Genome-wide characterization of the bHLH gene family in Gynostemma pentaphyllum reveals its potential role in the regulation of gypenoside biosynthesis.

BACKGROUND: Gynostemma pentaphyllum, an ancient Chinese herbal medicine, serves as a natural source of gypenosides with significant medicinal properties. Basic helix-loop-helix (bHLH) transcription factors play pivotal roles in numerous biological processes, especially in the regulation of secondary metabolism in plants. However, the characteristics and functions of the bHLH genes in G. pentaphyllum remain unexplored, and their regulatory role in gypenoside biosynthesis remains poorly elucidated. RESULTS: This study identified a total of 111 bHLH members in G. pentaphyllum (GpbHLHs), categorizing them into 26 subgroups based on shared conserved motif compositions and gene structures. Collinearity analysis illustrated that segmental duplications predominately lead to the evolution of GpbHLHs, with most duplicated GpbHLH gene pairs undergoing purifying selection. Among the nine gypenoside-related GpbHLH genes, two GpbHLHs (GpbHLH15 and GpbHLH58) were selected for further investigation based on co-expression analysis and functional prediction. The expression of these two selected GpbHLHs was dramatically induced by methyl jasmonate, and their nuclear localization was confirmed. Furthermore, yeast one-hybrid and dual-luciferase assays demonstrated that GpbHLH15 and GpbHLH58 could bind to the promoters of the gypenoside biosynthesis pathway genes, such as GpFPS1, GpSS1, and GpOSC1, and activate their promoter activity to varying degrees. CONCLUSIONS: In conclusion, our findings provide a detailed analysis of the bHLH family and valuable insights into the potential use of GpbHLHs to enhance the accumulation of gypenosides in G. pentaphyllum.

Integrated transcriptomics and metabolomics analysis provides insights into aromatic volatiles formation in Cinnamomum cassia bark at different harvesting times.

BACKGROUND: Cinnamomum cassia Presl, classified in the Lauraceae family, is widely used as a spice, but also in medicine, cosmetics, and food. Aroma is an important factor affecting the medicinal and flavoring properties of C. cassia, and is mainly determined by volatile organic compounds (VOCs); however, little is known about the composition of aromatic VOCs in C. cassia and their potential molecular regulatory mechanisms. Here, integrated transcriptomic and volatile metabolomic analyses were employed to provide insights into the formation regularity of aromatic VOCs in C. cassia bark at five different harvesting times. RESULTS: The bark thickness and volatile oil content were significantly increased along with the development of the bark. A total of 724 differentially accumulated volatiles (DAVs) were identified in the bark samples, most of which were terpenoids. Venn analysis of the top 100 VOCs in each period showed that twenty-eight aromatic VOCs were significantly accumulated in different harvesting times. The most abundant VOC, cinnamaldehyde, peaked at 120 months after planting (MAP) and dominated the aroma qualities. Five terpenoids, α-copaene, ß-bourbonene, α-cubebene, α-funebrene, and δ-cadinene, that peaked at 240 MAP could also be important in creating C. cassia's characteristic aroma. A list of 43,412 differentially expressed genes (DEGs) involved in the biosynthetic pathways of aromatic VOCs were identified, including phenylpropanoids, mevalonic acid (MVA) and methylerythritol phosphate (MEP). A gene-metabolite regulatory network for terpenoid and phenylpropanoid metabolism was constructed to show the key candidate structural genes and transcription factors involved in the biosynthesis of terpenoids and phenylpropanoids. CONCLUSIONS: The results of our research revealed the composition and changes of aromatic VOCs in C. cassia bark at different harvesting stages, differentiated the characteristic aroma components of cinnamon, and illuminated the molecular mechanism of aroma formation. These foundational results will provide technical guidance for the quality breeding of C. cassia.

Metabolomics analysis reveals the differences between Abrus cantoniensis Hance and Abrus mollis Hance.

