Quorum-sensing System Research Articles

Bacillus quorum quenching shapes the citrus mycobiome through interkingdom signaling

Microbiomes are sustained through infinite yet mutually interacting microbial communities, with bacteria and fungi serving as the major constituents. In recent times, microbial interventions have become popular for microbiome manipulation to achieve sustainable goals. Whether and how the introduced biocontrol agent drives fungal microbial assemblages (mycobiome) and the role of interkingdom signaling in shaping the microbiome structure and function remain poorly understood. Here, we implemented wild-type (WT) Bacillus subtilis L1–21 and its quorum quenching (QQ) mutants (L1–21Δytnp, L1–21Δyxel) individually and as consortia to explore the enrichment patterns of key mycobiome members in Huanglongbing (HLB) infected citrus compartments including leaf endosphere, root endosphere, and rhizosphere soil. The application of WT and its QQ mutants produced differential mycobiome enrichment across citrus compartments. Our findings reveal that application of WT B. subtilis enriched beneficial fungi such as Trichoderma (15.82 %) in leaf endosphere. In contrast, pathogenic fungi Fusarium (47.5 %) and Gibberella (0.47 %) involved in citrus root decline were adundant in the L1–21Δytnp treated root endosphere while Nigrospora (11 %) was predominant in L1–21Δyxel treated leaf endosphere, affirming the role of bacterial quorum sensing (QS) molecules in shaping the fungal community composition. In general, based on the fungal functional prediction, fungal pathogens were highly abundant in mutant-treated plants, particularly in leaf endosphere (L1–21Δytnp: 25 %; L1–21Δyxel: 36.35 %) compared to WT (20.93 %). Additionally, some fungal members exhibited strong compartment specificity and both mutants induced distinct mycobiome shifts in rhizosphere soil, leaf, and root endopshere. In conclusion, B. subtilis QQ modifies bacterial QS networks facilitating beneficial fungi to establish, while loss of QQ leads to enrichment of pathogenic fungal groups. Our study provides a direct link of perception and regulation of mycobiome through bacterial-based QS and QQ system, and its association with disease outcomes.

Polidocanol inhibits Enterococcus faecalis virulence factors by targeting fsr quorum sensing system

BackgroundThe wide spread of antimicrobial resistance in Enterococcus faecalis is a critical global concern, leading to increasingly limited treatment options. The fsr quorum sensing (QS) plays a critical role in the pathogenicity of E. faecalis, allowing bacteria to coordinate gene expression and regulate many virulence factors. Therefore, fsr QS of E. faecalis represents a potential therapeutic target that provides an effective strategy to treat antibiotic-resistant infections induced by E. faecalis.MethodsIn this study, distribution of different virulence factors including, gelatinase, protease, cell surface hydrophobicity and biofilm formation in sixty clinical isolates of Enterococcus faecalis was investigated. Sixty-six compounds were tested for their activity against fsr QS. The minimal inhibitory concentration of the tested compounds was evaluated using the microbroth dilution method. The effect of sub-inhibitory concentrations of the tested compounds on fsr QS was investigated using the gelatinase assay method. Additionally, the effect of potential QS inhibitor on the virulence factors was estimated. Quantitative real-time PCR was used to investigate the effect of the potential inhibitor on fsr QS related genes (fsrB-fsrC) and (gelE-sprE) and virulence associated genes including, asa1 and epbA.ResultsThe assessment of polidocanol activity against the fsr QS system was demonstrated by studying its effect on gelatinase production in E. faecalis clinical isolates. Sub-lethal concentrations of polidocanol showed a significant reduction in gelatinase and protease production by 54% to 70% and 64% to 85%, respectively. Additionally, it significantly reduced biofilm formation (P < 0.01) and interrupted mature biofilm at concentrations of ½, 1 × and 2 × MIC. Furthermore, polidocanol significantly decreased cell surface hydrophobicity (P < 0.01). Polidocanol at ½ MIC showed a significant reduction in the expression of QS genes including fsrB, fsrC, gelE and sprE by 57% to 97% without affecting bacterial viability. Moreover, it reduced the expression of virulence associated genes (asa1 and epbA) (P < 0.01).ConclusionPolidocanol appears to be a promising option for treating of E. faecalis infections by targeting the fsr QS system and exhibiting anti-biofilm activity.

