INTERACTIONS OF LIGHT AND TEMPERATURE ON THE GERMINATION OF PLANTAGO MARITIMA L.

Transcription

1 New Phytol. (1973) 72, INTERACTIONS OF LIGHT AND TEMPERATURE ON THE GERMINATION OF PLANTAGO MARITIMA L. BY SYLVIA M. ARNOLD* Department of Biology, University of York, and Department of Physiology and Environmental Studies, University of Nottingham {Received 5 October 1972) SUMMARY The percentage of seeds of Plantago 7naritima whieh germinated in the light increased linearly with temperature between 8 and 18 C. The response to fluctuating temperatures could be accounted for entirely by this linear relation. Germination was inhibited when seeds were kept in darkness during imbibition and percentage germination was shown to depend on the temperature of the seeds as well as the length of the dark period. Exposure to light at 25 C released the inhibition of some of the dark treated seeds but exposure to low temperature was ineffectual. Most of the seed came from Upper Teesdale but seed from Devon responded to germination tests in the same way. The germination of seeds of Kobresia simpliciuscula from Teesdale was also inhibited by darkness during imbibition. This type of response may promote the survival of seedlings in their natural environment. INTRODUCTION Plantago maritima occurs around most of the coastline of the British Isles and locally beside streams on the mountains of the Snowdon Range and the Highlands of Scotland (Clapham, Tutin and Warburg, 1962). It has been suggested that Plantago spread postglacially from the coast to the mountains and has subsequently been displaced from most intermediate areas (Gregor, 1938). P. maritima is found on the Pennines and seed for the present study came from plants collected from Upper Teesdale, Co. Durham, which is well known for the presence of species which are rare or unusually distributed (Pigott, 1956)- The work reported here forms part of an investigation into the effects of climate on germination and seedling growth of several plants characteristic of Teesdale. The interpretation of germination and seedling survival in the field depends on a detailed knowledge of the physiological responses of seeds to changing microclimatic conditions. Although the full complexity of field conditions cannot be reproduced in growth rooms, physical conditions such as temperature can be modulated to resemble diurnal changes in the field and the interaction of light and temperature can be examined. MATERIALS AND METHODS Seed used in germination experiments was harvested from plant material transplanted to Sutton Bonington from Widdybank Fell, Teesdale, during the winter of Seeds * Formerly S. M. Old. 583

2 584 SYLVIA M. ARNOLD were collected in the summers of 1970 and An additional supply of Plantago maritima seeds was obtained from Start Bay Devon. From harvest until use, seeds were stored dry at approximately 3 C. In all experiments, seeds were germinated on the surface of sieved (2.5 mm mesh) John Innes No. i compost contained in a temperature-controlled germination tank, in seed trays or in 76-mm pots. The compost was wetted to a water content of approximately 25% by weight and evaporation from the surface was prevented by a sealed cover of clear polythene. A cover of aluminium foil under black polythene was used to exclude light in some treatments. In other treatments, light was supplied for 18 hours per day from warm white fluorescent tubes giving 60 w/m^ visible radiation. Emergence of the radicle was taken as the criterion of germination in all experiments. (a) O O O O O Q O O Q i o Polystyrene ;.; [ Compost -- Woter I 1 3 mm aiuminium (b) Fig. I. Germination tank: (a) surface view; (b) end view. Germination tank A Jumo temperature control unit was used to produce sinusoidal oscillations of temperature in an aluminium tank containing of stirred water. The oscillations had a period of 24 hours and amplitude of + 10 or +20 C about a mean value set between o and 20 C. Three aluminium fins attached externally to one wall of the water bath (see Fig. i) were buried in a second tank containing John Innes compost. These fins were 560 mm long, 120 mm tall and 85 mm apart so that they enclosed two parallel channels of soil. To ensure that the temperature of the soil would be dictated by the temperature of the fins and not by the walls of the soil tank, the compost was separated from the walls by a 25-mm layer of expanded polystyrene and a clearance of 10 mm was left between the bottom of the flns and the base of the tank. The whole assembly was installed in a growth room. At appropriate distances from the water bath, the temperature of the compost was monitored with thermistor probes within 4 mm of the surface, attached to a Grant recorder. Fig. 2 shows how the amplitude of the oscillations decreased along the tank although the mean temperature at each station was similar. In germination experiments

