Skip to main content
NCBI home page
ACS Omega logoLink to ACS Omega
. 2023 Feb 8;8(7):6411–6422. doi: 10.1021/acsomega.2c06665

Effect of Hydrogen Peroxide on the Soil Microbial Community Structure and Growth of Malus hupehensis Rehd. Seedlings under Replant Conditions

Xin Xu †, Yifan Zhou ‡, Xiaoqi Wang †, Weitao Jiang †, Lei Qin †, Jian Wang †, Haijun Yu §, Xuesen Chen †, Xiang Shen †, Chengmiao Yin †,*, Zhiquan Mao †,*
PMCID: PMC9947989  PMID: 36844530

Abstract

graphic file with name ao2c06665_0011.jpg

Apple replant disease (ARD) is common in apple production, which seriously affects the growth and development of apples. In this study, hydrogen peroxide with a bactericidal effect was used to treat the replanted soil, and the effects of different concentrations of hydrogen peroxide on replanted seedlings and soil microbiology were investigated in order to seek a green, clean way to control ARD. Five treatments were set up in this study: replanted soil (CK1), replanted soil with methyl bromide fumigation (CK2), replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3). The results showed that hydrogen peroxide treatment improved replanted seedling growth and also inactivated a certain number of Fusarium, while the Bacillus, Mortierella, and Guehomyces also became more abundant in relative terms. The best results were obtained with replanted soil + 4.5% hydrogen peroxide (H3). Consequently, hydrogen peroxide applied to the soil can effectively prevent and control ARD.

1. Introduction

Apple replant disease (ARD) commonly refers to a situation where the old trees were removed, and they become weak or even die when young trees are replanted in the same orchard soil.1 Due to frequent tree species optimization, but limited land resources, it is difficult to avoid replanting cultivation in the original site during orchard renovation, which in turn leads to the widespread occurrence of ARD, and significantly restricts the sustainable development of the apple industry.2 Replanted young apple trees usually exhibit such phenomena as show delayed root extension, slow plant metabolism, poor resistance to stress, and even death of the whole plant.3 ARD causes serious economic losses throughout the whole life cycle of an orchard.4 Thus, it is urgent to develop alternative clean and green measures to control ARD.

ARD was caused by a combination of reasons,5,6 for example, nematodes, oomycetes, and chemosensitive autotoxic substance imbalance in the structure of soil microbiology,7−9 among which soil microbiological imbalance is known to be the main reason for ARD.10,11 When fruit trees are planted in orchards for successive years, beneficial bacteria decrease and the number of soil pathogenic fungi increases, which eventually results in a decrease in the yield of fruit trees.12 In South Africa, Washington, Italy, and other apple producing countries, Fusarium, Pythium, Phytophthora, and Cryptococcus were considered to be harmful pathogens that cause ARD.13,14 Liu15 found that the frequency of Fusarium was high when isolating and identifying harmful fungi in the soil of replanted orchards. Fusarium showed high pathogenicity to the M. hupehensis Rehd. seedlings.16

In the United States, the use of brassicaceae seed meal to mitigate ARD has achieved better results.4,17 However, in China, due to various reasons, soil fumigation is still a common way to control ARD.18,19 However, although traditional soil chemical fumigants have obvious effects, they also have disadvantages such as easy residue, pollution of the natural environment, and potential hazards to human health.20,21 Therefore, it is very important to seek a green, efficient, and reliable measure to solve ARD. Hydrogen peroxide is a simple, safe, economical, and environmentally friendly oxidizing agent, and the by-product of hydrogen peroxide is water, which is often called the “Green Oxidant”. Therefore, the use of hydrogen peroxide has been concerned by researchers for a long time.22 It is widely used in medicine, agriculture, food safety, and clinical and environmental applications.23,24 For example, in the medical field, hydrogen peroxide was often used as a bactericide or disinfectant.25 In the food processing industry, hydrogen peroxide was often used to kill microorganisms on food packaging bags, containers, and disinfectants such as drinking water, which could effectively inhibit the growth of microorganisms.26 In the agricultural field, Kyeong-Hwan et al27 used hydrogen peroxide vapor that interfered with the growth of disease-causing microorganisms. This was because hydrogen peroxide could react with lipid double bonds in the microbial cell wall and enter the microbial interior, acting on proteins and lipids and polysaccharides, altering cell permeability, and leading to cell lysis and death.28 In addition, traditional chemical fumigation was time-consuming and requires film covering. In contrast, hydrogen peroxide to kill harmful microorganism in replanted soils may be a clean, green measure. However, there was little literature describing the application of hydrogen peroxide in affecting the severity of ARD by killing soil microorganisms.

In the present experiment, we evaluated the feasibility of hydrogen peroxide to mitigate ARD under replanting conditions. We aimed to (1) determine the optimal concentration of hydrogen peroxide; (2) how different concentrations of hydrogen peroxide affect the soil microbial community structure; (3) response of replanted seedlings to various concentrations of hydrogen peroxide, thus providing new insights into the ARD mitigation.

2. Materials and Methods

2.1. Experimental Materials

The replanted soil was obtained from an old orchard in Manzhuang Town where apple trees have been planted for 34 years. The soil type was sandy loam. After the top soil was cleaned off, multiple points soil was randomly selected within a depth of 20–40 cm and mixed well. Supplementary Table 1 describes soil basic physicochemical properties.

The experiment material was M. hupehensis Rehd. seedlings, and it is a very widely planted rootstock of apple in China. M. hupehensis Rehd. seeds were stratificated for roughly 40 days at 4 °C. The seeds were planted in seedling cups contained with seedling substrates when white radicles were seen. When the seedling grew to 5–6 true leaves, the seedlings with similar growth, complete leaves, and no pests and diseases were transplanted into tile pots with different soil treatments (pot diameter 24 cm, height 18 cm, and soil weight 7 kg).

Methyl bromide fumigation products are provided by Jiangsu Lianyungang Dead Sea Bromide Co.

Hydrogen peroxide was purchased from Jinan Kunfeng Chemical Co., Ltd., with a concentration of 27.5%.

2.2. Experimental Design

The experiment was performed from April to October 2021 at the Science and Technology Innovation Park of Shandong Agricultural University (36.16°N, 117.1 6°E). A total of 5 treatments were examined: replanted soil (CK1), replanted soil with methyl bromide fumigation (CK2), replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3).

Methyl bromide fumigation treatment and hydrogen peroxide treatment were performed 15 days before the planting of M. hupehensis Rehd. seedlings (performed in mid-April 2021). Methyl bromide fumigant was mixed with replanting soil and put into the sealed trellis film for sealing, the soil layer is controlled about 20 cm, and 50 g of methyl bromide fumigation per square meter of soil is applied. If converted to pot, 0.125 g/kg is applied to each pot, and the temperature is controlled at 15 °C.19 Hydrogen peroxide was diluted in water in three different proportions (1.5, 3.0, and 4.5%). The soil was irrigated until the upper-middle layer of soil reached a saturated state (518 mL). Equal amounts of sterile water were added to CK1 and CK2 treatments as control. Five repetitions were set up for each treatment.

