Biofilm Dispersal Research Articles

Interface-Engineered CuxO@Bi2MoO6 Heterojunctions to Inhibit Piezoelectric Screening Effect and Promote Double-Nanozyme Catalysis for Antibacterial Treatment.

Sonodynamic therapy is confronted with the low acoustic efficiency of sonosensitizers, and nanozymes are accompanied by intrinsic low catalytic activity. Herein, to increase the piezopotential of N-type piezoelectric semiconductors, the P-N heterojunction is designed to inhibit the piezoelectric screening effect (PSE) and increase electron utilization efficiency to enhance nanozyme activity. P-type CuxO nanoparticles are in situ grown on N-type piezoelectric Bi2MoO6 (BMO) nanoflakes (NFs) to construct heterostructured CuxO@BMO by interface engineering. CuxO deposition leads to lattice distortion of BMO NFs to improve piezoelectric response, and the strong interface electric field (IEF) suppresses PSE and increases piezopotential. The nonlocal piezopotential, local IEF, and glutathione (GSH) inoculation enhances electron-hole separation and increases peroxidase (POD)-like activity of BMO and GSH oxidase (GSHOx)-like activity of CuxO with high selectivity. The heterojunction formation causes the transfer and rearrangement of interface electrons, and the increased piezopotential accelerates electron transfer at interfaces with bacteria, thus increasing the production of reactive oxidative species and interfering with adenosine triphosphate synthesis. The heterostructured nanozymes produce abundant intracellular ·OH and achieve 4log magnitude reductions in viable bacteria and effective biofilm dispersion. This study elucidates integral mechanisms of nanozyme and acoustic catalysis and opens up a new way to synergize high piezopotential and nanozyme-catalyzed therapy.

Microplastics colonized by Hafnia paralvei through biofilm formation regulated by c-di-GMP and cAMP promote its spread in water

Microplastics (MPs) colonized by pathogens pose significant risks to the environment and health of animals and humans, however, the strategies for pathogens colonization in MPs and the effects of its colonization on spread of pathogens have not been fully characterized. Here, we investigated the biofilm formation mechanism regulated by c-di-GMP in Hafnia paralvei Z11, and determined the effect of MPs colonized by H. paralvei Z11 on the spread of strain Z11. Overexpression of yhjH, a c-di-GMP phosphodiesterase gene, attenuated intracellular c-di-GMP level in strain Z11, leading to an increase in biofilm dispersal and a decrease in biofilm formation. Meanwhile, the decline of c-di-GMP inhibited the expression of cAMP phosphodiesterase genes, increasing the cAMP content and promoting bacterial motility, that was responsible for the increase of biofilm dispersal. Furthermore, the formation of biofilms by strain Z11 on MPs promotes its colonization, which contributes to its vertical and horizontal spread in water after colonizing polyvinyl chloride-MPs and polypropylene-MPs, respectively. Therefore, this study reveals, for the first time, MPs colonized by H. paralvei Z11 through biofilms regulated by crosstalk between c-di GMP and cAMP promote the spread of strain Z11 in water, which expands the understanding of colonization strategy of pathogens on MPs and its risk on spread of pathogens.

Active-bromide and surfactant synergy for enhanced microfouling control.

