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IntroductionIn mountainous tea plantations, which are the primary mode of tea cultivation in China, issues such as soil erosion and declining soil fertility are particularly severe. Although green manure cover is an effective agricultural measure for restoring soil fertility, its application in mountainous tea plantations has been relatively understudied.MethodsThis study investigated the effects of continuous green manure cover using the slope-protecting plant Ophiopogon japonicus on tea plant growth and soil microbial community structure. We implemented three treatments: 1 year of green manure coverage, 2 years of coverage, and a control, to study their effects on tea plant growth, soil physicochemical properties, and soil bacterial and fungal communities.ResultsResults demonstrate that green manure coverage significantly promote the growth of tea plants, enhanced organic matter and pH levels in soil, and various enzyme activities, including peroxidases and cellulases. Further functional prediction results indicate that green manure coverage markedly promoted several carbon cycling functions in soil microbes, including xylanolysis, cellulolysis, degradation of aromatic compounds, and saprotrophic processes. LEfSe analysis indicated that under green manure cover, the soil tends to enrich more beneficial microbial communities with degradation functions, such as Sphingomonas, Sinomonas, and Haliangium (bacteria), and Penicillium, Apiotrichum, and Talaromyce (fungi). In addition. Random forest and structural equation models indicated that carbon cycling, as a significant differentiating factor, has a significant promoting effect on tea plant growth.DiscussionIn the management practices of mountainous tea plantations, further utilizing slope-protecting plants as green manure can significantly influence the soil microbial community structure and function, enriching microbes involved in the degradation of organic matter and aromatic compounds, thereby positively impacting tea tree growth and soil nutrient levels.

As sustainable forest management gains increasing attention, comprehending the impact of stand density on soil properties and microbial communities is crucial for optimizing forest ecosystem functions. This study employed high-throughput sequencing in conjunction with soil physicochemical analysis to assess the effects of stand density on soil physicochemical properties and microbial community characteristics in Chinese fir plantations, aiming to elucidate the influence of density regulation on ecosystem services. Our results suggested that changes in soil physicochemical properties and microenvironmental conditions were key drivers of soil microbial diversity. Total carbon (TC), soluble nitrogen (SN), and light fraction organic matter decreased with increasing stand density, while total potassium (TK) and available phosphorus (AP) concentrations increased. The plot with a density of 900 trees ha−1 exhibited the highest bacterial diversity, in contrast to the plot with 1500 trees ha−1, which showed the lowest. The dominant microbial taxa were similar across different stand retention densities, with Acidobacteria, Proteobacteria, and Chloroflexi being the predominant bacterial phyla and Ascomycota and Basidiomycota being the main fungal groups. Significant positive correlations were observed between soil microbial community structures and environmental factors, particularly with respect to soil phosphorus and nitrogen content. The present study demonstrated that reduced stand densities modulated soil nutrient content and enhanced bacterial diversity, thereby contributing to a more complex and stable soil ecosystem structure. These insights provide a scientific foundation for optimizing the management of Chinese fir plantations, thereby supporting the sustainable development of forest ecosystems.

BackgroundThe consecutive monoculture of Rehmannia glutinosa leads to a serious decrease in its production and quality. Previous studies have demonstrated that intercropping altered species diversity and rhizosphere microbial diversity. However, it remained unknown whether the impaired growth of monocultured plants could be restored by enhanced belowground interspecific interactions.MethodIn the present research, a continuous cropping facilitator Achyranthes bidentata was intercropped with R. glutinosa under pot conditions, and three different types of root barrier treatments were set, including that complete belowground interaction (N), partial belowground interaction (S), and no belowground interspecies interaction (M), with the aims to investigate belowground interaction and the underlying mechanism of alleviated replanting disease of R. glutinosa by intercropping with A. bidentata.ResultsThe results showed that the land equivalent ratio (LER) of the two years was 1.17, and the system productivity index (SPI) increased by 16.92 % under S treatment, whereas no significant difference was found in N and M regimes. In the rhizosphere soil, intercropping systems had significantly increased the contents of sugars and malic acid in the soil of R. glutinosa, together with the content of organic matter and the invertase and urease activities. Meanwhile, intercropping increased the community diversity of fungi and bacteria, and the relative abundance of potential beneficial bacteria, such as Bacillus, Nitrospira, and Sphingomonas, despite the pathogenic Fusarium oxysporum was still the dominant genus in the rhizospheric soil of R. glutinosa under various treatments. The results of antagonism experiments and exogenous addition of specific bacteria showed that Bacillus spp. isolated from rhizosphere soil had a significant antagonistic effect on the pathogen of R. glutinosa.ConlusionTaken together, our study indicated that the R. glutinosa//A. bidentata intercropping systems alleviate the consecutive monoculture problem of R. glutinosa by recruiting beneficial bacteria. The studies we have conducted have a positive effect on sustainable agricultural development.