On May 01, 2021, replanted seedlings with consistent growth were planted in each treatment (two seedlings per pot). Each treatment received uniform pruning, irrigation, and management. Plant and soil samples were taken from the 5 treatments on August 15. Each treatment group was sampled by randomly chosen three seedlings. Rhizosphere soil was collected from the pot by removing soil at the top and around the pot, and it was mixed thoroughly. The rhizosphere soil samples were sealed in a resealable bag and divided into three parts after sieving through 20 mesh; one part was air-dried under natural conditions and performed for the measurement of soil enzyme activity; one part was put into a 4 °C refrigerator for the determination of soil microorganisms; and the other was placed at −80 °C for Illumina MiSeq sequencing. When the plant samples were collected, the M. hupehensis Rehd. seedlings were washed. Vigorous white roots were excised and stored in liquid nitrogen for the determination of root enzymes and viability. Fresh and intact leaves were selected from the plant samples and stored at −20 °C to determine the chlorophyll content.

2.3. Indicators for Plant and Soil Measurements

The plant heights and ground diameters were measured with conventional methods such as a pylon ruler and dial calipers. The soil on the seedlings was washed off by water, excess water was wiped off, and the fresh seedlings were weighed with an electronic balance. After the determination, it was quenched at 105 °C for 30 min and dried at 65 °C, and the dry mass was weighed. The determination of the chlorophyll content was carried out by extraction with ethanol29 (Supplementary Material 1.1). The determination of photosynthetic parameters was carried out on August 14 (sunny day, no wind) from 9:00 am to 11:00 am (Supplementary Material 1.2). The TTC method was used to determine the root respiration rate30 (Supplementary Material 1.3). The determination of root antioxidant enzyme activity refers to the method of Singh et al.31 (Supplementary Material 1.4).

The number of culturable microorganisms in the soil was measured by the dilution plate counting method,32 and the culturable bacteria and fungi were measured by Luria-Bertani, potato dextrose agar (Supplementary Material 1.5). Soil enzyme activity was measured by referring to the method determined by Chen et al.33 Soil sucrase activity was determined by the 3-amino-5-nitrosalicylic acid colorimetric method. Soil urease activity was determined by the sodium phenol–sodium hypochlorite colorimetric method. Soil neutral phosphatase activity was performed with the phenyl disodium phosphate colorimetric method (Supplementary Material 1.6). DNA was obtained from 0.5 g of fresh soil by the E.Z.N.A. soil DNA extraction kit.34 The gene copy number of F. oxysporum was determined by real-time fluorescence quantitative polymerase chain reaction using a Bio-Rad CFX96 quantitative PCR instrument (Supplementary Material 1.7). JR (5′-GGCCTGAGGG TTGTAATG-3′) and JF (5′-CG AGTTATACAACTCATCAACC-3′) were the primers employed for the reaction. Soil microbial communities in Illumina MiSeq sequencing: 338F (5′-ACTCCTACGGGAGGCAGCAG-3′) and 806R (5′-GGACTACHVGGGTWTCTAAT-3′) were 16S rRNA primer sequences,35 ITS1F (5′-CTTGGTCATTTAGAGGAAGTAA-3′) and ITS2R (5′-GCTGCGTTCTTCATCGATGC-3′) were ITS primer sequences.36

2.4. Data Analysis

The original sequencing was filtered out using fastp software to eliminate the ones smaller than 50 bp. According to their overlapped sequences, sequences over 10 bp were assembled using flash software. Operational taxonomic unit (OTU) clustering analysis was usually conducted with 97% similarity. Based on OTU results, the alpha diversity index was calculated in different treatments by using mothur (https://mothur.org/wiki/calculators/.version 1.30.2) and plotted with origin 2018 software (Origin Lab Corporation, USA). The rarefaction curves were plotted by R language, and the R language (version 3.3.1) vegan package was employed for Cluster heat map analysis. Principal coordinate analysis (PCoA) was calculated for each sample at the genus level. The significance of the difference between multiple groups was tested by the Kruskal–Wallis H test. Functional annotation and prediction of bacterial and fungal communities were performed using PICRUSt1 and FUNGuild prediction analysis, respectively. SPSS 26.0 (IBM SPSS Statistics, IBM Corporation, USA) was used for mean comparison and single-factor analysis of variance. The Duncan-style new multiple range method was used for significant difference comparison, and data were composed of mean ± standard error. Origin 2018 software was used to construct the figures.

3. Results

3.1. Influences of Hydrogen Peroxide on the Growth of Replanted Seedlings

There were significant differences in the growth status of replanted seedlings under different concentrations of hydrogen peroxide treatment (Figure 1). H2 and H3 treatments obviously improved the growth of replanted seedlings; among them, H3 treatment had the best effect. The plant height, ground diameter, fresh weight, and dry weight of the H3 treatment increased 0.63, 1.27, 8.47, and 9.69 times, respectively, compared to CK1. Compared with CK2, H3 treatment increased 0.23, 0.06, 0.82, and 0.87 times in the plant height, ground diameter, fresh weight, and dry weight, respectively (Figure 1).

Figure 1.

Figure 1

Open in a new tab

Growth status of replanted seedlings. (a) Plant height; (b) ground diameter; (c) fresh weight; (d) dry weight; CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide; letters reflect different degrees of treatment variation (P < 0.05).

3.2. Influences of Hydrogen Peroxide on the Chlorophyll Content and Photosynthetic Parameters of Replanted Seedlings

Replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) all significantly increased the chlorophyll a and b contents (Table 1). Among them, the treatment with replanted soil + 4.5% hydrogen peroxide (H3) had the best effect, followed by CK2. Compared with CK1, chlorophyll a, b increased by 15.8 and 11.0% in the replanted soil + 1.5% hydrogen peroxide (H1) treatment; 23.6 and 24.5% in the replanted soil + 3.0% hydrogen peroxide (H2) treatment; and 40.0 and 79.3% in replanted soil + 4.5% hydrogen peroxide (H3) treatment, respectively. The differences in chlorophyll a, b between H1 and H2 were not significant.

Table 1. Effects of Different Treatments on Chlorophyll a and Chlorophyll b in the Leaves of Replanted Seedlingsa.

treatment chlorophyll a (mg·g–1·FW) chlorophyll b (mg·g–1·FW)
CK1 18.45 ± 0.77d 5.46 ± 0.54d
CK2 23.44 ± 0.20b 8.23 ± 0.45ab
H1 21.37 ± 0.68c 6.06 ± 0.35d
H2 22.81 ± 0.56bc 6.80 ± 0.49cd
H3 25.83 ± 0.68a 9.79 ± 0.74a
Open in a new tab
a

Note: letters reflect different degrees of treatment variation (P < 0.05).

Replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments all significantly improved replanted seedling photosynthetic parameters (Figure 2). The application of replanted soil + 4.5% hydrogen peroxide (H3) was shown to be significantly different from all other treatments. Compared with CK1, intercellular carbon dioxide concentration, net photosynthetic rate, stomatal conductance, and transpiration rate of replanted soil + 4.5% hydrogen peroxide (H3) treatment increased by 14.7, 106.3, 81.8, and 56.7%, respectively. The difference in the intercellular carbon dioxide concentration between replanted soil + 1.5% hydrogen peroxide (H1) and replanted soil + 3.0% hydrogen peroxide (H2) treatments was not significant.