Novel motif associated with carbon catabolite repression in two major Gram-positive pathogen virulence regulatory proteins.

Carbon catabolite repression (CCR) is a widely conserved regulatory process that ensures enzymes and transporters of less-preferred carbohydrates are transcriptionally repressed in the presence of a preferred carbohydrate. This phenomenon can be regulated via a CcpA-dependent or CcpA-independent mechanism. The CcpA-independent mechanism typically requires a transcriptional regulator harboring a phosphotransferase regulatory domain (PRD) that interacts with phosphotransferase system (PTS) components. PRDs contain a conserved histidine residue that is phosphorylated by the PTS-associated HPr-His15~P protein. PRD-containing regulators often harbor additional domains that resemble PTS-associated EIIB protein domains with a conserved cysteine residue that can be phosphorylated by cognate PTS components. We noted that Mga, the PRD-containing central virulence regulator of Streptococcus pyogenes, has an EIIBGat domain containing a cysteine that, based on the presence of a similar motif in glycerol kinase, could be a target for phosphorylation. Using site-directed mutagenesis, we constructed phospho-ablative and phospho-mimetic substitutions of this cysteine and found that these substitutions modify the CCR of the Rgg2/3 quorum-sensing system. Moreover, we provide genetic evidence that the phospho-donor of this cysteine residue is likely to be ManL, the EIIA/B subunit of the mannose PTS system. Interestingly, a structurally distinct virulence gene regulator, PrfA of Listeria monocytogenes, harbors a similar cysteine-containing motif, and phospho-ablative and phospho-mimetic substitutions of the cysteine-altered CCR of PrfA-dependent virulence gene expression. Collectively, our data suggest that phosphorylation of a cysteine within the shared novel motif in Mga and PrfA may be a heretofore missing link between cellular metabolism and virulence.IMPORTANCEIn this study, we identified a novel cysteine-containing motif within the amino acid sequence of two structurally distinct transcriptional regulators of virulence in two Gram-positive pathogens that appears to link carbon metabolism with virulence gene expression. The results also highlight the potential post-translational modification of cysteine in bacterial species, a rare and understudied modification.

Transcriptome analysis expands underlying mechanisms of quorum sensing mediating heterotrophic nitrification-aerobic denitrification process at low temperature

Quorum sensing (QS) could regulate the behavior of microbial communities and help them resist adverse low-temperature environments. A newly isolated heterotrophic nitrification-aerobic denitrification (HN-AD) bacterium, strain YB1107, exhibited strong tolerance to harsh cold environments, removing 93.5 % of ammonia within 36 h and achieving a maximum specific growth rate of 0.28 h−1 at 10 °C. Strain YB1107 secreted large amounts of N-butanoyl-L-homoserine or N-octanoyl-L-homoserine lactones in response to cold stimuli. The add-back experiments indicated that these two signaling molecules jointly manipulated microbial physiological behavior by improving ammonia oxidation and biofilm formation, while inhibiting aerobic denitrification. The transcriptome analysis revealed that QS systems enhanced the cold resistance of HN-AD bacteria by promoting nitrogen assimilation and reducing dissimilation through regulating related genes. This study provided new molecular insights into how QS mediated HN-AD at low temperatures and laid the foundation for the potential applications of psychrophilic HN-AD bacteria in wastewater treatment.

Mechanisms of the Quorum Sensing Systems of Pseudomonas aeruginosa: Host and Bacteria.

