A new surface molecularly imprinted polyacrylamide nanoprobe for trace Cr(VI) with RRS technique.

This article was used potassium dichromate as the template molecule, silver nanoclusters as the nano matrix, acrylamide as the monomer, ethylene glycol dimethacrylate (EGDMA) as the crosslinking agent, and azodiisobutyronitrile (AIBN) as the initiator to prepare a new silver nanocluster surface MIP (AgNCs@MIP) nanoprobe for chromate. Upon addition of Cr(VI), it selectively adsorbs on the surface of AgNCs@MIP nanoprobes. The dichromate ion absorption peak at 350 nm overlaps with the AgNCs@MIP RRS peak at 370 nm, resulting in strong RRS energy transfer (RRS-ET) and a decrease in the RRS intensity. The decreased RRS intensity is directly proportional to the concentration of dichromate ions in the range of 0.0025-0.015 µmol/L, with a detection limit of 0.8 nmol/L. Therefore, a simple, fast, sensitive and selective RRS method for the determination of trace Cr(VI) in mineral water has been established, with a relative standard deviation of 9.2-9.8 % and recovery of 95.20 %-103.60 %.

Glutathione S-transferase directly metabolizes imidacloprid in the whitefly, Bemisia tabaci.

The whitefly Bemisia tabaci poses a significant threat to various crops and ornamental plants and causes severe damage to the agricultural industry. Over the past few decades, B. tabaci has developed resistance to several pesticides, including imidacloprid. Therefore, elucidating the mechanism that leads to insecticide detoxification is very important for controlling B. tabaci and managing whitefly resistance to neonicotinoid insecticides. Among insect detoxification enzymes, glutathione S-transferase (GST) is an important phase II detoxification enzyme that helps detoxify exogenous toxic substances. In this study, we cloned the BtGSTz1 gene and observed that its expression level was greater in imidacloprid-resistant populations than sensitive populations of B. tabaci. By silencing BtGSTz1 via RNA interference, we found a significant increase in the mortality of imidacloprid-resistant B. tabaci. Additionally, prokaryotic expression and in vitro metabolism studies revealed that the recombinant BtGSTz1 protein could metabolize 36.36% of the total imidacloprid, providing direct evidence that BtGSTz1 plays a crucial role in the detoxification of imidacloprid. Overall, our study elucidated the role of GSTs in physiological activities related to insecticide resistance, which helps clarify the resistance mechanisms conferred by GSTs and provides useful insights for sustainable integrated pest management.

A new PdMOF-loaded molecularly imprinted polyaniline nanocatalytic probe for ultratrace oxytetracycline with SERS technique.

In this paper, a new Pd metal organic framework (PdMOF) surface molecularly imprinted polyaniline nanocatalytic probe (PdMOF@MIP) with dual functions of recognition and catalysis was synthesized. It is found that the PdMOF@MIP nanoprobe can not only identify OTC but also catalyze the new nanoreaction of NaH2PO2-HAuCl4 to generate gold nanoparticles (AuNPs), and the generated AuNPs could be traced by surface-enhanced Raman scattering (SERS). When OTC specifically binds to PdMOF@MIP to generate PdMOF@MIP-OTC conjugate, its catalytic effect is weakened and the analytical signal is reduced linaerly. Accordingly, a new, highly sensitive, selective and simple SERS/RRS/Abs trimode detection platform for OTC was constructed. The linear range of SERS was 0.0625 ng/mL âˆ¼ 1.75 ng/mL and the limit of detection was 0.015 ng/mL. This new nanocatalytic probe detection strategy can also be used for the selective detection of other antibiotics such as tetracycline and doxycycline, respectively. In addition, the nanocatalytic mechanism has been investigated.

Thioredoxins o1 and h2 jointly adjust mitochondrial dihydrolipoamide dehydrogenase-dependent pathways towards changing environments.

