Next Article in Journal
Forage Plant Ecophysiology under Different Stress Conditions
Previous Article in Journal
The Taxonomic Circumscription and Nomenclatural History of Pilosella suecica (Asteraceae): A Special Case of Grey Literature in Taxonomic Botany
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Plants
Volume 13
Issue 10
10.3390/plants13101303
plants-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Microbiological Mechanisms of Collaborative Remediation of Cadmium-Contaminated Soil with Bacillus cereus and Lawn Plants

by
Beibei Zhou
*,
Zehao Yang
,
Xiaopeng Chen
,
Ruonan Jia
,
Shaoxiong Yao
,
Bin Gan
,
Dongliang Fan
,
Xie Yang
,
Wenqian Li
and
Yunhan Chen
State Key Laboratory of Eco-Hydraulics in Northwest Arid Region of China, Xi’an University of Technology, Xi’an 710048, China
*
Author to whom correspondence should be addressed.
Plants 2024, 13(10), 1303; https://doi.org/10.3390/plants13101303
Submission received: 9 April 2024 / Revised: 3 May 2024 / Accepted: 7 May 2024 / Published: 9 May 2024
(This article belongs to the Special Issue New Insight into Cadmium Phytoremediation – Volume II)
Browse Figures
Versions Notes

Abstract

:
Severe cadmium contamination poses a serious threat to food security and human health. Plant–microbial combined remediation represents a potential technique for reducing heavy metals in soil. The main objective of this study is to explore the remediation mechanism of cadmium-contaminated soil using a combined approach of lawn plants and microbes. The target bacterium Bacillus cereus was selected from cadmium-contaminated soil in mining areas, and two lawn plants (Festuca arundinacea A‘rid III’ and Poa pratensis M‘idnight II’) were chosen as the target plants. We investigated the remediation effect of different concentrations of bacterial solution on cadmium-contaminated soil using two lawn plants through pot experiments, as well as the impact on the soil microbial community structure. The results demonstrate that Bacillus cereus promotes plant growth, and the combined action of lawn plants and Bacillus cereus improves soil quality, enhancing the bioavailability of cadmium in the soil. At a bacterial suspension concentration of 105 CFU/mL, the optimal remediation treatment was observed. The removal efficiency of cadmium in the soil under Festuca arundinacea and Poa pratensis treatments reached 33.69% and 33.33%, respectively. Additionally, the content of bioavailable cadmium in the rhizosphere soil increased by up to 13.43% and 26.54%, respectively. Bacillus cereus increased the bacterial diversity in the non-rhizosphere soil of both lawn plants but reduced it in the rhizosphere soil. Additionally, the relative abundance of Actinobacteriota and Firmicutes, which have potential for heavy metal remediation, increased after the application of the bacterial solution. This study demonstrates that Bacillus cereus can enhance the potential of lawn plants to remediate cadmium-contaminated soil and reshape the microbial communities in both rhizosphere and non-rhizosphere soils.

1. Introduction

Industrial production activities (such as mining, sewage irrigation, and metal smelting) have made soil heavy metal pollution one of the most important global environmental problems [1]. Heavy metal pollution has been observed to reduce the total biomass, diversity, and activity of soil microbial communities and may alter their composition [2]. The contamination of soil by cadmium has emerged as a significant and pervasive environmental issue [3]. Cadmium (Cd) is a toxic metal that threatens the human food chain [4]. Cadmium contamination not only diminishes soil productivity, leading to soil degradation, but also poses a severe threat to food security, human health, and the natural environment. Therefore, the remediation of cadmium-contaminated soil is an urgent issue [5].
Numerous technologies for the remediation of contaminated soil have been reported, such as isolation/containment technologies, solidification/stabilization technologies, and electrokinetic remediation; however, these technologies have many drawbacks, including high input, secondary pollution, and soil deterioration [2]. Additionally, most of these engineering-based technologies are not suitable for repairing large areas of contaminated soil [6]. As traditional remediation techniques like physical and chemical methods are increasingly limited in addressing soil heavy metal pollution, phytoremediation has gained significant attention as an alternative due to its ecological, economic, and sustainable advantages [7], emerging as one of the most promising remediation approaches. The key advantages of phytoremediation are lower maintenance costs, ease of application, and a public perception of this technique as a ‘green’ approach. Phytoremediation is beneficial when dealing with larger areas or when other restoration approaches might be too costly or impractical [8]. In addition, plants play a vital role in improving soil quality and optimizing the soil microbial community. Various microorganisms living in the rhizosphere can have beneficial effects on plant growth and health and increase plant biomass production [9]. Therefore, it is very important to study the changes in soil nutrients and microbial community structure during phytoremediation.
Plants are known to have different tolerances to environmental stress [10]. Festuca arundinacea and Poa pratensis are grasses belonging to the Poaceae family. They possess characteristics such as rapid growth, large biomass, wide distribution, a strong regenerative ability, and well-developed underground rhizomes and root systems. They can effectively reduce the loss and spread of pollutants [11]. In addition, Festuca arundinacea and Poa pratensis not only contribute to the remediation and beautification of the environment but also provide landscape and ecological benefits. This approach results in both esthetic and ecological benefits. The characteristic of phytoremediation is to utilize green plants to transfer/transform pollutants from the environment or render them harmless. However, the effectiveness of phytoremediation is limited due to low biomass, slow plant growth, and the low bioavailability of metals in soil [12].
Bioremediation of contaminated soil through microbial activity is a promising strategy [13]. In recent years, a large number of microorganisms with the potential to absorb Cd2+ have been discovered. The microorganisms detected in various studies include Staphylococcus cohnii L1-N1, Bacillus cereus CKN12 [14], Microbacterium sp. D2-2, and Bacillus sp. C9-3 [15], as well as Enterobacter bugandensis XY1 and Serratia marcescens X43 [16]. Various microbial detoxification mechanisms, including bioadsorption, bioreduction, and bioaccumulation, have been proven effective in reducing the toxicity of Cd [17]. However, similar to phytoremediation techniques, microbial remediation is also greatly influenced by external environmental factors, exhibits poor genetic stability, and demonstrates strong host specificity, leading to incomplete treatment effectiveness against various types of pollutants.
From this perspective, it is evident that individual remediation methods have limitations. Therefore, utilizing combined remediation technologies is gradually becoming a forefront hotspot. Microbially assisted phytoremediation represents a highly promising technological approach that has the potential to significantly augment the capacity of plants to uptake heavy metals [18,19]. According to a study conducted by Lanping et al. [20], Lolium perenne combined with heavy-metal-tolerant microorganisms can enhance the absorption of cadmium, which suggest that heavy-metal-tolerant bacteria may have the potential to reduce cadmium toxicity or enhance the tolerance of Lolium perenne to cadmium. Additionally, other microorganisms were proven to alter the solubility and bioavailability of soil heavy metals, thereby significantly increasing the metal concentrations in roots and shoots [21], which has great potential in the application of phytoremediation. However, the synergistic remediation of heavy-metal-contaminated soil by lawn plants and Bacillus cereus was not fully studied.
Therefore, in this study, we focus on the impact of microbial application on the remediation of heavy-metal-contaminated soil by lawn plants. Festuca arundinacea and Poa pratensis as green plants, alongside the cadmium-tolerant strain Bacillus cereus, are employed as a bacterial inoculant. The purpose of this study is (I) to evaluate the remediation effect of lawn plants combined with Bacillus cereus on cadmium-contaminated soil by assessing changes in soil nutrient contents and cadmium bioavailability, as well as the distribution of Cd in lawn plants; (II) to investigate the metabolic functional diversity of microbial communities in rhizosphere and non-rhizosphere soils under conditions of bacterial liquid application; and (III) to examine the relationship between plant remediation and soil microbial ecological characteristics using redundancy analysis and cluster analysis.

