Effect of storage of achenes of Bidens gardneri Baker on light sensitivity during germination

Efeito do armazenamento dos aquênios de Bidens gardneri Baker na sensibilidade à luz durante a germinação

Effect of storage of achenes of Bidens gardneri Baker on light sensitivity during germination

REGINA M. SASSAKI¹, LILIAN B.P. ZAIDAN¹ and G.M. FELIPPE¹

(recebido em 10/06/98; aceito em 13/11/98)

ABSTRACT - (Effect of storage of achenes of Bidens gardneri Baker on light sensitivity during germination). Bidens gardneri is a very common herbaceous species in the cerrados of the state of São Paulo, whose seeds become light sensitive at 25°C only. Achenes of this species were stored in refrigerator at 4°C and in cerrado soil and in forest soil. The field experiments were carried out in the cerrado at the Reserva Biológica e Estação Experimental de Moji Guaçu, in Moji Guaçu and in the forest of the Instituto de Botânica, in São Paulo, Brazil. Achenes of B. gardneri vary in size and achenes from 7 to 12 mm long were used. Achenes stored for up to 6 months at 4°C showed light sensitivity; after 9 months storage, the difference in germination between light and darkness had disappeared for the smallest and the largest achenes used. Seeds of B. gardneri germinated during the period of storage in soil; the number of germinated seeds increased over the storage time, while the number of intact achenes decreased for the same period, no matter if the experiment was being carried out in the cerrado or in the forest. Therefore, the achenes germinated in soil in darkness. Light sensitivity was lost in intact achenes that had been stored in soil for three months.

RESUMO - (Efeito do armazenamento dos aquênios de Bidens gardneri Baker na sensibilidade à luz durante a germinação). Bidens gardneri é uma planta herbácea muito freqüente nos cerrados do estado de São Paulo, cujas sementes só germinam em luz na temperatura de 25°C. Aquênios foram armazenados a 4°C e enterrados em solo de cerrado e solo de mata. Os experimentos de campo foram realizados no cerrado na Reserva Biológica e Estação Experimental de Moji Guaçu, em Moji Guaçu e na floresta do Instituto de Botânica de São Paulo, em São Paulo, Brasil. Os aquênios de B. gardneri variam em tamanho e aquênios de 7 a 12 mm de comprimento foram usados. Os aquênios armazenados até 6 meses a 4°C apresentaram sensibilidade à luz, que foi perdida depois de 9 meses de armazenamento, tanto para os menores como para os maiores aquênios usados nos experimentos. Aquênios da espécie germinaram durante o período de armazenamento no solo; o número de aquênios germinados aumentou com o tempo de armazenamento e o número de aquênios intactos diminuiu com o período de armazenamento; estes resultados foram independentes do fato de o experimento ter sido feito em cerrado ou na floresta. Isto mostra que aquênios germinaram no escuro quando armazenados em solo. Os aquênios intactos perderam a sensibilidade à luz após armazenamento no solo por três meses.

Key words - Bidens gardneri, longevity, light sensitivity

Introduction

The natural cover of about 25% of the land area of Brazil was savannah vegetation given the generic name of cerrado. The cerrado vegetation is being quickly destroyed for the use of the land for short term agriculture. As the cerrado soil is characteristically poor in mineral nutrients (Alvim et al. 1968) frequently the land, after the removal of the original vegetation, is then left to regenerate. Thus seed bank and its features, like longevity of seeds, are very important.

Dormancy can be a determining factor for seed longevity in soil. Two types of longevity can be distinguished, ecological and potential longevity. The ecological longevity refers to the duration of the dormancy in natural conditions, the interval between maturation and dispersal of the seeds and germination or death in the soil; the potential longevity (viability) is the maximum duration of capacity to germinate of the dormant seeds stored in optimal conditions (Bewley & Black 1985). The study of the longevity of seeds is very important when considering species that can form a seed bank.