Figure 2.

Figure 2

Open in a new tab

Effect of hydrogen peroxide treatment on photosynthetic parameters of replanted seedlings. (a) Net photosynthetic rate; (b) intercellular carbon dioxide concentration; (c) transpiration rate; (d) stomatal conductance; CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide; letters reflect different degrees of treatment variation (P < 0.05).

3.3. Influences of Hydrogen Peroxide on Root Antioxidant Enzyme Activities and the Root Respiration Rate of Replanted Seedlings

Replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments all significantly improved root antioxidant enzyme activity (Figure 3). Compared with CK2, the superoxide dismutase (SOD) and catalase (CAT) activities of H3 increased by 21.55 and 4.93%, respectively. Significant differences were observed in the root antioxidant enzyme activities of replanted soil (CK1) and hydrogen peroxide treatment. The H1, H2, and H3 treatments also significantly increased the root respiration rates by 26.1, 51.3, and 96.4%, respectively, compared to replanted soil (CK1).

Figure 3.

Figure 3

Open in a new tab

Effects of root antioxidant enzymes and the root respiration rate of replanted seedlings under hydrogen peroxide treatment. (a) Superoxide dismutase activity; (b) catalase activity; (c) peroxidase activity; (d) root respiration rate; letters reflect different degrees of treatment variation (P < 0.05).

3.4. Influences of Hydrogen Peroxide on Soil Enzyme Activity

Replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments all significantly reduced the activity of soil enzymes (Figure 4). Among them, replanted soil with CK2 had the largest decrease, followed by replanted soil + 4.5% hydrogen peroxide (H3) treatment. Compared to CK1, sucrase, urease, and phosphatase activity were reduced by 3.80, 22.2, and 14.1%, in replanted soil + 1.5% hydrogen peroxide (H1); 17.5, 33.3, and 25.3%, in replanted soil + 3.0% hydrogen peroxide (H2); and 24.9, 44.4, and 36.5% in replanted soil + 4.5% hydrogen peroxide (H3), respectively.

Figure 4.

Figure 4

Open in a new tab

Effect of hydrogen peroxide treatment on soil enzyme activity. (a) Sucrase activity; (b) urease activity; (c) phosphatase activity. CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide; letters reflect different degrees of treatment variations (P < 0.05).

3.5. Analysis of Culturable Microorganisms and RT-qPCR Analysis of Fusarium oxysporum

The addition of different concentrations of hydrogen peroxide can significantly improve the environment of replanted soil (Figure 5). In particular, replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments significantly reduced the bacteria, fungi, and Fusarium oxysporum numbers in the soil. Replanted soil + 4.5% hydrogen peroxide (H3) treatment had the largest decrease, followed by replanted soil + 3.0% hydrogen peroxide (H2) and finally replanted soil + 1.5% hydrogen peroxide (H1). Compared with the replanted soil (CK1), soil bacteria and fungi were decreased by 34.6 and 58.9% in the replanted soil + 1.5% hydrogen peroxide (H1) treatment; 73.1 and 70.5% in the replanted soil + 3.0% hydrogen peroxide (H2) treatment; and 91.0 and 87.6% in the replanted soil + 4.5% hydrogen peroxide (H3) treatment, respectively. In addition, compared with replanted soil (CK1), Fusarium oxysporum of replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments decreased by 44.5, 66.6, and 90.8%, respectively. The gene copy number of F. oxysporum in the other treatments was lower than that in replanted soil (CK1).

Figure 5.

Figure 5

Open in a new tab

Changes in soil microbial population under different treatments. (a) Number of soil bacteria; (b) number of soil fungi; (c) gene copy number of F. oxysporum; CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide; letters reflect different degrees of treatment variations (P < 0.05).

3.6. Analysis of Soil Microbial Community Composition at the Genus Level

By sequencing, the dilution curve of fungi and bacteria tend to be flat, which indicated that the data volume of microbial communities of soil environmental samples was close to saturation, and it indicated that the current sequencing quantities could reflect the majority of microbial diversity information in the samples (Figure S1). The abundance heatmap for bacteria and fungi genera was constructed from the top 10 dominant species in the sample. Arthrobacter, Sphingomona, RB41, Bacillus, and Gaiella were the main bacterial species, except for unclassified bacteria (Figure 6a). Mortierella, Pseudallescheria, Fusarium, Lophiostoma, Guehomyces, Humicota, Trichoderma, and Kermia were the dominant fungal species except for the unclassified fungi (Figure 6b). The species of bacterial and fungal genera were basically the same between treatments, but their relative abundance differed significantly. The comparative abundance of Bacillus increased by 14.03% for the H3 treatment compared to CK1; the comparative abundance of Fusarium decreased by 40.29% for the H3 treatment compared to CK1. In addition, compared with CK1, CK2, H1, H2, and H3 treatments obviously increased the comparative abundance of Mortierella, and the largest increase was observed in the treatment of replant soil + 4.5% hydrogen peroxide (H3), followed by CK2 treatment. Compared to CK1, replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments significantly increased the comparative abundance of Trichoderma, with CK2 showing the largest increase.

Figure 6.

Figure 6

Open in a new tab

Differences between species of bacterial genus (a) and fungal genus (b) for different treatments. CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide.

3.7. Differences in the Microbial Community Composition in Different Treatments

The PCoA plot showed that PC1 and PC2 explained 75.85% of the changes in bacterial communities and 61.98% of the changes in fungal communities, respectively (Figure 7). Microbial communities were significantly different between treatments, CK2 being the most distant from CK1, meaning that the differences were the greatest. The three different hydrogen peroxide concentration treatments also had a certain distance from the CK1, and the application of hydrogen peroxide in the replanted soil might have an impact on the change of the microbial community structure. The treatments were subjected to the Kruskal–Wallis H test based on the different microbial community structures, at the level of bacterial genera, Gaiella, Pedomicrobium, Romboutsia, and Turicibacter that were more abundant in the hydrogen peroxide-treated soils, with CK2 treatment being the most pronounced (Figure S2). At the fungal genus level, the comparative abundance of Mortierella, Guehomyces in the soil treated with hydrogen peroxide was significantly higher, while the comparative abundance of Fusarium and Lophiostoma was significantly lower (Figure S2).

Figure 7.

Figure 7

Open in a new tab

PCoA was calculated for each sample on the basis of the Bray–Curtis distance matrix at the bacterial genus (a) and fungal genus (b) level. CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide.

3.8. Analysis of Microbial Alpha Diversity under Different Concentrations of Hydrogen Peroxide Treatment

The Simpson and Chao indices of bacteria and fungi in soil treatments with different concentrations of hydrogen peroxide were significantly different. In general, the Simpson index often estimated microbial diversity in samples, and the smaller the Simpson index, the higher the community diversity. The Simpson indices of replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments were all higher than those of CK1, indicating that the microbial diversity of soil treated with hydrogen peroxide was lower (Figure 8a,c). The Chao index was used to reflect the abundance of microorganisms, and the comparative abundance of bacteria and fungi in the hydrogen peroxide-treated soil showed a decreasing trend, with significant differences in the Chao index of bacterial genera between treatments. The Chao index of fungi genera in CK2 and H3 treatments showed some differences (Figure 8b,d).

