Biosynthesis Of Secondary Metabolites Research Articles

Recent advances in understanding the regulation of plant secondary metabolite biosynthesis by ethylene-mediated pathways.

Plants produce a large repertoire of secondary metabolites. The pathways that lead to the biosynthesis of these metabolites are majorly conserved in the plant kingdom. However, a significant portion of these metabolites are specific to certain groups or species due to variations in the downstream pathways and evolution of the enzymes. These metabolites show spatiotemporal variation in their accumulation and are of great importance to plants due to their role in development, stress response and survival. A large number of these metabolites are in huge industrial demand due to their potential use as therapeutics, aromatics and more. Ethylene, as a plant hormone is long known, and its biosynthetic process, signaling mechanism and effects on development and response pathways have been characterized in many plants. Through exogenous treatments, ethylene and its inhibitors have been used to manipulate the production of various secondary metabolites. However, the research done on a limited number of plants in the last few years has only started to uncover the mechanisms through which ethylene regulates the accumulation of these metabolites. Often in association with other hormones, ethylene participates in fine-tuning the biosynthesis of the secondary metabolites, and brings specificity in the regulation depending on the plant, organ, tissue type and the prevailing conditions. This review summarizes the related studies, interprets the outcomes, and identifies the gaps that will help to breed better varieties of the related crops and produce high-value secondary metabolites for human benefits.

Transcriptomic and biochemical analyses reveal the antifungal mechanism of cell-free supernatant from Aspergillus luchuensis YZ-1 against Aspergillus flavus

Aspergillus flavus (A. flavus) and its metabolites, aflatoxins, are widespread in crops and foods, threatening public health. Using chemical fungicides to control A. flavus can pose safety hazards and cause environmental pollution. Therefore, there is an urgent need for safe and effective antifungal agents to control A. flavus. In this study, we isolated Aspergillus luchuensisYZ-1 cell-free supernatant (CFS), volatiles, and mycelial extracts to investigate their inhibitory effects on A. flavus. Results indicate that the CFS exhibited greater efficacy in impeding the growth and synthesis of aflatoxin B1 (AFB1) in A. flavus. Transcriptomic analysis revealed a significant downregulation of lipid metabolism, biosynthesis of other secondary metabolites, and glycan metabolism in A. flavus treated with CFS. Approximately 94% of the genes in the aflatoxin synthesis pathway were significantly downregulated. Furthermore, the CFS treatment significantly increased reactive oxygen species (ROS) production in A. flavus, affected its energy metabolism, and upregulated methane metabolism and the pentose phosphate pathway. We then investigated the inhibitory mechanism of YZ-1 against A. flavus by biochemical analysis. It was further confirmed that CFS had destructive effects in the cell wall and cell membrane of A. flavus, as indicated by leakage of nucleic acids, proteins, alkaline phosphatase (AKP), as well as decreased chitin and ergosterol content. In summary, this study provides novel insights into the mechanisms of CFS against A. flavus, suggesting its potential as a biocontrol agent in food and agriculture.

Complex Interplay of Metabolic Pathways in Grafting of Ziziphus Species: Transcriptomic Insights into Regulatory Networks of Carbohydrates and Secondary Metabolite Biosynthesis

Grafting serves as a pervasive methodology in the propagation of jujube plants, yet the nuanced molecular mechanisms that dictate rootstock-scion interactions remain inadequately understood. We examined the transcriptomic landscapes of jujube heterograft combinations. Contrary to self-grafting conditions, early-stage heterografting yielded no discernible advantageous effects on scion biomass accretion. Interestingly, the rootstock’s biomass was significantly impacted by the scion, varying by species. The differentially expressed genes (DEGs) across graft combinations were mainly enriched for the vegetative growth of rootstocks, secondary metabolism, and resistance improvement of scions. Weighted gene co-expression network analysis (WGCNA) identified 27 hub genes which were negatively correlated with plant growth and biomass enlargement, serving as negative regulators, while the genes, L484_001734, ATHB-15, and BPC1, were involved in positive regulation. With biomass measurements, the transcriptomic data supported that an incomplete vascular recovery during early grafting led to nutrient accumulation at the graft junction, temporarily limiting plant growth while providing development resources for callus. In summary, our work has demonstrated that the intricate biological connections between the rootstock and scion guarantee the effective jujube grafting process by elucidating the molecular processes involved in the process.

Metabolomics Characterization of Phenolic Compounds in Colored Quinoa and Their Relationship with In Vitro Antioxidant and Hypoglycemic Activities.

