ppGpp is a dual-role regulator involved in balancing iron absorption and prodiginine biosynthesis in Pseudoalteromonas.

Iron is an essential element for microbial survival and secondary metabolism. However, excess iron availability and overloaded secondary metabolites can hinder microbial growth and survival. Microorganisms must tightly control iron homeostasis and secondary metabolism. Our previous studies have found that the stringent starvation protein A (SspA) positively regulates prodiginine biosynthesis by activating iron uptake in Pseudoalteromonas sp. strain R3. It is believed that the interaction between SspA and the small nucleotide ppGpp is important for iron to exert regulation functions. However, the roles of ppGpp in iron absorption and prodiginine biosynthesis, and the underlying relationship between ppGpp and SspA in strain R3 remain unclear. In this study, we found that ppGpp accumulation in strain R3 could be induced by limiting iron. In addition, ppGpp not only positively regulated iron uptake and prodiginine biosynthesis via increasing the SspA level but also directly repressed iron uptake and prodiginine biosynthesis independent of SspA, highlighting the finding that ppGpp can stabilize both iron levels and prodiginine production. Notably, the abolishment of ppGpp significantly increased prodiginine production, thus providing a theoretical basis for manipulating prodiginine production in the future. This dynamic ppGpp-mediated interaction between iron uptake and prodiginine biosynthesis has significant implications for understanding the roles of nutrient uptake and secondary metabolism for the survival of bacteria in unfavorable environments.

Development of a Quorum Sensing-Mediated Bacterial Autolytic System in Escherichia coli for Automatic Release of Intracellular Products.

Escherichia coli, one of the most efficient expression hosts for recombinant proteins, is widely used in chemical, medical, food, and other industries. De novo engineering of gene regulation circuits and cell density-controlled E. coli cell lysis are promising directions for the release of intracellular bioproducts. Here, we developed an E. coli autolytic system, named the quorum sensing-mediated bacterial autolytic (QS-BA) system, by incorporating an acyl-homoserine lactone (AHL)-based YasI/YasR-type quorum sensing circuit from Pseudoalteromonas into E. coli cells. The results showed that the E. coli QS-BA system can release the intracellular bioproducts into the cell culture medium in terms of E. coli cell density, which offers an environmentally-friendly, economical, efficient, and flexible E. coli lysis platform for production of recombinant proteins. The QS-BA system has the potential to serve as an integrated system for the large-scale production of target products in E. coli for medical and industrial applications.

Biosynthetic mechanism of the yellow pigments in the marine bacterium Pseudoalteromonas sp. strain T1lg65.

The Pseudoalteromonas genus marine bacteria have attracted increasing interest because of their abilities to produce bioactive metabolites. The pigmented Pseudoalteromonas group encodes more secondary metabolite biosynthetic gene clusters (BGCs) than the non-pigmented group. Here, we report a yellow pigmented bacterium Pseudoalteromonas sp. strain T1lg65, which was isolated from a mangrove forest sediment. We showed that the yellow pigments of T1lg65 belong to the group of lipopeptide alterochromides. Further genetic analyses of the alterochromide BGC revealed that the yellow pigments are biosynthesized by aryl-polyene synthases and nonribosomal peptide synthases. Within the gene cluster, altA encodes a tyrosine ammonia acid lyase, which catalyzes synthesis of the precursor 4-hydroxycinnamic acid (4-HCA) from tyrosine in the alterochromide biosynthetic pathway. In addition, altN, encoding a putative flavin-dependent halogenase, was proven to be responsible for the bromination of alterochromides based on gene deletion, molecular docking, and site mutagenesis analyses. In summary, the biosynthetic pathway, precursor synthesis, and bromination mechanism of the lipopeptide alterochromides were studied in-depth. Our results expand the knowledge on biosynthesis of Pseudoalteromonas pigments and could promote the development of active pigments in the future.IMPORTANCEThe marine bacteria Pseudoalteromonas spp. are important biological resources because they are producers of bioactive natural products, including antibiotics, pigments, enzymes, and antimicrobial peptides. One group of the microbial pigments, alterochromides, holds a great value for their novel lipopeptide structures and antimicrobial activities. Previous studies were limited to the structural characterization of alterochromides and genome mining for the alterochromide biosynthesis. This work focused on the biosynthetic mechanism for alterochromide production, especially revealing functions of two key genes within the gene cluster for the alterochromide biosynthesis. On the one hand, our study provides a target for metabolic engineering of the alterochromide biosynthesis; on the other hand, the 4-HCA synthase AltA and brominase AltN show potential in the biocatalyst industry.