Figure 8.

Figure 8

Open in a new tab

Diversity of microorganisms under different treatments. Simpson index for species of the genera bacteria (a) and fungi (c); Chao index for species of the genera bacteria (b) and fungi (d). CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide; letters reflect different degrees of treatment variations (P < 0.05).

3.9. Microbial Community Function Prediction

PICRUSt1 and FUNGuild prediction analyses were made to analyze the possible functions of bacterial and fungal communities in the rhizosphere soil, respectively. PICRUSt1 results indicated that a total of 24 taxon functions were obtained in the bacterial communities. Except for the unclassified functions and prediction-only functions, the top three functions were amino acid transport and metabolism, energy production and conversion, and signal transduction mechanisms, and their relative abundance in the bacterial community was 4.68, 4.01, and 3.72%, respectively (Figure 9a). Among the fungal communities, undefined saprotroph, endophyte-litter saprotroph-soil saprotroph-undefined saprotroph, animal pathogen-endophyte-lichen parasite-plant pathogen-soil saprotroph-wood saprotroph were the three function taxa with higher abundance (Figure 9b). Among them, the only functional group that corresponds to the genus Mortierella belongs to is endophyte-litter saprotroph-soil saprotroph-undefined saprotroph, and it had the highest abundance in the H3 treatment. The only fungal genera corresponding to animal pathogen-endophyte-lichen parasite-plant pathogen-soil saprotroph-wood saprotroph was Fusarium, and the abundance of Fusarium was significantly lower in the replanted soil + 3.0% hydrogen peroxide (H2) and replanted soil + 4.5% hydrogen peroxide (H3) treatments than in CK1(Table S2).

Figure 9.

Figure 9

Open in a new tab

Predicted function of bacterial (a) community and fungal (b) community. CK1: replanted soil; CK2: replanted soil with methyl bromide fumigation; H1: replanted soil + 1.5% hydrogen peroxide; H2: replanted soil + 3.0% hydrogen peroxide; H3: replanted soil + 4.5% hydrogen peroxide.

4. Discussion

ARD can lead to stunted tree growth, root rot, reduced fruit production, and then result in tree mortality, ultimately shortening the life span of replanted apple orchards.37,38 The main causes of ARD include increased numbers of harmful fungi and changes in the microbial community structure in the soil under replanting conditions.19,39 This study showed that different concentrations of hydrogen peroxide could promote the replanted seedling growth from the physiological indicators, and both the plant biomass of replanted soil + 3.0% hydrogen peroxide (H2) and replanted soil + 4.5% hydrogen peroxide (H3) treatments were obviously higher than that of CK1. This status was likely due to hydrogen peroxide killing pathogenic fungi in the soil, improving the soil environment for replanting and promoting plant growth.40 Plant growth cannot be achieved without photosynthesis, and chlorophyll is the main photosensitive pigment for photosynthesis to absorb light energy, which has an important impact on plant growth.41 Therefore, the chlorophyll content affects photosynthetic parameters, which in turn affects the growth of M. hupehensis Rehd. seedlings.42 Ozaki et al.43 found that the photosynthetic rate of melon leaves was also enhanced when treated with hydrogen peroxide. Compared with CK1, the chlorophyll content and photosynthetic parameters were obviously varied among different hydrogen peroxide treatments, with replanted soil + 4.5% hydrogen peroxide (H3) treatment being the most effective. Hydrogen peroxide impacts the structure of soil microbial communities and changes the microbial-mediated soil nutrient conversion process to facilitate nutrient uptake by crops. Therefore, the growth of the plants was enhanced and their chlorophyll content and photosynthetic parameters were also enhanced.44

Most studies have shown that in general, free radical production and elimination in plants often remain in relative balance. When confronted with adversity conditions, the production rate of free radicals was far greater than the elimination rate, and the plant was damaged.45 Under the condition of long-term continuous cropping, the deteriorating soil environment could create a threat of adversity in apple seedlings. The relative balance of free radicals was broken, and seedling growth was threatened. Superoxide dismutase, peroxidase, and catalase were the three important factors in the protection of plants from excessive free radicals. The three work synergistically to protect the inner membrane structure of the plant body and reduce the damage to the membrane structure by free radicals.46 In this study, it was found that hydrogen peroxide treatment enhanced the level of antioxidant enzymes in plants. The main reason is that hydrogen peroxide has a strong oxidizing and sterilizing effect, which purifies the soil environment and reduces microbial damage to the root system, thus promoting the growth of plants.47 The second may have a small amount of hydrogen peroxide surviving in the soil that can directly or indirectly activate antioxidant enzymes during stress, thus increasing the induced resistance of the plant.48 Studies have demonstrated that hydrogen peroxide provides a more vigorous root system in wheat.49 Hydrogen peroxide can also promote rooting and root growth of ground cover chrysanthemum plugs.50

A good rhizosphere microbial community structure could maintain the balance of soil microhabitats and ensure normal growth of plants.51 Soil microorganisms were important for plant growth as an important indicator for assessing soil ecosystems.52 Generally speaking, beneficial soil microorganisms have a positive effect on plant development, while harmful microorganisms can hinder plant development, and even result in death. For example, Fusarium wilt in many important crops worldwide is caused by Fusarium.32 RT-PCR analysis illustrated that that CK2, H1, H2, H3 treatments obviously decreased the gene copy number of F. oxysporium. The methyl bromide fumigant has great potential to kill pests and pathogens in the soil and also reduces the population of Fusarium.(32) Hydrogen peroxide possesses microbicidal and sporicidal activity.53

Research shows that Mortierella has a symbiotic or reciprocal relationship with plants.54Mortierella converts insoluble phosphorus and potassium in the soil into available phosphorus and potassium for plant uptake and utilization, thus improving plant resistance.55Mortierella was shown to have a negative relationship with the occurrence of ARD and could play a vital role in suppressing ARD by competing for nutrients or resisting nutrients.56Fusarium was severely positively interrelated with ARD in China,57,58 and it exhibits strong pathogenicity.56Bacillus not only inhibited the reproduction of pathogenic fungi, improved microbial community structure, and enhanced plant disease resistance but also secreted catabolic phytase to increase the amount of free phosphorus in the soil, promoted phosphorus uptake by plants, and improved the crop yield.59 Making fungi fertilizer from Trichoderma asperellum strain both promoted replanted seeding growth and development, optimized the soil microhabitats, and effectively alleviated ARD.60 In aquatic and terrestrial habitats, Lophiostom commonly occurs as sapwood on branches, stems, or bark in woody and herbaceous plants.61 Intaraudom et al.62 found that the secondary metabolites of Lophiostoma bipolare BCC25910 had no obvious inhibitory effect on disease-causing bacteria. In this experiment, the soil microbial community after hydrogen peroxide treatment was studied using amplicon sequencing technology. We found that soil treatment with hydrogen peroxide significantly improved the comparative abundance of Bacillus, Pedomicrobium, Mortierella, Guehomyces, and Trichoderma and significantly reduced the comparative abundance of Fusarium and Lophiostoma, with the most obvious effect of replanted soil + 4.5% hydrogen peroxide (H3) treatment. Hydrogen peroxide treatment significantly altered the soil microhabitats and distinctly reduced the number of Fusarium. Li63 found that the accumulation of hydrogen peroxide has the effect of directly poisoning and killing pathogenic bacteria.