Quorum-sensing is a communication mechanism between bacteria with the ability to activate signaling pathways in the bacterium and in the host cells. Pseudomonas aeruginosa is a pathogen with high clinical relevance due to its vast virulence factors repertory and wide antibiotic resistance mechanisms. Due to this, it has become a pathogen of interest for developing new antimicrobial agents in recent years. P. aeruginosa has three major QS systems that regulate a wide gene range linked with virulence factors, metabolic regulation, and environment adaption. Consequently, inhibiting this communication mechanism would be a strategy to prevent the pathologic progression of the infections caused by this bacterium. In this review, we aim to overview the current studies about the signaling mechanisms of the QS system of P. aeruginosa and its effects on this bacterium and the host.

Furazolidone reduces the pathogenesis of Trueperella pyogenes and Pseudomonas aeruginosa co-infection in a mouse model

The prevalence of abscess disease significantly limits the population expansion of captive forest musk deer, which is an endangered species protected by the legislation of China. Our prior work had demonstrated that Trueperella pyogenes and Pseudomonas aeruginosa are two important microorganisms in causing the abscess disease of forest musk deer, and furazolidone could inhibit the growth and virulence of the pathogens in vitro. In this study, the in vivo protection activity of furazolidone was evaluated by using mouse models chronically infected with T. pyogenes and P. aeruginosa. The results showed that furazolidone treatment significantly increased the survival rates of mice in the co-infection group, all the mice survived at 14 days post-infection. The damage degree of the lung tissues caused by bacterial infection was ameliorated by the treatment of furazolidone from 7 to 14 days post-infection, which also reduced the residual bacterial burden in the lungs. Compared to the untreated control group, the expression levels of genes activated by the quorum-sensing system of P. aeruginosa and the core virulence regulatory genes of T. pyogenes were significantly suppressed by furazolidone. In addition, the results of transcriptomic analyses showed that 270 DEGs were identified in the co-infection group. This finding further revealed that the immune responses of mice could be enhanced by the treatment of furazolidone, and this might also contribute to the clearance of bacteria from the lungs. Therefore, this study clearly reveals the protection activity of furazolidone against P. aeruginosa and T. pyogenes infection, and thus provides a promising candidate in the treatment of abscess disease.

Exploring Phytochemical Compounds Against Pseudomonas Aeruginosa Using QSAR, Molecular Dynamics, and Free Energy Landscape

AbstractPseudomonas aeruginosa is a versatile opportunistic bacterium that presents a considerable risk in medical environments because of its strong adaptability and resistance to multiple medications. Targeting the LasR quorum sensing system, which plays a crucial role in controlling virulence factors and biofilm formation, is a key intervention point. In this study, in silico molecular docking, machine learning‐based Quantitative Structure‐Activity Relationship (QSAR) techniques along with molecular dynamics simulation were employed to screen phytochemical compounds for their ability to inhibit the LasR QS system, a key regulator of virulence in Pseudomonas aeruginosa. This study screened 1652 phytochemicals using the ML‐based QSAR model to identify 52 phytochemicals that had better activity than the control (N‐{[3,5‐dibromo‐2‐(methoxymethoxy)phenyl]methyl}‐2‐nitrobenzamide). The in silico molecular docking approach that targeted LasR identified compounds 5281647, 57331045, and 5281672 with high binding affinity and hydrogen bonds that were comparable to the control (docking score=−10.3 kcal/mol and hydrogen bonds=4). In the 200 ns post‐molecular dynamics simulation, 5281647 exhibited a stable RMSD of 0.25 nm, which was comparable to the control. The maximum number of hydrogen bonds was exhibited by 57331045, while 5281647 and 5281672 consistently exhibited four hydrogen bonds. Overall, the Principal component analysis (PCA) and Free Energy Landscape (FEL) analyses of the complexes demonstrated that these three compounds were in stable states. In comparison to the control (ΔGTOTAL=−39.58 kcal/mol), the cumulative binding free energy (ΔGTOTAL) for 5281647 and 57331045 was −39.95 kcal/mol and −39.25 kcal/mol, respectively. This further confirms the superior binding affinity of the two compounds. Both 5281647 and 57331045 were identified as potent inhibitors of the LasR transcription factor, which is essential for the quorum sensing of Pseudomonas aeruginosa, in the present investigation. These findings underscore the importance of further exploration and optimization of phytochemicals for combating bacterial infections, offering promising avenues for future drug discovery efforts targeting this resilient pathogen.

