Plant Soil (2008) 313:83–87 DOI 10.1007/s11104-008-9681-6
REGULAR ARTICLE
Effect of salinity and chemical factors on seed germination in the halophyte Crithmum maritimum L. Laetitia Meot-Duros & Christian Magné
Received: 20 March 2008 / Accepted: 4 June 2008 / Published online: 5 July 2008 # Springer Science + Business Media B.V. 2008
Abstract Seeds of the halophyte Crithmum maritimum L. were exposed to salt stress and chemical pretreatments in order to improve germination. Seeds submitted to salt stress did not germinate but they recovered rapidly upon transfer to distilled water, recovery being higher after a low salinity pretreatment. Chemical treatments resulted in differential effects on seed germination. Known dormancy breakers such as potassium nitrate and thiourea had no effect on sea fennel seed germination. Conversely, L-ascorbic acid (40 or 60 mM) and ethanol (96%) significantly improved germination rate by 10, 30 and 30%, respectively. Pretreatment of seeds with L-ascorbic acid at 40 mM was shown to alleviate the negative effects of low NaCl concentration on germination. These findings indicate that the application of ascorbic acid may be used to improve sea fennel seed germination, which is of great interest for cultivating this plant.
Responsible Editor: Hans Lambers. L. Meot-Duros (*) : C. Magné Laboratoire d’Ecophysiologie et de Biotechnologie des Halophytes et des Algues Marines, EA 3877 (LEBHAM), Institut Universitaire Européen de la Mer, Université de Bretagne Occidentale, Technopôle Brest Iroise, Place Nicolas Copernic, 29280 Plouzané, France e-mail:
[email protected] C. Magné e-mail:
[email protected]
Keywords Crithmum maritimum L. . Dormancy . Effectors . Salt stress . Seed germination
Introduction Salinity is one of the most detrimental factors affecting germination of halophyte seeds and plant establishment (Katembe et al. 1998). An increase in salinity induces both a reduction in the percentage of germinating seeds and a delay in the initiation of the germination process. It can also cause a complete inhibition of the germination process at NaCl concentrations beyond the tolerance limits of the species (Pujol et al. 2000). In most halophyte seeds, high salinity-induced dormancy is controlled by compounds that regulate germination activity (Khan and Ungar 1996; Pujol et al. 2000). For example, seed dormancy can be broken by physical or chemical treatments, including scarification, maceration in strong acid or incubation in gibberellins or ethanol (Miyoshi and Sato 1997; Yang et al. 2007). Crithmum maritimum L. (Apiaceae), commonly known as sea fennel or rock samphire, is an edible halophyte growing on maritime cliffs and sometimes in sand. It is distributed widely along coastal areas of the Mediterranean Sea and the Atlantic Ocean. This species has long been used as a food ingredient and in folk medicine (Coiffard 1991). Moreover, it contains high levels of flavonoids, carotenoids, vitamin C, essential oils (Ruberto et al. 1991; Senatore and de
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Feo 1994), and substances with medicinal and antimicrobial properties (Ben Amor et al. 2005). Therefore, there is growing interest in sea fennel cultivation, highlighting the need for more knowledge of seed germination in that species. Sea fennel seed germination has been studied by various authors, mainly regarding light requirements (Okusanya 1977; Marchioni-Ortu and Bocchieri 1984; Thanos et al. 1991). Results often differed in these studies, suggesting that light requirement depends on the plant biotope (sand hill or rock). Two studies have described the effects of salinity on Crithmum maritimum seed germination. Okusanya (1977) found that seeds from southwest England presented a reduced germination rate and an increased time of germination as salinity increased. The author also showed that C. maritimum seed germination could recover after pretreatment with NaCl. MarchioniOrtu and Bocchieri (1984) studied the germination process of a Sardinian population of sea fennel, confirming, in complete dark experiments, the previous observations of Okusanya (1977). However, to our knowledge, no study has been made on the effect of dormancy breakers on sea fennel seed germination. In the present work, we studied the effectors of germination in a sea fennel population from Brittany. Preliminary observations carried out in our laboratory showed that the presence of light was essential for C. maritimum seed germination. The aim of this study was to find a suitable (pre)treatment that improves germination and overcomes the effects of NaCl.
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2 paper soaked in 5 ml solution. Incubation was carried out under a thermoperiod of 23/18°C (light/ dark) and an 18/6 h photoperiod (light/dark, 50 µmol sec−1 m−2, Sylvania GRO-LUX F15W/GRO-T8). The seeds were checked daily and the criterion for germination was visible radical emergence. The germination tests lasted 75 days for salt treatment, and 40 days for the chemical treatments tests. For each Petri dish, germination percentage (GP) was calculated as [number of germinated seeds/ number of sampled seeds] × 100. The recovery percentage was determined by the following formula: ða bÞ=ðc bÞ 100 where a is the number of seeds germinated after being transferred to distilled water, b is the number of seeds germinated in saline solution, and c is the total number of seeds. The rate of germination was estimated using a modified Timson index of germination velocity: X Germination velocity ¼ G=t where G is the percentage of seed germination at each measurement and t is the number of measurements (Khan and Ungar 1996). The effect of each treatment compared to the control was also expressed as [(number of germinated treated seeds/number of germinated control seeds) x 100]−100. Treatments
Materials and methods
Salt
Seeds of Crithmum maritimum L. were collected during autumn 2006 along the shoreline of Brittany at “Pointe du Toulinguet” (France). After removing immature seeds and unwanted materials, seeds were dried at ambient laboratory temperature and stored in a cold room at 5°C until germination tests began. In all experiments, treatments were made using analytical grade chemicals in water.
Seeds were sown in Petri dishes on filter paper soaked with NaCl solution (50, 250 or 500 mM). After 50 days of incubation, ungerminated salt-treated seeds were desalinated for 1 day, and finally tranferred to filter paper soaked with distilled water to study germination recovery. Control seeds were treated continuously with distilled water. Chemical effectors
Germination tests Three replicates of 20 seeds were used for each condition. Each set of seeds was placed in 7-cm diameter tightly sealed Petri plates with Whatman No.
Five chemical treatments to improve C. maritimum seed germination were tested. Four consisted of pretreating seeds at room temperature for 3 h in Lascorbic acid (at 20, 40 or 60 mM) or for 24 h in the
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following solutions: 96% ethanol, 0.1 mM GA3, or 50 mM thiourea. Seeds were then rinsed in deionised water prior to germination tests. A fifth treatment consisted of soaking seeds in 0.01 M KNO3 solution during the whole germination test. The effect of ascorbic acid pretreatment on seed germination in saline conditions was also studied. After 3 h of pretreatment, seeds were rinsed in distilled water and immersed in water (control) or NaCl solutions (50, 250 or 500 mM). Statistical analysis The transformed GP and germination velocity index were subjected to one-way analysis of variance (ANOVA) using Statgraphics Plus 5.1 (Statsoft, Cary, NC), and an LSD test was used to determine whether differences among treatment means were significant (P