The diversity and richness of species were commonly expressed by Alpha diversity. In this study, the simpson index of replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments was higher than that of CK1, indicating that the microbial diversity of replanted soil + 1.5% hydrogen peroxide (H1), replanted soil + 3.0% hydrogen peroxide (H2), and replanted soil + 4.5% hydrogen peroxide (H3) treatments was lower than that of CK1. Li64 found a decreased microbial community diversity after soil fumigant was applied, and this is the same result as that in our study. Hydrogen peroxide is a broad-spectrum disinfectant that inhibits harmful microorganisms while also having an effect on beneficial microorganisms, causing a “vacuum” in the soil.65 Soil enzymes are the results of soil microbial metabolism and decomposition of plant and animal residues,66 and it is involved in a series of biochemical reactions in the soil.67 In this study, hydrogen peroxide treatment reduced the soil enzyme activity, and the higher the application concentration, the more obvious the reduction in soil enzyme activity. At the same time, the soil microbial population also decreased to different degrees. This is probably due to the application of hydrogen peroxide that killed some of the microorganisms associated with soil enzyme activity in the soil, which in turn led to a decrease in soil enzyme activity. This is in agreement with the findings of Klose et al.68 who used bromomethane fumigation of soil to cause a decrease in soil microbial population and soil enzyme activity.

In the present study, 24 bacterial taxa functions were obtained by PICRUSt1 for bacteria community prediction analysis, among which amino acid transport and metabolism functions were more abundant, which was consistent with the results of Yang et al.69 FUNGuild prediction analysis showed that animal pathogen-endophyte-lichen parasite-plant pathogen-soil saprotroph-wood saprotroph corresponded to the fungi genus was only Fusarium, and the higher functional abundance of Fusarium corresponded to its higher abundance in the functional group composition. The high functional abundance of Fusarium was in line with the higher abundance of its functional taxa. Correspondingly, the Fusarium abundance in replanted soil (CK1) was higher. FUNGuild suggests that Fusarium was the key causal agent of ARD. The abundance of Mortierella in soil after hydrogen peroxide treatment was obviously high, compared to replanted soil, and the most obvious treatment was with replanted soil + 4.5% hydrogen peroxide (H3) treatment. Free radicals produced by hydrogen peroxide are known to damage DNA of spores in one of the species.70 This indicates that hydrogen peroxide reduced the abundance of Fusarium and increased the number of beneficial microorganisms, which effectively improved the soil microhabitat and alleviated ARD.

5. Conclusions

In this experiment, we found that the hydrogen peroxide treatment promoted the growth and development of replanted seedlings and improved the chlorophyll content, photosynthetic parameters, and root antioxidant enzyme activities of seedlings. Soil microhabitat also changed to different degrees, especially, increased the comparative abundance of Bacillus, Mortierella, decreased the comparative abundance of Fusarium. In summary, the effect of replanted soil + 4.5% hydrogen peroxide (H3) treatment was the most significant, which effectively alleviated ARD.

Acknowledgments

The research was supported by the China Agriculture Research System of MOF and MARA (Grant No. CARS-27), the National Natural Science Foundation of China (Grant No. 32072510), the Qingchuang Science and Technology Support Project of Shandong Colleges and Universities (No. 2019KJF020), the Natural Science Foundation of Shandong Province (Grant No. ZR2020MC131), Shandong Agricultural Major Applied Technology Innovation Project (Grant No. SD2019ZZ008), the National Key Research and Development Program of China (Grant No. 2020YFD1000201), Taishan Scholars Funded Project (No. ts20190923), and the Fruit Innovation Team in Shandong Province, China (Grant No. SDAIT-06-07).

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.2c06665.

  • Description of methods for the determination of indicators such as root antioxidant enzyme activity and soil culturable microorganisms; information about soil physicochemical properties; dilution curves; the Kruskal–Wallis test used to statistically compare the relative abundance of bacterial and fungal genera on different treatments; and distribution of OTUs of functional taxa (PDF)

Author Contributions

X.X. and Y.F.Z. contributed equally to this work as first author. All authors read and approved the manuscript.

The authors declare no competing financial interest.

Supplementary Material

ao2c06665_si_001.pdf (2.8MB, pdf)

