Application of valencene and prospects for its production in engineered microorganisms.

Valencene, a sesquiterpene with the odor of sweet and fresh citrus, is widely used in the food, beverage, flavor and fragrance industry. Valencene is traditionally obtained from citrus fruits, which possess low concentrations of this compound. In the past decades, the great market demand for valencene has attracted considerable attention from researchers to develop novel microbial cell factories for more efficient and sustainable production modes. This review initially discusses the biosynthesis of valencene in plants, and summarizes the current knowledge of the key enzyme valencene synthase in detail. In particular, we highlight the heterologous production of valencene in different hosts including bacteria, fungi, microalgae and plants, and focus on describing the engineering strategies used to improve valencene production. Finally, we propose potential engineering directions aiming to further increase the production of valencene in microorganisms.

Fe-CP-based Catalytic Oxidation and Dissipative Self-Assembly of a Ferrocenyl Surfactant Applied in DNA Capture and Release.

Dissipative self-assembly plays a vital role in fabricating intelligent and transient materials. The selection and design of the molecular structure is critical, and the introduction of valuable stimuli-responsive motifs into building blocks would bring about a novel perspective on the fuel driven nonequilibrium assemblies. For redox-responsive surfactants, novel methods of catalytic oxidation are very important for their activation/deactivation process through designing fuel input/energy dissipation. As an enzyme with a fast catalytic rate, Fe-based coordination polymers (Fe-CPs) are found to be highly effective oxidase-like enzymes to induce a reversible switch of a ferrocene-based surfactant over a wide range of temperatures and pH. This builds a bridge between the CPs materials and surfactants. Furthermore, glucose oxidase can also induce a switchable transition of a ferrocene-based surfactant. The GOX-catalyzed, glucose-fueled transient surfactant assemblies have been fabricated for many cycles, which has a successful application in a time-controlled and autonomous DNA capture and release process. The intelligent use of enzymes including CPs and GOX in ferrocene-based surfactants will pave the way for the oxidation of redox surfactants, which extends the application of stable or transient ferrocenyl self-assemblies.

Synthesis and structure-activity relationship of novel thiazole aminoguanidines against MRSA and Escherichia coli.

Novel lead thiazole aminoguanidines exhibited strong activity against Gram-positive bacteria. The potential targets of these substances are undecaprenyl diphosphate synthase (UPPS) and undecaprenyl diphosphate phosphatase (UPPP). Here, we report the synthesis and antibacterial evaluation of a library of thiazole aminoguanidines analogues, wherein the rotatable bond is inserted between the C2 position of thiazole and hydrophobic group. The molecular flexibility is increased, and new analogues with strong activity against MRSA and E. coli are produced. The best compound 4i showed rapid sterilization and low tendency to induce bacterial resistance. The IC50 of compound 4i to EcUPPS enzyme is 145 µmol L-1 (58 µg mL-1). Compound 4i can also inhibit and destroy bacterial biofilms. These thiazole aminoguanidines can be developed as potential therapeutic candidates in the future.

Nutrient condition modulates the antibiotic tolerance of Pseudomonas aeruginosa.

The variation in nutrient content across diverse environments has a significant impact on the survival and metabolism of microorganisms. In this study, we examined the influence of nutrients on the antibiotic tolerance of the PAO1 strain of Pseudomonas aeruginosa. Our findings indicate that under nutrient-rich conditions, this strain exhibited relatively high tolerance to ceftazidime, chloramphenicol, and tetracycline, but not aminoglycosides and fluoroquinolones. Transcriptome analysis revealed that genes associated with antibiotic tolerance were expressed more efficiently in nutrient-rich media, including ribosomal protein genes and multidrug efflux pump genes, which conferred higher tetracycline tolerance to the strain. Furthermore, the genes responsible for translation, biosynthesis, and oxidative phosphorylation were suppressed when nutrients were limited, resulting in decreased metabolic activity and lower sensitivity to ciprofloxacin. Artificial interference with ATP synthesis utilizing arsenate confirmed that the curtailment of energy provision bolstered the observed tolerance to ciprofloxacin. In general, our results indicate that this strain of P. aeruginosa tends to activate its intrinsic resistance mechanisms in nutrient-rich environments, thereby enhancing resistance to certain antibiotics. Conversely, in nutrient-limited environments, the strain is more likely to enter a dormant state, which enables it to tolerate antibiotics to which it would otherwise be sensitive. These findings further suggest that antibiotics released in environments with varying eutrophication levels may have divergent effects on the development of bacterial antibiotic resistance.