Efficient degradation of aqueous dichloromethane by an enhanced microbial electrolysis cell: Degradation kinetics, microbial community and metabolic mechanisms.

Dichloromethane (DCM) has been listed as a toxic and harmful water pollutant, and its removal needs attention. Microbial electrolysis cells (MECs) are viewed as a promising alternative for pollutant removal, which can be strengthened from two aspects: microbial inoculation and acclimation. In this study, the MEC for DCM degradation was inoculated with the active sludge enhanced by Methylobacterium rhodesianum H13 (strain H13) and then acclimated in the form of a microbial fuel cell (MFC). Both the introduction of strain H13 and the initiation in MFC form significantly promoted DCM degradation. The degradation kinetics were fitted by the Haldane model, with Vmax, Kh, Ki and vmax values of 103.2 mg/L/hr, 97.8 mg/L, 268.3 mg/L and 44.7 mg/L/hr/cm2, respectively. The cyclic voltammogram implies that DCM redox reactions became easier with the setup of MEC, and the electrochemical impedance spectrogram shows that the acclimated and enriched microbes reduced the charge transfer resistance from the electrode to the electrolyte. In the biofilm, the dominant genera shifted from Geobacter to Hyphomicrobium in acclimation stages. Moreover, Methylobacterium played an increasingly important role. DCM metabolism mainly occurred through the hydrolytic glutathione S-transferase pathway, given that the gene dcmA was identified rather than the dhlA and P450/MO. The exogenous electrons facilitated the reduction of GSSG, directly or indirectly accelerating the GSH-catalyzed dehalogenation. This study provides support for the construction of an efficient and stable MEC for DCM removal in water environment.

Deep Q-Learning-Based Buffer-Aided Relay Selection for Reliable and Secure Communications in Two-Hop Wireless Relay Networks.

This paper investigates the problem of buffer-aided relay selection to achieve reliable and secure communications in a two-hop amplify-and-forward (AF) network with an eavesdropper. Due to the fading of wireless signals and the broadcast nature of wireless channels, transmitted signals over the network may be undecodable at the receiver end or have been eavesdropped by eavesdroppers. Most available buffer-aided relay selection schemes consider either reliability or security issues in wireless communications; rarely is work conducted on both reliability and security issues. This paper proposes a buffer-aided relay selection scheme based on deep Q-learning (DQL) that considers both reliability and security. By conducting Monte Carlo simulations, we then verify the reliability and security performances of the proposed scheme in terms of the connection outage probability (COP) and secrecy outage probability (SOP), respectively. The simulation results show that two-hop wireless relay network can achieve reliable and secure communications by using our proposed scheme. We also performed comparison experiments between our proposed scheme and two benchmark schemes. The comparison results indicate that our proposed scheme outperforms the max-ratio scheme in terms of the SOP.

Discovery of Subcellular-Targeted Aza-BODIPY Photosensitizers for Efficient Photodynamic Antitumor Therapy.

In this study, we linked classical organelle-targeting groups, such as triphenylphosphonium, pentafluorobenzene, and morpholine, to our previously reported potent monoiodo Aza-BODIPY photosensitizer (BDP-15). They were conveniently prepared and retained the advantages of Aza-BODIPY PS with intense NIR absorption, moderate quantum yield, potent photosensitizing efficiency, and good stability. The in vitro antitumor assessment indicated that mitochondria-targeting and lysosome-targeting groups were more effective than ER-targeting groups. Considering undesirable dark toxicity of triphenylphosphonium-modified PSs, compound 6 containing amide-linked morpholine possessed a favorable dark/phototoxicity ratio (>6900 for tumor cells) and was localized in lysosomes with Pearson's coefficient of 0.91 to Lyso-Tracker Green DND-26. 6 exhibited significantly increased intracellular ROS production and resulted in early/late apoptosis and necrosis to disrupt tumor cells. Moreover, in vivo antitumor efficacy exploration suggested that even under a slightly low dose of light (30 J/cm2) and single-time photoirradiation, 6 retarded tumor growth dramatically and displayed much better PDT activity over BDP-15 and Ce6.

A FadR-Type Regulator Activates the Biodegradation of Polycyclic Aromatic Hydrocarbons by Mediating Quorum Sensing in Croceicoccus naphthovorans Strain PQ-2.

