- Email: [email protected]
Imbibition phases and germination response of Mimosa bimucronata (Fabaceae: Mimosoideae) to water submersion
Aquatic Botany 91 (2009) 105–109
Contents lists available at ScienceDirect
Aquatic Botany journal homepage: www.elsevier.com/locate/aquabot
Imbibition phases and germination response of Mimosa bimucronata (Fabaceae: Mimosoideae) to water submersion De´bora Kestring a,*, Jeferson Klein a, Luciana Cristina Candido Ribeiro de Menezes a, Marcelo Nogueira Rossi b a b
Unesp – Sa˜o Paulo State University, Departamento de Botaˆnica, IB, Botucatu 18618-000, SP, Brazil Universidade Federal de Sa˜o Paulo (Unifesp), Departamento de Cieˆncias Biolo´gicas, Diadema 09941-510, SP, Brazil
A R T I C L E I N F O
A B S T R A C T
Article history: Received 11 June 2008 Received in revised form 6 March 2009 Accepted 10 March 2009 Available online 17 March 2009
Mimosa bimucronata is a pioneering tree that occurs predominantly in moist lowlands, floodplains and on margins of rivers and lakes in Latin America. The effect of submergence on seed germination in M. bimucronata was firstly studied. Patterns of water absorption by M. bimucronata seeds were investigated thereafter to assess the imbibition phases of scarified and unscarified seeds. The germination percentage was significantly higher in scarified than in unscarified seeds, and the velocity of seed germination also increased considerably in scarified seeds. Submergence duration did not significantly affect germination percentages of scarified and unscarified seeds. Therefore, seed viability after submersion suggests that M. bimucronata may display hydrochorous dispersal and also that seeds are able to germinate successfully in areas with frequent seasonal flooding. With respect to imbibition phases, phase II was very short or even absent for scarified and unscarified seeds; therefore, a plateau, where water absorption by seeds is established, was not observed. Finally, we verified that the passage from phase I to III was very tenuous and took a long time in seeds without scarification. ß 2009 Elsevier B.V. All rights reserved.
Keywords: Germination Mimosa bimucronata Flooding Imbibition phases Dispersal
1. Introduction 1.1. The role of flooding on seed germination The frequency, extension, and duration of floods are determinant variables in flood-prone habitats (Guo et al., 1998; Bunn and Arthington, 2002; Scarano et al., 2003; Gomes et al., 2006), which may affect vegetation composition and structure in several systems (Nilsson, 1999; Ho¨lzel and Otte, 2004a; Vogt et al., 2007; Wittmann et al., 2007). For some plant species flooding may induce seed dormancy resulting in poor seed germination (Mollard et al., 2007). However, seeds of various other species come out of dormancy when submerged (e.g. Baskin et al., 1993, 1996, 2000) or simply need low oxygen pressure in order to germinate (Leck, 1996; Lorenzen et al., 2000). Plants can also take advantage of water to disperse seeds (hydrochorous dispersal) and establish their populations in suitable patches far away from maternal plants. Therefore, seed transport by river can be important for the establishment and maintenance of wetland vegetation, as long as seeds are deposited at sites safe for the germination, establishment
* Corresponding author. Tel.: +55 14 3811 6265; fax: +55 14 3815 3744. E-mail address: [email protected] (D. Kestring). 0304-3770/$ – see front matter ß 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.aquabot.2009.03.004
and development of plants (e.g. Andersson and Nilsson, 2002; Merritt and Wohl, 2006; Vogt et al., 2007). Although water regimes may significantly affect seed germination in swamp and riparian vegetation (Insausti et al., 1995; Brock and Rogers, 1998; Hroudova´ and Za´kravsky´, 2003; Warwick and Brock, 2003; Gomes et al., 2006), information about seed tolerance after inundation is rather scarce for many tropical tree species. Moreover, many pioneering plants living along riverbanks and in flood-prone areas are potentially important for habitat restoration. Consequently, the understanding of germination responses of their seeds after long periods of inundation is relevant. 1.2. Imbibition phases and seed germination It is known that seeds need to be hydrated to stimulate metabolism because dry mature seeds, in most cases, exhibit very low metabolic rates (Gallardo et al., 2001). Imbibition is the first step in the succession of events that ends with primary root emission (Bradford, 1995; Pereira et al., 2007). According to Bewley and Black (1994) water uptake by the seeds is triphasic. Phase I presents fast water uptake, followed by phase II, where water absorption by the seeds is established, reaching a plateau. Desiccation tolerance is verified in phases I and II, which actually defines the germination process (sensu stricto) (Bradford, 1990; Bewley and Black, 1994; Bewley, 1997; Gallardo et al., 2001). Phase
106
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109
III is known as a post-germination phase of water uptake, which is determined by primary root protrusion with an expressive increase in seed moisture, and only viable seeds can reach this phase (Bewley and Black, 1994; Manz et al., 2005; Pereira et al., 2007). As dormant seeds do not enter the phase III because they do not complete germination, the application of scarification methods allows the transition from germination (phases I and II) to postgermination (phase III) in dormant seeds otherwise (Manz et al., 2005). Therefore, scarified and non-scarified seeds usually present different temporal patterns of water uptake.