Research on optimization of control parameters of gravity shaking table.

When image processing and machine vision technology are used to extract features from the image of the ore belt of the shaking table, so as to realize the analysis of the processing indictors and mapping of control parameters. To realize the adaptive optimization of the multiple control parameters of the shaking table, it is necessary to have thorough access to the parameters of the internal and external properties of the gravity shaker, such as internal control parameters and external ore zone characteristics, as well as the processing indicators. In this study, information on the multi-scale characteristics of the zone is obtained through a visual experimental system, and the data-driven model of the separation process is constructed to characterize the relationship between the properties of the internal and external parameters of the shaking table, eventually, an adaptive optimization method of control parameters of the shaking table based on maximizing beneficiation efficiency is proposed. The research results show that the data from the geometric characteristics of the ore belts obtained from practical experiments all satisfy the statistical distribution requirements. In the three optimized support vector regression (SVR) models, the sparrow search algorithm optimized SVR (SSA-SVR) has the best comprehensive performance, which overcomes the limits of data samples under objective conditions and basically meets the existing industrial requirements. With these helps, the proposed optimization method has realized the continuous optimization of multiple control parameters of the shaking table, and the optimization results have a good guarantee.

Outer membrane protein of OmpF contributes to swimming motility, biofilm formation, osmotic response as well as the transcription of maltose metabolic genes in Citrobacter werkmanii.

Bacterial outer membrane proteins (Omps) are essential for environmental sensing, stress responses, and substance transport. Our previous study discovered that OmpA contributes to planktonic growth, biocide resistance, biofilm formation, and swimming motility in Citrobacter werkmanii, whereas the molecular functions of OmpF in this strain are largely unknown. Thus, in this study, the ompF gene was firstly knocked out from the genome of C. werkmanii using a homologous recombination method, and its phenotypical alternations of ∆ompF were then thoroughly characterized using biochemical and molecular approaches with the parental wild type (WT) and complementary (∆ompF-com) strains. The results demonstrated that the swimming ability of ∆ompF on semi-solid plates was reduced compared to WT due to the down-regulation of flgC, flgH, fliK, and fliF. Meanwhile, ompF deletion reduces biofilm formation on both glass and polystyrene surfaces due to decreased cell aggregation. Furthermore, ompF inactivation induced different osmotic stress (carbon sources and metal ions) responses in its biofilms when compared to WT and ∆ompF-com. Finally, a total of 6 maltose metabolic genes of lamB, malE, malK, malG, malM, and malF were all up-regulated in ∆ompF. The gene knockout and HPLC results revealed that the MalEFGK2 cluster was primarily responsible for maltose transport in C. werkmanii. Furthermore, we discovered for the first time that the upstream promoter of OmpF and its transcription can be combined with and negatively regulated by MalT. Overall, OmpF plays a role in a variety of biochemical processes and molecular functions in C. werkmanii, and it may even act as a targeted site to inhibit biofilm formation.

Optimization of dewatering process of concentrate pressure filtering by support vector regression.

This work studies the mechanism and optimization methods of the filter press dehydration process to better improve the efficiency of the concentrate filter press dehydration operation. Machine learning (ML) models of radial basis function (RBF)-OLS, RBF-generalized regression neural network, and support vector regression (SVR) are constructed, and laboratory and industrial simulations are performed separately, finally, optimization methods for the filtration dewatering process are designed and applied. In laboratory, all the machine learning models have obvious mistakes, but it can be seen that SVR has the best simulation effect. In order to achieve the optimization of the entire filtration and dewatering process, we obtained enough data from the industrial filtration and dewatering system, and in the industrial simulation results all the machine learning models performed considerably, SVR achieves the best accuracy in industrial simulation, and the simulated mean relative error of moisture and processing capacity are 1.57% and 3.81%, the model was tested with newly collected industrial data to verify the credibility. The optimal simulation results are obtained by optimization method based on control variables. Results show that the ML method of SVR and optimization methods of control variables applied to the industry not only can save energy consumption and cost but also can improves the efficiency of filter press operation fundamentally, which will provide some options for intelligent dewatering process and other industrial production optimization.

