Alleviating effects of silicon on cadmium toxicity in ginger ( Zingiber officinale Roscoe)

Background: This study assessed the effects of varying concentrations of silicon (Si) on the reduction of cadmium (2mg/kg Cd) toxicity in Zingiber officinale Rosc. via growth modulation, photosynthetic efficiency, and antioxidant defense. Result: As the Cd level increased, the physiological indexes of ginger exhibited a decreasing trend. This trend was especially noticeable when the Cd level was 2mg/kg. Next, the effect of different content of soil conditioner Si on ginger under 2mg/kg Cd soil background was explored. This effect was assessed under 2mg/kg Cd soil background. The three treatment Si0 (0), Si1 (1g/kg), Si2 (2g/kg) were examined. Morphology indexes, Cd content, Cd transfer and absorption coefficient, photosynthesis, and antioxidant enzyme activities under Cd stress were determined. On day 120 of the experiment, plant height had increased by 18.8% and 24.7% under the Si1 and Si2 treatments compared with the CK treatment. Meanwhile, the fresh weight of the rhizomes in Si1 and Si2 had increased by 14.3% and 19.5% in comparison with the Si0 treatment. Also, the yield of ginger improved dramatically under the Si1 and Si2 treatments. These treatments had increased by 14.3% and 19.5% compared with the CK treatment. The growth of ginger increased after the addition of Si under Cd stress, especially under the Si2 treatment. The Cd content of the mother-ginger, son-ginger, and grandson-ginger decreased by 27.6%, 35.5%, and 51.2%, respectively, under 2mg/kg Cd stress in the Si2 treatment. The Cd transfer was inhibited from the underground to the aboveground in the high-concentration Si treatment (Si2). When 1 g/kg and 2 g/kg Si was added to the soil, the leaf photosynthetic rate increased by 15.1% and 26.9% compared with the CK treatment at 11:00, respectively. Conclusion : Above all, soil conditioner Si could alleviate the negative effects of Cd stress on ginger by improving growth parameters, photosynthetic efficiency and the antioxidative defense. Si2 (2g/kg) oxide contributes to cadmium toxicity cadmium up-20

3 Background Cadmium (Cd) is one of the most poisonous heavy metals and widespread environmental pollutants [1]. The concentration of Cd in soil has been increasing in recent years. It is ranked as number seven among the top 20 toxins [2]. Its absorption coefficient is higher compared with other heavy metals such as copper and zinc. Absorption of Cd relies on the Cd concentration in soil [3]. As a result of industrial wastewater, exhaust emissions, and inappropriate use of pesticides and fertilizers, Cd pollution in the soil has become increasingly common. It thus poses a serious health risk to humans. Cd can immobilize in soil by binding to organic matter, and it is easily taken up and accumulated by plants [4].
Thus, dietary intake of the biotoxic crops could be a severe threat to humans, and the most likely risk is chronic toxicity in humans [5]. Besides being carcinogenic, Cd inhalations could harm the lungs, liver, kidneys, and bones [6]. In taller plants, Cd could accumulate in any part of the plant since it is highly mobile in phloem tissue. Cd toxicity changes chloroplast ultrastructure, decreases the net photosynthetic rate, leaf transpiration, and stomatal conductance [7]. Cd toxicity is usually accompanied by oxidative stress and DNA breakdown [8] and, ultimately, causes cellular damage or apoptosis [9, 10].
Recent studies indicate that a variety of strategies have been developed to lessen the toxic effects of Cd pollution. Nitric oxide (NO), as a signaling molecule, has a pivotal role in plant resistance to various stresses, including heavy metal stresses, such as Cd [11].
NO could interact with ROS, especially O 2− , because of the presence of an unpaired ewithin the NO molecule [12]. NO improved tolerance to Cd toxicity by reducing oxidative stress that is considered as the leading cause of cell death [13]. In Lupinus, when roots were grown in 50 µM Cd, NO could stimulate SOD activity to counteract the overproduction 4 of O 2− [ 14]. One study found that salicylic acid alleviates Cd-induced photosynthetic damage and cell death by inhibiting reactive oxygen overproduction [15]. The improvement of signal transduction also increased plant tolerance to Cd, and decreased cell death resulted from Cd [10,16]. The soil amendment, Biochar, has been known to protect plants against heavy metal stress [17]. In wheat, Biochar reduced cadmium toxicity for plants growing in Cd-contaminated saline soil [18].Biochar application decreased the oxidative stress in plants and recovered the antioxidant enzyme activities.
Silicon is the second-most abundant element in the world, and it could promote plant growth and weaken both biological and non-biological stresses [19,20]. It has been reported that Si plays a vital role in the transfer and accumulation of Cd in plants [21]. Si alleviates Cd toxicity in several ways. It activates the antioxidant system in plants [22], forms Si and Cd precipitants, and restrains Cd translocation, which weakens its biological activity [23]. In rice, 120 mg L − 1 Si decreased Cd accumulation and also reduced the ratio of Cd, transferring from roots to shoots, which lessened the Cd toxicity [24]. In Pisum sativum L, the application of Si decreased Cd accumulation and increased the absorption of macronutrients and micronutrients in shoots and roots, which alleviated Cd toxicity