Cieˆncias Agra´rias (Unesp/Botucatu Campus) (228500 5200 S; 488250 4600 W; area ffi 6700 m2) and the other population is situated near the Rubia˜o Ju´nior District (228530 0700 S; 488290 2300 W; area ffi 10000 m2), where they grow along the margin of a small river and a lake, respectively. Populations are separated by a distance of 8794 m (straight line). Most plants selected for this study were situated on the edge of populations, and 15 and 10 plants were randomly selected from each population, respectively. 2.2. Assessment of different submersion durations on seed germination
1.3. Study species and goals Mimosa bimucronata (DC.) Kuntze (Fabaceae: Mimosoideae) is a pioneering tree endemic to Brazil, Paraguay, Argentina, and Uruguay (Burkart, 1959). This species can reach a height of 10 m and occurs predominantly in moist lowlands, floodplains and on margins of rivers and lakes, where a few individuals can form dense aggregations (Carvalho, 1994; Lorenzi, 2000). M. bimucronata is cited as an important agent for restoration of degraded areas (Lorenzi, 2000), even though this plant is also an important weed when dense populations grow in pasture areas (Lorenzi, 2000), because their large thorny branches can hamper the access of cattle to water and fresh grass. In extreme conditions, because of shading, only a few grasses can grow below the canopy of these plants, thus causing a loss of grazing area. M. bimucronata produces craspedium fruits among which Tomaz et al. (2007) observed an average of 6.39 seeds per fruit. The production of mature M. bimucronata fruits takes place throughout most of the year (from March to December), but ripe fruits will fall off the trees from August to December (M.N. Rossi, personal communication). Therefore, for those M. bimucronata plants living in floodplains, most seeds are submerged during the rainy season (from December to March). For plants living along riverbanks and lakefronts, many seeds will undergo immediate inundation, because several fruits drop onto the water surface, and in rare situations lower branches of trees can be totally submerged. Despite the ecological and economic importance of M. bimucronata, it is not known whether, or to what extent, germination of submerged seeds is affected. Therefore, the first goal of the present study was to investigate the extent to which submergence affects germination of M. bimucronata seeds. While most M. bimucronata seeds are not dormant, the use of scarification (mechanical or chemical) enhances seed germination performance because these seeds have a hard coat, which is a barrier to water uptake (Ferreira et al., 1992; Ribas et al., 1996; Fowler and Carpanezzi, 1998; Tomaz et al., 2007). Under adverse germination conditions, seed scarification is an advantage because germination and seedling emergence are improved and synchronized, which have valuable implications for plants like M. bimucronata, with both agronomic (when it is a weed) and restoration importance. However, there are no studies aimed at determining the temporal pattern of water uptake by M. bimucronata seeds, while the extent to which scarification influences water imbibition process also remains unknown. Thus, the second objective of this study was to determine the imbibition phases of M. bimucronata seeds, comparing water absorption by scarified and unscarified seeds.
Mature fruits were randomly collected from each selected plant on May 24 and 25, and on June 1 and 2, 2006, and then transported in white paper bags to the laboratory (25 8C under 12-h light). Immediately after the last collection, fruits from both populations were mixed and randomly placed in six cylindrical plastic containers (1000 ml). Each container had about 500 fruits, and 500 ml of sterilized water was added to five of the six containers, leaving all fruits submerged. The container without water was the control group. Firstly, all recipients were left uncovered, and secondly, fruits were submerged instead of seeds to simulate natural conditions. Mature fruits of M. bimucronata do not open to release seeds when they fall off the tree. Therefore, when seeds reach the soil or water surface, they are still enclosed within the fruit exocarp. Germination tests were carried out on days 0 (control group), 1, 2, 15, 30, and 60 after submersion. For each germination test, 200 seeds were manually removed from fruits and separated into two groups of 100 seeds. In one group, sulfuric acid (98%) was applied to all seeds for 10 min (chemical scarification), and seeds from the other group were used as control. Although scarification can enhance germination on M. bimucronata seeds, the application of sulfuric acid on submerged seeds may reduce seed viability by damaging the embryo because the rigidity of the seed coat is expected to decrease after long periods of submergence. To investigate this question, in our submergence experiment we compared seed germination capacity between scarified and unscarified seeds. All seeds were placed in transparent plastic boxes (Gerbox— 11 cm 11 cm 4 cm) lined with two soaked filter papers (10 ml of distilled water) and kept for 21 days in an incubator under 18 h of white light at temperatures alternating between 20 and 30 8C (the light period corresponded to the highest temperature; Tomaz et al., 2007). All treatments (submersion durations in days) were carried out with five replicates of 20 seeds each (each Gerbox corresponded to one replicate). The germination percentage of each replicate was calculated daily for 21 days, with germination defined as the emission of 2 mm of primary root. To compare mean germination percentages among water treatments and between seeds with or without scarification in the end of the germination process, a two-way analysis of variance (Zar, 1999) was run (6 2 factorial design). To attain normality, germination percentage data were transformed as arcsin Hproportion; however, mean values computed from the raw data are presented in Section 3. The cumulative percentage of germinated seeds (mean values) over a 21-day period was graphically compared between scarified and unscarified seeds.
2. Methods
2.3. Assessment of water absorption by seeds
2.1. Plant population sites
For determination of water absorption by seeds, 1000 mature fruits of M. bimucronata were randomly collected from plants on May 29 and on June 6, 2006. Fruits were taken to the laboratory and 200 viable seeds were extracted manually from fruits. All seeds classified as unviable (dark colored seeds with irregular develop-
Mature fruits were collected from two M. bimucronata populations located in the city of Botucatu, state of Sa˜o Paulo, Brazil. One of the populations is located at the Faculdade de
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109
107
Fig. 1. Cumulative percentage of germinated seeds over a 21-day period for scarified and unscarified seeds.
ment) were discarded. In order to compare imbibition phases between scarified and unscarified seeds, 100 seeds underwent chemical scarification with sulfuric acid (98%) for 10 min (Fowler and Carpanezzi, 1998; Tomaz et al., 2007), and the remaining 100 seeds were not scarified. Four replications with 25 seeds each were used for each treatment, and each Gerbox container (see below) represented one replicate. Seeds were put into transparent plastic boxes (Gerbox— 11 cm 11 cm 4 cm) lined with two soaked filter papers (10 ml of distilled water). The Gerbox boxes were kept in an incubator under 18 h of white light in temperatures alternating between 20 and 30 8C, and the light period corresponded to the highest temperature (Tomaz et al., 2007). All seeds were dried with a paper towel at rigorous 1-h intervals and then weighed on an analytical scale, recording the number of germinated seeds in each treatment. After weighing, seeds were re-added into their respective Gerbox and then returned to the germination chamber. This process was interrupted when seeds reached a germination rate of at least 50%. To determine whether scarified and unscarified M. bimucronata seeds followed the three usual phases of water imbibition, linear and non-linear (exponential) adjustments to mean values of seed weights were made to obtain the best fit for the data. 3. Results
considerably on scarified seeds. For example, after the first week of evaluation, mean germination percentages on scarified and unscarified seeds were 81.2% and 14.7%, respectively (Fig. 1). Therefore, it was verified that scarified seeds germinated rapidly, reaching then a plateau with low variation in germination percentages, and a gradual increase in cumulative germination percentages over the 21-day period was observed for unscarified seeds (Fig. 1). For determination of the water imbibition phases, it was clearly verified that phase I was more accelerated on scarified seeds than on seeds without scarification (Fig. 2); therefore, water uptake occurred faster on scarified than on unscarified seeds (Fig. 2). However, it is interesting to note that phase II was very short or even absent for both seed groups because a plateau, where water absorption is established, was not verified in Fig. 2. The change from phase I to III on scarified and unscarified seeds occurred 11 and 22 h after the start of water absorption, respectively, and these phase changes were very tenuous on seeds without scarification (Fig. 2). 4. Discussion In the current study, we observed that submergence duration did not significantly affect the germination capacity of M. bimucronata seeds; therefore, seeds remained viable even after
Results of ANOVA showed that mean germination percentages among submersion durations (days) were not statistically significant (Table 1). However, as expected, the germination percentage was significantly different between scarified and unscarified seeds (Table 1). Interaction between submersion and scarification was not significant (Table 1). Scarification increased significantly the germination percentages, reaching the mean value of 92% in the end of the germination period (Fig. 1), while unscarified seeds showed mean germination percentage of 56.5% (Fig. 1). The velocity of seed germination also increased Table 1 Results of analysis of variance (two-way ANOVA) comparing germination percentage among submersion durations (days), between seeds with and without scarification, and considering interaction between submersion and scarification. Effect
DF
SS
F
P
Submersion duration (days) Scarification Submersion duration (days) scarification Error
5 1 5 48
0.14 3.16 0.13 1.32
0.98 114.66 0.97
0.438 <0.001 0.444
Percentage values were arcsin (Hproportion) transformed prior to analysis.
