Pseudostellaria heterophylla improves intestinal microecology through modulating gut microbiota and metabolites in mice.

BACKGROUND: Pseudostellaria heterophylla is a Chinese medicine and healthy edible that is widely used to for its immunomodulatory, antioxidant, antidiabetic and antitussive properties. However, the potential function of P. heterophylla in intestinal microecology remains unclear. In this study, we investigated the impact of P. heterophylla on immune functions and evaluated its potential to regulate the gut microbiota and metabolome. RESULTS: The results showed that P. heterophylla significantly increased the content of red blood cells, total antioxidant capacity and expression of immune factors, and decreased platelet counts when compared to the control under cyclophosphamide injury. In addition, P. heterophylla altered the diversity and composition of the gut bacterial community; increased the abundance of potentially beneficial Akkermansia, Roseburia, unclassified Clostridiaceae, Mucispirillum, Anaeroplasma and Parabacteroides; and decreased the relative abundance of pathogenic Cupriavidus and Staphylococcus in healthy mice. Metabolomic analyses showed that P. heterophylla significantly increased the content of functional oligosaccharides, common oligosaccharides, vitamins and functional substances. Probiotics and pathogens were regulated by metabolites across 11 pathways in the bacterial-host co-metabolism network. CONCLUSION: We demonstrated that P. heterophylla increased the abundance of probiotics and decreased pathogens, and further stimulated host microbes to produce beneficial secondary metabolites for host health. Our studies highlight the role of P. heterophylla in gut health and provide new insights for the development of traditional Chinese medicine in the diet. © 2024 Society of Chemical Industry.

Niche differentiation modulates metabolites abundance and composition in silicon fertilizer amended soil during sugarcane growth.

BACKGROUND: As one of the vital crops globally, sugarcane (Saccharum officinarum L.) has been one of model crops for conducting metabolome research. Although many studies have focused on understanding bioactive components in specific sugarcane tissues, crucial questions have been left unanswered about the response of metabolites to niche differentiation such as different sugarcane tissues (leaf, stem and root), and soil regions (rhizosphere and bulk) under silicon (Si) amended soils. Here, nontargeted metabolite profiling method was leveraged to assess the similarities and differences in the abundance and community composition of metabolites in the different sugarcane and soil compartments. Identify the compartment-specific expression patterns of metabolites, and their association with cane agronomic traits and edaphic factors. We also investigated the response of sugarcane agronomic traits and edaphic factors to Si amended soil. RESULTS: We found that Si fertilizer exhibited the advantages of overwhelmingly promoting the height and theoretical production of cane, and profoundly increased soil Si content by 24.8 and 27.0%, while soil available potassium (AK) was enhanced by 3.07 and 2.67 folds in the bulk and rhizosphere soils, respectively. It was also noticed that available phosphorus (AP) in the rhizosphere soil tremendously increased by 105.5%. We detected 339 metabolites in 30 samples using LC-MS/MS analyses, 161 of which were classified and annotated, including organooxygen compounds (19.9%), carboxylic acids and derivatives (15.5%), fatty acyls (15.5%), flavonoids (4.4%), phenols (4.4%), and benzene and substituted derivatives (3.7%). In addition, the total percentages covered by these core metabolites in each compartment ranged from 94.0% (bulk soil) to 93.4% (rhizosphere soil), followed by 87.4% (leaf), 81.0% (root) and 80.5% (stem), suggesting that these bioactive compounds may have migrated from the belowground tissues and gradually filtered in various aboveground niches of the plant. We also observed that the variations and enrichment of metabolites abundance and community were compartment-specific. Furthermore, some key bioactive compounds were markedly associated with plant growth parameters and soil edaphic. CONCLUSION: Taken together, we hypothesized that Si utilization can exhibit the advantage of enhancing edaphic factors and cane agronomic traits, and variations in metabolites community are tissue-specific.

[Changes in physicochemical property and microbial community of Pseudostellaria heterophylla soil at diffe-rent fallow ages]. / 太子参不同休耕年限土壤理化特征和微生物群落变化.