Chenopodium quinoa Willd. is rich in phenolic compounds and exhibits diverse biological activities. Few studies have focused on the effect of colored quinoa's phenolic profile on potential biological activity. This study used a UPLC-MS/MS-based metabolomic approach to examine the quinoa phenolics and their association with in vitro antioxidant and hypoglycemic properties. In total, 430 polyphenols, mainly phenolic acids, flavonoids, and flavonols, were identified. Additionally, 121, 116, and 148 differential polyphenols were found between the white and black, white and red, and black and red comparison groups, respectively; 67 polyphenols were screened as shared key differential metabolites. Phenylalanine, tyrosine, and the biosynthesis of plant secondary metabolites were the main differently regulated pathways. Black quinoa had better total phenolic contents (643.68 mg/100 g DW) and antioxidant capacity, while white quinoa had better total flavonoid contents (90.95 mg/100 g DW) and in vitro α-amylase (IC50 value of 3.97 mg/mL) and α-glucosidase (IC50 value of 1.08 mg/mL) inhibition activities. Thirty-six polyphenols, including epicatechin and linarin, etc., were highly correlated with in vitro antioxidant activity, while six polyphenols, including tiliroside and chrysoeriol, etc., were highly correlated with in vitro hypoglycemic activity. This study may provide important information for colored quinoa resources to develop their healthy food applications.

The landscape and predicted roles of structural variants in Fusarium graminearum genomes.

Structural rearrangements, such as inversions, translocations, duplications, and large insertions and deletions, are large-scale genomic variants that can play an important role in shaping phenotypic variation and in genome adaptation and evolution. We used chromosomal-level assemblies from eight Fusarium graminearum isolates to study structural variants and their role in fungal evolution. We generated the assemblies of four of these genomes after Oxford Nanopore sequencing. A total of 87 inversions, 159 translocations, 245 duplications, 58,489 insertions, and 34,102 deletions were detected. Regions of high recombination rate are associated with structural rearrangements, and a significant proportion of inversions, translocations, and duplications overlap with the repeat content of the genome, suggesting recombination and repeat elements are major factors in the origin of structural rearrangements in F. graminearum. Large insertions and deletions introduce presence-absence polymorphisms for many genes, including secondary metabolite biosynthesis cluster genes and predicted effectors genes. Translocation events were found to be shuffling predicted effector-rich regions of the genomes and are likely contributing to the gain and loss of effectors facilitated by recombination. Breakpoints of some structural rearrangements fall within coding sequences and are likely altering the protein products. Structural rearrangements in F. graminearum thus have an important role to play in shaping pathogen-host interactions and broader evolution through genome reorganization, the introduction of presence-absence polymorphisms, and changing protein products and gene regulation.

Multiple omics revealed the growth-promoting mechanism of Bacillus velezensis strains on ramie.

Beneficial bacteria that promote plant growth can shield plants from negative effects. Yet, the specific biological processes that drive the relationships between soil microbes and plant metabolism are still not fully understood. To investigate this further, we utilized a combination of microbiology and non-targeted metabolomics techniques to analyze the impact of plant growth-promoting bacteria on both the soil microbial communities and the metabolic functions within ramie (Boehmeria nivea) tissues. The findings indicated that the yield and traits of ramie plants are enhanced after treatment with Bacillus velezensis (B. velezensis). These B. velezensis strains exhibit a range of plant growth-promoting properties, including phosphate solubilization and ammonia production. Furthermore, strain YS1 also demonstrates characteristics of IAA production. The presence of B. velezensis resulted in a decrease in soil bacteria diversity, resulting in significant changes in the overall structure and composition of soil bacteria communities. Metabolomics showed that B. velezensis significantly altered the ramie metabolite spectrum, and the differential metabolites were notably enriched (P < 0.05) in five main metabolic pathways: lipid metabolism, nucleotide metabolism, amino acid metabolism, plant secondary metabolites biosynthesis, and plant hormones biosynthesis. Seven common differential metabolites were identified. Correlation analysis showed that the microorganisms were closely related to metabolite accumulation and yield index. In the B. velezensis YS1 and B. velezensis Y4-6-1 treatment groups, the relative abundances of BIrii41 and Bauldia were significantly positively correlated with sphingosine, 9,10,13-TriHOME, fresh weight, and root weight, indicating that these microorganisms regulate the formation of various metabolites, promoting the growth and development of ramie. Conclusively, B. velezensis (particularly YS1) played an important role in regulating soil microbial structure and promoting plant metabolism, growth, and development. The application of the four types of bacteria in promoting ramie growth provides a good basis for future application of biological fertilizers and bio-accelerators.