Pseudomonas aeruginosa heteroresistance to levofloxacin caused by upregulated expression of essential genes for DNA replication and repair.

Pseudomonas aeruginosa (P. aeruginosa), a common cause of severe chronic infections, has developed heteroresistance to several antibiotics, thus hindering successful treatment. In this study, we aimed to investigate the characteristics and mechanisms underlying levofloxacin (LVX) heteroresistance in P. aeruginosa PAS71 and PAS81 clinical isolates using a combination of physiological and biochemical methods, bacterial genomics, transcriptomics, and qRT-PCR. The six P. aeruginosa strains, namely PAS71, PAS72, PAS81, PAS82, ATCC27853, and PAO1, were studied. The Kirby-Bauer (K-B), minimum inhibitory concentration (MIC) test, and population analysis profile (PAP) experimental results showed that PAS71, PAS81, ATCC27853, and PAO1 were heteroresistant to LVX, with MIC of 0.25, 1, 0.5, and 2 µg/ml, respectively; PAS72 and PAS82 were susceptible to LVX with a MIC of 0.25 and 0.5 µg/ml, respectively. The resistance of PAS71 and PAS81 heteroresistant subpopulations was unstable and had a growth fitness cost. Genomic and transcriptomic results proved that the unstable heteroresistance of PAS71 and PAS81 was caused by elevated expression of essential genes involved in DNA replication and repair, and homologous recombination, rather than their genomic single-nucleotide polymorphism (SNP) and insertion-deletion (InDel) mutations. Additionally, PAS71 and PAS81 enhanced virulence and physiological metabolism, including bacterial secretion systems and biosynthesis of siderophore group nonribosomal peptides, in response to LVX stress. Our results suggest that the upregulation of key genes involved in DNA replication and repair, and homologous recombination causes unstable heteroresistance in P. aeruginosa against LVX. This finding provides novel insights into the occurrence and molecular regulation pathway of P. aeruginosa heteroresistant strains.

Metagenomic analysis of microbial communities and antibiotic resistance genes in spoiled household chemicals.

Numerous attempts have been utilized to unveil the occurrences of antibiotic resistance genes (ARGs) in human-associated and non-human-associated samples. However, spoiled household chemicals, which are usually neglected by the public, may be also a reservoir of ARGs because of the excessive and inappropriate uses of industrial drugs. Based upon the Comprehensive Antibiotic Research Database, a metagenomic sequencing method was utilized to detect and quantify Antibiotic Resistance Ontology (AROs) in six spoiled household chemicals, including hair conditioner, dishwashing detergent, bath shampoo, hand sanitizer, and laundry detergent. Proteobacteria was found to be the dominant phylum in all the samples. Functional annotation of the unigenes obtained against the KEGG pathway, eggNOG and CAZy databases demonstrated a diversity of their functions. Moreover, 186 types of AROs that were members of 72 drug classes were identified. Multidrug resistance genes were the most dominant types, and there were 17 AROs whose resistance mechanisms were categorized into the resistance-nodulation-cell division antibiotic efflux pump among the top 20 AROs. Moreover, Proteobacteria was the dominant carrier of AROs with the primary resistance mechanism of antibiotic efflux. The maximum temperature of the months of collection significantly affected the distributions of AROs. Additionally, the isolated individual bacterium from spoiled household chemicals and artificial mixed communities of isolated bacteria demonstrated diverse resistant abilities to different biocides. This study demonstrated that there are abundant microorganisms and a broad spectrum profile of AROs in spoiled household chemicals that might induce a severe threat to public healthy securities and merit particular attention.

