MISSISSIPPI SOYBEAN PROMOTION BOARD PROJECT NO (YEAR 1) 2013 ANNUAL REPORT

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1 MISSISSIPPI SOYBEAN PROMOTION BOARD PROJECT NO (YEAR 1) 2013 ANNUAL REPORT Title of project: Development of a seedling inoculation technique to evaluate soybean for resistance to Phomopsis seed decay: proof of concept Principal investigator: Shuxian Li, Research Plant Pathologist, USDA-ARS, Crop Genetics Research Unit (CGRU), Stoneville, MS Phone: shuxian.li@ars.usda.gov. BACKGROUND AND OBJECTIVE Phomopsis seed decay (PSD) can severely affect soybean seed quality due to reduction in seed viability and oil content, alteration of seed composition, and increased frequencies of moldy and/or split beans. Hot and humid environmental conditions, especially during the period from pod fill through harvest stages, favor pathogen growth and disease development. PSD has resulted in significant economic losses. In 2009, due to the prevalence of hot and humid environmental conditions during the pod fill to harvest period in the southern United States, PSD caused over 12 million bushels of yield loss in 16 states, including 2.2 million bushels of yield loss in Mississippi. There are few management strategies for this disease, and these strategies have not completely protected soybean against PSD. Resistant varieties can provide some protection at no additional cost to the grower above the price of the planting seed. Soybean varieties may vary for resistance or tolerance to PSD. The disease development is very sensitive to environment, and the severity varies by years and location. Finding new sources of resistance to PSD requires reliable disease evaluation techniques. Development of fast inoculation and seedling assay techniques under controlled conditions will facilitate identification of genotypes with resistance to PSD without having to wait the whole growing season and being concerned with environment effects. PSD is primarily caused by Phomopsis longicola. This fungal pathogen has been reported to infect all soybean tissues. Therefore, we hypothesize that results from inoculation of soybean seedlings under controlled conditions, and the measurement of stem lesions after inoculation, will be comparable to the seed assay from the whole season of the field tests. We propose to develop a fast seedling inoculation technique at seedling (V2-V4) stage in a growth chamber to evaluate soybean for resistance to Phomopsis seed decay. The seedling cut-stem inoculation approach was used to compare the aggressiveness of P. longicolla isolates from different geographic origins (Li, et al., 2010). Apr

2 The objectives of this research are to (1) develop a seedling inoculation technique under controlled conditions to evaluate soybean for resistance to Phomopsis seed decay; (2) apply the seedling inoculation technique to test selected soybean varieties from our previous field trails at Stoneville (MSPB funded project # ); (3) analyze the correlation between seed assays from whole-season field trails and the seedling assays; and (4) provide information about soybean varieties with Phomopsis seed decay (PSD) resistance and high seed quality that are identified by both seed and cut-stem assays to soybean growers and seed industry personnel interested in disease resistance and seed quality. REPORT OF PROGRESS/ACTIVITY During FY2013, a pilot study was conducted to test a well-documented PSD-susceptible cultivar, Williams 82. We tested the effect of plant ages (2, 3, 4, and 5 weeks old) on the stem lesion after inoculation with Phomopsis culture pug on cut seedlings. Three trials were finished and results are summarized in Table 1. Results show that plants that were cut but not inoculated did not have stem lesions. However, all inoculated plants showed lesions on the stem. There were differences in different trials, and this may due to the age of the pathogen and its viability. However, stem lesions from plants that were inoculated at different ages within the same trial were not significantly different. We selected 3-week-old plants for the tests of six maturity group IV commercial soybean varieties that we have Phomopsis seed infection data for from field trials in Stem lesion length was measured at 4, 7 and 9 days after inoculation. Experiments were repeated three times. Differences in stem lesion length were found among varieties (Table 2). By analysis of the Pearson correlation coefficients and significance for stem lesion length in the seedling assays in a growth chamber, and percentage Phomopsis-infection of seeds from field trials at Stoneville, MS in 2013, we found significant (P 0.05) correlations between stem lesion length measured at 7days after inoculation in the seedling cut-stem inoculation assays and the Phomopsis seed infection of both inoculated (positively correlated) and non-inoculated (negatively correlated) data from the 2013 field trials (Table 3). In addition, the stem lesion length measured at 9 days after inoculation in the seedling assay was also significantly correlated with the field assay data from an inoculated and delayed harvest trial (Table 3). The correlations between seedling and field assay data are very encouraging. The seedling cut-stem inoculation method is easy and fast. It will facilitate identification of genotypes with resistance to PSD without waiting the whole growing season and having to be concerned with environment effects. Further study will be conducted to test the effects of the pathogen age on the stem lesion length after inoculation. We will also test selected MG V soybean varieties and different genotypes, including soybean plant introductions. Apr

3 Table 1. Means of stem lesion length of soybean variety Williams 82 inoculated with Phomopsis longicolla at different plant ages using a seedling cut-stem inoculation technique in three replicated trials in a growth chamber at Stoneville, MS. Stem lesion Length (mm) Plant age Trial (wk) 4 dai a 7 dai 9 dai 11 dai 14 dai Mean LSD (P 0.05) b a Days after inoculation. b Fisher s protected lease significant difference for means within the column. Apr

4 Table 2. Mean s o f stem lesion length of six maturity group IV soybean varieties inoculated with Phomopsis longicolla in three replicated seedling cut-stem inoculation trials in a growth chamber at Stoneville, MS. Stem lesion length (mm) Variety Trial 4 dai a 7 dai 9 dai Armor ARX Morsoy R Morsoy R2s Pioneer 94Y SS AP Armor ARX Morsoy R Morsoy R2s Pioneer 94Y SS AP Armor ARX Morsoy R Morsoy R2s Pioneer 94Y SS AP Mean LSD (P 0.05) b a Days after inoculation. b Fisher s protected lease significant difference for means within the column. Apr

5 Table 3. Pearson correlation coefficients and significance for stem lesion length of six maturity group IV soybean varieties inoculated with Phomopsis longicolla using a seedling cutstem inoculation technique in three replicated growth chamber trials, and percentage Phomopsis infection of seeds from field trials with inoculated and non-inoculated treatments and normal and delayed harvest times at Stoneville, MS in Values a LL4dai b LL7dai c LL9dai d PSD/Inoc1 e PSD/Non2 f PSD/Inoc2 g LL4dai (0.610) (0.311) (0.945) (0.663) (0.716) LL7dai (0.005) (0.228) (0.073) (0.055) LL9dai (0.363) (0.177) (0.062) PSD/Inoc (0.582) (0.176) PSD/Non (0.425) PSD/Inoc2 1 a Pearson correlation coefficients were calculated based on overall means of stem lesion lengths from three trials of the cut-stem inoculation tests and percentage Phomopsisinfection of seeds harvested from field trials from inoculated and non-inoculated treatments, and normal and delayed harvest times. Values in parentheses are probabilities. Percent Stem lesion length (mm) at: b 4 days after inoculation using a seedling cut-stem inoculation technique. c 7 days after inoculation using a seedling cut-stem inoculation technique. d 9 days after inoculation using a seedling cut-stem inoculation technique. e Percentage seed infected by Phomopsis longicolla in field tests harvested at normal time (R8 growth stage) from inoculated treatments in Percentage seed infected by P. longicolla in field tests harvested at delayed time (R8+2 wk): f from non-inoculated treatments in g from inoculated treatment in Apr