BnAP2-12 overexpression delays ramie flowering: evidence from AP2/ERF gene expression.

The APETALA2/ethylene response factor (AP2/ERF) superfamily plays a significant role in regulating plant gene expression in response to growth and development. To date, there have been no studies into whether the ramie AP2/ERF genes are involved in the regulation of flower development. Here, 84 BnAP2/ERF members were identified from the ramie genome database, and various bioinformatics data on the AP2/ERF gene family, structure, replication, promoters and regulatory networks were analysed. BnAP2-12 was transferred into Arabidopsis through the flower-dipping method. Phylogenetic analysis classified the 84 BnAP2/ERF members into four subfamilies: AP2 (18), RAV (3), ERF (42), and DREB (21). The functional domain analysis of genes revealed 10 conserved motifs. Genetic mapping localised the 84 members on 14 chromosomes, among which chromosomes 1, 3, 5, and 8 had more members. Collinearity analysis revealed that 43.37% possibly resulted from replication events during the evolution of the ramie genome. Promoter sequence analysis identified classified cis-acting elements associated with plant growth and development, and responses to stress, hormones, and light. Transcriptomic comparison identified 3,635 differentially expressed genes (DEGs) between male and female flowers (1,803 and 1,832 upregulated and downregulated genes, respectively). Kyoto Encyclopaedia of Genes and Genomes pathway analysis categorised DEGs involved in metabolic pathways and biosynthesis of secondary metabolites. Gene Ontology enrichment analysis further identified enriched genes associated with pollen and female gamete formations. Of the 84 BnAP2/ERFs genes, 22 and 8 upregulated and downregulated genes, respectively, were present in female flowers. Co-expression network analysis identified AP2/ERF members associated with flower development, including BnAP2-12. Subcellular localisation analysis showed that the BnAP2-12 protein is localised in the nucleus and cell membrane. Overexpression BnAP2-12 delayed the flowering time of Arabidopsis thaliana. These findings provide insights into the mechanism of ramie flower development.

VvWRKY5 positively regulates wounding-induced anthocyanin accumulation in grape by interplaying with VvMYBA1 and promoting jasmonic acid biosynthesis.

Wounding stress induces the biosynthesis of various secondary metabolites in plants, including anthocyanin. However, the underlying molecular mechanism remains elusive. Here, we reported that a transcription factor, VvWRKY5, promotes wounding-induced anthocyanin accumulation in grape (Vitis vinifera). Biochemical and molecular analyses demonstrated that wounding stress significantly increased anthocyanin content, and VvMYBA1 plays an essential role in this process. VvWRKY5 could interact with VvMYBA1 and amplify the activation effect of VvMYBA1 on its target gene VvUFGT. The transcript level of VvWRKY5 was notably induced by wounding treatment. Moreover, our data demonstrated that VvWRKY5 could promote the synthesis of jasmonic acid (JA), a phytohormone that acts as a positive modulator in anthocyanin accumulation, by directly binding to the W-box element in the promoter of the JA biosynthesis-related gene VvLOX and enhancing its activities, and this activation was greatly enhanced by the VvWRKY5-VvMYBA1 protein complex. Collectively, our findings show that VvWRKY5 plays crucial roles in wounding-induced anthocyanin synthesis in grape and elucidates the transcriptional regulatory mechanism of wounding-induced anthocyanin accumulation.

Soil Application of Bacillus subtilis Regulates Flavonoid and Alkaloids Biosynthesis in Mulberry Leaves.

Flavonoids and alkaloids are the major active ingredients in mulberry leaves that have outstanding medicinal value. Bacillus subtilis can effectively activate the plants defense response and regulate the plant secondary metabolism. In this study, we explored the effects of soil application of B. subtilis on the content of flavonoids and the most important alkaloids (1-deoxynojirimycin, DNJ) in mulberry leaves. Significant decreases in flavonoid content were observed in tender leaves and mature leaves after treatment with B. subtilis; at the same time, significant increases in DNJ content were observed in tender leaves. Based on widely targeted LC-MS/MS and high-throughput approaches, we screened out 904 differentially synthesized metabolites (DSMs) and 9715 differentially expressed genes (DEGs). KEGG analyses showed that these DSMs and DEGs were both significantly enriched in the biosynthesis of secondary metabolites, flavonoid synthesis and plant hormone signal transduction. Further correlation analysis of DEMs and DEGs showed that 40 key genes were involved in flavonoid biosynthesis, with 6 key genes involved in DNJ biosynthesis. The expression of CHS, CHI, F3H, F3'H, FLS, UGT and AOC significantly responded to B. subtilis soil application. This study broadens our understanding of the molecular mechanisms underlying the accumulation of flavonoids and alkaloids in mulberry leaves.

