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© 2005 Plant Management Network. Cold Tolerance Evaluation of Improved Diploid and Tetraploid Cultivars of Perennial Ryegrass Deborah L. Warnock, Graduate Student, Richard H. Leep, Professor, Suleiman S. Bughrara, Assistant Professor, Department of Crop and Soil Sciences, Michigan State University, Plant and Soil Science Building, East Lansing 48824; and Doo-Hong Min, Research and Extension Forage Specialist, Michigan State University, Upper Peninsula Experiment Station, Chatham 49816 Corresponding author: Deborah L. Warnock. warnock1@msu.edu Warnock, D. L., Leep, R. H., Bughrara, S. S., and Min, D.-H. 2005. Cold tolerance evaluation of improved diploid and tetraploid cultivars of perennial ryegrass. Online. Crop Management doi:10.1094/CM-2005-0221-01-RS. Abstract Livestock and hay producers in temperate climates could greatly benefit from more high-quality, cold-tolerant forage cultivars. Currently in the Midwest, no rotational grazing experiments have been done studying perennial ryegrass (Lolium perenne L.) cold tolerance in both a grass monoculture and a binary mixture with ladino white clover (Trifolium repens L.). The objective of this study was to evaluate cold tolerance of diploid (‘Aries’ and ‘Mara’) and tetraploid (‘Barfort’ and ‘Quartet’) cultivars of perennial ryegrass. Aries and Quartet are from New Zealand, Mara is from Romania, and Barfort is from Holland. Germination across a range of temperatures was evaluated in the laboratory, using a thermogradient plate to determine if cultivars would germinate equally well as temperatures declined. In the field, grasses were grown as monocultures and as binary mixtures with ladino white clover and rated for winter hardiness at three Michigan field locations. The field trials were monitored during the establishment year (2001-2002 growing season) for winter injury, ground cover, and yield. Both the laboratory and field results showed Barfort as one of the better-adapted cultivars to cold temperatures. Mara was also adapted to cold temperatures in the field, but was poorly adapted in laboratory studies. Both autumn ground cover and first harvest yields correlated closely with amount of winter injury. As the season progressed and the grasses recovered from the winter injury, differences between cultivars were not as apparent. At all three Michigan locations (Hickory Corners or KBS, Lake City, and Chatham), however, Mara was one of the highest yielding cultivars for both first and total yields. Introduction The goal of forage growers is to maximize yield while ensuring stand persistence. Stand persistence depends on good management practices such as controlling weeds, maintaining proper soil fertility, and using a proper harvest schedule, but it also requires inherent winter hardiness of the forage species and cultivar. Producers who live in more temperate climates could greatly lengthen the stand life by growing more winter-hardy cultivars for hay or pasture production. Perennial ryegrass (PR) is the most common perennial grass grown in temperate regions throughout the world (17). In Michigan, PR is grown on approximately 10,000 acres and is primarily used in pastures (1). Michigan producers like PR because of its high quality and yield, but it has low winter hardiness (ability of a plant to survive winter). Humphreys and Eagles (7) reported that the cold tolerance (ability of plants to tolerate stresses when exposed to temperatures below 32°F) of PR needed to be improved before the species could be used in the United Kingdom and northern continental climates (7). A study by Frame (6) compared herbage productivity of several grass species, including PR, in the United Kingdom and determined that PR performed poorly compared to the other grass species mainly because production was affected by winter damage (6). Research in Canada evaluated the hardiness of thirty PR cultivars and found a large variability for tolerance to subfreezing temperatures; however, cultivars from Canada and northern Europe were the most winter-hardy (8). Studies in Wisconsin, however, suggested that PR is able to over-winter in colder climates, even where snow cover is unreliable (2,10). Norwegian studies found that PR populations from the Swiss Alps, adapted to long-lasting snow cover, were not any better adapted to Norwegian climates than the indigenous populations tested (12). Perennial ryegrass cultivars are either diploid (contains two sets of chromosomes) or tetraploid (contains four sets of chromosomes). Tetraploid plants can occur naturally, but more often chromosome doubling is induced by chemical means. Perennial ryegrass tetraploids were first developed in Holland (4) in the 1960s. In 1969, Michigan State University developed a tetraploid ryegrass hybrid called ‘Tetrelite,’ but it was only moderately winter-hardy. Past research results indicate diploid cultivars of PR have a higher cold tolerance than tetraploid cultivars (5,13). Tetraploid cultivars of other forage species, such as red clover (Trifolium pratense L.) and meadow fescue (Festuca pratensis Huds.), have also been shown to have lower cold tolerance than diploids (11,14). Currently, seed companies are offering improved PR cultivars with increased winter hardiness. The objective of this study was to evaluate