References

  1. Tewoldemedhin Y. T.; Mazzola M.; Labuschagne I.; Adéle M. A multiphasic approach reveals that apple replant disease is caused by multiple biological agents, with some agents acting synergistically. Soil Biol. Biochem. 2011, 43, 1917–1927. 10.1016/j.soilbio.2011.05.014. [DOI] [Google Scholar]
  2. Mazzola M.; Manici L. M. Apple replant disease: role of microbial ecology in cause and conteol. Annu. Rev. Phytopathol. 2012, 50, 45–65. 10.1146/annurev-phyto-081211-173005. [DOI] [PubMed] [Google Scholar]
  3. Wang X. Q.; Yao Y. Y.; Wang G. W.; Ma J. Z.; Yin C. M.; Chen X. S.; Mao Z. Q. Comprehensive Analysis of the Influence of Fulvic Acid from Paper Mill Effluent on Soil Properties, Soil Microbiome, and Growth of Malus hupehensis Rehd. Seedlings under Replant Conditions. ACS Omega 2021, 6, 24027–24038. 10.1021/acsomega.1c03201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. DuPont S.; Hewavitharana S.; Mazzola M. Field scale application of Brassica seed meal and anaerobic soil disinfestation for the control of apple replant disease. Appl. Soil Ecol. 2021, 166, 104076 10.1016/j.apsoil.2021.104076. [DOI] [Google Scholar]
  5. Rumberger A.; Yao S.; Merwin I. A.; Nelson E. B.; Thies J. E. Rootstock genotype and orchard replant position rather than soil fumigation or compost amendment determine tree growth and rhizosphere bacterial community composition in an apple replant soil. Plant Soil 2004, 264, 247–260. 10.1023/B:PLSO.0000047760.13004.94. [DOI] [Google Scholar]
  6. Pan F. B.; Xiang L.; Wang S.; Li J. J.; Shen X.; Chen X. S.; Yin C. M.; Mao Z. Q. Effects of short-term rotation and Trichoderma application on the soil environment and physiological characteristics of Malus hupehensis Rehd. seedlings under replant conditions. Acta Ecol. Sin. 2017, 37, 315–321. 10.1016/j.chnaes.2017.09.003. [DOI] [Google Scholar]
  7. Reardon C. L.; Strauss S. L.; Mazzola M. Changes in available nitrogen and nematode abundance in response to Brassica seed meal amendment of orchard soil. Soil Biol. Biochem. 2013, 57, 22–29. 10.1016/j.soilbio.2012.10.011. [DOI] [Google Scholar]
  8. Manici L. M.; Lazzeri L.; Palmieri S. In vitro fungitoxic activity of some glucosinolates and their enzyme-derived products toward plant pathogenic fungi. J. Agric. Food Chem. 1997, 45, 2768–2773. 10.1021/jf9608635. [DOI] [Google Scholar]
  9. Yin C. M.; Hu Y. L.; Wang G. S.; Zhang X. F.; Zhou H.; Shen X.; Chen X. S.; Mao Z. Q. Effects of major phenolic acids in apple replanting soil on the root system of Malus hupehensis Rehd. Seedlings. China Agric. Sci. 2016, 49, 961–969. [Google Scholar]
  10. Mazzola M.; Graham D.; Wang L.; Leisso R.; Hewavitharana S. S. Application sequence modulates microbiome composition, plant growth and apple replant disease control efficiency upon integration of anaerobic soil disinfestation and mustard seed meal amendment. Crop Protect. 2020, 132, 105125 10.1016/j.cropro.2020.105125. [DOI] [Google Scholar]
  11. Tilston E. L.; Deakin G.; Bennett J.; Passey T.; Harrison N.; Fernández F.; Xu X. M. Effect of fungal, oomycete and nematode interactions on apple root development in replant soil. CABI Agric. Biosci. 2020, 1, 14. 10.1186/s43170-020-00014-7. [DOI] [Google Scholar]
  12. Spath M.; Insam H.; Peintner U.; Kelderer M.; Kuhnert R.; Franke-Whittle I. H. Linking soil biotic and abiotic factors to apple replant disease: a greenhouse approach. J. Phytopathol. 2015, 163, 287–299. 10.1111/jph.12318. [DOI] [Google Scholar]
  13. Manici L. M.; Ciavatta C.; Kelderer M.; Erschbaumer G. Replant problems in South Tyrol: role of fungal pathogens and microbial population in conventional and organic apple orchards. Plant Soil 2003, 256, 315–324. 10.1023/A:1026103001592. [DOI] [Google Scholar]
  14. Van-Schoor L.; Denman S.; Cook N. C. Characterisation of apple replant disease under South African conditions and potential biological management strategies. Sci. Hortic. 2009, 119, 153–162. 10.1016/j.scienta.2008.07.032. [DOI] [Google Scholar]
  15. Liu Z.The isolation and identification of fungi pathogen from apple replant disease in Bohai Bay reaior and the screening of the antagonistic Trichoderma strains; Shandong Agricultural University. 2013. [Google Scholar]
  16. Franke-Whittle I. H.; Manici L. M.; Insam H.; Stres B. Rhizosphere bacteria and fungi associated with plant growth in soils of three replanted apple orchards. Plant Soil 2015, 395, 317–333. 10.1007/s11104-015-2562-x. [DOI] [Google Scholar]
  17. Mazzola M. Advances in Brassicaceae seed meal formulation and application for replant disease control in organic apple orchards. Plant Sci. 2011, 101, S117. [Google Scholar]
  18. Chen R.; Jiang W. T.; Xu S. Z.; Fan H.; Chen X. S.; Shen X.; Yin C. M.; Mao Z. Q. An emerging chemical fumigant: two-sided effects of dazomet on soil microbial environment and plant response. Environ. Sci. Pollut. Res. 2021, 29, 3022–3036. 10.1007/s11356-021-15401-4. [DOI] [PubMed] [Google Scholar]
  19. Jiang W. T.; Chen R.; Zhao L.; Qin L.; Fan H.; Chen X. S.; Wang Y. F.; Yin C. M.; Mao Z. Q. Chemical fumigants control apple replant disease: Microbial community structure-mediated inhibition of Fusarium and degradation of phenolic acids. J. Hazard. Mater. 2022, 440, 129786 10.1016/j.jhazmat.2022.129786. [DOI] [PubMed] [Google Scholar]
  20. Zhu Y.; Fazio G.; Mazzola M. Elucidating the molecular responses of apple rootstock resistant to ARD pathogens: challenges and opportunities for development of genomics-assisted breeding tools. Hortic. Res. 2014, 1, 14043. 10.1038/hortres.2014.43. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Somera T. S.; Freilich S.; Mazzola M. Comprehensive analysis of the apple rhizobiome as influenced by different Brassica seed meals and rootstocks in the same soil/plant system. Appl. Soil Ecol. 2021, 157, 103766 10.1016/j.apsoil.2020.103766. [DOI] [Google Scholar]
  22. He Y.Study on green oxidation reaction using hydrogen peroxide as oxidant; Nanjing University of Science and Technology, 2014. [Google Scholar]
  23. Chen M. N.; Li X.; Yang Q.; Chi X. Y.; Pan L. J.; Chen N.; Yang Z.; Wang T.; Wang M.; Yu S. L. Soil eukaryotic microorganism succession as affected by continuous cropping of peanut-pathogenic and beneficial fungi were selected. PLoS One 2012, 7, e40659 10.1371/journal.pone.0040659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Zhu X. X.; Chen W.; Wu K. L.; Li H. Y.; Fu M.; Liu Q.; Zhang X. A colorimetric sensor of H2O2 based on Co3O4-montmorillonite nanocomposites with peroxidase activity. New J. Chem. 2018, 42, 1501–1509. 10.1039/C7NJ03880A. [DOI] [Google Scholar]