Bidens gardneri (Asteraceae) is a common herb of the cerrados of the state of São Paulo, producing ornamental flowers and a great number of achenes. The species has been the subject of a few papers, relating to the nutrient conditions (Felippe & Dale 1990), flowering (Klein et al. 1992) and nutrient conditions and photoperiod (Klein et al. 1996). Felippe (1990) showed that the species presents light sensitivity when germination is studied at 25°C; with a long storage period in dry conditions at 4°C (up to 14 months) the achenes lose their light sensitivity and can easily germinate in dark conditions. But is this also true when the seeds are stored in moist soil? Felippe (1990) also showed that germination in light was high when long achenes were used and very low with short achenes. The objective of this paper was to try to find out if light dormancy is maintained when achenes of B. gardneri are artificially stored in the soil.

Material and methods

The achenes (seeds) of Bidens gardneri Baker were collected in 1996 in the cerrado of Itirapina, state of São Paulo, Brazil (22°51'S and 47°52'W). A capitulum of Bidens gardneri carries 39 achenes of different sizes (measured in 15 capitula). The distribution of achenes according to their lengths is shown in table 1, most being between 8 and 12 mm long. The relative mass is smaller the longer the achene as can be seen in the ratio mass/length shown in table 1. Changes in viability and light sensitivity during storage were followed in achenes of different sizes in length, four groups of achenes were used: 7 - 8 (1st group); 9 (2nd group); 10 (3rd group) and 11- 12 mm long (4th group).

The achenes were stored in three conditions:

1. in closed glass bottles, at 4°C (± 2C) in darkness (refrigerator);

2. mixed with soil and buried in the cerrado at the Reserva Biológica e Estação Experimental de Moji Guaçu, in Moji Guaçu, state of São Paulo, Brazil (22°18'S and 47°11'W). A description of this area and its vegetation is given in Mantovani & Martins (1993);

3. mixed with soil and buried in the forest at the Instituto de Botânica, in São Paulo, state of São Paulo, Brazil (23°39'S and 46°37'W). Description of the area and its vegetation can be seen in Struffaldi-De Vuono (1985).

The chemical and physical analyses of the soils used and the rainfall and the temperature of the two regions are presented in Simabukuro et al. (1998) and Sassaki et al. (1999).

The achenes mixed with soil were placed in small bags (10 x 15 cm) made of very fine black nylon gauze ("Sombrite"). In general 500 achenes were used in each bag. The bags were buried at 10 cm deep in the cerrado or in the forest at the beginning of April 1996. The bags were removed from the soil 1, 3, 6 and 9 months afterwards. At removal, the number of intact and germinated achenes was checked and the moisture content (%) of the intact achenes was determined (three replicates of 20 achenes):

MC % = [(FM - DM).FM^-1)] . 100,

where FM is fresh mass and DM is dry mass obtained after drying the achenes for 48 hours at 80°C (ISTA 1976).

Germination of intact achenes was analysed after 1 and 3 months storage in the soil but with longer storage periods there were not enough intact achenes for the tests. Germination was carried out at 25°C in 5 cm diameter Petri dishes on moistened filter paper in darkness or in white fluorescent light at 437µW.cm^-2.s^-1 at Petri dish level (Ruggiero & Zaidan 1997); three replicates of 30 seeds were used. Darkness was obtained by enclosing the Petri dishes in two black plastic bags and seeds were examined under a green safe light (Labouriau & Costa 1976).

Imbibition was determined using five replicates of 20 achenes, according to: % imbibition = [(Mt - Mi) . Mi^-1)] . 100, where Mt is the mass after imbibition and Mi the initial mass (before imbibition started).

Germination and moisture content were also measured for achenes stored at 4°C for 1, 3, 6, 9 and 12 months. Germination rate (t) was calculated as the period of time in days when 50% germination was reached (Labouriau 1967).

The germination percentage was subjected to angular transformation and different treatments were compared by analysis of variance (Snedecor 1962).