3 Light and temperature on germination 585 the tank was divided lengthwise into eight 60 x go mm plots each sown with forty uniformly spaced seeds and the temperature at the centre of each plot was taken as a representative average value. RESULTS Plantago maritima Temperature response in the light Fig. 3 shows the effect of a constant temperature on germination in the light after 14 days. Highest germination of Plantago maritima was recorded at 18 C; and no germination was observed below 8 C even after 30 days. The relation between germination in the light G (%) and (constant) temperature T ( C) was linear over the range of temperatures studied (G = T). 12,00 Fig. 2. Variation in the amplitude of diurnal soil temperature oscillations with distance along the germination tank. These results were obtained from germination tests in the tank, but experiments in seed trays or pots in constant temperature growth rooms gave equivalent results. At 18, 14 and 9 C, the percentage of seeds germinating in 76-mm pots was 90, 55 and 25% respectively after 14 days, figures consistent with the relation between germination and constant temperature. The effect of oscillating temperatures on germination in the light was also investigated. In most experiments the temperature amplitude was set between +3.0 and ±4.5 as shown in Table i, but a range of amplitudes up to ±8.5 C was run at a mean temperature of 18 C. Fig. 3 shows germination percentage after 14 days plotted against mean temperature. Again, the relation between germination percentage and temperature was linear {G = '). The difference between the two regression lines was not

4 586 SYLVIA M. ARNOLD signifieant {P = 0.4). Over the lower temperature ranges the regression for oscillating temperatures is shifted to the right of that for constant temperatures. This difference can be explained on the basis of the 8 C threshold for germination. An oseillating temperature regime with a low mean temperature will include periods when temperatures fall below the threshold. A higher mean temperature will then be required to acbieve the percentage of germination recorded at a constant temperature. The temperature response of loor- 9 II Mean femperature ( C) Fig. 3. The relation between temperature and pereentage germination of Plantago maritima after 14 days in the hght. A, Fluctuating temperature (r 0.98***);, constant temperature (r = 0.95***) Mean temperature of darkness ( C) Fig. 4. Relation between the temperature during 7 days of imbibition in darkness and subse.- quent percentage germination of Plantago maritima after 14 days at 18 C in the light. A, Fluctuating temperatures;, constant temperatures.

5 Light and temperature on germination 587 freshly harvested seeds was not significantly different from the response for seeds which were stored dry for 12 months. Effect of imbibition in darkness germination in light Duration and temperature. Germination percentage was recorded for seeds sown in compost and kept in darkness for 7 days at all the temperatures mentioned above. No germination occurred in the dark at any temperature. When seeds had been kept in the dark for 7 days at different temperatures the covers were removed. A constant day and night temperature of 18 C was then maintained at a Mean ( C) ^ Amplitude ( C) Table i. Range of temperature amplitude and means produced in the gernmiation tank 7.0 ± ± ± ±3-O 9-5 ±3-o 10.o ±3-5 I I.O ± ± ±4.2 to ± Days of illumination Fig. 5. The relation between the duration of imbibition at 18 C 2-8 days in darkness and subsequent germination oi Plantago maritima maintained at 18 C in the light. Seeds collected from Devon ( ) and Teesdale ( ). radiation of 60 W/m'^. Fig. 4 shows percentage germination with cubic curves fitted to these results. The percentage of germination in the light decreased when the temperature of the dark period exceeded 6-8 C. Little further increase in germination was recorded when the temperature during imbibition was decreased below 6 C, nor did germination decrease when temperature in the dark exceeded about 16 C. The effect of fluctuating temperatures during dark imbibition was also examined using regimes already described