Biofilms are structured microbial communities encased in a matrix of self-produced extracellular polymeric substance (EPS) and pose significant challenges in various industrial cooling systems. A nuclear power plant uses a biocide active-bromide for control of biological growth in its condenser cooling system. This study is aimed at evaluating the anti-bacterial and anti-biofilm efficacy of active-bromide against planktonic and biofilm-forming bacteria that are commonly encountered in seawater cooling systems. The results demonstrated that active-bromide at the concentration used at the power plant (1 ppm) exhibited minimal killing activity against Pseudomonas aeruginosa planktonic cells. The bacterial cell surface hydrophobicity assay using Staphylococcus aureus and P. aeruginosa indicated that Triton-X 100 significantly decreased the hydrophobicity of planktonic cells, enhancing the susceptibility of the cells to active-bromide. Biofilm inhibition assays revealed limited efficacy of active-bromide at 1 ppm concentration, but significant inhibition at 5 ppm and 10 ppm. However, the addition of a surfactant, Triton-X 100, in combination with 1 ppm active-bromide displayed a synergistic effect, leading to significant biofilm dispersal of pre-formed P. aeruginosa biofilms. This observation was substantiated by epifluorescence microscopy using a live/dead bacterial assay that showed the combination treatment resulted in extensive cell death within the biofilm, as indicated by a marked increase in red fluorescence, compared to treatments with either agent alone. These findings suggest that active bromide alone may be insufficient for microfouling control in the seawater-based condenser cooling system of the power plant. Including a biocompatible surfactant that disrupts established biofilms (microfouling) can significantly improve the efficacy of active bromide treatment.

Genome-Centric Metagenomics Insights into the Plastisphere-Driven Natural Degradation Characteristics and Mechanism of Biodegradable Plastics in Aquatic Environments.

Biodegradable plastics (BPs) are pervasively available as alternatives to traditional plastics, but their natural degradation characteristics and microbial-driven degradation mechanisms are poorly understood, especially in aquatic environments, the primary sink of plastic debris. Herein, the three-month dynamic degradation process of BPs (the copolymer of poly(butylene adipate-co-terephthalate) and polylactic acid (PLA) (PBAT/PLA) and single PLA) in a natural aquatic environment was investigated, with nonbiodegradable plastics polyvinyl chloride, polypropylene, and polystyrene as controls. PBAT/PLA showed the weight loss of 47.4% at 50 days and severe fragmentation within two months, but no significant decay for other plastics. The significant increase in the specific surface area and roughness and the weakening of hydrophobicity within the first month promoted microbial attachment to the PBAT/PLA surface. Then, a complete microbial succession occurred, including biofilm formation, maturation, and dispersion. Metagenomic analysis indicated that plastispheres selectively enriched degraders. Based on the functional genes involved in BPs degradation, a total of 16 high-quality metagenome-assembled genomes of degraders (mainly Burkholderiaceae) were recovered from the PBAT/PLA plastisphere. These microbes showed the greatest degrading potential at the biofilm maturation stage and executed the functions by PLA_depolymerase, polyesterase, hydrolase, and esterase. These findings will enhance understanding of BPs' environmental behavior and microbial roles on plastic degradation.

Holliday junction resolvase RuvC targets biofilm eDNA and confers plant resistance to vascular pathogens.

A biofilm lifestyle is critical for bacterial pathogens to colonize and protect themselves from host immunity and antimicrobial chemicals in plants and animals. The formation and regulation mechanisms of phytobacterial biofilm are still obscure. Here we found that the protein Ralstonia solanacearum resistance to ultraviolet C (RuvC) is highly abundant in biofilm and positively regulates pathogenicity by controlling systemic movement in tomato xylem. RuvC protein accumulates at the later stage of biofilm development and specifically targets Holliday junction (HJ)-like structures to disrupt the biofilm extracellular DNA (eDNA) lattice, thus facilitating biofilm dispersal. Recombinant RuvC protein can resolve extracellular HJ to prevent bacterial biofilm formation. Heterologous expression of R. solanacearum or Xanthomonas oryzae pv. oryzae RuvC with plant secretion signal in tomato or rice confers resistance to bacterial wilt or bacterial blight disease, respectively. Plant chloroplast-localized HJ resolvase monokaryotic chloroplast 1 (MOC1), which shares structural similarity with bacterial RuvC, shows a strong inhibitory effect on bacterial biofilm formation. Relocalization of SlMOC1 to apoplast in tomato roots leads to increased resistance to bacterial wilt. Our novel finding reveals a critical pathogenesis mechanism of R. solanacearum and provides an efficient biotechnology strategy to improve plant resistance to bacterial vascular disease.