Thioredoxins (TRXs) are central to redox regulation, modulating enzyme activities to adapt metabolism to environmental changes. Previous research emphasized mitochondrial and microsomal TRX o1 and h2 influence on mitochondrial metabolism, including photorespiration and the tricarboxylic acid (TCA) cycle. Our study aimed to compare TRX-based regulation circuits towards environmental cues mainly affecting photorespiration. Metabolite snapshots, phenotypes and CO2 assimilation were compared among single and multiple TRX mutants in the wild-type and the glycine decarboxylase T-protein knockdown (gldt1) background. Our analyses provided evidence for additive negative effects of combined TRX o1 and h2 deficiency on growth and photosynthesis. Especially metabolite accumulation patterns suggest a shared regulation mechanism mainly on mitochondrial dihydrolipoamide dehydrogenase (mtLPD1)-dependent pathways. Quantification of pyridine nucleotides, in conjunction with 13C-labelling approaches, and biochemical analysis of recombinant mtLPD1 supported this. It also revealed mtLPD1 inhibition by NADH, pointing at an additional measure to fine-tune it's activity. Collectively, we propose that lack of TRX o1 and h2 perturbs the mitochondrial redox state, which impacts on other pathways through shifts in the NADH/NAD+ ratio via mtLPD1. This regulation module might represent a node for simultaneous adjustments of photorespiration, the TCA cycle and branched chain amino acid degradation under fluctuating environmental conditions.

Cytochrome P450 CYP6EM1 Underpins Dinotefuran Resistance in the Whitefly Bemisia tabaci.

Being a destructive pest worldwide, the whitefly Bemisia tabaci has evolved resistance to neonicotinoid insecticides. The third-generation neonicotinoid dinotefuran has commonly been applied to the control of the whitefly, but its underlying mechanism is currently unknown. On the base of our transcriptome data, here we aim to investigate whether the cytochrome P450 CYP6EM1 underlies dinotefuran resistance in the whitefly. Compared to the susceptible strain, the CYP6EM1 gene was found to be highly expressed in both laboratory and field dinotefuran-resistant populations. Upon exposure to dinotefuran, the mRNA levels of CYP6EM1 were increased. These results demonstrate the involvement of this gene in dinotefuran resistance. Loss and gain of functional studies in vivo were conducted through RNAi and transgenic Drosophila melanogaster assays, confirming the role of CYP6EM1 in conferring such resistance. In a metabolism assay in vitro, the CYP6EM1 protein could metabolize 28.11% of dinotefuran with a possible dinotefuran-dm-NNO metabolite via UPLC-QTOF/MS. Docking of dinotefuran to the CYP6EM1 protein showed a good binding affinity, with an energy of less than -6.0 kcal/mol. Overall, these results provide compelling evidence that CYP6EM1 plays a crucial role in the metabolic resistance of B. tabaci to dinotefuran. Our work provides new insights into the mechanism underlying neonicotinoid resistance and applied knowledge that can contribute to sustainable control of a global pest such as whitefly.

Identification and RNAi-based function analysis of trehalase family genes in Frankliniella occidentalis (Pergande).

BACKGROUND: Insects utilize trehalases (TREs) to regulate energy metabolism and chitin biosynthesis, which are essential for their growth, development, and reproduction. TREs can therefore be used as potential targets for future insecticide development. However, the roles of TREs in Frankliniella occidentalis (Pergande), a serious widespread agricultural pest, remain unclear. RESULTS: Three TRE genes were identified in F. occidentalis and cloned, and their functions were then investigated via feeding RNA interference (RNAi) and virus-induced gene silencing (VIGS) assays. The results showed that silencing FoTRE1-1 or FoTRE1-2 significantly decreased expression levels of FoGFAT, FoPGM, FoUAP, and FoCHS, which are members of the chitin biosynthesis pathway. Silencing FoTRE1-1 or FoTRE2 significantly down-regulated FoPFK and FoPK, which are members of the energy metabolism pathway. These changes resulted in 2-fold decreases in glucose and glycogen content, 2-fold increases in trehalose content, and 1.5- to 2.0-fold decreases in chitinase activity. Furthermore, knocking down FoTRE1-1 or FoTRE1-2 resulted in deformed nymphs and pupae as a result of hindered molting. The VIGS assay for the three FoTREs revealed that FoTRE1-1 or FoTRE2 caused shortened ovarioles, and reduced egg-laying and hatching rates. CONCLUSION: The results suggest that FoTRE1-1 and FoTRE1-2 play important roles in the growth and development of F. occidentalis, while FoTRE1-1 and FoTRE2 are essential for its reproduction. These three genes could be candidate targets for RNAi-based management and control of this destructive agricultural pest. © 2024 Society of Chemical Industry.

Dual mutations in the whitefly nicotinic acetylcholine receptor ß1 subunit confer target-site resistance to multiple neonicotinoid insecticides.

