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We focused on the possibility that pathogenic microorganisms might produce immune suppressors to evade the action of immune cells. Based on this possibility, we have recently developed new co-culture method of pathogenic actinomyces and immune cells, however, the interaction mechanism between pathogens and cells was still unclear. In this report, co-culturing pathogenic fungi and immune cells were investigated. Pathogenic fungus Aspergillus terreus IFM 65899 and THP-1 cells were co-cultured and isolated a co-culture specific compound, butyrolactone Ia (1). 1 inhibits the production of nitric oxide by RAW264 cells and exhibits regulatory effects on autophagy, suggesting 1 plays a defensive role in the response of A. terreus IFM 65899 to immune cells. Furthermore, dialysis experiments and micrographs indicated that "physical interaction" between A. terreus IFM 65899 and THP-1 cells may be required for the production of 1. This is the first report of co-culture method of fungi with immune cells and its interaction mechanism., Competing Interests: Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

γ-Hydroxybutyrate (GHB) is an important C4 platform chemical, serving as a crucial precursor for the synthesis of various bulk chemicals, including γ-butyrolactone (GBL) and 1,4-butanediol (1,4-BDO). In this study, we report the systematic metabolic engineering of Corynebacterium glutamicum for the biological production of GHB from glucose via the introduction of a glutamate-derived pathway. We showed that C. glutamicum is a promising host for producing GHB due to its higher tolerance to GHB as compared to other chassis. By screening key enzymes capable of converting glutamate into GHB and blocking byproduct synthesis pathways, an engineered C. glutamicum strain was developed that achieved a GHB production titer of 30.6 g/L. Comparative transcriptome analysis was subsequently employed to identify previously uncharacterized aldehyde dehydrogenases responsible for succinate accumulation, and knockout of the corresponding genes led to an increased GHB titer of 33.7 g/L. Ultimately, the integration of a phosphoketolase-mediated nonoxidative glycolysis (NOG) pathway further enhanced GHB production, resulting in an accumulation of 38.3 g/L of GHB with a yield of 0.615 mol/mol glucose during batch fermentation. The GHB in the fermentation broth can be efficiently converted into GBL by acid treatment with a yield of 0.970 mol/mol.

The transition from planktonic to biofilm growth in bacteria is often accompanied by greater resistance to antibiotics and other stressors, as well as distinct alterations in physical traits, genetic activity, and metabolic restructuring. In many species, the heme nitric oxide/oxygen binding proteins (H-NOX) play an important role in this process, although the signaling mechanisms and pathways in which they participate are quite diverse and largely unknown. In Paracoccus denitrificans , deletion of the hnox gene results in a severe biofilm-deficient phenotype. Quantitative proteomics was used to assemble a comprehensive data set of P. denitrificans proteins showing altered abundance of those involved in several important metabolic pathways. Further, decreased levels of pyruvate and elevated levels of C 16 homoserine lactone were detected for the Δhnox strain, associating the biofilm deficiency with altered central carbon metabolism and quorum sensing, respectively. These results expand our knowledge of the important role of H-NOX signaling in biofilm formation.

Human intestinal microbiota plays a crucial role in converting secoisolariciresinol diglucoside, a lignan found in flaxseed, to enterodiol, which has a range of health benefits: antioxidative, antitumor, and estrogenic/anti-estrogenic effects. Given the high secoisolariciresinol diglucoside content in flaxseed cake, this study investigated the potential of co-fermenting flaxseed cake with fermented soybean product to isolate bacterial strains that effectively convert secoisolariciresinol diglucoside to enterodiol in a controlled environment (in vitro). The co-fermentation process with stinky tofu microbiota significantly altered the lignan, generating 12 intermediate lignan metabolites as identified by targeted metabolomics. One particular promising strain, ZB26, demonstrated an impressive ability to convert secoisolariciresinol diglucoside. It achieved a conversion rate of 87.42 ± 0.33%, with secoisolariciresinol and enterodiol generation rates of 94.22 ± 0.51% and 2.91 ± 0.03%, respectively. Further optimization revealed, under specific conditions (0.5 mM secoisolariciresinol diglucoside, pH 8, 30 °C for 3 days), ZB26 could convert an even higher percentage (97.75 ± 0.05%) of the secoisolariciresinol diglucoside to generate secoisolariciresinol (103.02 ± 0.16%) and enterodiol (3.18 ± 0.31%). These findings suggest that the identified strains ZB26 have promising potential for developing functional foods and ingredients enriched with lignans., Competing Interests: Declaration of competing interest We have no conflicts of interest concerning the work described in this manuscript., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Flax ( Linum usitatissimum L.) is the richest plant source of lignin secondary metabolites. Lignans from flax have been applied in the fields of food, medicine, and health due to their significant physiological activities. The most abundant lignan is secoisolariciresinol, which exists in a glycosylated form in plants., Results: After ingestion, it is converted by human intestinal flora into enterodiol and enterolactone, which both have physiological roles. Here, the basic structures, contents, synthesis, regulatory, and metabolic pathways, as well as extraction and isolation methods, of flax lignans were reviewed. Additionally, the physiological activity-related mechanisms and their impacts on human health, from the biosynthesis of lignans in plants to the physiological activity effects observed in animal metabolites, were examined., Conclusions: The review elucidates that lignans, as phenolic compounds, not only function as active substances in plants but also offer significant nutritional values and health benefits when flax is consumed.

Signaling systems of microorganisms are responsible for regulating the physiological and metabolic processes and also play vital roles in the communications of cells. Identifying signaling molecules mediating the cross-talks is challenging yet highly desirable for comprehending the microbial interactions. Here, we demonstrate that a pathogenic Gram-negative Chromobacterium violaceum exerts significant influence on the morphological differentiation and secondary metabolism of Gram-positive Streptomyces. The physiological metabolisms are directly modulated by three novel cinnamoyl lipids (CVCL1, 2, and 3) from C. violaceum CV12472, whose biosynthesis is under the control of N-acylhomoserine lactone signaling system. Furthermore, a receptor of CVCLs in Streptomyces ansochromogenes 7100 is determined to be SabR1, the cognate receptor of γ-butenolide signaling molecules. This study reveals an unprecedented mode of microbial interactions, and the quorum sensing signaling systems in these two groups of bacteria can be bridged via CVCLs, suggesting that CVCLs can modulate the physiological metabolism of cross-phylum microorganisms., (© 2024. The Author(s).)

Signaling molecules, particularly acyl-homoserine lactones (AHLs), can enhance microbial activity under low-temperature stress. However, the specific mechanisms underlying this effect remain unclear. This study identified a typical activated sludge functional bacterium that is sensitive to low temperatures and regulated by hexanoyl-L-homoserine lactone (C6-HSL), a representative of AHLs. It elucidates how C6-HSL modulates the bacterium's resistance to low-temperature stress. Experimental results indicated that C6-HSL significantly increased the levels of adenosine triphosphate (ATP), superoxide dismutase (SOD), peroxidase (POD) and glutathione peroxidase (GSH-Px) in strain LB-001 under low-temperature stress, while also decreasing the levels of reactive oxygen species (ROS). Additionally, C6-HSL markedly repaired the damage to cell membrane structure caused by low-temperature stress. At the genetic level, C6-HSL upregulated the expression of 20 key genes related to energy metabolism, antioxidation, and fatty acid synthesis. At the metabolic level, C6-HSL increased the levels of metabolites related to energy metabolism and antioxidation, boosted the content of unsaturated fatty acids, and reduced the content of saturated fatty acids. This study utilized C6-HSL and low-temperature induction in conjunction with 16S microbial diversity sequencing, genomics, transcriptomics, and metabolomics. These methods were employed to elucidate the molecular mechanisms by which exogenous C6-HSL regulates the resistance of activated sludge microbial communities to low-temperature stress. This research lays the foundation for the application of AHLs and cell communication in wastewater biological treatment, fostering deeper exploration and further innovation in related academic research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium that remains a prevalent clinical and environmental challenge. Quorum-sensing (QS) molecules are effective biomarkers in pinpointing the presence of P. aeruginosa . This study aimed to develop a convenient-to-use, whole-cell biosensor using P. aeruginosa reporters individually encapsulated within alginate-poly-L-lysine (alginate-PLL) microbeads to specifically detect the presence of bacterial autoinducers. The PLL-reinforced microbeads were prepared using a two-step method involving ionic cross-linking and subsequent coating with thin layers of PLL. The alginate-PLL beads showed good stability in the presence of a known cation scavenger (sodium citrate), which typically limits the widespread applications of calcium alginate. In media containing synthetic autoinducers-such as N-(3-oxo dodecanoyl) homoserine lactone (3-oxo-C 12 -HSL) and N -butanoyl-L-homoserine lactone (C 4 -HSL), or the cell-free supernatants of planktonic or the flow-cell biofilm effluent of wild P. aeruginosa (PAO1)-the encapsulated bacteria enabled a dose-dependent detection of the presence of these QS molecules. The prepared bioreporter beads remained stable during prolonged storage at 4 and -80 °C and were ready for on-the-spot sensing without the need for recovery. The proof-of-concept, optical fiber-based, and whole-cell biosensor developed here demonstrates the practicality of the encapsulated bioreporter for bacterial detection based on specific QS molecules.