Pseudostellaria heterophylla is authentic traditional Chinese herbal medicine in Fujian Province. P. hete-rophylla suffers from serious consecutive monoculture problems. Fallow can alleviate such problems, but the mecha-nism is still unclear. In this study, high-throughput sequencing was performed to analyze the changes in soil microbial community structure and diversity in the P. heterophylla soil at different fallow ages as well as their relationships with soil physicochemical properties and phenolic acids. The results showed that fungal community diversity decreased but bacterial community diversity increased in fallow soils compared with the control soil of P. heterophy-lla. For bacterial communities, the relative abundance of Acidobacteria increased, while that of Proteobacteria and Actinobacteria decreased in fallow soils. For fungal communities, the relative abundance of dominant phyla had no significant difference between fallow and control soils. Soil acidity and organic matter content showed a trend of weakening and decreasing, respectively, with the increases of fallow years. In addition, with the increases of fallow years, the content of phenolic acids in soil, including benzoic acid and salicylic acid, showed significant decrease, while some other phenolic acids such as p-coumaric acid were accumulated obviously. Taken together, fallow could efficiently ameliorate the structure of soil microbial community and soil properties of P. heterophylla, and thus alleviate the effects of continuous cropping.

Deciphering the Molecular Mechanisms of Chilling Tolerance in Lsi1-Overexpressing Rice.

Improving tolerance to low-temperature stress during the rice seedling stage is of great significance in agricultural science. In this study, using the low silicon gene 1 (Lsi1)-overexpressing (Dular-OE) and wild-type rice (Dular-WT), we showed that Lsi1 overexpression enhances chilling tolerance in Dular-OE. The overexpression of the Lsi1 increases silicon absorption, but it was not the main reason for chilling tolerance in Dular-OE. Instead, our data suggest that the overexpression of a Lsi1-encoding NIP and its interaction with key proteins lead to chilling tolerance in Dular-OE. Additionally, we show that the high-mobility group protein (HMG1) binds to the promoter of Lsi1, positively regulating its expression. Moreover, Nod26-like major intrinsic protein (NIP)'s interaction with α and ß subunits of ATP synthase and the 14-3-3f protein was validated by co-immunoprecipitation (Co-IP), bimolecular fluorescent complementary (BiFC), and GST-pulldown assays. Western blotting revealed that the overexpression of NIP positively regulates the ATP-synthase subunits that subsequently upregulate calcineurin B-like interacting protein kinases (CIPK) negatively regulating 14-3-3f. Overall, these NIP-mediated changes trigger corresponding pathways in an orderly manner, enhancing chilling tolerance in Dular-OE.

Bio-fertilizer Affects Structural Dynamics, Function, and Network Patterns of the Sugarcane Rhizospheric Microbiota.