Bacteria employ multiple transcriptional regulators to orchestrate cellular responses to adapt to constantly varying environments. The bacterial biodegradation of polycyclic aromatic hydrocarbons (PAHs) has been extensively described, and yet, the PAH-related transcriptional regulators remain elusive. In this report, we identified an FadR-type transcriptional regulator that controls phenanthrene biodegradation in Croceicoccus naphthovorans strain PQ-2. The expression of fadR in C. naphthovorans PQ-2 was induced by phenanthrene, and its deletion significantly impaired both the biodegradation of phenanthrene and the synthesis of acyl-homoserine lactones (AHLs). In the fadR deletion strain, the biodegradation of phenanthrene could be recovered by supplying either AHLs or fatty acids. Notably, FadR simultaneously activated the fatty acid biosynthesis pathway and repressed the fatty acid degradation pathway. As intracellular AHLs are synthesized with fatty acids as substrates, boosting the fatty acid supply could enhance AHL synthesis. Collectively, these findings demonstrate that FadR in C. naphthovorans PQ-2 positively regulates PAH biodegradation by controlling the formation of AHLs, which is mediated by the metabolism of fatty acids. IMPORTANCE Master transcriptional regulation of carbon catabolites is extremely important for the survival of bacteria that face changes in carbon sources. Polycyclic aromatic hydrocarbons (PAHs) can be utilized as carbon sources by some bacteria. FadR is a well-known transcriptional regulator involved in fatty acid metabolism; however, the connection between FadR regulation and PAH utilization in bacteria remains unknown. This study revealed that a FadR-type regulator in Croceicoccus naphthovorans PQ-2 stimulated PAH biodegradation by controlling the biosynthesis of the acyl-homoserine lactone quorum-sensing signals that belong to fatty acid-derived compounds. These results provide a unique perspective for understanding bacterial adaptation to PAH-containing environments.

[Advances in bacterial adsorption and transport of aromatic compounds].

Aromatic compounds are a class of organic compounds with benzene ring(s). Aromatic compounds are hardly decomposed due to its stable structure and can be accumulated in the food cycle, posing a great threat to the ecological environment and human health. Bacteria have a strong catabolic ability to degrade various refractory organic contaminants (e.g., polycyclic aromatic hydrocarbons, PAHs). The adsorption and transportation are prerequisites for the catabolism of aromatic compounds by bacteria. While remarkable progress has been made in understanding the metabolism of aromatic compounds in bacterial degraders, the systems responsible for the uptake and transport of aromatic compounds are poorly understood. Here we summarize the effect of cell-surface hydrophobicity, biofilm formation, and bacterial chemotaxis on the bacterial adsorption of aromatic compounds. Besides, the effects of outer membrane transport systems (such as FadL family, TonB-dependent receptors, and OmpW family), and inner membrane transport systems (such as major facilitator superfamily (MFS) transporter and ATP-binding cassette (ABC) transporter) involved in the membrane transport of these compounds are summarized. Moreover, the mechanism of transmembrane transport is also discussed. This review may serve as a reference for the prevention and remediation of aromatic pollutants.

Stringent Starvation Protein SspA and Iron Starvation Sigma Factor PvdS Coordinately Regulate Iron Uptake and Prodiginine Biosynthesis in Pseudoalteromonas sp. R3.