2. Results and Discussion

2.1. The Impact of Bacillus cereus Application Rate on Cd-Contaminated Soil

2.1.1. The Effects of Bacillus cereus Application Rate on Soil Physicochemical Properties

Figure 1a summarizes the changes in soil physicochemical properties under different treatments. The application of Bacillus cereus increased soil pH, organic matter, total nitrogen, and available phosphorus. From Figure 1a, it can be observed that compared to the control treatments G0 and Z0, the soil pH values in all treatment groups slightly increased, indicating that the application of Bacillus cereus in the soil could generate more alkaline substances due to its intrinsic characteristics, thereby raising the soil pH values. After planting Festuca arundinacea and Poa pratensis, the soil total nitrogen and available phosphorus contents showed a trend of increasing first and then decreasing with the increase in the bacterial liquid concentration, but they were all higher than those in the control treatment. The application of Bacillus cereus can increase the soil available phosphorus content, possibly due to the enhancement of soil phosphatase activity by Bacillus cereus by promoting the conversion of organic phosphorus to available phosphorus in the soil, which is similar to the findings of Haroun et al. [22]. Many studies have reported that soil nutrient content is closely related to plant growth. In general, high soil nutrient levels regulate the plant photosynthetic rate, stomatal conductance, and transpiration rate, promoting plant growth [23]. The inoculation of Bacillus cereus increased the soil available potassium, available phosphorus, and organic matter [24]. In this study, the application of Bacillus cereus increased the soil pH, organic matter, total nitrogen, and available phosphorus (Figure 1a). This study suggests a positive correlation between available phosphorus concentration and carbon source utilization rate [25]; rhizosphere bacteria with high metabolic activity can metabolize phosphorus-containing organic compounds in the soil, converting ineffective organic phosphorus into usable forms. Previous studies have also shown that inoculation with certain microorganisms promotes soil fertility after 60 days of greenhouse incubation [26]. The increase in soil organic matter and available phosphorus is beneficial for plant growth, especially under heavy metal stress, as nutrient-deficient environments hinder plant growth and development, limiting the progress of phytoremediation.

2.1.2. The Effect of Bacillus cereus Application Rate on Soil Cd Content

From Figure 1b, it can be observed that different treatments are able to reduce soil Cd content to varying degrees, with overall performance indicating that Festuca arundinacea is less than Poa pratensis. In the treatment with Festuca arundinacea, compared to the control group G0, the soil Cd content decreased by 6.7%, 10.28%, 9.29%, and 6.94% in treatments G1, G2, G3, and G4, respectively; the soil Cd content was lowest at 1.46 mg/kg when the bacterial suspension concentration was 105 CFU/mL. In the Poa pratensis planting treatment, compared to control Z0, treatments Z1, Z2, Z3, and Z4, respectively, reduced the soil Cd content by 11.34%, 13.49%, 9.83%, and 5.96%. The application of bacterial solution can enhance the removal efficiency of plants for the soil heavy metal Cd, and the optimal removal effect is observed at a bacterial solution concentration of 105 CFU/mL; specifically, in the treatments with Festuca arundinacea (G2) and Poa pratensis (Z2), the removal rates reached 33.69% and 33.33%, respectively.

2.1.3. The Effect of Bacillus cereus Application Rate on the Forms of Cd in Rhizosphere Soil

The bioavailability of heavy metals in soil directly affects the adsorption efficiency of plants. The heavy metal Cd in soil commonly exists in four forms: acid-soluble (F1), reducible (F2), oxidizable (F3), and residual (F4) forms. As shown in Figure 1c, the percentage content of acid-soluble cadmium is highest in the rhizosphere soil under different treatments, while the percentage content of residual cadmium is lowest. The total proportion of acid-soluble and reducible cadmium ranges from 57.76 to 70.81%, while the total proportion of oxidizable and residual cadmium ranges from 29.19 to 42.24%. With the increase in bacterial liquid concentration, the total proportion of acid-soluble and reducible cadmium in the rhizosphere soil of both types of lawn plants showed a trend of first increasing and then decreasing, and it was higher than that of the control treatment.
It is worth noting that although the total cadmium concentration in the soil decreased, the application of Bacillus cereus actually increased the bioavailability of cadmium. The acid-soluble and reducible forms are highly mobile and readily utilizable by plants. After inoculation with Bacillus cereus, the total proportion of acid-soluble cadmium and reducible cadmium in the rhizosphere soil of the two lawn plants increased by up to 13.43% and 19.82%, respectively. The increase in the percentage of bioavailable cadmium can stimulate the accumulation of cadmium in plants because the limited availability of metals in contaminated soil often hinders the efficiency of phytoremediation. In many contaminated soils, most heavy metals are bound to various organic and inorganic compounds, making them inaccessible to plants, thus becoming the primary limiting factor in the phytoremediation process [27].

2.2. The Effect of Bacillus cereus Application Rate on Plant Growth and Cd Absorption

During the 60-day growth period, both types of lawn plants grew well in the cadmium-contaminated soil, especially when the bacterial solution concentration was 105 CFU/mL. As shown in Figure 2, the application of the bacterial solution significantly increased the aboveground and root biomass of both lawn plants. However, due to the differences in plant species and their tolerance to heavy metals, the changes in biomass varied among different plant parts. The aboveground and root biomass of Festuca arundinacea and Poa pratensis reached their maximum values at a bacterial solution concentration of 105 CFU/mL; in the Festuca arundinacea treatment, the aboveground biomass increased by 46.7% compared to the control, while the root biomass increased by 33.8%. In the Poa pratensis treatment, the biomass increased by 81.2% and 76.1%, respectively. It can be inferred from this that Festuca arundinacea exhibits better tolerance to Cd compared to Poa pratensis.
Additionally, the Cd concentrations in both lawn plants significantly increased (Table 1), with the Cd content in the aboveground parts of Festuca arundinacea being up to 2.61 times higher than that in control group G0, and the Cd concentration in the roots was up to 1.41 times higher; Poa pratensis showed a similar trend to Festuca arundinacea under bacterial inoculation treatment, with the highest Cd content observed in the aboveground and root parts when the bacterial suspension concentration was 105 CFU/mL, reaching 2.63 and 1.33 times that of control group Z0, respectively. The application of Bacillus cereus also facilitated the transport of Cd in both lawn plants and significantly increased the bioconcentration factor (BCF) of the two lawn plants. The bioconcentration factor of Festuca arundinacea increased to 0.607~0.906, while that of Poa pratensis increased to 0.423~0.519. This study demonstrates that the application of Bacillus cereus can enhance the transport of Cd to the aboveground parts of lawn plants and increase the storage of Cd within plant tissues, thereby achieving a reduction in soil Cd content through harvesting plant stems and leaves.
In a study conducted by Farh et al. [28], Staphylococcus and Bacillus were identified as plant growth-promoting bacteria (PGPB). These PGPB secrete growth hormones, including IAA, ACC, dehydrogenase, and iron carriers, which promote plant development [29]. After 60 days of cultivation, the application of Bacillus cereus indeed increased the biomass of both plant species, thereby enhancing the plants’ remediation efficiency (Table 1). The observed increase in lawn plant biomass in this study can be attributed to the secretion of IAA or the phosphate solubilization by Bacillus cereus. In a study conducted by Li et al. [30], it was observed that the inoculation of Bacillus cereus into alfalfa significantly increased plant biomass and enhanced the availability of cadmium (Cd) in soil, leading to improved efficiency of Cd removal. This study demonstrates that the application of Bacillus cereus can alter the bioavailability of metals and regulate metal uptake and translocation, thereby enhancing the absorption and accumulation of cadmium (Cd) in both lawn plants. The removal efficiency of heavy metals in soil is strongly influenced by the biomass of plants and the bioavailability of heavy metals. When the bioavailability of heavy metals increases, it can promote the uptake of these metals by plants, leading to higher concentrations of heavy metals in plant tissues. Therefore, this process enhances the ability of plants to extract heavy metals from the soil [31,32].
Previous studies have reported that the accumulation of Cd in plants largely depends on the bioavailable concentration of Cd. Ma et al. [33] demonstrated that inoculation with metal-resistant bacteria significantly promoted the accumulation of heavy metals in plants by improving the mobility of heavy metals in the soil. The application of Bacillus cereus can increase the content of acid-soluble and reducible cadmium in the soil, which may stimulate the accumulation of cadmium in both lawn plants (Figure 1c). The application of Bacillus cereus can activate heavy metals in rhizosphere soil, enhance the bioavailability of heavy metals in rhizosphere soil, and increase the migration ability of heavy metals, thereby enhancing the adsorption and accumulation of heavy metals by plants. Additionally, Sharma et al. [34] demonstrated that Bacillus spp. enhance the bioavailability of heavy metals by producing organic acids and polysaccharides. Previous studies have shown that the accumulation and uptake of heavy metals (HMs) by plants highly depend on their bioavailability in the soil [35]. Due to the increased bioavailability of HMs, the phytoremediation efficiency of sunflowers for uranium (U)- and cadmium (Cd)-contaminated soil significantly improved in pot experiments [36].