Transcriptome analysis provides insights into preservation mechanism of postharvest Muscat Hamburg grapes treated with SO2

Sulfur dioxide (SO2) is the most frequently used preservative for table grapes, yet the preservation mechanism remains unclear. To ascertain the specific genes and pathways involved in SO2 preservation, RNA-seq technology was employed to characterize the transcriptome profile of grapes during postharvest storage. A total of 22,288 genes were identified, of which 377 genes were differentially expressed (≥ 2-fold change) between SO2 and control groups, mainly enriched in secondary metabolism, plant-pathogen interactions, plant hormone signaling, etc. Numerous genes encoding pathogenesis-related (PR) proteins exhibited higher expression levels in SO2 group, while the activities of disease-resistant enzymes, such as β-1,3-glucanase (PR2) and chitinase (PR3) significantly increased by 28.9 % and 29.3 % (P < 0.05), indicating SO2-induced plant resistance. Exposure to SO2 also enhanced the expression of genes encoding essential enzymes of secondary metabolism, and significantly increased both the activities of critical enzymes for the biosynthesis of secondary metabolites, such as phenylalanine ammonia-lyase (PAL) and 4-coumarate-CoA ligase (4CL), and the contents of secondary metabolites such as total phenol, flavonoid, anthocyanin, and lignin, demonstrating an enhanced chemical and physical barriers in SO2-fumigated grapes. Meanwhile, the genes associated with ethylene signaling and cell wall degradation were down-regulated by SO2 preservative, and some ethylene-responsive elements were identified in the promoter regions of cell wall hydrolase genes, suggesting that SO2-inhibited ethylene signaling might contribute to maintaining cell wall integrity. Altogether, gene expression patterns and cellular physiological processes were altered in grapes exposed to SO2, which helped maintain fruit quality and prolong postharvest life by regulating the defense responses and fruit ageing.

Integrated Transcriptome and Proteome Analysis Reveals the Regulatory Mechanism of Root Growth by Protein Disulfide Isomerase in Arabidopsis.

Protein disulfide isomerase (PDI, EC 5.3.4.1) is a thiol-disulfide oxidoreductase that plays a crucial role in catalyzing the oxidation and rearrangement of disulfides in substrate proteins. In plants, PDI is primarily involved in regulating seed germination and development, facilitating the oxidative folding of storage proteins in the endosperm, and also contributing to the formation of pollen. However, the role of PDI in root growth has not been previously studied. This research investigated the impact of PDI gene deficiency in plants by using 16F16 [2-(2-Chloroacetyl)-2,3,4,9-tetrahydro-1-methyl-1H-pyrido[3,4-b]indole-1-carboxylic acid methyl ester], a small-molecule inhibitor of PDI, to remove functional redundancy. The results showed that the growth of Arabidopsis roots was significantly inhibited when treated with 16F16. To further investigate the effects of 16F16 treatment, we conducted expression profiling of treated roots using RNA sequencing and a Tandem Mass Tag (TMT)-based quantitative proteomics approach at both the transcriptomic and proteomic levels. Our analysis revealed 994 differentially expressed genes (DEGs) at the transcript level, which were predominantly enriched in pathways associated with "phenylpropane biosynthesis", "plant hormone signal transduction", "plant-pathogen interaction" and "starch and sucrose metabolism" pathways. Additionally, we identified 120 differentially expressed proteins (DEPs) at the protein level. These proteins were mainly enriched in pathways such as "phenylpropanoid biosynthesis", "photosynthesis", "biosynthesis of various plant secondary metabolites", and "biosynthesis of secondary metabolites" pathways. The comprehensive transcriptome and proteome analyses revealed a regulatory network for root shortening in Arabidopsis seedlings under 16F16 treatment, mainly involving phenylpropane biosynthesis and plant hormone signal transduction pathways. This study enhances our understanding of the significant role of PDIs in Arabidopsis root growth and provides insights into the regulatory mechanisms of root shortening following 16F16 treatment.