The Oxylipin Dependent Quorum Sensing System enhances Pseudomonas aeruginosa dissemination during burn-associated infection.

Following severe burn injury, Pseudomonas aeruginosa is the leading cause of life-threatening infection. Herein, we unveil how P. aeruginosa strategically employs host-derived oleic acid, released as consequence of burn-injury, to induce a hypervirulent phenotype via its Oxylipin Dependent Quorum Sensing system (ODS). ODS activation enhanced P. aeruginosa invasion of burned skin and promoted its dissemination to distant organs in vivo . ODS regulation of P. aeruginosa virulence involved the control of nitic oxide levels, a key signaling molecule in bacteria, through upregulation of the nitric oxide reductases NorCB. Immunization with OdsA, one of the enzymes involved in oxylipin generation, or treatment with a pharmacological inhibitor of OdsA, protected mice against lethal P. aeruginosa infection following burn-injury. Our findings reveal a new mechanism underlying P. aeruginosa hypervirulence in burn wounds and identifies OdsA as a promising target for preventing disseminated infections following burns.

Impact of Quorum Sensing on the Virulence and Survival Traits of Burkholderia plantarii.

Quorum sensing (QS) is a mechanism by which bacteria detect and respond to cell density, regulating collective behaviors. Burkholderia plantarii, the causal agent of rice seedling blight, employs the LuxIR-type QS system, common among Gram-negative bacteria, where LuxI-type synthase produces QS signals recognized by LuxR-type regulators to control gene expression. This study aimed to elucidate the QS mechanism in B. plantarii KACC18965. Through whole-genome analysis and autoinducer assays, the plaI gene, responsible for QS signal production, was identified. Motility assays confirmed that C8-homoserine lactone (C8-HSL) serves as the QS signal. Physiological experiments revealed that the QS-defective mutant exhibited reduced virulence, impaired swarming motility, and delayed biofilm formation compared to the wild type. Additionally, the QS mutant demonstrated weakened antibacterial activity against Escherichia coli and decreased phosphate solubilization. These findings indicate that QS in B. plantarii significantly influences various pathogenicity and survival traits, including motility, biofilm formation, antibacterial activity, and nutrient acquisition, highlighting the critical role of QS in pathogen virulence and adaptability.

The quorum-sensing TraI/TraR system contributes to the release of Fe and Al from biotite by Agrobacterium pusense SX41

The quorum-sensing (QS) system in Rhizobiaceae has been studied, however, the mechanisms of QS system-mediated mineral dissolution by bacteria remain poorly understood. Here, the mineral-solubilizing Agrobacterium pusense SX41 harboring the QS system (responsible for N-acyl homoserine lactone (AHL) production) was characterized for QS system-mediated Al and Fe release from biotite. Whole-genome sequence analysis revealed that SX41 contains the QS TraI/TraR system. SX41 and the SX41∆traI, SX41∆traR, and SX41∆traI/R mutants were constructed from SX41 and compared for QS TraI/R system-mediated Fe and Al release from biotite in the solution and the underlying mechanisms. The SX41∆traI, SX41∆traR, and SX41∆traI/R mutants significantly decreased the Al and Fe concentrations in the medium by 24 % to 54 % on day 1 and 25 % to 42 % on day 5, while these mutants significantly reduced exopolysaccharide (EPS) concentrations in the medium by 21 % to 55 % on day 5, compared with those of SX41. Furthermore, these mutants demonstrated decreased cell counts and biofilm formation on biotite surfaces compared with SX41. Notably, the SX41∆traR mutant exhibited more pronounced effects on Al and Fe release from biotite, EPS production, and cell adsorption and biofilm formation on biotite surfaces compared with the SX41∆traI mutant. Moreover, the addition of biotite significantly upregulated the expression of traI and traR in SX41 and the complemented strains SX41∆traIR and SX41∆traRR. These findings underscore the pivotal role of the TraI/R system in the interactions between SX41 and biotite and highlight the distinct influences of traI and traR on Al and Fe release from biotite by SX41.