Organisms need sufficient intracellular iron to maintain biological processes. However, cells can be damaged by excessive iron-induced oxidation stress. Therefore, iron homeostasis must be strictly regulated. In general, bacteria have evolved complex mechanisms to maintain iron homeostasis. In this study, we showed that Pseudoalteromonas sp. R3 has four sets of iron uptake systems. Among these, the siderophore pyoverdine-dependent iron uptake system and the ferrous iron transporter Feo system are more important for iron uptake and prodiginine biosynthesis. Stringent starvation protein SspA positively controls iron uptake and iron-dependent prodiginine biosynthesis by regulating the expression of all iron uptake systems. In turn, the expression of SspA can be induced and repressed by extracellular iron deficiency and excess, respectively. Interestingly, extracytoplasmic function sigma factor PvdS also regulates iron uptake and prodiginine production and responds to extracellular iron levels, exhibiting a similar phenomenon as SspA. Notably, not only do SspA and PvdS function independently, but they can also compensate for each other, and their expression can be affected by the other. All of these findings demonstrate that SspA and PvdS coordinate iron homeostasis and prodiginine biosynthesis in strain R3. More importantly, our results also showed that SspA and PvdS homologs in Pseudomonas aeruginosa PAO1 have similar functions in iron uptake to their counterparts in Pseudoalteromonas, suggesting that coordination between SspA and PvdS on iron homeostasis could be conserved in typical Gram-negative bacteria. Since master regulation of iron homeostasis is extremely important for cell survival, this cross talk between SspA and PvdS may be environmentally significant. IMPORTANCE Both deficiency and excess of intracellular iron can be harmful, and thus, the iron homeostasis needs to be tightly regulated in organisms. At present, the ferric uptake regulator (Fur) is the best-characterized regulator involved in bacterial iron homeostasis, while other regulators of iron homeostasis remain to be further explored. Here, we demonstrated that the stringent starvation protein SspA and the extracytoplasmic function sigma factor PvdS coordinate iron uptake and iron-dependent prodiginine biosynthesis in Pseudoalteromonas sp. R3. These two regulators work independently, but their functions can compensate for the other and their expression can be affected by the other. Moreover, their expression can be activated and repressed by extracellular iron deficiency and excess, respectively. Notably, SspA and PvdS homologs in Pseudomonas aeruginosa PAO1 exhibit similar functions in iron uptake to their counterparts in Pseudoalteromonas, suggesting that this novel fine-tuned mode of iron homeostasis could be conserved in typical Gram-negative bacteria.

The stringent starvation protein SspA modulates peptidoglycan synthesis by regulating the expression of peptidoglycan synthases.

The peptidoglycan (PG) layer of bacterial cells is essential for maintaining the cell shape and survival of cells; therefore, the synthesis of PG needs to be spatiotemporally controlled. While it is well established that PG synthesis is mediated posttranslationally through interactions between PG synthases and their cognate partners, much less is known about the transcriptional regulation of genes encoding these synthases. Based on a previous finding that the Gram-negative bacterium Shewanella oneidensis lacking the prominent PG synthase exhibits impaired cell wall integrity, we performed genetic selections to isolate the suppressors. We discovered that disrupting the sspA gene encoding stringent starvation protein A (SspA) is sufficient to suppress compromised PG. SspA serves as a transcriptional repressor that regulates the expression of the two types of PG synthases, class A penicillin-binding proteins and SEDS/bPBP protein complexes. SspA is an RNA polymerase-associated protein, and its regulation involves interactions with the σ70 -RNAP complex and an antagonistic effect of H-NS, a global nucleoid-associated protein. We also present evidence that the regulation of PG synthases by SspA is conserved in Escherichia coli, adding a new dimension to the current understanding of PG synthesis and its regulation.

Development of Whole-Cell Biosensors for Screening of Peptidoglycan-Targeting Antibiotics in a Gram-Negative Bacterium.

There is an urgent need to develop novel antibiotics since antibiotic resistance is an increasingly serious threat to global public health. Whole-cell biosensors are one of the promising strategies for new antibiotic discovery. The peptidoglycan (PG) of the bacterial cell wall is one of the most important targets for antibiotics. However, the biosensors for the detection of PG-targeting antibiotics in Gram-negative bacteria have not been developed, mainly because of the lack of the regulatory systems that sense and respond to PG stress. Recently, we identified a novel two-component signal transduction system (PghKR) that is responsible for sensing and responding to PG damage in the Gram-negative bacterium Shewanella oneidensis. Based on this system, we developed biosensors for the detection of PG-targeting antibiotics. Using ampicillin as an inducer for PG stress and the bacterial luciferase LuxCDABE as the reporter, we found that the PghKR biosensors are specific to antibiotics targeting PG synthesis, including ß-lactams, vancomycin, and d-cycloserine. Deletion of genes encoding PG permease AmpG and ß-lactamase BlaA improves the sensitivity of the biosensors substantially. The PghKR biosensor in the background of ΔblaA is also functional on agar plates, providing a simple method for screening bacteria that produce PG-targeting antibiotics. IMPORTANCE The growing problem of antibiotic resistance in Gram-negative bacteria urgently needs new strategies so that researchers can develop novel antibiotics. Microbial whole-cell biosensors are capable of sensing various stimuli with a quantifiable output and show tremendous potential for the discovery of novel antibiotics. As the Achilles' heel of bacteria, the synthesis of the peptidoglycan (PG) is targeted by many antibiotics. However, the regulatory systems that sense and respond to PG-targeting stress in Gram-negative bacteria are reported rarely, restricting the development of biosensors for the detection of PG-targeting antibiotics. In this study, we developed a highly sensitive and specific biosensor based on a novel two-component system in the Gram-negative bacterium Shewanella oneidensis that is responsible for the sensing and responding to PG stress. Our biosensors have great potential for discovering novel antibiotics and determining the mode of action of antibiotics.