Transcriptome analysis of Bipolaris sorokiniana - Hordeum vulgare provides insights into mechanisms of host-pathogen interaction.

Spot blotch disease incited by Bipolaris sorokiniana severely affects the cultivation of barley. The resistance to B. sorokiniana is quantitative in nature and its interaction with the host is highly complex which necessitates in-depth molecular analysis. Thus, the study aimed to conduct the transcriptome analysis to decipher the mechanisms and pathways involved in interactions between barley and B. sorokiniana in both the resistant (EC0328964) and susceptible (EC0578292) genotypes using the RNA Seq approach. In the resistant genotype, 6,283 genes of Hordeum vulgare were differentially expressed out of which 5,567 genes were upregulated and 716 genes were downregulated. 1,158 genes of Hordeum vulgare were differentially expressed in the susceptible genotype, out of which 654 genes were upregulated and 504 genes were downregulated. Several defense-related genes like resistant gene analogs (RGAs), disease resistance protein RPM1, pathogenesis-related protein PRB1-2-like, pathogenesis-related protein 1, thaumatin-like protein PWIR2 and defensin Tm-AMP-D1.2 were highly expressed exclusively in resistant genotype only. The pathways involved in the metabolism and biosynthesis of secondary metabolites were the most prominently represented pathways in both the resistant and susceptible genotypes. However, pathways involved in MAPK signaling, plant-pathogen interaction, and plant hormone signal transduction were highly enriched in resistant genotype. Further, a higher number of pathogenicity genes of B. sorokiniana was found in response to the susceptible genotype. The pathways encoding for metabolism, biosynthesis of secondary metabolites, ABC transporters, and ubiquitin-mediated proteolysis were highly expressed in susceptible genotype in response to the pathogen. 14 and 11 genes of B. sorokiniana were identified as candidate effectors from susceptible and resistant host backgrounds, respectively. This investigation will offer valuable insights in unraveling the complex mechanisms involved in barley- B. sorokiniana interaction.

The evolution of the gliotoxin biosynthetic gene cluster in Penicillium fungi.

Fungi biosynthesize diverse secondary metabolites, small organic bioactive molecules with key roles in fungal ecology. Fungal secondary metabolites are often encoded by physically clustered genes known as biosynthetic gene clusters (BGCs). Fungi in the genus Penicillium produce a cadre of secondary metabolites, some of which are useful (e.g. the antibiotic penicillin and the cholesterol-lowering drug mevastatin) and others harmful (e.g. the mycotoxin patulin and the immunosuppressant gliotoxin) to human affairs. Fungal genomes often also encode resistance genes that confer protection against toxic secondary metabolites. Some Penicillium species, such as Penicillium decumbens, are known to produce gliotoxin, a secondary metabolite with known immunosuppressant activity. To investigate the evolutionary conservation of homologs of the gliotoxin BGC and of genes involved in gliotoxin resistance in Penicillium, we analyzed 35 Penicillium genomes from 23 species. Homologous, lesser fragmented gliotoxin BGCs were found in 12 genomes, mostly fragmented remnants of the gliotoxin BGC were found in 21 genomes, whereas the remaining 2 Penicillium genomes lacked the gliotoxin BGC altogether. In contrast, broad conservation of homologs of resistance genes that reside outside the BGC across Penicillium genomes was observed. Evolutionary rate analysis revealed that BGCs with higher numbers of genes evolve slower than BGCs with few genes, suggestive of constraint and potential functional significance or more recent decay. Gene tree-species tree reconciliation analyses suggested that the history of homologs in the gliotoxin BGC across the genus Penicillium likely involved multiple duplications, losses, and horizontal gene transfers. Our analyses suggest that genes encoded in BGCs can have complex evolutionary histories and be retained in genomes long after the loss of secondary metabolite biosynthesis.

De Novo Transcriptome Analysis by PacBio SMRT-Seq and Illumina RNA-Seq Provides New Insights into Polyphenol Biosynthesis in Chinese Olive Fruit