The Connection between Czc and Cad Systems Involved in Cadmium Resistance in Pseudomonas putida.

Heavy metal pollution is widespread and persistent, and causes serious harm to the environment. Pseudomonas putida, a representative environmental microorganism, has strong resistance to heavy metals due to its multiple efflux systems. Although the functions of many efflux systems have been well-studied, the relationship between them remains unclear. Here, the relationship between the Czc and Cad systems that are predominantly responsible for cadmium efflux in P. putida KT2440 is identified. The results demonstrated that CzcR3, the response regulator of two-component system CzcRS3 in the Czc system, activates the expression of efflux pump genes czcCBA1 and czcCBA2 by directly binding to their promoters, thereby helping the strain resist cadmium stress. CzcR3 can also bind to its own promoter, but it has only a weak regulatory effect. The high-level expression of czcRS3 needs to be induced by Cd2+, and this relies on the regulation of CadR, a key regulator in the Cad system, which showed affinity to czcRS3 promoter. Our study indicates that the Cad system is involved in the regulation of the Czc system, and this relationship is important for maintaining the considerable resistance to cadmium in P. putida.

Nanoemulsion fluorescent inks for anti-counterfeiting encryption with dual-mode, full-color, and long-term stability.

An oil-in-water nanoemulsion (O/W NE) is selected as the carrier to encapsulate hydrophobic dual-mode luminescent upconversion nanoparticles (UC NPs) and downconversion (DC) carbon quantum dots (CQDs) inside the oil droplets for forming water-based fluorescent inks. The NE inks conform well to the requirements of inkjet printing for anti-counterfeiting encryption applications.

Correction: Cepharanthine hydrochloride reverses the mdr1 (P-glycoprotein)-mediated esophageal squamous cell carcinoma cell cisplatin resistance through JNK and p53 signals.

[This corrects the article DOI: 10.18632/oncotarget.22676.].

Pseudomonas putida actively forms biofilms to protect the population under antibiotic stress.

Antibiotics are frequently used for clinical treatment and by the farming industry, and most of these are eventually released into the surrounding environment. The impact of these antibiotic pollutants on environmental microorganisms is a concern. The present study showed that after Pseudomonas putida entered the logarithmic growth phase, tetracycline strongly stimulated its biofilm formation in a dose-dependent manner. This was supported by the increased expression of the key adhesin gene lapA in response to tetracycline treatment. Tetracycline treatment also changed the expression levels of the exopolysaccharide gene clusters alg, bcs and pea and the adhesin gene lapF. However, these genes did not participate in the tetracycline-induced biofilm formation. When a biofilm had been established, the P. putida population became more tolerant to tetracycline. Confocal laser scanning microscopic images showed that the interior of the biofilm provided favorable conditions that protected bacterial cells from tetracycline. Besides, biofilm formation of P. putida was also promoted by several other antibiotics, including oxytetracycline, fluoroquinolones, rifampicin, and imipenem, but not aminoglycosides. Susceptibility tests suggested that biofilm conferred a higher tolerance on P. putida cells to specific antibiotics (e.g., tetracyclines and fluoroquinolones). These antibiotics exerted a stronger inducing effect on biofilm formation. Together, our results indicate that P. putida actively forms robust biofilms in response to antibiotic stress, and the biofilms improve the survival of bacterial population under such stress.

Solvent Vapor Strengthened Polyimide Nanofiber-Based Aerogels with High Resilience and Controllable Porous Structure.

Owing to the hierarchically three-dimensional (3D) network, ultralow density, and high porosity, nanofiber-based aerogels (NFAs) have drawn great attention recently. However, precise control of the porous structure and mechanical properties of NFAs, which have been proved to be extremely essential to the applications, still remains a major challenge. Herein, electrospun polyimide (PI) nanofibers were utilized as building blocks to construct NFAs through the solid-templating technique. The porous structure of PI nanofiber-based aerogels (PI-NFAs) could be adjusted by changing the processing parameters. By further welding the adjacent nanofibers at the contact sites with solvent vapor, high-resilience PI-NFAs were successfully prepared with comparable or higher recoverable, under compression, folding and torsion relative to other NFAs. The welded PI-NFAs showed ultralow density (minimum of 0.96 mg/cm3), high porosity (maximum of 99.93%), and tunable hierarchical structure. Therefore, this study brought a new perspective on the simple preparation of high-resilience nanofiber-based aerogels with tunable porous structures.

