Sweet corn (Zea mays L. saccharata Sturt) is an important commercial crop in China, which is mainly eaten fresh because of its rich nutrition and good palatability, and is loved by the majority of consumers (Zheng, 2019). Guangdong Province is the main producing area of sweet corn, and the planting area accounts for about 60% of the total area of China (Wang et al., 2014). However, in recent years, with the continuous development of industrialization, urbanization and agricultural modernization, a large number and various kinds of heavy metal ions have been continuously input and accumulated into the soil (Yang et al., 2018). The investigation and research showed that about 1/5 of China’s cultivated land was polluted by heavy metals to varying degrees, among which cadmium ion pollution was the most serious (Chen et al., 2017). Due to the high degree of soil acidification in southern China, the relative enrichment of cadmium ions in soil has been often increased, and the content of available cadmium has been at a high level for a long time. So, the farmland in this region was facing a serious problem of exceeding the standard of cadmium (Chen et al., 2017). Therefore, how to develop the production and industry of sweet corn with the characteristics of “high quality, high yield, high efficiency, ecology and safety” has become the key problem to be solved urgently in China’s agricultural safety production.

Cadmium (Cd) is not an essential element for plant growth, and has strong biological toxicity (He et al., 2019). Cadmium ions have high hydrophilicity and migration ability, and can be enriched in plants even at low concentrations (Liu et al., 2020). When cadmium accumulates to a certain extent in plants, cells will be damaged, resulting in various toxic symptoms, such as leaf wilting, stem shortening, lateral root reduction, chlorophyll content reduction, cell membrane permeability change, antioxidant enzyme activity reduction and so on, which has a strong inhibitory effect in the whole process of corn growth (Lin et al., 2012), and seriously restricts the healthy and sustainable development of corn industry (Shen et al., 2019). Studies have shown that too high cadmium ion concentration will reduce the cortical cells and vascular tissue of corn coleoptile, deform the xylem and phloem, and then affect seed germination, resulting in a significant reduction in germination rate (Wahid et al., 2015). After cadmium ions enter plant cells, they will combine with intracellular solutes and destroy their normal structure, resulting in blocked absorption of water and mineral elements, and significant inhibition of photosynthesis, respiration and transpiration (Fan et al., 2018; Anjum et al., 2016). With the increase of cadmium concentration, the enrichment of cadmium ions in plant cells will lead to the production of free radicals in plants, which will trigger oxidative stress response and affect the normal metabolic process of plants (Yu et al., 2010). In addition, high concentration of cadmium ions will also lead to the rupture of plant cell membrane, cell disintegration and the reduction of water absorption capacity of roots, which will seriously limit the fresh weight and water content of plants (Yu et al., 2010). It can be seen that cadmium ions in soil seriously restrict the growth of plants, and the degree of harm is positively correlated with the enrichment of cadmium ions.

Previous studies have shown that the tolerance of corn seeds of different genotypes to cadmium ions was quite different during germination (Meng et al., 2016), but there were relatively few studies on the cadmium tolerance response characteristics of fresh eating corn and the cadmium tolerance differences among different sweet corn varieties in the existing literature. Corn seedlings are relatively sensitive to the external environment, and their growth are greatly affected by stress. In addition, the evaluation of cadmium tolerance in corn seedling stage has the advantages of easy operation, strong repeatability and low environmental pollution (Yang et al., 2019). Based on this, this study selected 12 sweet corn varieties mainly promoted in South China for cadmium stress treatment at seedling stage, and screened out sweet corn varieties with strong cadmium tolerance by comparing and analyzing the differences between different varieties before and after cadmium ion treatment, in order to provide some reference for guiding the planting and popularization of sweet corn in South China.

1 Results and Analysis

1.1 Different responses of different sweet corn varieties to cadmium stress

Under the same cadmium stress treatment, the variation coefficients of cadmium tolerance index corresponding to plant height, root number, root length, shoot fresh weight, shoot dry weight, root fresh weight, root dry weight and SPAD value of 12 sweet corn varieties were between 7.02%~68.21%, among which the variation coefficient of cadmium tolerance index of five phenotypic traits including root related indexes such root number, root length, root fresh weight and root dry weight and SPAD value of the first leaf was large (CV>10%) (Table 1). Among the SPAD indexes with the largest variation coefficient of cadmium tolerance index, the cadmium tolerance indexes of ‘Zhongtian 7’, ‘Zhongtian 9’, ‘Ruizhen’, ‘Weitian 1’ and ‘Yuetian 28’ were all less than 0.35, while the cadmium tolerance indexes of ‘Zhuyutian 1’, ‘Zhongxiantian 102’ and ‘Fotian 10’ were all more than 0.95, and the difference among varieties reached a significant level (p<0.05). Among the cadmium tolerance indexes corresponding to the root number traits, the cadmium tolerance index of ‘Jinzhongyu’ was the lowest (0.76), and it was also significantly different from ‘Fotian 10’ (1.02), which had the highest cadmium tolerance index (p<0.05).