Impact of AbaI mutation on virulence, biofilm development, and antibiotic susceptibility in Acinetobacter baumannii

The quorum sensing (QS) system mediated by the abaI gene in Acinetobacter baumannii is crucial for various physiological and pathogenic processes. In this study, we constructed a stable markerless abaI knockout mutant (ΔabaI) strain using a pEXKm5-based allele replacement method to investigate the impact of abaI on A. baumannii. Proteomic analysis revealed significant alterations in protein expression between the wild type (WT) and ΔabaI mutant strains, particularly in proteins associated with membrane structure, antibiotic resistance, and virulence. Notably, the downregulation of key outer membrane proteins such as SurA, OmpA, OmpW, and BamA suggests potential vulnerabilities in outer membrane integrity, which correlate with structural abnormalities in the ΔabaI mutant strain, including irregular cell shapes and compromised membrane integrity, observed by scanning and transmission electron microscopy. Furthermore, diminished expression of regulatory proteins such as OmpR and GacA-GacS highlights the broader regulatory networks affected by abaI deletion. Functional assays revealed impaired biofilm formation and surface-associated motility in the mutant strain, indicative of altered colonization capabilities. Interestingly, the mutant showed a complex antibiotic susceptibility profile. While it demonstrated increased susceptibility to membrane-targeting antibiotics, its response to beta-lactams was more nuanced. Despite increased expression of metallo-beta-lactamase (MBL) superfamily proteins and DcaP-like protein, the mutant unexpectedly showed lower MICs for carbapenems (imipenem and meropenem) compared to the wild-type strain. This suggests that abaI deletion affects antibiotic susceptibility through multiple, potentially competing mechanisms. Further investigation is needed to fully elucidate the interplay between quorum sensing, antibiotic resistance genes, and overall antibiotic susceptibility in A. baumannii. Our findings underscore the multifaceted role of the abaI gene in modulating various cellular processes and highlight its significance in A. baumannii physiology, pathogenesis, and antibiotic resistance. Targeting the abaI QS system may offer novel therapeutic strategies for this clinically significant pathogen.

The role of male hormones in bacterial infections: enhancing Staphylococcus aureus virulence through testosterone-induced Agr activation.

Staphylococcus aureus is a notorious pathogen predominantly involved in skin and soft tissue infections, exhibiting a distinct innate sex bias. This study explores the influence of testosterone on the virulence of S. aureus and elucidates its underlying mechanisms. Utilizing a skin abscess model in intact and castrated male mice, we assessed the effects of testosterone on S. aureus pathogenicity. Compared to controls, castrated mice showed significantly reduced abscess sizes and decreased bacterial loads, highlighting the role of testosterone in modulating the severity of S. aureus infections. In vitro experiments revealed that testosterone enhances the hemolytic activity, cytotoxicity, and oxidative stress resistance of S. aureus. Real-time quantitative PCR analysis showed a significant upregulation of the genes encoding α-hemolysin (hla) and phenol-soluble modulin (psmα). Importantly, testosterone treatment significantly enhanced the expression of the accessory gene regulator (Agr) quorum-sensing system components (agrC, agrA, agrB, agrD), while the SaeRS system (saeR, saeS, and sbi) exhibited only slight changes. Gene knockout experiments revealed that deletion of agrC, rather than saeRS and agrBD, abolishes the testosterone-induced enhancement of hemolysis and gene expression, underscoring the key role of AgrC. Molecular docking simulations indicated a direct interaction between testosterone and AgrC protein, with a strong binding affinity at the active site residue SER201. This study provides new insights into the mechanistic basis of how testosterone enhances the pathogenicity of S. aureus, potentially contributing to increased male susceptibility to S. aureus infections and offering a targeted approach for therapeutic interventions.

Erythromycin disrupts Acinetobacter baumannii biofilms through destruction of the quorum sensing system.