Neonicotinoid insecticides, which target insect nicotinic acetylcholine receptors (nAChRs), have been widely and intensively used to control the whitefly, Bemisia tabaci, a highly damaging, globally distributed, crop pest. This has inevitably led to the emergence of populations with resistance to neonicotinoids. However, to date, there have been no reports of target-site resistance involving mutation of B. tabaci nAChR genes. Here we characterize the nAChR subunit gene family of B. tabaci and identify dual mutations (A58T&R79E) in one of these genes (BTß1) that confer resistance to multiple neonicotinoids. Transgenic D. melanogaster, where the native nAChR Dß1 was replaced with BTß1A58T&R79E, were significantly more resistant to neonicotinoids than flies where Dß1 were replaced with the wildtype BTß1 sequence, demonstrating the causal role of the mutations in resistance. The two mutations identified in this study replace two amino acids that are highly conserved in >200 insect species. Three-dimensional modelling suggests a molecular mechanism for this resistance, whereby A58T forms a hydrogen bond with the R79E side chain, which positions its negatively-charged carboxylate group to electrostatically repulse a neonicotinoid at the orthosteric site. Together these findings describe the first case of target-site resistance to neonicotinoids in B. tabaci and provide insight into the molecular determinants of neonicotinoid binding and selectivity.

Silencing of catalase reduces unfavorable low-temperature tolerance capacity in whiteflies.

BACKGROUND: Temperature is a primary factor that determines the eco-geographical distribution and population development of invasive insects. Temperature stress leads to various negative effects, including excess reactive oxygen species (ROS), and catalase (CAT) is a key enzyme against ROS in the antioxidant pathway. The whitefly Bemisia tabaci MED is a typical invasive pest that causes damage worldwide. Our previous studies have shown that CAT promotes whitefly adaptation to high temperature by eliminating ROS. However, the mechanism underlying the low-temperature adaptation of whiteflies is still unknown. RESULTS: In this study, we investigated the role of CAT in the low-temperature tolerance of B. tabaci MED by analyzing its survival rate, reproduction, and ROS levels at 25 °C (as a control, suitable temperature), 20 °C (moderately decreased temperature), and 4 °C (severely decreased temperature). Silencing of BtCAT1, BtCAT2, or BtCAT3 reduced the viability of whiteflies under a short-term severely decreased temperature (4 °C), which manifested as decreases in survival and fecundity accompanied by significant increases in ROS levels. Moreover, even at a moderately decreased temperature (20 °C), silencing of BtCAT1 led to high ROS levels and low survival rates in adults. CONCLUSION: Silencing of BtCATs significantly increased the sensitivity of B. tabaci MED to low temperatures. BtCAT1 is likely more essential than other BtCATs for low-temperature tolerance in whiteflies. © 2024 Society of Chemical Industry.

Two horizontally acquired bacterial genes steer the exceptionally efficient and flexible nitrogenous waste cycling in whiteflies.

Nitrogen is an essential element for all life on earth. Nitrogen metabolism, including excretion, is essential for growth, development, and survival of plants and animals alike. Several nitrogen metabolic processes have been described, but the underlying molecular mechanisms are unclear. Here, we reveal a unique process of nitrogen metabolism in the whitefly Bemisia tabaci, a global pest. We show that it has acquired two bacterial uricolytic enzyme genes, B. tabaci urea carboxylase (BtUCA) and B. tabaci allophanate hydrolase (BtAtzF), through horizontal gene transfer. These genes operate in conjunction to not only coordinate an efficient way of metabolizing nitrogenous waste but also control B. tabaci's exceptionally flexible nitrogen recycling capacity. Its efficient nitrogen processing explains how this important pest can feed on a vast spectrum of plants. This finding provides insight into how the hijacking of microbial genes has allowed whiteflies to develop a highly economic and stable nitrogen metabolism network and offers clues for pest management strategies.

Uridine diphosphate glucosyltransferases are involved in spinosad resistance in western flower thrips Frankliniella occidentalis (Pergande).