1.2 Comprehensive evaluation of cadmium tolerance of different sweet corn varieties

The correlation analysis of cadmium tolerance indexes of 8 traits showed that except that there was no significant correlation between two pairs of traits, that is, relative root number and relative root length, relative shoot dry weight and relative root number, there was significant or extremely significant correlation between cadmium tolerance indexes of the other 26 pairs of traits (Table 2), which showed that there was a large degree of overlap between the information provided by cadmium tolerance indexes of various traits. So evaluating cadmium resistance only through a single index existed some limitations. Therefore, the principal component analysis (PCA) method was used to convert multiple traits into comprehensive indicators before evaluation.

Through the principal component analysis of the cadmium tolerance index of 8 single indexes of 12 sweet corn varieties, the eigenvalue of the extracted first principal component was the highest (5.87), the contribution rate was 73.42%, and the eigenvector of each single index was between 0.74~0.96 (Table 3), which can comprehensively reflect the comprehensive situation of the test materials, Therefore, it can be used as a comprehensive evaluation index of cadmium tolerance of 12 sweet corn varieties selected in this experiment. Through further cluster analysis of the scores of the principal components of all sweet corn varieties, the 12 sweet corn varieties were divided into three categories (Figure 1): sensitive varieties (Zhongtian 7), intermediate varieties (Yuetian 28, Zhongtian 9, Ruizhen, Weitian 1, Fotian 2, Zhongxiantian 3, Kupula, Jinzhongyu) and tolerant varieties (Zhongxiantian 102, Zhuyutian 1, Fotian 10).

1.3 Cadmium stress destroyed the chlorophyll stability of sweet corn seedlings

In order to further explore the response differences of sweet corn varieties with different cadmium tolerance to cadmium stress, ‘Zhuyutian 1’ (tolerant varieties) and ‘Zhongtian 7’ (sensitive varieties) were taken as examples. After 3 days’ treatment, the phenotypes of the two varieties were significantly different, and there was no significant difference between the phenotypes of the tolerant variety ‘Zhuyutian 1’ before and after cadmium treatment; while after cadmium stress treatment, the seedlings of sensitive variety ‘Zhongtian 7’ showed obvious toxicity, mainly including growth retardation, slender stem, poor root development and obvious yellowing of the first leaf, and the control group grew well and had normal leaf color (Figure 2A). The changes of relative chlorophyll content (SPAD value) in the first leaf of different sweet corn varieties before and after cadmium stress treatment were further analyzed. The results showed that the SPAD value of tolerant variety ‘Zhuyutian 1’ had no significant difference before and after treatment (p>0.05) (Figure 2B). The SPAD value of sensitive variety ‘Zhongtian 7’ decreased gradually with the increase of cadmium treatment time, and basically reached the lowest point on the third day, which was very significantly different from the control group (p<0.01). The change law of relative chlorophyll content among different varieties was consistent with the phenotype of the change of leaf color, that is, the first leaf of sensitive variety ‘Zhongtian 7’ turned yellow and began to wilt after 3 days of cadmium treatment (Figure 2).

1.4 Cadmium stress inhibited the biomass accumulation of sweet corn seedlings

The decrease of relative chlorophyll content limited photosynthesis and the accumulation of photosynthetic products to a certain extent. The difference of biomass accumulation between the two varieties before and after cadmium stress treatment was analyzed (Figure 3). The results showed that both fresh weight (shoot fresh weight, root fresh weight and total fresh weight) and dry weight (shoot dry weight, root dry weight and total dry weight) showed a significant downward trend in sensitive variety ‘Zhongtian 7’ after cadmium treatment for 3 days, with a decrease range of 15.29%~25.95%, and reached a significant level (p<0.05). Among them, the root dry weight was the most affected by cadmium stress, with an average value of 0.032 g, while the average value of the control group was 0.044 g, which decreased by 27.3% (Figure 3B). In contrast, the tolerant variety ‘Zhuyutian 1’ was not affected by cadmium stress, the difference was not significant, and even showed a certain degree of promotion, among which the shoot dry weight was the most obvious, with an increase of 11.45%. It can be seen that cadmium stress destroyed the stability of chlorophyll to a certain extent, affected the normal photosynthesis of plants, and then reduced the accumulation of plant biomass and inhibited the process of growth.