BACKGROUND: Abrus cantoniensis Hance. (Ac) and Abrus mollis (Am), two edible and medicinal plants with economic value in southern China, belong to the Abrus genus. Due to its growth characteristics, Am often replaces Ac in folk medicine. However, the latest National Pharmacopeia of China only recommends Ac. The differences in the metabolite composition of the plants are directly related to the differences in their clinical efficacy. RESULTS: The difference in metabolites were analyzed using an untargeted metabolomic approach based on ultrahigh-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC‒ESI‒MS/MS). The roots (R), stems (S) and leaves (L) of the two varieties were examined, and 635 metabolites belonging to 8 classes were detected. A comparative study revealed clear variations in the metabolic profiles of the two plants, and the AmR group had more active ingredients (flavonoids and terpenoids) than the AcR group. The metabolites classified as flavonoids and triterpene saponins showed considerable variations among the various samples. Both Ac and Am had unique metabolites. Two metabolites (isovitexin-2''-xyloside and soyasaponin V) specifically belong to Ac, and nine metabolites (vitexin-2"-O-galactoside, ethyl salicylate, 6-acetamidohexanoic acid, rhein-8-O-glucoside, hederagenin-3-O-glucuronide-28-O-glucosyl(1,2)-glucoside, methyl dioxindole-3-acetate, veratric acid, isorhamnetin-3-O-sophoroside-7-O-rhamnoside, and isorhamnetin-3-O-sophoroside) specifically belong to Am. CONCLUSIONS: The metabolite differences between Ac and Am cause the differences in their clinical efficacy. Our findings serve as a foundation for further investigation of biosynthesis pathways and associated bioactivities and provide guidance for the clinical application of traditional Chinese medicine.

Matrine-producing endophytic fungus Galactomyces candidum TRP-7: screening, identification, and fermentation conditions optimization for Matrine production.

Matrine (MA) is an alkaloid extracted from the root of genus Sophora with various pharmacological activities. Production of MA by endophytic fungi offers an alternative challenge to reduce the massive consumption to meet the increasing demand of MA. In the current study, the positive strains with MA producing ability were screened from endophytic fungal isolated from the root of Sophora tonkinensis Gagnep. Chromatographic analyses verified the identity of the produced MA. Among these fungi, Galactomyces candidum strain TRP-7 was the most valuable strain for MA production with the initial yield 8.26 mg L-1. The MA production was efficiently maximized up to 17.57 mg L-1 of fermentation broth, after optimization of eight process parameters using Plackett-Burman and Box-Behnken designs. The statistical optimization resulted in a 1.127 times increase in MA production as compared to the initial yield of TRP-7. This is the first report to isolate endophytic fungi with MA-producing activity from S. tonkinensis Gagnep., and to identify an endophytic fungus G. candidum TRP-7 as a new promising start strain for a higher MA yield.

Characterization of Volatile Organic Compounds Emitted from Endophytic Burkholderia cenocepacia ETR-B22 by SPME-GC-MS and Their Inhibitory Activity against Various Plant Fungal Pathogens.

The use of antagonistic microorganisms and their volatile organic compounds (VOCs) to control plant fungal pathogens is an eco-friendly and promising substitute for chemical fungicides. In this work, endophytic bacterium ETR-B22, isolated from the root of Sophora tonkinensis Gagnep., was found to exhibit strong antagonistic activity against 12 fungal pathogens found in agriculture. Strain ETR-B22 was identified as Burkholderia cenocepacia based on 16S rRNA and recA sequences. We evaluated the antifungal activity of VOCs emitted by ETR-B22. The VOCs from strain ETR-B22 also showed broad-spectrum antifungal activity against 12 fungal pathogens. The composition of the volatile profiles was analyzed based on headspace solid phase microextraction (HS-SPME) gas chromatography coupled to mass spectrometry (GC-MS). Different extraction strategies for the SPME process significantly affected the extraction efficiency of the VOCs. Thirty-two different VOCs were identified. Among the VOC of ETR-B22, dimethyl trisulfide, indole, methyl anthranilate, methyl salicylate, methyl benzoate, benzyl propionate, benzyl acetate, 3,5-di-tert-butylphenol, allyl benzyl ether and nonanoic acid showed broad-spectrum antifungal activity, and are key inhibitory compounds produced by strain ETR-B22 against various fungal pathogens. Our results suggest that the endophytic strain ETR-B22 and its VOCs have high potential for use as biological controls of plant fungal pathogens.

Novel Substituted Thiophenes and Sulf-Polyacetylene Ester from Echinops ritro L.