Fig. 2. Cumulative mean weights of seeds with and without scarification (imbibition phases). Seeds probably changed directly from phase I to III, and phase I on seeds without scarification was very tenuous. The change from phase I to III on seeds with and without scarification occurred after 11 and 22 h, respectively. Straight lines were not shown for phase III in both cases because unexplained variances were not found (r2 = 1.0).
108
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109
being submerged for 60 days. This finding corroborates with other studies where water submergence did not affect the viability and/ or germinability of seeds (e.g. Ho¨lzel and Otte, 2004b; Geissler and Gzik, 2008). The viability after submersion suggests that M. bimucronata may present hydrochorous dispersal, a very important finding given that water may favor adequate seed dispersal for those plants living along riverbanks (e.g. Andersson and Nilsson, 2002; Merritt and Wohl, 2006; Vogt et al., 2007). Our results show that despite the lack of seed dispersion, M. bimucronata seeds may be able to germinate successfully even when plants are located in areas associated with lakes and rivers where seasonal flooding frequently occurs. While dissecting submerged fruits for the germination experiments, low amounts of water were verified inside fruits submerged for 48 h, keeping seeds relatively dry. Therefore, it is possible that seeds gained protection by fruit exocarp during the first 2 days of submersion. However, after 48 h of submersion, all seeds within fruits were totally soaked. Many Mimosa species have tough seed coats, which are impermeable to water (Orozco-Almanza et al., 2003). Water content in seeds with impermeable seed coats has important implications for germination, because impermeable coats prevent germination until environmental conditions promote water absorption by seeds followed by germination (Cervantes et al., 1996; Orozco-Almanza et al., 2003). Our results suggest that M. bimucronata seeds have a hard seed coat because the germination experiment was more successful with scarified seeds, which germinated rapidly, in higher percentages and with less variability than unscarified seeds (Fig. 1). However, when seeds were not scarified, germination percentages were also relatively high, indicating that embryo dormancy mechanisms are not present for this species. As the result from interaction between days of submersion and scarification was not statistically significant (Table 1), the application of sulfuric acid on submerged seeds did not reduce seed viability as expected. It is important to state that in a pilot experiment, we also submerged fruits for 120 days, but under this condition only a few viable seeds could be extracted from fruits. For this reason, we decided to exclude this treatment from the definitive experiments. However, we believe that germination may occur over longer flooding periods than those observed herein even though a decrease in seed viability may be significant. The imbibition experiments showed that seeds without scarification presented a smooth and long phase I, which clearly supports our previous conclusion that M. bimucronata seeds have hard coats. It is interesting to note that phase II was not verified for both scarified and unscarified seeds. Therefore, it is possible that this phase is not present or it is very short on M. bimucronata seeds. This result is important because theory and some experiments suggest that water absorption occurs according to a triphasic pattern (e.g. Bewley and Black, 1994; Pereira et al., 2007). However, such triphasic pattern was not clearly observed here, and it is possible that this does not necessarily occur for many other plant species, especially for those living under disturbed conditions, as do many Mimosa species (Orozco-Almanza et al., 2003). As water absorption is usually a limiting factor for seeds with hard coats, either the absence or the very short phase II could be an adaptive strategy, which would confer an increased germination velocity, thereby reducing competition and unfavorable effects during seedling establishment. Such strategy might also be related to the pioneering nature of M. bimucronata plants, which would confer a successful colonization of degraded areas. Nevertheless, the life history strategies and the physiological mechanisms of seed germination of many Mimosa species remain unknown. Finally, the results of the present study encouraged us to aim future experiments at laboratory and field conditions, with longer submergence durations, in order to understand the physiological
basis of M. bimucronata seed germination and also to determine to what extent its germination capacity is affected under natural conditions. Acknowledgements We thank Prof. James Welsh for assistance with the English version of the manuscript. D. Kestring and J. Klein thank CNPq (Conselho Nacional de Desenvolvimento Cientı´fico e Tecnolo´gico) and Capes (Coordenac¸a˜o de Aperfeic¸oamento de Pessoal de Nı´vel Superior), respectively. L.C.C.R. Menezes (05/57999-3) and M.N. Rossi (04/06737-6) are particularly grateful to Fapesp (Fundac¸a˜o de Amparo a` Pesquisa do Estado de Sa˜o Paulo) for financial support. References Andersson, E., Nilsson, C., 2002. Temporal variation in the drift of plant litter and propagules in a small boreal river. Freshwater Biol. 47, 1674–1684. Baskin, C.C., Chester, E.W., Baskin, J.M., 1993. Germination ecology of Leptochloa panicoides, a summer annual grass of seasonally dewatered mudflats. Acta Oecol. 