Integrated Biofilm Dispersion and Virulence Responsiveness for Targeted Treatment of Pseudomonas aeruginosa Infection in Lungs

AbstractThe self‐organization of microbes into biofilms provides multiple benefits including tolerance to mechanical stress and resistance to immune defences and antibiotics. Coupled to a compromised mucociliary function, these traits have dire consequences in cystic fibrosis patients – persistent infections are the main reason for morbidity and mortality. Thereby, disease progression is associated with universal colonization by Pseudomonas aeruginosa, which selects for a slimy phenotype to adapt to the lung microenvironment. Recognizing this, drug‐delivery vehicles that break down the mucoid extracellular matrix made of alginate are designed to enable better penetration and biofilm dispersion. In parallel, a protective layer responds to the proteolytic activity of the pathogen and thus controls drug availability. To realize this architecture, silica nanoparticles are loaded with imipenem, and then coated with elastin and alginate lyase in a layer‐by‐layer fashion using ultrasound. The nanoscale formulations eradicate up to 80% of the total biomass and reduce the bacterial viability in biofilms by 3 logs, considerably outperforming the bulk antibiotic in vitro, whereby the effects are correlated to changes in the viscoelasticity. Furthermore, the stimuli‐responsive nanocarriers are safe and effective in animal models of P. aeruginosa infection, presenting a considerable therapeutic promise in the challenging context of lung diseases.

Chemical signaling in biofilm-mediated biofouling.

Biofouling is the undesirable accumulation of living organisms and their metabolites on submerged surfaces. Biofouling begins with adhesion of biomacromolecules and/or microorganisms and can lead to the subsequent formation of biofilms that are predominantly regulated by chemical signals, such as cyclic dinucleotides and quorum-sensing molecules. Biofilms typically release chemical cues that recruit or repel other invertebrate larvae and algal spores. As such, harnessing the biochemical mechanisms involved is a promising avenue for controlling biofouling. Here, we discuss how chemical signaling affects biofilm formation and dispersion in model species. We also examine how this translates to marine biofouling. Both inductive and inhibitory effects of chemical cues from biofilms on macrofouling are also discussed. Finally, we outline promising mitigation strategies by targeting chemical signaling to foster biofilm dispersion or inhibit biofouling.

New commercial wipes inhibit the dispersion and adhesion of Staphylococcus aureus and Pseudomonas aeruginosa biofilms.

Bacterial biofilms can form on surfaces in hospitals, clinics, farms, and food processing plants, representing a possible source of infections and cross-contamination. This study investigates the effectiveness of new commercial wipes against Staphylococcus aureus and Pseudomonas aeruginosa biofilms (early attachment and formed biofilms), assessing LH SALVIETTE wipes (Lombarda H S.r.l.) potential for controlling biofilm formation. The wipes efficacy was studied against the early attachment phase and formed biofilm of S. aureus ATCC 6538 and P. aeruginosa ATCC 15442 on a polyvinyl chloride (PVC) surface, following a modified standard test EN 16615:2015, measuring Log10 reduction and cell viability using live/dead staining. It was also evaluated the wipes anti-adhesive activity over time (3 h, 2 4h), calculating CFU.mL-1 reduction. Data were analyzed using t-student test. The wipes significantly reduced both early phase and formed S. aureus biofilm, preventing dispersion on PVC surfaces. Live/dead imaging showed bacterial cluster disaggregation and killing action. The bacterial adhesive capability decreased after short-time treatment (3 h) with the wipes compared to 24 h. Results demonstrated decreased bacterial count on PVC surface both for early attachment phase and formed biofilms, also preventing the bacterial biofilm dispersion.

Comprehensive Genomic and Evolutionary Analysis of Biofilm Matrix Clusters and Proteins in the Vibrio Genus.