Fertilizers and microbial communities that determine fertilizer efficiency are key to sustainable agricultural development. Sugarcane is an important sugar cash crop in China, and using bio-fertilizers is important for the sustainable development of China's sugar industry. However, information on the effects of bio-fertilizers on sugarcane soil microbiota has rarely been studied. In this study, the effects of bio-fertilizer application on rhizosphere soil physicochemical indicators, microbial community composition, function, and network patterns of sugarcane were discussed using a high-throughput sequencing approach. The experimental design is as follows: CK: urea application (57 kg/ha), CF: compound fertilizer (450 kg/ha), BF1: bio-fertilizer (1500 kg/ha of bio-fertilizer + 57 kg/ha of urea), and BF2: bio-fertilizer (2250 kg/ha of bio-fertilizer + 57 kg/ha of urea). The results showed that the bio-fertilizer was effective in increasing sugarcane yield by 3-12% compared to the CF treatment group, while reducing soil acidification, changing the diversity of fungi and bacteria, and greatly altering the composition and structure of the inter-root microbial community. Variance partitioning canonical correspondence (VPA) analysis showed that soil physicochemical variables explained 80.09% and 73.31% of the variation in bacteria and fungi, respectively. Redundancy analysis and correlation heatmap showed that soil pH, total nitrogen, and available potassium were the main factors influencing bacterial community composition, while total soil phosphorus, available phosphorus, pH, and available nitrogen were the main drivers of fungal communities. Volcano plots showed that using bio-fertilizers contributed to the accumulation of more beneficial bacteria in the sugarcane rhizosphere level and the decline of pathogenic bacteria (e.g., Leifsonia), which may slow down or suppress the occurrence of diseases. Linear discriminant analysis (LDA) and effect size analysis (LEfSe) searched for biomarkers under different fertilizer treatments. Meanwhile, support vector machine (SVM) assessed the importance of the microbial genera contributing to the variability between fertilizers, of interest were the bacteria Anaerolineace, Vulgatibacter, and Paenibacillus and the fungi Cochliobolus, Sordariales, and Dothideomycetes between CF and BF2, compared to the other genera contributing to the variability. Network analysis (co-occurrence network) showed that the network structure of bio-fertilizers was closer to the network characteristics of healthy soils, indicating that bio-fertilizers can improve soil health to some extent, and therefore if bio-fertilizers can be used as an alternative to chemical fertilizers in the future alternative, it is important to achieve green soil development and improve the climate.

The appearance of volatile aromas in Tieguanyin tea with different elevations.

Chinese people have consistentlypreferred high mountain tea because specific flavors are memorable for them, and also, people have traditionally considered this type of tea to be good for health. Tieguanyin is one of the famous traditional Chinese tea that has ever-changing aromas. To illustrate the various characteristics in volatile fragment compounds from Tieguanyin tea, fresh tea leaves collected from different elevations (450, 650, and 900 m) were detected using GC-MS by solid-liquid extraction. The results showed that volatile aromatic compounds, such as benzyl alcohol, phenyl ethanol, and acetophenone, were the most abundant in tea leaves located with high elevation. Meanwhile, 1-hexanol, 1-nananol, and nanoic acid, as a type of aliphatic aroma, were more prevalent in low-elevation tea orchards. Catechols and alkaloids are largely cumulated in low- and high-elevation tea leaves, respectively. Our findings also showed that elemene was widely consisted of high-elevation tea metabolites. It provided practicality for the preparation of tea manufacturing in major Tieguanyin tea-producing regions.

First Report of Mucor spp. Causing Root Rot of Radix pseudstellariae in Guizhou Province of China.