14, 693–704. Baskin, C.C., Chester, E.W., Baskin, J.M., 1996. Effect of flooding on annual dormancy cycles in buried seeds of two wetland Carex species. Wetlands 16, 84–88. Baskin, C.C., Baskin, J.M., Chester, E.W., 2000. Effect of flooding on the annual dormancy cycle and on germination of seeds of the summer annual Schoenoplectus purshianus (Cyperaceae). Aquat. Bot. 67, 109–116. Bewley, J.D., 1997. Seed germination and dormancy. Plant Cell 9, 1055–1066. Bewley, J.D., Black, M., 1994. Seeds: Physiology of Development and Germination. Plenum Press, New York. Bradford, K.J., 1990. A water relation analysis of seed germination rates. Plant Physiol. 94, 840–849. Bradford, K.J., 1995. Water relations in seed germination. In: Kigel, J., Galili, G. (Eds.), Seed Development and Germination. Marcel Dekker Inc., New York, pp. 351–398. Brock, M.A., Rogers, K.H., 1998. The regeneration potential of the seed bank of an ephemeral floodplain in South Africa. Aquat. Bot. 61, 123–135. Bunn, S.E., Arthington, A.H., 2002. Basic principles and ecological consequences of altered flow regimes for aquatic biodiversity. Environ. Manage. 30, 492–507. Burkart, A., 1959. Leguminosae. In: Parodi, L.R. (Ed.), Enciclopedia Argentina de Agricultura y Jardineria. ACME, Buenos Aires, Argentina, pp. 443–512. Carvalho, P.E.R., 1994. Espe´cies Florestais Brasileiras: Recomendac¸o˜es Silviculturais, Potencialidades e Uso da Madeira. Embrapa-CNPF, Colombo, PR. Cervantes, V., Carabias, J., Va´zquez-Yanes, C., 1996. Seed germination of woody legumes from deciduous tropical forest of southern Mexico. Forest Ecol. Manage. 82, 171–184. Ferreira, A.G., Lipp-Joa˜o, K.H., Heuser, E.D., 1992. Efeitos de escarificac¸a˜o sobre a germinac¸a˜o e do ph no crescimento de Acacia bonariensis Gill e Mimosa bimucronata (D.C.) O.K. Rev. Bras. Fisiol. Veg. 4, 63–65. Fowler, J.A.P., Carpanezzi, A.A., 1998. Tecnologia de sementes de Marica´ Mimosa bimucronata (DC) O. Ktze. Bol. Pesq. Flor. 36, 47–56. Gallardo, K., Job, C., Groot, S.P.C., Puype, M., Demol, H., Vandekerckhove, J., Job, D., 2001. Proteomic analysis of Arabidopsis seed germination and priming. Plant Physiol. 126, 835–848. Geissler, K., Gzik, A., 2008. The impact of flooding and drought on seeds of Cnidium dubium, Gratiola officinalis, and Juncus atratus, three endangered perennial river corridor plants of Central European lowlands. Aquat. Bot. 89, 283–291. Gomes, P.B., Va´lio, I.F.M., Martins, F.R., 2006. Germination of Geonoma brevispatha (Arecaceae) in laboratory and its relation to the palm spatial distribution in a swamp forest. Aquat. Bot. 85, 16–20. Guo, Y., Shelton, M.G., Lockhart, B.R., 1998. Effects of flood duration and season on germination of black, cherrybark, northern red, and water oak acorns. New Forest 15, 69–76. Ho¨lzel, N., Otte, A., 2004a. Ecological significance of seed germination characteristics in flood-meadow species. Flora 199, 12–24. Ho¨lzel, N., Otte, A., 2004b. Inter-annual variation in the soil seed bank of floodmeadows over two years with different flooding patterns. Plant Ecol. 174, 279– 291. Hroudova´, Z., Za´kravsky´, P., 2003. Germination responses of diploid Butomus umbellatus to light, temperature and flooding. Flora 198, 37–44. Insausti, P., Soriano, A., Sanchez, R.A., 1995. Effects of flood-influenced factors on seed germination of Ambrosia tenuifolia. Oecologia 103, 127–132. Leck, M.A., 1996. Germination of macrophytes from a Delaware River tidal freshwater wetland. Bull. Torrey Bot. Club 123, 48–67. Lorenzen, B., Brix, H., McKee, K.L., Mendelssohn, I.A., Miao, S.L., 2000. Seed germination of two Everglades species, Cladium jamaicense and Typha domingensis. Aquat. Bot. 66, 169–180. Lorenzi, H., 2000. Plantas Daninhas do Brasil: Terrestres, Aqua´ticas, Parasitas e To´xicas. Instituto Plantarum, Nova Odessa, SP. Manz, B., Mu¨ller, K., Kucera, B., Volke, F., Leubner-Metzger, G., 2005. Water uptake and distribution in germinating tobacco seeds investigated in vivo by nuclear magnetic resonance imaging. Plant Physiol. 138, 1538–1551.
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109 Merritt, D.M., Wohl, E.E., 2006. Plant dispersal along rivers fragmented by dams. River Res. Appl. 22, 1–26. Mollard, F.P.O., Insausti, P., Sanchez, R.A., 2007. Flooding induces secondary dormancy in Setaria parviflora seeds. Seed Sci. Res. 17, 55–62. Nilsson, C., 1999. Rivers and streams. In: Rydin, H. (Ed.), Acta Phytogeogr. Suec., vol. 84, pp. 135–148. Orozco-Almanza, M.S., Le´on-Garcı´a, L.P., Grether, R., Garcı´a-Moya, E., 2003. Germination of four species of the genus Mimosa (leguminosae) in a semi-arid zone of Central Mexico. J. Arid Environ. 55, 75–92. Pereira, M.D., Dias, D.C.F.S., Dias, L.A.S., Arau´jo, E.F., 2007. Hydration of carrot seeds in relation to osmotic potential of solution and conditioning method. Rev. Bras. Sem. 29, 144–150. Ribas, L.L.F., Fossati, L.C., Nogueira, A.C., 1996. Superac¸a˜o de dormeˆncia de sementes de Mimosa bimucronata (D.C.) O. Kuntze (Marica´). Rev. Bras. Sem. 18, 98–101.
109
Scarano, F.R., Pereira, T.S., Roˆc¸as, G., 2003. Seed germination during floatation and seedling growth of Carapa guianensis, a tree from flood-prone forest of the Amazon. Plant Ecol. 168, 291–296. Tomaz, C.A., Kestring, D., Rossi, M.N., 2007. Effects of the seed predator Acanthoscelides schrankiae on viability of its host plant Mimosa bimucronata. Biol. Res. 40, 281–290. Vogt, K., Rasran, L., Jensen, K., 2007. Seed deposition in drift lines: opportunity or hazard for species establishment? Aquat. Bot. 86, 385–392. Warwick, N.W.M., Brock, M.A., 2003. Plant reproduction in temporary wetlands: the effects of seasonal timing, depth, and duration of flooding. Aquat. Bot. 77, 153– 167. Wittmann, A.O., Piedade, M.T.F., Parolin, P., Wittmann, F., 2007. Germination in four low-va´rzea tree species on Central Amazonia. Aquat. Bot. 86, 197–203. Zar, J.H., 1999. Biostatistical Analysis. Prentice Hall, Upper Saddle River, NJ.