Preparation of PI/PTFE-PAI Composite Nanofiber Aerogels with Hierarchical Structure and High-Filtration Efficiency.

Electrospun nanofiber, showing large specific area and high porosity, has attracted much attention across various fields, especially in the field of air filtration. The small diameter contributes to the construction of filters with high-filtration efficiency for fine particulate matter (PM), however, along with an increase in air resistance. Herein, composited nanofiber aerogels (NAs), a truly three-dimensional (3D) derivative of the densely compacted electrospun mat, were constructed with the blocks of polytetrafluoroethylene-polyamideimide (PTFE-PAI) composite nanofiber and polyimide (PI) nanofiber. PI/PTFE-PAI NAs with hierarchically porous architecture and excellent mechanical properties have been obtained by thermally induced crosslink bonding. Results indicated that sintering at 400 °C for 30 min could complete the decomposition of polyethylene (PEO) and imidization of polyamic acid (PAA) into PI, as well as generate sufficient mechanical bonding between adjacent nanofibers in the NAs without extra additive. The well-prepared PI/PTFE-PAI NAs could withstand high temperature up to 500 °C. In addition, the filtration tests illustrated that the composite NAs had an excellent performance in PM filtration. More importantly, the filtration behavior could be adjusted to meet the requirements of various applications. The excellent thermal stability and high-filtration efficiency indicated its great potential in the field of high-temperature air filtration.

DNA thermotropic liquid crystals controlled by positively charged catanionic bilayer vesicles.

We report DNA thermotropic liquid crystal (TLC) formation by positively charged catanionic surfactant bilayer vesicles. The properties of DNA TLCs were found to be manipulated by both the chemical structures of cationic and anionic surfactants and the DNA amount. Positively charged catanionic bilayer vesicles bond to negative DNA sites resulting in the transition from vesicles to long range ordered lamellar crystals of DNA-catanionic surfactants, as confirmed by cryo- and freeze-fracture (FF) TEM observations and small-angle X-ray scattering (SAXS) measurements.

A crosstalk between c-di-GMP and cAMP in regulating transcription of GcsA, a diguanylate cyclase involved in swimming motility in Pseudomonas putida.

The ubiquitous bacterial second messenger c-di-GMP is synthesized by diguanylate cyclase (DGC) and degraded by phosphodiesterase (PDE). Pseudomonas putida has dozens of DGC/PDE-encoding genes in its genome, but the phenotypical-genotypical correlation and transcriptional regulation of these genes are largely unknown. Herein, we characterize function and transcriptional regulation of a P. putida c-di-GMP-metabolizing enzyme, GcsA. GcsA consists of two per-ARNT-sim (PAS) domains, followed by a canonical conserved central sequence pattern (GGDEF) domain and a truncated EAL domain. In vitro analysis confirmed the DGC activity of GcsA. The phenotypic observation revealed that GcsA inhibited swimming motility in an FlgZ-dependent manner. In terms of transcriptional regulation, gcsA was found to be cooperatively regulated by c-di-GMP and cAMP via their effectors, FleQ and Crp respectively. The transcription of gcsA was promoted by c-di-GMP and inhibited by cAMP. In vitro binding analysis revealed that FleQ indirectly regulated the transcription of gcsA, while Crp directly regulated the transcription of gcsA by binding to its promoter. Besides, an inverse relationship between the cellular c-di-GMP and cAMP levels in P. putida was confirmed. These findings provide basic knowledge regarding the function and transcriptional regulation of GcsA and demonstrate a crosstalk between c-di-GMP and cAMP in the regulation of the expression of GcsA in P. putida.