Vibrio cholerae pathogens cause cholera, an acute diarrheal disease resulting in significant morbidity and mortality worldwide. Biofilm formation by V. cholerae enhances its survival in natural ecosystems and facilitates transmission during cholera outbreaks. Critical components of the biofilm matrix are the Vibrio polysaccharide (VPS) produced by the vps-1 and vps-2 gene clusters, and biofilm matrix proteins encoded in the rbm cluster. However, the biofilm matrix clusters and associated matrix proteins in other Vibrio species remain under investigated, and their evolutionary patterns are largely unknown. In this study, we systematically annotated the biofilm matrix clusters across 6,121 Vibrio genomes, revealing their distribution, diversity, and evolution. We found that biofilm matrix clusters not only exist in V. cholerae but also in phylogenetically distant Vibrio species. Additionally, vps-1 clusters tend to co-locate with rbmABC genes, while vps-2 clusters are often adjacent to rbmDEF genes in various Vibrio species, which helps explain the separation of these clusters by the rbm cluster in well-characterized V. cholerae strains. Evolutionary analysis of RbmC and Bap1 reveals that these two major biofilm matrix proteins are sequentially and structurally related and have undergone domain/modular alterations during their evolution. RbmC genes are more prevalent, while bap1 likely resulted from an ancient duplication event of rbmC and is only present in a major clade of species containing rbmC counterparts. Notably, a novel loop-less Bap1 variant, identified in two subspecies clades of V. cholerae, was found to be associated with altered biofilm formation and the loss of antibiotic efflux pumps and chemotaxis. Another rbm cluster gene, rbmB, involved in biofilm dispersal, was found to share a common ancestor with Vibrio prophage pectin lyase-like tail proteins, indicating its functional and evolutionary linkages to Vibriophage proteins. In summary, our findings establish a foundational understanding of the proteins and gene clusters that contribute to Vibrio biofilm formation from an evolutionary perspective across a broad taxonomic scale. This knowledge paves the way for future strategies aimed at engineering and controlling biofilms through genetic modification.

Bioemulsifier from sponge-associated bacteria reduces staphylococcal biofilm

Biofilm formation is a major health concern and studies have been pursued to find compounds able to prevent biofilm establishment and remove pre-existing biofilms. While biosurfactants (BS) have been well-known for possessing antibiofilm activities, bioemulsifiers (BE) are still scarcely explored for this purpose. The present study aimed to evaluate the bioemulsifying properties of cell-free supernatants produced by Bacillaceae and Vibrio strains isolated from marine sponges and investigate their antiadhesive and antibiofilm activities against different pathogenic Gram-positive and Gram-negative bacteria. The BE production by the marine strains was confirmed by the emulsion test, drop-collapsing, oil-displacement, cell hydrophobicity and hemolysis assays. Notably, Bacillus cereus 64BHI1101 displayed remarkable emulsifying activity and the ultrastructure analysis of its BE extract (BE64-1) revealed the presence of structures typically observed in macromolecules composed of polysaccharides and proteins. BE64-1 showed notable antiadhesive and antibiofilm activities against Staphylococcus aureus, with a reduction of adherence of up to 100 % and a dispersion of biofilm of 80 %, without affecting its growth. BE64-1 also showed inhibition of Staphylococcus epidermidis and Escherichia coli biofilm formation and adhesion. Thus, this study provides a starting point for exploring the antiadhesive and antibiofilm activities of BE from sponge-associated bacteria, which could serve as a valuable tool for future research to combat S. aureus biofilms.

Synthetic mucus biomaterials synergize with antibiofilm agents to combat Pseudomonas aeruginosa biofilms.