  25. Checinska A.; Fruth I. A.; Green T. L.; Crawford R. L.; Paszczynski A. J. Sterilization of biological pathogens using supercritical fluid carbon dioxide containing water and hydrogen peroxide. J. Microbiol. Methods 2011, 87, 70–75. 10.1016/j.mimet.2011.07.008. [DOI] [PubMed] [Google Scholar]
  26. Salazar P.; Rico V.; González-Elipe A. R. Non-enzymatic hydrogen peroxide detection at NiO nanoporous thin film-electrodes prepared by physical vapor deposition at oblique angles. Electrochim. Acta 2017, 235, 534–542. 10.1016/j.electacta.2017.03.087. [DOI] [Google Scholar]
  27. Kyeong-Hwan B.; Jae-Won H.; Dong-Hyun K. Effect of hydrogen peroxide vapor treatment for inactivating Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes on organic fresh lettuce. Food Control 2014, 44, 78–85. 10.1016/j.foodcont.2014.03.046. [DOI] [Google Scholar]
  28. Mcdonnell G.; Russell A. D. Antiseptics and disinfectants: activity, action, and resistance. Clin. Microbiol. Rev. 2001, 14, 227–227. 10.1128/CMR.14.1.227-227.2001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Wang M.; Yin C. M.; Sun M. M.; Yang M. F.; Feng F. L.; Chen X. S.; Shen X.; Mao Z. Q. Effect of xanthic acid microbicides on photosynthetic characteristics of continuous Malus hupehensis Rehd. J. Plant Physiol. 2019, 55, 99–106. [Google Scholar]
  30. Zheng J.; Ying Q.; Fang C.; Sun N.; Si M. L.; Yang J.; Zhu B.; Ruan Y. L.; Zhu Z. J.; He Y. Alternative oxidase pathway is likely involved in waterlogging tolerance of watermelon. Sci. Hortic. 2021, 278, 109831 10.1016/j.scienta.2020.109831. [DOI] [Google Scholar]
  31. Singh B. K.; Sharma S. R.; Singh B. Antioxidant enzymes in cabbage: variability and inheritance of superoxide dismutase, peroxidase and catalase. Sci. Hortic. 2010, 124, 9–13. 10.1016/j.scienta.2009.12.011. [DOI] [Google Scholar]
  32. Zhang R.; Yan Z.; Wang Y. K.; Chen X. S.; Yin C. M.; Mao Z. Q. Effects of Trichoderma harzianum fertilizer on the soil environment of Malus hupehensis Rehd. seedlings under replant conditions. Hortscience 2021, 56, 1073–1079. 10.21273/HORTSCI15970-21. [DOI] [Google Scholar]
  33. Chen R.; Jiang W. T.; Duan Y. N.; Qiao H. Y.; Fan H.; Chen X. S.; Shen X.; Yin C. M.; Mao Z. Q. Effect of Emerging Soil Chemical Amendments on the Replant Soil Environment and Growth of Malus hupehensis Rehd. Seedlings. ACS Omega 2021, 6, 20445–20454. 10.1021/acsomega.1c02447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wang M.; Duan Y. N.; Sun S. Y.; Xiang L.; Wang G. S.; Chen X. S.; Shen X.; Yin C. M.; Mao Z. Q.. Effects of Different Nitrogen Forms on the Growth of Malus hupehensis Rehd. Seedlings and the Number of Fusarium oxysporum in Soil; 2017, 23, 1014–1021.
  35. Xu N.; Tan G. C.; Wang H. Y.; Gai X. P. Effect of biochar additions to soil on nitrogen leaching, microbial biomass and bacterial community structure. Eur. J. Soil Biol. 2016, 74, 1–8. 10.1016/j.ejsobi.2016.02.004. [DOI] [Google Scholar]
  36. Adams R. I.; Miletto M.; Taylor J. W.; Bruns T. D. Dispersal in microbes: Fungi in indoor air are dominated by outdoor air and show dispersal limitation at short distances. ISME J. 2013, 7, 1262–1273. 10.1038/ismej.2013.28. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Laurent A. S.; Merwin I. A.; Thies J. E. Long-term orchard groundcover management systems affect soil microbial communities and apple replant disease severity. Plant Soil 2008, 304, 209–225. 10.1007/s11104-008-9541-4. [DOI] [Google Scholar]
  38. Wang Y. F.; Pan F. B.; Wang G. S.; Zhang G. D.; Wang Y. L.; Chen X. S.; Mao Z. Q. Effects of biochar on photosynthesis and antioxidative system of Malus hupehensis Rehd. seedlings under replant conditions. Sci. Hortic. 2014, 175, 9–15. 10.1016/j.scienta.2014.05.029. [DOI] [Google Scholar]
  39. Mazzola M. Elucidation of the microbial complex having a causal role in the development of apple replant disease in Washington. Phytopathology 1998, 88, 930–938. 10.1094/PHYTO.1998.88.9.930. [DOI] [PubMed] [Google Scholar]
  40. Szpunar-Krok E.; Jańczak-Pieniążek M.; Skrobacz K.; Bobrecka-Jamro D.; Balawejder M. Response of Potato (Solanum Tuberosum L.) Plants to Spraying by Hydrogen Peroxide. Sustainability 2020, 12, 2469. 10.3390/su12062469. [DOI] [Google Scholar]
  41. Wang S.; Wang X. D.; Shi X. B.; Wang B. L.; Zheng X. C.; Wang H. B.; Liu F. Z. Red and blue lights significantly affect photosynthetic properties and ultrastructure of mesophyll cells in senescing grape leaves. Hortic. Plant J. 2016, 2, 82–90. 10.1016/j.hpj.2016.03.001. [DOI] [Google Scholar]
  42. Li Y.; He N. P.; Hou J. H.; Xu L.; Liu C. C.; Zhang J. H.; Wang Q. F.; Zhang X. M.; Wu X. Q. Factors Influencing Leaf Chlorophyll Content in Natural Forests at the Biome Scale. Front. Ecol. Evol. 2018, 6, 64. 10.3389/fevo.2018.00064. [DOI] [Google Scholar]
  43. Ozaki K.; Uchida A.; Takabe T.; Shinagawa F.; Tanaka Y.; Takabe T.; Hayashi T.; Hattori T.; Rai A. K.; Takabe T. Enrichment of sugar content in melon fruits by hydrogen peroxide treatment. J. Plant Physiol. 2008, 166, 569–578. 10.1016/j.jplph.2008.08.007. [DOI] [PubMed] [Google Scholar]
  44. Huang B.Effect of fumigation on soil carbon and phosphorus and its microbiological mechanism. Ph.D Thesis, Chinese Academy of Agricultural Science: Beijing, 2020. [Google Scholar]
  45. Srivastava R. K.; Pandey P.; Rajpoot R.; Rani A.; Dubey R. S. Cadmium and lead interactive effects on oxidative stress and antioxidative responses in rice seedlings. Protoplasma 2014, 251, 1047–1065. 10.1007/s00709-014-0614-3. [DOI] [PubMed] [Google Scholar]
  46. Ahmad P.; Jaleel C. A.; Salem M. A.; Nabi G.; Sharma S. Roles of enzymatic and nonenzymatic antioxidants in plants during abiotic stress. Crit. Rev. Biotechnol. 2010, 30, 161–175. 10.3109/07388550903524243. [DOI] [PubMed] [Google Scholar]
  47. Li J. Y.; Sui M. H.; Yang J. R.; Xu M. M. Prospects for the development of H2O2 catalytic oxidation sterilization. Water Treat. Technol. 2015, 41, 9–14. [Google Scholar]
  48. Neto A.; Prisco J. T.; Enéas-Filho J.; Medeiros J.; Gomes-Filho E. Hydrogen peroxide pre-treatment induces salt stress acclimation in maize plants. J. Plant Physiol. 2005, 162, 1114–1122. 10.1016/j.jplph.2005.01.007. [DOI] [PubMed] [Google Scholar]