2.3. The Effect of Bacillus cereus Application Rate on the Microbial Structure of Heavy-Metal-Contaminated Soil

2.3.1. The Variation in Bacterial Community Alpha Diversity between Rhizosphere and Non-Rhizosphere Soil

Heavy metal pollution can continue to influence soil physicochemical properties, thereby affecting the community structure and functionality of soil microorganisms [37,38]. By calculating the Alpha diversity indices of OTU sequences in Cd-contaminated soil under different treatments, we analyzed the changes in microbial diversity and richness in both rhizosphere and non-rhizosphere soil induced by different bacterial liquid concentrations. As shown in Table 2, in non-rhizosphere soil of both lawn plants, the Chao1 index, Shannon index, and Simpson index increased overall under each bacterial treatment, indicating an improvement in the metabolic diversity of the treated samples. This might be attributed to the availability of soil nutrients and other soil environmental conditions, including higher nutrient content and enzyme activity in the soil [39]. In rhizosphere soil, under bacterial treatment for both lawn plants, there was no significant change in the Simpson index, but the Chao1 index and Shannon index decreased compared to the control treatment, indicating that the application of Bacillus cereus might reduce the richness of bacterial communities in rhizosphere soil. It is speculated that Bacillus cereus can rapidly adapt to the rhizosphere environment of plants, becoming a dominant microbial community, and inhibiting the growth and reproduction of most other microorganisms.

2.3.2. The Changes in the Relative Abundance of Bacterial Species

The relative abundance results of the top ten bacterial species at the phylum level across all samples indicate significant changes in the composition of microbial communities in both rhizospheric and non-rhizospheric soils following Bacillus cereus inoculation. The phyla Proteobacteria, Actinobacteriota, Bacteroidota, Firmicutes, and Patescibacteria are the most abundant across all samples. In the non-rhizospheric soil of Festuca arundinacea (Figure 3a), they collectively account for 90.27~95.48% of the total abundance. In the rhizospheric soil, they collectively account for 77.32~90.78% of the total abundance. In the non-rhizospheric soil of Poa pratensis (Figure 3b), their total abundance collectively ranges from 80.5 to 90.7%, while in the rhizospheric soil, their total abundance collectively ranges from 81.5 to 93.2%. However, the distribution of each phylum varies depending on different soil locations and treatments. The phylum Proteobacteria tends to be more enriched in the rhizosphere across all treatments. Proteobacteria, which are Gram-negative bacteria, are abundant in harsh environments. They are characterized by an outer layer of lipopolysaccharides in their cell wall, which helps to sequester heavy metals from the external environment, thereby reducing the harmful toxicity of heavy metals and conferring a high degree of tolerance to them [40,41,42]. Under different treatments, the changes in various phyla differ. As shown in Figure 3a, compared to control G0, the relative abundance of Actinobacteriota decreased in both rhizosphere and non-rhizosphere soils of Festuca arundinacea after the application of the bacterial solution. Conversely, the relative total abundance of Proteobacteria, Firmicutes, and Bacteroidota increased. With the increasing bacterial solution concentration, the relative total abundance showed a trend of initially increasing and then decreasing. As depicted in Figure 3b, the relative abundance of Actinobacteriota decreased in both rhizosphere and non-rhizosphere soils of Poa pratensis after the application of the bacterial solution, similar to Festuca arundinacea. This may be attributed to the decrease in the soil cadmium concentration after the remediation treatment, as the abundance of Actinobacteriota in the soil is positively correlated with the concentration of heavy metals [43]. Actinobacteria, as rhizospheric bacteria, have been widely studied. They are capable of decomposing various organic compounds and utilizing different carbohydrates as energy sources [44].
In order to further understand the correlation within species, a hierarchical clustering heatmap analysis was conducted to illustrate the microbial community profile at the genus level (Figure 4). The heatmap displays an overview of the 30 most abundant genera, illustrating the impact of the inoculation on the composition of both rhizospheric and non-rhizospheric bacterial communities. Based on the relative abundance results of bacterial phyla, most bacteria in each treatment belong to Proteobacteria and Firmicutes; the composition of bacterial communities changed significantly with the application of bacterial liquid treatments. In the non-rhizospheric area of Festuca arundinacea (Figure 4a), compared to the G0 treatment, the relative abundance of Pseudarthrobacter, Klebiella, and Lactobacillus decreased after the application of bacterial liquid. Conversely, the relative abundance of genera such as Sphingomonas, Mucilaginibacter, Ramlibacter, Massilia, Gemmatimonas, and Microvirga increased. Research has found that Sphingomonas can promote the secretion of plant hormones such as indole-3-acetic acid, participate in nitrogen cycling, and be applied to the degradation of pollutants and the remediation of heavy-metal-contaminated soils [45,46]. In the rhizospheric area of Festuca arundinacea, following the application of the bacterial solution, there was an increase in the relative abundance of genera such as Lactobacillus (which produces lactic acid as a byproduct of glucose metabolism), Pseudarthrobacter (known for pollutant degradation), Sinomonas, and Bifidobacterium (beneficial bacteria capable of synthesizing various vitamins).
Additionally, the distribution of the microbial community structure responded differently to the simultaneous application of different concentrations of bacterial solution. In the non-rhizospheric area of Poa pratensis (Figure 4b), after the application of the bacterial solution, the relative abundance of dominant genera such as Bacteroides, Faecalibacterium, Bifidobacterium, and Calorithrix decreased in the Z0 treatment. The distribution of the microbial community structure in the Z1 treatment mainly focused on Chitinophagaceae and Parasegetibacter (Cd-tolerant, promoting plant growth), while in the Z2 treatment, it mainly focused on Ramlibacter and Massilia. In the Z3 treatment, it mainly concentrated on Pseudarthrobacter, WPS-2, Mucilaginibacter (Cd-resistant strains), Clostridia_UCG-014, and Ruminococcus. In the rhizospheric area of Poa pratensis, after the Z1 treatment, the relative abundance of Microvirga and Lactobacillus increased compared to the Z0 control. Following the Z2 treatment, Massilia (with a phosphorus-solubilizing ability, metal-tolerant) and Ramlibacter (beneficial bacteria) showed increased abundance compared to the control. After the Z3 and Z4 treatments, the abundance of Acinetobacter, Pseudarthrobacter (pollutant-degrading bacteria), Sinomonas, and Mucilaginibacter (metal-tolerant strains, involved in the remediation of heavy metal pollution) increased compared to the control. These research findings indicate that the application of Bacillus cereus leads to increased diversity among bacterial populations. This diversity promotes the stability of microbial communities, thereby aiding in alleviating the toxicity of heavy-metal-contaminated soils.

2.3.3. Biomarker Identification and LEfSe Analysis

Using the LEfSe 1.1.01 software, bacterial biomarkers at the phylum to genus level were identified for different treatments, including the cladogram (Figure 5a) and the LDA distribution histogram (Figure 5b). The results indicate significant differences in the enriched species communities between the rhizospheric and non-rhizospheric zones of Festuca arundinacea and Poa pratensis. Taxa such as Firmicutes, Clostridia, Negativicutes, Clostridiales, Lachnospiraceae, Prevotellaceae, Oscillospirales, Clostridiaceae, Lachnospiraceae, Tannerellaceae, Prevotella, Parabacteroides, Roseburia, Blautia, and Clostridium_sensu_stricto_1 were specifically enriched in the non-rhizospheric soil of Festuca arundinacea. Actinobacteriota and Chloroflexi phyla, as well as the Actinobacteria class, the TK10 order, the Catenulisporales order, the TK10 family, the Dysgonomonadaceae family, the Catenulisporaceae family, the Lactobacillaceae family, the Catenulispora genus, the Dysgonomonas genus, the Lactobacillus genus, and the Gemmatimonas genus are specifically enriched in the rhizosphere soil of Festuca arundinacea. The family Spirosomaceae and the genera Chitinophagaceae, Dyadobacter, and Sphingobacterium were specifically enriched in the non-rhizospheric soil of Poa pratensis. The genera Polyangia and Lysobacter, along with the class Alphaproteobacteria, the order Haliangiales, the family Haliangiaceae, and the family Xanthomonadaceae, were specifically enriched in the rhizospheric soil of Poa pratensis. These results indicate that different treatments lead to differences in the indigenous microbial taxa and their activity expression between rhizospheric and non-rhizospheric soils.