Uridine diphosphate glucosyltransferases (UGTs) play crucial roles in the insect detoxification system and are associated with pesticide resistance. Our previous transcriptomic analysis of spinosad-susceptible (Ivf03) and resistant (NIL-R) Frankliniella occidentalis revealed numerous upregulated UGT genes in the NIL-R strain, suggesting their potential contribution to spinosad resistance. To investigate this hypothesis, here we conducted UGT activity assays and spinosad induction experiments, employing RNA interference (RNAi) techniques for gene function validation. We found significantly elevated UGT activity in the NIL-R strain compared to Ivf03, with 5-nitrouracil showing a substantial synergistic effect on the resistant strain. Eighteen UGT genes were identified in F. occidentalis, with gene expansion and duplication observed within families UGT466, 467, and 468. Ten out of the eighteen UGTs exhibited higher expression levels in NIL-R, specifically FoUGT466B1, FoUGT468A3, and FoUGT468A4 consistently being upregulated across nymphs, males, and females. RNAi-based functional validation targeting these three UGT genes led to increased susceptibility to spinosad in a life stage-, sex-, and dose-dependent manner. These results indicate that UGTs are indeed involved in spinosad resistance in F. occidentalis, and the effects are dependent on life stage, sex, and dose. Therefore, sustainable control for F. occidentalis resistance should always consider these differential responses.

RNA m6 A Methylation Suppresses Insect Juvenile Hormone Degradation to Minimize Fitness Costs in Response to A Pathogenic Attack.

Bioinsecticides and transgenic crops based on the bacterial pathogen Bacillus thuringiensis (Bt) can effectively control diverse agricultural insect pests, nevertheless, the evolution of resistance without obvious fitness costs has seriously eroded the sustainable use of these Bt products. Recently, it has been discovered that an increased titer of juvenile hormone (JH) favors an insect host (Plutella xylostella) to enhance fitness whilst resisting the Bt pathogen, however, the underlying regulatory mechanisms of the increased JH titer are obscure. Here, the involvement of N6 -methyladenosine (m6 A) RNA modification in modulating the availability of JH in this process is defined. Specifically, it is found that two m6 A methyltransferase subunit genes, PxMettl3 and PxMettl14, repress the expression of a key JH-degrading enzyme JH esterase (JHE) to induce an increased JH titer, mitigating the fitness costs associated with a robust defense against the Bt pathogen. This study identifies an as-yet uncharacterized m6 A-mediated epigenetic regulator of insect hormones for maintaining fitness during pathogen defense and unveils an emerging Bt resistance-related m6 A methylation atlas in insects, which further expands the functional landscape of m6 A modification and showcases the pivotal role of epigenetic regulation in host-pathogen interactions.

Dynamic monitoring of the insecticide resistance status of Bemisia tabaci across China from 2019-2021.

BACKGROUND: Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a major agricultural insect pest that causes severe economic losses worldwide. Several insecticides have been applied to effectively control this key pest. However, owing to the indiscriminate use of chemical insecticides, B. tabaci has developed resistance against these chemical compounds over the past several years. RESULTS: From 2019 to 2021, 23 field samples of B. tabaci were collected across China. Twenty species were identified as the Mediterranean 'Q' type (MED) and three were identified as MED/ Middle East-Asia Minor 1 mixtures. Subsequently, resistance of the selected populations to different insecticides was evaluated. The results showed that 13 populations developed low levels of resistance to abamectin. An overall upward trend in B. tabaci resistance toward spirotetramat, cyantraniliprole and pyriproxyfen was observed. In addition, resistance to thiamethoxam remained low-to-moderate in the 23 field populations. CONCLUSION: These findings suggest that the overall resistance of the field-collected B. tabaci populations has shown an upward trend over the years in China. We believe our study can provide basic data to support integrated pest management and insecticide resistance management of field B. tabaci in China. © 2023 Society of Chemical Industry.

CYP4CS5-mediated thiamethoxam and clothianidin resistance is accompanied by fitness cost in the whitefly Bemisia tabaci.