Successive planting and monoculture, as common forest management methods, are widely used globally, especially in Chinese fir plantations in the subtropical areas of southern China. Although soil fertility depletion and productivity decline caused by successive planting have been widely reported, the underlying mechanism is still ambiguous. In this study, the composition and diversity of soil microorganisms (rhizosphere and bulk soils) in Chinese fir seedlings exposed to successive planting soils (first-generation Chinese fir seedings, FCP. second-generation Chinese fir seedings, SCP. third-generation Chinese fir seedings, TCP) and broadleaf tree species soil (Phoebe zhennan S. Lee et F. N. Wei, CK) were examined with high-throughput sequencing technology. Our findings revealed that the diversity and richness of bacterial and fungal communities were remarkably reduced in TCP than FCP and SCP, and were remarkably different between FCP and SCP. At the phylum level, the fungi with greatest relative abundance were Basidiomycota (5.74–32.88%) and Ascomycota (57.63–87.38%), while the bacteria with the greatest relative abundance were Acidobacteria (23.16–31.17%) and Proteobacteria (24.71–29.32%) for all treatments in both soil types. Additionally, the relative abundance of some pathogens (Penicillium and Burkholderia) was significantly higher in TCP than in FCP and SCP, suggesting that the presence of pathogens is an important factor in increasing the incidence of soil-borne sickness. Moreover, changes in fungal and bacterial communities were predominantly driven by soil dissolved organic carbon (DOC), DOC/DON ratio (DOCN), NO3–-N, microbial biomass carbon (MBC), and MBC/MBN ratio (MBCN). Overall, the long-term monoculture of Chinese fir promotes the microecological imbalance of rhizosphere and bulk soil, and remarkably reduced soil microbial community diversity. These results can provide a scientific support for the implementation of future management measures for fir plantations (e.g., fertilization, addition of microbial fungicides, and construction of mixed forests).

The complex composition and interaction of root-associated microbes are critical to plant health and performance. In this study, we presented a detailed characterization of three rhizocompartment (rhizosphere, rhizoplane, and root) microbiomes of Achyranthes bidentata under different years of consecutive monoculture by deep sequencing in order to determine keystone microorganisms via co-occurrence network analysis. The network analysis showed that multiple consecutive monoculture (MCM, represented 5Y and 10Y) soils generated some distinct beneficial bacterial taxa such as Bacillus, Fictibacillus, Bradyrhizobium, Shinella, and Herbaspirillum. For fungi, Mortierella substituted for Fusarium in occupying an important position in different rhizocompartments under A. bidentate monoculture. Quantitative PCR analysis confirmed a significant increase in Bacillus, Pseudomonas, and Burkholderia spp. The results of the inoculation assay showed that addition of beneficial bacteria Bacillus subtilis 74 and Bacillus halodurans 75 significantly increased the root length and fresh weight of A. bidentata. Furthermore, three types of phytosterones, as the main allochemicals, were identified both in the rhizosphere soil and in culture medium under sterile conditions by LC-MS/MS. When looking at in vitro interactions, it was found that phytosterones displayed a positive interaction with dominant beneficial species (Bacillus amyloliquefaciens 4 and B. halodurans 75) and had a negative effect on the presence of the pathogenic fungi Fusarium solani and Fusarium oxysporum. Overall, this study demonstrated that consecutive monoculture of A. bidentata can alter the bacterial and fungal community by secreting root exudates, leading to recruitment of beneficial microbes and replacement of plant-specific pathogenic fungi with plant beneficial fungi.