Overexpression of carboxyl/cholinesterase (CCE) genes has been reported to be associated with many cases of pesticide resistance in arthropods. However, it has been rarely documented that CCE genes participate in spirodiclofen resistance in Panonychus citri . In previous research, we found that spirodiclofen resistance is related to increased P450 and CCE enzyme activities in P. citri . In this study, we identified two CCE genes, PcCCE3 and PcCCE5 , which were significantly upregulated in spirodiclofen-resistant strain and after exposure to spirodiclofen. RNA interference of PcCCE3 and PcCCE5 increased the spirodiclofen susceptibility in P. citri . In vitro metabolism indicated that Pc CCE3 and Pc CCE5 could interact with spirodiclofen, but metabolites were detected only in the Pc CCE3 treatment. Our results indicated that Pc CCE3 participates in spirodiclofen resistance through direct metabolism, and Pc CCE5 may be involved in the spirodiclofen resistance by passive binding and sequestration, which provides new insights into spirodiclofen resistance in P. citri .

Ralstonia solanacearum species complex (RSSC) includes soilborne bacterial plant pathogens with worldwide distribution and wide host ranges. Virulence factors are regulated via four hierarchically organized cell-cell contact independent quorum-sensing (QS) signalling systems: the Phc, which uses as signals (R)-methyl 3-hydroxypalmitate [(R)-3-OH PAME] or (R)-methyl 3-hydroxymyristate [(R)-3-OH MAME], the N-acyl homoserine lactone (AHL)-dependent RasI/R and SolI/R systems, and the recently identified anthranilic acid-dependent system. The unique Phc QS system has been extensively studied; however, the role of the two AHL QS systems has only recently been addressed. In this microreview, we present and discuss current data of the SolI/R and RasI/R QS systems in the RSSC. We also present the distribution and frequency of these AHL QS systems in the RSSC, discuss possible ecological roles and evolutive implications. The complex QS hierarchical networks emphasizes the crucial role of cell-cell signalling in the virulence of the RSSC., (© 2024 The Author(s). Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Pseudomonas aeruginosa (P. aeruginosa) biofilm formation is a crucial cause of enhanced antibiotic resistance. Quorum sensing (QS) is involved in regulating biofilm formation; QS inhibitors block the QS signaling pathway as a new strategy to address bacterial resistance. This study investigated the potential and mechanism of L-HSL (N-(3-cyclic butyrolactone)-4-trifluorophenylacetamide) as a QS inhibitor for P. aeruginosa. The results showed that L-HSL effectively inhibited the biofilm formation and dispersed the pre-formed biofilm of P. aeruginosa. The production of extracellular polysaccharides and the motility ability of P. aeruginosa were suppressed by L-HSL. C. elegans infection experiment showed that L-HSL was non-toxic and provided protection to C. elegans against P. aeruginosa infection. Transcriptomic analysis revealed that L-HSL downregulated genes related to QS pathways and biofilm formation. L-HSL exhibits a promising potential as a therapeutic drug for P. aeruginosa infection. KEY POINTS: • Chemical synthesis of N-(3-cyclic butyrolactone)-4-trifluorophenylacetamide, named L-HSL. • L-HSL does not generate survival pressure on the growth of P. aeruginosa and can inhibit the QS system. • KEGG enrichment analysis found that after L-HSL treatment, QS-related genes were downregulated., (© 2024. The Author(s).)

Aims : Plant-derived lignans may protect against obesity, while their bioactivity needs gut microbial conversion to enterolignans. We used repeated measures to identify enterolignan-predicting microbial species and investigate whether enterolignans and enterolignan-predicting microbial species are associated with obesity. Methods : Urinary enterolignans, fecal microbiota, body weight, height, and circumferences of the waist (WC) and hips (HC) were repeatedly measured at the baseline and after 1 year in 305 community-dwelling adults in Huoshan, China. Body composition and liver fat [indicated by the controlled attenuation parameter (CAP)] were measured after 1 year. Multivariate-adjusted linear models and linear mixed-effects models were used to analyze single and repeated measurements, respectively. Results : Enterolactone and enterodiol levels were both inversely associated with the waist-to-hip ratio, body fat mass (BFM), visceral fat level (VFL), and liver fat accumulation (all P < 0.05). Enterolactone levels were also associated with lower WC ( β = -0.0035 and P = 0.013) and HC ( β = -0.0028 and P = 0.044). We identified multiple bacterial genera whose relative abundance was positively associated with the levels of enterolactone (26 genera) and enterodiol (22 genera, all P false discovery rate < 0.05), and constructed the enterolactone-predicting microbial score and enterodiol-predicting microbial score to reflect the overall enterolignan-producing potential of the host gut microbiota. Both these scores were associated with lower body weight and CAP (all P < 0.05). The enterolactone-predicting microbial score was also inversely associated with the BFM ( β = -0.1128 and P = 0.027) and VFL ( β = -0.1265 and P = 0.044). Conclusion : Our findings support that modulating the host gut microbiome could be a potential strategy to prevent obesity by enhancing the production of enterolignans.

In recent years, as the paradigm of communication between cells has been clarified, the ability of bacteria to change their gene expression patterns in response to various extracellular signals has attracted great interest. In particular, intracellular and intercellular communication between bacterial populations, called quorum sensing (QS), is essential for coordinating physiological and genetic activities. QS studies are critical, particularly in elucidating the regulatory mechanisms of infectious processes in food-borne pathogens. Elucidating the QS mechanisms in Salmonella is effective in silencing the virulence factors in the fight against this bacterium. The aims of this study were; to create luxS gene mutants that play a vital role in the QS activity of Salmonella and to determine the effect of this mutation on the expression of virulence genes in the bacteria and to determine the impact of synthetic N-hexanoyl-homoserine lactone (C6HSL) on biofilm formation and AI-2 signaling pathway of Salmonella wild strain and luxS gene mutants. luxS gene mutants were constructed by recombining the gene region with the chloramphenicol gene cassette based on homologous region recombination. In the luxS mutants obtained in this way, the expression of eight different virulence genes (hilA, invA, inv, glgC, fimF, fliF, lpfA, gyrA), which have essential roles in Salmonella pathogenicity, was determined by quantitative real-time reverse transcriptase polymerase chain reaction (rRT-qPCR) method and compared with natural strains. As a result of these studies, it was determined that the expression of each gene examined was significantly reduced in luxS mutant strains. The relative AI-2 activities of Salmonella strains were analyzed depending on time. It was determined that the highest activity occurred at the fourth hour and the AI-2 activities of luxS mutants were reduced compared to the wild strain. Finally, it was determined that C6HSL increased the biofilm activity of Salmonella Typhimurium DMC4, SL1344 wild strains, and mutants, mainly at the 72nd hour. In conclusion, our results proved that C6HSL stimulated QS communication in all strains and increased biofilm of Salmonella formation and autoinducer activity. This situation determines that Salmonella responds to external signals by using QS systems. In addition, this research contributed to provide additional information on interspecies communication mechanisms to develop strategies to prevent biofilm formation of this pathogen.