BACKGROUND: Understanding the trade-offs between insecticide resistance and the associated fitness is of particular importance to sustainable pest control. One of the most devastating pest worldwide, the whitefly Bemisia tabaci, has developed resistance to various insecticides, especially the neonicotinoid group. Although neonicotinoid resistance often is conferred by P450s-mediated metabolic resistance, the relationship between such resistance and the associated fitness phenotype remains largely elusive. By gene cloning, quantitative reverse transcription (qRT)-PCR, RNA interference (RNAi), transgenic Drosophila melanogaster, metabolism capacity in vitro and 'two sex-age stage' life table study, this study aims to explore the molecular role of a P450 gene CYP4CS5 in neonicotinoid resistance and to investigate whether such resistance mechanism carries fitness costs in the whitefly. RESULTS: Our bioassay tests showed that a total of 13 field-collected populations of B. tabaci MED biotype displayed low-to-moderate resistance to thiamethoxam and clothianidin. Compared to the laboratory susceptible strain, we then found that an important P450 CYP4CS5 was remarkably upregulated in the field resistant populations. Such overexpression of CYP4CS5 had a good match with the resistance level among the whitefly samples. Further exposure to the two neonicotinoids resulted in an increase in CYP4CS5 expression. These results implicate that overexpression of CYP4CS5 is closely correlated with thiamethoxam and clothianidin resistance. RNAi knockdown of CYP4CS5 increased mortality of the resistant and susceptible populations after treatment with thiamethoxam and clothianidin in bioassay, but obtained an opposite result when using a transgenic line of D. melanogaster expressing CYP4CS5. Metabolic assays in vitro revealed that CYP4CS5 exhibited certain capacity of metabolizing thiamethoxam and clothianidin. These in vivo and in vitro assays indicate an essential role of CYP4CS5 in conferring thiamethoxam and clothianidin resistance in whitefly. Additionally, our life-table analysis demonstrate that the field resistant whitefly exhibited a prolonged development time, shortened longevity and reduced fecundity compared to the susceptible, suggesting an existing fitness cost as a result of the resistance. CONCLUSION: Collectively, in addition to the important role of CYP4CS5 in conferring thiamethoxam and clothianidin resistance, this resistance mechanism is associated with fitness costs in the whitefly. These findings not only contribute to the development of neonicotinoids resistance management strategies, but also provide a new target for sustainable whitefly control. © 2023 Society of Chemical Industry.

Depicting the genetic and metabolic panorama of chemical diversity in the tea plant.

As a frequently consumed beverage worldwide, tea is rich in naturally important bioactive metabolites. Combining genetic, metabolomic and biochemical methodologies, here, we present a comprehensive study to dissect the chemical diversity in tea plant. A total of 2837 metabolites were identified at high-resolution with 1098 of them being structurally annotated and 63 of them were structurally identified. Metabolite-based genome-wide association mapping identified 6199 and 7823 metabolic quantitative trait loci (mQTL) for 971 and 1254 compounds in young leaves (YL) and the third leaves (TL), respectively. The major mQTL (i.e., P < 1.05 × 10-5, and phenotypic variation explained (PVE) > 25%) were further interrogated. Through extensive annotation of the tea metabolome as well as network-based analysis, this study broadens the understanding of tea metabolism and lays a solid foundation for revealing the natural variations in the chemical composition of the tea plant. Interestingly, we found that galloylations, rather than hydroxylations or glycosylations, were the largest class of conversions within the tea metabolome. The prevalence of galloylations in tea is unusual, as hydroxylations and glycosylations are typically the most prominent conversions of plant specialized metabolism. The biosynthetic pathway of flavonoids, which are one of the most featured metabolites in tea plant, was further refined with the identified metabolites. And we demonstrated the further mining and interpretation of our GWAS results by verifying two identified mQTL (including functional candidate genes CsUGTa, CsUGTb, and CsCCoAOMT) and completing the flavonoid biosynthetic pathway of the tea plant.

Bibliometric and literature review of the development of mineral fertilizers.

Mineral fertilizers are a new type of sustainable fertilizers, containing natural ores as the primary raw material with various nutrients and organic matters. This study combines two methods of bibliometric analysis to comprehensively review the progress of mineral fertilizers from 2000 to 2021. The results showed that the research on mineral fertilizers has increased in the past 21 years, especially after 2014. Developed countries studied mineral fertilizers more extensively than developing countries, but some developing countries, such as China and India, are also paying attention to this area in recent years. Chinese Academic of Sciences, Agriculture and Agri-Food Canada, and Chinese Academy of Agricultural Sciences were the main publishing institutions. Nutrient elements, changes in soil properties, and the effects on promoting crop growth were the main contents of the research. Still, such issues as bioremediation, soil environment improvement, and crop resistance are becoming hot spots. The field of mineral fertilizers showed a strong interdisciplinary nature and an increasingly comprehensive research perspective. The goal is that this synthesis will be used as a starting point for a broader study on responsible environmental management and research on improving fertilizer use efficiency.