Under consecutive monoculture, the abundance of pathogenic fungi, such as Fusarium oxysporum in the rhizosphere of Radix pseudostellariae, negatively affects the yield and quality of the plant. Therefore, it is pertinent to explore the role of antagonistic fungi for the management of fungal pathogens such as F. oxysporum. Our PCR-denatured gradient gel electrophoresis (DGGE) results revealed that the diversity of Trichoderma spp. was significantly declined due to extended monoculture. Similarly, quantitative PCR analysis showed a decline in Trichoderma spp., whereas a significant increase was observed in F. oxysporum. Furthermore, seven Trichoderma isolates from the R. pseudostellariae rhizosphere were identified and evaluated in vitro for their potentiality to antagonize F. oxysporum. The highest and lowest percentage of inhibition (PI) observed among these isolates were 47.91 and 16.67%, respectively. In in vivo assays, the R. pseudostellariae treated with four Trichoderma isolates, having PI > 30%, was used to evaluate the biocontrol efficiency against F. oxysporum in which T. harzianum ZC51 enhanced the growth of the plant without displaying any disease symptoms. Furthermore, the expression of eight defense-related genes of R. pseudostellariae in response to a combination of F. oxysporum and T. harzianum ZC51 treatment was checked, and most of these defense genes were found to be upregulated. In conclusion, this study reveals that the extended monoculture of R. pseudostellariae could alter the Trichoderma communities in the plant rhizosphere leading to relatively low level of antagonistic microorganisms. However, T. harzianum ZC51 could inhibit the pathogenic F. oxysporum and induce the expression of R. pseudostellariae defense genes. Hence, T. harzianum ZC51 improves the plant resistance and reduces the growth inhibitory effect of consecutive monoculture problem.

Rehmannia glutinosa, a perennial medicinal plant, suffers from severe replant disease under consecutive monoculture. The rhizosphere microbiome is vital for soil suppressiveness to diseases and for plant health. Moreover, N-acyl homoserine lactone (AHL)-mediated quorum sensing (QS) regulates diverse behavior in rhizosphere-inhabiting and plant pathogenic bacteria. The dynamics of short-chain AHL-mediated QS bacteria driven by consecutive monoculture and its relationships with R. glutinosa replant disease were explored in this study. The screening of QS bacteria showed that 65 out of 200 strains (32.5%) randomly selected from newly planted soil of R. glutinosa were detected as QS bacteria, mainly consisting of Pseudomonas spp. (55.4%). By contrast, 34 out of 200 (17%) strains from the diseased replant soil were detected as QS bacteria, mainly consisting of Enterobacteriaceae (73.5%). Functional analysis showed most of the QS bacteria belonging to the Pseudomonas genus showed strong antagonistic activities against Fusarium oxysporum or Aspergillus flavus, two main causal agents of R. glutinosa root rot disease. However, the QS strains dominant in the replant soil caused severe wilt disease in the tissue culture seedlings of R. glutinosa. Microbial growth assays demonstrated a concentration-dependent inhibitory effect on the growth of beneficial QS bacteria (i.e., Pseudomonas brassicacearum) by a phenolic acid mixture identified in the root exudates of R. glutinosa, but the opposite was true for harmful QS bacteria (i.e., Enterobacter spp.). Furthermore, it was found that the population of quorum quenching (QQ) bacteria that could disrupt the beneficial P. brassicacearum SZ50 QS system was significantly higher in the replant soil than in the newly planted soil. Most of these QQ bacteria in the replant soil were detected as Acinetobacter spp. The growth of specific QQ bacteria could be promoted by a phenolic acid mixture at a ratio similar to that found in the R. glutinosa rhizosphere. Moreover, these quorum-quenching bacteria showed strong pathogenicity toward the tissue culture seedlings of R. glutinosa. In conclusion, consecutive monoculture of R. glutinosa contributed to the imbalance between beneficial and harmful short-chain AHL-mediated QS bacteria in the rhizosphere, which was mediated not only by specific root exudates but also by the QQ bacterial community.