Three new substituted bithiophenes (1⁻3), and one new sulf-polyacetylene ester, ritroyne A (16) were isolated from the whole plant of Echinops ritro together with twelve known substituted thiophenes. The structures were elucidated on the basis of extensive spectroscopic analysis including 1D and 2D NMR as well as MS. Furthermore, the absolute configuration of ritroyne A (16) was established by computational methods. In bioscreening experiments, four compounds (2, 4, 12, 14) showed similar antibacterial activity against Staphylococcus aureus ATCC 2592 with levofloxacin (8 µg/mL). Five compounds (2, 4, 9, 12, 14) exhibited antibacterial activities against Escherichia coli ATCC 25922, with minimum inhibitory concentration (MIC) values of 32⁻64 µg/mL. Three compounds (2, 4, 12) exhibited antifungal activities against Candida albicans ATCC 2002 with MIC values of 32⁻64 µg/mL. However, compound 16 did not exhibit antimicrobial activities against three microorganisms.

Development and Characterization of High-Throughput EST-Based SSR Markers for Pogostemon cablin Using Transcriptome Sequencing.

Simple sequence repeats (SSRs) or microsatellite markers derived from expressed sequence tags (ESTs) are routinely used for molecular assisted-selection breeding, comparative genomic analysis, and genetic diversity studies. In this study, we investigated 54,546 ESTs for the identification and development of SSR markers in Pogostemon cablin (Patchouli). In total, 1219 SSRs were identified from 1144 SSR-containing ESTs. Trinucleotides (80.8%) were the most abundant SSRs, followed by di- (10.8%), mono- (7.1%), and hexa-nucleotides (1.3%). The top six motifs were CCG/CGG (15.3%), AAG/CTT (15.0%), ACC/GGT (13.5%), AGG/CCT (12.4%), ATC/ATG (9.9%), and AG/CT (9.8%). On the basis of these SSR-containing ESTs, a total of 192 primer pairs were randomly designed and used for polymorphism analysis in 38 accessions collected from different geographical regions of Guangdong, China. Of the SSR markers, 45 were polymorphic and had allele variations from two to four. Furthermore, a transferability analysis of these primer pairs revealed a 10⁻40% cross-species transferability in 10 related species. This report is the first comprehensive study on the development and analysis of a large set of SSR markers in P. cablin. These markers have the potential to be used in quantitative trait loci mapping, genetic diversity studies, and the fingerprinting of cultivars of P. cablin.

Bacteria from nodules of Abrus mollis Hance: genetic diversity and screening of highly efficient growth-promoting strains.

Introduction: Abrus mollis Hance. (AM) is an important species used in southern Chinese medicine. It is mainly found in Guangdong and Guangxi provinces in China, and it is effective in the treatment of hepatitis. Endophytic bacteria are known to affect the growth and quality of medicinal plants. However, there are limited reports describing endophytic bacteria related to AM. Methods: In the present study, Illumina-based 16S rRNA gene sequencing was used to investigate the endophytic bacterial communities of root nodules of AM at five sampling sites in Guangxi. In addition, 179 strains of endophytic bacteria were isolated and categorized into 13 haplotypes based on recA sequence analysis. Results: The phylogeny of the 16S rRNA gene sequences revealed a predominance of nonrhizobial endophytes. Microbial diversity analysis showed that Proteobacteria was the dominant phylum in all samples, while Bradyrhizobium was the dominant genus in different samples. An efficient strain, Rhizobium tropici FM-19, was screened and obtained through greenhouse experiments. The AM plants inoculated with this strain showed the best growth performance and high nitrogen fixation and nodulation capacity. Notably, total phenols and total flavonoids, important active components in AM, increased by 30.9 and 42.7%, respectively, after inoculation with Rhizobium tropici FM-19. Discussion: This study provides insights into the complex microbial diversity of AM nodules and provides strain information for the efficient cultivation of AM.

Evaluation of different drying methods on the quality of Cinnamomum cassia barks by analytic hierarchy process method.

