Germination and vigor responses to UV-B irradiation

The UV-B radiation caused differences in the variables germinated seeds (%), normal seedlings (%) and dry weight (g), on the other hand there were no differences in the variables length of hypocotyl (cm) and length of root (cm).In the variables where differences between the treatments and the control (p≤ 0.05) were observed, a tendency to increase with a maximum in the treatment with 5.4 kJ m^-2 is observed, then a decrease that reaches the lowest values with 16.2 kJ m^-2, then again a positive trend reaching another peak in the treatment with 27 kJ m^-2 (Figure 1).

Figure 1 Different doses of UV-B radiation applied to Big Rio hybrid tomato seeds and their effect on germination and vigor of seedlings. The bars in each graph indicate mean values for each variable and different literals on each bar indicate significant differences according to a Tukey test (p≤ 0.05).  

In the germinated seeds variable (Figure 1a), the treatment with 5.4 kJ m^-2 of UV-B radiation is statistically superior (p≤ 0.05) to the control, this kind of response has been explained by ^{Luckey (1980)}, who Mentions that the sublethal application of an agent capable of inducing physical or chemical stress can generate positive responses. Apparently at 5.4 kJ m^-2, a response threshold was reached in the seed in which certain receptors that trigger a response aimed at accelerating germination could act. Doses greater than 5.4 kJ m^-2 resulted in responses not different from the control, but with the treatment of 27 kJ m^-2 a value 21% higher than the control was reached again. Possibly with this higher amount of energy a second response threshold could be reached in the seed, in that threshold could be involved different receptors.

These kinds of responses with several maxima, and perhaps dependent on different receptors, are very valuable for the seeds from an adaptive point of view, since they allow a greater flexibility in the response to different environmental contexts (examples: different depths under the soil in a seed bank or different types or canopy coverings on a seed bank) and ensure that it is not only a certain threshold value or individual stimulus that triggers the germination process and the growth of the embryo (^{Long et al., 2015}).

Similar results were found in the percentage of normal seedlings (Figure 1b), treatments with 1.8 and 5.4 kJ m^-2 were superior to the control, perhaps as a consequence of a hormone-induced radiation effect mentioned by ^{Luckey (1980)}. Normally these responses are positive since the formation of a normal seedling ensures optimum growth and better productivity (^{Delibaltova and Ivanova, 2006}). The treatment with 27 kJ m^-2 showed a new response peak as in germinated seeds variable, which could indicate the action of different signaling receptors or cascades activated with different energy thresholds.

In the hypocotyl length variable (Figure 1c) as well as in the root length (Figure 1d) there was no significant difference (p≥ 0.05). Other authors found response to irradiation in these variables, ^{Kacharava et al. (2009)} irradiated bean seeds for 30 minutes and found an increase in plant height, but with 90 min the height decreased.

Similarly, ^{Hidema et al. (2006)} found that supplementary UV-B radiation decreased plant growth and grain yield in rice. It is not available until an explanation of why stimulus against UV-B occur different, positive or negative, or neutral responses, between species even in the same family (^{Kossuth and Biggs, 1981}). The same happens with the different responses exhibited by different variables in the same species, as shown in this study.

In the dry weight (Figure 1e), the same response already described for the percentage of germinated seedlings and normal seedlings, with maxima at 5.4 and 27 kJ m^-2, is repeated. An analogous result was described by ^{Kacharava et al. (2009)}, who reported an increase in dry matter by applying 60 minutes of ultraviolet radiation to bean seed.

Germination and vigor responses to UV-C irradiation

In contrast to UV-B irradiation in the case of UV-C, no positive responses were reported. This difference probably arises from the fact that, unlike UV-B, UV-C radiation is not a natural component in ecosystems (^{Nawkar et al., 2013}). In agreement with ^{Promyou and Supapvanich, (2012)} it is possible that the observed responses in germinated seeds and seedlings are the result of the oxidative damage caused by the exposure even to the smaller doses of UV-C. This explains in part why the negative responses were evident in the processes most sensitive to oxidative damage on germination and normal seedling production after germination (Figure 2a and 2b). While the subsequent processes that gave rise to hypocotyl length, root length and dry weight, which are variables evidenced only in normal seedlings, they were not affected (Figures 2c, 2d and 2e). The latter results coincide with those reported by ^{Foroughbakhch-Pournavab et al. (2015)} in wheat and sunflower.

Figure 2 Different doses of UV-C radiation applied to Big Rio hybrid tomato seeds and their effect on germination and vigor of seedlings. The bars in each graph indicate mean values for each variable and different literals on each bar indicate significant differences according to a Tukey test (p≤ 0.05).  