the cold tolerance of improved PR cultivars (both tetraploid and diploid) in the laboratory and over a wide range of climatic field conditions in Michigan at sites ranging from 42°N to 46°N latitude. Laboratory Test for Germination On 1 October 2001 and 18 February 2002, germination response to a range of temperatures was evaluated for four PR cultivars using a one-way thermogradient plate. The procedure used was a modified version of that described by Wade et al. (15). The improved PR cultivars used were Aries and Mara (diploid), and Barfort and Quartet (tetraploid). The experiment was a completely randomized block design, with three replications of each cultivar. The thermogradient plate was marked such that petri dishes could be placed equidistantly across the apparatus, allowing seeds to be tested at temperatures of approximately 40, 50, 60, 70, and 80°F. At each of the five temperature regimes, three petri dishes were randomly placed in three blocks for a total of nine petri dishes per temperature regime. The petri dishes were lined with Whatman Number 4 filter paper, which was divided into four equal quadrants. Five seeds were placed in each quadrant and covered with another layer of the filter paper. One week constituted one run of the experiment. At the end of the week, seeds were scored as germinated or not germinated. Germination was defined as the emergence of a radicle from the seed coat. Data were statistically analyzed using the PROC MIXED procedure of SAS version 8e (SAS Institute Inc., Cary, NC) to test for statistical significance of cultivar, temperature, and cultivar × temperature interaction. Run and replication were considered random variables with run and temperature nested within replication. Unequal variances for temperature were accounted for using the repeated/group function in SAS. No data transformation was necessary. Cold Tolerance Testing In the Field Field trials were established in August 2001 at three locations in Michigan. The first seeding occurred on 7 August 2001 at the Upper Peninsula Experiment Station at Chatham (46.7°N) on a Chatham stony loam (coarse-loamy, mixed, frigid Typic Haplorthods). The second seeding was planted on 14 August 2001 at the Lake City Experiment Station at Lake City (44.2°N) on a Nester sandy loam (fine, mixed, semiactive, frigid Oxyauic Glossudalfs). The third location was at W. K. Kellogg Biological Station (KBS) at Hickory Corners (42.3°N) on a Kalamazoo loam (fine-loamy, mixed, semiactive, mesic Typic Hapludalfs) and was planted on 21 August 2001. Four improved cultivars of PR were planted as monocultures and as binary mixtures with ladino white clover. The PR cultivars were the same as those used in the laboratory experiment. In the monoculture, PR was seeded at a rate of 30 lb/acre. In the binary mixture, PR was seeded at a rate of 25 lb/acre and ladino white clover was seeded at 3 lb/acre. Field plot size was 6 × 15 ft and plots were seeded with a Carter forage plot seeder (Carter Manufacturing Co. Inc., Brookston, IN). The experiment had a randomized complete block design with three replications. Cultivars in the grass monoculture were maintained with split applications of ammonium sulfate fertilizer (21-0-0). Each of the four applications consisted of 50 lb of actual nitrogen per acre, for a total of 200 lb of nitrogen per year. Applications were in early spring, following the first and second grazing periods, and in late August. No nitrogen fertilization was applied to the binary mixture because this contributes to a decline in the clover component of the mixture (17). Data collection for the field study established in the fall of 2001 began in the spring of 2002 and included visual assessments for winter injury (on a 1 to 9 scale where 1 = no winter injury, 9 = winter-killed) and ground cover (on a 1 to 9 scale where 1 = no ground cover, 9 = complete ground cover). Forage yield was determined at each grazing event by randomly placing a 1.5-×-1.5-ft quadrat per plot. The forage inside the quadrat was clipped at a 3.0-inch height and removed. All removed samples were dried at 140°F for 48 h and the dry weight was recorded. Field data were statistically analyzed by analysis of variance (ANOVA), using the PROC GLM procedure of SAS version 8e (SAS Institute Inc., Cary, NC). No data transformation was necessary. Cultivar means were compared using Fischer’s LSD. Laboratory Germination Results and Discussion The germination results over a range of temperatures are presented in Table 1. At one of the five temperature regimes (40°F) no germination was observed. When germination occurred, germination among the cultivars significantly differed (P < 0.05) at only two of the five temperatures. At a temperature of 60°F, Aries, Barfort, and Quartet showed the highest germination percentages. The germination of Mara was significantly lower. At 50°F the only significant difference was between Barfort and Mara. Diploid PR is thought to have a greater cold tolerance, but this experiment showed Barfort, a tetraploid cultivar, to have greater cold tolerance than Mara, a diploid cultivar. Table 1. Germination results from thermogradient plate experiment. Data includes two runs of the experiment with each run lasting one week. Germination was defined as the emergence of a radicle from the seed coat.