Cinnamomum cassia Presl is a major food spice as well as traditional herbal medicine with anti-inflammatory, analgesic, and stomachic properties, which must be dried to preserve its quality, but mostly by using traditional, ineffective drying method. In order to find a scientific drying method by evaluating different drying methods that could influence the quality of C. cassia, ten indices were employed to evaluate different drying methods in C. cassia using the Analytic Hierarchy Process (AHP) method though calculating the total scores and ranking the priority. Four quality markers (Q-Markers) (coumarin, cinnamyl alcohol, cinnamaldehyde and o-methoxycinnamaldehyde) were isolated from the samples and analyzed by high performance liquid chromatography (HPLC) method under different drying methods. The results showed that various drying methods had multiple effects on the physicochemical qualities, essential oil content, and Q-Marker contents. Compared with other drying methods, oven-drying of 45 °C (45OD) maintained optimal levels of color and aroma, it also significantly shortened the drying time by 225 h than traditionally shade-drying (SHD) method with the drying rate (48.35 %), and obtained the highest essential oil content (3.05 %) and Q-Marker contents (30.23 mg g-1). Furthermore, the ash content (4.22 %) were satisfied with the stipulation of Chinese pharmacopoeia in 45OD samples. Applying AHP allowed us to identify 45OD as the optimal drying method with the highest total score (9.00), followed by the traditional shade-drying (SHD) method (7.88). The present study is the first report to apply the AHP method for quality evaluation of drying processing in C. cassia. It can provide the theoretical basis for evaluating an excellent method for C. cassia drying processing, as well as the rational use of different drying methods to furtherly develop the high quality C. cassia industry.

Genome-wide identification and characterization of the JAZ gene family in Gynostemma pentaphyllum reveals the COI1/JAZ/MYC2 complex potential involved in the regulation of the MeJA-induced gypenoside biosynthesis.

The Jasmonate ZIM domain (JAZ) proteins, functioning as critical suppressors for jasmonic acid (JA) signal transduction in plants, occupy crucial roles in multiple biological processes, particularly in the orchestration of secondary metabolic pathways. However, the mechanism underlying the JA-induced gypenosides accumulation in Gynostemma pentaphyllum remains poorly elucidated. Our research led to the identification of 11 distinct JAZ members in G. pentaphyllum (GpJAZs). According to the classification approach of AtJAZ, we allocated these members into five subgroups that shared similar conserved motif compositions. Subsequently, we identified the presence of various cis-acting elements associated with light stimuli, hormone responses, and stress signals within the promoter regions of the GpJAZ gene family. The expression levels of GpJAZ genes in different tissues were quite different, and the majority of GpJAZ genes exhibited varying degrees of response to methyl jasmonate (MeJA) induction. Yeast two-hybrid (Y2H) assays revealed interactions between GpJAZ1/2/4/5/7/9/10 and GpMYC2, whereas GpCOI1 protein was found to interact with GpJAZ1/2/4/5, thereby forming the COI1/JAZ/MYC2 complex. Furthermore, as an activator of gypenoside metabolic pathway, GpMYC2 could activate the promoter activity of the gypenoside metabolism-related genes to varying degrees by binding to their promoters, indicating that the COI1/JAZ/MYC2 module involved in the MeJA-induced regulation of gypenosides. In summary, our findings present an exhaustive examination of the JAZ gene family, furnishing a significant lead for delving deeper into the molecular mechanisms that drive the MeJA-induced enhancement of gypenosides accumulation in G. pentaphyllum.

Alkaloid variations within the genus Stephania (Menispermaceae) in China.

The genus Stephania, which is rich in alkaloids, has been used as a traditional medicine or folklore herb against numerous ailments in China. However, the understanding of the variation within the genus Stephania is obscure, which limits the optimal utilization of the genus. An evaluation of the variation within the genus Stephania would help screen the ideal Stephania genotypes for drug utilization. In the present study, alkaloids in the tubers of four commonly cultivated Stephania species in China, i.e., the genotype Stephania kwangsiensis Lo. (SK-guangxi) from Guangxi Province and three Stephania yunnanensis H.S. Lo. genotypes (SY-xueteng, SY-hongteng, and SY-lvteng) sourced from Yunnan Province, were investigated, and the genus variations were compared. The results revealed significant variations in the abundance of alkaloids in tubers within the genus Stephania. The Stephania genotypes SY-xueteng and SY-hongteng showed a relatively high abundance of total alkaloids compared with the Stephania genotypes SK-guangxi and SY-lvteng. Specifically, the Stephania genotype SY-xueteng had a relatively high abundance of palmatine in tubers, and the Stephania genotype SY-hongteng exhibited a high abundance of stephanine in tubers. Our study provides foundations for further utilization of ideal Stephania genotypes by clarifying the variations in the alkaloid contents within the genus in China.

Variation in flavonoid and antioxidant activities of Pyrrosia petiolosa (Christ) Ching from different geographic origins.