Results and Discussion

As shown previously by Felippe (1990) the newly collected achenes of B. gardneri showed light sensitivity during germination (figure 1). The results of germination for achenes stored at 4°C up to 12 months are also shown in figure 1. The achenes showed light sensitivity, germination being statistically higher in light than in darkness up to 6 months storage at 4°C. By 9 months storage in this condition the difference in germination between light and darkness had disappeared for the smallest and the largest achenes (7 and 8 mm long; 11 and 12mm long). Thus it seems that light sensitivity in B. gardneri decreases with storage for a long period at 4°C. This is clearly observed when achenes of all sizes were used (figure 2). There was a positive correlation between period of storage and germination in darkness. Bidens odorata (Corkidi et al. 1991) and Bidens pilosa (Forsyth & Brown 1982) show light sensitivity and also showed an increase in germination in dark conditions with long period of storage. Both germination in light and in darkness were affected by the size of the achenes (figure 1). There was a very high positive correlation between size and germination for achenes stored for 12 months (r = 0.9972 for germination in light and r = 0.9758 for germination in darkness). The size of the achenes and the period of storage at 4°C also had an effect on the rate of germination as measured by the medium time of germination, t (table 2). In general, the higher the size of the achene and the longer the period of storage the faster was the germination (lower values of t). Germination in light conditions was higher in the longer achenes of Bidens odorata than in the shorter ones (Corkidi et al. 1991).

Figure 1. Germination in light (open symbols) and in darkness (closed symbols) of Bidens gardneri achenes (7-8; 9; 10; 11-12 mm long) stored at 4°C for 0, 1, 3, 6, 9 and 12 months. The results of analysis of variance are shown by letters. Small letters (a, b) compare germination in light and darkness in each period of storage and for each size of achene. Capital letters (A,B,C) compare germination in light for each period of storage. Capital letters with apostrophe (A', B', C') compare germination in darkness for each period of storage. Results shown up to day 20 (values were the same afterwards, up to day 30).

Figure 2. Correlation between period of storage at 4°C and maximum germination in darkness (closed symbols) or in light (open symbols) in achenes of Bidens gardneri.

When stored in soil some seeds of B. gardneri germinated during the period of storage; the number of germinated seeds increased and number of intact achenes decreased with period of storage in the soil. Thus, there was a negative correlation between number of intact achenes collected from the soil and period of storage and a positive one between germinated achenes in the soil and period of storage, as can be seen in figure 3. When storage was longer than 3 months very few intact achenes were recovered from burial in the soil; because of this it was impossible to carry germination experiments. Achenes of Bidens polylepis behave in a different way when stored in soil: only 2% of them germinated while buried in soil in a non heated greenhouse (Baskin et al. 1995). The moisture content (%) was much higher in the intact achenes removed from the soil than for achenes stored at 4°C (table 3); this is not surprising as the achenes over the storage period in the soil imbibed water from the rain (see rain data in Simabukuro et al. 1998 and Sassaki et al. 1999).

Figure 3. Correlation between period of storage in the soil in the cerrado of Moji Guaçu and in the forest of the Instituto de Botânica and number of intact achenes (open symbols) or germinated achenes of Bidens gardneri (closed symbols) when the containers were removed from the soil.

The germination of the intact achenes stored in the cerrado for 1 and 3 months is shown in figure 4. After 1 month storage in the soil the higher germination values were observed in the longer achenes; this was also shown for achenes stored for 1 month at 4°C but the difference in germination values in darkness and in light was reduced, compared with achenes stored for 1 month at 4°C. The statistical analysis (small letters) showed differences between germination in light against darkness. In light there was no difference in germination values when the length of the achenes was considered. In darkness germination was higher in the 11-12 mm long achenes and the lowest germination value was detected in the 7-8mm long achenes (see A' and C'). By 3 months storage in the soil the achenes had lost their sensitivity to light. Germination of the achenes stored in forest for 1 and 3 months is shown in figure 4. With achenes stored for 1 month germination was higher in light for the longer achenes and the lowest values were found in the 7-8 mm long achenes; by the third month the achenes had lost their sensitivity to light. Thus, light sensitivity is lost after storage for 9 to 12 months at 4°C while with storage in the soil (cerrado or forest) this happens with storage for one month. Thus, soil storage radically changes the behaviour of the seeds of this species to their sensitivity to light.