2.4. The Response Relationship between Plant Remediation and Soil Microbial Ecological Characteristics

The relationship between plant remediation and plant microecological characteristics is closely associated with soil physicochemical properties, heavy metal content, and bacterial community structure. To fully evaluate the relative impacts of soil physicochemical parameters (pH, OM, TN, AP) and heavy metal Cd concentration, as well as heavy metal forms (F1, F2, F3, F4), on the abundance and diversity of microbial communities in heavy-metal-contaminated farmland, redundancy analysis (RDA) was conducted (Figure 6). The results indicate that pH, OM, TN, AP, and Cd, as well as Cd forms F3 and F4, are the main factors influencing the bacterial community structure in Festuca arundinacea (p < 0.05) (Figure 6a,b). The main factors influencing the bacterial community structure in Poa pratensis are pH, TN, AP, and Cd, as well as Cd forms F2, F3, and F4 (p < 0.05) (Figure 6c,d). The results indicate that the soil microbial community structure is driven by soil nutrients such as TN and AP [47].
By combining the Spearman correlation heatmap with UPGMA clustering analysis, we investigated the relationship between soil properties and soil microbial community composition at the phylum level in non-rhizosphere and rhizosphere soils (Figure 7). In Festuca arundinacea soil, TN, pH, and AP were significantly positively correlated with Bacteroidota (p < 0.05), while soil AP and OM were significantly positively correlated with Firmicutes. Soil Cd content showed a significant negative correlation with Firmicutes and Gemmatimonadota (Figure 7a). In Poa pratensis soil, TN, pH, and AP were identified as key environmental factors positively influencing Firmicutes and Patescibacteria (p < 0.05). Soil T-Cd content showed a negative impact on Bacteroidota, exhibiting a significant negative correlation (Figure 7b). In the rhizospheric soil of Festuca arundinacea, oxidizable Cd content (F2) and reducible Cd content (F3) were significantly positively correlated with Proteobacteria. Residual Cd content (F4) showed a significant positive correlation with Firmicutes and Bacteroidota (Figure 7c). In the rhizospheric soil of Poa pratensis, the oxidizable Cd content (F2) showed a significantly positive correlation with Proteobacteria (p < 0.01) and a significant negative correlation with Bacteroidota (Figure 7d).
The interaction between roots and microbes plays a significant role in the phytoremediation of heavy-metal-contaminated soil. Microbes are pivotal players in the bioremediation of polluted soil, with certain functional microbes capable of reducing soil contamination levels [48]. Several growth parameters of the plant H. Pennisetum, including shoot length, biomass, and dry weight, indicate that inoculation with Bacillus significantly improves plant growth in the soil [49]. Compared to the CK treatment, the application of Bacillus cereus increased the diversity of bacterial communities in the non-rhizospheric soil of the lawn plants but reduced the richness of bacterial communities in the rhizospheric soil (Table 2). Meanwhile, the application of Bacillus cereus significantly increased the relative abundance of the phyla Proteobacteria and Firmicutes in the soil (Figure 3). Proteobacteria and Firmicutes have also been found to be dominant phyla in mine soil due to their possession of heavy-metal-tolerant gene clusters [50]; the ability of these phyla to coexist in extreme environments is believed to have the potential for remediating heavy metal pollution. In conclusion, Bacillus cereus significantly increased the relative abundance of dominant genera associated with improving soil nutrients, reducing soil Cd concentration, and altering the form of heavy metals. These factors may partly explain how the application of Bacillus cereus promotes the growth of two types of lawn plants and enhances their uptake of Cd, thus contributing to the increased efficiency of microbial-enhanced phytoremediation of Cd-contaminated soil.

3. Materials and Methods

3.1. Experimental Materials

The soil used in this experiment was collected from the 0–20 cm layer of the heavily metal-contaminated area in Dafanshan Mining Area, Lujiang County, Anhui Province. The soil texture is silty clayey loam (13.25% clay, 52.84% silt, and 22.48% sand). The soil was passed through a 2 mm sieve and then placed in sealed bags for storage. The basic physicochemical properties of the initial soil samples at the sampling points are shown in Table 3. The strains were screened from the soil samples of this experiment and identified as Bacillus cereus [51]. Our experiment shows that this strain can grow well in a medium with a cadmium concentration of 50 mg/L and exhibits excellent tolerance to high cadmium concentrations.
In the pot experiment, two common lawn plants were selected as the test subjects: Festuca arundinacea ‘Arid Ⅲ’ and Poa pratensis ‘Midnight Ⅱ’.

3.2. Experimental Design and Sample Collection

The pot experiment was conducted in the greenhouse of Xi’an University of Technology, located in Xi’an City, Shaanxi Province, China. The tested bacterial strains were inoculated into beef extract peptone broth. The inoculated broth was then incubated at 30 °C and 150 r/min for 72 h. After incubation, it was centrifuged at 5000 r/min for 10 min, and the supernatant was discarded, preparing suspensions of the test strains at different concentrations. The bacterial pellets were resuspended in 20 mL of sterile water. The duration of the pot experiment was 60 days after seeding, with temperature settings of 25 °C during the day (12 h) and 15 °C at night (12 h) in the growth chamber. There were 10 treatments in total, with each treatment replicated 3 times, resulting in a total of 30 pots. In total, 5.5 kg of contaminated soil from the mining area was weighed and placed into polyethylene plastic pots (43 cm long, 19 cm wide, and 14 cm high, with four drainage holes at the bottom). Each pot was then fertilized with 1.21 g of potassium dihydrogen phosphate and 1.28 g of urea as basal fertilizer. Additionally, 0.5 g of lawn plant seeds were sown and covered with a thin layer of soil on the surface. Throughout the entire growth period, water was periodically supplemented using the weighing method to maintain soil moisture at 60~70% of field capacity. Additionally, the positions of the pots were regularly changed to eliminate the effects of light exposure on the growth of the lawn plants. After the seeds germinated, different concentrations of the tested bacterial suspension were inoculated into the root zone of the plants, with the concentrations specified in Table 4.
After 60 days of growth, soil and plant samples were collected from the lawn plants. We cleaned the aboveground parts and root systems of the plants, then weighed them separately. The plant samples were then oven-dried at 105 °C for 30 min, followed by drying at 80 °C until a constant weight was achieved. The dried plant samples were pulverized, ground, and stored in bags. Soil samples were divided into rhizosphere soil and non-rhizosphere soil. Whilst separating the roots, the rhizosphere soil was collected in a plastic bag by shaking the roots vigorously. The soil outside the root zone was collected as the non-rhizosphere soil [52]. A portion of the soil samples was air-dried and passed through a 1 mm nylon sieve for the determination of soil organic matter, available phosphorus, and other nutrient contents. Another portion of the soil samples was passed through a 0.15 mm nylon sieve for the determination of heavy metal content in the soil. Another portion of the soil samples underwent freeze-drying at low temperature (Beijing Songyuan, Beijing, China); the soil samples were ground using a fully automatic cryogenic grinder (JXFSTPRP-CL, Shanghai Jingxin, Shanghai, China), sieved, and stored in a −80 °C freezer for subsequent high-throughput sequencing analysis of soil microbial communities.

3.3. Measurement Methods

3.3.1. Soil Physicochemical Properties

The pH of the collected soil samples was measured using a pH meter (Mettler Toledo, Switzerland); the suspension had a soil/water ratio of 1:2.5 (mass/volume) [53]. Soil organic matter (OM) was determined using the potassium dichromate volumetric method with external heating. Total nitrogen (TN) is determined using the Kjeldahl method, available phosphorus (AP) in soil is measured with a SmartChem fully automated discrete chemistry analyzer (Smartchem 450, AMS Alliance, Rome, Italy), and the total Cadmium (T-Cd) content (C1)in soil is determined by the digestion method using hydrochloric acid, nitric acid, hydrofluoric acid, and perchloric acid., while the modified BCR sequential extraction method was employed to determine the content of different forms of cadmium (C2), which were then measured using graphite furnace atomic absorption spectrometry. The calculation formula for the relative content of each form of Cd (C3) is as follows:
C3 = C2 × 100%/C1

3.3.2. Plant Biomass and Heavy Metal Accumulation

The plant samples were oven-dried at 105 °C for 30 min to terminate biological activity, followed by drying at 80 °C to a constant weight, and then the plant biomass was determined. The determination of heavy metal content was divided into two parts: aboveground parts and roots. In total, 0.200 g (accurate to 0.0001 g) of dried plant samples was weighed and transferred into digestion tubes. After soaking overnight in 10 mL of HNO3 and HClO4 (7:1, v/v) solution, the digestion tubes were placed in a graphite digestion instrument (EH-2542, Beijing Auwii Instrument Co., Ltd., Beijing, China) and an automatic heating program was set as follows: digestion was carried out at 120 °C for 60 min, followed by digestion at 160 °C for 30 min and, finally, at 180 °C for 15 min. The Cd content in the plant samples was determined using graphite furnace atomic absorption spectrometry, with each treatment replicated three times. The bioconcentration factor (BCF) and translocation factor (TF) of heavy metal Cd in plants were calculated using the following formulas:
BCF = Ct/Cs
TF = Caboveground/Croot
where Ct and Cs are the Cd concentration in plant tissues and soils; Caboveground and Croot are the Cd concentration in the plants’ aboveground part and root part.