Bacterial biofilms are often highly resistant to antimicrobials causing persistent infections which when not effectively managed can significantly worsen clinical outcomes. As such, alternatives to standard antibiotic therapies have been highly sought after to address difficult-to-treat biofilm-associated infections. We hypothesized a biomaterial-based approach using the innate functions of mucins to modulate bacterial surface attachment and virulence could provide a new therapeutic strategy against biofilms. Based on our testing in Pseudomonas aeruginosa biofilms, we found synthetic mucus biomaterials can inhibit biofilm formation and significantly reduce the thickness of mature biofilms. In addition, we evaluated if synthetic mucus biomaterials could work synergistically with DNase and/or α-amylase for enhanced biofilm dispersal. Combination treatment with these antibiofilm agents and synthetic mucus biomaterials resulted in up to 3 log reductions in viability of mature P. aeruginosa biofilms. Overall, this work provides a new bio-inspired, combinatorial approach to address biofilms and antibiotic-resistant bacterial infections.

Combatting Antibiotic-Resistant Pathogens: Disrupting Pseudomonas Aeruginosa Biofilm Viscoelasticity Using Carbon Dot Nanoparticles

Bacterial infection is a global burden, killing one in every eight people worldwide, primarily due to antibiotic resistance. Pseudomonas aeruginosa is one of the top five bacteria responsible for around 7.7 million fatalities caused by bacterial infections. P. aeruginosa secretes biofilm, a biological substance with viscoelastic characteristics to avoid antimicrobial treatments particularly, in chronic and serious wound infections. In the bid to create topical treatments, we investigated the mechano-active potentials of cellulase-based carbon dots as a therapeutic agent against P. aeruginosa biofilm where we envisage the retaining of the cellulase hydrolytic activity in the cellulase-based carbon dots. Cellulase is a glycoside hydrolase that has already been shown to be effective in causing biofilm dispersal as well as reduction in biofilm stiffness and elasticity.The cellulase carbon dots were synthesized using the microwave while citric acid carbon dots (CA-CDs) were hydrothermally synthesized to serve as non-protein-based carbon dots and then characterized for surface properties, size, elemental compositions, and fluorescence profile. P. aeruginosa biofilms were grown in a microfluidic environment for 24 hours and then treated with these carbon dots at a range of concentrations. Using microrheology, it was seen that biofilms treated with cellulase-based carbon dots were more viscous and less stiff compared to those treated with citric acid-based carbon dots. This appears to be because the cellulase carbon dots impact PSL within the biofilm based on microrheology studies examining bacteria strains with specific EPS components removed. This is consistent with the impact of molecular cellulase on biofilms. In general, initial studies suggest that the cellulase-based carbon dots have the potential to act as an excellent multi-antibiofilm therapeutic in the future.

A rare biofilm dispersion strategy demonstrated by Staphylococcus aureus under oxacillin stress

Staphylococcus aureus (S. aureus), a versatile Gram-positive bacterium, is implicated in a spectrum of infections, and its resilience is often attributed to biofilm formation. This study investigates the effect of sub-inhibitory doses of oxacillin on biofilm formation by methicillin-resistant S. aureus (MRSA). Specifically, it examines how these doses influence biofilms' development, maturation, and dispersal. The biofilm's zenith reached 48 h of incubation, followed by a noteworthy decline at 96 h and a distinctive clearance zone around biofilm-positive cells exposed to oxacillin. Scanning electron micrographs unveiled an intriguing active biofilm dispersal mechanism, a rarity in this species. Among 180 isolates, only three carrying the elusive icaD gene exhibited this phenomenon. icaD gene was absent in their counterparts. Notably, the icaD gene emerges as a distinctive marker, crucial in regulating biofilm dispersion and setting these isolates apart. The captivating interplay of oxacillin, biofilm dynamics, and genetic signatures disintegrate novel dimensions in understanding MRSA's adaptive strategies and underscores the importance of the icaD gene in engineering biofilm resilience.

Neutrophil Extracellular Traps-Inspired Bismuth-Based Polypeptide Nanonets for Synergetic Treatment of Bacterial Infections.