Contents lists available at ScienceDirect
Aquatic Botany journal homepage: www.elsevier.com/locate/aquabot
Imbibition phases and germination response of Mimosa bimucronata (Fabaceae: Mimosoideae) to water submersion De´bora Kestring a,*, Jeferson Klein a, Luciana Cristina Candido Ribeiro de Menezes a, Marcelo Nogueira Rossi b a b
Unesp – Sa˜o Paulo State University, Departamento de Botaˆnica, IB, Botucatu 18618-000, SP, Brazil Universidade Federal de Sa˜o Paulo (Unifesp), Departamento de Cieˆncias Biolo´gicas, Diadema 09941-510, SP, Brazil
A R T I C L E I N F O
A B S T R A C T
Article history: Received 11 June 2008 Received in revised form 6 March 2009 Accepted 10 March 2009 Available online 17 March 2009
Mimosa bimucronata is a pioneering tree that occurs predominantly in moist lowlands, floodplains and on margins of rivers and lakes in Latin America. The effect of submergence on seed germination in M. bimucronata was firstly studied. Patterns of water absorption by M. bimucronata seeds were investigated thereafter to assess the imbibition phases of scarified and unscarified seeds. The germination percentage was significantly higher in scarified than in unscarified seeds, and the velocity of seed germination also increased considerably in scarified seeds. Submergence duration did not significantly affect germination percentages of scarified and unscarified seeds. Therefore, seed viability after submersion suggests that M. bimucronata may display hydrochorous dispersal and also that seeds are able to germinate successfully in areas with frequent seasonal flooding. With respect to imbibition phases, phase II was very short or even absent for scarified and unscarified seeds; therefore, a plateau, where water absorption by seeds is established, was not observed. Finally, we verified that the passage from phase I to III was very tenuous and took a long time in seeds without scarification. ß 2009 Elsevier B.V. All rights reserved.
Keywords: Germination Mimosa bimucronata Flooding Imbibition phases Dispersal
1. Introduction 1.1. The role of flooding on seed germination The frequency, extension, and duration of floods are determinant variables in flood-prone habitats (Guo et al., 1998; Bunn and Arthington, 2002; Scarano et al., 2003; Gomes et al., 2006), which may affect vegetation composition and structure in several systems (Nilsson, 1999; Ho¨lzel and Otte, 2004a; Vogt et al., 2007; Wittmann et al., 2007). For some plant species flooding may induce seed dormancy resulting in poor seed germination (Mollard et al., 2007). However, seeds of various other species come out of dormancy when submerged (e.g. Baskin et al., 1993, 1996, 2000) or simply need low oxygen pressure in order to germinate (Leck, 1996; Lorenzen et al., 2000). Plants can also take advantage of water to disperse seeds (hydrochorous dispersal) and establish their populations in suitable patches far away from maternal plants. Therefore, seed transport by river can be important for the establishment and maintenance of wetland vegetation, as long as seeds are deposited at sites safe for the germination, establishment
* Corresponding author. Tel.: +55 14 3811 6265; fax: +55 14 3815 3744. E-mail address: [email protected] (D. Kestring). 0304-3770/$ – see front matter ß 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.aquabot.2009.03.004
and development of plants (e.g. Andersson and Nilsson, 2002; Merritt and Wohl, 2006; Vogt et al., 2007). Although water regimes may significantly affect seed germination in swamp and riparian vegetation (Insausti et al., 1995; Brock and Rogers, 1998; Hroudova´ and Za´kravsky´, 2003; Warwick and Brock, 2003; Gomes et al., 2006), information about seed tolerance after inundation is rather scarce for many tropical tree species. Moreover, many pioneering plants living along riverbanks and in flood-prone areas are potentially important for habitat restoration. Consequently, the understanding of germination responses of their seeds after long periods of inundation is relevant. 1.2. Imbibition phases and seed germination It is known that seeds need to be hydrated to stimulate metabolism because dry mature seeds, in most cases, exhibit very low metabolic rates (Gallardo et al., 2001). Imbibition is the first step in the succession of events that ends with primary root emission (Bradford, 1995; Pereira et al., 2007). According to Bewley and Black (1994) water uptake by the seeds is triphasic. Phase I presents fast water uptake, followed by phase II, where water absorption by the seeds is established, reaching a plateau. Desiccation tolerance is verified in phases I and II, which actually defines the germination process (sensu stricto) (Bradford, 1990; Bewley and Black, 1994; Bewley, 1997; Gallardo et al., 2001). Phase
106
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109
III is known as a post-germination phase of water uptake, which is determined by primary root protrusion with an expressive increase in seed moisture, and only viable seeds can reach this phase (Bewley and Black, 1994; Manz et al., 2005; Pereira et al., 2007). As dormant seeds do not enter the phase III because they do not complete germination, the application of scarification methods allows the transition from germination (phases I and II) to postgermination (phase III) in dormant seeds otherwise (Manz et al., 2005). Therefore, scarified and non-scarified seeds usually present different temporal patterns of water uptake.
Cieˆncias Agra´rias (Unesp/Botucatu Campus) (228500 5200 S; 488250 4600 W; area ffi 6700 m2) and the other population is situated near the Rubia˜o Ju´nior District (228530 0700 S; 488290 2300 W; area ffi 10000 m2), where they grow along the margin of a small river and a lake, respectively. Populations are separated by a distance of 8794 m (straight line). Most plants selected for this study were situated on the edge of populations, and 15 and 10 plants were randomly selected from each population, respectively. 2.2. Assessment of different submersion durations on seed germination
1.3. Study species and goals Mimosa bimucronata (DC.) Kuntze (Fabaceae: Mimosoideae) is a pioneering tree endemic to Brazil, Paraguay, Argentina, and Uruguay (Burkart, 1959). This species can reach a height of 10 m and occurs predominantly in moist lowlands, floodplains and on margins of rivers and lakes, where a few individuals can form dense aggregations (Carvalho, 1994; Lorenzi, 2000). M. bimucronata is cited as an important agent for restoration of degraded areas (Lorenzi, 2000), even though this plant is also an important weed when dense populations grow in pasture areas (Lorenzi, 2000), because their large thorny branches can hamper the access of cattle to water and fresh grass. In extreme conditions, because of shading, only a few grasses can grow below the canopy of these plants, thus causing a loss of grazing area. M. bimucronata produces craspedium fruits among which Tomaz et al. (2007) observed an average of 6.39 seeds per fruit. The production of mature M. bimucronata fruits takes place throughout most of the year (from March to December), but ripe fruits will fall off the trees from August to December (M.N. Rossi, personal communication). Therefore, for those M. bimucronata plants living in floodplains, most seeds are submerged during the rainy season (from December to March). For plants living along riverbanks and lakefronts, many seeds will undergo immediate inundation, because several fruits drop onto the water surface, and in rare situations lower branches of trees can be totally submerged. Despite the ecological and economic importance of M. bimucronata, it is not known whether, or to what extent, germination of submerged seeds is affected. Therefore, the first goal of the present study was to investigate the extent to which submergence affects germination of M. bimucronata seeds. While most M. bimucronata seeds are not dormant, the use of scarification (mechanical or chemical) enhances seed germination performance because these seeds have a hard coat, which is a barrier to water uptake (Ferreira et al., 1992; Ribas et al., 1996; Fowler and Carpanezzi, 1998; Tomaz et al., 2007). Under adverse germination conditions, seed scarification is an advantage because germination and seedling emergence are improved and synchronized, which have valuable implications for plants like M. bimucronata, with both agronomic (when it is a weed) and restoration importance. However, there are no studies aimed at determining the temporal pattern of water uptake by M. bimucronata seeds, while the extent to which scarification influences water imbibition process also remains unknown. Thus, the second objective of this study was to determine the imbibition phases of M. bimucronata seeds, comparing water absorption by scarified and unscarified seeds.