Radix pseudostellariae L. is one of the most common and highly-prized Chinese medicinal plants with various pharmacological effects, and mainly produced in acid soils in the Guizhou and Fujian provinces of southwestern and southeastern China, respectively (Wu et al. 2020). However, consecutive monoculture of R. pseudostellariae results in severe root rot and decline in biomass and quality of underground tubers. Root tubers of R. pseudostellariae are typically planted in December and harvested in next June. Root rot commonly starts developing in May. The disease incidence of root rot was ranging from 37 to 46% in root portions and basal stem of R. pseudostellariae under the consecutive monoculture fields in Shibing County, Guizhou Province, China (108°12'E, 27°03'N) (Li et al. 2017). Severe root rot was observed in Shibing County in May 2018. Infected plants displayed curly, withered, and yellow leaves, blight, retarded growth, root rot, and yield losses. Abundant whitish mycelia were observed on roots and surrounding soil. Two fungal isolates, designated GZ20190123 and GZ20190124, were obtained from symptomatic roots cultured on potato dextrose agar (PDA). The optimum temperature range for growth of the two isolates was 25 to 30°C. The optimum pH range for the growth of GZ20190123 was 5 to 5.5, whereas GZ20190124 grew better between pH 5 to 8.5. The mean mycelial growth rates of GZ20190123 and GZ20190124 at 30°C were 2.1 and 1.5 cm/day, respectively. Conidia of the two isolates were ovoid or obclavate and were produced in single or branched chains. The internal transcribed spacer (ITS) region was amplified with primers ITS1 and ITS4 (White et al. 1990). The sequences were deposited in GenBank as accession No. MN726736 for GZ20190123 and MN726738 for GZ20190124. Sequence comparison revealed 99% (GZ20190123) and 97% (GZ20190124) identity with previously reported isolate xsd08071 of Mucor racemosus Bull. (accession No. FJ582639.1) and isolate BM3 of Mucor fragilis Bainier (accession No. MK910058.1), respectively, which was confirmed by phylogenetic analysis. The two isolates were tested for pathogenicity on R. pseudostellariae. Six roots of R. pseudostellariae were surface-sterilized with 75% ethanol and stab inoculated with mycelia using a sterile toothpick for each isolate. Sterile distilled water was stab inoculated to twelve roots to serve as the control. Treated roots were incubated in a greenhouse with 16 h day length [light intensity 146.5 µmol/(m2·s)] and day/night temperature 26°C/18°C. The inoculated roots showed the expected symptoms on roots and sprouts 7 days after inoculation, whereas the control roots with sprouts did not show any symptom. The fungi were re-isolated from the diseased roots and confirmed as expected M. racemosus or M. fragilis based on the ITS sequences, which satisfied Koch's postulates. Thus, isolate GZ20190123 was identified as M. racemosus and GZ20190124 as M. fragilis. Previously, M. racemosus and M. fragilis have been reported as a pathogen on tomato (Kwon and Hong 2005) and grape (Ghuffar et al. 2018), respectively. To our knowledge, this is the first report of M. racemosus and M. fragilis causing root rot of R. pseudostellariae in southwestern China, where the disease could cause a significant loss to production of this important medicinal plant.

Linking Short-Chain N-Acyl Homoserine Lactone-Mediated Quorum Sensing and Replant Disease: A Case Study of Rehmannia glutinosa.

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.

[Effects of Rehmannia glutinosa consecutive monoculture on the community structure and diversity of phyllosphere bacteria]. / åœ°é»„è¿žä½œå¯¹å¶é™ ç»†èŒç¾¤è½ç»“æž„åŠå¤šæ ·æ€§çš„å½±å“.

Rehmannia glutinosa, a perennial herbaceous species, belongs to the family Scrophularia-ceae. As a staple medicinal material, its tuberous roots are highly valued in traditional Chinese medicine. However, R. glutinosa suffers from serious consecutive monoculture problems in production, which leads to a decline in both productivity and quality. Phyllosphere bacteria, the most abundant component of phyllosphere microorganisms, play crucial roles in plant growth and health. Characterization of phyllosphere bacteria could provide new insights into the mechanisms of consecutive monoculture problems and their control measures. Meanwhile, the varied taxa could be served as an important indicator of consecutive monoculture problems. The barcoded pyrosequencing of 16S rDNA genes combined with a culture-dependent approach was applied to characterize the shifts of bacterial community structure and diversity in the phyllosphere under consecutive monoculture of R. glutinosa. The results showed that consecutive monoculture clearly affected bacterial community structure in the phyllosphere. The phyllosphere bacterial communities of the two-year monocultured (TY) and the diseased plants (DP) were more similar, and different from the one-year monocultured (OY). The evenness, Shannon and Simpson diversity indices were significantly lower in TY and DP than in OY. Species annotation showed that bacterial community in R. glutinosa phyllosphere mainly consisted of Proteobacteria (91.2%), Firmicutes (5.1%) and Actinobacteria (3.7%). There was no significant difference in the number of detected bacterial taxa. However, Proteobacteria was significantly increased while Firmicutes and Actinobacteria were significantly decreased under consecutive monoculture. At the genus level, the relative abundances of genera Exiguobacterium, Bacillus and Arthrobacter, potentially beneficial microorganisms, were significantly higher in OY than that in TY and DP, but it was opposite for the genus Pseudomonas. The results from the culture-dependent approach and pathogenicity test showed that Pseudomonas plecoglossicida D9, widely isolated from the diseased leaves, was highly pathogenic to leaves. In conclusion, R. glutinosa monoculture resulted in distinct phyllosphere bacterial community variation with the accumulation of pathogen loads at the expense of beneficial microorganisms, which could contribute to the occurrence of leaf disease symptoms,and aggravate R. glutinosa replant disease in a monoculture regime.