Soil temperature affects both the rate of germination and growth of seedlings. Cool season grasses grow actively when soil temperatures are between 60 and 70°F. Too high or low a temperature, however, can prevent germination or greatly reduce the rate of seed germination. Germination did not occur at 40°F because the temperature was too extreme and the seeds were most likely dormant. The four temperatures where germination did occur have large differences among coefficient of variation (CV) values (Table 1). Variation was among seeds tested at each temperature. This is probably due to the design of the thermogradient plate. One end of the metal plate was heated with hot water, while the opposite end of the plate was cooled with antifreeze. Although the plate was insulated to minimize temperature variation, the heating and cooling sources were subjected to varying air temperatures in the laboratory. This resulted in larger temperature variations at each end of the metal plate and more stable temperatures at the center of the plate. In fact, the lowest CV values occurred at marked regions of the plate corresponding to temperatures of 70 and 80°F with the exact center of the plate falling between the two regions. Winter Injury Winter injury evaluations were taken in April 2002. Winter injury differences could be due to cultivar, location, or an interaction of these two factors. An interaction would indicate the cultivars’ winter injury did not remain constant across the three locations. No interaction was present in the binary mixture or in the monoculture. Since there was no interaction, factors could be examined individually. Monoculture winter injury ratings were significantly different among the three locations. There was no significant difference in binary mixture winter injury ratings among the three locations which indicates that growing perennial ryegrass with clovers may reduce winter injury. Chatham had reliable snow cover throughout the entire winter, while Lake City had snow cover mostly in January and February of 2002, and KBS had snow cover from late December 2001 to February 2002. Due to differences in snow cover (snow insulates the forage, minimizing temperature fluctuations and reducing the frequency and severity of freezing-thawing events), results will be discussed by location. Daily air temperatures, both lows (Fig. 1) and highs (Fig. 2), were recorded from September 2001 through March 2002. Monthly averages for minimum and maximum air temperatures, along with thirty-year averages, can be seen for KBS (Table 2), Lake City (Table 3), and Chatham (Table 4).
Table 2. Monthly averages for minimum and maximum air temperatures at W. K. Kellogg Biological Station (KBS), MI. Monthly averages for the trial year, September 2001 to March 2002, are compared to the 30-year average (1971-2000).
Table 3. Monthly averages for minimum and maximum air temperatures at Lake City, MI. Monthly averages for the trial year, September 2001 to March 2002, are compared to the 30-year average (1971-2000).
Table 4. Monthly averages for minimum and maximum air temperatures at Chatham, MI. Monthly averages for the trial year, September 2001 to March 2002, are compared to the 30-year average (1971-2000).
No significant difference in winter injury at the KBS location was observed among any of the cultivars when grown as a binary mixture (Table 5). This may be due to the clover helping to "catch" the snow and providing insulation. When grown as a monoculture, Aries had a significantly greater (P < 0.05) amount of injury (Table 6). At Lake City in the binary mixture, Aries and Quartet exhibited significantly more injury (Table 5). In the monoculture, Mara and Barfort resulted in the least amount of winter injury. Quartet exhibited a significantly greater (P < 0.05) amount of injury, and Aries had significantly more injury than Quartet (Table 6). It was expected that Quartet would not perform as well in a colder climate without snow cover because it was tetraploid. Barfort, even though it is tetraploid, probably performed well because it is from Holland, whereas Quartet is from New Zealand. Kunelius and Castonguay (8) also found that cultivars originating from a similar climate as the climate they were being tested at were better adapted than cultivars originating from a different climate (8). At Chatham, the binary mixture needed to be reseeded due to a poor stand of clover. For this reason, data was only collected from the grass monoculture. At Chatham, no difference in winter injury was observed among cultivars in the monoculture (Table 6). Table 5. Winter injury results for cultivars grown as binary mixtures at W.