Mature fruits were randomly collected from each selected plant on May 24 and 25, and on June 1 and 2, 2006, and then transported in white paper bags to the laboratory (25 8C under 12-h light). Immediately after the last collection, fruits from both populations were mixed and randomly placed in six cylindrical plastic containers (1000 ml). Each container had about 500 fruits, and 500 ml of sterilized water was added to five of the six containers, leaving all fruits submerged. The container without water was the control group. Firstly, all recipients were left uncovered, and secondly, fruits were submerged instead of seeds to simulate natural conditions. Mature fruits of M. bimucronata do not open to release seeds when they fall off the tree. Therefore, when seeds reach the soil or water surface, they are still enclosed within the fruit exocarp. Germination tests were carried out on days 0 (control group), 1, 2, 15, 30, and 60 after submersion. For each germination test, 200 seeds were manually removed from fruits and separated into two groups of 100 seeds. In one group, sulfuric acid (98%) was applied to all seeds for 10 min (chemical scarification), and seeds from the other group were used as control. Although scarification can enhance germination on M. bimucronata seeds, the application of sulfuric acid on submerged seeds may reduce seed viability by damaging the embryo because the rigidity of the seed coat is expected to decrease after long periods of submergence. To investigate this question, in our submergence experiment we compared seed germination capacity between scarified and unscarified seeds. All seeds were placed in transparent plastic boxes (Gerbox— 11 cm 11 cm 4 cm) lined with two soaked filter papers (10 ml of distilled water) and kept for 21 days in an incubator under 18 h of white light at temperatures alternating between 20 and 30 8C (the light period corresponded to the highest temperature; Tomaz et al., 2007). All treatments (submersion durations in days) were carried out with five replicates of 20 seeds each (each Gerbox corresponded to one replicate). The germination percentage of each replicate was calculated daily for 21 days, with germination defined as the emission of 2 mm of primary root. To compare mean germination percentages among water treatments and between seeds with or without scarification in the end of the germination process, a two-way analysis of variance (Zar, 1999) was run (6 2 factorial design). To attain normality, germination percentage data were transformed as arcsin Hproportion; however, mean values computed from the raw data are presented in Section 3. The cumulative percentage of germinated seeds (mean values) over a 21-day period was graphically compared between scarified and unscarified seeds.
2. Methods
2.3. Assessment of water absorption by seeds
2.1. Plant population sites
For determination of water absorption by seeds, 1000 mature fruits of M. bimucronata were randomly collected from plants on May 29 and on June 6, 2006. Fruits were taken to the laboratory and 200 viable seeds were extracted manually from fruits. All seeds classified as unviable (dark colored seeds with irregular develop-
Mature fruits were collected from two M. bimucronata populations located in the city of Botucatu, state of Sa˜o Paulo, Brazil. One of the populations is located at the Faculdade de
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109
107
Fig. 1. Cumulative percentage of germinated seeds over a 21-day period for scarified and unscarified seeds.
ment) were discarded. In order to compare imbibition phases between scarified and unscarified seeds, 100 seeds underwent chemical scarification with sulfuric acid (98%) for 10 min (Fowler and Carpanezzi, 1998; Tomaz et al., 2007), and the remaining 100 seeds were not scarified. Four replications with 25 seeds each were used for each treatment, and each Gerbox container (see below) represented one replicate. Seeds were put into transparent plastic boxes (Gerbox— 11 cm 11 cm 4 cm) lined with two soaked filter papers (10 ml of distilled water). The Gerbox boxes were kept in an incubator under 18 h of white light in temperatures alternating between 20 and 30 8C, and the light period corresponded to the highest temperature (Tomaz et al., 2007). All seeds were dried with a paper towel at rigorous 1-h intervals and then weighed on an analytical scale, recording the number of germinated seeds in each treatment. After weighing, seeds were re-added into their respective Gerbox and then returned to the germination chamber. This process was interrupted when seeds reached a germination rate of at least 50%. To determine whether scarified and unscarified M. bimucronata seeds followed the three usual phases of water imbibition, linear and non-linear (exponential) adjustments to mean values of seed weights were made to obtain the best fit for the data. 3. Results
considerably on scarified seeds. For example, after the first week of evaluation, mean germination percentages on scarified and unscarified seeds were 81.2% and 14.7%, respectively (Fig. 1). Therefore, it was verified that scarified seeds germinated rapidly, reaching then a plateau with low variation in germination percentages, and a gradual increase in cumulative germination percentages over the 21-day period was observed for unscarified seeds (Fig. 1). For determination of the water imbibition phases, it was clearly verified that phase I was more accelerated on scarified seeds than on seeds without scarification (Fig. 2); therefore, water uptake occurred faster on scarified than on unscarified seeds (Fig. 2). However, it is interesting to note that phase II was very short or even absent for both seed groups because a plateau, where water absorption is established, was not verified in Fig. 2. The change from phase I to III on scarified and unscarified seeds occurred 11 and 22 h after the start of water absorption, respectively, and these phase changes were very tenuous on seeds without scarification (Fig. 2). 4. Discussion In the current study, we observed that submergence duration did not significantly affect the germination capacity of M. bimucronata seeds; therefore, seeds remained viable even after
Results of ANOVA showed that mean germination percentages among submersion durations (days) were not statistically significant (Table 1). However, as expected, the germination percentage was significantly different between scarified and unscarified seeds (Table 1). Interaction between submersion and scarification was not significant (Table 1). Scarification increased significantly the germination percentages, reaching the mean value of 92% in the end of the germination period (Fig. 1), while unscarified seeds showed mean germination percentage of 56.5% (Fig. 1). The velocity of seed germination also increased Table 1 Results of analysis of variance (two-way ANOVA) comparing germination percentage among submersion durations (days), between seeds with and without scarification, and considering interaction between submersion and scarification. Effect
DF
SS
F
P
Submersion duration (days) Scarification Submersion duration (days) scarification Error
5 1 5 48
0.14 3.16 0.13 1.32
0.98 114.66 0.97
0.438 <0.001 0.444
Percentage values were arcsin (Hproportion) transformed prior to analysis.
