Abstract
Intercropping of ornamental flowering plants like Lycoris radiata Herb. and Cuphea hookeriana Walp. with tea trees can enhance the visibility and esthetic appeal of tea gardens. However, there has been limited research of the impact of intercropping ornamental flowering plants with tea trees on the soil in tea gardens. During this study, our objective was to analyze the effects of intercropping systems on tea garden soil by examining the physicochemical properties of rhizosphere soil samples from tea gardens intercropped with L. radiata and C. hookeriana. We also performed rhizosphere microbial metagenomic sequencing to assess the microbial community structure. The results revealed significant improvements in soil physicochemical indicators, particularly pH. Although intercropping systems had minor impacts on bacterial diversity and abundance, they had more pronounced effects on the community structure of microorganisms at the phylum and genus levels. Furthermore, an analysis of microbial functions using Functional Annotation of Prokaryotic Taxa (FAPROTAX) revealed enrichment of carbon and nitrogen cycling pathways in the tea garden soil. Our findings indicated that intercropping practices have the potential to enhance the visual appeal of tea gardens while improving soil fertility and modulating the microbial community structure. These results contribute to our understanding of intercropping strategies and the implications of intercropping for tea tree growth and ecosystem functioning.
Intercropping in tea gardens has a long history. Ancient mountain dwellers developed advanced intercropping soil management techniques as a result of their harmonious coexistence with nature. These practices have surpassed traditional slash-and-burn agriculture. Proper intercropping methods have the potential to promote tea tree growth, increase yield, and improve overall quality (Brooker et al. 2015; Le et al. 2021; Xiong et al. 2024), thus allowing for the economic utilization of land and space. However, it is important to note that improper intercropping can lead to competition for nutrients, water, and light between tea trees and intercrops (Trenbath 1976; Wu et al. 2020). This competition can negatively affect tea tree growth by increasing their susceptibility to pests, such as mites, aphids, and tea mosquito bugs, and diseases, such as tea gray blight and tea anthracnose, potentially resulting in a decline or even the death of tea trees. Various intercropping patterns have been implemented for different purposes (Boudreau 2013). For example, tea–bean intercropping enhances soil fertility primarily through the nitrogen fixation provided by the rhizobium of legume crops. This nitrogen fixation increases the number of soil microorganisms and soil enzyme activity, thus accelerating soil nutrient cycling (Sun et al. 2022). Tea–rubber intercropping has shown promising results by generating a comprehensive economic yield that surpasses planting a single crop on the same land area (Le et al. 2021). Additionally, tea–chestnut intercropping influences the microclimate, reduces soil bulk density and soil erosion, and improves soil nutrients and water availability. This includes an increase in soil carbon and nutrient availability (Duan et al. 2021). Intercropping tea with aromatic plants such as Catsia tora Linn., Medicagosativa sativa L., Leonurus heterophyllus Sweet., and Mentha haplocalyx Briq. promotes a significant increase in natural enemies (such as Aphelinus sp.) of tea pests. This alternative strategy for sustainable pest management in tea cultivation offers promising results (Iqbal et al. 2006; Zhang et al. 2017). Furthermore, tea–corn intercropping has improved tea quality and flavor through ecological shading, along with the effective control of major pests such as mites and aphids (Wu et al. 2021; Zou et al. 2022). Therefore, intercropping functional plants in tea plantations serves ecological functions such as enhancing soil health, improving the microenvironment of tea plantations, increasing pest control efficiency, enhancing the quality and yield of tea products, and boosting the ecological service functions for tea tourism.
Wuyishan is situated on the border of Jiangxi and northwestern Fujian provinces and is renowned for its typical Danxia landforms. Additionally, it is a well-known tourist destination and summer retreat in China. Wuyishan has a significant position as one of China’s renowned tea-producing regions, with Wuyi Rock Tea being its specialty (Huo et al. 2021; Xu et al. 2021). As of 2023, the tea plantation area in Wuyishan had expanded to 9867 ha (Li et al. 2023; Zhao et al. 2023). However, the majority of tea gardens primarily cultivate tea trees, resulting in the mountaintops appearing barren with limited vegetation. Intercropping ornamental plants within tea gardens emerges as a viable solution to enhance the landscape of tea plantations. Strategically combining tea trees with flowers allows the tea garden to showcase blooming flowers year-round, thus creating unique views during every season. This approach significantly enhances the visual appeal of the tea garden.
