3.1. Distribution of Cadmium in Various Parts of Rice
The yield of the rice grain is lower under flooding condition than that under intermittent condition. The yield of the rice grain was increased and changed slightly with liming, compared to the control (Figure 1
The allocation of Cd concentration in rice is shown in Figure 1
. Compared to intermittent conditions, the Cd concentration in various parts of rice was decrease slightly under flooding conditions. After liming, the allocation of Cd in rice was significantly different during the different growth stages of rice. At the filling stage of rice, compared to control, the Cd concentration in root was significantly decreased by 33.3% in the intermittent combined with lime treatment (IL), while the corresponding figure was 30.3% in the flooding combined with lime treatment(FL). The Cd concentration of straw was decreased and changed slightly among treatments. At the maturity stage of rice, Cd uptake by rice straw significantly reduced by 40.3% in the IL treatment and 41.7% in the FL treatment, respectively, compared to control. Similarly, compared to control, rice Cd concentration reduced by 34.9% (decreasing from 0.86 to 0.56 mg/kg) in the IL treatment and 55.8% (decreasing from 0.77 to 0.34 mg/kg) in the FL treatment, respectively.
Normally, potentially toxic element concentrations in brown rice are significantly affected by straw and rice root concentrations [51
]. Previous studies have shown that the Cd concentration in rice root and straw was significantly decreased by application of water regimes or amendment [19
]. With liming, Cd concentration in the root decreased significantly at the filling stages of rice, whereas at maturity stage there was lower Cd accumulation in straw than that in root. The results show that the pregnant stage of rice is the critical periods for controlling rice Cd concentration. Meanwhile, to reduce rice Cd concentration, controlling Cd accumulation in root at the filling stage of rice should be prior considered. In the current study, liming at tillering stage of rice is an effective way to decrease rice Cd concentration under flooding condition.
3.2. Soil pH and Availability of Cadmium
Soil pH is an important factor governing solid-solution equilibria of potentially toxic elements [55
]. As shown in Table 1
, the value of pH in flooding soil was higher than that in intermittent soil. Compared to control, soil pH significantly increased (p
< 0.05) with liming under intermittent and flooding conditions, respectively, because the release of hydroxyl ions through the hydrolysis of lime neutralized the acidity soil. Soil pH was correlated negatively with the availability of soil Cd. The degree of Cd uptake by plants depends on their availability, while DTPA-extractable Cd is suitable for predicting the availability of Cd in soil [24
Soil DTPA-extractable Cd concentration decreased during growth stages of rice (Table 3
). Flooding soil had lower DTPA-extractable Cd concentration than intermittently wetted soil. Compared to intermittent conditions, the DTPA-extractable Cd concentration under flooding treatment was reduced significantly by 24.5% at the filling stage of rice (p
< 0.05). At filling stage of rice, DTPA-extractable Cd reduced significantly by 18.4% in IL treatment compared to control. At maturity stage of rice, soil DTPA-extractable Cd reduced significantly by 23.0% in IL treatment and 21.6% in FL treatment compared to control, respectively.
Previous studies also stated that flooding conditions generally decreased Cd availability in tested soil [18
]. Lime combined with flooding conditions was a more suitable way to reduce soil Cd availability than flooding conditions alone. Several mechanisms have been attributed to the soil Cd availability. First, the increase of pH led to an increase in negative charges of soil under the flooding condition alone or combined with liming and it could also hydrolyze Cd2+
, which Cd in soil precipitates as hydroxides or carbonates and adsorbs tightly to soil colloid, ultimately, leading to lower availability [34
]. Second, the concentrations of iron and manganese oxides in flooding soil have been decreased, while that of mobile Cd in soil increased, which can lead to the immobilization of Cd by readsorption or precipitation [61
]. Finally, microorganisms in flooded soil, such as sulfur-reducing bacteria that can reduce sulfates to sulfide or S2−
which then reacts with Cd2+
to form CdS precipitates, can also reduce the availability of Cd [63
In addition, after liming, the decrease amplitude of DTPA-extractable Cd concentrations is higher at the maturity stage of rice than that filling stage under flooding conditions. The Cd concentration is low at the maturity stage of rice, indicating that a new equilibrium was established between the different Cd forms in soil, which may be closely associated with soil properties, temperature and rhizosphere environment at different stages of rice. The specific reason needs further research.