Result
The effect of cadmium stress on the growth and quality of ginger There is an impact on ginger growth when 1, 2, and 4 mg/kg of Cd are applied during the growth process of ginger (Table 1). At 40d, the plant height and fresh weight were significantly lower than the CK at four mg/kg Cd level. At 80d and 120d after the treatment, the physiological indexes of ginger decreased as Cd levels increased. Above all, the physiological indexes under 2 mg/kg and 4 mg/kg of Cd stress were significantly lower than those of the CK. This indicates that the root growth was significantly inhibited, the number of branches was reduced, and the biomass of the above-ground parts was reduced when the Cd level was higher than 2 mg/kg ( Table 1).
The quality indexes of ginger rhizomes indicated there were significant differences among the treatments. The yield and dry weight of ginger showed a decreasing trend with the increasing of Cd levels. The yield decreased by 9.1%, and dry weight decreased by 7.8% 6 compared with the CK at 2 mg/kg of Cd. Gingerols act as the primary indicators when evaluating ginger flavor and quality. As seen in Table 2, the content of gingerols significantly decreased as the Cd level increased. The content of gingerols was reduced by 12.1%, 31.0%, and 38.0% compared with the CK. This showed that the ginger quality was significantly reduced at 2 mg/kg Cd stress ( Table 2).
The effect of silicon on the growth and quality of ginger under Cd stress Based on the effect of Cd stress on the growth and quality of ginger, the physiology indexes significantly decreased compared with the CK from the 2 mg/kg of Cd. Because of this, 2 mg/kg of Cd was chosen for the background value in the subsequent research. To observe the effect of silicon on the growth and quality of ginger under Cd stress, we used 0 g/kg, 1 g/kg, 2 g/kg Si to explore the alleviation effect of varying Si concentrations under 2 mg/kg of Cd stress. The growth of ginger was promoted after the addition of Si under Cd stress (Fig. 1). There is no significant difference among different treatments at 40d, but the difference was significant at the rhizome stage.  (Table 3). Also, the yield of ginger improved dramatically under the Si1 and Si2 treatments, which showed an increase of 14.3% and19.5% compared with the CK. Soluble sugar, crude cellulose, soluble protein, free amino acid, and vitamin C increased as the application of Si increased. The effect was the greatest in the Si2 treatment. Application of Si could improve ginger flavor quality. The content of gingerols was improved by 36.8% and 63.2% under Si1 and Si2, compared with the Si0 (Table 4).