The selection of functional plants for intercropping in tea plantations should meet requirements such as simple cultivation management, broad adaptability, the absence of common pests and diseases with tea trees, noninvasive weeds, long-lasting and universal impacts on habitats, and appropriate timing alignment with the growth and harvest periods of tea trees (Chen et al. 2019; Wu et al. 2021). Furthermore, soil quality greatly affects plant growth, with microorganisms serving as key indicators (Ding et al. 2018). In tea gardens, these microorganisms play a vital role in regulating plant growth and development. They facilitate nitrogen fixation, withstand acidity and aluminum, combat pathogens, and promote nutrient activation and absorption (Dubey et al. 2019; Ramesh et al. 2019; Yang et al. 2020, 2021). Some microorganisms directly influence tea plant growth through the synthesis of growth regulators like indole-3-acetic acid, abscisic acid, and ethylene inhibitors (Xiang et al. 2020). Studies have demonstrated that plant age, season, soil characteristics, and intercropping methods impact the species and quantity of rhizosphere microorganisms (Deng et al. 2019; Guo et al. 2018; Liu et al. 2010; Maron et al. 2018; Pajares and Ramos 2019). Consequently, it is essential to consider soil quality and the presence of beneficial microorganisms when selecting functional plants for intercropping in tea plantations because these factors greatly influence tea plant quality and yield.
Although the primary goal of incorporating flowering landscape plants in intercropping is to enhance the visual esthetics of tea gardens in Wuyishan, the impact of this practice on the soil health of tea gardens remains uncertain. Therefore, the aim of this research is to analyze the impact of intercropping on the physicochemical properties of tea garden soil and the structure of the microbial community. This study aimed to evaluate the effects of intercropping with L. radiata and C. hookeriana on soil fertility, microbial diversity, and microbial functions in tea gardens to provide recommendations for establishing eco-friendly tea gardens that enhance the tea garden landscape while maintaining tea production.
Materials and Methods
Experimental site and soil sample collection.
The current research was conducted in a tea garden located in Wuyishan City, Fujian Province, China (lat. 27°7′N, long. 118°1′E), which is renowned for its production of rock tea. This region experienced a typical northern subtropical monsoon climate, with an average annual temperature of 19 °C and an average annual rainfall exceeding 2 km. The tea garden consisted of both single cultivation and intercropping systems and has been cultivated for 1.5 years using a unified agricultural management strategy. The Spring tea picking period typically spanned from mid-April to mid-May. Following each tea harvesting season, fertilization was applied in the tea garden, with additional fertilization performed during late October.
Soil samples were collected from three different cultivation systems in the tea garden on 26 Sep 2022, during the Fall season. The temperatures during collection ranged from 20 to 25 °C, and the weather was sunny. Soil samples were collected from the rhizosphere soil of tea trees in the single-cultivation system, as well as from the C. hookeriana–tea intercropping system and L. radiata–tea intercropping system. Each sample was collected in triplicate and labeled as CK (tea monoculture), Y (C. hookeriana–tea intercropping), and W (L. radiata–tea intercropping), with careful consideration given to selecting samples at similar altitudes, slope positions, and aspects. The surface soil was removed, and soil samples were collected from a depth of 5 to 20 cm along the tea tree roots. A total of nine soil samples were collected and promptly transported to the laboratory in a cooler box. Each soil sample was divided into two parts. One part was air-dried, ground, and sieved through a 100-mesh screen for chemical analysis. The other part was temporarily stored at −80 °C in a freezer for DNA extraction. Subsequently, the samples were sent to Shanghai Personalbio Biotechnology Co., Ltd., for DNA extraction, amplification, and high-throughput sequencing to determine the diversity and community composition of soil bacteria.
Testing of soil chemical properties.
Soil pH was determined using a pH meter, whereas soil organic matter was determined using the potassium dichromate method. Available nitrogen levels were measured using the alkaline diffusion method, and available phosphorus was determined using the 0.5 mol/L NaHCO3 extraction and molybdenum antimony anti-colorimetric method. Additionally, available potassium was determined using the NH4OAc extraction and flame photometry method (Sun et al. 2022; Zhang et al. 2017).
DNA extraction, polymerase chain reaction amplification, and sequencing.
