[Objective]To clarify the material basis of sourness in heated tobacco and the"tobacco raw material-chemical components-sensory"correlation mechanism.[Method]57 representative tobacco leaves of heated tobacco products were selected.39 chemical compositions of the raw tobacco leaves was determined using methods such as continuous flow analysis,UPLC,and GC-MS;heated tobacco samples were prepared using a roller-pressing method,and 54 chemical composition of the smoke was qualitatively and quantitatively analyzed using central-cut two-dimensional gas chromatography-mass spectrometry(GC-GC/MS).The sourness was evaluated using a 0-5 point scale.Kruskal-Wallis tests,Spearman's correlation analysis,Random forest model,and cluster analysis were employed to identify the key chemical components associated with sourness.[Result]57 heated tobacco product samples were classified into three groups based on sensory evaluation of sourness:slightly sour(38),moderately sour(10),and sour(9).Analysis of differences indicated that 47 of the 93 detected components showed significant differences between groups(P＜0.05).Correlation analysis indicated that components such as chlorogenic acid(Rho=0.84),sucrose(Rho=0.82),rutin(Rho=0.81),and acetic acid(Rho=0.56)showed a highly significant positive correlation with the sourness score;components such as,isovaleramide(Rho=-0.77),oxalic acid(Rho=-0.72),and nicotine(Rho=-0.50)showed a significant negative correlation with the sourness score.Cluster analysis results indicate that the 57 samples can be divided into two clusters.The right-hand cluster(37 samples),consisting primarily of flue-cured tobacco,is enriched with components positively correlated with sourness,such as chlorogenic acid and sucrose.These samples exhibit a sensory profile of"sour"or"slightly sour",forming the material basis for the strong sourness profile of heated tobacco products;the left cluster(20 samples),composed of air-cured tobaccos such as Burley and cigar tobacco,was highly enriched in negatively correlated components such as nitrogen-containing compounds and was perceived as"slightly sour".The classification results objectively reflect how typical metabolic differences shape sensory profiles:flue-cured tobacco dominated by carbon metabolism extensively accumulates sugars and polyphenols,laying the material basis for strong sourness,whereas air-cured tobacco dominated by nitrogen metabolism is enriched in alkaline substances such as alkaloids,significantly suppressing sourness expression.[Conclusion]The sourness of heated tobacco products is highly correlated with the chemical components of both raw materials and smoke.This study identified polyphenols and sugars represented by chlorogenic acid,sucrose,and acetic acid as the key positive acid-contributing components driving the expression of sourness in heated tobacco products,while nitrogen-containing compounds and specific dicarboxylic acids represented by isovaleramide,oxalic acid,and nicotine were identified as the negative regulatory components that inhibit sourness.The synergistic accumulation of carbon metabolites and the appropriate reduction of nitrogenous alkaline substances constitute the shared material basis for forming the strong sourness profile in heated tobacco products,providing a scientific basis for raw material screening and sensory regulation.

The WD40 protein is a key regulator of the flavonoid biosynthetic pathway. However, its role in the flavonoid-rich aquatic cereal Chinese wild rice (Zizania latifolia) remains unclear. Here, we systematically characterised the WD40 gene family in Z. latifolia. In total, 38 ZlWD40 genes were identified and mapped to 15 chromosomes. Among them, ZlTTG1 (Zla08G018110) was localised to the nucleus. ZlTTG1-overexpression (ZlTTG1) in rice changed the pericarp colour from light brown to dark purple but did not significantly affect agronomic traits. ZlTTG1 overexpression increased flavonoid content and antioxidant activity and enhanced enzyme inhibitory effects in rice seeds. Compared with the control, 155 flavonoids and 269 genes were upregulated in ZlTTG1-overexpressing rice seeds, which may contribute to the dark purple pericarp phenotype. Consistently, ZlTTG1 rice seeds showed higher expression of flavonoid biosynthetic genes (OsCHS, OsCHIL1, OsCHIL2, OsF3H-1, OsF3'H, OsDFR, OsANS, and OsUGT707A3) and increased activities of key biosynthetic enzymes, including CHS, F3H, F3'H, DFR, and ANS. This study provides a foundation for the functional analysis of ZlWD40 genes and identifies new genetic resources for developing flavonoid-rich functional rice.