Programmable intercellular signaling using components of naturally occurring quorum sensing can allow for coordinated functions to be engineered in microbial consortia. LuxR-type transcriptional regulators are widely used for this purpose and are activated by homoserine lactone (HSL) signals. However, they often suffer from imperfect molecular discrimination of structurally similar HSLs, causing misregulation within engineered consortia containing multiple HSL signals. Here, we studied one such example, the regulator LasR from Pseudomonas aeruginosa . We elucidated its sequence-function relationship for ligand specificity using targeted protein engineering and multiplexed high-throughput biosensor screening. A pooled combinatorial saturation mutagenesis library (9,486 LasR DNA sequences) was created by mutating six residues in LasR's β5 sheet with single, double, or triple amino acid substitutions. Sort-seq assays were performed in parallel using cognate and noncognate HSLs to quantify each corresponding sensor's response to each HSL signal, which identified hundreds of highly specific variants. Sensor variants identified were individually assayed and exhibited up to 60.6-fold ( p = 0.0013) improved relative activation by the cognate signal compared to the wildtype. Interestingly, we uncovered prevalent mutational epistasis and previously unidentified residues contributing to signal specificity. The resulting sensors with negligible signal crosstalk could be broadly applied to engineer bacteria consortia.

A universal biochemical signal for bacterial cell-cell communication could facilitate programming dynamic responses in diverse bacterial consortia. However, the classical quorum sensing paradigm is that Gram-negative and Gram-positive bacteria generally communicate via homoserine lactones (HSLs) or oligopeptide molecular signals, respectively, to elicit population responses. Here, we create synthetic HSL sensors for Gram-positive Bacillus subtilis 168 using allosteric LuxR-type regulators (RpaR, LuxR, RhlR, and CinR) and synthetic promoters. Promoters were combinatorially designed from different sequence elements (-35, -16, -10, and transcriptional start regions). We quantified the effects of these combinatorial promoters on sensor activity and determined how regulator expression affects its activation, achieving up to 293-fold activation. Using the statistical design of experiments, we identified significant effects of promoter regions and pairwise interactions on sensor activity, which helped to understand the sequence-function relationships for synthetic promoter design. We present the first known set of functional HSL sensors (≥20-fold dynamic range) in B. subtilis for four different HSL chemical signals: p -coumaroyl-HSL, 3-oxohexanoyl-HSL, n -butyryl-HSL, and n -(3-hydroxytetradecanoyl)-HSL. This set of synthetic HSL sensors for a Gram-positive bacterium can pave the way for designable interspecies communication within microbial consortia.

Acyl-homoserine lactones (AHLs) are quorum-sensing signaling molecules in Gram-negative bacteria and positively regulate biofilm formation in Salmonella under specific conditions. In this study, biofilm formation in Salmonella enterica was evaluated at 28 and 37 °C, under aerobic and anaerobic conditions. Additionally, the influence of the N -dodecanoyl-DL-homoserine lactone (C12-HSL) on biofilm formation and the expression of genes related to the synthesis of structural components, regulation, and quorum sensing was assessed under anaerobiosis at 28 and 37 °C. Biofilm formation was found not to be influenced by the atmospheric conditions at 28 °C. However, it was reduced at 37 °C under anaerobiosis. C12-HSL enhanced biofilm formation at 37 °C under anaerobiosis and increased the expression of the adrA and luxS genes, suggesting an increase in c-di-GMP, a second messenger that controls essential physiological functions in bacteria. These results provide new insights into the regulation of biofilm formation in Salmonella under anaerobic conditions.

Importance: The bacterium Pseudomonas aeruginosa colonizes and thrives in many environments, in which it is typically found in surface-associated polymicrobial communities known as biofilms. Adaptation to this social behavior is aided by quorum sensing (QS), an intercellular communication system pivotal in the expression of social traits. Regardless of its importance in QS regulation, the loss of function of the master regulator LasR is now considered a conserved adaptation of P. aeruginosa , irrespective of the origin of the strains. By investigating the QS circuitry in surface-grown cells, we found an accumulation of QS signal 3-oxo-C 12 -HSL in the absence of its cognate receptor and activator, LasR. The current understanding of the QS circuit, mostly based on planktonic growing cells, is challenged by investigating the QS circuitry of surface-grown cells. This provides a new perspective on the beneficial aspects that underline the frequency of LasR-deficient isolates., Competing Interests: The authors declare no conflict of interest.

The symbioses that animals form with bacteria play important roles in health and disease, but the molecular details underlying how bacterial symbionts initially assemble within a host remain unclear. 1 , 2 , 3 The bioluminescent bacterium Vibrio fischeri establishes a light-emitting symbiosis with the Hawaiian bobtail squid Euprymna scolopes by colonizing specific epithelium-lined crypt spaces within a symbiotic organ called the light organ. 4 Competition for these colonization sites occurs between different strains of V. fischeri, with the lancet-like type VI secretion system (T6SS) facilitating strong competitive interference that results in strain incompatibility within a crypt space. 5 , 6 Although recent studies have identified regulators of this T6SS, how the T6SS is controlled as symbionts assemble in vivo remains unknown. 7 , 8 Here, we show that T6SS activity is suppressed by N-octanoyl-L-homoserine lactone (C8 HSL), which is a signaling molecule that facilitates quorum sensing in V. fischeri and is important for efficient symbiont assembly. 9 , 10 We find that this signaling depends on the quorum-sensing regulator LitR, which lowers expression of the needle subunit Hcp, a key component of the T6SS, by repressing transcription of the T6SS regulator VasH. We show that LitR-dependent quorum sensing inhibits strain incompatibility within the squid light organ. Collectively, these results provide new insights into the mechanisms by which regulatory networks that promote symbiosis also control competition among symbionts, which in turn may affect the overall symbiont diversity that assembles within a host., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

While pioneering methods have demonstrated that bacterial N-acyl homoserine lactone (AHL) signaling molecules can influence the growth and self-aggregation of suspended microalgae, whether AHLs can affect the initial adhesion to a carrier has remained an open question. Here we revealed that the microalgae exhibited different adhesion potential under AHL mediation, where the performance was affiliated to both AHL types and concentrations. The result can be well explained by the interaction energy theory, where the energy barrier between the carriers and the cells varied due to AHL mediation. Depth analyses revealed that AHL acted through modifying the properties of the surface electron donor of the cells, which were dependent upon three major components, i.e., extracellular protein (PN) secretion, the PN secondary structure, and the PN amino acid composition. These findings expand the known diversity of AHLs mediation on microalgal initial adhesion and metabolisms, which may interface with other major cycles and become helpful to theoretically guide the application of AHLs in microalgal culture and harvesting., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The host transcriptional activator Early growth response 1 (EGR1) plays a vital role in cell cycle and differentiation, cell proliferation, and regulation of cytokines and several growth factors. It is an immediate-early gene that is expressed as an initial response to various environmental stimuli. Bacterial infection is one such factor that can trigger the expression of EGR1 in host. Therefore, it is imperative to understand expression of EGR1 during early stages of host-pathogen interaction. Streptococcus pyogenes is an opportunistic bacteria causing skin and respiratory tract infections in humans. The quorum-sensing molecule, N-(3-oxododecanoyl)-l-homoserine lactone (Oxo-C12), not synthesised by S. pyogenes, can be sensed by S. pyogenes leading to molecular changes in the pathogen. In this study, we investigated the role of Oxo-C12 on EGR1 regulation in lung epithelial and murine macrophage cell line upon S. pyogenes infection. We report that Oxo-C12 sensitised S. pyogenes upregulates the transcriptional expression of EGR1 through ERK1/2 pathway. It was observed that EGR1 was not involved in the intial attachment of S. pyogenes to A549 cells. However, inhibition of EGR1 in macrophage cell line, J774A.1, through the ERK1/2 pathway resulted in decreased adhesion of S. pyogenes. The EGR1 upregulation by Oxo-C12 sensitised S. pyogenes plays a vital role in enhancing the survival of S. pyogenes in murine macrophages, leading to persistent infection. Thus, understanding the molecular modulation in the host during bacterial infection will further help develop therapeutics to target specific sites., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