In relation to germinated seeds (Figure 2a) the response to radiation was practically the same with all doses of UV-C, showing lower values to the control with the doses of 0.9, 2.7, 8.1 and 10.8 kJ m^-2, whereas with the doses of 5.4, 13.5 and 16.2 there were no significant differences (p≥ 0.05), these results indicate the great sensitivity of the germinative response to the UV-C stimulus and are very similar to those reported by ^{Brown et al. (2001)}, who irradiated lettuce seeds and found responses of this type, depending on the dose, in the incidence variables of Xanthomonas campestris pv. campestris and in that of colony forming units.

In the percentage of normal seedlings (Figure 2b) only the highest dose of 16.2 kJ m^-2 showed a lower value than the control, again supporting the explanation that unlike the UV-B stimulus, which resembles a response to a photomorphogenic stimulus, the response to UV-C is due to oxidative or other damage caused by the large amount of energy carried by UV-C photons. These results coincide with those of ^{Brown et al. (2001)}, who reports that the diameter of the head of lettuce plants obtained from seeds treated with low doses of UV-C radiation was not different from the control, whereas in seeds treated with higher doses the diameter was reduced considerably. Other negative responses to UV-C radiation applied to plant tissues are low protein content, dysfunctions in chloroplasts, DNA damage, and in various cellular processes (^{Danon and Gallois, 1998}; ^{Rastogi et al., 2010}; ^{Ruiz-López et al., 2010}).

Plant responses from seeds irradiated with UV-B and UV-C

In the second stage, the SPAD and stomatal density variables of the beam and underside were different to the control, indicating that the radiation applied to the seed is capable of generating changes that become visible in later stages (Figure 3), such as described by ^{Musil et al. (1998)} and ^{Magliano and Casal (2004)} for environmental signals perceived in pre-germination by phytochromes.

Figure 3 Response of the plants obtained from Big Rio hybrid seeds irradiated with UV-B and UV-C and planted in pots under greenhouse. The bars in each graph indicate mean values for each variable and different literals on each bar indicate significant differences according to a Tukey test (p≤ 0.05).  

For the variables plant height (Figure 3a) and stem diameter (Figure 3d) no treatment was different from the control. This behavior of tomato plants was different from that reported for other species such as cabbage, which showed higher growth when the seeds were irradiated with 3.6 kJ m^-2 of UV-C (^{Brown et al., 2001}), Arabidopsis with decreased petiole vs. UV-B radiation (^{Gruber et al., 2009}) or soybean showed lower seedling length when seeds were irradiated with UV-C (^{Foroughbakhch-Pournavab et al., 2015}). Again, the different response of each species seems to apply (^{Kossuth and Biggs, 1981}), considering factors such as seed size, seed thickness, chemical composition or other intrinsic seed properties of each species.

In the stomatal beam density (Figure 3c) and the underside (Figure 3d), changes induced by irradiation of the seeds with UV-B and UV-C were found determining that the response in the plants was to raise the density stomatal in the beam, whereas for the stomatal density in the back an inverse trend was found against the higher dose of energy applied with UV-B, but an opposite response when applying UV-C. ^{Wargent et al. (2009)} indicated that these adaptive responses to UV radiation impact on plant performance, modifying the interaction with different environmental factors through changes in form, physiology and biochemistry.

Similar results were obtained by ^{Benavides-Mendoza et al. (2003)} when irradiating wheat seeds with a low-intensity laser (AsAlGa), observing that at a dose of 6 kJ m^-2 the seedlings obtained from the irradiated seeds showed higher values of stomatal density in the bundle and at the back to grow in an environment conducive to growth, while germinating in a medium with different levels of salinity irradiated seedlings presented a clear adaptive response decreasing the stomatal density, a response that was not observed in the control. It is probable that the tomato plants obtained from the seeds irradiated with UV show responses with some background of adaptation to an environment where a greater amount of radiant energy is detected, which would imply the necessity of adjustments in the characters of the epidermis.

In the variables of fresh weight (Figure 3e) and dry weight (Figure 3-F) no difference was found in any of the treatments with the control. Again these results seem to indicate a specific response for tomato plants, since ^{Kacharava et al. (2009)} observed increases in the fresh weight of some varieties of Phaseolus vulgaris with applications of 60 and 90 min of UV-B; as well as an increase in the weight of white beet with 60 min of UV-B applied to the seed. In the lettuce irradiation of the seeds with a low intensity laser caused increase in the radical biomass but did not change the weight of the leaves and the stem (^{Benavides-Mendoza et al., 2003}).

The SPAD units (Figure 3g) decreased with UV-B treatments 5.8 and 10.5 kJ m^-2. The response can be explained by the fact that SPAD units indirectly measure chlorophyll concentration in foliar sheets (^{Rodríguez-Mendoza et al., 1998}) and that this pigment responds inversely to the amount of radiant energy detected in the environment (^{Vu et al., 1984}; ^{Strid et al., 1990}). The results obtained clearly indicate that such signaling occurs from the seed. In contrast to UV-B in the case of UV-C irradiation there were no differences with the control, but Figure 3g shows a positive trend in data different from that exhibited by UV-B.