Chill seeds of non-woody plants before sowing: freshwater wetlands

Supporting evidence from individual studies

1. A replicated, controlled study in the early 1990s in a laboratory in Colorado, USA (Thullen & Eberts 1995) found that chilling hardstem bulrush Scirpus acutus seeds increased their germination rate. In all five of five comparisons, germination rates were higher for seeds that had been chilled before incubation (reaching 32–88% after 12 weeks) than for seeds that had not been chilled (reaching 0–7% after 12 weeks). Longer chilling periods typically increased germination rates (seven of nine comparisons; other two comparisons no significant effect; see original paper for data). Methods: Ninety-six sets of 10–50 hardstem bulrush seeds were incubated in flasks of water at 10/25°C or 18/22°C (night/day temperatures). Of these, 80 sets had been chilled in water (4°C; for 2–12 weeks) before incubation. The remaining 16 sets did not receive this chilling treatment. All seeds had been collected in August 1991 from two wild populations, stored in the laboratory for >5 months, and sterilized immediately before the experiment. Seed germination was monitored weekly during 12 weeks of incubation.

   Study and other actions tested

   Referenced paper

   Thullen J.S. & Eberts D.R. (1995) Effects of temperature, stratification, scarification, and seed origin on the germination of Scirpus acutus Muhl. seeds for use in constructed wetlands. Wetlands, 15, 298-304.

2. A replicated, randomized, controlled study in 1995–1996 in a greenhouse in Florida, USA (Ponzio 1998) found that chilling sawgrass Cladium jamaicense seeds had no significant effect on their germination rate, whilst freezing sawgrass seeds reduced their germination rate. Germination success did not significantly differ between seeds chilled in water (50% germinated) and seeds stored dry at room temperature (57% germinated). However, germination success of seeds that were frozen then chilled in water was significantly lower (41% germinated) than for the chilled-only seeds and the seeds stored at room temperature. Methods: In September 1995, freshly-collected sawgrass seeds were sprinkled onto 12 trays of sterilized soil (100 seeds/tray). Four trays were planted with seeds that had been chilled (stored in tap water at 4–10°C for one month before planting). Four trays were planted seeds that had been frozen then chilled (kept at 0°C for 25 days, then stored in tap water at 4–10°C for three days). The final four trays were planted with seeds that had not been soaked, chilled or frozen. All trays were placed in random positions in a greenhouse and watered daily until no more germination occurred.

   Study and other actions tested

   Referenced paper

   Ponzio K.J. (1998) Effects of various treatments on the germination of sawgrass, Cladium jamaicense Crantz, seeds. Wetlands, 18, 51-58.

3. A replicated, randomized, paired, controlled study in 1997 in a laboratory in Utah, USA (Jones et al. 2004) found that chilling sedge Carex spp. seeds before sowing typically had no significant effect on their germination rate, although the precise effect depended on the species and the light/temperature regime after sowing. The germination rate of chilled and unchilled seeds was statistically similar in 22 of 36 comparisons (for which chilled: 0–98%; unchilled: 0–96%). This included 13 of 18 comparisons of seeds germinating in the dark, and 11 of 12 comparisons of seeds germinating in summer temperatures. Chilling significantly increased the germination rate in the other 14 of 36 comparisons (for which chilled: 17–86%; unchilled: 0–32%). This included six of six comparisons of seeds germinating in the light in spring temperatures. Methods: Two-year-old seeds of beaked sedge Carex utriculata and Nebraska sedge Carex nebracensis were sown into a total of 384 petri dishes (192 dishes/species; 32 seeds/dish), then incubated in the laboratory. Seeds in 288 dishes were chilled before sowing (moist incubation at 5°C for 7, 30 or 150 days). Seeds in the other 96 dishes were not chilled. Dishes were allocated to various other treatments (including germination in light vs dark and different post-sowing temperature regimes). Each incubator shelf received four random dishes of each treatment combination. Germination rates, as a percentage of viable seeds, were recorded for each dish 36 days after sowing.

   Study and other actions tested

   Referenced paper

   Jones K.L., Roundy B.A., Shaw N.L. & Taylor J.R. (2004) Environmental effects on germination of Carex utriculata and Carex nebrascensis relative to riparian restoration. Wetlands, 24, 467-479.