Pyrrosia petiolosa (Christ) Ching has both medicinal and health benefits in China. The potential antioxidant activities of P. petiolosa, which are mainly attributed to its flavonoids, have attracted much attention in recent years. The present study aimed to determine the concentration of flavonoid components and evaluate the relative antioxidant activities of P. petiolosa from different geographic origins using a UPLC-MRM-MS-based metabolomics approach. In total, 97 flavonoid components were identified, and their concentrations in the samples from different geographic locations showed significant variation. Thirteen flavonoid components were identified as potential biomarkers for distinguishing between the two major regions, Guizhou (GZ) and Guangxi (GX). The GZ group showed higher total flavonoid content, free radical scavenging activities, and ferric reducing antioxidant power. The well positive correlations were found between the antioxidant capacities and some flavonoid markers. The ecogeographic factors, namely altitude and longitude, play a crucial role in the difference of antioxidant activities and flavonoids concentration. These results indicate that P. petiolosa is rich in flavonoid compounds and is a promising source of natural antioxidants, providing a basis for the quality control of P. petiolosa.

[Study on Camellia Sect. Chrysantha Chang species identification by FTIR technology].

FTIR spectra from 16 kinds of Camellia Sect. Chrysantha by Fourier transform infrared spectroscopy (FTIR) method combined with the system clustering and correlation coefficient method were used to analyze and compare these spectral data. The results, show that: Camellia Sect. Chrysantha of 16 kinds were divided into three groups, the first kind was: C. longzhouensis etc, in all eleven kinds; The second kind was: C. achrysantha, C. limonia, C. pingguoensis and C. chuongtsoensis; The third kind was C. microcarpa, which for a class alone. According to the difference in related anatomy and morphology, this study supported that C. longgangensis and C. ptilosperma should be incorporated into one kind; C. multipetala, C. longgangensis, C. parvipetala, C. tunghinensis and C. limonia, C. achrysantha, C. microcarpa, C. nitidissima, C. terminali and C. pingguoensis should be divided into separate species. FTIR-cluster analysis can be used as a possible means for the identification of Camellia Sect. Chrysantha.

[Calibration methods of infrared spectra for geographical origins determination of Radix Zanthoxyli].

The instrument and the experimental environment influence the infrared spectra, which may limited the identification of the samples by a prediction model. Based on the Fourier transform infrared spectroscopy (FTIR) technology, the authors performed different infrared spectral calibration methods for Radix Zanthoxyli geographical origins determination, the SIMCA was used to establish an identification models, and the model was used to distinguish samples from four different regions of Guangxi. According to the result of prediction, the authors could obtain the most suitable calibration method for the identification model. The results showed that, respectively, by the multiple scattering correction and standard normal variation, their PCA data distribution and the distance between models is ideal, suggesting that we can eliminate the interference from the environmental and human factors by these two correction methods, and also separate each samples of different habitats. The test using the method to measure the geographical origins of Radix Zanthoxyli proved that the recognition rate and rejection rate are both at or near 100%. Visible, and both the multiplicative scatter correction and the standard normal variation are all the ideal calibration methods for Radix Zanthoxyli infrared spectral geographical origins determination.

Autotoxicity in Panax notoginseng of root exudatesand their allelochemicals.

The growth of Panax notoginseng (Burk.) F. H. Chen is frequently hindered due to replanting failure. In the present study, the objective is to determine whether root exudates from P. notoginseng have autotoxicity and identification of allelochemicals from root exudates or rhizosphere soil. We investigated autotoxicity in P. notoginseng using seedling emergence bioassays and hydroponic culture. The allelochemicals in the soils and root exudates were identified with GC-MS, and the autotoxicity of the identified key allelochemicals was investigated by bioassay. The results showed that the root exudates, and extracts from consecutively cultivated soils also showed significant autotoxicity against seedling emergence and growth. In the non-renewed culture solution without activated charcoal (AC), the fresh and dry mass of P. notoginseng tubers of roots was reduced by about half compared to the addition with AC. A total of 44 different components from all samples were defined by GC-MS analyses. Furthermore, the results of multiple statistical analysis showed a t the difference among cultivated soil, uncultivated soil and root exudates. Bioassay of the identified allelochemicals revealed that benzoic acid, phthalic acid, palmitic acid, and stearic acid significantly affected the root growth of P. notoginseng. These substances at 100 µM more significantly decreased the number of lateral roots. Our results demonstrated that autotoxicity results in replant failure of P. notoginseng.