The effects of chemical and organic fertilizer usage on rhizosphere soil in tea orchards.

Sustainable agriculture is an important global issue. The use of organic fertilizers can enhance crop yield and soil properties while restraining pests and diseases. The objective of this study was to assess the effects of long-term use of chemical and organic fertilizers on tea and rhizosphere soil properties in tea orchards. Inductively coupled plasma mass spectrometry (ICP-MS) and high-throughput sequencing technology analyses were used to investigate heavy metals content and bacterial composition in rhizosphere soils. Our results indicated that organic fertilizer treatment significantly decreased Cu, Pb and Cd contents in rhizosphere soil sample. The results also showed that treatment with organic fertilizer significantly decreased the contents of Cd, Pb and As in tea leaves. Furthermore, organic fertilizer significantly increased the amino acids content of tea and the pH of the soil. The use of organic fertilizer significantly increased in the relative abundance of Burkholderiales, Myxococcales, Streptomycetales, Nitrospirales, Ktedonobacterales, Acidobacteriales, Gemmatimonadales, and Solibacterales, and decreased the abundance of Pseudonocardiales, Frankiales, Rhizobiales, and Xanthomonadales. In conclusion, organic fertilizer can help to shape the microbial composition and recruit beneficial bacteria into the rhizosphere of tea, leading to improved tea quality and reduced heavy metals content in rhizosphere soil and tea leaves.

In-Silico Prediction and Modeling of the Quorum Sensing LuxS Protein and Inhibition of AI-2 Biosynthesis in Aeromonas hydrophila.

luxS is conserved in several bacterial species, including A. hydrophila, which causes infections in prawn, fish, and shrimp, and is consequently a great risk to the aquaculture industry and public health. luxS plays a critical role in the biosynthesis of the autoinducer-2 (AI-2), which performs wide-ranging functions in bacterial communication, and especially in quorum sensing (QS). The prediction of a 3D structure of the QS-associated LuxS protein is thus essential to better understand and control A. hydrophila pathogenecity. Here, we predicted the structure of A. hydrophila LuxS and characterized it structurally and functionally with in silico methods. The predicted structure of LuxS provides a framework to develop more complete structural and functional insights and will aid the mitigation of A. hydrophila infection, and the development of novel drugs to control infections. In addition to modeling, the suitable inhibitor was identified by high through put screening (HTS) against drug like subset of ZINC database and inhibitor ((-)-Dimethyl 2,3-O-isopropylidene-l-tartrate) molecule was selected based on the best drug score. Molecular docking studies were performed to find out the best binding affinity between LuxS homologous or predicted model of LuxS protein for the ligand selection. Remarkably, this inhibitor molecule establishes agreeable interfaces with amino acid residues LYS 23, VAL 35, ILE76, and SER 90, which are found to play an essential role in inhibition mechanism. These predictions were suggesting that the proposed inhibitor molecule may be considered as drug candidates against AI-2 biosynthesis of A. hydrophila. Therefore, (-)-Dimethyl 2,3-O-isopropylidene-l-tartrate inhibitor molecule was studied to confirm its potency of AI-2 biosynthesis inhibition. The results shows that the inhibitor molecule had a better efficacy in AI-2 inhibition at 40 µM concentration, which was further validated using Western blotting at a protein expression level. The AI-2 bioluminescence assay showed that the decreased amount of AI-2 biosynthesis and downregulation of LuxS protein play an important role in the AI-2 inhibition. Lastly, these experiments were conducted with the supplementation of antibiotics via cocktail therapy of AI-2 inhibitor plus OXY antibiotics, in order to determine the possibility of novel cocktail drug treatments of A. hydrophila infection.