This study was conducted to explore the effects of erythromycin on biofilms comprising Acinetobacter baumannii (A baumannii). To clarify the effect of erythromycin on the biofilms of A baumannii, we collected pure Ab strains isolated and identified from a variety of sample types extracted from patients in the microbiological laboratory of our hospital from April to August 2023, and divided them into an experimental group (treated with erythromycin) and a control group (without erythromycin). The morphology and quantity of A baumannii biofilm were observed at 24h, 48h, 72h, and 5d post-treatment, respectively, and the expression of quorum sensing (QS) system gene (abaI, abaR) mRNA was detected by fluorescence quantitative PCR. The results showed that A baumanniis are prone to form multiple drug-resistant (MDR) bacteria, against which the most commonly used clinical antibiotics are ineffective. Overall, we found that the number of bacteria, the number of bacteria in the biofilm, and the number of biofilms formed gradually increased over time, with a statistical difference (P < .05). After the addition of erythromycin, significant improvements in biofilm formation were achieved, indicating that erythromycin can destroy A baumannii biofilms, inhibiting bacterial growth to a certain extent. The expression levels of abaI and abaR gradually increased over time, indicating that the role of the QS system became more apparent over time. Biofilm formation is related to the QS system of A baumanniis. After erythromycin treatment, abaI and abaR mRNA expression was downregulated in the experimental group. Erythromycin disrupts A baumannii biofilms by destroying the quorum sensing system.

The mntH gene of Burkholderia cenocepacia influences motility and quorum sensing to control virulence.

Quorum sensing (QS) signals widely exist in bacteria to control biological functions in response to populations of cells. Burkholderia cenocepacia, an important opportunistic pathogen in patients with cystic fibrosis (CF), is commonly found in the environment and mostly utilizes the N-acylhomoserine lactone (AHL) and cis-2-dodecenoic acid (BDSF) QS systems to control biological functions. Our previous study illuminated the detailed mechanism by which B.cenocepacia integrates BDSF and cyclic diguanosine monophosphate (c-di-GMP) signals to control virulence. Here, we employed Tn5 transposon mutagenesis to identify genes related to the BDSF QS system. One of the most significantly attenuated mutants had an insertion in the mntH gene. Here, we showed that MntH (Bcam0836), a manganese transport protein, controls QS-regulated phenotypes, including motility, biofilm formation and virulence. We also found that. BDSF production was attenuated at both the gene and signaling levels in the Bcam0836 mutant, and that Bcam0836 influenced the expression of some genes regulated by the BDSF receptor RpfR and the downstream regulator GtrR. These results show that the manganese transport protein. MntH modulates a subset of genes and functions regulated by the QS system in B. cenocepacia.

(p)ppGpp interacts with TrmE and regulates the PcoI/PcoR quorum-sensing system of Pseudomonas fluorescens 2P241

The efficient colonization of plant-beneficial Pseudomonas spp. is a prerequisite for their biocontrol capacity. Prior work revealed that the PcoI/PcoR quorum-sensing (QS) system plays a pivotal role in the root colonization of P. fluorescens 2P24. During the colonization, strain 2P24 has faced diverse impacts from plant-derived reactive oxygen species and other environmental stress. However, the molecular mechanism by which the PcoI/PcoR QS system is regulated under unfavored conditions remains unclear. Thus, in this study, the role of the (p)ppGpp synthetase RelA and the bifunctional (p)ppGpp synthase/hydrolase SpoT in the PcoI/PcoR QS system of P. fluorescens was investigated. Our data indicated that the deficiency of relA and spoT genes remarkably improved the expression of the pcoI gene, whereas the mutation of the spoT gene significantly repressed the expression of the pcoI gene. We further demonstrated that the regulation of the PcoI/PcoR QS system by (p)ppGpp was dependent on the function of the trmE gene, which encodes a tRNA modification GTPase. Furthermore, the mutation of relA, spoT, or both significantly influenced the motility, biofilm formation, oxidative stress, osmotic tolerance, and rhizosphere colonization. Collectively, our data indicated that the (p)ppGpp signaling pathway mediated by the relA gene and spoT gene was important to the function of the PcoI/PcoR QS system and had important implications for the understanding of the molecular mechanism of (p)ppGpp in epiphytic fitness via TrmE of P. fluorescens.

