Impact of γ-rays on seed germination/short-term storage in four native alpine species: Correlation with free radical and antioxidant profiles
Introduction
The effect of gamma (γ) radiation on seeds changes depending on strength (dose) and duration (dose-rate) of the imposed stress as well as the target species. Recently, Macovei et al. (2014) observed that in rice (Oryza sativa L.) seeds the response to γ-rays was dependent on total dose. High dose rate (HDR; 5.15 Gy min−1) irradiation was harmful while low total doses delivered at low dose rate (LDR; 0.28 Gy min−1) increased survival and growth. Similarly, wheat (Triticum aestivum L.) germination was inhibited by total doses in the 100–400 Gy range (Borzouei et al., 2010) and enhanced at lower total doses (10–100 Gy) (Singh and Datta, 2010). However, Wi et al. (2007) reported that exposure to 50 Gy total dose was sufficient to suppress growth in Arabidopsis seedlings. Thus, the selection of the highest threshold for γ-rays treatments is influenced by inter-species variability.
Germination tests and biometric measurements are routinely used to to evaluate the impact of γ-rays in seeds (Chaudhuri, 2002), however additional parameters need to be considered since the dose-dependent response of seeds to γ-rays correlates with the extent of free radical accumulation (Macovei and Tuteja, 2013). Gamma-rays cause water ionization which generates reactive oxygen species (ROS), including superoxide radical (O2−), hydrogen peroxide (H2O2) and hydroxyl radical (OH−), and secondary free radicals, such as organic hydroperoxide (ROOH), hydrochlorous acid (HOCl) and peroxynitrite (ONOO−) (Esnault et al., 2010). Electron Paramagnetic Resonance (EPR) was successfully used to highlight the linear correlation between free radical levels (paramagnetic species) and the absorbed γ-ray doses in maize (Zea mays L.) seeds (Marcu et al., 2013). Furthermore, it has been demonstrated that the activity of endogenous ROS-scavenging systems can significantly reduce the amount of free radical species generated by γ-irradiation (Kim et al., 2012, Ventura et al., 2012).
The aim of this study was to elucidate the relationship between γ-irradiation, ROS accumulation and seed quality in four native alpine species, Campanula barbata L., Cirsium spinosissinum (L.) Scop., Plantago alpina L., and Silene vulgaris (Moench) Garcke. The effects of γ-rays delivered at different doses on germination efficiency and antioxidant activity were assessed in dry seeds while changes in seedling length and weight were also determined. EPR spectroscopy was used to monitor the free radical profiles in seeds and their correlation with germination rates. The impact of standard (seed bank) dry storage conditions was also investigated. Different degrees of radiosensitivity were observed, depending on the tested species. A deeper understanding of the radiobiological responses occurring in seeds and seedlings of native species will help developing improved invigoration protocols which are currently needed to overcome the constrains of seed bank operators and improve long-term ex situ conservation of endangered plants (Araújo et al., 2016). This is especially relevant for alpine species, known to be short-lived in storage (Mondoni et al., 2011, Mondoni et al., 2014).
Section snippets
Wild population seed collection and γ-irradiation treatments
Seeds of Campanula barbata L. (Campanulaceae), Cirsium spinosissinum (L.) Scop. (Asteraceae), Plantago alpina L. (Plantaginaceae) and Silene vulgaris (Moench) Garcke (Caryophillaceae) were collected at the point of natural dispersal from four different alpine locations in Valtellina (Sondrio Province, Italy) at 46° 27' 3.5"N, 10° 13' 25.2"E (1910 m a.sl.), 46° 25' 43,8"N, 10° 11' 18.0"E (2173 m a.sl.), 46° 25' 59"N10° 11' 32"E (2142 m a.sl.) and 46° 30' 12,4"N, 10 18 43,4"E (1928 m a.sl.),
Germination response and seedling growth performance in γ-irradiated and dehydrated seeds
C. barbata revealed significant differences in seed germination in response to γ-irradiation and dehydration treatments (Fig. 1A). The maximum germination percentages were recorded in control (non-irradiated seeds) at 0 days (86.00±5.02%), 7 days (90.60±8.50%) and 14 days (96.55±0.00%). Germination efficiency declined as the total dose and dehydration period increased. The maximum decrease (89%) in germination percentage was observed with 100 Gy after 7 days of dehydration (10.76±5.77%) while
Discussion
The present work focuses on four native alpine species, C. barbata, C. spinosissinum, P. alpina and S. vulgaris, providing for them profiles of γ-rays tolerance in the attempt to gather knowledge useful to improve long-term ex situ conservation. Biometric parameters allowed to discriminate between ‘pattern 1’ (decreased growth, C. barbata and C. spinosissimum), and ‘pattern 2’ (normal growth, P. alpina and S. vulgaris). As for ‘pattern 1’, the observed decrease in plant biomass suggests for a
Acknowledgments
This work was supported by Regione Lombardia D.G. Attività Produttive Ricerca e Innovazione - Struttura Asse 1 POR FSE 2007–2013, Project ID 4344853 ‘Advanced Priming Technologies for the Lombardy Agro-Seed Industry-PRIMTECH’. S.A. has been awarded by a Research Contract within the same project sponsored by CARIPLO Foundation (Action 3, Code 2013-1727). The financial support from Fundação para a Ciência e a Tecnologia (Lisbon, Portugal) is acknowledged through research unit GREEN-it
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