3.3.3. Soil Microbial Structure

DNA extraction was performed on soil samples from metal-contaminated layers (fresh samples) and soil samples after different remediation treatments. The bacterial 16S rRNA gene amplicon sequences were determined through sequencing. Total DNA was extracted from the filter membranes using the FastDNA Spin Kit (MP Biomedicals, Santa Ana, CA, USA), following the instructions provided in the manual. The quality and concentration of the extracted DNA were measured using the NanoDrop ND-1000 ultraviolet–visible spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA). The primers 515F (5′-CCTAYGGGRBGCASCAG-3′) and 806R (5′-GGACTACNNGGG TATCTAAT-3′) were used to amplify the V3-V4 region of the 16S rRNA gene via PCR. The PCR products were sent to Novogene Co., Ltd. (Tianjin, China) for subsequent library construction and sequencing on the Illumina MiSeq high-throughput sequencing platform provided by the company (Illumina, Kapa Biosciences, Woburn, MA, USA). The remaining sequences after filtering chimeras using Usearch 11 software were clustered into operational taxonomic units (OTUs) based on a 97% similarity threshold. Subsequently, data from each sample were normalized, and the Alpha diversity indices such as Chao1, Shannon, and Simpson were calculated using Qiime software (Version 1.9.1).

3.4. Statistical Analysis

All experimental data were statistically processed using Excel 2016 software; the data were subjected to a one-way analysis of variance (ANOVA) using the SPSS 26.0 software (SPSS, Inc., Chicago, IL, USA) at a significance level of p ≤ 0.05. Graphs were generated using Origin 2021 software. The correlation between microbial communities and soil environmental factors was analyzed using R 4.2.2. The Canoco 4.5 software was employed for redundancy analysis (RDA) to determine the important environmental factors influencing the microbial community structure.

4. Conclusions

Bacillus cereus inoculation promotes the growth and Cd accumulation of lawn plants (Festuca arundinacea A‘rid III’ and Poa pratensis M‘idnight II’). This study demonstrated that the application of Bacillus cereus reduced the Cd content in soil to some extent and improved the soil quality. Additionally, the application of Bacillus cereus promoted the transformation of Cd in rhizospheric soil from the oxidizable and residual states to the acid-soluble and reducible states. In the Festuca arundinacea and Poa pratensis treatments, the bioavailable Cd content in rhizospheric soil increased by up to 13.43% and 26.54%, respectively. Moreover, the application of Bacillus cereus enhanced the absorption of cadmium by plants. Both lawn plants demonstrated the ability to absorb significant amounts of cadmium from the soil and translocate it into their tissues after the application of Bacillus cereus. The optimal remediation treatment was observed when the bacterial liquid concentration was 105 CFU/mL. The bioconcentration factor (BCF) of Festuca arundinacea and Poa pratensis increased by 0.906 and 0.519 times, respectively, and the translocation factor (TF) of cadmium in plant tissues reached its maximum values of 0.467 and 0.339, respectively. Additionally, Bacillus cereus increased the diversity of bacterial communities in non-rhizospheric soil, while reducing diversity in rhizospheric soil. The application of Bacillus cereus treatment reduced the proportion of Actinobacteriota in both the rhizospheric and non-rhizospheric soil of lawn plants, while enhancing the relative abundance of ecologically beneficial microorganisms in the soil. This study confirms the significant potential and prospects of using lawn plants in conjunction with Bacillus cereus for the remediation of cadmium-contaminated soil.

Author Contributions

B.Z.: writing—review and editing, supervision, resources, methodology, data curation, and conceptualization. Z.Y.: writing—review and editing, writing—original draft, visualization, investigation, formal analysis, data curation, and conceptualization. X.C.: methodology, monitoring, validation, and conceptualization. R.J.: conceptualization, data management, and formal analysis. S.Y.: conceptualization, methodology, and supervision. B.G.: investigation and verification. D.F.: survey and software. X.Y.: survey. Y.C.: supervision and data management. W.L.: verification. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the National Natural Science Foundation of China (52222903; 41977007), a research project of the State Key Laboratory of Eco-hydraulics in Northwest Arid Region of China (QJNY-2019-01).

Data Availability Statement

Data are contained within the article.