Polished rice is highly favoured for excellent taste; however, its processing can lead to the loss of flavonoids. To address this issue, this study achieved endosperm-specific expression of ZlRc and ZlMYB1 (Rc-MYB1) in wild-type (WT) rice. Rc-MYB1 rice exhibited a reduction in yield; however, it showed significantly increased flavonoid content, antioxidant activity, and enzyme-inhibitory effects in seeds. Flavonoids accumulated in the endosperm of Rc-MYB1 rice seeds. Comparative analyses revealed significant upregulation of 15 flavonoids and 732 genes in Rc-MYB1 seeds relative to those in the WT. Expression of flavonoid biosynthesis-, transport-, and accumulation-associated genes (OsF5HL2, OsVSR6, and Os1-CysPrxB, respectively) and ferulate-5-hydroxylase activity were significantly higher in Rc-MYB1 rice seeds than in the WT. Overall, this study demonstrates successful enrichment of flavonoids in Rc-MYB1 endosperm, providing a promising strategy for improving the nutritional quality of polished rice and advancing the development of functional staple foods.

The high-value carotenoid astaxanthin is biosynthesized through a dual-enzyme-catalyzed cascade and is getting increased attention for engineered biosynthesis in plants. When developing astaxanthin-producing tobacco by expressing 2A-peptide-linked CBFD (carotenoid β-ring-4-dehydrogenase) and HBFD (carotenoid 4-hydroxy-β-ring-4-dehydrogenase) from Adonis aestivalis, this work discovered an in-enzyme splicing site at the N-terminus of HBFD that has potentials for multiple protein expression in plant using monocistronic cassette. Based on this finding, we generated astaxanthin-producing tobacco plants expressing a directly fused protein of CBFD and HBFD with a monocistronic cassette. Further integrated IP (immunoprecipitation) and LC-MS/MS assays revealed the presence of an in-enzyme splicing site at the N-terminus of HBFD. Nevertheless, the obtained astaxanthin-producing tobacco plants exhibited a growth retardation as observed by previous researches. Subsequent studies revealed that the astaxanthin-producing caused growth retardation of tobacco was correlated with chloroplast disruption and chlorophyll reduction, and it could be alleviated by expressing a chlorophyll biosynthetic enzyme identified by proteomics. Additionally, crossing the astaxanthin-producing tobacco with a variety having higher chlorophyll content also alleviated the growth retardation caused by astaxanthin production, and improved the total astaxanthin yield per plant by at least threefold along with the biomass increase. This work provides novel approaches for expressing multiple proteins in tobacco and for engineering efficient astaxanthin-producing tobacco.

Northern wild rice (NWR; Zizania palustris L.), an annual aquatic plant in the Poaceae family, has high economic importance due to its nutrient-rich grains. However, the existing NWR genome assembly for this species has severe fragmentation and incomplete gene representation. A near-complete genome was assembled in this study to provide a high-quality genomic reference for NWR-associated research. The assembled genome exhibited a total contig length of 1.41 Gb and a contig N50 of 109.22 Mb. Overall, a 73.60% repetitive sequence content was identified and 47,804 genes predicted. Phylogenetic analysis indicated that Z. palustris was most closely related to Zizania latifolia, with an estimated divergence time of 4.57-8.15 Mya. Meanwhile, Z. palustris underwent a recent, species-specific long terminal repeat (LTR) expansion, associated with its larger genome size. We identified two genomic blocks in the Z. palustris and Z. latifolia genomes that exhibit strong synteny with the rice phytocassane biosynthetic gene cluster. The centromeric satellite repeats in Z. palustris identified in this study primarily comprised a 145 bp repetitive unit. The findings also revealed centromere homogenisation and rearrangement accompanied by LTR invasion in NWR. Among the genes missing in the previous NWR genome, we observed LTR insertion events that resulted in expanded gene lengths in our updated NWR genome. The present updated NWR genome provides a valuable resource for crop genetic improvement, functional gene discovery, and research on critical biological processes. (c) 2025 Crop Science Society of China and Institute of Crop Science, CAAS. Production and hosting by Elsevier B.V. on behalf of KeAi Communications Co., Ltd. This is an open access article under the CC BY-NCND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Microplastics and various other contaminants are frequently present in soil. Here we investigated the long-term integrated responses of changes in the microbial community, metabolomics, heavy metal availability, and nutritional properties of the cadmium-cuprum-zinc-contaminated coastal saline soil to the three different microplastics. Various categories of microplastics had notable impacts on the available potassium, organic matter, availability of cadmium and cuprum, as well as the enzymatic activity in soil. Microplastics contamination caused diverse changes in microbial diversity and the composition of bacterial and fungal communities, resulting in the enrichment of Mortierella and a decrease of Bacillus abundance. The metabolites in soil primarily affected by microplastics contamination were the pathways involved organic acids and their derivatives, organoheterocyclic compounds, as well as lipids and lipid-like substances. Therefore, the addition of microplastics to soil may influence soil fertility, metal mobility, and alter the structure and metabolic processes of the microbial community in soil.