Genomic DNA was extracted from 0.5-g soil samples using the Soil DNA Isolation Kit (Omega Bio-tek, Norcross, GA, USA) according to the manufacturer’s instructions. To assess the quality and concentration of the extracted DNA, three independent DNA extracts from each sample were pooled, and their absorbance values were measured at 260 nm and 280 nm using a fluorescence spectrophotometer [Quantifluor-ST fluorometer E6090 (Promega, Madison, WI, USA); Quant-iT PicoGreen dsDNA Assay Kit P7589 (Invitrogen, Waltham, MA, USA)]. Additionally, the DNA quality was evaluated through 1% agarose gel electrophoresis. The DNA solution concentration was adjusted accordingly and stored at 4 °C, whereas the storage solution was kept at −20 °C.
The polymerase chain reaction amplification program consisted of an initial denaturation step at 95 °C for 2 min, followed by 20 cycles of denaturation at 98 °C for 10 s, annealing at 62 °C for 30 s, extension at 68 °C for 30 s, and a final extension at 68 °C for 10 min. The amplification products of the bacterial 16S rRNA gene and fungal ITS gene were visualized on a 2% agarose gel and purified using the AxyPrep DNA Gel Extraction Kit (Axygen Biosciences, Union City, CA, USA). To determine the DNA concentration, quantification was performed using the QuantiFluor-ST (Promega) assay. The purified amplicons were pooled equimolarly and subjected to paired-end sequencing (2 × 250) on the Illumina platform following standard protocols. The constructed sequencing library was performed using Genedenovo on the Illumina HiSeq2500 PE250 platform (Genedenovo Co., Ltd., Guangzhou, China).
Data analysis.
A variance analysis (least significant difference test, P < 0.05) for multiple indicators such as soil chemical properties, different bacterial genera, and microbial functional diversity was conducted using SPSS 26.0 software. The taxonomic composition was analyzed using QIIME2 (2019.4) with a rarefaction curve generated. Boxplots for the Chao1 index, observed species index, Simpson index, and Shannon index were created using GraphPad Prism 8. The relative abundance of taxa at the phylum and genus levels was plotted using the ggplot2 package of R 4.3.1 software. Operational taxonomic unit Venn diagrams were generated using the VennDiagram package of R 4.3.1. A principal component clustering analysis was performed using the Vegan package of R 4.3.1 to understand the clustering pattern of the samples. Correlation heat maps were plotted using TBtools to visualize the correlations between different variables or taxa.
Results
Changes in soil physicochemical properties under intercropping treatments
A one-way analysis of variance revealed the significant effects of the planting modes on soil properties. The analysis results are shown in Table 1. Compared with the CK, both intercropping methods significantly increased pH (P = 0.001), available phosphorus (P < 0.001), and available potassium (P < 0.05). Additionally, W significantly improved soil organic matter (P < 0.05). Furthermore, there was a highly significant difference among the three groups in terms of available potassium. However, there were no significant differences among the different intercropping treatments in terms of pH, organic matter, and available potassium. Overall, both intercropping treatments had a positive effect on improving the fertility of tea garden soil, with W intercropping showing a more significant effect.
Effects of different planting patterns on soil chemical properties.
Changes in soil microbial community diversity
Alpha diversity analysis.
Soil bacterial communities were evaluated using high-throughput 16S rDNA amplicon sequencing. The sequencing depth per sample ranged from 86,865 to 187,660 sequences, with an average of 127,500 sequences. To ensure comprehensive coverage of the bacterial population, the dilution curve was examined (Fig. 1E), thus affirming the sufficiency of the sequencing depth. To evaluate the richness of the bacterial communities, the Chao1 index (Fig. 1A) and the observed species index (Fig. 1B) were used. The richness was characterized by these indices, whereas the diversity was represented by the Shannon index (Fig. 1C) and Simpson index (Fig. 1D). Interestingly, no increases in the richness and diversity of soil microbiota were observed when different plants were intercropped with tea. No significant differences in the richness of soil microbiota among different intercropping patterns were found, and the diversity of soil microbiota was not enhanced. Moreover, a significant decrease in the diversity of soil microbiota was detected after intercropping with L. radiata (P < 0.05). The coverage of soil bacteria in terms of diversity was consistently more than 0.99, suggesting that the true composition of bacterial communities in the soil samples was adequately reflected by the sequencing depth.
CHAO1 index (A). Observed specifications index (B). Shannon index (C). Simpson index (D). Rarefied fraction curves (E). Principal coordinate analysis (PCoA) of the soil bacterial community (F). Venn diagrams of core operational taxonomic units (OTUs) (G) of soil bacteria in different planting modes.
Citation: HortScience 59, 9; 10.21273/HORTSCI17790-24
Beta diversity analysis.
The similarities and dissimilarities between treatments were assessed using a beta diversity analysis that specifically used the Jaccard distance algorithm. A principal coordinate analysis was conducted to visually represent the results (Fig. 1F), thus revealing the clustering of bacterial members according to sampling areas. Within each treatment group, the sample points were relatively concentrated and distributed in different quadrants, indicating that the composition of soil bacterial communities in tea plantations can be altered by intercropping treatments, with the structure varying depending on the different combinations of intercropping treatments.
Composition of soil bacterial communities under different treatments.
The Venn diagram (Fig. 1G) revealed that a total of 19,056 soil bacterial operational taxonomic units (OTUs) were identified. The control group (CK) had 8386 OTUs, whereas Y and W had 7581 and 7927 OTUs, respectively. However, intercropping treatments did not significantly impact the number of soil bacterial OTUs. Only 1037 were shared among all treatment groups, accounting for only 5.4% of the total. Additionally, there were 4779 unique OTUs in group Y, 5134 in group CK, and 5342 in group W. Significant differences were observed among group Y, group W, and group CK, suggesting that intercropping treatments have the potential to modify the structure and diversity of bacterial communities.
Changes in the composition and structure of soil microbial communities.
Figure 2A illustrates the relative abundance of soil bacteria at the phylum level, whereas Figure 2B represents the relative abundance at the genus level. Upon analyzing the phylum-level data, we found that the top 10 most abundant phyla in all soil samples were Proteobacteria, Acidobacteria, Chloroflexi, Actinobacteria, Planctomycetes, WPS-2, Verrucomicrobia, Bacteroidetes, Gemmatimonadetes, and GAL15. Regarding the intercropping of C. hookeriana and tea plants, a significant decrease in the relative abundance of Acidobacteria was observed, whereas the relative abundance of Actinobacteria significantly increased. Similarly, regarding the intercropping of L. radiata and tea plants, there was a notable decrease in the relative abundance of Proteobacteria, Bacteroidetes, and Gemmatimonadetes, whereas the relative abundance of Chloroflexi and WPS-2 significantly increased. Comparing the two intercropping modes, we observed significant differences in the relative abundance of bacteria at the phylum level, namely, Proteobacteria, Chloroflexi, Acidobacteria, Actinobacteria, Planctomycetes, Bacteroidetes, and Gemmatimonadetes. Notably, the intercropping of L. radiata and tea plants exerted a greater influence on the phylum-level composition of soil bacteria in the tea garden than that of the intercropping mode of C. hookeriana and tea plants.
Relative abundance of soil bacteria at the phylum level in different planting modes (A). Relative abundance of soil bacteria at the genus level in different planting modes (B). Significance analysis of the top 30 different genera at the genus level (C). Correlation analysis heat map of different genera present in the C. hookeriana–tea intercropping system and soil chemical properties (D). Correlation analysis heat map of different genera appearing in the L. radiata–tea intercropping system and soil chemical properties (E).
Citation: HortScience 59, 9; 10.21273/HORTSCI17790-24
Significant differences in the top 30 genera (Fig. 2C) were identified by a genus-level analysis. In comparison with the control group (CK), a significant increase in the relative abundance of the genera Acidothermus, HSB_OF53-F07, and FCPS473 was observed in both intercropping modes. Conversely, the genera Subgroup_6 and Bryobacter experienced a notable decrease in the relative abundance. Moreover, a significant decrease in the relative abundance of the genera Bradyrhizobium, Rhodanobacter, Gemmatimonas, Roseiarcus, and Reyranella was observed within the L. radiata group. Conversely, a significant increase in the relative abundance of the genera AD3, WPS-2, Methylobacterium, and IMCC26256 was noted. Only the C. hookeriana group displayed a significant increase in the relative abundance of the genera Bradyrhizobium, Conexibacter, and Mycobacterium. These findings demonstrated that the L. radiata group had a greater impact on soil microorganisms at the genus level.
At the genus level, the effects of the two intercropping patterns on tea garden soil bacteria align with general trends. However, a significant increase in the relative abundance of the C. hookeriana group was observed at the Bradyrhizobium level, whereas the L. radiata group showed a significant decrease in the relative abundance. Through a correlation analysis (Fig. 2D, 2E) of different genera appearing in the two intercropping patterns and soil chemical properties, it was found that most of the different genera in the C. hookeriana group were significantly positively correlated with the following four soil chemical properties: pH, alkali hydrolyzed nitrogen, available potassium, and available phosphorus. However, the different genera in the L. radiata group showed significant correlations with the following four soil chemical properties: pH, organic matter, available potassium, and available phosphorus. However, a positive correlation with soil chemical properties was observed in approximately half of the different genera, whereas the other half showed a negative correlation. Among these factors, pH, available phosphorus, and available potassium were identified as the main influencers of the different genera; of the top 30 genera with the highest abundance, 18 were significantly affected by these factors.
Changes in the functional characteristics of soil microorganisms.
To investigate the effects of different intercropping treatments on the various aspects of bacterial communities, a Functional Annotation of Prokaryotic Taxa (FAPROTAX) analysis was conducted, which assigned 55 microbial functional categories of the 80 functions available in the FAPROTAX database. Among them, 10 microbial functions (Fig. 3A) were clustered in the carbon cycle, whereas 14 microbial functions (Fig. 3B) were clustered during the nitrogen cycle. Significant differences (P < 0.05) in seven microbial functions related to the carbon cycle (methanotrophy, methylotrophy, cellulolysis, fermentation, aromatic hydrocarbon degradation, hydrocarbon degradation, chemolithotrophy) and three microbial functions related to the nitrogen cycle (aerobic nitrite oxidation, nitrate reduction, ureolysis) were observed in the L. radiata–tea intercropping system compared with the control group (Fig. 3E). In the C. hookeriana–tea intercropping system, significant differences (P < 0.05) were observed in two microbial functions related to the carbon cycle (cellulolysis, aromatic compound degradation) and two microbial functions related to the nitrogen cycle (nitrate reduction, ureolysis) (Fig. 3F).
Relative abundance heat map of microorganisms involved in soil carbon cycling (A) and soil nitrogen cycling (B) under different planting modes. Bacterial abundance involved in carbon cycling (C). Bacterial abundance involved in nitrogen cycling (D). Analysis of significant differences in soil carbon cycling microbial functions (E). Analysis of significant differences in soil nitrogen cycling microbial functions (F).
Citation: HortScience 59, 9; 10.21273/HORTSCI17790-24
It can be concluded that various C and N cycling functions were impacted by intercropping. The L. radiata group, compared with the C. hookeriana group, has a greater influence on soil carbon and nitrogen cycling functions, particularly in the carbon cycle. The combination of the previous microbial species composition analyses of the two intercropping systems revealed that the L. radiata group had a stronger impact on the microbial community structure, thereby resulting in more alterations in the microbial functions. Additionally, a statistical analysis of the abundance of microorganisms involved in carbon and nitrogen cycling was performed (Fig. 3C, 3D). The box plots showed that the C. hookeriana group exhibited the most significant increase in the abundance of microorganisms involved in the carbon cycling function, whereas the L. radiata group exhibited the larges increase in microorganisms involved in the nitrogen cycling function.
Correlation analysis of soil carbon and nitrogen cycling functions and genus-level bacterial communities.
The correlation analysis of significantly different genera within the top 30 and carbon and nitrogen cycling functions (Fig. 4A, 4B) revealed that these genera displayed a stronger correlation with carbon cycling. The heat map (Fig. 4A) illustrated the correlation between different genera and carbon cycling functions. Notably, Bradyrhizobium, Rhodanobacter, Gemmatimonas, Reyranella, Burkholderia-Caballeronia-Paraburkholderia, and Mycobacterium exhibited significant positive correlations with fermentation and aromatic compound degradation functions while displaying significant negative correlations with methanotrophy and methylotrophy functions. Interestingly, the genera AD3 and Methylobacterium demonstrated contrasting results compared with the aforementioned genera. They exhibited significant positive correlations with methanotrophy and methylotrophy functions while displaying significant negative correlations with fermentation and aromatic compound degradation functions. The heat map (Fig. 4B) exhibited the correlation between different genera and nitrogen cycling functions. These different genera displayed a strong correlation with aerobic ammonia oxidation, aerobic nitrite oxidation, nitrogen fixation, and ureolysis. Furthermore, various genera demonstrated opposite correlations between aerobic ammonia oxidation and aerobic nitrite oxidation. Among these, three genera (AD3, Acidothermus, Methylobacteriaceae) showed a notable positive correlation with airborne ammonia oxidation, but a significant negative correlation with airborne nitrate oxidation. These genera also exhibited consistent correlations with aerobic nitrite oxidation and ureolysis.
Correlation analysis of bacterial genera with significant differences in the top 30 and soil carbon cycling (A) and nitrogen cycling (B).
Citation: HortScience 59, 9; 10.21273/HORTSCI17790-24
Discussion
Effects of intercropping modes on soil physicochemical properties and bacterial communities.
Compared with monocropping systems, intercropping systems have been shown to exhibit superior chemical properties in the rhizosphere soil, as supported by previous research (Li et al. 2018; Yang et al. 2021). One significant effect of intercropping is the improvement of soil pH, which is an essential factor in the soil environment that greatly influences the composition and structure of soil bacterial communities. Despite using the same management methods in tea gardens, it is intercropping that leads to changes in the chemical properties of the soil within a single season. To investigate these effects, we used 16S rRNA sequencing to analyze the rhizosphere soil microbiota within monocropping and intercropping systems. Our finding revealed that although the intercropping system did not result in increased bacterial community diversity, there were significant transformations in species composition at the phylum and genus levels. Notably, the L. radiata group exhibited the most pronounced alterations. By conducting a correlation analysis of different bacterial genera observed in the two intercropping systems and soil chemical properties, we were able to identify significant correlations between these different bacterial genera and various properties. Among them, pH, available phosphorus, and available potassium emerged as the primary factors that influence the variation of these bacterial genera.
Effects of intercropping patterns on soil carbon cycling.
Carbon is a fundamental component of all organic compounds, and the cycling of soil carbon plays a crucial role in the cycling of nitrogen, sulfur, and phosphorus. Only under favorable conditions for the accumulation of soil organic carbon will there be increases in organic nitrogen, phosphorus, and sulfur contents (Shu et al. 2021; Tang et al. 2023). In our study, through the functional prediction of microorganisms, we observed a significant enhancement in the degradation of organic matter in both intercropping systems. Bacterial genera such as Candidatus, Solibacter, Conexiactor, and Acidothermus play key roles in the decomposition of organic matter and the utilization of carbon sources (Chen et al. 2023; Luo et al. 2023; Siyao et al. 2017; Zhang et al. 2016). Notably, there was significant improvement in cellulolysis. Acidothermus, the second most abundant genus at the genus level, showed a substantial increase in relative abundance in both intercropping systems. Acidothermus is known for producing highly thermostable cellulases (Jiang et al. 2017; Tucker et al. 1989), making it highly efficient in the degradation of lignocellulosic biomass, and its increased abundance under intercropping conditions enhances cellulolysis.
Furthermore, we observed a significant enhancement in hydrocarbon degradation within the L. radiata intercropping group and aromatic compound degradation in the C. hookeriana intercropping group. Degradation of aromatic compounds, with their benzene ring structures, is generally more complex and challenging than degradation of simple hydrocarbons. The genus Sphingomonas exhibited exceptional degradation ability by breaking down a wide range of organic compounds, including polycyclic aromatic hydrocarbons, pesticides, and organic waste (Fagervold et al. 2021; Zhou et al. 2022). For example, Sphingomonas aeruginosa possesses a wide array of degradation-related genes, such as those involved in phenanthrene degradation. This gene pool triggers robust upregulation of extradiol cleavage pathway enzymes in sphingomonads, showing genetic and enzymatic similarity to Sphingobium yanoikuyae B1. The enzymes encoded by genes such as bphA2cA1c, bphA1[a-e]A2[a-e], and bphC include ring-hydroxylating dioxygenase, putative biphenyl-2,3-diol 1,2-dioxygenase, and catechol 2,3-dioxygenase, respectively. Sphingomonads exhibited well-adapted capabilities because of their utilization of meta-cleavage and ortho-cleavage pathways. Additionally, certain species within this genus have shown the potential to enhance plant growth in agricultural soils under various stresses such as drought, salinity, and heavy metals. This effect is attributed to their ability to produce plant growth hormones such as gibberellins and indole-3-acetic acid (Asaf et al. 2020).
Burkholderia species have the unique ability to break down organic compounds, contributing to soil improvement and providing essential nutrients for plants (Lee et al. 2016; Morya et al. 2020). Furthermore, these bacteria are known as rhizosphere-promoting bacteria that produce various hormones and metabolites such as indole-3-acetic acid, gluconic acid, and glucosamine. These compounds stimulate root development and enhance plant biomass and yield. Additionally, Burkholderia synthesizes antibacterial substances, such as antibiotics, bacteriocins, and phenolic compounds. These substances can effectively inhibit or eliminate certain plant pathogens like Ralstonia solanacearum, Glomerella cingulata, and Didymella bellidis. Consequently, they enhance the resistance of plants to diseases (Elshafie and Camele 2021; Heo et al. 2022). Burkholderia AY001 has a good control effect on tomato wilt and bacterial spot disease. Burkholderia gladioli pv. agaricicola strain ICMP 12322 was able to enhance disease protection and improve the consistency of biological control against tomato wilt disease caused by Verticillium dahliae. Nevertheless, it is worth noting that some research has suggested that specific strains within the Burkholderia genus have the potential to function as novel pathogens (Ham et al. 2011).
A correlation analysis identified four genera, Bradyrhizobium, Rhodanobacter, Gemmatimonas, and Reyranella, that exhibit a significant positive correlation with the degradation of aromatic compound. Specifically, Rhodanobacter possesses antagonistic properties against certain fungal pathogens. Furthermore, some strains within the Rhodanobacter genus have the ability to denitrify and break down aromatic compounds like benzyl ether (Zhong et al. 2022). Bradyrhizobium and Gemmatimonas are well-known as rhizosphere-promoting bacteria, and Gemmatimonas also demonstrates the ability to degrade complex compounds. However, limited research of the characteristics and contributions of Reyranella has been performed. Although the correlation analysis suggested a strong positive relationship between these genera and aromatic compound degradation, further studies are necessary to ascertain whether this enhancement is a result of the inherent capacity to degrade aromatic compounds or other influential factors.
Additionally, our research has demonstrated that the L. radiata intercropping group exhibited a substantial enhancement in the functions of methanotrophy and methylotrophy. Methanotrophy, a form of methylotrophy, involves the participation of methylotrophic bacteria. These bacteria use volatile organic compounds such as methane, dichloromethane, formaldehyde, methanol, and formic acid as carbon sources and energy for their growth. Therefore, they also contribute to improving air quality (Kumar et al. 2016). Methylotrophic bacteria can generally be classified into two categories: obligate methylotrophic bacteria and facultative methylotrophic bacteria. Obligate methylotrophic bacteria, known as methanotrophs, can use methane for their growth and belong to genera such as Methylobacterium and Methylovirgula. Facultative methylotrophic bacteria, however, cannot use methane for growth and encompass genera such as Hyphomicrobium (Kang et al. 2022; Kumar et al. 2016; Nysanth et al. 2023).
Among the top 30 abundant genera identified at the genus level, we observed a higher relative abundance of Methylobacterium in the L. radiata intercropping group compared with the control group. Moreover, during the correlation analysis of microbial functions, Methylobacterium showed a significant positive correlation with both methanotrophy and methylotrophy. It is worth noting that methylotrophic bacteria also play important roles in phosphorus, nitrogen, and carbon cycling, thereby contributing to the mitigation of global warming. Previous studies have emphasized the significance of methylotrophic bacteria in the rhizosphere and their potential applications in various commercial settings. Notably, methylotrophic bacteria have gained attention as bioinoculants in agriculture. They can be used as seed coatings or seed inoculants to enhance seed germination. Additionally, the application of methylotrophic bacteria to potted plants promotes growth and increases yield (Gamit and Amaresan 2023; Gamit et al. 2023; Nysanth et al. 2023). Recent research has even unveiled a fourth branch of Rhizobiales that exhibits both methylotrophy and nitrogen fixation capabilities (Kumar et al. 2019; Nysanth et al. 2023). In summary, beneficial methylotrophic bacteria have the potential to enhance plant growth through nitrogen fixation, phosphate solubilization, the production of plant hormones, and the facilitation of balanced carbon cycling, thereby reducing land pollution caused by the application of fertilizers and pesticides and playing a vital role in the advancement of sustainable agriculture.
Effects of intercropping modes on soil nitrogen cycling.
Nitrogen primarily exists in the form of N2, which is not directly available for utilization. Nitrogen fixation is necessary to convert it to reactive nitrogen. Whether through biological or nonbiological nitrogen fixation, microbial processes are essential for facilitating nitrogen transformation (Nysanth et al. 2023). Therefore, microbes act as the driving force behind the conversion of nitrogen in different forms, thus playing a crucial role in maintaining the stability of ecosystem structure and function (Liu et al. 2020).
In the soil nitrogen cycle under two intercropping modes, nitrogen fixation and ureolysis play significant roles in enhancing nitrogen cycling. Nitrogen fixation converts atmospheric nitrogen into ammonia or ammonium ions, which can be readily absorbed by plants. Rhizobium bacteria contribute to this process by converting atmospheric nitrogen to plant-accessible ammonium or nitrite (Lindström and Mousavi 2020). This provides plants with a necessary nitrogen source, thus promoting their growth and development. Urea decomposition by soil microorganisms leads to the formation of plant-accessible ammonium and nitrate, which provide abundant substrates for soil nitrogen cycling and facilitates the overall nitrogen cycling process in the soil. Aerobic ammonia oxidation is a biochemical process whereby bacteria or other microorganisms convert ammonia into nitrite in the presence of oxygen. Ammonia oxidation serves as the initial step in the nitrogen cycle by providing substrates for subsequent nitrification and denitrification processes by converting ammonia into nitrite (Lehtovirta Morley 2018; Stein and Klotz 2016). Nitrifying bacteria, such as Rhodanobacter, can participate in soil ammonia oxidation. Under anaerobic conditions, Rhodanobacter exhibits denitrification capabilities by using nitrate, nitrite, or nitrous oxide as electron acceptors (Xu et al. 2020). Sphingomonas, another bacterial genus, also contributes to nitrogen nitrification and denitrification processes in the soil by providing the necessary nitrogen source for plants (Xie and Yokota 2006). Both nitrate and nitrite are crucial nitrogen sources that play significant roles in plant growth and development. Plants primarily absorb nitrate as their main nitrogen source, which is then converted into organic nitrogen compounds, such as amino acids, for participation in protein synthesis and other physiological processes. Nitrite, although relatively toxic, can still promote plant growth and increase yield at moderate levels (Crawford 1995; Iqbal et al. 2020). Functional predictions suggest that both intercropping modes facilitate the conversion between nitrite and nitrate and have more significant roles in the oxidation of nitrite to nitrate. This process enables effective absorption and utilization of nitrogen compounds in the soil by plants, thereby maintaining soil nitrogen cycling.
Conclusion
In summary, both intercropping modes improved several chemical properties (such as available phosphorus and available potassium) of the soil, altered the structure of the soil bacterial community, and enhanced the utilization of carbon and nitrogen sources by rhizospheric microorganisms. The rhizospheric microorganisms in the L. radiata intercropping group showed a significantly stronger carbon metabolism ability than that in the C. hookeriana intercropping group; however, the C. hookeriana intercropping group exhibited significantly stronger nitrogen metabolism ability compared with that of the L. radiata intercropping group. The soil bacterial community structure was significantly altered in both intercropping modes, with most the significantly different genera belonging to beneficial bacteria that facilitate plant growth, enhance soil carbon and nitrogen cycling, and possess antagonistic properties against soil-borne diseases and pests. Therefore, these two intercropping modes not only enhance the visual appeal of the tea plantation but also improve soil fertility, regulate the structure of the soil microbial community, and promote better tea plant growth. Each intercropping mode has its distinct advantages, for instance, the L. radiata intercropping group excels in degrading hydrocarbon compounds, whereas the C. hookeriana intercropping group excels in degrading aromatic compounds. Further studies and monitoring are essential to a better understanding of the specific mechanisms and long-term effects of intercropping ornamental plants in tea gardens. This knowledge can guide future decisions and practices that could maximize the benefits of intercropping for tea garden ecosystems.
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