Acknowledgments

The authors also gratefully acknowledge the Xi’an University of Technology for providing lab facilitation and experimental measurements.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Yang, X.; Wang, D.; Tao, Y.; Shen, M.; Wei, W.; Cai, C.; Ding, C.; Li, J.; Song, L.; Yin, B.; et al. Effects of elevated CO2 on the Cd uptake by rice in Cd-contaminated paddy soils. J. Hazard. Mater. 2023, 442, 130140. [Google Scholar] [CrossRef]
  2. Liu, S.; Ali, S.; Yang, R.; Tao, J.; Ren, B. A newly discovered Cd-hyperaccumulator Lantana camara L. J. Hazard. Mater. 2019, 371, 233–242. [Google Scholar] [CrossRef] [PubMed]
  3. Fan, D.; Wang, S.; Guo, Y.; Zhu, Y.; Agathokleous, E.; Ahmad, S.; Han, J. Cd induced biphasic response in soil alkaline phosphatase and changed soil bacterial community composition: The role of background Cd contamination and time as additional factors. Sci. Total. Environ. 2020, 757, 143771. [Google Scholar] [CrossRef] [PubMed]
  4. Khatereh, N.; Ali, A.; Behnam Asgari, L.; van Hullebusch, E.D. Effects of biochar dose on cadmium accumulation in spinach and its fractionation in a calcareous soil. Arab. J. Geosci. 2022, 15, 336. [Google Scholar] [CrossRef]
  5. Rizwan, M.; Ali, S.; Adrees, M.; Ibrahim, M.; Tsang, D.C.W.; Zia-Ur-Rehman, M.; Zahir, Z.A.; Rinklebe, J.; Tack, F.M.G.; Ok, Y.S. A critical review on effects, tolerance mechanisms and management of cadmium in vegetables. Chemosphere 2017, 182, 90–105. [Google Scholar] [CrossRef] [PubMed]
  6. Grobelak, A.; Napora, A. The Chemophytostabilisation Process of Heavy Metal Polluted Soil. PLoS ONE 2015, 10, e0129538. [Google Scholar] [CrossRef]
  7. Xiao, L.; Bi, Y.; Du, S.; Wang, Y.; Guo, C.; Christie, P. Response of ecological stoichiometry and stoichiometric homeostasis in the plant-litter-soil system to re-vegetation type in arid mining subsidence areas. J. Arid. Environ. 2020, 184, 104298. [Google Scholar] [CrossRef]
  8. Nayeri, S.; Dehghanian, Z.; Asgari Lajayer, B.; Thomsson, A.; Astatkie, T.; Price, G.W. CRISPR/Cas9-Mediated genetically edited ornamental and aromatic plants: A promising technology in phytoremediation of heavy metals. J. Clean. Prod. 2023, 428, 139512. [Google Scholar] [CrossRef]
  9. Zheng, K.; Liu, Z.; Liu, C.; Liu, J.; Zhuang, J. Enhancing remediation potential of heavy metal contaminated soils through synergistic application of microbial inoculants and legumes. Front. Microbiol. 2023, 14, 1272591. [Google Scholar] [CrossRef]
  10. Gerami, Z.; Lakzian, A.; Hemati, A.; Amirifar, A.; Asgari Lajayer, B.; van Hullebusch, E.D. Effect of cadmium on sorghum root colonization by glomeral fungi and its impact on total and easily extractable glomalin production. Environ. Sci. Pollut. Res. 2021, 28, 34570–34583. [Google Scholar] [CrossRef]
  11. Hou, J.; Wang, Q.; Liu, W.; Zhong, D.; Ge, Y.; Christie, P.; Luo, Y. Soil microbial community and association network shift induced by several tall fescue cultivars during the phytoremediation of a petroleum hydrocarbon-contaminated soil. Sci. Total. Environ. 2021, 792, 148411. [Google Scholar] [CrossRef] [PubMed]
  12. Rajkumar, M.; Ae, N.; Freitas, H. Endophytic bacteria and their potential to enhance heavy metal phytoextraction. Chemosphere 2009, 77, 153–160. [Google Scholar] [CrossRef] [PubMed]
  13. Haque, S.; Srivastava, N.; Pal, D.B.; Alkhanani, M.F.; Almalki, A.H.; Areeshi, M.Y.; Naidu, R.; Gupta, V.K. Functional microbiome strategies for the bioremediation of petroleum-hydrocarbon and heavy metal contaminated soils: A review. Sci. Total. Environ. 2022, 833, 155222. [Google Scholar] [CrossRef] [PubMed]
  14. Fan, L.; Kailu, Z.; Yang, Z.; Dong, L.; Xiaoyan, S.; Li, L.; Huiping, F.; Qing, H.; Zhiqiang, Z. Screening of cadmium-chromium-tolerant strains and synergistic remediation of heavy metal-contaminated soil using king grass combined with highly efficient microbial strains. Sci. Total. Environ. 2023, 912, 168990. [Google Scholar] [CrossRef] [PubMed]
  15. Long, J.; Yu, M.; Xu, H.; Huang, S.; Wang, Z.; Zhang, X.-X. Characterization of cadmium biosorption by inactive biomass of two cadmium-tolerant endophytic bacteria Microbacterium sp. D2-2 and Bacillus sp. C9-3. Ecotoxicology 2021, 30, 1419–1428. [Google Scholar] [CrossRef]
  16. Wang, X.; Cai, D.; Ji, M.; Chen, Z.; Yao, L.; Han, H. Isolation of heavy metal-immobilizing and plant growth-promoting bacteria and their potential in reducing Cd and Pb uptake in water spinach. Sci. Total. Environ. 2022, 819, 153242. [Google Scholar] [CrossRef] [PubMed]
  17. Lin, C.-C.; Lin, H.-L. Remediation of soil contaminated with the heavy metal (Cd2+). J. Hazard. Mater. 2005, 122, 7–15. [Google Scholar] [CrossRef] [PubMed]
  18. Xie, Y.; Bu, H.; Feng, Q.; Wassie, M.; Amee, M.; Jiang, Y.; Bi, Y.; Hu, L.; Chen, L. Identification of Cd-resistant microorganisms from heavy metal-contaminated soil and its potential in promoting the growth and Cd accumulation of bermudagrass. Environ. Res. 2021, 200, 111730. [Google Scholar] [CrossRef]
  19. Manoj, S.R.; Karthik, C.; Kadirvelu, K.; Arulselvi, P.I.; Shanmugasundaram, T.; Bruno, B.; Rajkumar, M. Understanding the molecular mechanisms for the enhanced phytoremediation of heavy metals through plant growth promoting rhizobacteria: A review. J. Environ. Manag. 2019, 254, 109779. [Google Scholar] [CrossRef]
  20. Lanping, S.; Xiaoyang, X.; Yuanyuan, Z.; Wanyue, H.; Xiaoping, L.; Yanmei, A.; Yang, W.; Zekun, Z.; Hong, C.; Yongjie, H.; et al. Dynamic mechanisms of cadmium accumulation and detoxification by Lolium perenne grown in soil inoculated with the cadmium-tolerant bacterium strain Cdq4-2. Sci. Total. Environ. 2023, 873, 162314. [Google Scholar] [CrossRef]
  21. Alves, A.R.A.; Yin, Q.; Oliveira, R.S.; Silva, E.F.; Novo, L.A.B. Plant growth-promoting bacteria in phytoremediation of metal-polluted soils: Current knowledge and future directions. Sci. Total. Environ. 2022, 838, 156435. [Google Scholar] [CrossRef] [PubMed]
  22. Haroun, M.; Xie, S.; Awadelkareem, W.; Wang, J.; Qian, X. Influence of biofertilizer on heavy metal bioremediation and enzyme activities in the soil to revealing the potential for sustainable soil restoration. Sci. Rep. 2023, 13, 20684. [Google Scholar] [CrossRef]
  23. Chen, B.; Tan, S.; Zeng, Q.; Wang, A.; Zheng, H. Soil nutrient heterogeneity affects the accumulation and transfer of cadmium in Bermuda grass (Cynodon dactylon (L.) pers.). Chemosphere 2019, 221, 342–348. [Google Scholar] [CrossRef] [PubMed]
  24. Liu, C.; Li, B.; Dong, Y.; Lin, H. Endophyte colonization enhanced cadmium phytoremediation by improving endosphere and rhizosphere microecology characteristics. J. Hazard. Mater. 2022, 434, 128829. [Google Scholar] [CrossRef]
  25. Li, K.-T.; Yang, Y.; Zhang, S.-W.; Cheng, X. Dynamics of the Bacterial Community’s Soil during the In-Situ Degradation Process of Waste Chicken Feathers. Indian J. Microbiol. 2022, 62, 225–233. [Google Scholar] [CrossRef]
  26. Daraz, U.; Li, Y.; Sun, Q.; Zhang, M.; Ahmad, I. Inoculation of Bacillus spp. Modulate the Soil Bacterial Communities and Available Nutrients in the Rhizosphere of Vetiver Plant Irrigated with Acid Mine Drainage. Chemosphere 2020, 263, 128345. [Google Scholar] [CrossRef]
  27. Li, B.; Wu, B.; Dong, Y.; Lin, H.; Liu, C. Endophyte inoculation enhanced microbial metabolic function in the rhizosphere benefiting cadmium phytoremediation by Phytolacca acinosa. Chemosphere 2023, 338, 139421. [Google Scholar] [CrossRef] [PubMed]
  28. Farh, M.E.-A.; Kim, Y.-J.; Sukweenadhi, J.; Singh, P.; Yang, D.-C. Aluminium resistant, plant growth promoting bacteria induce overexpression of Aluminium stress related genes in Arabidopsis thaliana and increase the ginseng tolerance against Aluminium stress. Microbiol. Res. 2017, 200, 45–52. [Google Scholar] [CrossRef] [PubMed]
  29. Li, X.; Zhang, J.; Ma, J.; Liu, Q.; Shi, T.; Gong, Y.; Yang, S.; Wu, Y. Status of chromium accumulation in agricultural soils across China (1989–2016). Chemosphere 2020, 256, 127036. [Google Scholar] [CrossRef]
  30. Li, Q.; Xing, Y.; Fu, X.; Ji, L.; Li, T.; Wang, J.; Chen, G.; Qi, Z.; Zhang, Q. Biochemical mechanisms of rhizospheric Bacillus subtilis-facilitated phytoextraction by alfalfa under cadmium stress—Microbial diversity and metabolomics analyses. Ecotoxicol. Environ. Saf. 2021, 212, 112016. [Google Scholar] [CrossRef]
  31. Jeong, S.; Moon, H.S.; Shin, D.; Nam, K. Survival of introduced phosphate-solubilizing bacteria (PSB) and their impact on microbial community structure during the phytoextraction of Cd-contaminated soil. J. Hazard. Mater. 2013, 263, 441–449. [Google Scholar] [CrossRef] [PubMed]