Abstract Background Pseudostellaria heterophylla (P. heterophylla), a herbaceous perennial, belongs to Caryophyllaceae family and is one of the Chinese herbal medicine with high pharmacodynamic value. It can be used to treat the spleen deficiency, anorexia, weakness after illness and spontaneous perspiration symptoms. Our previous study found that consecutive monoculture of Pseudostellaria heterophylla could lead to the deterioration of the rhizosphere microenvironment. The specialized forms of pathogenic fungus Fusarium oxysporum f.Sp. heterophylla (F. oxysporum) in rhizosphere soils of P. heterophylla plays an important role in the consecutive monoculture of P. heterophylla. Results In this study, F. oxysporum was used to infect the tissue culture plantlets of P. heterophylla to study the responding process at three different infection stages by using RNA-sequencing. We obtained 127,725 transcripts and 47,655 distinct unigenes by de novo assembly and obtained annotated information in details for 25,882 unigenes. The Kyoto Encyclopedia of Genes and Genomes pathway analysis and the real-time quantitative PCR results suggest that the calcium signal system and WRKY transcription factor in the plant-pathogen interaction pathway may play an important role in the response process, and all of the WRKY transcription factor genes were divided into three different types. Moreover, we also found that the stimulation of F. oxysporum may result in the accumulation of some phenolics in the plantlets and the programmed cell death of the plantlets. Conclusions This study has partly revealed the possible molecular mechanism of the population explosion of F. oxysporum in rhizosphere soils and signal response process, which can be helpful in unraveling the role of F. oxysporum in consecutive monoculture problems of P. heterophylla.

Abstract A three-year field monoculture trial of Radix pseudostellariae and complementary laboratory studies were conducted to further elucidate the underlying mechanism responsible for significant decreases in the biomass yield and quality of R. pseudostellariae under continuous monoculture regimes. HPLC analysis indicated that continuous monoculture soil was rich in organic acids, which had cumulative effects over time. Further analysis suggested that the application of a mixture of organic acids significantly promoted growth of pathogenic fungi, and increased the expression of chemotaxis-related gene (cheA) and biofilm formation of the specific pathogenic Kosakonia sacchari. However, opposite reactions were observed in the case of Bacillus megaterium and Bacillus pumilus. Concurrently, the present results revealed that the mixed organic acids stimulated the production of toxins, as well as H2O2 in the pathogenic fungi. Furthermore, the presence of organic acids reflecting environmental conditions under monocropping had negative effects on the expression of the biocontrol-related genes, which resulted in attenuated antagonistic activities of plant growth-promoting rhizobacteria (PGPR) to suppress mycelial growth of the pathogenic fungi. These results help to unveil the mechanisms associated with how accumulated organic acids differentially mediate deterioration of soil microbial composition and structure in monocropping system.

Availability of nitrogen (N) in soil changes the composition and activities of microbial community, which is critical for the processing of soil organic matter and health of crop plants. Inappropriate application of N fertilizer can alter the rhizosphere microbial community and disturb the soil N homeostasis. The goal of this study was to assess the effect of different ratio of N fertilizer at various early to late growth stages of rice, while keeping the total N supply constant on rice growth performance, microbial community structure, and soil protein expression in rice rhizosphere. Two different N regimes were applied, i.e., traditional N application (NT) consists of three sessions including 60, 30 and 10% at pre-transplanting, tillering and panicle initiation stages, respectively, while efficient N application (NF) comprises of four sessions, i.e., 30, 30, 30, and 10%), where the fourth session was extended to anthesis stage. Soil metaproteomics combined with Terminal Restriction Fragment Length Polymorphism (T-RFLP) were used to determine the rhizosphere biological process. Under NF application, soil enzymes, nitrogen utilization efficiency and rice yield were significantly higher compared to NT application. T-RFLP and qPCR analysis revealed differences in rice rhizosphere bacterial diversity and structure. NF significantly decreased the specific microbes related to denitrification, but opposite result was observed for bacteria associated with nitrification. Furthermore, soil metaproteomics analysis showed that 88.28% of the soil proteins were derived from microbes, 5.74% from plants, and 6.25% from fauna. Specifically, most of the identified microbial proteins were involved in carbohydrate, amino acid and protein metabolisms. Our experiments revealed that NF positively regulates the functioning of the rhizosphere ecosystem and further enabled us to put new insight into microbial communities and soil protein expression in rice rhizosphere.