7
The effect of silicon on Cd content, Cd accumulation in different organ of ginger under Cd stress Cd content was significantly reduced after the addition of Si under 2 mg/kg Cd stress (Table 5). At 40d and 80d, the Cd content of the rhizome and the root was significantly reduced after the addition of Si. At the same time, the Cd content of the stem and the leaves had reduced, but not significantly so. At 120d, Cd content of each part was dramatically decreased after applying Si. The Cd content had reduced from 0.2566, 0.1070, and 0.0580 µg/g, respectively, to 0.1861, 0.0686, and 0.0277 µg/g, respectively, in mother-ginger, son-ginger, and grandson-ginger under the Si2 treatment (Table 5).
As Table 6 shows, Cd accumulation in the root and rhizome was less than the CK under the Si1 treatment. The Cd accumulation in the aboveground part changed insignificantly and showed a slightly increasing trend, indicating that Cd accumulation in the aboveground part was promoted after applying Si. The Cd accumulation under the Si2 treatment significantly decreased compared with the CK, indicating that high-concentration Si treatment significantly inhibited Cd accumulation. In conclusion, Si in ginger could promote or inhibit Cd accumulation, depending on the concentration of Si in the soil (Table 6).
The effect of silicon on cadmium absorption coefficient and transfer coefficient under Cd stress The primary transfer coefficient under the Si2 treatment was significantly lower than the Si0 and Si1 treatments, implying that higher concentrations of Si inhibited Cd migration from the root to the rhizome ( Table 7). The secondary transfer coefficient under the Si1 treatment was significantly higher than in the Si0 and Si2 treatments, which indicates that the lower concentration of silicon promotes the transfer of Cd from the rhizome to the ground. The root absorption coefficient of the Si2 treatment was significantly lower than in 8 the Si0 and Si1 treatments, which indicates that a high concentration of silicon could dramatically inhibit the root from absorbing Cd from the soil. The aboveground absorption coefficient of the Si2 treatment (0.023) was significantly lower than the Si0 (0.040) and Si1 (0.038) treatments, indicating that a high concentration of silicon inhibited the plant from absorbing Cd from the soil ( Table 7). The ROS level exhibited a rising trend as the treatment days increased (Fig. 4) (Fig. 4).
The SOD, POD, and CAT increased as the Si content increased under two mg/kg Cd (Fig. 5).
There was a significant difference in the three treatments, and the Si2 treatment was the highest. With the extension of time, the SOD activity of the Si0 treatment gradually decreased. The SOD activity of the Si1 treatment and the Si2 treatment increased and then decreased. At 120d, the SOD activity of the Si2 treatment was the highest, the Si1treatment was second, and the Si0 treatment was the lowest, suggesting that the addition of Si inhibited the SOD activity declining (Fig. 5). The most minor damage to the membrane system in the ginger leaf was in the Si2 treatment, while the Si1 treatment was second. The Si0 treatment showed the most critical damage to the membrane system.
During the whole growth stage, the extent of damage was increasingly severe as time went on. At 120d, the MDA content reached the maximum. The MDA of the Si1 treatment and the Si2 treatment was reduced by 14.6% and 20.0% compared with the Si0 treatment which indicates that the damage of the membrane system in the ginger leaf was lower when silicon-containing fertilizer was applied. The Proline content significantly decreased from the Si0 treatment to the Si1 treatment, to the Si2 treatment during the whole growth period (Fig. 6).

Silicon augmented plant growth and biomass
Heavy metal stress can destroy the physiological and biochemical processes in plants Silicon restored photosynthetic efficiency Cd could reduce chlorophyll synthesis, the photochemical quantum yield of ΦPSII, and the CO 2 fixation rate. It was reported that in maize, chlorophyll synthesis and the photochemical quantum yield of ΦPSII decreased under Cd stress [42]. In durum wheat, Cd affected chlorophyll fluorescence [43]. However, the exogenous addition of Si under Cd stress had powerful effects on chlorophyll synthesis and photosynthetic machinery. In our study, the Pn was the least at 13:00, and the Pn of the Si1 and the Si2 treatments were improved by 14.4% and 24.6%, respectively, compared with the CK. Therefore, Si addition under Cd stress alleviated the damage to the ginger plants by weakening the decreasing trend of the Pn under Cd stress (Figure 2). The results were similar in peas, cotton, and maize. In the pea plant, Si addition promoted the contents of chlorophyll pigment and carotene [44]. The same condition occurred in cotton seedlings; the contents of photosynthetic pigments were increased with the addition of exogenous Si [45]. The positive effects of Si on photosynthesis could be due to the destroyed uptake of heavy metals, which could enhance PSI and PSII activation [46]. In maize, Si alleviates Cd toxicity by increasing photosynthetic rate in the modified bundle sheath cells [47].

Silicon modulated antioxidant activity
Plants could use a series of strategies against the toxic effects of heavy metals under heavy metal stress conditions. The activities of SOD, POD, and CAT are of considerable significance to scavenge the ROS caused by heavy metals [48,49]. Previous studies have reported that Si mediates up-regulation of the antioxidant defense system by increasing the SOD, POD, CAT, and GR activity [50,51].Alleviation of heavy metals toxicity by Si was correlated with protection against oxidative damage. In a previous report, Si could alleviate Cd stress because of a noticeable increase in antioxidant activity and a decrease in MDA in pakochi [52]. In cotton, Si addition could significantly improve the plant's defense capacity against oxidative damage caused by Cd stress. MDA, H 2 O 2 , and electrolyte leakage were reduced, and SOD, POD, APX, and CAT activities were enhanced under Cd stress [45]. In cucumber, the application of Si could eliminate heavy metal Mn toxicity by improving antioxidant activity, according to Shi et al. [53].
Our study showed that SOD, POD, and CAT activities were increased as the Si content 13 increased under Cd stress (Fig.5). At 120d, MDA of the Si1 treatment and the Si2 treatment was reduced by 14.6% and 20.0% compared with the Si0 treatment, indicating that the damage of the membrane system in the ginger leaf was lower when siliconcontaining fertilizer was applied. Proline content was markedly decreased from the Si0 treatment to the Si1 treatment to the Si2 treatment during the whole growth period (Fig.6). At 120d, after applying Si, the Cd content in various organs of the ginger plant was reduced. The Pn of ginger leaves appeared in double peaks in one day, and the diurnal variation trend of each treatment was similar, ranked as Si2 > Si1 > Si0. Overall, our results suggest that Si could alleviate the negative effect of Cd stress on ginger, and the Si2 treatment was the most effective.

