Volume 20, No. 1, 1998

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(pp. 5-17)
Testing Tree Seeds for Vigor: A Review
F. T. Bonner
This review examines the use of vigor tests for tree seeds. It suggests that precise evaluations of these tests and their application with seeds of woody plants is not yet possible. This is due to the wide genetic variation, primarily manifested in variable maturity and dormancy, that exists in most tree seed lots. Sensitive measurements of germination rate during standard germination tests have proved to be just as good, if not better, than any vigor test in judging the quality of seed lots. Accelerated aging, leachate conductivity, and germination rate all show promise, but extensive tests of field  emergence are needed to validate the laboratory test procedures and interpretation.
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(pp. 18-22)
Initiatives to Improve the International Seed Health System: A Review
Denis C.McGee
Seeds can be efficient means of introducing plant diseases into geographical regions by transmitting seed-borne plant pathogens. Numerous examples exist of imports of diseased seeds that have had serious economic consequences, such as karnal bunt (Tilletia indica Mitra) from Asia to Mexico and the US (Brennan et al, 1992; Neergaard, 1977). The need for international quarantine controls to prohibit introductions of economically important plant diseases by seeds is clearly justified.
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(pp. 23-31)
Soybean Genetic Resistance and Benomyl for Phomopsis Seed Decay Control
RogerW. Elmore,* Harry C.Minor, and Ben. L. Doupnik, Jr.
Phomopsis seed decay (PSD) is the seed infection phase of an important soybean [Glycine max (L.) Merr.] disease complex and is caused by several species of Phomopsis and Diaporthe. The complex (which also includes pod and stem blight and stem canker) reduces seed quality in all major soybean growing areas of the world. Farmers use cultural and chemical methods to partially control PSD. Genetic resistance is an important defense against plant disease and an important alternative to chemical control.

Widely grown cultivars with genetic resistance to PSD are not available; however, resistance to PSD froma plant introduction (PI 417479) is now incorporated into an improved breeding line. The objective of this study was to ascertain the relative effectiveness of genetic resistance and fungicide application (benomyl) for the control of PSD. Six lines from crosses with PI 417479 were compared to two cultivars and PI 417479 at Missouri (MO) and Nebraska (NE) in 1989 and 1990. Benomyl reduced PSD by 55% atMO. The PSD resistant line had 1.3% and 3.3% PSD with and without benomyl, respectively, at MO; whereas PSD of themoderately resistant cultivar, ‘Williams 82’,was 5.3% and 18.0%, respectively. In environments favoring high PSD levels like those encountered inMO, lines resistant to PSD can provide effective control without fungicide application. In these same environments fungicide application is necessary to reduce PSD in susceptible and moderately resistant lines. Incorporating PSD resistance into cultivars with high yield potential may enhance soybean profitability by reducing the need for chemical control.
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(pp. 32-42)
Physiology and Biochemistry of Recalcitrant Guarea guidonia (L.) Sleumer Seeds
K. F. Connor* and F. T. Bonner
Investigations of recalcitrant, or desiccation-sensitive, seeds have as yet failed to identify the causes of this phenomenon. Experiments with Guarea guidonia (L.) Sleumer (American muskwood) were initiated to determine the effects of desiccation on the physiology and biochemistry of the seeds of this tropical tree species. Seeds were air-dried at room temperature for 7 days. At intervals, germination was tested, moisture content determined, and lipids extracted.

The bulk lipids, nonpolar lipids, monoglycerides, and phospholipids were analyzed by gas chromatography (GC), thermal characteristics of whole tissue samples were examined using differential scanning calorimetry (DSC), and moisture content was determined using the Karl Fisher analysis. DSC thermograms showed that as moisture content and germinability of seeds declined, so did enthalpy values and onset temperatures of cotyledon tissue and embryonic axes. GC analyses determined that unsaturated fatty acids accounted for approximately 70% of the bulk lipids; however, ratios of unsaturated/saturated fatty acids and amounts of individual fatty acids fluctuated between test periods. Palmitic acid was the most common saturatedfatty  acid, and linoleic acid was the most prevalent unsaturated fatty acid. Generally, Karl Fisher analyses of seed moisture content offered the best possibility of monitoring seed deterioration during drying.
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(pp. 43-55)
Detection and Removal of Off-colored Bean Seeds by Color Sorting
P. C. Lee, D. H. Paine, and A. G. Taylor*
Off-colored seeds are routinely present in a seed lot, and subtle differences in seed coat color are most easily observed in white-seeded crops. Off-colored seeds not only affect the general appearance of a seed lot, but also may be indicative of decreased seed quality. Seeds of a white-seeded cultivar of snap bean (Phaseolus vulgaris L.) were manually sorted into brown, tan, green and white fractions under white light. Germination tests were conducted on each sorted fraction; nonsorted beans served as the control.