Whole grains represent key components of a healthy diet, helping to meet the nutritional needs of consumers and playing a crucial role in preventing chronic diseases. Whole grains are rich in various types of flavonoids with antioxidants and health-promoting properties at varying levels. This article defines and elucidates different whole grain types, analyses the advantages and disadvantages of commonly used metabolomics instruments, and systematically organises and classifies flavonoids detected in whole grains. Additionally, we mapped flavonoid biosynthetic pathways and discussed the usefulness of metabolomic techniques in elucidating the functions of key genes involved in flavonoid biosynthesis. The MYB-bHLH-WD40 (MBW) complex regulates flavonoid biosynthesis during seed development, regulating seed colour and flavonoid content. In addition, MBW complex expression is highly tissue-specific; it is preferentially expressed in purple or black tissues. This review describes flavonoid diversity and biosynthetic pathways in whole grains and provides a theoretical foundation for functional whole grain development and usage.

In metabolically engineered plants, the target products are usually uniformly distributed in the whole plant or specific tissues. When engineering tobacco to produce astaxanthin, a ketocarotenoid with strong antioxidant activity and multiple bioactivities, a scattered distribution of astaxanthin-producing regions was observed in a small portion of astaxanthin-producing tobacco plants, which caused mosaic-like red and green spots on the leaves (ASTA-mosaic). A physiological assay showed that the non-astaxanthin green region (Mosaic_G) had relatively higher chlorophyll content and better chloroplast structure than the astaxanthin-producing red region (Mosaic_R). Then, metabolomics, proteomics, and small RNA transcriptomics were employed to analyze the uneven distribution of astaxanthin-producing regions in tobacco leaves. The results of metabolomics and proteomics revealed a decrease in carotenoid metabolism, chlorophyll biosynthesis, and chlorophyll degradation in the Mosaic_G region. Pheophorbide a, an intermediate of chlorophyll degradation, was found to be significantly reduced in the Mosaic_G region, which was accompanied by the attenuation of chlorophyllase and pheophytinase, which catalyze the formation of pheophorbide a in chlorophyll degradation. Reductions in photosynthetic antenna proteins and photosystem-associated proteins were observed in the Mosaic_R region, consistent with the better chloroplast structure of the Mosaic_G region. Small RNA transcriptomics showed that several small RNAs could target chlorophyll-degradative genes, but they were more effective in targeting the astaxanthin biosynthetic genes. This finding was supported by the fact that the Mosaic_G region can remain green up to the senescence of tobacco leaves. This work provides insights into the mechanism of the uneven distribution of astaxanthin-producing regions in tobacco leaves and may contribute to the specialized utilization of tobacco plants for metabolic engineering.

Pigmented rice (e.g., black or purple rice), rich in flavonoids and other active compounds, has lower yields than non-pigmented rice. The study aimed to breed pigmented rice with stable yields. To this end, we created transgenic rice by inserting the ZlRc and ZlRd genes from Chinese wild rice (Zizania latifolia) into rice (Oryza sativa) and investigated the resulting yield and flavonoid content. Rc encodes a transcription factor that regulates flavonoid biosynthesis, while dihydroflavonol 4-reductase (DFR/Rd) is an important flavonoid biosynthetic enzyme. The dark brown colour of the pericarp rice was attributed to the co-overexpression of ZlRc and ZlRd (ZlRcRd), which did not influence the agronomic traits or yield. No significant difference was observed in the chlorophyll content or photosynthetic gas exchange parameters between the ZlRcRd and wild-type (WT) rice. The flavonoid content in the ZlRcRd rice seeds was significantly enhanced, and so were antioxidant activity and inhibitory effects on α-glucosidase, α-amylase, pancreatic lipase, and tyrosinase. Compared with WT rice, 1080 genes and 82 flavonoids were upregulated in the ZlRcRd rice. The expression of key genes involved in flavonoid biosynthesis (CHS, F3'H, and F3'5'H) and the activities of related key enzymes were significantly higher in ZlRcRd than WT rice. This study proposes new germplasm resources and technical approaches for breeding pigmented rice with stable yields and high flavonoid content, which will address the challenge of low yields of pigmented rice and increase farmers' enthusiasm for planting pigmented rice.