Sequential extraction is often to study the relative bioavailability of soil-sorbed potentially toxic elements by revealing the speciation of the elements in soil [42
]. Compared to intermittent wetting, at the maturity stage of rice the proportion of acid extractable Cd decreased significantly by 44.4% in flooded soil while reducible Cd and oxidizable Cd increased by 59.8% and 78.6%, respectively (Figure 2
). At the filling stage of rice, the proportions of acid extractable Cd decreased significantly by 22.6% in IL soil and 5.4% in FL soil, while reducible Cd increased by 40.4% and 13.6%, compared to control, respectively. At the maturity stage of rice, the proportions of acid extractable Cd decreased significantly by 47.1% in IL soil and 23.0% in FL soil while residual Cd increased 15.0% and 16.6%, compared to control, respectively. The results indicate that the Cd fractions in soil were closely related to the duration of flooding. Liming promoted the transformation of Cd in soil from acid-extractable to reducible form at the filling stage of rice and to residual fraction at the maturity stage of rice. These results were consistent with Chen, who report that liming can was a suitable way to decrease Cd availability and increased stable fractions under flooding condition [34
]. Huang et al. [66
] also reported that the combination of moisture management and amendment promoted the transformation of Cd in red paddy soil from acid-extractable to reducible fraction.
3.3. Soil Enzyme Activity and Microbial Characteristics
Soil enzyme activity and microbial community have been used to evaluate the soil quality following soil remediation activities [37
]. Soil urease and invertase activities were reduced by 15.8% and 6.5% under flooding conditions, respectively, compared to intermittent conditions (Table 4
). After liming, soil enzyme activity was increased. Phosphatase, urease, and invertase activities in the IL soil were significantly increased by 116.7%, 61.4% and 28.8%, compared to control, respectively. Similarly, in the FL soil, soil urease activity increased by 46.5%, that of acid phosphatase was 41.3%, and that of invertase was 20.8% compared to control, respectively.
Previous studies had also reported that soil enzyme activities were negatively correlated with soil moisture, which was due to the low redox potential and anaerobic soil conditions [67
]. The activity of soil enzymes was higher after lime treatment, indicating that a certain degree of metabolic recovery was related to the liming of Cd-contaminated soil. Sun et al. [38
] reported that application of sepiolite significantly increased soil enzyme activity and presumed the changes in pH may be primarily responsible for this behavior. However, enzyme activity may also change under potentially toxic element stress [68
]. In our study, liming changed significantly Cd stress level in the soil, which is another factor that can influence enzyme activity. In addition, the high invertase and urease activities in soils indicated the rich functional state of the soil. Urease activity increased significantly with liming while the invertase changed slightly (Table 4
). The results can contribute to the reasons as follows: one the hand, urease activity was significantly affected by the level of contamination due to urease could be combined with soil main component of humus to form stable compounds outside the cells [70
]. On the other hand, urease is an extracellular enzyme and inhibited by metal ions through reaction with the sulfhydryl groups, synthesis of metal-sequestering saccharides or proteins and trapping or precipitation of metals on microbial surfaces [72
]. Therefore, urease has the potential to be used to assess soil recovery for the remediation of potentially toxic elements in contaminated soil.
The composition of the bacterial community plays a role in determining the intrinsic stability of soil microbial communities [74
]. DGGE, as a microbial diversity screening method, can monitor the changes of microbial community response at the molecular level [75
]. The DGGE band pattern of 16S rDNA amplified by primers 357f-gc and 517R amplification of was used to determine the bacterial community, as shown in Figure 3
. The DGGE profiles of bacteria were basically similar after the four treatments, suggesting that the microorganisms with these bands were relatively stable and less affected by the treatments such as liming or water regimes. However, there were still a few bands that emerged or vanished with liming. The changes of bacterial community in soil were presented by the DGGE profiles, and the number of bands in the DGGE patterns increased with liming. In particular, the band number significantly increased with the IL treatment, as shown in Table 2
. The Shannon index indicated that bacterial community diversity was slightly reduced in flooding soil, compared to intermittent, while liming significantly increased bacterial community diversity. Soil moisture has consistently been shown to be strongly correlated with the variation in the microbial community [67
]. The highest band number and Shannon index can be obtained by application of lime, which could be attributed to the high pH and low toxicity of potentially toxic elements [67
] or might be relevant for the replication of new bacterial species. A study of the changes of the specific bacteria in soil will be determined by high-throughput sequencing technique. The results indicate that lime combined with various water regimes is favorable to improve soil environmental quality.