Polyphenols play a crucial role in fruit flavor. To elucidate the mechanism of fruit polyphenol metabolism, we constructed a transcriptome atlas through PacBio single-molecule real-time (SMRT) sequencing and Illumina next-generation sequencing (NGS) using Canarium album (Lour.) Raeusch., which is a fantastic fruit rich in polyphenolic compounds. In this work, PacBio full-length transcriptome assembly generated 135,439 isoforms with an average length of all isoforms of 2687.94 bp and an N50 length of 3224 bp. To gain deeper insights into the molecular mechanisms of polyphenol biosynthesis in C. album, we constructed twelve RNA-Seq libraries from four developmental stages of the fruits. We identified a total of 28,658 differentially expressed genes (DEGs). We found that many DEGs were involved in metabolic pathways, biosynthesis of secondary metabolites, biosynthesis of antibiotics, starch and sucrose metabolism, and plant hormone signal transduction. Here, we report the expression profiles of 215 DEGs encoding 27 enzymes involved in the polyphenol biosynthesis pathway in C. album. In addition, 285 differentially expressed transcription factors (TFs) continuously down-regulated in four developmental periods of C. album fruit, which may indicate their potential role in the response to polyphenol metabolism and phenylpropanoid biosynthesis pathways. This report will help us understand polyphenol biosynthesis’s functions and metabolic mechanism in C. album. The transcriptome data provide a valuable resource for genetic and genomics research. They will facilitate future work exploiting C. album and other fruits used as medicine and food.

Chromosome-level genome assembly provides insights into the genome evolution and functional importance of the phenylpropanoid–flavonoid pathway in Thymus mongolicus

BackgroundThymus mongolicus (family Lamiaceae) is a Thyme subshrub with strong aroma and remarkable environmental adaptability. Limited genomic information limits the use of this plant.ResultsChromosome-level 605.2 Mb genome of T. mongolicus was generated, with 96.28% anchored to 12 pseudochromosomes. The repetitive sequences were dominant, accounting for 70.98%, and 32,593 protein-coding genes were predicted. Synteny analysis revealed that Lamiaceae species generally underwent two rounds of whole genome duplication; moreover, species-specific genome duplication was identified. A recent LTR retrotransposon burst and tandem duplication might play important roles in the formation of the Thymus genome. Using comparative genomic analysis, phylogenetic tree of seven Lamiaceae species was constructed, which revealed that Thyme plants evolved recently in the family. Under the phylogenetic framework, we performed functional enrichment analysis of the genes on nodes that contained the most gene duplication events (> 50% support) and of relevant significant expanded gene families. These genes were highly associated with environmental adaptation and biosynthesis of secondary metabolites. Combined transcriptome and metabolome analyses revealed that Peroxidases, Hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferases, and 4-coumarate-CoA ligases genes were the essential regulators of the phenylpropanoid–flavonoid pathway. Their catalytic products (e.g., apigenin, naringenin chalcone, and several apigenin-related compounds) might be responsible for the environmental tolerance and aromatic properties of T. mongolicus.ConclusionThis study enhanced the understanding of the genomic evolution of T. mongolicus, enabling further exploration of its unique traits and applications, and contributed to the understanding of Lamiaceae genomics and evolutionary biology.

Comprehensive genomic analysis of Bacillus subtilis and Bacillus paralicheniformis associated with the pearl millet panicle reveals their antimicrobial potential against important plant pathogens

BackgroundPlant microbiome confers versatile functional roles to enhance survival fitness as well as productivity. In the present study two pearl millet panicle microbiome member species Bacillus subtilis PBs 12 and Bacillus paralicheniformis PBl 36 found to have beneficial traits including plant growth promotion and broad-spectrum antifungal activity towards taxonomically diverse plant pathogens. Understanding the genomes will assist in devising a bioformulation for crop protection while exploiting their beneficial functional roles.ResultsTwo potential firmicute species were isolated from pearl millet panicles. Morphological, biochemical, and molecular characterization revealed their identities as Bacillus subtilis PBs 12 and Bacillus paralicheniformis PBl 36. The seed priming assays revealed the ability of both species to enhance plant growth promotion and seedling vigour index. Invitro assays with PBs 12 and PBl 36 showed the antibiosis effect against taxonomically diverse plant pathogens (Magnaporthe grisea; Sclerotium rolfsii; Fusarium solani; Alternaria alternata; Ganoderma sp.) of crops and multipurpose tree species. The whole genome sequence analysis was performed to unveil the genetic potential of these bacteria for plant protection. The complete genomes of PBs 12 and PBl 36 consist of a single circular chromosome with a size of 4.02 and 4.33 Mb and 4,171 and 4,606 genes, with a G + C content of 43.68 and 45.83%, respectively. Comparative Average Nucleotide Identity (ANI) analysis revealed a close similarity of PBs 12 and PBl 36 with other beneficial strains of B. subtilis and B. paralicheniformis and found distant from B. altitudinis, B. amyloliquefaciens, and B. thuringiensis. Functional annotation revealed a majority of pathway classes of PBs 12 (30) and PBl 36 (29) involved in the biosynthesis of secondary metabolites, polyketides, and non-ribosomal peptides, followed by xenobiotic biodegradation and metabolism (21). Furthermore, 14 genomic regions of PBs 12 and 15 of PBl 36 associated with the synthesis of RiPP (Ribosomally synthesized and post-translationally modified peptides), terpenes, cyclic dipeptides (CDPs), type III polyketide synthases (T3PKSs), sactipeptides, lanthipeptides, siderophores, NRPS (Non-Ribosomal Peptide Synthetase), NRP-metallophone, etc. It was discovered that these areas contain between 25,458 and 33,000 secondary metabolite-coding MiBiG clusters which code for a wide range of products, such as antibiotics. The PCR-based screening for the presence of antimicrobial peptide (cyclic lipopeptide) genes in PBs 12 and 36 confirmed their broad-spectrum antifungal potential with the presence of spoVG, bacA, and srfAA AMP genes, which encode antimicrobial compounds such as subtilin, bacylisin, and surfactin.ConclusionThe combined in vitro studies and genome analysis highlighted the antifungal potential of pearl millet panicle-associated Bacillus subtilis PBs12 and Bacillus paralicheniformis PBl36. The genetic ability to synthesize several antimicrobial compounds indicated the industrial value of PBs 12 and PBl 36, which shed light on further studies to establish their action as a biostimulant for crop protection.