Tobacco is an excellent chassis for metabolic engineered biosynthesis of astaxanthin, a high-value carotenoid, yet efficient cultivation and processing methods remain to be established for enhancing its astaxanthin production. This work developed astaxanthin-producing tobacco plants utilizing the astaxanthin biosynthetic pathway from Adonis aestivalis and carried out metabolomics analyses to reveal the limits of astaxanthin-production efficiency in tobacco. The results showed that astaxanthin production altered the abundance of amino acids as well as a set of compounds involved in carbohydrate metabolism. The abundance of some downstream nitrogenous substances, such as nicotine, spermidine, uridine phosphate and guanosine phosphate, and organic acids in the tricarboxylic acid cycle were also changed in the astaxanthin-producing plants. These findings prompted us to determine the effects of additional nitrogen and carbon supply on astaxanthin production, which revealed that adding either or both of nitrogen and sucrose notably increased the biomass and astaxanthin production of tobacco. As high biomass plants, drying of the raw tobacco materials is required for further extraction. Efforts to optimize the processing method of raw-materials from astaxanthin-producing tobacco revealed that a direct vacuum-drying method could reduce the astaxanthin loss in tobacco leaves by similar to 20 % in comparison with shadedrying or freeze-drying. This method also well retained the astaxanthin in tobacco stems and made them worthy of astaxanthin extraction. Integration of the high-nitrogen/sucrose cultivation and direct vacuum-drying methods essentially improved the whole-biomass utilization efficiency of astaxanthin-producing tobacco and resulted in similar to 2 folds of astaxanthin yield, which may contribute greatly to cost effective astaxanthin production.

Cembranoids and labdanes are two important types of diterpenes in tobacco (Nicotiana genus) that are predominantly found in the leaf and flower glandular trichome secretions. This is the first systematic review of the biosynthesis, chemical structures, bioactivities, and utilisation values of cembranoid and labdane diterpenes in tobacco. A total of 131 natural cembranoid diterpenes have been reported in tobacco since 1962; these were summarised and classified according to their chemical structure characteristics as isopropyl cembranoids (1-88), seco-cembranoids (89-103), chain cembranoids (104-123), and polycyclic cembranoids (124-131). Forty natural labdane diterpenes reported since 1961 were also summarised and divided into epoxy side chain labdanes (132-150) and epoxy-free side chain labdanes (151-171). Tobacco cembranoid and labdane diterpenes are both formed via the methylerythritol 4-phosphate pathway and are synthesised from geranylgeranyl diphosphate. Their biosynthetic pathways and the four key enzymes (cembratrienol synthase, cytochrome P450 hydroxylase, copalyl diphosphate synthase, and Z-abienol cyclase) that affect their biosynthesis have been described in detail. A systematic summary of the bioactivity and utilisation values of the cembranoid and labdane diterpenes is also provided. The agricultural bioactivities associated with cembranoid and labdane diterpenes include antimicrobial and insecticidal activities as well as induced resistance, while the medical bioactivities include cytotoxic and neuroprotective activities. Further research into the cembranoid and labdane diterpenes will help to promote their development and utilisation as plant-derived pesticides and medicines.

Tobacco cembranoids, known for their anti-insect and antifungal properties, were shown to be mainly present on the surface of leaves and flowers, being biosynthesized by their trichomes. It remains unclear whether they could be biosynthesized in other organs without trichomes. Cembratrien-ol synthases (CBTSs) catalyze the conversion of GGPP to CBT-ols and thus play an important role in cembranoid biosynthesis. This study identified the CBTS family genes in tobacco and examined their spatiotemporal expression patterns. The CBTS genes showed diverse expression patterns in tobacco organs, with the majority highly expressed in leaves and a few highly expressed in flowers. The expression of CBTS genes were also correlated with the development of tobacco plants, and most of them showed the highest expression level at the budding stage. Furthermore, their expression is mediated by the JA (jasmonate) signaling in all tobacco organs. Several CBTS genes were found to be highly expressed in tobacco roots that have no trichomes, which prompted us to determine the cembranoid production in roots and other organs. GC-MS and UPLC assays revealed that cembranoids were produced in all tobacco organs, which was supported by the bioactivity assay results that almost all these CBTS enzymes could catalyze CBT-ol biosyntheis in yeast, and that the content ratio of CBT-ols and CBT-diols in tobacco roots was different to that in leaves. This work sheds insights into the expression profiles of tobacco CBTS genes and provides a feasibility to engineer tobacco roots for industrial production of cembranoids.