Figure 4. Germination in light (open symbols) and in darkness (closed symbols) of Bidens gardneri achenes (7-8; 9; 10; 11-12 mm long) stored in cerrado soil in the cerrado of Moji Guaçu for 1 and 3 months and in forest soil in the forest of the Instituto de Botânica for 1 and 3 months. The results of analysis of variance are shown by letters. Small letters (a, b) compare germination in light and darkness in each period of storage and for each size of achene. Capital letters (A,B,C) compare germination in light for each period of storage. Capital letters with apostrophe (A', B', C') compare germination in darkness for each period of storage.

Felippe (1980) has shown for Rumex obtusifolius that for seeds which only germinate in the light at 25°C, alternating temperatures could have the same promotive effect as light. Bidens gardneri, another light sensitive seed, may behave in the same way as Rumex. Ortolani & Pinto (1972) have shown that in Ribeirão Preto, state of São Paulo (21°10'S and 47°50'W) in summer the median air temperature was 26°C. In our experiments, in April, the air max temperature was around 30°C and the mininum around 20°C for Moji Guaçu and Instituto de Botânica (Simabukuro et al. 1998, Sassaki et al. 1999). According to Ortolani & Pinto (1972) alternating temperatures occurred in the soil at 5 cm deep (22°C at 8.00 h and 35°C at 16.00 h) and at 10 cm deep (23°C at 8.00 h and 34°C at 16.00 h, but the temperatures were nearly constant at 20 cm deep (25°C and 27°C). As the seeds in this work were buried at 10 cm deep, some alternating temperatures occurred. Although the achenes were stored in the soil in a relatively dry April, some rain occurred soon after the beginning of storage (see Simabukuro et al. 1998, Sassaki et al. 1999) and the alternating temperatures could have affected the imbibed seeds. This could explain the fact that seeds germinated when buried, but does not explain the lack of light sensitivity of the intact achenes removed from the soil.

The soil of the cerrado of Moji Guaçu is very porous presenting a high content of sand (Simabukuro et al. 1998) and thus the water from the rain can easily reach the buried seeds. The soil of the forest of the Instituto de Botânica contains about half the sand of the cerrado soil but a higher level of clay and in this case can retain the water (Sassaki et al. 1999). The rainfall occurring by the beginning of the storage could have been enough for the seeds to imbibe, as they take up water very fast: in an experiment to check imbibition the percentage of imbibition of the achenes of B. gardneri (stored for 12 months at 4°C or newly collected) was around 40% after 5 hours from the beginning of the test (data not shown). The differences in germination (see figure 1) between longer and shorter achenes is not a result of imbibition rates as imbibition was the same for the achenes of different sizes used here (data not shown). Thus, the differences in germination can not be explained by differences in relation to water permeability.

However, shorter achenes have higher mass/length ratio than longer ones (table 1). This could be due to a thicker coat which is permeable to water, although the thickness of the achenes coat was not measured in the present work. One possibility is that this thicker seed coat could present some difficulty for the radicle to go through thus reducing germination. This seems to be the case as Felippe (1990) has shown that coat removal promoted germination of the shorter achenes of Bidens gardneri. Short achenes of Bidens bipinnata also show dormancy (Dakshini & Aggarwall 1974). Buried short achenes of Bidens gardneri may have slowly degraded their coats to allow seed germination only later in the season compared to the faster germination of the long achenes. Thus the requirements of the short achenes for germination will be responsible for the formation of a transitory seed bank. In contrast, the long achenes that will germinate soon after dispersal do not contribute to a seed bank.

Acknowledgements - Are due to CNPq for financial support and research grants for the authors. Thanks are also due to Josimara N. Rondon for technical assistance. Thanks are also due to Prof. John E. Dale (University of Edinburgh) for correcting our mistakes in English and his comments on the manuscript.

References

ALVIM, P.T. SANTANA, C.J. & MIRANDA, E.R. 1968. Avaliação da fertilidade em alguns solos de cerrado em Brasília por meio de ensaios de microparcelas. Ci. & Cult. 20:613-619.

BASKIN, C.C. BASKIN, J.M. & CHESTER, E.W. 1995. Role of temperature in the germination ecology of the summer annual Bidens polylepis Blake (Asteraceae). Bull. Torrey bot. Cl. 122:275-281.