1.5 Cadmium stress affected the growth of sweet corn seedlings

In order to further explore the reasons for the difference of biomass, the seedling height, main root length and number of total roots were measured. Compared with the control group, the sensitive variety ‘Zhongtian 7’ was significantly inhibited in both plant height and root length after 3 days of cadmium treatment, reducing by 17.23% and 29.89% respectively (Table 4). Due to the large variation range of the number of total roots, although it also decreased by 16.88%, the statistical analysis was not significant. However, the differences of seedling height, root length and number of total roots of tolerant variety ‘Zhuyutian 1’ before and after cadmium treatment did not reach a significant level (Table 4). These results showed that the tolerant materials had a certain ability to adapt to cadmium ion stress and could maintain normal growth in a short time, while the sensitive materials showed yellowing of leaves and inhibition of growth after cadmium stress, resulting in the reduction of biomass accumulation.

2 Discussion

Cadmium is a kind of heavy metal. Previous studies have shown that it has strong toxicity to a variety of plants, and different varieties of the same plant often show different degrees of tolerance due to differences in genetic characteristics (Hu et al., 2019). This study found that the variation coefficient of cadmium tolerance index of number of total roots, root length, root fresh weight, root dry weight and SPAD value of 12 sweet corn varieties was large (Table 1), which was similar to the research results of Gu et al. (2014) on ordinary corn. These results showed that the genetic differences among varieties were also important factors affecting the tolerance of corn to cadmium stress. Therefore, it is possible to screen excellent sweet corn varieties with strong cadmium tolerance among the existing varieties to solve the problem of cadmium toxicity in polluted farmland.

A large number of studies have found that plants are extremely sensitive to cadmium stress at seedling stage, and a series of abnormal changes in growth will occur after cadmium poisoning, for example, the absorption, transportation and metabolism of mineral elements and water will be hindered (Fan et al., 2018), the structure of coleoptile tissue will be necrotic (Wahid and Khaliq, 2005), the structure of mitochondria and enzymes related to respiration may be irreversibly damaged, and soluble sugar content and peroxidase activity in cells will also change significantly (He et al., 2020). It can be seen that cadmium toxicity to plants and plant resistance to cadmium stress is a complex physiological process, which can not be evaluated by a single index. Tian et al. (2018) found that there was a considerable overlap in the information provided by the cadmium tolerance index of each investigation index when studying the cadmium tolerance response characteristics of Iris lactea var. chinensis at seedling stage. The comprehensive evaluation index generated by multiple indexes should be used to evaluate the cadmium tolerance to improve the reliability of the conclusion. In this study, through the correlation analysis of cadmium tolerance indexes of 8 characters, such as seedling height, root length, number of total roots, shoot fresh weight, shoot dry weight, root fresh weight, root dry weight and relative chlorophyll content, it was found that there was a generally significant or extremely significant correlation between cadmium tolerance indexes of each character (Table 2). Further principal component analysis of the tolerance indexes of these 8 indexes showed that the eight single indexes in the seedling stage of sweet corn under cadmium stress were transformed into one comprehensive index. The contribution rate of the comprehensive index and the load of the original eight single indexes were higher, which objectively and comprehensively reflected the differences in cadmium tolerance of different varieties of sweet corn (Table 3). Then, using the scores of principal components for cluster analysis, 12 sweet corn varieties can be divided into three types: sensitive varieties (1), intermediate varieties (8) and tolerant varieties (3) (Figure 1).

Anjum et al. (2016) found that cadmium stress significantly inhibited the shoot growth of corn. Vaculík et al. (2015) observed that the photosynthetic organ structure of leaves was damaged after cadmium treatment, the synthesis of chlorophyll a and chlorophyll b was affected, and the chlorophyll content decreased with the increase of cadmium treatment concentration. Fan et al. (2018) believed that cadmium stress would lead to suberization and dark brown of corn root tissue and inhibit the differentiation and development of lateral root primordia. The results of this study showed that the seedlings of sensitive sweet corn varieties were greatly affected by cadmium stress, the phenomenon of leaf chlorosis was the most obvious, and the relative content of chlorophyll was significantly reduced, resulting in the inhibition of photosynthesis, then affecting the accumulation of biomass and restricting the growth (Figure 2; Figure 3; Table 4). Among them, seedling height and root length were the most sensitive to cadmium stress. Compared with the control group, the root length was affected more than the seedling height, and the possible reason was that the root, as an organ in direct contact with heavy metal ions, was first injured. In addition, some studies have shown that the roots produce ethylene and transport it to the shoot under the stress of cadmium. The ethylene stress is highly toxic to cells, and the roots, as the organ producing ethylene, may also be the first to be damaged (Dubois et al., 2018). In addition, the total number of roots of plants also decreased by 16.88% after cadmium stress (Table 4), but it has not reached a significant level, which may be due to the short treatment time of 3 days in this experiment, resulting in the failure of inhibition.