Excessive use of antibiotics and the formation of bacterial biofilms can lead to persistent infections caused by drug-resistant bacteria, rendering ineffective immune responses and even life-threatening. There is an urgent need to explore synergistic antibacterial therapies across all stages of infection. Drawing inspiration from the antibacterial properties of neutrophil extracellular traps (NETs) and integrating the bacterial biofilm dispersal mechanism involving boronic acid-catechol interaction, the multifunctional bismuth-based polypeptide nanonets (PLBA-Bi-Fe-TA) are developed. These nanonets are designed to capture bacteria through a coordination complex involving cationic polypeptides (PLBA) with boronic acid-functionalized side chains, alongside metal ions (bismuth (Bi) and iron (Fe)), and tannic acid (TA). Leveraging the nanoconfinement-enhanced high-contact network-driven multiple efficiency, PLBA-Bi-Fe-TA demonstrates the excellent ability to swiftly capture bacteria and their extracellular polysaccharides. This interaction culminates in the formation of a highly hydrophilic complex, effectively enabling the rapid inhibition and dispersion of antibiotic-resistant bacterial biofilms, while Fe-TA shows mild photothermal ability to further assist fluffy mature biofilm. In addition, Bi is beneficial to regulate the polarization of macrophages to pro-inflammatory phenotype to further kill escaping biofilm bacteria. In summary, this novel approach offers a promising bionic optimization strategy for treating bacterial-associated infections at all stages through synergetic treatment.

Epsilon-poly-l-lysine inhibits biofilm formation and aids dispersion in Acinetobacter baumannii

Acinetobacter baumannii is a prominent hospital-associated bacterium whose eradication is increasingly challenging due to its remarkable ability to resist antibiotics. The most common illnesses caused by antibiotic-resistant A. baumannii are biofilm-associated. Therefore, novel methods to combat A. baumannii are urgently needed. Application of antimicrobial peptides (AMPs) is one of the avenues to be explored. Epsilon poly-l-lysine (ε-PL) is an antimicrobial peptide with low mammalian toxicity. It is commonly used as a food preservative and has advantages such as biodegradability, good water solubility, and thermal stability.Therefore, the antimicrobial activity of ε-PL against clinical isolates of A. baumannii was investigated. The effect of ε-PL on antimicrobial sensitivity was determined by broth dilution assay. The effect of ε-PL on biofilm formation and dispersion was studied using a crystal violet assay. The changes in the expression of quorum sensing related and virulence genes (abaI, csuE, pilT, bap, and luxI) were analyzed using qPCR by the Δ Δ CT method.All the A. baumannii clinical isolates (n = 28) tested, were resistant to multiple drugs. The treatment with ε-PL resulted in a significant (p < 0.05) reduction in the biofilm formation abilities of all the clinical isolates of A. baumannii. Also, ε-PL caused a significant (p < 0.05) decrease in the dispersion of preformed biofilms. The reduction in the biofilm formation could be attributed to the inhibition of autoinducer synthase (abaI) which is required for biofilm development in A. baumannii. Also, it could be due to altering of expression of biofilm-related genes like csuE, pilT, bap, and luxI. These results suggest that ε-PL could be effective in the elimination of A. baumannii biofilms and decreasing its virulence.

Legionella pneumophila cell surface RtxA release by LapD/LapG and its role in virulence