BEWLEY, J.D. & BLACK, M. 1985. Seed physiology of development and germination. Plenum, New York.

CORKIDI, L., RINCON, E. & VÁSQUEZ-YANES, C. 1991. Effects of light and temperature on germination of heteromorphic achenes of Bidens odorata (Asteraceae). Can. J. Bot. 69:574-579.

DAKSHINI, K.M.M. & AGGARWALL, S.K. 1974. Intracapitular cypsele dimorphism and dormancy in Bidens bipinnata. Biol. Plant. 16:469-471.

FELIPPE, G.M. 1980. Germination of the light-sensitive seeds of Cucumis anguria and Rumex obtusifolius: effects of temperature. New Phytol. 84:439-448.

FELIPPE, G.M. 1990. Germinação de Bidens gardneri Baker, uma planta anual dos cerrados. Hoehnea 17:7-11.

FELIPPE, G.M. & DALE, J.E. 1990. The effects of phosphorus supply on growth of plants from the Brazilian cerrado: experiments with seedlings of the annual weed Bidens gardneri Baker (Compositae) and the tree Qualea grandiflora (Mart.) (Vochysiaceae). Oecologia 82:81-86.

FORSYTH, C. & BROWN, N.A.C. 1982. Germination of the dimorphic fruits of Bidens pilosa L. New Phytol. 90:151-164.

ISTA (INTERNATIONAL SEED TESTING ASSOCIATION) 1976. International rules for seed testing; rules 1976. Seed Sci. & Technol. 41:1-180.

KLEIN, A. L., ZAIDAN, L.B.P. & FELIPPE, G.M. 1992. Flowering and heterophylly in Bidens gardneri Baker. Revta brasil. Bot. 15:139-144.

KLEIN,A.L., ZAIDAN, L.B.P. & FELIPPE, G.M. 1996. Interaction between soil and photoperiod on development of Bidens gardneri Baker (Asteraceae), a herbaceous species from the Brazilian cerrado. Revta brasil. Bot. 19:1-5.

LABOURIAU, L.F.G. 1967. Sobre a fisiologia da germinação das sementes de Vicia graminea SM. Universidade Rural do Brasil, Rio de Janeiro.

LABOURIAU, L.F.G. & COSTA, J.A.F. 1976. Objetivos e instalações básicas de um laboratório de fisiologia vegetal. Academia Brasileira de Ciências, Rio de Janeiro.

MANTOVANI, W. & MARTINS, F.R. 1993. Florística do cerrado na Reserva Biológica de Moji Guaçu, SP. Acta bot. brasil. 7:33-60.

ORTOLANI, A.A. & PINTO, H.S. 1972. Temperatura do solo. In Elementos de pedologia (A.A. Moniz, ed.). Editora Polígono, São Paulo, p.59.

RUGGIERO, P.G.C. & ZAIDAN, L.B.P. 1997. Estudos de desenvolvimento de Viguiera robusta Gardn., uma Asteraceae do cerrado. Revta brasil. Bot. 20:1-9.

SASSAKI, R.M., RONDON, J.N., ZAIDAN, L.B.P. & FELIPPE, G.M. 1999. Longevity of seeds from legume tree species artificially stored in cerrado and forest soils. Hoehnea 26: (in press).

SIMABUKURO, E.A., ESTEVES, L.M. & FELIPPE, G.M. 1998. Analysis of a fern spore bank in Southeast Brazil. Hoehnea 25:45-57.

SNEDECOR, G.W. 1962. Statistical methods. Iowa State University Press, Iowa.

STRUFFALDI-DE VUONO, Y. 1985. Fitossociologia do estrato arbóreo da floresta do Instituto de Botânica (São Paulo, SP). Tese de doutorado. Universidade de São Paulo, São Paulo.