Table 6. Winter injury results for grass monoculture cultivars at W. K.
With little or no snow cover, forage grasses can be damaged by subfreezing temperatures, standing water, ice encasement, heaving, and desiccation. At Chatham, there was no significant difference among the cultivars, because all four cultivars were rated as having no winter injury. This is probably due in part to the good snow cover at Chatham throughout the entire winter. Ground Cover Ground cover evaluations were taken in early spring (middle of April to early May) and again in late fall (end of October to early November), depending on location. In the spring at KBS, ground cover rankings in the binary mixture were as follows: Barfort > Mara, Quartet > Aries. Mara and Quartet had equal ground cover ratings. Barfort produced significantly more (P < 0.05) ground cover than the other cultivars (data not shown). In the monoculture, ground cover rankings were: Barfort > Aries, Mara > Quartet. The only significant difference was between Barfort and Quartet, with Barfort having a greater amount of ground cover than Quartet (P < 0.05, data not shown). At Lake City, spring ground cover rankings in the binary mixture were: Barfort > Mara > Aries > Quartet; Barfort and Mara exhibited a statistically greater amount of ground cover than Quartet (P < 0.05, data not shown). In the monoculture, ground cover rankings were: Mara > Barfort > Quartet > Aries. Barfort and Mara had significantly greater (P < 0.05) ground cover than the other two cultivars. Quartet exhibited a significantly greater amount of ground cover than Aries (data not shown). At Chatham, fall ground cover rankings of the grass monoculture were: Barfort, Quartet > Aries > Mara, but ground cover of these cultivars was not significantly different in the spring (P < 0.05, data not shown). In the binary mixture at KBS, there was no significant difference in the amount of fall ground cover among cultivars (P < 0.05, data not shown). In the monoculture rankings were: Mara > Quartet > Barfort > Aries. Mara exhibited significantly more ground cover in the fall than Barfort and Aries (P < 0.05, data not shown). For the binary mixture study in Lake City, ground cover rankings were as follows: Mara > Aries > Quartet > Barfort; significant differences occurred only between Mara and Barfort (P < 0.05, data not shown). Rankings for the monoculture at Lake City were Mara > Barfort > Quartet > Aries. Mara exhibited significantly greater (P < 0.05) ground cover than both Quartet and Aries. Barfort also exhibited a significantly greater amount of ground cover than Aries (data not shown). At Chatham, the grass monoculture rankings were all equal and there was no significant difference in spring ground cover between cultivars (P < 0.05, data not shown). Spring ground cover loosely paralleled winter injury results. In both the binary mixture and monoculture, cultivars which had more winter injury had less spring ground cover as well, but analyses of fall ground cover was not as predictable. As cultivars in both the binary mixture and monoculture recovered from the winter injury, the difference in cultivar means became smaller so more winter injury did not necessarily mean less fall ground cover. Yield Yield was determined, but only the first and total yields are discussed here. Five harvests were collected at KBS, three at Lake City, and four at Chatham. At KBS, the cultivar yields from the first cutting were not significantly different (P < 0.05) in either the binary mixture (Table 7) or the monoculture (Table 9). At Lake City, Aries yielded more than Quartet in the binary mixture (Table 7), while Mara yielded more Aries and Quartet in the monoculture (Table 9). Aries and Mara had greater first yields than the tetraploid cultivars at Chatham in the monoculture (Table 9). Table 8 shows first harvest botanical composition results (biomass separated into grass, clover, and weed components) for the binary mixture at KBS and LC. Table 7. First yield (DM tons/acre) for cultivars grown as binary mixtures
Table 8. Botanical separation data at the first harvest (DM tons/acre) for binary mixtures at W. K. Kellogg Biological Station (KBS) at Hickory Corners and the Lake City Experiment Station (LC) at Lake City, MI. There is no yield data from the Upper Peninsula Experiment Station at Chatham because it was reseeded.