d-(-)-Pantolactone (DPL) is a key intermediate for the production of d-(+)-pantothenate (vitamin B5). Deracemization of d,l-pantolactone (D,L-PL) through oxidizing l-(+)-pantolactone (LPL) to ketopantoyl lactone (KPL) and subsequently reducing KPL to DPL is a promising route for synthesizing DPL. Herein, a newly mined l-pantolactone dehydrogenase from Rhodococcus hoagie (RhoLPLDH) was used for the oxidative dehydrogenation of LPL. To alleviate inclusion bodies formed by membrane-bound RhoLPLDH intracellular expression in E. coli, strategies involving chaperone assistance and decreasing induction temperature were used to achieve RhoLPLDH soluble expression. To enhance its activity, directed evolution and hydrophilicity-based engineering yielded increased catalytic activity and thermostability. 1 M LPL was efficiently converted to KPL by engineering strain CM5 co-expressing RhoLPLDH L254I/V241I/I156L/F224Q/N164K and chaperone. A "two stages in one-pot" method was employed in deracemization of 1 M D,L-PL with 91.2% yield. These results demonstrated that CM5 catalyst exhibits great potential in enzyme cascade deracemization for the production of DPL., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The parasitic weed Striga has led to billions of dollars' worth of agricultural productivity loss worldwide. Striga detects host plants using compounds of the strigolactone class of phytohormones. Early steps in the strigolactone signaling pathway involve substrate binding and hydrolysis followed by a conformational change to an "active" or "closed" state, after which it associates with a MAX2-family downstream signaling partner. The structures of the inactive and active states of strigolactone receptors are known through X-ray crystallography, and the transition pathway from the inactive to active state in apo receptors has previously been characterized using molecular dynamics simulations. However, it also has been suggested that a covalent butenolide modification of the receptor on the catalytic histidine through substrate hydrolysis promotes formation of the active state. Using molecular dynamics simulations, we show that the presence of the covalent butenolide enhances activation in both AtD14, a receptor found in Arabidopsis, and ShHTL7, a receptor found in Striga, but the enhancement is ∼50 times greater in ShHTL7. We also show that several conserved interactions with the covalent butenolide modification promote transition to the active state in both AtD14 (non-parasite) and ShHTL7 (parasite). Finally, we demonstrate that the enhanced activation of ShHTL7 likely results from disruption of ShHTL7-specific histidine interactions that inhibited activation in the apo case. These results provide a possible explanation for difference in strigolactone sensitivity seen between different strigolactone-sensitive proteins and can be used to aid the design of selective modulators to control Striga parasites., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

In Pseudomonas aeruginosa , quorum sensing (QS) depends on an interconnected regulatory hierarchy involving the Las, Rhl and Pq s systems, which are collectively responsible for the co-ordinated synthesis of a diverse repertoire of N -acylhomoserine lactones (AHLs) and 2-alkyl-4-quinolones (AQs). Apparent population density-dependent phenomena such as QS may, however, be due to growth rate and/or nutrient exhaustion in batch culture. Using continuous culture, we show that growth rate and population density independently modulate the accumulation of AHLs and AQs such that the highest concentrations are observed at a slow growth rate and high population density. Carbon source (notably succinate), nutrient limitation (C, N, Fe, Mg) or growth at 25 °C generally reduces AHL and AQ levels, except for P and S limitation, which result in substantially higher concentrations of AQs, particularly AQ N -oxides, despite the lower population densities achieved. Principal component analysis indicates that ~26 % variation is due to nutrient limitation and a further 30 % is due to growth rate. The formation of N -(3-oxododecanoyl)-l-homoserine lactone (3OC12-HSL) turnover products such as the ring opened form and tetramic acid varies with the limiting nutrient limitation and anaerobiosis. Differential ratios of N -butanoyl-homoserine lactone (C4-HSL), 3OC12-HSL and the AQs as a function of growth environment are clearly apparent. Inactivation of QS by mutation of three key genes required for QS signal synthesis ( lasI , rhlI and pqsA ) substantially increases the concentrations of key substrates from the activated methyl cycle and aromatic amino acid biosynthesis, as well as ATP levels, highlighting the energetic drain that AHL and AQ synthesis and hence QS impose on P. aeruginosa .

The lichen Cetraria islandica is traditionally used as a demulcent for the symptomatic treatment of irritations of the mouth and throat and associated dry cough, as well as for the treatment of temporary loss of appetite. In addition to depsides and depsidones, thalli contain paraconic acids, a group of secondary metabolites commonly found in lichens and fungi. Among those, protolichesterinic acid has shown promising pharmacological activities. However, the efficient isolation of paraconic acids is quite complex due to their very similar chemical structures and their weak ultraviolet absorption. In the present work, a two-step isolation protocol of protolichesterinic acid and lichesterinic acid from a complex paraconic acid mixture is described using Sephadex LH20 column chromatography and fast centrifugal partition chromatography. Final purities higher than 95% and recoveries above 50% are achieved. Additionally, reliable qualitative techniques for detecting and differentiating paraconic acids are described. Finally, some data on compound stability and enantiomeric purity are shown., (© 2023 The Authors. Journal of Separation Science published by Wiley-VCH GmbH.)

Bacterial expression systems play an indispensable role in the biosynthesis of recombinant proteins. Different proteins and the tasks associated with them may require different systems. The purpose of this work is to make an expression vector that allows switching on and off the expression of the target gene during cell incubation. Several expression vectors for use in Escherichia coli cells were developed using elements of the luxR/luxI type quorum sensing system of psychrophilic bacterium Aliivibrio logei. These vectors contain A. logei luxR2 and (optionally) luxI genes and LuxR2-regulated promoter, under the control of which a target gene is intended to be inserted. The synthesis of the target protein depends directly on the temperature: gene expression starts when the temperature drops to 22 °C and stops when it rises to 37 °C, which makes it possible to fix the desired amount of the target protein in the cell. At the same time, the expression of the target gene at a low temperature depends on the concentration of the autoinducer (L-homoserine N-(3-oxohexanoyl)-lactone, AI) in the culture medium in a wide range from 1 nM to 10 μM, which makes it possible to smoothly regulate the rate of target protein synthesis. Presence of luxI in the vector provides the possibility of autoinduction. Constructed expression vectors were tested with gfp, ardA, and ardB genes. At maximum, we obtained the target protein in an amount of up to 33% of the total cellular protein. KEY POINTS: • A. logei quorum sensing system elements were applied in new expression vectors • Expression of target gene is inducible at 22 °C and it is switched off at 37 °C • Target gene expression at 22 °C is tunable by use different AI concentrations., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

N-acyl homoserine lactones (N-HLs) are signaling molecules used by Gram-negative bacteria in a phenomenon called quorum sensing. Bacteria will detect N-HLs as a way of monitoring their population which, upon reaching a critical level, will express a specific phenotype. An example is the expression of bioluminescence by Vibrio fischeri. Most studies have not considered the chirality of these molecules nor have they used highly sensitive detection methods. Here, the production of d,l-N-HLs are monitored for V. fischeri, Burkholderia cepacia, Pseudomonas fluorescens, and P. putida, using highly sensitive tandem mass spectrometry analysis. Novel N-HLs are reported for both V. fischeri and B. cepacia, including a plethora of previously unknown d-N-HLs, including the first d-N-HLs containing oxo and hydroxy functionalities. Anomalously, N-HLs were not detected in any cultures of P. fluorescens and P. putida, which are species that previously were reported to produce N-HLs. However, it is apparent that differences in the reported occurrence and levels of N-HLs can result from (a) different strains of bacteria, (b) different growth media and environmental conditions, and (c) sometimes false-positive results from detection methodologies. Time studies of V. fischeri suggest the possibility that separate synthetic and elimination pathways exist between d- and l-N-HLs. Possible biological processes that could be the source of d-N-HL production are considered., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

Functional metagenomics is an essential and effective approach to recover new enzymes from the environment. In this chapter, we describe a procedure to construct metagenomic library to discover new N-acyl homoserine lactone (AHL) degrading enzymes based on a direct method or an indirect enrichment procedure. Applicable to any bacterial ecosystem, it enables rapid identification of functional enzymes effective to degrade AHLs., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

The N -acyl homoserine lactone (AHL) acylases are widely used as quorum sensing (QS) blockers to inhibit bacterial food spoilage. However, their substrate specificity for long-chain substrates weakens their efficiency. In this study, a computer-assisted design of AHL acylase PF2571 was performed to modify its substrate scope. The results showed that the variant PF2571 H194Y, L221R could effectively quench N -hexanoyl-l-homoserine lactone and N -octanoyl-l-homoserine lactone without impairing its activity against long-chain AHLs. Kinetic analysis of the enzymatic activities further corroborated the observed substrate expansion. The inhibitory activities of this variant were significantly enhanced against the QS phenotype of Aeromonas veronii BY-8, with inhibition rates of 45.67, 78.25, 54.21, and 54.65% against proteases, motility, biofilms, and extracellular polysaccharides, respectively. Results for molecular dynamics simulation showed that the steric hindrance, induced by residue substitution, could have been responsible for the change in substrate scope. This study dramatically improves the practicability of AHL acylase in controlling food spoilage.

N -acyl-homoserine lactones (AHLs) as autoinducers of Gram-negative bacteria for quorum sensing regulation have shown positive effects on the production of aromatic proteins in extracellular polymeric substances (EPSs) during bioflocculation. To investigate the role of AHLs in aromatic protein production, a Chlorella -bacteria system with great bioflocculation was established via fed-batch cultivation. Tryptophan and aromatic proteins as the main compounds in the EPS of bioflocs showed an increasing trend during fed-batch cultivation. The Chlorella cells only secreted tryptophan rather than aromatic proteins during axenic cultivation. N-dodecanoyl-l-homoserine lactone (C12-HSL) was correlated with the flocculation activity and extracellular protein content of bioflocs during fed-batch cultivation. The addition of exogenous C12-HSL enhanced the flocculation activity of the Chlorella -bacteria system and aromatic protein production in the EPS. Chlorella cells sensed exogenous C12-HSL and significantly upregulated the aromatic protein synthesis pathway during axenic cultivation. In addition, vanillin as a quorum-sensing inhibitor suppressed the positive effect of C12-HSL on flocculation activity and aromatic protein production and synthesis. This result indicated that vanillin intercepts the response of Chlorella cells to C12-HSL. Overall, C12-HSL is supposed to be an important signal molecule to achieve communication between Chlorella and Gram-negative bacteria and subsequently induce Chlorella cells to produce aromatic proteins for biofloc formation.

In the gut ecosystem, microorganisms regulate group behaviour and interplay with the host via a molecular system called quorum sensing (QS). The QS molecule 3-oxo-C12:2-HSL, first identified in human gut microbiota, exerts anti-inflammatory effects and could play a role in inflammatory bowel diseases where dysbiosis has been described. Our aim was to identify which signalling pathways are involved in this effect. We observed that 3-oxo-C12:2-HSL decreases expression of pro-inflammatory cytokines such as Interleukine-1β (- 35%) and Tumor Necrosis Factor-α (TNFα) (- 40%) by stimulated immune RAW264.7 cells and decreased TNF secretion by stimulated PBMC in a dose-dependent manner, between 25 to 100 µM. Transcriptomic analysis of RAW264.7 cells exposed to 3-oxo-C12:2-HSL, in a pro-inflammatory context, highlighted JAK-STAT, NF-κB and TFN signalling pathways and we confirmed that 3-oxo-C12:2-HSL inhibited JAK1 and STAT1 phosphorylation. We also showed through a screening assay that 3-oxo-C12:2-HSL interacted with several human bitter taste receptors. Its anti-inflammatory effect involved TAS2R38 as shown by pharmacologic inhibition and led to an increase in intracellular calcium levels. We thus unravelled the involvement of several cellular pathways in the anti-inflammatory effects exerted by the QS molecule 3-oxo-C12:2-HSL., (© 2022. The Author(s).)

The γ-butyrolactone motif is found in many natural signaling molecules and other specialized metabolites. A prominent example is the potent aquatic phytotoxin cyanobacterin, which has a highly functionalized γ-butyrolactone core structure. The enzymatic machinery that assembles cyanobacterin and structurally related natural products (herein termed furanolides) has remained elusive for decades. Here, we elucidate the biosynthetic process of furanolide assembly. The cyanobacterin biosynthetic gene cluster was identified by targeted bioinformatic screening and validated by heterologous expression in Escherichia coli. Full functional evaluation of the recombinant key enzymes in vivo and in vitro, individually and in concert, provided in-depth mechanistic insights into a streamlined C-C bond-forming cascade that involves installation of compatible reactivity at seemingly unreactive Cα positions of amino acid precursors. Our work extends the biosynthetic and biocatalytic toolbox for γ-butyrolactone formation, provides a general paradigm for furanolide biosynthesis and sets the stage for their targeted discovery, biosynthetic engineering and enzymatic synthesis., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Vibrio fluvialis is a marine opportunistic pathogen that frequently causes diseases in aquatic animals and humans. V. fluvialis can produce quorum sensing signaling molecules to coordinate cell density-dependent behavioral changes, including N-acyl homoserine lactone (AHL), which acts as a vital mediator of virulence-associated gene expression. Currently, several AHL molecules in V. fluvialis have been detected via biological and physicochemical methods, although different detection approaches have generated diverse AHL profiles. Here, we describe the AHL-producing bacterium, V. fluvialis BJ-1, which was isolated from marine sediments from the East China Sea. V. fluvialis BJ-1 could stimulate AHL-mediated β-galactosidase synthesis of the biosensor Agrobacterium tumefaciens NTL4 (pZLR4) but could not induce violacein production in the AHL reporter strain, Chromobacterium violaceum CV026. This bacterial isolate exhibited strong AHL-producing activity at low cell density; however, the AHL activity declined when population density remained at high levels. Analysis of the AHLs by Ultra-High-Performance Liquid Chromatography tandem Mass Spectrometry demonstrated that V. fluvialis BJ-1 produced five different AHL signaling molecules, including two linear chain AHL products (C 8 - and C 10 -HSL), and three β-carbon-oxidative AHL products (3-O-C 8 -, 3-O-C 10 - and 3-O-C 12 -HSL). Significantly, the present study is the first to accurately define the AHL profile of marine V. fluvialis. In future, the coupling of UHPLC to ESI-MS/MS is expected to be utilized for the accurate determination of AHL profiles in marine Vibrio., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: The conversion of plant lignans to bioactive enterolignans in the gastrointestinal tract is mediated through microbial processing. The goal of this study was to examine the relationships between lignan intake, plasma enterolactone concentrations, gut microbiome composition, and metabolic risk in free-living male adults., Results: In 303 men participating in the Men's Lifestyle Validation Study (MLVS), lignan intake was assessed using two sets of 7-day diet records, and gut microbiome was profiled through shotgun sequencing of up to 2 pairs of fecal samples (n = 911). A score was calculated to summarize the abundance of bacteria species that were significantly associated with plasma enterolactone levels. Of the 138 filtered species, plasma enterolactone levels were significantly associated with the relative abundances of 18 species at FDR < 0.05 level. Per SD increment of lignan intake was associated with 20.7 nM (SEM: 2.3 nM) higher enterolactone concentrations among participants with a higher species score, whereas the corresponding estimate was 4.0 nM (SEM: 1.7 nM) among participants with a lower species score (P for interaction < 0.001). A total of 12 plasma metabolites were also significantly associated with these enterolactone-predicting species. Of the association between lignan intake and metabolic risk, 19.8% (95%CI: 7.3%-43.6%) was explained by the species score alone, 54.5% (95%CI: 21.8%-83.7%) by both species score and enterolactone levels, and 79.8% (95%CI: 17.7%-98.6%) by further considering the 12 plasma metabolites., Conclusion: We identified multiple gut bacteria species that were enriched or depleted at higher plasma levels of enterolactone in men. These species jointly modified the associations of lignan intake with plasma enterolactone levels and explained the majority of association between lignan intake and metabolic risk along with enterolactone levels and certain plasma metabolites., (© 2022. The Author(s).)

The development of novel and safe insecticides remains an important need for a growing world population to protect crops and animal and human health. New chemotypes modulating the insect nicotinic acetylcholine receptors have been recently brought to the agricultural market, yet with limited understanding of their molecular interactions at their target receptor. Herein, we disclose the first crystal structures of these insecticides, namely, sulfoxaflor, flupyradifurone, triflumezopyrim, flupyrimin, and the experimental compound, dicloromezotiaz, in a double-mutated acetylcholine-binding protein which mimics the insect-ion-channel orthosteric site. Enabled by these findings, we discovered novel pharmacophores with a related mode of action, and we describe herein their design, synthesis, and biological evaluation.

Cell-cell communication is critical for bacterial survival in natural habitats, in which miscellaneous regulatory networks are encompassed. However, elucidating the interaction networks of a microbial community has been hindered by the population complexity. This study reveals that γ-butyrolactone (GBL) molecules from Streptomyces species, the major antibiotic producers, can directly bind to the acyl-homoserine lactone (AHL) receptor of Chromobacterium violaceum and influence violacein production controlled by the quorum sensing (QS) system. Subsequently, the widespread responses of more Gram-negative bacterial AHL receptors to Gram-positive Streptomyces signaling molecules are unveiled. Based on the cross-talk between GBL and AHL signaling systems, combinatorial regulatory circuits (CRC) are designed and proved to be workable in Escherichia coli (E. coli). It is significant that the QS systems of Gram-positive and Gram-negative bacteria can be bridged via native Streptomyces signaling molecules. These findings pave a new path for unlocking the comprehensive cell-cell communications in microbial communities and facilitate the exploitation of innovative regulatory elements for synthetic biology., (© 2021. Science China Press and Springer-Verlag GmbH Germany, part of Springer Nature.)

The highly conserved ATP-dependent Lon protease plays important roles in diverse biological processes. The lon gene is usually nonessential for viability; however, lon mutants of several bacterial species, although viable, exhibit cellular defects. Here, we show that a lack of Lon protease causes pleiotropic effects in the rice pathogen Burkholderia glumae. The null mutation of lon produced three colony types, big (BLONB), normal (BLONN), and small (BLONS), in Luria-Bertani (LB) medium. Colonies of the BLONB and BLONN types were re-segregated upon subculture, while those of the BLONS type were too small to manipulate. The BLONN type was chosen for further studies, as only this type was fully genetically complemented. BLONN-type cells did not reach the maximum growth capacity, and their population decreased drastically after the stationary phase in LB medium. BLONN-type cells were defective in the biosynthesis of quorum sensing (QS) signals and exhibited reduced oxalate biosynthetic activity, causing environmental alkaline toxicity and population collapse. Addition of excessive N-octanoyl-homoserine lactone (C8-HSL) to BLONN-type cell cultures did not fully restore oxalate biosynthesis, suggesting that the decrease in oxalate biosynthesis in BLONN-type cells was not due to insufficient C8-HSL. Co-expression of lon and tofR in Escherichia coli suggested that Lon negatively affects the TofR level in a C8-HSL-dependent manner. Lon protease interacted with the oxalate biosynthetic enzymes, ObcA and ObcB, indicating potential roles for the oxalate biosynthetic activity. These results suggest that Lon protease influences colony morphology, growth, QS system, and oxalate biosynthesis in B. glumae., Competing Interests: The authors have declared that no competing interests exist.

N -acyl homoserine lactones (AHLs) are small signaling molecules used by many Gram-negative bacteria for coordinating their behavior as a function of their population density. This process, based on the biosynthesis and the sensing of such molecular signals, and referred to as Quorum Sensing (QS), regulates various gene expressions, including growth, virulence, biofilms formation, and toxin production. Considering the role of QS in bacterial pathogenicity, its modulation appears as a possible complementary approach in antibacterial strategies. Analogues and mimics of AHLs are therefore biologically relevant targets, including several families in which heterocyclic chemistry provides a strategic contribution in the molecular design and the synthetic approach. AHLs consist of three main sections, the homoserine lactone ring, the central amide group, and the side chain, which can vary in length and level of oxygenation. The purpose of this review is to summarize the contribution of heterocyclic chemistry in the design of AHLs analogues, insisting on the way heterocyclic building blocks can serve as replacements of the lactone moiety, as a bioisostere for the amide group, or as an additional pattern appended to the side chain. A few non-AHL-related heterocyclic compounds with AHL-like QS activity are also mentioned.

Microbes employ autoinducers of quorum sensing (QS) for population communication. Although the autoinducer of Pseudomonas aeruginosa LasI-LasR system, N-(3-oxododecanoyl)- l-homoserine lactone (3OC12), has been reported with deleterious effects on host cells, its biological effects on integrity of the intestinal epithelium and epithelial barrier are still unclear and need further investigation. In the present study, flow cytometry, transcriptome analysis and western blot technology have been adopted to investigate the potential molecular mechanisms of 3OC12 and its structurally similar analogs damage to intestinal epithelial cells. Our results indicated that 3OC12 and 3OC14 trigger apoptosis rather than necrosis and ferroptosis in intestinal epithelial cells. RNA-sequencing combined with bioinformatics analysis showed that 3OC12 and 3OC14 reduced the expression of genes from extracellular matrix (ECM)-receptor interaction pathway. Consistently, protein expressions from ECM and tight junction-associated pathway were significantly reduced after 3OC12 and 3OC14 challenge. In addition, 3OC12 and 3OC14 led to blocked cell cycle, decreased mitochondrial membrane potential, increased reactive oxygen species level and elevated Ca 2+ concentration. Reversely, the antioxidant NAC could effectively mitigate the reduced expression of ECM and tight junction proteins caused by 3OC12 and 3OC14 challenge. Collectively, this study demonstrated that QS autoinducer exposure to intestinal epithelial cells ablates the ECM and tight junctions by triggering oxidative stress and apoptosis, and finally disrupts the intestinal epithelial barrier. These findings provide a rationale for defensing QS-dependent bacterial infections and potential role of NAC for alleviating the syndrome., (© 2020 Wiley Periodicals LLC.)

Quorum sensing interference (QSI), the disruption and manipulation of quorum sensing (QS) in the dynamic control of bacteria populations could be widely applied in synthetic biology to realize dynamic metabolic control and develop potential clinical therapies. Conventionally, limited QSI molecules (QSIMs) were developed based on molecular structures or for specific QS receptors, which are in short supply for various interferences and manipulations of QS systems. In this study, we developed QSIdb (http://qsidb.lbci.net/), a specialized repository of 633 reported QSIMs and 73 073 expanded QSIMs including both QS agonists and antagonists. We have collected all reported QSIMs in literatures focused on the modifications of N-acyl homoserine lactones, natural QSIMs and synthetic QS analogues. Moreover, we developed a pipeline with SMILES-based similarity assessment algorithms and docking-based validations to mine potential QSIMs from existing 138 805 608 compounds in the PubChem database. In addition, we proposed a new measure, pocketedit, for assessing the similarities of active protein pockets or QSIMs crosstalk, and obtained 273 possible potential broad-spectrum QSIMs. We provided user-friendly browsing and searching facilities for easy data retrieval and comparison. QSIdb could assist the scientific community in understanding QS-related therapeutics, manipulating QS-based genetic circuits in metabolic engineering, developing potential broad-spectrum QSIMs and expanding new ligands for other receptors., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: Accumulating evidence has suggested that human gut microbiota metabolize certain dietary compounds and subsequently produce bioactive metabolites that may exert beneficial or harmful effects on coronary artery disease (CAD) risk., Objectives: This study examined the joint association of 2 gut microbiota metabolites, enterolactone and trimethylamine N-oxide (TMAO), that originate from intake of plant-based foods and animal products, respectively, in relation to CAD risk., Methods: A prospective nested case-control study of CAD was conducted among participants who were free of diabetes, cardiovascular disease, and cancer in the Nurses' Health Study II and the Health Professionals Follow-up Study. Plasma concentrations of enterolactone and TMAO, as well as choline and L-carnitine, were assayed among 608 CAD case-control pairs., Results: A high enterolactone and low TMAO profile was associated with better diet quality, especially higher intake of whole grains and fiber and lower intake of red meats, as well as lower concentrations of plasma triglycerides and C-reactive protein. Participants with a high enterolactone/low TMAO profile had a significantly lower risk of CAD: the multivariate-adjusted OR was 0.58 (95% CI: 0.38, 0.90), compared with participants with a low enterolactone/high TMAO profile. No significant interaction between enterolactone and TMAO on CAD risk was observed. Neither TMAO nor enterolactone alone were associated with CAD risk in pooled analyses. In women, a higher enterolactone concentration was significantly associated with a 54% lower CAD risk (P trend = 0.03), although the interaction by sex was not significant., Conclusions: Our results show that a profile characterized by high enterolactone and low TMAO concentrations in plasma is linked to a healthful dietary pattern and significantly associated with a lower risk of CAD. Overall, these data suggest that, compared with individual markers, multiple microbiota-derived metabolites may facilitate better differentiation of CAD risk and characterization of the relations between diet, microbiota, and CAD risk., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Recent studies suggest that quorum-sensing molecules may play a role in gut microbiota-host crosstalk. However, whether microbiota produces quorum-sensing molecules and whether those molecules can trans-kingdom transport to the host are still unknown. Here, we develop a UPLC-MS/MS-based assay to screen the 27 N-acyl homoserine lactones (AHLs) in the gut microbiota and host. Various AHL molecules are exclusively detected in the cecal contents, sera and livers from conventionally-raised mice but cannot be detected in germ-free mice. Pathogen-produced C4-HSL is detected in the cecal contents and sera of Citrobacter rodentium (C. rodentium)-infected mice, but not found in uninfected controls. Moreover, C. rodentium infection significantly increases the level of multiple AHL molecules in sera. Our findings demonstrate that both commensal and pathogenic bacteria, can produce AHLs that can be detected in host bodies, suggesting that quorum-sensing molecules could be a group of signaling molecules in trans-kingdom microbiota-host crosstalk.

In December 2020, the U.K. authorities reported to the World Health Organization (WHO) that a new COVID-19 variant, considered to be a variant under investigation from December 2020 (VUI-202012/01), was identified through viral genomic sequencing. Although several other mutants were previously reported, VUI-202012/01 proved to be about 70% more transmissible. Hence, the usefulness and effectiveness of the newly U.S. Food and Drug Administration (FDA)-approved COVID-19 vaccines against these new variants are doubtfully questioned. As a result of these unexpected mutants from COVID-19 and due to lack of time, much research interest is directed toward assessing secondary metabolites as potential candidates for developing lead pharmaceuticals. In this study, a marine-derived fungus Aspergillus terreus was investigated, affording two butenolide derivatives, butyrolactones I ( 1 ) and III ( 2 ), a meroterpenoid, terretonin ( 3 ), and 4-hydroxy-3-(3-methylbut-2-enyl)benzaldehyde ( 4 ). Chemical structures were unambiguously determined based on mass spectrometry and extensive 1D/2D NMR analyses experiments. Compounds ( 1 - 4 ) were assessed for their in vitro anti-inflammatory, antiallergic, and in silico COVID-19 main protease (M pro ) and elastase inhibitory activities. Among the tested compounds, only 1 revealed significant activities comparable to or even more potent than respective standard drugs, which makes butyrolactone I ( 1 ) a potential lead entity for developing a new remedy to treat and/or control the currently devastating and deadly effects of COVID-19 pandemic and elastase-related inflammatory complications.

We developed a bioelectronic communication system that is enabled by a redox signal transduction modality to exchange information between a living cell-embedded bioelectronics interface and an engineered microbial network. A naturally communicating three-member microbial network is 'plugged into' an external electronic system that interrogates and controls biological function in real time. First, electrode-generated redox molecules are programmed to activate gene expression in an engineered population of electrode-attached bacterial cells, effectively creating a living transducer electrode. These cells interpret and translate electronic signals and then transmit this information biologically by producing quorum sensing molecules that are, in turn, interpreted by a planktonic coculture. The propagated molecular communication drives expression and secretion of a therapeutic peptide from one strain and simultaneously enables direct electronic feedback from the second strain, thus enabling real-time electronic verification of biological signal propagation. Overall, we show how this multifunctional bioelectronic platform, termed a BioLAN, reliably facilitates on-demand bioelectronic communication and concurrently performs programmed tasks.

Despite extensive studies on the actin regulators that direct microfilament dynamics, how these regulators are combinatorially utilized in organismal tissues to generate 3D structures is an unresolved question. Here, we present an in-depth characterization of cortical actin cap dynamics and their regulation in vivo. We identify rapid phases of initiation, expansion, duplication, and disassembly and examine the functions of seven different actin and/or nucleator regulators (ANRPs) in guiding these behaviors. We find ANRPs provide distinct activities in building actin cap morphologies - specifically, while DPod1 is a major regulator of actin intensities, Cortactin is required for continued cortical growth, while Coronin functions in both growth and intensity and is required for Cortactin localization to the cap periphery. Unexpectedly, cortical actin populations recover more rapidly after regulator disruption, suggestive of a deep competition for limited G-actin pools, and we measure in vivo Arp2/3 recruitment efficiencies through an ectopic relocalization strategy. Our results illustrate how the coordination of multiple actin regulators can orchestrate organized and dynamic actin structures in a developmental system., Competing Interests: YX, RB, JB No competing interests declared, (© 2021, Xie et al.)

Acinetobacter baumannii possesses a single divergent luxR / luxRI -type quorum-sensing (QS) locus named abaR / abaI This locus also contains a third gene located between abaR and abaI , which we term abaM , that codes for an uncharacterized member of the RsaM protein family known to regulate N -acylhomoserine lactone (AHL)-dependent QS in other beta- and gammaproteobacteria. Here, we show that disruption of abaM via a T26 insertion in A. baumannii strain AB5075 resulted in increased production of N -(3-hydroxydodecanoyl)-l-homoserine lactone and enhanced surface motility and biofilm formation. In contrast to the wild type and the abaI ::T26 mutant, the virulence of the abaM ::T26 mutant was completely attenuated in a Galleria mellonella infection model. Transcriptomic analysis of the abaM ::T26 mutant revealed that AbaM differentially regulates at least 76 genes, including the csu pilus operon and the acinetin 505 lipopeptide biosynthetic operon, that are involved in surface adherence, biofilm formation and virulence. A comparison of the wild type, abaM ::T26 and abaI ::T26 transcriptomes, indicates that AbaM regulates ∼21% of the QS regulon including the csu operon. Moreover, the QS genes ( abaI and abaR ) were among the most upregulated in the abaM ::T26 mutant. A. baumannii lux -based abaM reporter gene fusions revealed that abaM expression is positively regulated by QS but negatively autoregulated. Overall, the data presented in this work demonstrates that AbaM plays a central role in regulating A. baumannii QS, virulence, surface motility, and biofilm formation. IMPORTANCE Acinetobacter baumannii is a multiantibiotic-resistant pathogen of global health care importance. Understanding Acinetobacter virulence gene regulation could aid the development of novel anti-infective strategies. In A. baumannii , the abaR and abaI genes that code for the receptor and synthase components of an N -acylhomoserine (AHL) lactone-dependent quorum sensing system (QS) are separated by abaM Here, we show that although mutation of abaM increased AHL production, surface motility, and biofilm development, it resulted in the attenuation of virulence. AbaM was found to control both QS-dependent and QS-independent genes. The significance of this work lies in the identification of AbaM, an RsaM ortholog known to control virulence in plant pathogens, as a modulator of virulence in a human pathogen., (Copyright © 2021 López-Martín et al.)

A fast, sensitive, and reliable analytical method was developed and validated for simultaneous identification and quantification of spirodiclofen, spiromesifen, and spirotetramat and their relevant metabolites in edible fungi by ultra-performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS). First, sample extraction was done with acetonitrile containing 1% formic acid followed by phase separation with the addition of MgSO 4 :NaOAc. Then, the supernatant was purified by primary secondary amine (PSA), octadecylsilane (C18), and graphitized carbon black (GCB). The linearities of the calibrations for all analytes were excellent (R 2  ≥ 0.9953). Acceptable recoveries (74.5-106.4%) for all analytes were obtained with good intra- and inter- relative standard deviations of less than 14.5%. The limit of quantification (LOQs) for all analytes was 10 μg kg -1 . For accurate quantification, matrix-matched calibration curve was applied to normalize the matrix effect. The results indicated that the method was suitable for detecting the three acaricides and their relevant metabolites in edible fungi.

Mammalian lignans are phytoestrogens with important bioactivities, and their concentrations in livestock milk may influence the health of consumers. This research aimed to establish a method to quantify multiple mammalian lignans in the biofluids of dairy sheep using ultra-HPLC-triple quadropole mass spectrometry with multiple-reaction monitoring. Secoisolariciresinol, 2-[(4-hydroxy-3-methoxyphenyl)methyl]-3-[(3-hydroxyphenyl)methyl]-1,4-butanediol, enterodiol (ED), enterolactone (EL), ED-sulfate (ED-S), and EL-sulfate (EL-S) were purified from the urine of flaxseed cake-fed dairy sheep. The structures of these lignans were identified by a combination of mass and nuclear magnetic resonance spectra. These purified lignans were used as standards to optimize their quantification conditions in urine, milk, and plasma of dairy sheep. On this basis, the lignan metabolites in biofluids were quantified. To improve analysis sensitivity, plasma and milk were pretreated with acetonitrile containing 1% formic acid and passed through a HybridSPE-PL 55261-U column (Supelco, Bellefonte, PA). The limit of quantification of the lignans ranged from 1.43 to 18.3 ng/mL in plasma, and from 1.01 to 18.7 ng/mL in milk. The linearity of the calibration curves ranged from their limit of quantification to at least 217 ng/mL in plasma, and 217 ng/mL in milk. Regression coefficient of the calibration curves were above 0.99 for secoisolariciresinol, 2-[(4-hydroxy-3-methoxyphenyl)methyl]-3-[(3-hydroxyphenyl)methyl]-1,4-butanediol, ED, EL, ED-S, and EL-S, indicating satisfactory relationships between the peak areas and concentrations in the quantification range. The relative concentrations of ED-glucuronide and EL-glucuronide (EL-G) in different biofluids were compared based on their chromatogram peak areas. The sheep plasma contained all forms of mammalian lignans (i.e., ED, EL, ED-S, EL-S, ED-glucuronide, and EL-G.); the urine contained ED, EL, ED-S, and EL-S; and the milk contained ED, EL, ED-S, EL-S, and EL-G. Milk-to-plasma concentration ratios of the mammalian lignans indicated that the free forms were more permeable than the sulfated conjugates. Mammalian lignans found in sheep plasma and milk may provide health benefits to the sheep and sheep-product consumers. The analytical method established in this work could be used to quantify mammalian lignans in livestock products., (Copyright © 2021 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Context: Recent studies have outlined the potential role of dietary factors in patients who have survived cancer., Objective: The aim of this study was to summarize the evidence of the relation between dietary intake of phytoestrogens and their blood biomarkers and, overall, cancer-specific mortality and recurrence in patients with cancer., Data Sources: A systematic search of PubMed, EMBASE, and Web of Science databases of studies published up to September 2019 was performed. Databases were searched for prospective and retrospective cohort studies reporting on dietary phytoestrogen intake and/or blood biomarkers and the outcomes investigated., Data Extraction: Data were extracted from each identified study using a standardized form., Data Analysis: Twenty-eight articles on breast, lung, prostate, and colorectal cancer, and glioma were included for systematic review. Given the availability of studies, a quantitative meta-analysis was performed solely for breast cancer outcomes. A significant inverse association among higher dietary isoflavone intake, higher serum/plasma enterolactone concentrations, and overall mortality and cancer recurrence was found. Among other cancer types, 2 studies reported that higher serum enterolactone and higher intake of lignans were associated with cancer-specific survival for colorectal cancer and glioma, respectively., Conclusions: Dietary phytoestrogens may play a role in survival from breast cancer ; evidence regarding other cancers is too limited to draw any conclusions., (© The Author(s) 2020. Published by Oxford University Press on behalf of the International Life Sciences Institute. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Although Psychrobacter strain M9-54-1 had been previously isolated from the microbiota of holothurians and shown to degrade quorum sensing (QS) signal molecules C6 and C10-homoserine lactone (HSL), little was known about the gene responsible for this activity. In this study, we determined the whole genome sequence of this strain and found that the full 16S rRNA sequence shares 99.78-99.66% identity with Psychrobacter pulmonis CECT 5989 T and P. faecalis ISO-46 T . M9-54-1, evaluated using the agar well diffusion assay method, showed high quorum quenching (QQ) activity against a wide range of synthetic N -acylhomoserine lactone (AHLs) at 4, 15, and 28 °C. High-performance liquid chromatography-mass-spectrometry (HPLC-MS) confirmed that QQ activity was due to an AHL-acylase. The gene encoding for QQ activity in strain M9-54-1 was identified from its genome sequence whose gene product was named AhaP. Purified AhaP degraded substituted and unsubstituted AHLs from C4- to C14-HSL. Furthermore, heterologous expression of ahaP in the opportunistic pathogen Pseudomonas aeruginosa PAO1 reduced the expression of the QS-controlled gene lecA , encoding for a cytotoxic galactophilic lectin and swarming motility protein. Strain M9-54-1 also reduced brine shrimp mortality caused by Vibrio coralliilyticus VibC-Oc-193, showing potential as a biocontrol agent in aquaculture.

Bacterial hormones, such as the iconic gamma-butyrolactone A-factor, are essential signaling molecules that regulate diverse physiological processes, including specialized metabolism. These low molecular weight compounds are common in Streptomyces species and display species-specific structural differences. Recently, unusual gamma-butyrolactone natural products called salinipostins were isolated from the marine actinomycete genus Salinispora based on their antimalarial properties. As the salinipostins possess a rare phosphotriester motif of unknown biosynthetic origin, we set out to explore its construction by the widely conserved 9-gene spt operon in Salinispora species. We show through a series of in vivo and in vitro studies that the spt gene cluster dually encodes the salinipostins and newly identified natural A-factor-like gamma-butyrolactones (Sal-GBLs). Remarkably, homologous biosynthetic gene clusters are widely distributed among many actinomycete genera, including Streptomyces , suggesting the significance of this operon in bacteria.