Influence of Bacillus subtilis strain Z-14 on microbial ecology of cucumber rhizospheric vermiculite infested with fusarium oxysporum f. sp. cucumerinum

Fusarium oxysporum (FO) is a typical soil-borne pathogenic fungus, and the cucumber wilt disease caused by F. oxysporum f. sp. cucumerinum (FOC) seriously affects crop yield and quality. Vermiculite is increasingly being used as a culture substrate; nevertheless, studies exploring the effectiveness and mechanisms of biocontrol bacteria in this substrate are limited. In this study, vermiculite was used as a culture substrate to investigate the control effect of Bacillus subtilis strain Z-14 on cucumber wilt and the rhizospheric microecology, focusing on colonization ability, soil microbial diversity, and rhizosphere metabolome. Pot experiments showed that Z-14 effectively colonized the cucumber roots, achieving a controlled efficacy of 61.32% for wilt disease. It significantly increased the abundance of Bacillus and the expression of NRPS and PKS genes, while reducing the abundance of FO in the rhizosphere. Microbial diversity sequencing showed that Z-14 reduced the richness and diversity of the rhizosphere bacterial community, increased the richness and diversity of the fungal community, and alleviated the effect of FO on the community structure of the cucumber rhizosphere. The metabolomics analysis revealed that Z-14 affected ABC transporters, amino acid synthesis, and the biosynthesis of plant secondary metabolites. Additionally, Z-14 increased the contents of phenylacetic acid, capsidol, and quinolinic acid, all of which were related to the antagonistic activity in the rhizosphere. Z-14 exhibited a significant control effect on cucumber wilt and influenced the microflora and metabolites in rhizospheric vermiculite, providing a theoretical basis for further understanding the control effect and mechanism of cucumber wilt in different culture substrates.

Transcriptomic Analysis for Diurnal Temperature Differences Reveals Gene-Regulation-Network Response to Accumulation of Bioactive Ingredients of Protocorm-like Bodies in Dendrobium officinale.