Fig. 2. Cumulative mean weights of seeds with and without scarification (imbibition phases). Seeds probably changed directly from phase I to III, and phase I on seeds without scarification was very tenuous. The change from phase I to III on seeds with and without scarification occurred after 11 and 22 h, respectively. Straight lines were not shown for phase III in both cases because unexplained variances were not found (r2 = 1.0).
108
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109
being submerged for 60 days. This finding corroborates with other studies where water submergence did not affect the viability and/ or germinability of seeds (e.g. Ho¨lzel and Otte, 2004b; Geissler and Gzik, 2008). The viability after submersion suggests that M. bimucronata may present hydrochorous dispersal, a very important finding given that water may favor adequate seed dispersal for those plants living along riverbanks (e.g. Andersson and Nilsson, 2002; Merritt and Wohl, 2006; Vogt et al., 2007). Our results show that despite the lack of seed dispersion, M. bimucronata seeds may be able to germinate successfully even when plants are located in areas associated with lakes and rivers where seasonal flooding frequently occurs. While dissecting submerged fruits for the germination experiments, low amounts of water were verified inside fruits submerged for 48 h, keeping seeds relatively dry. Therefore, it is possible that seeds gained protection by fruit exocarp during the first 2 days of submersion. However, after 48 h of submersion, all seeds within fruits were totally soaked. Many Mimosa species have tough seed coats, which are impermeable to water (Orozco-Almanza et al., 2003). Water content in seeds with impermeable seed coats has important implications for germination, because impermeable coats prevent germination until environmental conditions promote water absorption by seeds followed by germination (Cervantes et al., 1996; Orozco-Almanza et al., 2003). Our results suggest that M. bimucronata seeds have a hard seed coat because the germination experiment was more successful with scarified seeds, which germinated rapidly, in higher percentages and with less variability than unscarified seeds (Fig. 1). However, when seeds were not scarified, germination percentages were also relatively high, indicating that embryo dormancy mechanisms are not present for this species. As the result from interaction between days of submersion and scarification was not statistically significant (Table 1), the application of sulfuric acid on submerged seeds did not reduce seed viability as expected. It is important to state that in a pilot experiment, we also submerged fruits for 120 days, but under this condition only a few viable seeds could be extracted from fruits. For this reason, we decided to exclude this treatment from the definitive experiments. However, we believe that germination may occur over longer flooding periods than those observed herein even though a decrease in seed viability may be significant. The imbibition experiments showed that seeds without scarification presented a smooth and long phase I, which clearly supports our previous conclusion that M. bimucronata seeds have hard coats. It is interesting to note that phase II was not verified for both scarified and unscarified seeds. Therefore, it is possible that this phase is not present or it is very short on M. bimucronata seeds. This result is important because theory and some experiments suggest that water absorption occurs according to a triphasic pattern (e.g. Bewley and Black, 1994; Pereira et al., 2007). However, such triphasic pattern was not clearly observed here, and it is possible that this does not necessarily occur for many other plant species, especially for those living under disturbed conditions, as do many Mimosa species (Orozco-Almanza et al., 2003). As water absorption is usually a limiting factor for seeds with hard coats, either the absence or the very short phase II could be an adaptive strategy, which would confer an increased germination velocity, thereby reducing competition and unfavorable effects during seedling establishment. Such strategy might also be related to the pioneering nature of M. bimucronata plants, which would confer a successful colonization of degraded areas. Nevertheless, the life history strategies and the physiological mechanisms of seed germination of many Mimosa species remain unknown. Finally, the results of the present study encouraged us to aim future experiments at laboratory and field conditions, with longer submergence durations, in order to understand the physiological
basis of M. bimucronata seed germination and also to determine to what extent its germination capacity is affected under natural conditions. Acknowledgements We thank Prof. James Welsh for assistance with the English version of the manuscript. D. Kestring and J. Klein thank CNPq (Conselho Nacional de Desenvolvimento Cientı´fico e Tecnolo´gico) and Capes (Coordenac¸a˜o de Aperfeic¸oamento de Pessoal de Nı´vel Superior), respectively. L.C.C.R. Menezes (05/57999-3) and M.N. Rossi (04/06737-6) are particularly grateful to Fapesp (Fundac¸a˜o de Amparo a` Pesquisa do Estado de Sa˜o Paulo) for financial support. References Andersson, E., Nilsson, C., 2002. Temporal variation in the drift of plant litter and propagules in a small boreal river. Freshwater Biol. 47, 1674–1684. Baskin, C.C., Chester, E.W., Baskin, J.M., 1993. Germination ecology of Leptochloa panicoides, a summer annual grass of seasonally dewatered mudflats. Acta Oecol. 