6 588 SYLVIA M. ARNOLD (Table i), and the mean temperature of the dark period plotted against germination percentage (Fig. 4). The response to fluctuating and constant temperature was similar. The two curves show a tendency to diverge between 7 and 10 C, probably because the fluctuating temperatures fell below 8 C for several hours in the day. The corresponding curve would therefore be shifted towards a higher mean temperature to produce an equivalent reduction in germination. In experiments to examine the eftect of imbibition for 7 days in darkness at fluctuating temperatures with a mean between 8 and 12 C, a proportion of the seeds was maintained at their dark temperature after their light-excluding covers were removed. Their germination after 14 days in the light was small and not significantly different from corresponding levels in Fig. 3. The effect of the duration of dark imbibition at 18 and 14 C on subsequent germination in the light at 18 C was examined. The response of germination to imbibition for 1-8 days at 18 C in the dark is shown in Fig. 5. Seeds collected from both Teesdale and loor LSD Days of iuumination Fig. 6. Percentage germination of Plantago maritima exposed to light after imbibition in darkness at 18 C for less than 6 days ( ) or more than 6 days ( ). from Devon were used in these experiments, and their response to varying lengths of dark treatment could not be distinguished. A dark period of 5 days or less had no significant effect on subsequent germination, whereas reduced germination was recorded in seeds subjected to a longer dark treatment. Values for germination are allocated to each of these categories in Fig. 6. After 6 days of illumination, germination of seeds which had imbibed in the dark for 6 days or more was significantly less than in seeds kept dark for less than 6 days including those which were continuously in the light. When the dark period at 18 C was extended to 21 days the reduction of germination in the light was maintained and only 15% germination was recorded. The response of germination at 18 C to imbibition in darkness for 4, 7 and 11 days at 14 C is shown in Fig. 7. A longer period of imbibition was required at 14 C than at 18 C to reduce subsequent germination to approximately 30%. There was no significant difference between percentage germination for a 4-day or a 7-day exposure. The percentage was 27.5 after 21 days in darkness at 14 C. A dark imbibition period of i week at

7 Light and temperature on germination 589 g C resulted in 80% germination oi Plantago maritima seeds when they were exposed to light at 18 C. Figs. 5, 6 and 7 record measurements on seed planted on the surfaee of riddled John Innes No. i compost contained in 76-mm pots, and Fig. 4 refers to seed planted in the germination tank filled with this same medium. From pot experiments, imbibition at 18, 14 and 9 C for 7 days resulted in 25, 55, and 80% germination respeetively after 14 days' illumination at 18 C. When these results are compared with the curve for constant temperature in Fig. 4 it ean be seen that although germination in pots after a 14 C dark treatment was more than in the tank, germination after the 18 and 9 C treatments lie within the expected limits. Deductions about threshold temperatures for maximum and minimum reduction in germination following 7 days of darkness are therefore substantiated / _ -I 50 c 40 o D 30 (U o O.Oi 0.05 LSD 10 f Days of illumination Fig. 7. Percentage gerinination of Plantago maritima exposed to light at i8 C after imbibition in darkness at 14 C for 4 days ( ), 7 days ( ) and 11 days ( ). Temperature extremes following dark imbibition. During several experiments on the duration of darkness whieh redueed germination of P. maritima in the light, growth room failures resulted in a large unintentional increase in temperature extending up to 24 hours. It was noted that these high temperatures tended to increase germination above the expected level. To investigate this response, the effect of one day at 25 C after 7 days of darkness at 18 C was determined. All seeds were given a dark treatment of 7 days at 18 C, and the seeds (all ungerminated) were then placed in the following regimes: (a) light at 25 C for i day followed by transfer to an 18 C, 18-hour day; (b) darkness at 25 C for I day followed by transfer to an 18" C, 18-hour day; (c) darkness at 25 C for I day followed by transfer to darkness at 18 C for i week, and then an 18 C, 18-hour day. Results obtained in treatments (a) and (b) are shown in Fig. 8. Germination recorded in treatments (b) and (c) was not significantly higher than would be expected from seeds simply pretreated with 7 days of darkness at 18 C (i.e %). When the 25 C treatment included exposure to light (treatment (a)) germination increased significantly, compared with germination after exposure to 25 C in the dark. Germination after the

8 59 SYLVIA M. ARNOLD 25 C treatment in the light was not significantly different from germination obtained when seeds were exposed to light immediately. The effect of darkness for 1-5 weeks at 3 C on seeds pretreated with 7 days of darkness at 18 C was investigated. Variation in the duration of the cold treatment had no significant effect on subsequent germination in the light at 18 C. A comparison of the average germination of all seeds exposed to cold treatments with those simply given 7 days at 18 C in the dark is shown in Fig. 9. Darkness at 3 C, following darkness for 7 days at 18 C had no additional effect on subsequent germination in the light. Effect of storage Maximum percentage of germination under the most favourable temperature-light regime used, namely 18 C with an 18-hour day, varied slightly with different experi Days of illumination at i8''c 24 Fig. 8. Percentage germination of Plantago ynaritima at 18 C in light after imbibition for 7 days at i8 C in darkness followed by: i day at 25 C in light ( ); or i day at 25 C in dark ( ). loor Days of iiluminat ion Fig. y. Percentage germination of Plantago maritima at 18 C in light after imbibition for 7 days at i8 C in darkness followed by: an extended period at 3 C ( ); or no cold treatment ( ).