†Means in a column followed by a common letter are not significantly different within a location (P < 0.05). Table 9. First yield (DM tons/acre) for grass monoculture cultivars at W. K. Kellogg Biological Station (KBS) at Hickory Corners, the Lake City Experiment Station (LC) at Lake City, and the Upper Peninsula Experiment Station (UP) at Chatham, MI. First harvests occurred on 20 April 2002, 20 May 2002, and 4 June 2002, for KBS, LC, and UP, respectively.
No significant difference in total yield was observed at KBS for either the binary mixture (Table 10) or monoculture treatments (Table 12). At Lake City, Mara had a greater yield than Quartet in the binary mixture (Table 10). In the monoculture, Mara yielded more than both Quartet and Aries (Table 12). At Chatham, Mara had a significantly (P < 0.05) larger total yield than Aries and Barfort (Table 12). Table 11 shows total botanical composition results for the binary mixture at KBS and LC. At KBS, there was significantly (P < 0.05) more clover in mixtures of both Mara and Barfort compared to Aries and Quartet, indicating better compatibility of Mara and Barfort with ladino clover compared to Aries and Quartet. There were no differences in clover content between cultivars at Lake City, probably due to the low clover content overall in the study at Lake City. The overall low clover content may be due to winter injury of the clover component at Lake City. There was a greater amount of ladino clover in the binary mixture at KBS compared to Lake City. Table 10. Total yield (DM tons/acre) for cultivars grown as binary mixtures
Table 11. Total botanical separation data (DM tons/acre) for cultivars grown as binary mixtures at W. K. Kellogg Biological Station (KBS) at Hickory Corners and the Lake City Experiment Station (LC) at Lake City, MI. There is no yield data from the Upper Peninsula Experiment Station at Chatham because it was reseeded.
†Means in a column followed by a common letter are not significantly different within a location (P < 0.05). Table 12. Total yield (DM tons/acre) for grass monoculture cultivars at W. K. Kellogg Biological Station (KBS) at Hickory Corners, the Lake City Experiment Station (LC) at Lake City, and the Upper Peninsula Experiment Station (UP) at Chatham, MI. There were five harvests at KBS, three at LC, and four at UP for the establishment year.
Yields from the first harvest were closely related to winter injury. Research by Frame (6) reported that winter damaged PR had decreased first harvest yields, but the grass had largely recovered by the second harvest (6). In both the binary mixture and monoculture, cultivars with the least amount of winter injury typically had the highest yields. At Chatham, however, there was no significant difference between cultivars for either winter injury or spring ground cover, but the diploid cultivars had significantly (P < 0.05) higher first harvest yields compared to the tetraploid cultivars. Total yields, however, did not seem to follow a definite pattern. It was expected that cultivars with the most amount of winter injury would have lower total yields. At KBS, regardless of winter injury or ground cover, there were no significant differences in cultivar total yields. This was true in both the binary mixture and the monoculture. Total yield at Lake City seemed to depend on both winter injury and fall ground cover in both the binary mixture and the monoculture. There were significant differences in total yield at Chatham between monoculture treatments, even though there were no differences in winter injury, spring ground cover, or fall ground cover. Total yield probably did not follow the expected pattern (cultivars with the most winter injury yielding the least) because factors such as the amount of precipitation, amount of sunlight, and animal preference may also influence production. Implications Past research has shown that diploid cultivars have greater cold tolerance than tetraploid cultivars (5,13). Our results, however, showed Barfort (a tetraploid cultivar) was better adapted than the other cultivars to cold temperatures. This was true in both the laboratory and field experiments. The ability to germinate in cold temperatures was not indicative, however, of the cultivars’ performance in the field. Humphreys and Eagles (7) reported similar findings, where laboratory assessments of PR freezing tolerance did not always accurately predict survival under natural conditions (7). This research suggests that winter survival depends more on origin than ploidy level as both European cultivars, Mara and Barfort, performed