Under cadmium stress, plants have the effect of “high concentration inhibiting growth and low concentration promoting growth” (Zhang et al., 2019; Wang et al., 2019). In this study, sweet corn varieties with different tolerance were treated with the same concentration of cadmium ion. It was found that the concentration in this experiment had a significant inhibitory effect on sensitive varieties, and each biomass index of tolerant varieties increased to varying degrees, with the increase range of 0.99%~11.45% (Table 1; Table 4). This showed that the concentration of this experiment was high for sensitive varieties, but low for tolerant varieties. Therefore, it can inhibit the growth of sensitive varieties but promote the growth of tolerant varieties to a certain extent. It can be seen that the effect of cadmium stress on the growth of sweet corn seedlings was not only related to its treatment concentration and time, but also genetic factors. Therefore, screening and planting tolerant varieties is conducive to the promotion and quality improvement of sweet corn in southern China.

3 Materials and Methods

3.1 Test materials

The tested materials were 12 sweet corn varieties mainly popularized in South China (Table 5), with wide genetic background and strong representativeness. All seeds were provided by Guangzhou Key Laboratory for Research and Development of Crop Germplasm Resources.

3.2 Test design

The germination, growth and treatment of all corn seeds were carried out in an artificial climate chamber. The day and night temperature of the incubator was 28℃/25℃, the day and night duration was 16 h/8 h, and the relative humidity was 80%.

Firstly, the selected sweet corn seeds were disinfected with 1% sodium hypochlorite solution for 10 minutes, then washed with sterilized distilled water, soaked for 2 hours and placed in the germination plate. After germination for 48 hours, seedlings with consistent growth were selected for transplanting. Sand culture was selected for planting, and each culture box (specification: 17 cm×11 cm×5 cm) was filled with 700 g of quartz sand sterilized by high temperature, and 12 plants were transplanted in each pot. Corn seedlings were cultured to the third leaf stage and subjected to cadmium (CdCl₂·5/2 H₂O) stress treatment at the concentration of 0.01 mol/L (40 mL). Distilled water treatment was used as the control group. Each experiment was repeated three times and a randomized block design was used.

3.3 Phenotypic determination and methods

After cadmium treatment, take photos to record the growth changes of corn seedlings, and compare and analyze the phenotypic differences of the same variety before and after treatment and among different varieties. The SPAD value of the first fully expanded leaf of corn seedling was measured by SPAD-502Plus chlorophyll content tester, once a day, with 3 repetitions each time. After cadmium treatment for 3 days, wash the sandy soil, measure the seedling height and main root length with the position of seed embryo as the boundary, and count the total number of roots. All plants were divided into shoot and root parts. All roots were counted as root parts, and the rest were shoot parts. After absorbing all residual water with absorbent paper, weigh the fresh weight of the shoot part and the root part with an electronic balance with an accuracy of 0.01 g. Then put them into the oven for deactivation of enzymes at 105℃ for 20 minutes, dry them at 70℃ to constant weight, and measure the dry weight of the shoot and root parts.

3.4 Data processing

Due to the large inherent genetic differences among different varieties of sweet corn, the cadmium tolerance index proposed by Piotto et al. (2018) was used to measure the differences in cadmium tolerance among different varieties. The calculation formula was:

Cadmium tolerance index = measured value of cadmium treatment group/measured value of control group

Excel 2010 was used to sort out routine data; SPSS 21.0 was used for significance test, correlation analysis, principal component analysis and cluster analysis; Origin 8.0 was used for mapping.

Authors’ contributions

SW and LPF were the designers and the persons in charge of the project, guiding the experimental design, data analysis, manuscript writing and revision; XTC and SHY were the experimental designers and executors of this study; XTC and HY completed data analysis and wrote the first draft of the manuscript; CLJ, GZW and LSY participated in the experimental design, research implementation and experimental result analysis; All authors read and approved the final manuscript.

Acknowledgments

This study was jointly funded by the Research and Development Program in Key Areas of Guangdong Province (2018B020202013), the Project of Guangzhou Key Laboratory for Research and Development of Crop Germplasm Resources (202002010010) and the 2019 Provincial College Students’ Innovation and Entrepreneurship Training Program of Guangdong Province (S201911347037).

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