Innervated Reconstruction of Fingertip Degloving Injury Using a Dorsal Digital Perforator Flap Combined With a Cross-Finger Flap.

BACKGROUND: The reconstruction of a fingertip degloving injury presents a functional and aesthetic challenge. We used a dorsal digital perforator flap combined with a cross-finger flap to reconstruct this type of injury. The purposes of this retrospective study were to evaluate the efficacy of the combined flaps and to present our clinical experience. METHODS: From November 2016 to October 2019, 16 patients (13 men and 3 women) with fingertip degloving injuries were treated with a dorsal digital perforator flap combined with a cross-finger flap for innervated reconstruction. We used an innervated dorsal digital perforator flap for the reconstruction of the dorsal defect of the degloved fingertip and an innervated cross-finger flap for the volar defect. The average size of the defect was 4.2 × 1.9 cm. The average sizes of the flaps were 2.3 × 2.1 cm (the dorsal digital perforator flap) and 2.5 × 2.1 cm (the cross-finger flap). RESULTS: All flaps and skin grafts survived completely without ischemia or venous congestion. All wounds and their donor sites healed primarily without exudation and infection. Patients were followed up for a mean time of 11.3 ± 1.9 months (range, 9-15 months). At the final follow-up, no significant difference was seen in the averaged total active motion between the injured fingers and the contralateral fingers. No significant difference was found in the averaged total active motion between the donor fingers and the contralateral fingers. All flaps obtained excellent or good sensory performance. All flaps had mild cold intolerance. Thirteen patients had no pain, 2 reported mild pain, and 1 experienced moderate pain. Ten patients were very satisfied with the appearance of the reconstructed finger. CONCLUSIONS: The dorsal digital perforator flap combined with a cross-finger flap is an effective and reliable method for the reconstruction of fingertip degloving injuries.

Discovery of an Amino Acid-Modified Near-Infrared Aza-BODIPY Photosensitizer as an Immune Initiator for Potent Photodynamic Therapy in Melanoma.

The discovery of novel photosensitizers with potent phototoxicity and desirable water solubility is an urgent task for photodynamic therapy. Herein, a series of amino acid-modified aza-BODIPY photosensitizers were synthesized and evaluated. These new PSs exhibited enhanced aqueous solubility, increased 1O2 generation efficiency, and an improved photo-dark toxicity ratio. Aspartic acid-modified PS of 1a, which possessed intense NIR absorption and high 1O2 quantum yield, demonstrated the most potent efficacy toward the investigated tumor cell lines without using an emulsifier. Subcellular localization, cell-based ROS production, and cell death pathway of 1a were studied. In vivo fluorescence imaging and ex vivo organ distribution assays manifested that 1a possessed reasonable distribution and clearance. In vivo PDT studies indicated that 1a revealed advantages over Ce6 and our previously optimized PS of BDP-4. It not only afforded an excellent PDT effect with a low drug dose under only single-time photoirradiation but also induced an antitumor immunological response.

Development of a whole-cell biosensor for detection of antibiotics targeting bacterial cell envelope in Bacillus subtilis.

It is an urgent need to develop novel antibiotics to treat infections caused by multi-drug-resistant bacteria. One promising strategy could be the use of whole-cell biosensors, which have been extensively studied to monitor environmental pollutants and intracellular metabolites. Here, we used the σM-mediated regulatory system of Bacillus subtilis to construct a whole-cell biosensor for the detection of cell envelope-acting antibiotics. Using polymyxin B as the inducer for bacterial cell envelope stress and enhanced green fluorescent protein (EGFP) as the reporter, we found that the promoter of ypuA (PypuA) had the lowest background noise and the most significant changes in the fluorescence output. The whole-cell biosensor displayed dose-dependent and time-dependent responses in fluorescence signals. The detection range of this biosensor for polymyxin B was between 0.125 and 12 µg/mL. The response of the biosensor is specific to antibiotics that target the cell envelope. Besides determination in liquid cultures, the output signal of the biosensor can be easily determined on agar surfaces. Using this biosensor, we successfully detected polymyxins secreted by its producing strain and bacteria that produce cell envelope-acting antibiotics. KEY POINTS: • A whole-cell biosensor was constructed based on the σM-mediated regulatory system. • The response of the biosensor is specific to cell envelope-acting antibiotics. • The biosensor can be used to screen novel cell envelope-acting antibiotics.