Dendrobium officinale Kimura et Migo (D. officinale) is one of the most important traditional Chinese medicinal herbs, celebrated for its abundant bioactive ingredients. This study demonstrated that the diurnal temperature difference (DIF) (T1: 13/13 °C, T2: 25/13 °C, and T3: 25/25 °C) was more favorable for high chlorophyll, increased polysaccharide, and total flavonoid contents compared to constant temperature treatments in D. officinale PLBs. The transcriptome analysis revealed 4251, 4404, and 4536 differentially expressed genes (DEGs) in three different comparisons (A: 25/13 °C vs. 13/13 °C, B: 13/13 °C vs. 25/25 °C, and C: 25/13 °C vs. 25/25 °C, respectively). The corresponding up-/down-regulated DEGs were 1562/2689, 2825/1579, and 2310/2226, respectively. GO and KEGG enrichment analyses of DEGs showed that the pathways of biosynthesis of secondary metabolites, carotenoid biosynthesis, and flavonoid biosynthesis were enriched in the top 20; further analysis of the sugar- and flavonol-metabolism pathways in D. officinale PLBs revealed that the DIF led to a differential gene expression in the enzymes linked to sugar metabolism, as well as to flavonol metabolism. Certain key metabolic genes related to ingredient accumulation were identified, including those involved in polysaccharide metabolism (SUS, SUT, HKL1, HGL, AMY1, and SS3) and flavonol (UGT73C and UGT73D) metabolism. Therefore, these findings indicated that these genes may play an important role in the regulatory network of the DIF in the functional metabolites of D. officinale PLBs. In a MapMan annotation of abiotic stress pathways, the DEGs with significant changes in their expression levels were mainly concentrated in the heat-stress pathways, including heat-shock proteins (HSPs) and heat-shock transcription factors (HSFs). In particular, the expression levels of HSP18.2, HSP70, and HSF1 were significantly increased under DIF treatment, which suggested that HSF1, HSP70 and HSP18.2 may respond to the DIF. In addition, they can be used as candidate genes to study the effect of the DIF on the PLBs of D. officinale. The results of our qPCR analysis are consistent with those of the transcriptome-expression analysis, indicating the reliability of the sequencing. The results of this study revealed the transcriptome mechanism of the DIF on the accumulation of the functional metabolic components of D. officinale. Furthermore, they also provide an important theoretical basis for improving the quality of D. officinale via the DIF in production.

Secondary metabolite gene clusters from the phytopathogenic fungus Gaeumannomyces tritici

The take-all disease is one of the most important maladies in cereals and grasses, being caused by the fungus Gaeumannomyces tritici. Secondary metabolites are known to perform critical functions during the infection process of various phytopathogens. However, the current understanding of the biosynthesis of secondary metabolites in G. tritici is limited. Similarly, comprehensive analyses of the expression, conservation, and evolution of these biosynthesis-related genes are crucial for enhancing our knowledge of the molecular mechanisms that drive the development of the take-all disease. Here we have performed a deep survey and description of secondary metabolite biosynthetic gene clusters in G. tritici, analyzed a previously published RNA-seq of a mimicked infection condition, and assessed the conservation among 10 different Magnaporthales order members. Notably, the majority of the 35 putative gene clusters identified were conserved among these species, with GtPKS1, GtPKS3, and GtTERP4 uniquely identified in G. tritici. In the mimicked infection condition, seven gene clusters, including the GtPKS1 cluster, exhibited upregulated expression. Through comparative genomic analysis, GtPKS1 was associated with the production of dichlorodiaporthin, a metabolite with cytotoxic and antifungal activity. In addition, GtPKS10 and GtPKSNRPS3 showed similarities to already characterized biosynthetic pathways involved in the synthesis of ACR-toxin (phytotoxic) and trichosetin (phytotoxic and antibiotic), respectively. These three gene clusters were further scrutinized through phylogenetic inference, which revealed the distribution of orthologous sequences across various plant-associated fungi. Finally, the detailed identification of several genes enrolled in secondary metabolite biosynthesis provides the foundation for future in-depth research, supporting the potential impact of several small molecules on G. tritici lifecycle and host interactions.

The roles of SARP family regulators involved in secondary metabolism in Streptomyces.

Streptomyces species are best known for their ability to produce abundant secondary metabolites with versatile bioactivities and industrial importance. These metabolites are usually biosynthesized through metabolic pathways encoded by cluster-situated genes. These genes are also known as biosynthetic gene clusters (BGCs) of secondary metabolites. The expression of BGCs is intricately controlled by pyramidal transcriptional regulatory cascades, which include various regulators. Streptomyces antibiotic regulatory proteins (SARPs), a genus-specific family of regulators, are widely distributed and play important roles in regulating the biosynthesis of secondary metabolites in Streptomyces. Over the past decade, the biological functions of SARPs have been extensively investigated. Here, we summarized the recent advances in characterizing the roles of SARPs involved in Streptomyces secondary metabolism from the following three aspects. First, the classification and domain organization of SARPs were summarized according to their size variation. Second, we presented a detailed description of the regulatory mechanisms and modes of action of SARPs involved in secondary metabolism. Finally, the biotechnological application of SARPs was illustrated by improving the production of target secondary metabolites and discovering novel bioactive natural products. This review will help researchers to comprehensively understand the roles of SARPs in secondary metabolite biosynthesis in Streptomyces, which will contribute to building a solid foundation for their future application in synthetic biology.