Programmable Bacteria with Dynamic Virulence Modulation System for Precision Antitumor Immunity.

Engineered bacteria-mediated antitumor approaches have been proposed as promising immunotherapies for cancer. However, the off-target bacterial toxicity narrows the therapeutic window. Living microbes will benefit from their controllable immunogenicity within tumors for safer antitumor applications. In this study, a genetically encoded microbial activation strategy is reported that uses tunable and dynamic expression of surface extracellular polysaccharides to improve bacterial biocompatibility while retaining therapeutic efficacy. Based on screening of genes associated with Salmonella survival in macrophages, a novel attenuated Salmonella chassis strain AIS (htrA gene-deficient) highly enriched in tumors after administration and rapidly cleared from normal organs are reported. Subsequently, an engineered bacterial strain, AISI-H, is constructed based on the AIS strain and an optimized quorum-sensing regulatory system. The AISI-H strain can achieve recovery of dynamic tumor-specific bacterial virulence through a novel HTRA-RCSA axis-based and quorum-sensing synthetic gene circuit-mediated increase in extracellular polysaccharide content. These strains act "off" in normal organs to avoid unwanted immune activation and "on" in tumors for precise tumor suppression in mice. The AISI-H strain shows significant tumor inhibition and potent activation of anticancer immunity in a melanoma mouse model. The AISI-H strain exhibits excellent biocompatibility. This bacterial regulation strategy expands the applications of microbe-based antitumor therapeutics.

Metabolic activities of marine ammonia-oxidizing archaea orchestrated by quorum sensing.

Ammonia-oxidizing archaea (AOA) play crucial roles in marine carbon and nitrogen cycles by fixing inorganic carbon and performing the initial step of nitrification. Evaluation of carbon and nitrogen metabolism popularly relies on functional genes such as amoA and accA. Increasing studies suggest that quorum sensing (QS) mainly studied in biofilms for bacteria may serve as a universal communication and regulatory mechanism among prokaryotes; however, this has yet to be demonstrated in marine planktonic archaea. To bridge this knowledge gap, we employed a combination of metabolic activity markers (amoA, accA, and grs) to elucidate the regulation of AOA-mediated nitrogen, carbon processes, and their interactions with the surrounding heterotrophic population. Through co-transcription investigations linking metabolic markers to potential key QS genes, we discovered that QS molecules could regulate AOA's carbon, nitrogen, and lipid metabolisms under different conditions. Interestingly, specific AOA ecotypes showed a preference for employing distinct QS systems and a distinct QS circuit involving a typical population. Overall, our data demonstrate that QS orchestrates nitrogen and carbon metabolism, including the exchange of organic metabolites between AOA and surrounding heterotrophic bacteria, which has been previously overlooked in marine AOA research.

The activity of the quorum sensing regulator HapR is modulated by the bacterial extracellular vesicle (BEV)-associated protein ObfA of Vibrio cholerae.