14, 693–704. Baskin, C.C., Chester, E.W., Baskin, J.M., 1996. Effect of flooding on annual dormancy cycles in buried seeds of two wetland Carex species. Wetlands 16, 84–88. Baskin, C.C., Baskin, J.M., Chester, E.W., 2000. Effect of flooding on the annual dormancy cycle and on germination of seeds of the summer annual Schoenoplectus purshianus (Cyperaceae). Aquat. Bot. 67, 109–116. Bewley, J.D., 1997. Seed germination and dormancy. Plant Cell 9, 1055–1066. Bewley, J.D., Black, M., 1994. Seeds: Physiology of Development and Germination. Plenum Press, New York. Bradford, K.J., 1990. A water relation analysis of seed germination rates. Plant Physiol. 94, 840–849. Bradford, K.J., 1995. Water relations in seed germination. In: Kigel, J., Galili, G. (Eds.), Seed Development and Germination. Marcel Dekker Inc., New York, pp. 351–398. Brock, M.A., Rogers, K.H., 1998. The regeneration potential of the seed bank of an ephemeral floodplain in South Africa. Aquat. Bot. 61, 123–135. Bunn, S.E., Arthington, A.H., 2002. Basic principles and ecological consequences of altered flow regimes for aquatic biodiversity. Environ. Manage. 30, 492–507. Burkart, A., 1959. Leguminosae. In: Parodi, L.R. (Ed.), Enciclopedia Argentina de Agricultura y Jardineria. ACME, Buenos Aires, Argentina, pp. 443–512. Carvalho, P.E.R., 1994. Espe´cies Florestais Brasileiras: Recomendac¸o˜es Silviculturais, Potencialidades e Uso da Madeira. Embrapa-CNPF, Colombo, PR. Cervantes, V., Carabias, J., Va´zquez-Yanes, C., 1996. Seed germination of woody legumes from deciduous tropical forest of southern Mexico. Forest Ecol. Manage. 82, 171–184. Ferreira, A.G., Lipp-Joa˜o, K.H., Heuser, E.D., 1992. Efeitos de escarificac¸a˜o sobre a germinac¸a˜o e do ph no crescimento de Acacia bonariensis Gill e Mimosa bimucronata (D.C.) O.K. Rev. Bras. Fisiol. Veg. 4, 63–65. Fowler, J.A.P., Carpanezzi, A.A., 1998. Tecnologia de sementes de Marica´ Mimosa bimucronata (DC) O. Ktze. Bol. Pesq. Flor. 36, 47–56. Gallardo, K., Job, C., Groot, S.P.C., Puype, M., Demol, H., Vandekerckhove, J., Job, D., 2001. Proteomic analysis of Arabidopsis seed germination and priming. Plant Physiol. 126, 835–848. Geissler, K., Gzik, A., 2008. The impact of flooding and drought on seeds of Cnidium dubium, Gratiola officinalis, and Juncus atratus, three endangered perennial river corridor plants of Central European lowlands. Aquat. Bot. 89, 283–291. Gomes, P.B., Va´lio, I.F.M., Martins, F.R., 2006. Germination of Geonoma brevispatha (Arecaceae) in laboratory and its relation to the palm spatial distribution in a swamp forest. Aquat. Bot. 85, 16–20. Guo, Y., Shelton, M.G., Lockhart, B.R., 1998. Effects of flood duration and season on germination of black, cherrybark, northern red, and water oak acorns. New Forest 15, 69–76. Ho¨lzel, N., Otte, A., 2004a. Ecological significance of seed germination characteristics in flood-meadow species. Flora 199, 12–24. Ho¨lzel, N., Otte, A., 2004b. Inter-annual variation in the soil seed bank of floodmeadows over two years with different flooding patterns. Plant Ecol. 174, 279– 291. Hroudova´, Z., Za´kravsky´, P., 2003. Germination responses of diploid Butomus umbellatus to light, temperature and flooding. Flora 198, 37–44. Insausti, P., Soriano, A., Sanchez, R.A., 1995. Effects of flood-influenced factors on seed germination of Ambrosia tenuifolia. Oecologia 103, 127–132. Leck, M.A., 1996. Germination of macrophytes from a Delaware River tidal freshwater wetland. Bull. Torrey Bot. Club 123, 48–67. Lorenzen, B., Brix, H., McKee, K.L., Mendelssohn, I.A., Miao, S.L., 2000. Seed germination of two Everglades species, Cladium jamaicense and Typha domingensis. Aquat. Bot. 66, 169–180. Lorenzi, H., 2000. Plantas Daninhas do Brasil: Terrestres, Aqua´ticas, Parasitas e To´xicas. Instituto Plantarum, Nova Odessa, SP. Manz, B., Mu¨ller, K., Kucera, B., Volke, F., Leubner-Metzger, G., 2005. Water uptake and distribution in germinating tobacco seeds investigated in vivo by nuclear magnetic resonance imaging. Plant Physiol. 138, 1538–1551.
D. Kestring et al. / Aquatic Botany 91 (2009) 105–109 Merritt, D.M., Wohl, E.E., 2006. Plant dispersal along rivers fragmented by dams. River Res. Appl. 22, 1–26. Mollard, F.P.O., Insausti, P., Sanchez, R.A., 2007. Flooding induces secondary dormancy in Setaria parviflora seeds. Seed Sci. Res. 17, 55–62. Nilsson, C., 1999. Rivers and streams. In: Rydin, H. (Ed.), Acta Phytogeogr. Suec., vol. 84, pp. 135–148. Orozco-Almanza, M.S., Le´on-Garcı´a, L.P., Grether, R., Garcı´a-Moya, E., 2003. Germination of four species of the genus Mimosa (leguminosae) in a semi-arid zone of Central Mexico. J. Arid Environ. 55, 75–92. Pereira, M.D., Dias, D.C.F.S., Dias, L.A.S., Arau´jo, E.F., 2007. Hydration of carrot seeds in relation to osmotic potential of solution and conditioning method. Rev. Bras. Sem. 29, 144–150. Ribas, L.L.F., Fossati, L.C., Nogueira, A.C., 1996. Superac¸a˜o de dormeˆncia de sementes de Mimosa bimucronata (D.C.) O. Kuntze (Marica´). Rev. Bras. Sem. 18, 98–101.
109
Scarano, F.R., Pereira, T.S., Roˆc¸as, G., 2003. Seed germination during floatation and seedling growth of Carapa guianensis, a tree from flood-prone forest of the Amazon. Plant Ecol. 168, 291–296. Tomaz, C.A., Kestring, D., Rossi, M.N., 2007. Effects of the seed predator Acanthoscelides schrankiae on viability of its host plant Mimosa bimucronata. Biol. Res. 40, 281–290. Vogt, K., Rasran, L., Jensen, K., 2007. Seed deposition in drift lines: opportunity or hazard for species establishment? Aquat. Bot. 86, 385–392. Warwick, N.W.M., Brock, M.A., 2003. Plant reproduction in temporary wetlands: the effects of seasonal timing, depth, and duration of flooding. Aquat. Bot. 77, 153– 167. Wittmann, A.O., Piedade, M.T.F., Parolin, P., Wittmann, F., 2007. Germination in four low-va´rzea tree species on Central Amazonia. Aquat. Bot. 86, 197–203. Zar, J.H., 1999. Biostatistical Analysis. Prentice Hall, Upper Saddle River, NJ.