Comparative Metagenomic Analysis of Rhizosphere Microbial Community Composition and Functional Potentials under Rehmannia glutinosa Consecutive Monoculture.

Consecutive monoculture of Rehmannia glutinosa, highly valued in traditional Chinese medicine, leads to a severe decline in both quality and yield. Rhizosphere microbiome was reported to be closely associated with the soil health and plant performance. In this study, comparative metagenomics was applied to investigate the shifts in rhizosphere microbial structures and functional potentials under consecutive monoculture. The results showed R. glutinosa monoculture significantly decreased the relative abundances of Pseudomonadaceae and Burkholderiaceae, but significantly increased the relative abundances of Sphingomonadaceae and Streptomycetaceae. Moreover, the abundances of genera Pseudomonas, Azotobacter, Burkholderia, and Lysobacter, among others, were significantly lower in two-year monocultured soil than in one-year cultured soil. For potentially harmful/indicator microorganisms, the percentages of reads categorized to defense mechanisms (i.e., ATP-binding cassette (ABC) transporters, efflux transporter, antibiotic resistance) and biological metabolism (i.e., lipid transport and metabolism, secondary metabolites biosynthesis, transport and catabolism, nucleotide transport and metabolism, transcription) were significantly higher in two-year monocultured soil than in one-year cultured soil, but the opposite was true for potentially beneficial microorganisms, which might disrupt the equilibrium between beneficial and harmful microbes. Collectively, our results provide important insights into the shifts in genomic diversity and functional potentials of rhizosphere microbiome in response to R. glutinosa consecutive monoculture.

Rhizosphere Fungal Community Dynamics Associated with Rehmannia glutinosa Replant Disease in a Consecutive Monoculture Regime.

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 shorter-term 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 Lectera 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.

Alleviatory effect of spent Pleurotus eryngii Quel substrate on replant problem of Rehmannia glutinosa Libosch.

Rehmannia glutinosa Libosch. is a medicinal plant cultivated at a commercial scale in China. However, replanting problems result in a severe decline in both the biomass and quality of its roots, which are of greatest medicinal value. This study attempted to remediate the replant soil using spent Pleurotus eryngii Quel substrate for alleviating this issue, and to investigate the underlying mechanisms. Results showed that R. glutinosa grew successfully in fresh soil and remedial replant soil, while no roots were harvested in the unremedied replant soil. Overall, the nutritional status in the remedial soil was higher than that of the unremedied and fresh soil, while the concentration of allelopathic phenolic acids was lower. When planted in unremedied soil, the growth of five plant pathogens was induced and one beneficial fungus was suppressed. When planted in remedied soil, four out of the five pathogens were suppressed, while two beneficial fungi were identified in the remedial soil. This study suggests that the spent P. eryngii substrate significantly alleviates the replant problem of R. glutinosa, and that the alleviatory function reflects a synergetic effect, including the supplementation of soil nutrition, the degradation of allelochemicals, and the remediation of unbalanced microbial community.

Assessment of the Diversity of Pseudomonas spp. and Fusarium spp. in Radix pseudostellariae Rhizosphere under Monoculture by Combining DGGE and Quantitative PCR.

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.

Transcriptome analysis of Pseudostellaria heterophylla in response to the infection of pathogenic Fusarium oxysporum.

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.

Spatial Distribution Patterns of Root-Associated Bacterial Communities Mediated by Root Exudates in Different Aged Ratooning Tea Monoculture Systems.