Experimental materials
The experiment took place at Shandong Agricultural University at the experimental 14 horticulture station. The ginger variety used was "Laiwudajiang" (WanXing Food company, LaiWu, Shandong Province). The soil tested was pH=7.3, with 100.5 mg/kg alkalihydrolyzed nitrogen (N), 63.4 mg/kg available phosphorus (P 2 O 5 ), and 127.8 mg/kg available potassium (K 2 O). The background value of soil Cd was 0.14 mg/kg. The soil used for the study was air-dried and put into a plastic basin with a 30 cm diameter and a height of 28 cm. Each basin contained 8.0 kg of air-dried soil. The ginger was planted when it germinated to 1cm. Two plants were grown per pot.

Experimental design
The experiment followed the environmental quality standard GB15618-2009.The Cd level was set at zero, one, two, and four mg/kg (soil). There were 10 basins per treatment, and each treatment had three replicates. Varying concentrations of cadmium chloride (CdCl2·2.5 H2O) solution was added to the soil via sewage irrigation once the ginger emerged. The CdCl2·2.5 H2O was applied in this way to ensure the uniform distribution of heavy metals in the soil and to prevent loss from the plastic containers. Other aspects of the experiment were conducted according to the standard method. Samples were collected at the seedling stage, trilling stage, and rhizome expansion stage, respectively (i.e., 40 d, 80 d, and 120 d), after the sewage irrigation treatment, and relevant indexes were determined.
Based on the experiment done in the first year, the stress level of Cd was set at two mg/kg (soil), and the amount of silicon fertilizer was zero, one, and two g/kg (soil), respectively. There were 10 pots in each treatment, and each treatment had three replicates. When sowing ginger, the silicon fertilizer mixed with the 10 cm topsoil in a basin. Two mg/kg Cd (in terms of Cd 2+ ) were applied to the soil using the sewage irrigation method when the ginger emerged. The sewage irrigation method was used to 15 ensure that the heavy metals were evenly distributed in the soil and not lost from the pot.
Samples were taken at 40 d, 80 d, and 120 d after treatment, and relevant indexes were measured.

Determination of growth indexes
The plants were taken and rinsed with water. Plant height, stem diameter, branch number, leaf number, root, stem, leaf and fresh weight of rhizome were measured.
After 20 minutes in the drying oven at 105 ℃, the samples were dried to constant weight at 75 ℃. The dry matter of each organ was measured.

Determination of quality
The volume of soluble proteins, soluble sugars, free amino acids, crude fiber, and vitamin C were measured using various methods. Soluble proteins were measured by staining with Coomassie Brilliant Blue. Soluble sugar content was determined using the anthrone colorimetry technique [54]. Free amino acids were measured using the ninhydrin method [55]. Crude fiber volume was determined by the acid-wash method. And vitamin C was measured using the standard of 2,6-dichloro indophenol [56].
To measure the content of gingerols, 1 g of ginger powder was added to the 100-ml volumetric flask, then 70 ml acetone was mixed into the mixture and shaken for 1 h at 50°C. The mixture was then cooled and diluted with acetone to volume. Filter liquor was used for determining the content of gingerols [57].

Ethics approval and consent to participate
Not applicable

Consent for publication
Not applicable.

Availability of data and material
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Competing interests
The authors declare that they have no competing interests.

Funding
This study was supported by the National Characteristic Vegetable Industry Technology System Project (Grant No. CARS-24-A-09) and the Shandong Province's dual-class discipline construction project (Grant No. SYL2017YSTD06).The funding organizations provided the financial support to the research projects, but were not involved in the design of the study, data collection, analysis of the data, or the writing of the manuscript.

Authors' Contributions:
As designed and performed the experiment, ZC and JZ did the experiment.    Values followed with the same letter was not significant at P = 0.05. Error bars stand for the standard errors. Tables.pdf