Radix pseudostellariae is a perennial tonic medicinal plant, with high medicinal value. However, consecutive monoculture of this plant in the same field results in serious decrease in both yield and quality. In this study, a 3-year field experiment was performed to identify the inhibitory effect of growth caused by prolonged monoculture of R. pseudostellariae. DGGE analysis was used to explore the shifts in the structure and diversity of soil Fusarium and Pseudomonas communities along a 3-year gradient of monoculture. The results demonstrated that extended monoculture significantly boosted the diversity of Fusarium spp., but declined Pseudomonas spp. diversity. Quantitative PCR analysis showed a significant increase in Fusarium oxysporum, but a decline in Pseudomonas spp. Furthermore, abundance of antagonistic Pseudomonas spp. possessing antagonistic ability toward F. oxysporum significantly decreased in consecutively monocultured soils. Phenolic acid mixture at the same ratio as detected in soil could boost mycelial and sporular growth of pathogenic F. oxysporum while inhibit the growth of antagonistic Pseudomonas sp. CJ313. Moreover, plant bioassays showed that Pseudomonas sp. CJ313 had a good performance that protected R. pseudostellariae from infection by F. oxysporum. In conclusion, this study demonstrated that extended monoculture of R. pseudostellariae could alter the Fusarium and Pseudomonas communities in the plant rhizosphere, leading to relatively low level of antagonistic microorganisms, but with relatively high level of pathogenic microorganisms.

As potent allelochemicals, phenolic acids are believed to be associated with replanting disease and cause microflora shift and structural disorder in the rhizosphere soil of continuously monocultured Radix pseudostellariae. The transcriptome sequencing was used to reveal the mechanisms underlying the differential response of pathogenic bacterium Kosakonia sacchari and beneficial bacterium Bacillus pumilus on their interactions with phenolic acids, the main allelochemicals in root exudates of R. pseudostellariae in the monoculture system. The microbes were inoculated in the pots containing soil and the medicinal plant in this study. The results showed that the addition of beneficial B. pumilus to the 2-year planted soil significantly decreased the activity of soil urease, catalase, sucrase, and cellulase and increased the activity of chitinase compared with those in the 2nd-year monocropping rhizosphere soil without any treatment. However, opposite results were obtained when K. sacchari was added. Transcriptome analysis showed that vanillin enhanced glycolysis/gluconeogenesis, fatty acid biosynthesis, pentose phosphate, bacterial chemotaxis, flagellar assembly, and phosphotransferase system pathway in K. sacchari. However, protocatechuic acid, a metabolite produced by K. sacchari from vanillin, had negative effects on the citrate cycle and biosynthesis of novobiocin, phenylalanine, tyrosine, and tryptophan in B. pumilus. Concurrently, the protocatechuic acid decreased the biofilm formation of B. pumilus. These results unveiled the mechanisms how phenolic acids differentially mediate the shifts of microbial flora in rhizosphere soil, leading to the proliferation of pathogenic bacteria (i.e., K. sacchari) and the attenuation of beneficial bacteria (i.e., B. pumilus) under the monocropping system of R. pseudostellariae.

The biomass and quality of Pseudostellariae heterophylla suffers a significant decline under monoculture. Since rhizosphere microbiome plays crucial roles in soil health, deep pyrosequencing combined with qPCR was applied to characterize the composition and structure of soil bacterial community under monoculture and different amendments. The results showed compared with the first-year planted (FP), second-year monoculture of P. heterophylla (SP) led to a significant decline in yield and resulted in a significant increase in Fusarium oxysporum but a decline in Burkholderia spp. Bio-organic fertilizer (MT) formulated by combining antagonistic bacteria with organic matter could significantly promote the yield by regulating rhizosphere bacterial community. However, organic fertilizer (MO) without antagonistic bacteria could not suppress Fusarium wilt. Multivariate statistics analysis showed a distinct separation between the healthy samples (FP and MT) and the unhealthy samples (SP and MO), suggesting a strong relationship between soil microbial community and plant performance. Furthermore, we found the application of bio-organic fertilizer MT could significantly increase the bacterial community diversity and restructure microbial community with relatively fewer pathogenic F. oxysporum and more beneficial Burkholderia spp. In conclusion, the application of novel bio-organic fertilizer could effectively suppress Fusarium wilt by enriching the antagonistic bacteria and enhancing the bacterial diversity.

This paper presents an in-situ storage topology for ultra-high-speed burst mode imagers, enabling low noise operation while keeping a high frame depth. The proposed pixel architecture contains a 4T pinned photodiode, a correlated double sampling (CDS) amplification stage, and an in-situ memory bank. Focusing on the sampling noise, the system level trade-off of the proposed pixel architecture is discussed, showing its advantages on the noise, power, and scaling capability. Integrated with an AC coupling CDS stage, the amplification is obtained by exploiting the strong capacitance to the voltage relation of a single NMOS transistor. A comprehensive noise model is developed for optimizing the trade-off between the area and noise. As a proof-of-concept, a prototype imager with a 30 µm pixel pitch was fabricated in a CMOS 130 nm technology. A 108-cell memory bank is implemented allowing dense layout and parallel readout. Two types of CDS amplification stages were investigated. Despite the limited memory capacitance of 10 fF/cell, the photon transfer curves of both pixel types were measured over different operation speeds up to 20 Mfps showing a noise performance of 8.4 e−.

Soil microbes are active players in energy flow and material exchange of the forest ecosystems, but the research on the relationship between the microbial diversity and the vegetation types is less conducted, especially in the subtropical area of China. In this present study, the rhizosphere soils of evergreen broad-leaf forest (EBF), coniferous forest (CF), subalpine dwarf forest (SDF) and alpine meadow (AM) were chosen as test sites. Terminal-restriction fragment length polymorphisms (T-RFLP) analysis was used to detect the composition and diversity of soil bacterial communities under different vegetation types in the National Natural Reserve of Wuyi Mountains. Our results revealed distinct differences in soil microbial composition under different vegetation types. Total 73 microbes were identified in soil samples of the four vegetation types, and 56, 49, 46 and 36 clones were obtained from the soils of EBF, CF, SDF and AM, respectively, and subsequently sequenced. The Actinobacteria, Fusobacterium, Bacteroidetes and Proteobacteria were the most predominant in all soil samples. The order of Shannon-Wiener index (H) of all soil samples was in the order of EBF>CF>SDF>AM, whereas bacterial species richness as estimated by four restriction enzymes indicated no significant difference. Principal component analysis (PCA) revealed that the soil bacterial communities' structures of EBF, CF, SDF and AM were clearly separated along the first and second principal components, which explained 62.17% and 31.58% of the total variance, respectively. The soil physical-chemical properties such as total organic carbon (TOC), total nitrogen (TN), total phosphorus (TP) and total potassium (TK) were positively correlated with the diversity of bacterial communities.

Soil microbes play an essential role in the forest ecosystem as an active component. This study examined the hypothesis that soil microbial community structure and metabolic activity would vary with the increasing stand ages in long-term pure plantations of Pinus elliottii. The phospholipid fatty acids (PLFA) combined with community level physiological profiles (CLPP) method was used to assess these characteristics in the rhizospheric soils of P. elliottii. We found that the soil microbial communities were significantly different among different stand ages of P. elliottii plantations. The PLFA analysis indicated that the bacterial biomass was higher than the actinomycic and fungal biomass in all stand ages. However, the bacterial biomass decreased with the increasing stand ages, while the fungal biomass increased. The four maximum biomarker concentrations in rhizospheric soils of P. elliottii for all stand ages were 18:1ω9c, 16:1ω7c, 18:3ω6c (6,9,12) and cy19:0, representing measures of fungal and gram negative bacterial biomass. In addition, CLPP analysis revealed that the utilization rate of amino acids, polymers, phenolic acids, and carbohydrates of soil microbial community gradually decreased with increasing stand ages, though this pattern was not observed for carboxylic acids and amines. Microbial community diversity, as determined by the Simpson index, Shannon-Wiener index, Richness index and McIntosh index, significantly decreased as stand age increased. Overall, both the PLFA and CLPP illustrated that the long-term pure plantation pattern exacerbated the microecological imbalance previously described in the rhizospheric soils of P. elliottii, and markedly decreased the soil microbial community diversity and metabolic activity. Based on the correlation analysis, we concluded that the soil nutrient and C/N ratio most significantly contributed to the variation of soil microbial community structure and metabolic activity in different stand ages of P. elliottii plantations.