  32. Wood, J.L.; Tang, C.; Franks, A.E. Microbial associated plant growth and heavy metal accumulation to improve phytoextraction of contaminated soils. Soil Biol. Biochem. 2016, 103, 131–137. [Google Scholar] [CrossRef]
  33. Ma, Y.; Oliveira, R.S.; Nai, F.; Rajkumar, M.; Luo, Y.; Rocha, I.; Freitas, H. The hyperaccumulator Sedum plumbizincicola harbors metal-resistant endophytic bacteria that improve its phytoextraction capacity in multi-metal contaminated soil. J. Environ. Manag. 2015, 156, 62–69. [Google Scholar] [CrossRef]
  34. Sharma, P.; Tripathi, S.; Chaturvedi, P.; Chaurasia, D.; Chandra, R. Newly isolated Bacillus sp. PS-6 assisted phytoremediation of heavy metals using Phragmites communis: Potential application in wastewater treatment. Bioresour. Technol. 2020, 320, 124353. [Google Scholar] [CrossRef]
  35. Huang, G.; Zhou, X.; Guo, G.; Ren, C.; Rizwan, M.S.; Islam, M.S.; Hu, H. Variations of dissolved organic matter and Cu fractions in rhizosphere soil induced by the root activities of castor bean. Chemosphere 2020, 254, 126800. [Google Scholar] [CrossRef]
  36. Chen, L.; Yang, J.-y.; Wang, D. Phytoremediation of uranium and cadmium contaminated soils by sunflower (Helianthus annuus L.) enhanced with biodegradable chelating agents. J. Clean. Prod. 2020, 263, 121491. [Google Scholar] [CrossRef]
  37. Wyszkowska, J.; Borowik, A.; Kucharski, M.; Kucharski, J. Effect of cadmium, copper and zinc on plants, soil microorganisms and soil enzymes. J. Elem. 2013, 18, 769–796. [Google Scholar] [CrossRef]
  38. Gutiérrez, C.; Fernández, C.; Escuer, M.; Campos-Herrera, R.; Beltrán Rodríguez, M.E.; Carbonell, G.; Rodríguez Martín, J.A. Effect of soil properties, heavy metals and emerging contaminants in the soil nematodes diversity. Environ. Pollut. 2016, 213, 184–194. [Google Scholar] [CrossRef]
  39. Bettermann, A.; Zethof, J.H.T.; Babin, D.; Cammeraat, E.L.H.; Solé-Benet, A.; Lázaro, R.; Luna, L.; Nesme, J.; Sørensen, S.J.; Kalbitz, K.; et al. Importance of microbial communities at the root-soil interface for extracellular polymeric substances and soil aggregation in semiarid grasslands. Soil Biol. Biochem. 2021, 159, 108301. [Google Scholar] [CrossRef]
  40. Pereira, S.I.A.; Lima, A.I.G.; Figueira, E.M.d.A.P. Screening Possible Mechanisms Mediating Cadmium Resistance in Rhizobium leguminosarum bv. viciae Isolated from Contaminated Portuguese Soils. Microb. Ecol. 2006, 52, 176–186. [Google Scholar] [CrossRef]
  41. Ma, S.; Qiao, L.; Liu, X.; Zhang, S.; Zhang, L.; Qiu, Z.; Yu, C. Microbial community succession in soils under long-term heavy metal stress from community diversity-structure to KEGG function pathways. Environ. Res. 2022, 214, 113822. [Google Scholar] [CrossRef] [PubMed]
  42. Kou, B.; He, Y.; Wang, Y.; Qu, C.; Tang, J.; Wu, Y.; Tan, W.; Yuan, Y.; Yu, T. The relationships between heavy metals and bacterial communities in a coal gangue site. Environ. Pollut. 2023, 322, 121136. [Google Scholar] [CrossRef] [PubMed]
  43. Liu, B.; Yao, J.; Chen, Z.; Ma, B.; Li, H.; Wancheng, P.; Liu, J.; Wang, D.; Duran, R. Biogeography, assembly processes and species coexistence patterns of microbial communities in metalloids-laden soils around mining and smelting sites. J. Hazard. Mater. 2021, 425, 127945. [Google Scholar] [CrossRef] [PubMed]
  44. Xu, Z.; Wang, D.; Tang, W.; Wang, L.; Li, Q.; Lu, Z.; Liu, H.; Zhong, Y.; He, T.; Guo, S. Phytoremediation of cadmium-polluted soil assisted by D-gluconate-enhanced Enterobacter cloacae colonization in the Solanum nigrum L. rhizosphere. Sci. Total. Environ. 2020, 732, 139265. [Google Scholar] [CrossRef] [PubMed]
  45. Ahemad, M. Phosphate-solubilizing bacteria-assisted phytoremediation of metalliferous soils: A review. 3 Biotech 2014, 5, 111–121. [Google Scholar] [CrossRef]
  46. Manickam, N.; Bajaj, A.; Saini, H.S.; Shanker, R. Surfactant mediated enhanced biodegradation of hexachlorocyclohexane (HCH) isomers by Sphingomonas sp. NM05. Biodegradation 2012, 23, 673–682. [Google Scholar] [CrossRef] [PubMed]
  47. Li, Y.; Shi, X.; Ling, Q.; Li, S.; Wei, J.; Xin, M.; Xie, D.; Chen, X.; Liu, K.; Yu, F. Bacterial extracellular polymeric substances: Impact on soil microbial community composition and their potential role in heavy metal-contaminated soil. Ecotoxicol. Environ. Saf. 2022, 240, 113701. [Google Scholar] [CrossRef]
  48. Han, H.; Wu, X.; Yao, L.; Chen, Z. Heavy metal-immobilizing bacteria combined with calcium polypeptides reduced the uptake of Cd in wheat and shifted the rhizosphere bacterial communities. Environ. Pollut. 2020, 267, 115432. [Google Scholar] [CrossRef]
  49. Kamal, N.; Liu, Z.; Qian, C.; Wu, J.; Zhong, X. Improving hybrid Pennisetum growth and cadmium phytoremediation potential by using Bacillus megaterium BM18-2 spores as biofertilizer. Microbiol. Res. 2020, 242, 126594. [Google Scholar] [CrossRef]
  50. Zhao, X.; Sun, Y.; Huang, J.; Wang, H.; Tang, D. Effects of soil heavy metal pollution on microbial activities and community diversity in different land use types in mining areas. Environ. Sci. Pollut. Res. 2020, 27, 20215–20226. [Google Scholar] [CrossRef]
  51. Jia, R. Microbially-Enhanced Phytoremediation of Acidiccadmium-Contaminated Soil by Turfgrass and It Smechanism; Xi’an University of Technology: Xi’an, China, 2023. [Google Scholar]
  52. Jalali, M.; Jalali, M. Sewage Sludge Application Effects on Phosphorus Uptake by Cucumber and on Rhizosphere and Non-rhizosphere Soils Under Greenhouse Conditions. J. Soil Sci. Plant Nutr. 2021, 21, 596–611. [Google Scholar] [CrossRef]
  53. Wang, L.; Lin, H.; Dong, Y.; Li, B.; He, Y. Effects of endophytes inoculation on rhizosphere and endosphere microecology of Indian mustard (Brassica juncea) grown in vanadium-contaminated soil and its enhancement on phytoremediation. Chemosphere 2019, 240, 124891. [Google Scholar] [CrossRef] [PubMed]
Plants 13 01303 g001
Figure 1. Effects of different treatments on (a) physical and chemical properties; (b) Cd concentration in soil; and (c) the proportion of Cd morphology in rhizosphere soil. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis. F1, F2, F3, and F4 represent different forms of soil Cd: F1 as acid-soluble, F2 as reducible, F3 as oxidizable, and F4 as residual forms. Different letters represent notable differences between the treatments at p < 0.05. The bars are standard deviation in the figure.
Figure 1. Effects of different treatments on (a) physical and chemical properties; (b) Cd concentration in soil; and (c) the proportion of Cd morphology in rhizosphere soil. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis. F1, F2, F3, and F4 represent different forms of soil Cd: F1 as acid-soluble, F2 as reducible, F3 as oxidizable, and F4 as residual forms. Different letters represent notable differences between the treatments at p < 0.05. The bars are standard deviation in the figure.
Plants 13 01303 g001
Plants 13 01303 g002
Figure 2. Effects of different bacterial solution concentrations on biomass coefficient of lawn plants. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis. Different letters represent notable differences between the treatments at p < 0.05. The bars are standard deviation in the figure.
Figure 2. Effects of different bacterial solution concentrations on biomass coefficient of lawn plants. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis. Different letters represent notable differences between the treatments at p < 0.05. The bars are standard deviation in the figure.
Plants 13 01303 g002
Plants 13 01303 g003
Figure 3. Relative abundance of the top 10 dominant microorganisms of bacteria at the phylum level in rhizosphere soil and non-rhizosphere soils under different treatments (%): (a) Festuca arundinacea; (b) Poa pratensis. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis.
Figure 3. Relative abundance of the top 10 dominant microorganisms of bacteria at the phylum level in rhizosphere soil and non-rhizosphere soils under different treatments (%): (a) Festuca arundinacea; (b) Poa pratensis. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis.
Plants 13 01303 g003
Plants 13 01303 g004
Figure 4. Heatmap of the top 30 genera of rhizosphere and non-rhizosphere soil bacteria under different treatments: (a) Festuca arundinacea; (b) Poa pratensis. Abundance is expressed as color intensity, reflecting the proportion of the total number of valid sequences in each group. Vertical: treatment. Horizontal: annotation information for species. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis.