  49. Hameed A.; Malik S. A.; Iqbal N.; Arshad R.; Farooq S. Influence of hydrogen peroxide on initial leaf and coleoptile growth in etiolated wheat (triticum aestivum L.) seedlings. Asian J. Plant Sci. 2003, 2, 1121–1125. 10.3923/ajps.2003.1121.1125. [DOI] [Google Scholar]
  50. Liao W. B.; Zhang M. L.; Wu Y. H.; Xiao Y. L. Effect of nitric oxide and hydrogen peroxide on the rooting of ground cover chrysanthemum cuttings. Hortic. Plant J. 2009, 36, 1643–1650. [Google Scholar]
  51. Oldroyd G. E. D. Speak, friend, and enter: Signalling systems that promote beneficial symbiotic associations in plants. Nat. Rev. Microbiol. 2013, 11, 252–263. 10.1038/nrmicro2990. [DOI] [PubMed] [Google Scholar]
  52. Po-Ju K.; Wan J. Effects of soil microbes on plant competition: a perspective from modern coexistence theory. Ecol. Monogr. 2020, 90, e01391 10.1002/ecm.1391. [DOI] [Google Scholar]
  53. Oberländer J.; Mayer M.; Greeff A.; Keusgen M.; Schöning M. J. Spore-based biosensor to monitor the microbicidal efficacy of gaseous hydrogen peroxide sterilization processes. Biosens. Bioelectron. 2018, 104, 87–94. 10.1016/j.bios.2017.12.045. [DOI] [PubMed] [Google Scholar]
  54. Ozimek E.; Jolanta J. Ś.; Justyna B.; Teresa K. K.; Renata T.; Anna S.; Artur N.; Agnieszka H. Synthesis of Indoleacetic Acid, Gibberellic Acid and ACC-Deaminase by Mortierella Strains Promote Winter Wheat Seedlings Growth under Different Conditions. Int. J. Mol. Sci. 2018, 19, 3218. 10.3390/ijms19103218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Zhang H. S.; Wu X. H.; Li G.; Qin P. Interactions between arbuscular mycorrhizal fungi and phosphate-solubilizing fungus (Mortierella sp.) and their effects on Kostelelzkya virginica growth and enzyme activities of rhizosphere and bulk soils at different salinities. Biol. Fertil. Soils 2011, 47, 543–554. 10.1007/s00374-011-0563-3. [DOI] [Google Scholar]
  56. Wang G. S.; Yin C. M.; Pan F. B.; Wang X. B.; Li X.; Wang Y. F.; Wang J. Z.; Tian C. P.; Chen J.; Mao Z. Q. Analysis of the Fungal Community in Apple Replanted Soil Around Bohai Gulf. Hortic. Plant J. 2018, 4, 175–181. 10.1016/j.hpj.2018.05.003. [DOI] [Google Scholar]
  57. Sheng Y. F.; Wang H. Y.; Wang M.; Li H. H.; Xiang L.; Pan F. B.; Chen X. S.; Shen X.; Yin C. M.; Mao Z. Q. Effects of Soil Texture on the Growth of Young Apple Trees and Soil Microbial Community Structure Under Replanted Conditions. Hortic. Plant J. 2020, 6, 123–131. 10.1016/j.hpj.2020.04.003. [DOI] [Google Scholar]
  58. Pan L. H.; Zhao L.; Jiang W. T.; Wang M.; Chen X. S.; Shen X.; Yin C. M.; Mao Z. Q. Effect of Zinc Oxide Nanoparticles on the Growth of Malus hupehensis Rehd. Seedlings. Front. Environ. Sci. 2022, 10, 94. 10.3389/fenvs.2022.835194. [DOI] [Google Scholar]
  59. Shuang M.; Wang Y. J.; Teng W.; Jin G. H. Isolation and identification of an endophytic bacteria Bacillus sp. K-9 exhibiting biocontrol activity against potato common scab. Arch. Microbiol. 2022, 204, 483. 10.1007/s00203-022-02989-5. [DOI] [PubMed] [Google Scholar]
  60. Wang H. Y.; Zhang R.; Mao Y. F.; Jiang W. T.; Chen X. S.; Shen X.; Yin C. M.; Mao Z. Q. Effects of Trichoderma asperellum 6S-2 on Apple Tree Growth and Replanted Soil Microbial Environment. J. Fungi 2022, 8, 63. 10.3390/jof8010063. [DOI] [PMC free article] [PubMed] [Google Scholar]
  61. Urmas K.; Henrik N. R.; Kessy A.; Tedersoo L.; Taylor A. F. S.; Mohammad B.; Bates S. T.; Bruns T. D.; Bengtsson-Palme J.; Callaghan T. M.; Douglas B.; Drenkhan T.; Eberhardt U.; Dueñas M.; Grebenc T.; Griffith G. W.; Hartmann M.; Kirk P. M.; Kohout P.; Larsson E.; Lindahl B. D.; Lücking R.; Martín M. P.; Matheny P. B.; Nguyen N. H.; Niskanen T.; Oja J.; Peay K. G.; Peintner U.; Peterson M.; Põldmaa K.; Saag L.; Saag I.; Schuyler A.; Scott M.; Senes C.; Smith M. E.; Suija A.; Taylor D. L.; Telleria M. T.; Weiss M.; Larsson K. H. Towards a unified paradigm for sequence-based identification of fungi. Mol. Ecol. 2013, 22, 5271–5277. 10.1111/mec.12481. [DOI] [PubMed] [Google Scholar]
  62. Intaraudom C.; Nitthithanasilp S.; Rachtawee P.; Boonruangprapa T.; Prabpai S.; Kongsaeree P.; Pittayakhajonwut P. Phenalenone derivatives and the unusual tricyclic sesterterpene acid from the marine fungus Lophiostoma bipolare BCC25910. Phytochemistry 2015, 120, 19–27. 10.1016/j.phytochem.2015.11.003. [DOI] [PubMed] [Google Scholar]
  63. Li J.Effects of two fumigants on soil microbial diversity and community structure; Chinese Academy of Agricultural Sciences, 2017. [Google Scholar]
  64. Li L. L.Modulation of Poncirus trifoliata Root System Conformation by Hydrogen Peroxide and Glomus versiforme; Yangtze University, 2013. [Google Scholar]
  65. Chen L. S.; Shi Y. X.; Li L.; Chai A. L.; Guo N.; Fan T. F.; Xie X. W.; Li B. J. Efficacies of Different Soil Fumigants Against Fusarium spp. of Lettuce (Lactuca sativa) Planting Soil and Their Influences on Soil Microbial Community. Journal of Agricultural. Biotechnology 2021, 29, 2365–2374. 10.3969/j.issn.1674-7968.2021.12.010. [DOI] [Google Scholar]
  66. Zimmermann S.; Frey B. Soil respiration and microbial properties in an acid forest soil: effects of wood ash. Soil Biol. Biochem. 2002, 34, 1727–1737. 10.1016/S0038-0717(02)00160-8. [DOI] [Google Scholar]
  67. Zhang X. L.; Ma M.; Wu Z. Z.; Zhang Z. Z.; Lin H. Effects of Helianthus annuus and Glycyrrhiza glabrainter cropping on rhizosphere soil enzyme activities and soil microbes functional diversity. Soils 2016, 48, 1114–1119. [Google Scholar]
  68. Klose S.; Acosta M. V.; Ajwa H. A. Microbial community composition and enzyme activities in a sandy loam soil after fumigation with methyl bromide or alternative biocides. Soil Biol. Biochem. 2006, 38, 1243–1254. 10.1016/j.soilbio.2005.09.025. [DOI] [Google Scholar]
  69. Yang P.; Zhai Y. P.; Zhao X.; Wang S. X.; Liu H. L.; Zhang X. Effect of interactions between arbuscular mycorrhizal fungi and rhizobacteria on the bacterial community structure and predicted function of PICRUSt in rhizosphere soil of alfalfa. Microbiol. Bull. 2020, 47, 3868–3879. [Google Scholar]
  70. Cadet J.; Delatour T.; Douki T.; Gasparutto D.; Pouget J. P.; Ravanat J. L.; Sauvaigo S. Hydroxyl radicals and DNA base damage. Mutat. Res., Fundam. Mol. Mech. Mutagen. 1999, 424, 9–21. 10.1016/S0027-5107(99)00004-4. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

ao2c06665_si_001.pdf (2.8MB, pdf)

Articles from ACS Omega are provided here courtesy of American Chemical Society

RESOURCES