BACKGROUND: The consecutive monoculture for most of medicinal plants, such as Rehmannia glutinosa, results in a significant reduction in the yield and quality. There is an urgent need to study for the sustainable development of Chinese herbaceous medicine. METHODOLOGY/PRINCIPAL FINDINGS: Comparative metaproteomics of rhizosphere soil was developed and used to analyze the underlying mechanism of the consecutive monoculture problems of R. glutinosa. The 2D-gel patterns of protein spots for the soil samples showed a strong matrix dependency. Among the spots, 103 spots with high resolution and repeatability were randomly selected and successfully identified by MALDI TOF-TOF MS for a rhizosphere soil metaproteomic profile analysis. These proteins originating from plants and microorganisms play important roles in nutrient cycles and energy flow in rhizospheric soil ecosystem. They function in protein, nucleotide and secondary metabolisms, signal transduction and resistance. Comparative metaproteomics analysis revealed 33 differentially expressed protein spots in rhizosphere soil in response to increasing years of monoculture. Among them, plant proteins related to carbon and nitrogen metabolism and stress response, were mostly up-regulated except a down-regulated protein (glutathione S-transferase) involving detoxification. The phenylalanine ammonia-lyase was believed to participate in the phenylpropanoid metabolism as shown with a considerable increase in total phenolic acid content with increasing years of monoculture. Microbial proteins related to protein metabolism and cell wall biosynthesis, were up-regulated except a down-regulated protein (geranylgeranyl pyrophosphate synthase) functioning in diterpenoid synthesis. The results suggest that the consecutive monoculture of R. glutinosa changes the soil microbial ecology due to the exudates accumulation, as a result, the nutrient cycles are affected, leading to the retardation of plant growth and development. CONCLUSIONS/SIGNIFICANCE: Our results demonstrated the interactions among plant, soil and microflora in the proteomic level are crucial for the productivity and quality of R. glutinosa in consecutive monoculture system.

Consecutive monoculture of Rehmannia glutinosa in the same field leads to a severe decline in both quality and yield of tuberous roots, the most useful part in traditional Chinese medicine. Fungi are an important and diverse group of microorganisms in the soil ecosystem and play crucial roles in soil health. In this study, high-throughput pyrosequencing of internal transcribed spacer 2 ribosomal DNA amplicons was applied to gain insight into how consecutive monoculture practice influence and stimulate R. glutinosa rhizosphere and bulk soil fungal communities. The results from nonmetric multidimensional scaling ordination and clustering analysis revealed distinctive differences between rhizosphere and bulk soil fungal communities. However, longer-term monocultured bulk soils were more similar to the rhizosphere soils in comparison with the shorterterm monocultured bulk soils. Moreover, consecutive monoculture caused a gradual shift in the composition and structure of the soil fungal community. The cultivation of this plant led to the appearance of some exclusive operational taxonomic units in rhizosphere or bulk soils that were assigned to the genera Fusarium, Rhizoctonia, and so on. Furthermore, the sum of the relative abundance of species of Fusarium, Cylindrocarpon, and Gibberella (belonging to the family Nectriaceae); Rhizoctonia, Thanatephorus, and Ceratobasidium (belonging to the family Ceratobasidiaceae); and Lectern and Plectosporium (belonging to the family Plectosphaerellaceae) was significantly higher in consecutively monocultured (CM) than in newly planted (NP) soil in both rhizosphere and bulk soils. In particular, Fusarium abundance was significantly higher in CM than in NP in the rhizosphere, and higher in rhizosphere soils than in bulk soils for each treatment. A pathogenicity test showed that both Fusarium strains isolated were pathogenic to R. glutinosa seedlings. In addition, the culture filtrate and mycotoxins produced by Fusarium oxysporum significantly repressed the growth of the antagonistic bacterium. Pseudomonas aeruginosa. In conclusion, consecutive monoculture of R. glutinosa restructured the fungal communities in both rhizosphere and bulk soils but bulk effects developed more slowly over time in comparison with rhizosphere effects. Furthermore, microbial interactions might lead to a reduction in the abundance of beneficial microbes. [ABSTRACT FROM AUTHOR]

Intercropping has been widely used to control disease and improve yield in agriculture. In this study, maize and peanut were used for non-separation intercropping (NS), semi-separation intercropping (SS) using a nylon net, and complete separation intercropping (CS) using a plastic sheet. In field experiments, two-year land equivalent ratios (LERs) showed yield advantages due to belowground interactions when using NS and SS patterns as compared to monoculture. In contrast, intercropping without belowground interactions (CS) showed a yield disadvantage. Meanwhile, in pot experiments, belowground interactions (found in NS and SS) improved levels of soil-available nutrients (nitrogen (N) and phosphorus (P)) and enzymes (urease and acid phosphomonoesterase) as compared to intercropping without belowground interactions (CS). Soil bacterial community assay showed that soil bacterial communities in the NS and SS crops clustered together and were considerably different from the CS crops. The diversity of bacterial communities was significantly improved in soils with NS and SS. The abundance of beneficial bacteria, which have the functions of P-solubilization, pathogen suppression, and N-cycling, was improved in maize and peanut soils due to belowground interactions through intercropping. Among these bacteria, numbers of Bacillus, Brevibacillus brevis, and Paenibacillus were mainly increased in the maize rhizosphere. Burkholderia, Pseudomonas, and Rhizobium were mainly increased in the peanut rhizosphere. In conclusion, using maize and peanut intercropping, belowground interactions increased the numbers of beneficial bacteria in the soil and improved the diversity of the bacterial community, which was conducive to improving soil nutrient (N and P) supply capacity and soil microecosystem stability. [ABSTRACT FROM AUTHOR]

Radix pseudostellariae L. is a common and popular Chinese medication. However, continuous monoculture has increased its susceptibility to severe diseases. We identified two pathogenic microorganisms, Talaromyces helicus M. (KU355274) and Kosakonia sacchari W. (KU324465), and their antagonistic bacterium, Bacillus pumilus Z. in rhizosphere soil of continuously monocultured R. pseudostellariae. Nine types of phenolic acids were identified both in the rhizosphere soil and in culture medium under sterile conditions. A syringic acid and phenolic acid mixture significantly promoted the growth of T. helicus and K. sacchari. T. helicus could utilize eight types of phenolic acids, whereas K. sacchari could only use four phenolic acids. K. sacchari produced protocatechuic acid when consuming vanillin. Protocatechuic acid negatively affected the growth of B. pumilus. The 3A-DON toxin produced by T. helicus promoted the growth of K. sacchari and inhibited growth of B. pumilus at low concentrations. These data help explain why phenolic exudates mediate a microflora shift and structure disorder in the rhizosphere soil of continuously monocultured R. pseudostellariae and lead to increased replanting disease incidence. [ABSTRACT FROM AUTHOR]

Pot experiment was carried out at different nitrogen levels in maize and peanut intercropping system to investigate the mechanisms that govern interactions between intercropped species in soil rhizosphere. Three different nitrogen levels 100%, 75% and 50% of normal nitrogen application rate were used in maize monoculture. Two peanut plants were intercropped with single maize plant per pot. Effect of reduced nitrogen application was investigated in soil rhizosphere by measuring soil enzymatic activity, nutrition, soil microbial diversity and composition. Phospholipid fatty acid (PLFA) was carried out to understand about composition of viable soil micro flora and community level physiological profile (CLPP) was applied to work out the microbial diversity. The results revealed that soil nutrition and enzymatic activities were higher in intercropping treatments as compared to mono-cropping with similar level of nitrogen. The results of PLFA analysis showed that intercropping inhibited the fungal population and promoted the bacterial community. The results of AWCD showed that 4 kinds of carbon utilization sources (carboxylic acid, polymer, amines & carbohydrates) increased by the decrease of nitrogen levels in intercropping treatments and other 2 kinds of carbon sources (phenolic acids & amino acids) showed different trend. Cluster analysis (CA) and Principal component analysis (PCA) carried out from the results of PLFA and CLPP indicated distinct separation amongst all treatments of monocropping and intercropping as well as with different level of nitrogen application that reflected a variation in soil microbial activity and composition. However intercropping treatment with 75% nitrogen level performed similar to 100% monocropping treatment of maize. The results revealed that by intercropping of peanut with maize can be used to decrease the dependence on artificial fertilizer which provides a convenient organic farming model for growers across the world. [ABSTRACT FROM AUTHOR]