4. A replicated, paired, controlled study in 2003–2004 in six prairie pothole wet meadows in Iowa, USA (Kettenring & Galatowitsch 2011) found that chilling sedge Carex spp. seeds increased germination rates for seeds sown in natural meadows, but not for seeds sown in restored meadows. In natural meadows and averaged across all four sown species, the germination rate was higher for chilled seeds (3%) than seeds kept at room temperature (<0.3%). However, the difference was only significant for one of four species when analyzed separately (bristly sedge Carex comosa; chilled: 3%; room temperature: <0.2%). In recently rewetted meadows, the germination rate did not significantly differ between treatments, whether averaged across species (chilled: 13%; room temperature: 9%) or analyzed for individual species (chilled: 9–17%; room temperature: 8–11%). Methods: In late spring 2003, wild-collected seeds of four sedge species were sown into the wet meadow zone of six prairie potholes (three natural and three rewetted one year previously). Within each pothole, eighteen 50-cm diameter plots/species were sown with 300 seeds. Nine plots/pothole received chilled seeds (kept at 1–5°C over the previous winter) whilst the other nine received unchilled seeds (kept at room temperature over the previous winter). Seedlings were counted for two growing seasons.

   Study and other actions tested

   Referenced paper

   Kettenring K.M. & Galatowitsch S.M. (2011) Carex seedling emergence in restored and natural prairie wetlands. Wetlands, 31, 273-281.

5. A replicated, randomized, paired, controlled study in 2015 in a laboratory in Utah, USA (Marty & Kettenring 2017) found that chilling seeds of three bulrush species either increased or had no significant effect on their germination rate, depending on the duration of chilling. Seeds that had been chilled for 180 days before sowing had a higher germination rate than unchilled seeds in 14 of 18 comparisons (for which chilled: 22–90%; unchilled: 0–66%). However, seeds that had been chilled for 30 days before sowing had a statistically similar germination rate to unchilled seeds in 16 of 18 comparisons (for which chilled: 0–62%; unchilled: 0–77%). Chilling seeds never significantly increased the germination rate of one of the species, threesquare bulrush Schoenoplectus americanus (four of four comparisons; chilled: 61–77%; unchilled: 50–75%). Methods: Field-collected seeds of three bulrush species were sown into petri dishes filled with sand (16–64 dishes/species; ≥50 seeds/dish). There were 4–16 dishes/species for each of four pre-sowing temperature treatments: chilling (4°C) for 30 or 180 days, or room temperature for 30 or 180 days. Replicates were split across two incubators. After sowing, dishes were kept saturated or flooded and incubated at 18–35°C. Germination rates for each dish were recorded four weeks after sowing.

   Study and other actions tested

   Referenced paper

   Marty J.E. & Kettenring K.M. (2017) Seed dormancy break and germination for restoration of three globally important wetland bulrushes. Ecological Restoration, 35, 138-147.

6. A replicated, randomized, controlled study in 2015 in a greenhouse in Utah, USA (Marty & Kettenring 2017) found that chilling seeds of three bulrush species either increased or had no significant effect on their germination rate, with the precise effect depending on factors such as the species, source site and whether seeds were soaked in chemicals before chilling. Seeds that had been chilled before sowing had a higher germination rate than unchilled seeds in 17 of 30 comparisons (for which chilled: 35–88%; unchilled: 2–26%). This included 9 of 10 comparisons involving seeds that had not been chemically treated before chilling. Chilled seeds had a similar germination rate to unchilled seeds in the other 13 of 30 comparisons (for which chilled: 28–69%; unchilled: 2–75%). This included 10 of 10 comparisons involving seeds that had been soaked in bleach before chilling. Methods: Field-collected seeds of three bulrush species were sown onto sand in the greenhouse (36–72 sets of seeds/species; ≥100 seeds/set). Seeds in 18–36 random sets/species were chilled at 4°C for 30 days before sowing. The other sets were kept at room temperature. Some sets were also soaked in acid, bleach and/or water before or after chilling. After sowing, seeds were kept saturated and at 28–35°C. Germination rates for each set were recorded five weeks after sowing.

   Study and other actions tested

   Referenced paper

   Marty J.E. & Kettenring K.M. (2017) Seed dormancy break and germination for restoration of three globally important wetland bulrushes. Ecological Restoration, 35, 138-147.