The white fraction resulted in 6.5 percentage point higher germination than the nonsorted seeds, while the percent germination from brown, tan and green fractions was significantly lower than the control. Therefore, seeds with off-colored seed coats were of poor quality. Light reflectance was determined with a spectrophotometer from off-color fractions and compared with white seeds. The greatest difference in light reflectance difference between all off-colored and white seeds was observed from 275 to 450 nm with a 360 nm maximum. Little differences in light reflectance were measured between different colored seeds in the NIR (>750 nm). Similar differences in light reflectance were measured from rapidly aged seeds. Beans were also sorted by an electronic color sorter, Xeltron 10 RMP, with two sorting logics (absolute threshold logic and relative threshold logic). Electronic sorting was able to effectively remove off-colored seeds from the seed lot.
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(pp. 56-70)
Corn Cold Test Germination as Influenced by Soil Moisture, Temperature, and Pathogens
J. M.Woltz,* D. M. TeKrony, D. B. Egli, and P. Vincelli
The cold test is the most popular vigor test for corn (Zea mays L.) seed; however, standardization among seed testing laboratories has not been accomplished. This experiment determined the effect of soil moisture, cold period temperature and soil-borne pathogens on the germination of treated and untreated seed lots of five corn hybrids using the ‘shoe box’ cold test method.

Seeds were planted in microwaved (pathogen-free) and non-microwaved (raw) soil adjusted to moisture levels corresponding to soil water potentials of -30, -50, -400 and -1000 kPa and tested at 10°C for seven days followed by 25°C for five days. A second experiment utilized a cold period temperature range from 4.5 to 13°C at a soil water potential of -40 kPa. Germination of untreated seed in raw soil ranged from 85% at -1000 kPa soil water potential (driest) to 57% at -30 kPa (wettest), with best separation of seed lots occurring at -50 kPa (0.24 kg kg-1 soil moisture). Seed treatment with Captan increased germination and nearly eliminated differences among  soil moisture levels. The highest germination for treated and untreated seed lots in raw soil occurred at 4.5°C and 13°C, while the lowest germination (greatest stress) occurred at 9 and 10°C. No germination differences occurred among seed lots or among any experimental treatments (soil moisture, temperature, seed treatment) when pathogen-free soil was used. The presence of soil-borne plant pathogens was the primary factor influencing cold test germination. The major variables (soil moisture, temperature and seed treatment) that influence pathogen response and cold test germination can be controlled in the laboratory,which should aid in standardization.
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(pp. 71-82)
Effects of Desiccation on the Recalcitrant Seeds of Carapa guianensis Aubl. and Carapa procera DC.
K. F. Connor,* I. D. Kossmann Ferraz, F. T. Bonner, and J. A. Vozzo
This study was undertaken to determine if the seeds of Carapa guianensis Aubl. and Carapa procera DC. undergo physiological, biochemical, and ultrastructural changes when they are desiccated; and to find if these changes can be used to monitor viability in Carapa. Seeds were air-dried at room temperature for 7–11 days. Samples were taken at frequent intervals and germination was tested, moisture determined, lipids extracted, and samples taken for electron microscopy.

The moisture content (MC) of the embryonic axes remained high throughout the experiment. The cotyledons were drier and had a higher MC variation between individual seeds during desiccation. While Karl Fisher moisture analyses indicated no relationship between axis MC and seed viability, differential scanning calorimetry (DSC) thermograms showed a strong relationship between the melting endotherm peak onset values, enthalpy (heat content) and seed germinability. Both techniques were ineffective in determining changes in seed viability when viability remained above 50%. Analyses of the bulk lipids indicated that changes were taking place,  but gas chromatography (GC) results were inconsistent from year to year. Electron microscopy (EM) examinations found that cellular contents of Carapa showed little organization when seeds were fresh, but that spherosomes accumulated as desiccation progressed. These data and those from the moisture, DSC and GC analyses, add support to the hypothesis that storage problems of recalcitrant seeds are associated with intact seed MC and with lipid composition, metabolism, and distribution in the cells.
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(pp. 85-90)
Leaf Vernation Test for Perennial Ryegrass
Don F. Grabe*
A supplementary test procedure is described for improving the accuracy of the fluorescence test for perennial ryegrass. In practice, a fluorescence test is conducted as usual. Fluorescent seedlings are then transplanted and grown to the 3- to 4-leaf stage to be identified on the basis of leaf vernation.