9 Light and temperature on germination 591 ments. This variation was not consistent with a reduction in viability following extended dry storage. Table 2 shows that dry storage for i, 4, and 12 months had no significant effect on germination in the light at 8 and 18 C. The original experiment to determine the duration of imbibition in darkness at 18 C which would reduce germination in the light significantly was done on seeds after i month of dry storage following harvest (Fig. 4). This experiment was repeated using the same seed source but after an additional 3 months' storage. Results obtained were similar to those given in Fig. 4. Six days of darkness caused a highly significant (P<o.oi) reduction in germination, and this difference was apparent after 6 days of exposure to light- Table 2. The effect of storage on light germination in Plantago maritima Temperature ( C) 8 18 loor ±3.7 Storage (months) o± o o c E 0) o LSD O.OI Days of 16 lluminaf ion Fig. 10. Germination of Kobresia simplicitiscuta at i8 C in light after imbibition in darkness at 18 C for 3 days ( ), 6 days (O), 8 days ( ), 12 days (A); and immediate light ( ). Kobresia simpliciuscula Kobresia seed collected from Teesdale in the summer of 1970 did not germinate until the seeds had been stored dry for 8 months. Subsequently seeds needed light for high germination. Full germination was recorded at 18 C and an 18-hour day. The effect of imbibition in darkness at 18 C for 3, 6, 8, and 12 days is shown in Fig. 10. Darkness for 3 days did not reduce germination significantly below the level of controls (immediate light). Germination decreased when darkness was extended to 6 days, and fell even further when 8 days of darkness were imposed. Seeds kept in darkness for 12 days failed to germinate even after 28 days in light. Once seeds pretreated for 3 days of darkness were exposed to light, they germinated more rapidly than did seed exposed immediately to light. In the former case, seeds are in an imbibed state before exposure and

10 592 SYLVIA M. ARNOLD can respond to light by rapid germination. In the latter case imbibition must precede germination and consequently germination is slightly delayed. DISCUSSION There are many reports of seeds which germinate more easily when exposed to fluctuating rather than constant temperatures (e.g. Sayers and Ward, 1966; Toole and Toole, 1941); and Thompson (1970) has reported an essential requirement for fluctuating temperature in Lycopus europaeus. No stimulation of germination by fluctuating temperatures occurred in Plantago maritima. Over a range of mean temperatures from 18 to 8" C, diurnal soil temperature fluctuations with amplitudes from 17 to 5 C had no significant effect on the germination of P. marititna in the light. In the lower temperature ranges there was a tendency for germination at a given mean temperature to be lower in the fluctuating than in a constant temperature regime. This can be explained in terms of the threshold temperature for germination which was about 8 C. A similar response to fluctuating temperature was recorded when the effect of the temperature of imbibition in darkness was investigated. Overall, soil temperature fluctuations had no effect on the response of germination to mean temperature during imbibition in the dark. When this mean temperature approached threshold values for the imbibition reaction (6-8 C), the presence of soil temperature fluctuations tended to increase the mean temperature needed to achieve the response recorded at constant temperatures. Darkness consistently inhibited the germination of P. maritima. A light requirement for germination in this species was reported by Witte (1935). There are many examples where species whose seeds exhibit a need for light in constant temperatures lose this need when fluctuating temperature is used in germination tests (e.g. Andersen, 1947; Sifton, 1959). In the present study, however, none of the temperature regimes affected the light requirement for germination in P. maritima. This conclusion was not limited to plants from one geographical area. Seed collected from Co. Durham and Devon responded similarly to experimental conditions. Extended periods of dry storage did not materially affect germination responses to light and temperature. Although all the present results were from seeds germinating on the surface of John Innes No. i compost, filter paper floating on distilled water had been used as a germinating medium in preliminary experiments. The response of germination to a 7-day dark treatment at 18 C before exposure to light was not significantly different from that obtained using John Innes compost. When P. maritima seeds are exposed to darkness during imbibition subsequent germination in light may be inhibited. The extent of this inhibition depends both on the temperature and on the duration of imbibition. The level of inhibition increased as temperature rose from 8 to 16 C (Fig. 4). At 18 C, 6 days of darkness was sufficient to reduce germination in the light significantly, whereas at 14 C 11 days of darkness was required to produce a marked decrease in germination. The formation of a 'germinationdelaying factor' can be postulated for imbibed seed of P. maritima held at relatively warm temperatures in the dark and the concentration of this factor appears to be critical. It is formed more quickly as temperature in the dark is increased from 8 to 16 C and a reduction from 18 to 14 C in the dark temperature can be compensated for by an increase in the duration of darkness. Once the inhibitor is formed, its effectiveness is not significantly diminished by extended periods of cold (e.g. 3 C). A treatment of 25 C for