Figure 4. Heatmap of the top 30 genera of rhizosphere and non-rhizosphere soil bacteria under different treatments: (a) Festuca arundinacea; (b) Poa pratensis. Abundance is expressed as color intensity, reflecting the proportion of the total number of valid sequences in each group. Vertical: treatment. Horizontal: annotation information for species. Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis.
Plants 13 01303 g004
Plants 13 01303 g005
Figure 5. LEFse analysis of bacterial community of each group: (a) taxonomic cladistics of bacteria community; (b) LDA distribution of bacterial community. Note: G and C groups represent non-rhizospheric and rhizospheric soils of Festuca arundinacea; Z and B groups represent non-rhizospheric and rhizospheric soils of Poa pratensis. In the phylogenetic tree, nodes from inner to outer represent taxonomic categories of bacterial communities from phylum to genus, where p, c, o, f, g, and s represent phylum, class, order, family, genus, and species levels, respectively. The size of the nodes is proportional to their relative abundance. The length of the bars in the LDA value histogram reflects the contribution of species abundance to the differential effect size.
Figure 5. LEFse analysis of bacterial community of each group: (a) taxonomic cladistics of bacteria community; (b) LDA distribution of bacterial community. Note: G and C groups represent non-rhizospheric and rhizospheric soils of Festuca arundinacea; Z and B groups represent non-rhizospheric and rhizospheric soils of Poa pratensis. In the phylogenetic tree, nodes from inner to outer represent taxonomic categories of bacterial communities from phylum to genus, where p, c, o, f, g, and s represent phylum, class, order, family, genus, and species levels, respectively. The size of the nodes is proportional to their relative abundance. The length of the bars in the LDA value histogram reflects the contribution of species abundance to the differential effect size.
Plants 13 01303 g005
Plants 13 01303 g006
Figure 6. RDA of environmental factor counts in rhizosphere soils for bacteria: (a,b) Soil properties and soil bacterial communities of tall Festuca arundinacea. (c,d) Soil properties and soil bacterial communities of Poa pratensis. F1, F2, F3, and F4 represent different forms of soil Cd: F1 as acid-soluble, F2 as reducible, F3 as oxidizable, and F4 as residual forms. p ≤ 0.001 is represented by three symbols (***), 0.001 < p ≤ 0.01 is demonstrated by two asterisks (**), and 0.01 < p ≤ 0.05 is demonstrated by a single asterisk (*).
Figure 6. RDA of environmental factor counts in rhizosphere soils for bacteria: (a,b) Soil properties and soil bacterial communities of tall Festuca arundinacea. (c,d) Soil properties and soil bacterial communities of Poa pratensis. F1, F2, F3, and F4 represent different forms of soil Cd: F1 as acid-soluble, F2 as reducible, F3 as oxidizable, and F4 as residual forms. p ≤ 0.001 is represented by three symbols (***), 0.001 < p ≤ 0.01 is demonstrated by two asterisks (**), and 0.01 < p ≤ 0.05 is demonstrated by a single asterisk (*).
Plants 13 01303 g006
Plants 13 01303 g007
Figure 7. The relationship between soil physicochemical properties and microbial community composition at the phylum level for (a) non-rhizospheric soil of Festuca arundinacea, (b) non-rhizospheric soil of Poa pratensis, (c) rhizospheric soil of Festuca arundinacea, and (d) rhizospheric soil of Poa pratensis. p ≤ 0.01 is demonstrated by two asterisks (**), and 0.01 < p ≤ 0.05 is demonstrated by a single asterisk (*).
Figure 7. The relationship between soil physicochemical properties and microbial community composition at the phylum level for (a) non-rhizospheric soil of Festuca arundinacea, (b) non-rhizospheric soil of Poa pratensis, (c) rhizospheric soil of Festuca arundinacea, and (d) rhizospheric soil of Poa pratensis. p ≤ 0.01 is demonstrated by two asterisks (**), and 0.01 < p ≤ 0.05 is demonstrated by a single asterisk (*).
Plants 13 01303 g007
Table 1. Effects of different concentrations of bacterial solution on Cd concentrations in lawn plants.
Table 1. Effects of different concentrations of bacterial solution on Cd concentrations in lawn plants.
Lawn PlantsHandleConcentration (CFU/mL)Cd Concentration (mg/kg)Translocation Factor (TF)Bioconcentration Factor (BCF)
AbovegroundRoot
Festuca arundinacea ‘Arid Ⅲ’G000.481 ± 0.012 d1.907 ± 0.061 d0.255 ± 0.003 d0.431 ± 0.006 c
G11030.753 ± 0.013 c2.557 ± 0.08 b0.294 ± 0.006 c0.603 ± 0.009 b
G21051.256 ± 0.139 a2.682 ± 0.093 a0.467 ± 0.039 a0.898 ± 0.031 a
G31070.992 ± 0.018 b2.489 ± 0.039 c0.398 ± 0.007 b0.877 ± 0.010 a
G41090.756 ± 0.014 c2.557 ± 0.082 b0.308 ± 0.005 c0.673 ± 0.011 b
Poa pratensis
‘Midnight Ⅱ’		Z000.323 ± 0.027 e1.895 ± 0.031 d0.170 ± 0.012 d0.221 ± 0.006 d
Z11030.651 ± 0.038 b2.385 ± 0.069 b0.273 ± 0.014 b0.530 ± 0.013 b
Z21050.851 ± 0.043 a2.526 ± 0.136 a0.339 ± 0.031 a0.499 ± 0.029 a
Z31070.529 ± 0.018 c2.065 ± 0.045 c0.256 ± 0.014 c0.452 ± 0.015 c
Z41090.469 ± 0.041 d1.825 ± 0.026 d0.257 ± 0.021 c0.456 ± 0.017 c
Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis. Different letters represent notable differences between the treatments at p < 0.05. The bars are standard deviation in the table.
Table 2. Alpha diversity index of soil bacterial community.
Table 2. Alpha diversity index of soil bacterial community.
TreatmentChao1 IndexShannon IndexSimpson IndexCoverage
Festuca arundinacea ‘Arid Ⅲ’Non-rhizosphere soilG0649.76 ± 2.23 i7.27 ± 0.03 g0.956 ± 0.005 f0.987
G1687.29 ± 2.38 h7.40 ± 0.02 f0.977 ± 0.005 ab0.990
G2845.55 ± 4.93 d7.61 ± 0.04 d0.980 ± 0.001 a0.998
G3729.80 ± 5.27 g7.45 ± 0.01 e0.971 ± 0.002 c0.986
G4752.73 ± 3.89 f7.59 ± 0.06 de0.975 ± 0.003 b0.993
Rhizosphere soilG01025.64 ± 2.76 a8.57 ± 0.03 a0.966 ± 0.001 d0.988
G11003.04 ± 3.86 b8.56 ± 0.04 a0.960 ± 0.001 e0.997
G2820.59 ± 3.58 de8.48 ± 0.04 b0.961 ± 0.002 e0.998
G3795.01 ± 3.72 e7.95 ± 0.03 c0.958 ± 0.003 ef0.988
G4973.20 ± 4.06 c8.52 ± 0.02 ab0.965 ± 0.002 d0.989
Poa pratensis
‘Midnight Ⅱ’		Non-rhizosphere soilZ0601.36 ± 2.58 h6.67 ± 0.04 h0.957 ± 0.003 g0.990
Z1646.55 ± 3.40 f6.75 ± 0.02 g0.962 ± 0.002 f0.998
Z2761.03 ± 4.23 e6.94 ± 0.01 ef0.988 ± 0.001 a0.988
Z3816.12 ± 1.917 d7.33 ± 0.02 c0.965 ± 0.001 e0.993
Z4873.15 ± 2.99 b7.91 ± 0.03 a0.975 ± 0.001 d0.994
Rhizosphere soilZ0916.03 ± 7.91 a7.75 ± 0.02 b0.980 ± 0.003 b0.986
Z1828.77 ± 4.80 c7.72 ± 0.03 b0.981 ± 0.005 b0.989
Z2643.00 ± 2.74 f7.03 ± 0.02 e0.979 ± 0.002 bc0.985
Z3910.36 ± 4.69 ab6.89 ± 0.02 f0.975 ± 0.002 d0.991
Z4634.33 ± 2.71 g7.14 ± 0.04 d0.977 ± 0.002 c0.992
Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis. Different letters represent notable differences between the treatments at p < 0.05. The bars are standard deviation in the table.
Table 3. The physicochemical properties of the tested soil.
Table 3. The physicochemical properties of the tested soil.
pHOrganic Matter (OM, g/kg)Total Nitrogen (TN, g/kg)Total Potassium (TP, g/kg)Total Carbon (TC, g/kg)Available Phosphorus (AP, mg/kg)Total Cd
(T-Cd, mg/kg)
4.329.260.4025.066.174.582.31
Table 4. Design of pot experiments.
Table 4. Design of pot experiments.
Lawn PlantsExperimental CodeInoculation DosageViable Count
Poa pratensis
‘Midnight Ⅱ’		Z020 mL0 CFU/mL
Z120 mL103 CFU/mL
Z220 mL105 CFU/mL
Z320 mL107 CFU/mL
Z420 mL109 CFU/mL
Festuca arundinacea ‘Arid Ⅲ’G020 mL0 CFU/mL
G120 mL103 CFU/mL
G220 mL105 CFU/mL
G320 mL107 CFU/mL
G420 mL109 CFU/mL
Note: G0, G1, G2, G3, and G4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Festuca arundinacea, while Z0, Z1, Z2, Z3, and Z4 represent the concentrations of 0, 103, 105, 107, and 109 CFU/mL of Bacillus cereus inoculum applied to Poa pratensis.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Zhou, B.; Yang, Z.; Chen, X.; Jia, R.; Yao, S.; Gan, B.; Fan, D.; Yang, X.; Li, W.; Chen, Y. Microbiological Mechanisms of Collaborative Remediation of Cadmium-Contaminated Soil with Bacillus cereus and Lawn Plants. Plants 2024, 13, 1303. https://doi.org/10.3390/plants13101303

AMA Style

Zhou B, Yang Z, Chen X, Jia R, Yao S, Gan B, Fan D, Yang X, Li W, Chen Y. Microbiological Mechanisms of Collaborative Remediation of Cadmium-Contaminated Soil with Bacillus cereus and Lawn Plants. Plants. 2024; 13(10):1303. https://doi.org/10.3390/plants13101303

Chicago/Turabian Style

Zhou, Beibei, Zehao Yang, Xiaopeng Chen, Ruonan Jia, Shaoxiong Yao, Bin Gan, Dongliang Fan, Xie Yang, Wenqian Li, and Yunhan Chen. 2024. "Microbiological Mechanisms of Collaborative Remediation of Cadmium-Contaminated Soil with Bacillus cereus and Lawn Plants" Plants 13, no. 10: 1303. https://doi.org/10.3390/plants13101303

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop