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Outbreaks of leaf spot disease occurred in Rehmannia glutinosa fields in Henan Province, China, in 2019, with the incidence ranging from 20% to 40%. R. glutinosa plants with diseased leaves were collected, and 25 isolates were obtained. Pathogenicity tests, morphological observations, and phylogenetic analyses were conducted to identify the pathogens, and the biological characteristics and control agents of the pathogens were studied. Five isolates of pathogenic fungi were isolated. Three isolates were identified as Fusarium equiseti, which is a new pathogen causing R. glutinosa leaf disease; the other two isolates were identified as Fusarium acuminatum. The mycelia of F. equiseti grew fastest on Czapek medium, and the optimal temperature and pH were 25 °C and 10.0, respectively. The mycelia of F. equiseti grew from 5 °C t o 35 °C, and the lethal temperature was 55 °C. The optimal carbon and nitrogen sources were soluble starch and peptone, respectively. Eight fungicides had inhibitory effects on the mycelial growth of F. equiseti and F. acuminatum. Prochloraz had higher activities against F. equiseti and F. acuminatum, with EC50 values of 0.139 mg·L−1 and 0.123 mg·L−1, respectively. These results provide useful information that will aid the development of management strategies to control leaf diseases of R. glutinosa caused by F. equiseti and F. acuminatum. [ABSTRACT FROM AUTHOR]

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 FROM AUTHOR]

The production and quality of Rehmannia glutinosa can be dramatically reduced by replant disease under consecutive monoculture. The root-associated microbiome, also known as the second genome of the plant, was investigated to understand its impact on plant health. Culture-dependent and culture-independent pyrosequencing analysis was applied to assess the shifts in soil bacterial communities in the rhizosphere and rhizoplane under consecutive monoculture. The results show that the root-associated microbiome (including rhizosphere and rhizoplane microbiomes) was significantly impacted by rhizocompartments and consecutive monoculture. Consecutive monoculture of R. glutinosa led to a significant decline in the relative abundance of the phyla Firmicutes and Actinobacteria in the rhizosphere and rhizoplane. Furthermore, the families Flavobacteriaceae, Sphingomonadaceae, and Xanthomonadaceae enriched while Pseudomonadaceae, Bacillaceae, and Micrococcaceae decreased under consecutive monoculture. At the genus level, Pseudomonas, Bacillus, and Arthrobacter were prevalent in the newly planted soil, which decreased in consecutive monocultured soils. Besides, culture-dependent analysis confirmed the widespread presence of Pseudomonas spp. and Bacillus spp. in newly planted soil and their strong antagonistic activities against fungal pathogens. In conclusion, R. glutinosa monoculture resulted in distinct root-associated microbiome variation with a reduction in the abundance of beneficial microbes, which might contribute to the declined soil suppressiveness to fungal pathogens in the monoculture regime. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The perennial herbaceous plant, Rehmannia glutinosa Libosch, is one of traditional Chinese medicines with a long history of cultivation. However, replanted disease severely affects its yield and quality in production. In this study, a specific culture device was designed to accurately isolate the root exudates of R. glutinosa. In addition, the formation mechanism of replanted diseases mediated by root exudates was deeply studied in R. glutinosa. The results indicated that root exudates have obvious allelopathic activity, furthermore, metagenomics analysis found that the exudates were found to significantly induce the proliferation of harmful pathogenic fungal and the reduction of probiotics in rhizosphere of R. glutinosa. Further analysis found that, 8,758 genes were differentially expressed in root exudate-treated R. glutinosa plants. These genes mainly involved in critical cellular processes including immune response, hormone metabolism, signaling transduction and cell membrane transport. Of which, numerous genes were found to involve in immune response, such as PR (Pathogenesis-related protein), were highly expressed in root exudate-treated plants. Transiently overexpression experiments found that a PR1 could enhance the resistance of R. glutinosa to root exudates treatment. These results indicated that the interaction between root exudates and microbes altered the expression pattern of the genes related to immune pathway and signaling transduction mediated by it. These disordered genes finally severely affected the growth and development of R. glutinosa , and eventually formed the replanted disease. This study provides a novel approach to collect root exudates and a new data basis for revealing the molecular events occurring in replanted plants. • Constructed a special device to accurately collect root exudates and eliminate the interference in soils. • Root exudates promote the proliferation of pathogenic organisms such as Fusarium oxysporum. • The replanted diseases of R. glutinosa are accompanied by immune response and ROS accumulation. • PR1 genes played the key role in root exudate-mediated replanted disease of R. glutinosa. [ABSTRACT FROM AUTHOR]

The rapid accumulation of lignin in the cell wall is one of important immune defense mechanism in response to adversity stress in plants. In this study, we found that the enlargement of the root tubers of Rehmannia glutinosa (R. glutinosa) is arrested under consecutive monoculture stress, and this process is accompanied by abnormal accumulation of lignin. Meanwhile, the function of key catalytic enzyme genes in lignin biosynthetic pathway under consecutive monoculture stress was systematically analyzed, of which roles of core genes were validated using reverse genetics. We elucidated that an abnormal deposition of lignin in R. glutinosa roots, induced by consecutive monoculture stress, and arrested the enlargement of root tubers. Additionally, by manipulating the key catalytic enzyme gene RgCCR6 , we were able to alter lignin content of roots of R. glutinosa , thereby affecting tuber enlargement. We speculate that cell lignification is an important defense strategy in resistance against consecutive monoculture stress, but the overreacted defense hindered the normal enlargement of root tubers. The findings provide new insights for effectively improving yield reductions of root crops subjected to environmental stress. • Consecutive monoculture stress induced special deposition of lignin in the cambium cells. • Over-immunity led to abnormal deposition of lignin in cambium cells. • Abnormal deposition of lignin limited root tubers enlargement. • RgCCR in lignin biosynthesis was core node affected root tubers enlargement. [ABSTRACT FROM AUTHOR]

Consecutive monoculture of Rehmannia glutinosa results in severe replant disease and remarkably reduces the yield and quality. Rhizosphere microbiome plays a crucial role in soil health and plant performance. In this study, high-throughput pyrosequencing combined with a culture-dependent approach were applied to analyze the changes in rhizosphere microbiome structure and bacteria-fungi inter-kingdom associations under R. glutinosa consecutive monoculture. The results revealed a distinct separation between the newly planted (NP) and the two-year monocultured (SM). R. glutinosa consecutive monoculture resulted in a significant decline in the Shannon and Simpson diversity indices for the fungal community, and a significant decrease in relative abundances of the phyla Actinobacteria , Chloroflexi and Basidiomycota. The relative abundances of the genera Streptomyces , Arthrobacter , Bacillus , Nocardioides and Lysobacter were significantly higher in NP than in SM, while the opposite was true for Fusarium. Quantitative PCR confirmed a significant reduction in the phylum Actinobacteria and the genus Bacillus , and a significant increase in Fusarium oxysporum. Furthermore, the abundances of Actinobacteria and Bacillus strains with antagonistic activities against Fusarium oxysporum were lower in extended monoculture soils. Most of the isolated antagonistic bacteria were found to be positive for different plant growth promotion activities such as solubilization of phosphate and potassium, production of siderophore and indolic compounds. In addition, it was found that phenolic acid mixture and catalpol, identified in the root exudates of R. glutinosa , could selectively inhibit or stimulate different microbes. Moreover, the culture filtrate and volatile organic compounds produced by F. oxysporum could in turn suppress the growth of certain antagonistic bacteria. In conclusion, these findings highlight the close association between R. glutinosa replant disease and rhizosphere microbiome assembly, which was mediated by root exudates and microbe-microbe interactions. • Consecutive cropping of Rehmannia glutinosa affects rhizosphere microbiome assembly. • Consecutive cropping significantly decreased rhizosphere fungal community diversity. • Abundances of some plant-growth-promoting rhizobacteria decreased under monoculture. • Proliferation of Fusarium oxysporum inhibited the growth of beneficial bacteria. • Root exudates modulated the dysbiosis of rhizosphere microbiota. [ABSTRACT FROM AUTHOR]

Abstract: Rehmannia glutinosa, a widely used Chinese medicinal herb, has been shown to suffer from serious consecutive monoculture problems that cause significant decline in both yield and quality. The objective of this study was to evaluate the response of the soil microbial community and the effect on catabolic diversity to consecutive monoculture regime using three different techniques: substrate-induced respiration (SIR), phospholipid fatty acid (PLFA) and community-level physiological profiles (CLPP) analyses. We found that basal soil respiration (BSR) was significantly higher in the control and newly planted soils than in the second and third year consecutive monoculture soils. However, no significant difference was observed in SIR among the newly planted, second and third year consecutive monoculture soils. The PLFA signatures indicated that the bacterial biomass was larger than the fungal biomass in all four treatments and both enhanced with the increasing years of monoculture and attained the peak in SM. The ratio of cyclopropyl PLFAs to their metabolic precursors (cy/pre), a measure of physiological stress in microbial communities, in the second and third year consecutive monoculture soils was significantly greater than that in the control and newly planted soils. Biolog analysis results revealed that the consumption of carboxylic acids, phenolic acids and amines, especially acid carbon substrates, in the consecutively monocultured soil was significantly greater than that in the newly planted soil. Both PLFA- and CLPP-based principal component analysis (PCA) and cluster analysis revealed the distinct separation between the control, newly planted plots and the second, third year consecutive monoculture plots. Through our PLFA-based and Biolog analysis, together with microbial respiration determination, we were able to reveal characteristic differences in the microbial community composition and activities in the rhizosphere following R. glutinosa monoculture. [Copyright &y& Elsevier]