Comparative transcriptomic analysis identifies KcMYB1 as a R2R3-MYB anthocyanin activator in Kadsura coccinea.

Fruit color, as an important appearance attribute, is crucial for attracting consumers. However, the underlying mechanism regulating mature fruit color formation in Kadsura coccinea remains unclear. Here, a comprehensive metabolomics and transcriptomics analysis was performed to investigate the molecular mechanisms of anthocyanin accumulation between two K. coccinea cultivars with different mature fruit colors-'Dahong No. 1' (red) and 'Jinhu' (yellow). Targeted metabolomic analysis revealed high anthocyanin levels, most of which were cyanidin and delphinidin derivatives, in 'Dahong No. 1' mature fruit peel. The SNP analysis indicated that the two different cultivars had similar genetic background. Moreover, comparative transcriptomic analysis demonstrated that differentially expressed genes (DEGs) were related to flavonoid biosynthesis and metabolic process in the two K. coccinea cultivars. Gene expression profiling data showed that the structural and regulatory genes associated with anthocyanin biosynthesis were significantly upregulated in 'Dahong No. 1' mature fruit peel, which was verified by quantitative real-time polymerase chain reaction (qRT-PCR). Notably, the key anthocyanin activator KcMYB1 was identified, which was significantly upregulated in 'Dahong No. 1' compared with 'Jinhu'. We further confirmed that KcMYB1 actively regulated the accumulation of anthocyanin by ectopic expression in vivo. Furthermore, allelic constitution of KcMYB1 in K. coccinea were investigated. The present study can provide insights for understanding the regulatory mechanisms of anthocyanin differential accumulation in the mature fruits of K. coccinea.

Cultivable endophytic fungal community associated with the karst endemic plant Nervilia fordii and their antimicrobial activity.

Endophytic fungi from medicinal plants with specific pharmacological functions attract much attention to provide the possibility of discovering valuable natural drugs with novel structures and biological activities. Nervilia fordii is a rare and endangered karst endemic plant that is used as medicine and food homology in Guangxi, China. These plants have been reported to have antimicrobial, antitumor, antiviral, and anti-inflammatory activities. However, few studies have focused on the diversity and antibacterial activity of endophytic fungi from N. fordii. In the present study, 184 endophytic fungi were isolated from the healthy tissues of N. fordii, and their molecular diversity and antimicrobial activities were analyzed for the first time. These fungi were categorized into 85 different morphotypes based on the morphological characteristics and the similarity between the target sequence and the reference sequence in the GenBank database. With the exception of 18 unidentified fungi, the fungal isolates belonged to at least 2 phyla, 4 classes, 15 orders, 45 known genera, and 45 different species, which showed high abundance, rich diversity, and obvious tissue specificity. All isolates were employed to screen for their antimicrobial activities via the agar diffusion method against Escherichia coli, Staphylococcus aureus, and Candida tropicalis. Among these endophytes, eight strains (9.41%) displayed inhibitory activity against E. coli, 11 strains (12.94%) against S. aureus, and two strains (2.35%) against C. tropicalis, to some extent. In particular, our study showed for the first time that the fungal agar plugs of Penicillium macrosclerotiorum 1151# exhibited promising antibacterial activity against E. coli and S. aureus. Moreover, the ethyl acetate (EA) extract of P. macrosclerotiorum 1151# had antibacterial effects against E. coli and S. aureus with a minimum inhibitory concentration (MIC) of 0.5 mg ml-1. Further research also confirmed that one of the antimicrobial compounds of P. macrosclerotiorum 1151# was methyl chloroacetate and exhibited excellent antibacterial activity against E. coli and S. aureus up to 1.71-fold and 1.13-fold compared with tetracycline (TET) (5 mg ml-1), respectively. Taken together, the present data suggest that various endophytic fungi of N. fordii could be exploited as sources of novel natural antimicrobial agents.

[Raman spectra and structure analysis of 2,6-pyridine dicarboxylic acid in different states and single Bacillus spore].