Design and Synthesis of a Water-Based Nanodelivery Pesticide System for Improved Efficacy and Safety.

Developing an environmentally friendly and safe nanodelivery system is crucial to improve the efficacy of pesticides and minimize environmental and health risks. However, preparing a completely water-based nanopesticide without using harmful solvents is a technical challenge. In this study, a water-based nanodelivery pesticide system was constructed to improve the efficacy and safety of Emamectin Benzoate (EB). A specific surfactant, 29-(4-(5-hydroxynonan-5-yl)phenoxy)-3,6,9,12,15,18,21,24,27-nonaoxanonacosan-1-ol (SurEB) was designed and synthesized to form a water-based nanodelivery system (EBWNS) with EB. Molecular dynamics simulations revealed the self-assembly and interaction forces between SurEB and EB in water, providing insights into the formation mechanism of EBWNS nanoparticles. The nanodelivery system showed the prolonged effectivity of EB with reduced degradation and demonstrated a good control efficacy for multiple target pests, such as red spider mite, beet armyworm larvae (Lepidoptera: Noctuidae), and rice stem borers (Chilo suppressalis). Toxicology tests on various objects demonstrated that the EBWNS has low toxicity for seeds, HaCaT cells, zebrafish, earthworm, and E. coli. This study provides a distinctive perspective for developing environmentally friendly nanopesticide formulations, which clarified a water-based treatment method for specific lipid-soluble pesticides. The water-based nanodelivery pesticide system has the potential to improve the efficacy and safety of pesticides in the process of field applications.

A Horizontally Transferred Plant Fatty Acid Desaturase Gene Steers Whitefly Reproduction.

Polyunsaturated fatty acids (PUFAs) are essential nutrients for all living organisms. PUFA synthesis is mediated by Δ12 desaturases in plants and microorganisms, whereas animals usually obtain PUFAs through their diet. The whitefly Bemisia tabaci is an extremely polyphagous agricultural pest that feeds on phloem sap of many plants that do not always provide them with sufficient PUFAs. Here, a plant-derived Δ12 desaturase gene family BtFAD2 is characterized in B. tabaci and it shows that the BtFAD2-9 gene enables the pest to synthesize PUFAs, thereby significantly enhancing its fecundity. The role of BtFAD2-9 in reproduction is further confirmed by transferring the gene to Drosophila melanogaster, which also increases the fruit fly's reproduction. These findings reveal an extraordinary evolutionary scenario whereby a phytophagous insect acquired a family of plant genes that enables it to synthesize essential nutrients, thereby lessening its nutritional dependency and allowing it to feed and reproduce on many host plants.

Cucumber malate decarboxylase, CsNADP-ME2, functions in the balance of carbon and amino acid metabolism in fruit.

Central metabolism produces carbohydrates and amino acids that are tightly correlated to plant growth and thereby crop productivity. Malate is reported to link mitochondrial respiratory metabolism with cytosolic biosynthetic pathways. Although the function of malate metabolism-related enzymes in providing carbon has been characterized in some plants, evidence for this role in the fleshy fruit of cucumber is lacking. Here, radiolabeled bicarbonate fed into the xylem stream from the cucumber roots was incorporated into amino acids, soluble sugars, and organic acids in the exocarp and vasculature of fruits. The activities of decarboxylases, especially decarboxylation from NADP-dependent malic enzyme (NADP-ME), were higher in cucumber fruit than in the leaf lamina. Histochemical localization revealed that CsNADP-ME2 was mainly located in the exocarp and vascular bundle system of fruit. Radiotracer and gas-exchange analysis indicated that overexpression of CsNADP-ME2 could promote carbon flux into soluble sugars and starch in fruits. Further studies combined with metabolic profiling revealed that the downregulation of CsNADP-ME2 in RNA interference (RNAi) lines caused the accumulation of its substrate, malate, in the exocarp. In addition to inhibition of glycolysis-related gene expression and reduction of the activities of the corresponding enzymes, increased amino acid synthesis and decreased sugar abundance were also observed in these lines. The opposite effect was found in CsNADP-ME2-overexpressing lines, suggesting that there may be a continuous bottom-up feedback regulation of glycolysis in cucumber fruits. Overall, our studies indicate that CsNADP-ME2 may play potential roles in both central carbon reactions and amino acid metabolism in cucumber fruits.