Positive plant-soil feedback depends on beneficial interactions between roots and microbes for nutrient acquisition; growth promotion; and disease suppression. Recent pyrosequencing approaches have provided insight into the rhizosphere bacterial communities in various cropping systems. However; there is a scarcity of information about the influence of root exudates on the composition of root-associated bacterial communities in ratooning tea monocropping systems of different ages. In Southeastern China; tea cropping systems provide the unique natural experimental environment to compare the distribution of bacterial communities in different rhizo-compartments. High performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) was performed to identify and quantify the allelochemicals in root exudates. A high-throughput sequence was used to determine the structural dynamics of the root-associated bacterial communities. Although soil physiochemical properties showed no significant differences in nutrients; long-term tea cultivation resulted in the accumulation of catechin-containing compounds in the rhizosphere and a lowering of pH. Moreover; distinct distribution patterns of bacterial taxa were observed in all three rhizo-compartments of two-year and 30-year monoculture tea; mediated strongly by soil pH and catechin-containing compounds. These results will help to explore the reasons why soil quality and fertility are disturbed in continuous ratooning tea monocropping systems; and to clarify the associated problems.

Differential proteomic analysis of replanted Rehmannia glutinosa roots by iTRAQ reveals molecular mechanisms for formation of replant disease.

BACKGROUND: The normal growth of Rehmannia glutinosa, a widely used medicinal plant in China, is severely disturbed by replant disease. The formation of replant disease commonly involves interactions among plants, allelochemicals and microbes; however, these relationships remain largely unclear. As a result, no effective measures are currently available to treat replant disease. RESULTS: In this study, an integrated R. glutinosa transcriptome was constructed, from which an R. glutinosa protein library was obtained. iTRAQ technology was then used to investigate changes in the proteins in replanted R. glutinosa roots, and the proteins that were expressed in response to replant disease were identified. An integrated R. glutinosa transcriptome from different developmental stages of replanted and normal-growth R. glutinosa produced 65,659 transcripts, which were accurately translated into 47,818 proteins. Using this resource, a set of 189 proteins was found to be significantly differentially expressed between normal-growth and replanted R. glutinosa. Of the proteins that were significantly upregulated in replanted R. glutinosa, most were related to metabolism, immune responses, ROS generation, programmed cell death, ER stress, and lignin synthesis. CONCLUSIONS: By integrating these key events and the results of previous studies on replant disease formation, a new picture of the damaging mechanisms that cause replant disease stress emerged. Replant disease altered the metabolic balance of R. glutinosa, activated immune defence systems, increased levels of ROS and antioxidant enzymes, and initiated the processes of cell death and senescence in replanted R. glutinosa. Additionally, lignin deposition in R. glutinosa roots that was caused by replanting significantly inhibited tuberous root formation. These key processes provide important insights into the underlying mechanisms leading to the formation of replant disease and also for the subsequent development of new control measures to improve production and quality of replanted plants.

Insights into the Mechanism of Proliferation on the Special Microbes Mediated by Phenolic Acids in the Radix pseudostellariae Rhizosphere under Continuous Monoculture Regimes.

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.

Enhancement of trichothecene mycotoxins of Fusarium oxysporum by ferulic acid aggravates oxidative damage in Rehmannia glutinosa Libosch.

Rehmannia glutinosa is an important medicinal herb that cannot be replanted in the same field due to the effects of autotoxic substances. The effects of these substances on R. glutinosa in continuous cropping systems are unknown. In the present study, bioassays revealed that R. glutinosa exhibited severe growth restriction and higher disease indices in the FO+FA (F.oxysporum pretreated with ferulic acid) treatment. The increases in the contents of MDA and H2O2 were greater in the FA+FO treatment than in the FA or FO only treatments, respectively. Consistent with this result, the enzyme activities in the seedlings increased with treatment time. To identify the main factor underlying the increased pathogenicity of FO, macroconidia and trichothecene mycotoxins coproduced by FO were separated and used to treat R. glutinosa seedlings. The MDA and H2O2 contents were similar in the seedlings treated with deoxynivalenol and in the FA+FO treatment. Quantification of the relative expression of certain genes involved in Ca2+ signal transduction pathways suggested that trichothecene mycotoxins play an important role in the increased pathogenicity of FO. In conclusion, FA not only directly enhances oxidative damage in R. glutinosa but also increases wilting symptom outbreaks by promoting the secretion of trichothecene mycotoxins by FO.