Solanesol, which accumulates predominantly in the leaves of tobacco plants, has medically important bioactive properties. To investigate the genetic basis of solanesol in tobacco (Nicotiana tabacum), the solanesol contents of 222 accessions, 206 individuals from an N. tabacum Maryland609 (low-solanesol) × K326 (high-solanesol) F2 population and their corresponding F1 self-pollinations, were determined using ultra-performance liquid chromatography. Genome-wide quantitative trait locus (QTL) and association analysis were performed to identify QTLs and markers associated with solanesol content based on simple sequence repeat molecular markers. A total of 12 QTLs underlying solanesol content were mapped to seven linkage groups (LGs), with three of the QTLs (QTL3-1, QTL21-6, and QTL23-3) explaining 5.19–10.05% of the phenotypic variation. Association analysis revealed 38 significant marker-trait associations in at least one environment. The associations confirmed the QTLs located on LG3, LG10, LG14, LG21, and LG23, while new elite makers were located on 11 additional LGs, each explaining, respectively, 5.16–20.07% of the phenotypic variation. The markers LG14-PT54448, LG10-PT60114-2, LG10-PT60510, LG10-PT61061, and LG-21PT20388 may be useful for molecular-assisted selection of solanesol content in tobacco leaves. These results increase our understanding of the inheritance of solanesol-associated genes and will contribute to molecular-assisted breeding and further isolation of regulatory genes involved in solanesol biosynthesis in tobacco leaves.

Anthocyanins are natural flavonoids with a high antioxidant power and many associated health benefits, but most rice produce little amounts of these compounds. In this study, 141 MYB transcription factors in 15 chromosomes, including the nucleus-localised ZlMYB1 (Zla03G003370) and ZlMYB2 (Zla15G015220), were discovered in Zizania latifolia. Overexpression of ZlMYB1 or ZlMYB2 in rice seeds induced black pericarps, and flavonoid content, antioxidant capacity, and α-glucosidase and tyrosinase inhibition effects significantly increased compared to those in the control seeds. ZlMYB1 and ZlMYB2 overexpression induced the upregulation of 764 and 279 genes, respectively, and the upregulation of 162 and 157 flavonoids, respectively, linked to a black pericarp phenotype. The expression of flavonoid 3'-hydroxylase and UDP-glycose flavonoid glycosyltransferase, as well as the activities of these enzymes, increased significantly in response to ZlMYB1 or ZlMYB2 overexpression. This study systematically confirmed that the overexpression of ZlMYB1 and ZlMYB2 promotes flavonoid biosynthesis (especially of anthocyanins) in rice.

The fall armyworm Spodoptera frugiperda is considered one of the most destructive crop pests, posing a significant threat to food and crop security. In this study, we conducted a chemical investigation of the endophytic fungus Aspergillus sp. 1022LEF, leading to the identification of a previously unreported benzothiazole derivative, 6-(2-hydroxyethyl)benzo[d]thiazol-4-ol (HBT). Its structure was unambiguously characterized using extensive spectroscopic methods, including 1D and 2D NMR data, HRESIMS data, and single-crystal X-ray diffraction analysis. The insecticidal assay revealed that HBT possessed remarkable activity against S. frugiperda with an LC50 value of 0.24 mg/mL. Further transcriptomic and proteomic analyses revealed that HBT induced mortality in S. frugiperda by impeding DNA replication and protein synthesis, influencing mitochondria-mediated autophagy, and perturbing hormone synthesis, thereby disrupting the fundamental biosynthetic processes. HBT demonstrated high activity in controlling S. frugiperda, which highlighted its potential use as a lead in the development of biopesticides.