well in the field experiments compared to the New Zealand cultivars, Aries and Quartet. Research by Cooper (3) also found a similar relationship between cold survival and temperature at site of origin in ryegrass populations (3). Diploid cultivars are also thought to produce more biomass than the tetraploid cultivars. At KBS, there were no differences in cultivar yield for either the first yield or the total yield. Only at Chatham did the first yield of monoculture grown diploid cultivars yield more than the tetraploids. This study indicates that perennial ryegrass can be grown throughout Michigan if cultivars are chosen carefully based upon variety trials where winter injury is evaluated for different cultivars. Producers should select a cultivar that originates from a location with a similar climate and evaluated in a variety trial within the region instead of selecting a cultivar based on ploidy level. This is important in Michigan because choosing an adapted cultivar, such as Barfort or Mara, will decrease the amount of winter injury and increase stand persistence. By choosing an adapted cultivar, a producer could grow perennial ryegrass and still have both high quality and high yields even in a northern region of Michigan. Although grasses recovered from any winter injury, first harvest yields can be decreased by a severe winter. For this reason, producers should rely on a more winter-hardy species such as tall fescue or orchardgrass for the first harvest. Acknowledgments The authors want to thank Tim Dietz and Chris Kapp for their time and aid in data collection. The authors also thank the staff at W. K. Kellogg Biological Station and the Lake City Experiment Station for their care and management of the cattle used for grazing. Literature Cited 1. Anonymous. 2000. Perennial ryegrass adaptation and management for Michigan. Mich. Hay Grazier Newsl. 6:1-3. 2. Casler, M. D., and Walgenbach, R. P. 1990. Ground cover potential of forage grass cultivars mixed with alfalfa at divergent locations. Crop Sci. 30:825-831. 3. Cooper, J. P. 1964. Climatic variation in forage grasses. I. Leaf development in climatic races of Lolium and Dactylis. J. App. Ecol. 1:45-61. 4. Connolly, V. 2001. Breeding improved cultivars of perennial ryegrass. Teagasc Res. Rep. No. 3495. 5. Dvorak, J., and Fowler, D. B. 1978. Cold hardiness potential of triticale and tetraploid rye. Crop Sci. 18:477-478. 6. Frame, J. 1989. Herbage productivity of a range of grass species under a silage cutting regime with high fertilizer nitrogen application. Grass Forage Sci. 44:267-276. 7. Humphreys, M. O., and Eagles, C. F. 1988. Assessment of perennial ryegrass (Lolium perenne L.) for breeding 1. Freezing tolerance. Euphytica 38:75-84. 8. Kunelius, H. T., and Castonguay, Y. 2003. Performance of perennial ryegrass in Atlantic Canada. Am. Forage Grassland Counc. Proc. Rep. 12:26. 9. Nassiri, M., and Elgersma, A. 2002. Effects of nitrogen on leaves, dry matter allocation and regrowth dynamics in Trifolium repens L. and Lolium perenne L. in pure and mixed swards. Plant Soil 246:107-121. 10. Novy, E. M., Casler, M. D., and Hill, R. R., Jr. 1995. Selection for persistence of tetraploid ryegrasses and festulolium in a mixture with perennial legumes. Crop Sci. 35:1046-1051. 11. Sjoseth, H. 1971. Studies on frost hardiness in diploid and autotetraploid red clover (Trifolium pratense) and winter rye (Secale cereals). Hereditas 43:679-682. 12. Solberg, E. T., Rognil, O. A., and Østrem, L. 1994. Potential for improving adaptation of Lolium perenne L. to continental climates in Norway. Euphytica 77:183-191. 13. Sugiyama, S. 1998. Differentiation in competitive ability and cold tolerance between diploid and tetraploid cultivars in Lolium perenne. Euphytica 103:55-59. 14. Tyler, B. F., Borrill, M., and Chorlton, D. 1978. Studies in Festuca x. Observations on germination and seedling cold tolerance in diploid Festuca pratensis and tetraploid F. pratensis var. appennina in relation to their altitude distribution. J. Appl. Ecol. 15:219-226. 15. Wade, L. J., Hammer, G. L., and Davey, M. A. 1993. Response of germination to temperature amongst diverse sorghum hybrids. Field Crops Res. 31:295-308. 16. Wilkins, P. W. 1991. Breeding perennial ryegrass for agriculture. Euphytica 52:201-214. 17. Williams, T. A., Evans, D. R., Rhodes, I., and Abberton, M. T. 2003. Long-term performance of white clover varieties grown with perennial ryegrass under rotation grazing by sheep with different nitrogen applications. J. Agric. Sci. 140:151-159. |
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