Leaf vernation refers to the arrangement of the leaves as they emerge from the young shoot—leaves of annual ryegrass are rolled and leaves of perennial ryegrass are folded. Subtracting the number of fluorescent perennials from the total number of fluorescent seedlings before calculating test fluorescence provides a more accurate determination of the presence of annual ryegrass than does the fluorescence test alone.
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(pp. 91-93)
Non Peroxidase Oxidation of Guaiacol
Richard A. Vierling, Reid G. Palmer and James R.Wilcox
The peroxidase test for soybean [Glycine max (L.) Merrill] is a standard assay used in the identification of soybean cultivars. Cultivars are divided into two groups based on the presence of either high or low seed coat peroxidase activity. High or low activity is based on the observed color change associated with oxidized guaiacol.

Seed coat peroxidase activity also can be measured using the soybean peroxidase capture assay (SPCA), which uses a monoclonal antibody to isolate the enzyme from biological samples. Unlike the SPCA, the guaiacol assay does not separate the peroxidase enzyme from other enzymes or reactive oxygen species. Results showed that 7% of the stored soybean seeds tested, which showed high peroxidase activity using the guaiacol test, showed no measurable peroxidase activity using SPCA. This suggests that the initial guaiacol oxidation resulted from non enzymatic oxidation of guaiacol by reactive oxygen species or other oxidative enzymes and not from seed coat peroxidase enzymatic activity.
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(pp. 94-98)
A Bioassay for Detection and Quantification of Xanthomonas campestris in Sunflower Seeds
R. S. Romeiro,* A. B.Moura and A. J. A.Monteiro
The combined techniques of extraction and plating on culture medium with hypocotyl pricking inoculation was studied for detection of Xanthomonas campestris in naturally infected sunflower seeds. The method was efficient to detect the pathogen and allowed quantification of viable bacterial cells in association with two seeds lots, estimated as, respectively, 3.096 x 107 c.f.u. and 2.088 x 106 c.f.u. per gram of seeds. Parallel assays by direct planting indicated a percentage of transmission of, respectively, 18% and 5%. This is a simple method adaptable to routine testing of many samples.
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(pp. 99-105)
Rapid Germination of Eastern Dogwood (Cornus florida L. cv. Small) using Embryo Extraction, Cut Cotyledons and Gibberellic Acid
J.E. Laufmann* and L.E.Wiesner
Normal seed germination of Cornus florida L. cv. Small requires a moist prechill treatment at 3–5°C for 90–120 days followed by 20–30 days of germination at 20–30°C. Expected germination using this procedure is 30–45%. This low germination percentage is primarily due to slow seedling growth and high fungal contamination. To decrease the prechill period and increase germination, seeds from three locations were evaluated using embryo extraction, cut cotyledons, and gibberellic acid (GA3) treatments.

Storability in liquid nitrogen (LN2) was also examined. Viabilities based on tetrazolium test results were correlated with germination percentages obtained using these procedures for seeds from all locations. The prechill treatment (90–120 days) was eliminated and the germination period was decreased from 20–30 days to 6–14 days by performing embryo extraction, clipping cotyledons, and placing embryos on blotters soaked with 1mM GA3. Germination using this method was as high as 100%. Germination rates of LN2-exposed seeds ranged from 45–82%, and seed death was associated with fungus or bacteria and not due to mechanical damage from exposure to cryogenic temperature.
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(pp. 106-108)
Lactic Acid Clearing of Grass Seeds in Tetrazolium Tests
Don F. Grabe* and John A. Peters
Lactophenol is routinely used as a clearing agent for grass seeds in the tetrazolium test. Because of the toxic nature of lactophenol, a non-toxic replacement is highly desirable. Lactic acid was compared with lactophenol in over 200 samples of grass seeds, including many varieties of six seed kinds. Seven analysts participated in rating the two reagents for clearing ability, safety, ease of preparation, odor, and glare. In all factors evaluated, lactic acid was equal to or superior to lactophenol.We conclude that lactic acid may be substituted for lactophenol as a clearing agent for grass seeds in the tetrazolium test.
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