Characterization of differentiated autoregulation of LuxI/LuxR-type quorum sensing system in Pseudoalteromonas.

Gram-negative bacteria usually use acyl-homoserine lactones (AHLs)-mediated LuxI/LuxR-type quorum sensing (QS) systems for cell-cell cooperation and/or bacteria-environment communication. LuxI and LuxR are AHLs synthase and receptor, respectively. These two parts could form a positive regulatory feedback loop, controlling various types of group behaviors. However, the autoregulation mechanisms between them are fragmented and could be highly differentiated in different bacteria. Here, we clarified the autoregulation mechanism between LuxI and LuxR in Pseudoalteromonas sp. R3. YasI (LuxI in strain R3) synthesizes two types of AHLs, C8-HSL and 3-OH-C8-HSL. It is worth noting that YasR (LuxR in strain R3) only responds to C8-HSL rather than 3-OH-C8-HSL. YasR-C8HSL can activate the yasI transcription by recognizing "lux box" at yasI upstream. Interestingly, YasR can directly promote the yasR expression with AHL-independent manner, but AHL absence caused by the yasI-deficiency led to the significant decrease in the yasR expression. Further study demonstrated that the yasI-deficiency can result in the decrease in the yasR mRNA stability. Notably, both yasI-deficiency and yasR-deficiency led to the significant decrease in the expression of hfq encoding RNA chaperone. Therefore, it was speculated that not only YasR itself can directly regulate the yasR transcription, but YasR-C8HSL complex indirectly affects the yasR mRNA stability by regulating Hfq.

A Lightweight Method for Defense Graph Neural Networks Adversarial Attacks.

Graph neural network has been widely used in various fields in recent years. However, the appearance of an adversarial attack makes the reliability of the existing neural networks challenging in application. Premeditated attackers, can make very small perturbations to the data to fool the neural network to produce wrong results. These incorrect results can lead to disastrous consequences. So, how to defend against adversarial attacks has become an urgent research topic. Many researchers have tried to improve the model robustness directly or by using adversarial training to reduce the negative impact of an adversarial attack. However, the majority of the defense strategies currently in use are inextricably linked to the model-training process, which incurs significant running and memory space costs. We offer a lightweight and easy-to-implement approach that is based on graph transformation. Extensive experiments demonstrate that our approach has a similar defense effect (with accuracy rate returns of nearly 80%) as existing methods and only uses 10% of their run time when defending against adversarial attacks on GCN (graph convolutional neural networks).

Hfq and sRNA00002 positively regulate the LuxI/LuxR-type quorum sensing system in Pseudoalteromonas.

Pseudoalteromonas spp. are Gram-negative bacteria which are ubiquitous in marine environments. Our previous work found that there is a classic LuxI/LuxR-type quorum sensing (QS) system which was named YasI/YasR in Pseudoalteromonas sp. R3, but the factors that control QS in strain R3 are unclear yet. Here, we found that the deficiency of hfq encoding RNA chaperon Hfq down-regulated the transcription levels of yasI encoding acyl-homoserine lactones (AHLs) synthase and yasR encoding AHLs receptor in strain R3. The assay based on fusion reporter of yasI-lacZ showed that Hfq regulates the expression of yasR at both transcriptional and translational levels. In addition, Hfq affects the expression of yasI via yasR. Further analysis indicated that the 5'UTR region of yasR is necessary for Hfq to control QS. In addition, the deletion of hfq increases the unstability of the target yasR mRNA. Based on transcriptome sequencing and bioinformatic analysis together with molecular experiments, Hfq-dependent sRNA00002 was identified to be involved in positively regulating QS in Pseudoalternas sp. R3. It was found that sRNA00002 deficiency causes the decrease in expression of yasI and yasR, and thus abolishes the production of AHLs in strain R3. It was concluded that Hfq-dependent sRNA00002 regulates yasR expression by base-pairing with target yasR mRNA at 5'UTR region and altering the stability of yasR mRNA. Our work paves the way for understanding the regulation mechanism of Hfq-dependent sRNAs on QS in Pseudoalteromonas.

Microwave pretreatment of camellia (Camellia oleifera Abel.) seeds: Effect on oil flavor.

Microwave is a new pretreatment technology, and microwave processing time of camellia seeds is a factor affecting the flavor of camellia seed oil (CSO). Therefore, this study on the characteristic volatile compounds of CSO from microwaved seeds with different processing time was carried out by electronic nose (E-nose), headspace-solid phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) and headspace-gas chromatography-ion mobility spectrometry (HS-GC-IMS). The results of E-nose show that W1W, W2W and W5S were the main sensors to distinguish the flavor profile of CSOs. Through HS-SPME-GC-MS and odor activity value analysis, 80 volatile compounds were detected and 22 key aroma compounds were screened in CSOs. Compared with HS-SPME-GC-MS, 44 volatile compounds were detected by HS-GC-IMS, including 9 identical compounds and 35 different compounds. In general, the volatile compounds of 0, 2 and 3 min CSOs were mainly alcohols and esters, while the 4, 5 and 6 min CSOs were mainly heterocycles and aldehydes.

Heterologous expression of bacterial cytochrome P450 from Microbacterium keratanolyticum ZY and its application in dichloromethane dechlorination.

Dichloromethane (DCM) is a volatile halogenated hydrocarbon with teratogenic, mutagenic and carcinogenic effects. Biodegradation is generally regarded as an effective and economical approach of pollutant disposal. In this study, a novel strain was isolated and its cytochrome P450 was heterologously expressed for DCM degradation. The isolate, Microbacterium keratanolyticum ZY, was characterized as a Gram-positive, rod-shaped and flagella-existed bacterium without spores (GenBank No. SUB8814364; CCTCC M 2019953). After successive whole-genome sequencing, assembly and annotation, eight identified functional genes (encoding cytochrome P450, monooxygenase, dehalogenase and hydrolase) were successfully cloned and expressed in Escherichia coli BL21 (DE3). The recombinant strain expressing cytochrome P450 presented the highest degradation efficiency (90.6%). Moreover, the specific activity of the recombinant cytochrome P450 was more than 1.2 times that of the recombinant dehalogenase (from Methylobacterium rhodesianum H13) under their optimum conditions. The kinetics of DCM degradation by recombinant cytochrome P450 was well fitted with the Haldane model and the value of maximum specific degradation rate was determined to be 0.7 s-1. The DCM degradation might occur through successive hydroxylation, dehydrohalogenation, dechlorination and oxidation to generate gem-halohydrin, formyl chloride, formaldehyde and formic acid. The study helps to comprehensively understand the DCM dechlorination process under the actions of bacterial functional enzymes (cytochrome P450 and dehalogenase).

Enhanced dechlorination of an enzyme-catalyzed electrolysis system by ionic liquids: Electron transfer, enzyme activity and dichloromethane diffusion.

Enzyme-catalyzed electrolysis system (EES) is a promising technique for the efficient dechlorination of pollutants. In this study, ionic liquids (ILs) was first introduced to enhance the dichloromethane dechlorination performance of an EES. An imidazole-based IL, 1-ethyl-3-methylimidazole tetrafluoroborate ([EMIM][BF4]), was chosen due to its excellent performance on dechlorination enhancement than other three ILs. The cyclic voltammograms with different scan rates shows that the presence of IL increased the apparent electron transfer rate constant (ks) from 0.008 to 0.013 s-1. The calculated surface electroactive species concentration (τc) also increased from 7.8 × 10-9 to 9.5 × 10-9 mol cm-2. Electrochemical impedance spectroscopy analysis illustrates that the IL mainly weakened the interfacial resistance between electrolyte and cathode to accelerate the electron communication in the EES. The introduction of IL facilitated the regeneration of reduced glutathione from oxidized glutathione, whereas inhibited the catalytic activity of dehalogenase via the disruption of secondary structure shown in circular dichroism spectra. The presence of IL was also facilitated the dichloromethane diffusion from electrolyte to cathode. The mass transfer rate constants of dichloromethane (km,d) increased by 6.9 times after the addition of IL. The optimum volume concentration, pH value, reaction temperature and applied voltage were 20%, 7, 35 °C and -0.8 V vs Ag/AgCl, respectively. The study is helpful to understand the promotion mechanism of IL on the dechlorination performance of EES when it is adopted as a treatment technique.