The Raman spectra of 2,6-pyridine dicarboxylic acid (DPA) and their calcium salts(Ca-DPA) in different states and the Ca-DPA in a single bacterial spore have been recorded by laser tweezers Raman system (LTRS) and the spectra have been assigned. Raman spectra of different states of DPA and Ca-DPA are different evidently. Analysis leading to differences in the structure of spectrum may be due to that the Raman spectra of DPA crystalline reflected more precise characteristics information compared to DPA powder, in which the laser can penetrate through DPA crystalline and the Raman scatter from the crystalline interior is greater than that from DPA powder. The second reason is that DPA powder and Ca-DPA crystalline contain water molecules, and the intermolecular hydrogen bonding in the crystals of these molecules is extensive. The presence of calcium ions would affect the pyridine ring so that both sides of the carboxyl pyridine ring have a certain geometric deformation and the hydroxy carboxylic was damaged. The DPA2-anion is principal in Ca-DPA and the DPA solution. The calcium ion affects the stability of the pyridine ring structure in the Ca-DPA solution. The result from the spectra also showed that the DPA in single spores present Ca-DPA crystal state.

Integration of transcriptome and targeted metabolome profiling reveals hormone related genes involved in the growth of Bletilla striata.

Bletilla striata (Thunb.) Reichb.f. (BS) is a traditional Chinese medicine with numerous beneficial effects. In our previous study, Aspergillus flavus was isolated from B. striata. To explore the physiological and molecular mechanisms of Aspergillus flavus elicitor (1-G4) that promoted Bletilla striata growth, in this study, we performed the determination of growth indexes and transcriptomics and metabolomics analysis under 5% and 10% 1-G4 conditions. Results showed that 1-G4 elicitor could significantly promote the growth and development of B. striata. With the increasing concentration of 1-G4 elicitor, the contents of SA, ICAld, and ME-IAA significantly increased while the IP and ACC contents decreased dramatically. A total of 1657 DEGs (763 up-regulated and 894 down-regulated) between the control (CK) and 5% elicitor (CK vs G5) and 2415 DEGs (1208 up-regulated and 1207 down-regulated) between the control and 10% elicitor (CK vs G10) were identified. Further, we found that 22, 38, and 2 unigenes were involved in ME-IAA, IP, and ACC, respectively. It was indicated that these unigenes might be involved in B. striata growth. Overall, the current study laid a theoretical foundation for the effective utilization of endophytic fungi and the optimization of germplasm resources of B. striata.

Identification of Isoflavonoid Biosynthesis-Related R2R3-MYB Transcription Factors in Callerya speciosa (Champ. ex Benth.) Schot Using Transcriptome-Based Gene Coexpression Analysis.

The R2R3-MYB family is one of the largest plant transcription factor (TF) families playing vital roles in defense, plant growth, and secondary metabolism biosynthesis. Although this gene family has been studied in many species, isoflavonoid biosynthesis-related R2R3-MYB TFs in Callerya speciosa (Champ. ex Benth.) Schot, a traditional Chinese medicinal herb, are poorly understood. Here, a total of 101 R2R3-MYB TFs were identified from C. speciosa transcriptome dataset. 25 clades divided into five functional groups were clustered based on the sequence similarity and phylogenetic tree. Conserved motifs and domain distribution, expression patterns, and coexpression networks were also employed to identify the potential R2R3-MYB TFs in the regulation of isoflavonoid biosynthesis. In silico evaluation showed that the deduced R2R3-CsMYB proteins contain highly conserved R2R3 repeat domain at the N-terminal region, that is the signature motif of R2R3-type MYB TFs. Eight potential TFs (CsMYB17, CsMYB36, CsMYB41, CsMYB44, CsMYB45, CsMYB46, CsMYB72, and CsMYB81) had high degrees of coexpression with four key isoflavonoid biosynthetic genes (CsIFS, CsCHS7, CsHID-1, and CsCHI3), in which CsMYB36 as a potential regulator possessed the highest degree. HPLC analysis showed that formononetin and maackiain contents were significantly increased during the development of tuberous roots, which might be controlled by both related R2R3-CsMYBs and structural genes involved in the isoflavonoid biosynthesis pathway. The transcriptome data were further validated by reverse transcription real-time PCR (RT-qPCR) analysis, and similar expression profiles between TFs and key structural genes were identified. This study was the first step toward the understanding of the R2R3-MYB TFs regulating isoflavonoid biosynthesis in C. speciosa. The results will provide information for further functional analysis and quality improvement through genetic manipulation of these potential R2R3-CsMYB genes in C. speciosa.