• BASKIN, C.C. BASKIN, J.M. & CHESTER, E.W. 1995. Role of temperature in the germination ecology of the summer annual Bidens polylepis Blake (Asteraceae). Bull. Torrey bot. Cl. 122:275-281.
• BEWLEY, J.D. & BLACK, M. 1985. Seed physiology of development and germination. Plenum, New York.
• CORKIDI, L., RINCON, E. & VÁSQUEZ-YANES, C. 1991. Effects of light and temperature on germination of heteromorphic achenes of Bidens odorata (Asteraceae). Can. J. Bot. 69:574-579.
• DAKSHINI, K.M.M. & AGGARWALL, S.K. 1974. Intracapitular cypsele dimorphism and dormancy in Bidens bipinnata Biol. Plant. 16:469-471.
• FELIPPE, G.M. 1980. Germination of the light-sensitive seeds of Cucumis anguria and Rumex obtusifolius: effects of temperature. New Phytol. 84:439-448.
• FELIPPE, G.M. 1990. Germinaçăo de Bidens gardneri Baker, uma planta anual dos cerrados. Hoehnea 17:7-11.
• FELIPPE, G.M. & DALE, J.E. 1990. The effects of phosphorus supply on growth of plants from the Brazilian cerrado: experiments with seedlings of the annual weed Bidens gardneri Baker (Compositae) and the tree Qualea grandiflora (Mart.) (Vochysiaceae). Oecologia 82:81-86.
• FORSYTH, C. & BROWN, N.A.C. 1982. Germination of the dimorphic fruits of Bidens pilosa L. New Phytol. 90:151-164.
• KLEIN, A. L., ZAIDAN, L.B.P. & FELIPPE, G.M. 1992. Flowering and heterophylly in Bidens gardneri Baker. Revta brasil. Bot. 15:139-144.
• KLEIN,A.L., ZAIDAN, L.B.P. & FELIPPE, G.M. 1996. Interaction between soil and photoperiod on development of Bidens gardneri Baker (Asteraceae), a herbaceous species from the Brazilian cerrado. Revta brasil. Bot. 19:1-5.
• LABOURIAU, L.F.G. 1967. Sobre a fisiologia da germinaçăo das sementes de Vicia graminea SM. Universidade Rural do Brasil, Rio de Janeiro.
• LABOURIAU, L.F.G. & COSTA, J.A.F. 1976. Objetivos e instalaçőes básicas de um laboratório de fisiologia vegetal. Academia Brasileira de Cięncias, Rio de Janeiro.
• MANTOVANI, W. & MARTINS, F.R. 1993. Florística do cerrado na Reserva Biológica de Moji Guaçu, SP. Acta bot. brasil. 7:33-60.
• ORTOLANI, A.A. & PINTO, H.S. 1972. Temperatura do solo. In Elementos de pedologia (A.A. Moniz, ed.). Editora Polígono, Săo Paulo, p.59.
• RUGGIERO, P.G.C. & ZAIDAN, L.B.P. 1997. Estudos de desenvolvimento de Viguiera robusta Gardn., uma Asteraceae do cerrado. Revta brasil. Bot. 20:1-9.
• SASSAKI, R.M., RONDON, J.N., ZAIDAN, L.B.P. & FELIPPE, G.M. 1999. Longevity of seeds from legume tree species artificially stored in cerrado and forest soils. Hoehnea 26: (in press).
• SIMABUKURO, E.A., ESTEVES, L.M. & FELIPPE, G.M. 1998. Analysis of a fern spore bank in Southeast Brazil. Hoehnea 25:45-57.
• SNEDECOR, G.W. 1962. Statistical methods. Iowa State University Press, Iowa.
• STRUFFALDI-DE VUONO, Y. 1985. Fitossociologia do estrato arbóreo da floresta do Instituto de Botânica (Săo Paulo, SP). Tese de doutorado. Universidade de Săo Paulo, Săo Paulo.

1. Seção de Fisiologia e Bioquímica de Plantas, Instituto de Botânica, Caixa Postal 4005, 01016-970 São Paulo, SP, Brazil. Research Fellow, CNPq.

Seção de Fisiologia e Bioquímica de Plantas, Instituto de Botânica, Caixa Postal 4005, 01016-970 São Paulo, SP, Brazil. Research Fellow, CNPq.

Publication Dates

• Publication in this collection
  19 Aug 1999
• Date of issue
  Apr 1999

History

• Accepted
  13 Nov 1998
• Received
  10 June 1998