11 Light and temperature on germination 593 I day reduced the effectiveness of the inhibitor when the seeds were exposed to light during this treatment. Preliminary experiments with Kobresia simpliciuscula showed that a similar response to periods of dark imbibition occurs in this species (Fig. 10). Wesson and Wareing (1969) suggested a phenomenon similar to the inhibition reported here, which occurred in buried weed seeds, and they described this response to periods of dark imbibition as 'dark hardness'. Although it is not possible to postulate the nature of the germination-delaying factor from these experiments, the results describe time-temperature relationships which determine its formation and maintenance, and provide evidence for its occurrence in widely differing species. The formation of a germination-delaying factor in imbibed seeds buried in the soil may be widespread in species whose seeds require light for germination. Detailed interpretation of these responses in terms of the germination and seedling survival of Plantago maritima must await further work, but results suggest a mechanism which might stagger germination in natural environments. Seeds falling from plants in the late summer will germinate immediately given suitable external conditions. If seeds are temporarily buried during this time, the delaying factor might reduce germination if seeds later become exposed to light at a time of year when few seedlings would survive. The germination of seeds inhibited in this manner would await warming of the soil surface in spring when seedlings would have a much better chance of survival. The situation in Kobresia simpliciuscula would appear more complex since seeds require an extended period of after ripening before germination can occur. ACKNOWLEDGMENTS The work was undertaken while the author was working under a grant to Professor J. L. Monteith from the I.C.I. Teesdale Trust Fund. I would particularly like to thank Professor Monteith for advice and assistance throughout the study. Thanks are due to the University of Nottingham, and to Dr M. C. Lewis and the University of York for provision of working facilities; to Dr T. V. Callaghan and Miss G. E. Jones for assistance with computation ot results, to the former for use of his program for curvilinear regression, and finally to my husband for assistance with field work. REFERENCES ANDERSEN, A. M. (1947). The effect of alternating temperatures, light intensities and moistening agents of the substratum on the germination of freshly harvested seed of Oregon ryograss. Proc. Ass. off. Seed Analysts N. Am., 37, 152. CLAFHAM, A. R., TuTlN, T. G. & VVARiiriiG, K. F. (1962). Flora of the British Isles, 2nd edn. Camhridgc Uni\"ersity Press, l^ondon. GRHGOR, J. W. (1938). Experimental taxonomy. II. Initial population differentiation in Plantai;o maritima in Britain. New PliytoL, 37, 15. PiGOTT, C. D. (1956). The vegetation of Upper Teesdale in the North Pennines.J. Ecol., 44,!;45. SAYERS, R. L. & WARD, R. T. (1966). Germination responses in alpine species. Bot. Gti:::., SiFTON, H. 13. (1959). The germination of light-sensitive seeds of Typha latifolia L. Can. j. Bot'., 37, 719. THOMPSON, P. A. (1970). An analysis of tlie effect of alternating temperatures on germination of Lvcopus europaeus L. J. exp. Bot., 21, 808. ToOLE, E. H. & ToOLE, V. K. (1941). Progress of germination of seed of Digitaria as inhuenced by germination temperature and other factors, jf. agric. Res., 63, 65. WESSON, G. & WARI-ING, P. F. (1969). The role of light in the germination of naturally occurring populations of buried weed seeds, jf. e.xp. Bot., 20, 402. WiTTE, H. (1935). Some investigations regarding the germination of different species of the genus I'lantaeo Svensk. bot. Tidskr., 29, 532.

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