Five new sorbicillinoid derivatives, including (+/-)-aspersorbicillin A [(+/-)-1], a pair of enantiomers at C-9, and aspersorbicillins B-D (2-4), together with two known analogs (5 and 6) were isolated from the endophytic fungus Aspergillus aculeatus TE-65L. Their structures including absolute configurations were determined by detailed spectroscopic analyses and electronic circular dichroism calculations. The herbicidal activity of sorbicillinoids on the germ and radicle elongation of various weed types was reported for the first time. Compound 1 displayed significant herbicidal activity against Eleusine indica germ elongation (IC50 = 28.8 mu g/mL), while compound 6 inhibited radicle elongation (IC50 = 25.6 mu g/mL). Both were stronger than those of glyphosate (66.2 and 30.9 mu g/mL, respectively). Further transcriptomic and LC-MS/MS metabolomic analysis indicated that 6 induced the transcriptional expressions of genes related to the lignin biosynthetic pathway, resulting in lignin accumulation. Transmission electron microscopy confirmed the cell wall thickening of seeds treated with 6, suggesting weed growth inhibition. This study reveals new lead compounds for fabricating natural herbicides and expands the agricultural use of sorbicillinoid analogs.

The production of alternative proteins is of great significance in the mitigation of food problems. This study proposes an integrated approach including protein extraction, enzymatic hydrolysis, and fermentation to produce both plant proteins and single-cell proteins as alternative proteins from tobacco leaves, a highly-abundant and protein-rich agricultural waste. Alkaline extraction of proteins before polysaccharide hydrolysis was found to be preferable for increasing the yields of plant proteins and mono-sugars. The combined use of pectinase-rich enzymes from Aspergillus brunneoviolaceus and hemicellulase-rich enzymes from Penicillium oxalicum achieved the release of 80.7 % of the sugars after 72 h. Cutaneotrichosporon cutaneum could simultaneously utilize multiple sugars, including galacturonic acid, in the enzymatic hydrolysate to produce single-cell proteins. Via this approach, 43.54 g crude proteins of high protein contents and rich in essential amino acids can be produced from 100.00 g waste tobacco leaves, providing a promising strategy for its valorization.

Biomass-degrading enzymes produced by microorganisms have a great potential in the processing of agricultural wastes. In order to produce suitable biomass-degrading enzymes for releasing sugars and aroma compounds from tobacco scraps, the feasibility of directly using the scraps as a carbon source for enzyme production was investigated in this study. By comparative studies of ten fungal strains isolated from tobacco leaves, Aspergillus brunneoviolaceus Ab-10 was found to produce an efficient enzyme mixture for the saccharification of tobacco scraps. Proteomic analysis identified a set of plant biomass-degrading enzymes in the enzyme mixture, including amylases, hemicellulases, cellulases and pectinases. At a substrate concentration of 100 g/L and enzyme dosage of 4 mg/g, glucose of 17.6 g/L was produced from tobacco scraps using the crude enzyme produced by A. brunneoviolaceus Ab-10. In addition, the contents of 23 volatile molecules, including the aroma compounds 4-ketoisophorone and benzyl alcohol, were significantly increased after the enzymatic treatment. The results provide a strategy for valorization of tobacco waste by integrating the production of biomass-degrading enzymes into the tobacco scrap processing system.

Cembranoids produced by tobacco glandular trichomes have bioactivities in resistance to insect pests and pathogens. Cembratrien-ol synthase (CBTS) plays a key role in the biosynthesis of cembranoids and directly determines the cembranoid content in tobacco. This study examined the effect of changing CBTS1 expression on tobacco resistance to the insect pest Spodoptera frugiperda and oomycete pathogen Phytophthora nicotianae. The CDS sequence of CBTS1 was cloned into gene overexpression and silencing vectors and introduced into tobacco (Nicotiana tabacum L. cv. TN90) to obtain CBTS1-overexpression plants (CBTS1-OE) and CBTS1-silenced plants (CBTS1-RI). Compared with control plants, the content of cembratrien-ol (CBT-ol) was increased 4.48 times in the CBTS1-OE plants but decreased by 68% in the CBTS1-RI plants, while that of cembratrien-diol (CBT-diol) was increased 3.17 times in the CBTS1-OE plants but decreased by 76% in the CBTS1-RI plants. The S. frugiperda resistance of transgenic tobacco plants was evaluated by in vitro toxicity test, and the results showed that the resistance of CBTS1-OE plants to S. frugiperda was significantly improved but that of CBTS1-RI plants was reduced. The P. nicotianae resistance of transgenic tobacco plants was assessed by the detached leaf assay, and the results showed that the resistance of CBTS1-OE plants to P. nicotianae was enhanced, while that of CBTS1-RI plants was attenuated. Further gene expression analysis showed that overexpression of CBTS1 increased the expression of the pathogen-related gene PR-1a, while silencing of CBTS1 decreased its expression. This study demonstrated that manipulating the expression of CBTS1 could change the cembranoid content in tobacco plants and alter their resistance to both insect pests and oomycete pathogens.