Phenotypic-genotypic analysis of GGDEF/EAL/HD-GYP domain-encoding genes in Pseudomonas putida.

Cyclic diguanylate (c-di-GMP) is a broadly conserved bacterial signalling molecule that modulates diverse cellular processes, such as biofilm formation, colony morphology and swimming motility. The intracellular level of c-di-GMP is controlled by diguanylate cyclases (DGCs) with GGDEF domain and phosphodiesterases (PDEs) with either EAL or HD-GYP domain. Pseudomonas putida KT2440 has a large group of genes on its genome encoding proteins with GGDEF/EAL/HD-GYP domains. However, phenotypic-genotypic correlation and c-di-GMP metabolism of these genes were largely unknown. Herein, by systematically constructing deletion mutants/overexpression strains of the 42 predicted c-di-GMP metabolism-related genes and analysing the phenotypes, we preliminarily revealed the role of each gene in biofilm formation, colony morphology and swimming motility. Subsequent results from protein sequence alignments and cellular c-di-GMP assessment indicated that 25 out of the 42 genes were likely to encode DGCs, nine genes were predicted to encode PDEs, four genes encoded bifunctional enzymes and the other four genes encoded enzymatically inactive proteins. This study offers a basic understanding of the roles of these 42 genes and can serve as a toolkit for investigators to further elucidate the functions of these GGDEF and EAL/HD-GYP domain-containing proteins.

Magnetic and Biocompatible Fullerenol/Fe(III) Microcapsules with Antioxidant Activities.

Fullerene C60 (refers to C60 hereafter) has a unique three-dimensional architecture and intriguing physicochemical properties. It has great potential applications in materials chemistry and life science. However, a big obstacle for the widespread application of C60 lies in the limited strategies to make supramolecular structures with diverse morphologies and functions. Herein, we report a strategy to prepare C60-based, magnetic microcapsules which can be used as external antioxidants to effectively attenuate oxidative stress. The microcapsules are composed of fullerenol, a highly water-soluble C60 multiadduct, and iron ions (Fe3+) released from a rusty nail. They can be easily obtained through coordination between the hydrophilic functional groups in fullerenol and Fe3+ with polystyrene microspheres as templates. The fullerenol/Fe3+ microcapsules have good colloidal stability both in water and serum. Their biocompatibility has been confirmed by in vitro tests on HEK293 and Hela cells. Electron spin resonance measurements indicate that the fullerenol/Fe3+ microcapsules can effectively scavenge hydroxyl radicals (OH·-) produced by H2O2, which greatly improves the living environment of the cells. The fullerenol/Fe3+ microcapsules exhibit ferromagnetic properties and can respond to the external magnetic field, enabling magnetic manipulation, and/or separation in practical applications.

Copper(ii) complexes with NNO ligands: synthesis, crystal structures, DNA cleavage, and anticancer activities.

Three novel copper(ii) complexes, Cu(L1)2 (1), Cu(L2)2·2DMF (2), and Cu(L3)2·2DMF (3), were synthesized using three aroylhydrazone ligands, (E)-2-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide (HL1), (E)-3-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide (HL2) and (E)-4-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide (HL3). The complexes were characterized by elemental analysis, infrared (IR), and Ultraviolet-visible light (UV-vis) spectroscopy. The X-ray crystal structures of the complexes all possess a distorted octahedral coordination geometry. Both an absorption spectral titration and a competitive binding assay (ethidium bromide, 4',6-diamidino-2-phenylindole (DAPI), and methyl green) revealed that complexes 2 and 3 bind readily to calf thymus DNA (ctDNA) through intercalative and minor groove binding modes. Complexes 2 and 3 also exhibited oxidative cleavage of supercoiled plasmid DNA (pUC19) in the presence of ascorbic acid as an activator. Cytotoxicity studies showed that complexes 2 and 3 possessed high cytotoxicities toward the HeLa human cervical cancer cell line, but weak toxicities toward the L929 normal mouse fibroblast cell line. We therefore have reason to believe that complexes 2 and 3 both show potential as promising anticancer candidate drugs.