto fresh V8 resulted in normal growth. Isolates also grew on 25 and 50 µg a.i./ml V8 and on commercial grade metalaxyl (Ridomil; 4.68% a.i.) at 5 µg a.i./ml. Four day old mycelia produced in shake culture and washed to remove nutrients also grew on amended media, indicating that the V8 agar plug was not required for growth on the metalaxyl. Isolates also grew on amended Lima bean agar, but growth was slower and halos were not as distinct. Six isolates showed growth after 3 weeks on V8 with mefenoxam (Apron; 33.3% a.i.) at 5 µg a.i/ml. The biological implications of these results are unknown.
The role of light intensity in the soybean Sclerotinia sclerotiorum interaction. A. J. PELTIER and C. R. Grau. Dept. Plant Pathology, University of Wisconsin-Madison, Madison, WI 53706.
Interaction phenotypes (IP) observed in controlled environments often do not relate to field performance. Light intensity is regarded as important to the outcome of interactions and a source of variation. Our objectives were to determine 1) the role of light in accession environmental sensitivity, 2) whether light influences IP in controlled environments and the ability to predict IP in the field, and to 3) compare methods to achieve maximum resolution among accessions in pair-wise comparisons. Trials were conducted in a growth chamber and in a nursery at the West Madison ARS. Five light environments (266-393 µmol m(^-2) s(^-1)) were created. The cut petiole method was used for inoculations; lesion length (LL) was assessed 10 dpi. A disease severity index was used in field evaluations. The IP of two resistant accessions, W04.1000 and W04.1002, were stable across light environments. ANOVA-type statistics detected more phenotypic differences among accessions than traditional LSD mean comparisons, with W04.1002 superior to a standard resistant check. Rank correlations of field and light environment IP indicated that light influenced prediction of field performance with controlled environment data. Disease evaluations conducted in environments with 303 to 337 µmol m(^-2) s(^-1) of light were most predictive of field IP (R(^2) = 0.94-1.0).
Solute and osmolality analysis of apple cells infected with Erwinia amylovora. P. Teubig (1) and W. Fricke (2). (1) Department of Plant Pathology, Michigan State University, East Lansing, MI 48824 USA; (2) Plant Biology, University of Paisley, Paisley, PA1 2BE, UK.
Fire blight is a destructive disease of apple and pear. Energy dispersing X-ray (EDX) and osmolality analysis were adapted to evaluate the interactions of fire blight pathogen Erwina amylovora and apple. These methods rely on the ability to sample sap from single cells using a glass micro-capillary. EDX analysis of apple upper epidermal cell content revealed differential distribution of magnesium and chlorine in apple leaf tissue. The higher magnesium concentration has not been observed in other plant leaf tissue analysed. E. amylovora inoculated apple plants tended to have higher concentrations of calcium in the mesophyll than control plants. In vivo experiments showed that epidermal cells of apple plants inoculated with an avirulent E. amylovora strain had higher osmolality than those inoculated with the wild-type. E. amylovora. This potential change in cellular osmolality of plants inoculated with an avirulent pathogen is indicative of the translocation of solutes into epidermal cells and may represent an active mechanism by which the plant initiates defense.
Upregulation of Erwinia amylovora genes during host tissue colonization. Y. F. Zhao, S. E. BLUMER, and G. W. Sundin. Dept. Plant Pathology, Michigan State University, E. Lansing, MI 48842.
Erwinia amylovora is the causative agent of fire blight, a necrotic disease affecting apple, pear and other rosaceous plants. Control of fireblight in the U.S. consists of cultural methods and the use of the antibiotics streptomycin and sometimes tetracycline. Genetic assays to identify genes involved in E. amylovora infection and colonization can help in the development of future control methods. We have conducted a modified in vivo expression technology (IVET) assay based on the infection of immature pear fruit by E. amylovora. A library of 20,000 random genomic DNA clones was inserted in front of a promoterless uidA gene. E. amylovora (Ea110) containing the fusions were screened for GUS activity after being cultured on immature pear disks. A total of 394 unique in planta expressed genes were identified by sequence analysis and grouped into categories including host-microbe interactions, stress response, regulatory function and nutrient acquisition. Experiments are in progress to determine the contribution of possible virulence determinants, including type II secretion, to fireblight disease.
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