Vibrio cholerae, a facultative human pathogen and causative agent of the severe diarrheal disease cholera, transits between the human intestinal tract and aquatic reservoirs. Like other bacterial species, V. cholerae continuously releases bacterial extracellular vesicles (BEVs) from its surface, which have been recently characterised for their role during in vivo colonisation. However, between epidemic outbreaks, V. cholerae persists in the biofilm mode for extended periods in aquatic reservoirs, which enhances environmental fitness and host transition. In this study, we investigated the effect of V. cholerae BEVs on biofilm formation, a critical feature for ex vivo survival. In contrast to BEVs from planktonic cultures, our results show that physiological concentrations of BEVs from dynamic biofilm cultures facilitate V. cholerae biofilm formation, which could be linked to a proteinaceous factor. Comparative proteomic analyses of planktonic- and biofilm-derived BEVs identified a previously uncharacterised outer membrane protein as an abundant component of dynamic biofilm-derived BEVs, which was found to be responsible for the BEV-dependent enhancement of biofilm production. Consequently, this protein was named outer membrane-associated biofilm facilitating protein A (ObfA). Comprehensive molecular studies unravelled ObfA as a negative modulator of HapR activity. HapR is a key transcriptional regulator of the V. cholerae quorum sensing (QS) cascade acting as a potent repressor of biofilm formation and virulence. Consistently, obfA mutants not only exhibited reduced biofilm production but also reduced colonisation fitness. Surprisingly, our results demonstrate that ObfA does not affect HapR through the canonical QS system but via the Csr-cascade altering the expression of the small regulatory RNAs CsrC and CsrD. In summary, this study elucidates a novel intraspecies BEV-based communication in V. cholerae that influences biofilm formation and colonisation fitness via a new regulatory pathway involving HapR, Csr-cascade and the BEV-associated protein ObfA.

The absence of luxS reduces the invasion of Avibacterium paragallinarum but is not essential for virulence.

The contagious respiratory pathogen, Avibacterium paragallinarum, contributes to infectious coryza in poultry. However, commercial vaccines have not shown perfect protection against infectious coryza. To search for an alternative approach, this research aimed to investigate whether the quorum-sensing system of pathogens plays a crucial role in their survival and pathogenicity. The LuxS/AI-2 quorum-sensing system in many Gram-negative and Gram-positive bacteria senses environmental changes to regulate physiological traits and virulent properties, and the role of the luxS gene in Av. paragallinarum remains unclear. To investigate the effect of the luxS gene in the quorum-sensing system of Av. paragallinarum, we constructed a luxS mutant. Bioluminescence analysis indicated that the luxS gene plays a vital role in the LuxS/AI-2 quorum-sensing system. The analysis of the LuxS/AI-2 system-related genes showed the level of pfs mRNA to be significantly increased in the mutant strain; however, lsrR, lsrK, and lsrB mRNA levels were not significantly different compared with the wild type. The ability of the luxS mutant strain to invade HD11 and DF-1 cells was significantly decreased compared with the wild-type strain. In addition, all chickens challenged with various doses of the luxS mutant strain developed infections and symptoms, and those challenged with the lowest dose exhibited only minor differences compared to chickens challenged with the wild-type strain. Thus, the deletion of the luxS gene reduces the invasion, but the luxS gene does not play an essential role in the pathogenesis of A. paragallinarum.

Impact of Graphene Layers on Genetic Expression and Regulation within Sulfate-Reducing Biofilms.

Bacterial adhesion and biofilm maturation is significantly influenced by surface properties, encompassing both bare surfaces and single or multi-layered coatings. Hence, there is an utmost interest in exploring the intricacies of gene regulation in sulfate-reducing bacteria (SRB) on copper and graphene-coated copper surfaces. In this study, Oleidesulfovibrio alaskensis G20 was used as the model SRB to elucidate the pathways that govern pivotal roles during biofilm formation on the graphene layers. Employing a potent reporter green fluorescent protein (GFP) tagged to O. alaskensis G20, the spatial structure of O. alaskensis G20 biofilm on copper foil (CuF), single-layer graphene-coated copper (Cu-GrI), and double-layer graphene-coated copper (Cu-GrII) surfaces was investigated. Biofilm formation on CuF, Cu-GrI, and Cu-GrII surfaces was quantified using CLSM z-stack images within COMSTAT v2 software. The results revealed that CuF, Cu-GrI, and Cu-GrII did not affect the formation of the GFP-tagged O. alaskensis G20 biofilm architecture. qPCR expression showed insignificant fold changes for outer membrane components regulating the quorum-sensing system, and global regulatory proteins between the uncoated and coated surfaces. Notably, a significant expression was observed within the sulfate reduction pathway confined to dissimilatory sulfite reductases on the Cu-GrII surface compared to the CuF and Cu-GrI surfaces.