BackgroundLegionella pneumophila is a Gram-negative intracellular bacillus and is the causative agent of a severe form of pneumonia called Legionnaires’ disease which accounts for 2-9% of cases of community acquired pneumonia. It produces an extremely large protein belonging to the RTX (Repeats in ToXin) family, called RtxA, and we previously reported that RtxA is transported by a dedicated type 1 secretion system (T1SS) to the cell surface. RTX proteins have been shown to participate in the virulence or biofilm formation of various bacteria, the most studied models being the pore forming hemolysin A (HlyA) of Escherichia coli and the biofilm associated protein LapA of P. fluorescens. LapA localization depends on the enzymatic release by LapD/LapG complex activity. This study aimed to elucidate the dual localization (cell surface associated or released state) of L. pneumophila RTX protein (RtxA) and whether this released versus sequestered state of RtxA plays a role in L. pneumophila virulence.ResultsThe hereby work reveals that, in vitro, LapG periplasmic protease cleaves RtxA N-terminus in the middle of a di-alanine motif (position 108–109). Consistently, a strain lacking LapG protease maintains RtxA on the cell surface, whereas a strain lacking the c-di-GMP receptor LapD does not exhibit cell surface RtxA because of its continuous cleavage and release, as in the LapA-D-G model of Pseudomonas fluorescens. Interestingly, our data point out a key role of RtxA in enhancing the infection process of amoeba cells, regardless of its location (embedded or released); therefore, this may be the result of a secondary role of this surface protein.ConclusionsThis is the first experimental identification of the cleavage site within the RTX protein family. The primary role of RtxA in Legionella is still questionable as in many other bacterial species, hence it sounds reasonable to propose a major function in biofilm formation, promoting cell aggregation when RtxA is embedded in the outer membrane and facilitating biofilm dispersion in case of RtxA release. The role of RtxA in enhancing the infection process may be a result of its action on host cells (i.e., PDI interaction or pore-formation), and independently of its status (embedded or released).

Transcriptional responses of Neisseria gonorrhoeae to glucose and lactate: implications for resistance to oxidative damage and biofilm formation.

Understanding how bacteria adapt to different environmental conditions is crucial for advancing knowledge regarding pathogenic mechanisms that operate during infection as well as efforts to develop new therapeutic strategies to cure or prevent infections. Here, we investigated the transcriptional response of Neisseria gonorrhoeae, the causative agent of gonorrhea, to L-lactate and glucose, two important carbon sources found in the host environment. Our study revealed extensive transcriptional changes that gonococci make in response to L-lactate, with 37% of the gonococcal transcriptome being regulated, compared to only 9% by glucose. We found that L-lactate induces a transcriptional program that would negatively impact iron transport, potentially limiting the availability of labile iron, which would be important in the face of the multiple hydrogen peroxide attacks encountered by gonococci during its lifecycle. Furthermore, we found that L-lactate-mediated transcriptional response promoted aerobic respiration and dispersal of biofilms, contrasting with an anaerobic condition previously reported to favor biofilm formation. Our findings suggest an intricate interplay between carbon metabolism, iron homeostasis, biofilm formation, and stress response in N. gonorrhoeae, providing insights into its pathogenesis and identifying potential therapeutic targets.IMPORTANCEGonorrhea is a prevalent sexually transmitted infection caused by the human pathogen Neisseria gonorrhoeae, with ca. 82 million cases reported worldwide annually. The rise of antibiotic resistance in N. gonorrhoeae poses a significant public health threat, highlighting the urgent need for alternative treatment strategies. By elucidating how N. gonorrhoeae responds to host-derived carbon sources such as L-lactate and glucose, this study offers insights into the metabolic adaptations crucial for bacterial survival and virulence during infection. Understanding these adaptations provides a foundation for developing novel therapeutic approaches targeting bacterial metabolism, iron homeostasis, and virulence gene expression. Moreover, the findings reported herein regarding biofilm formation and L-lactate transport and metabolism contribute to our understanding of N. gonorrhoeae pathogenesis, offering potential avenues for preventing and treating gonorrhea infections.

Biofilm dispersal patterns revealed using far-red fluorogenic probes.

Bacteria frequently colonize niches by forming multicellular communities called biofilms. To explore new territories, cells exit biofilms through an active process called dispersal. Biofilm dispersal is essential for bacteria to spread between infection sites, yet how the process is executed at the single-cell level remains mysterious. Here, we characterize dispersal at unprecedented resolution for the global pathogen Vibrio cholerae. To do so, we first developed a far-red cell-labeling strategy that overcomes pitfalls of fluorescent protein-based approaches. We reveal that dispersal initiates at the biofilm periphery and ~25% of cells never disperse. We define novel micro-scale patterns that occur during dispersal, including biofilm compression and the formation of dynamic channels. These patterns are attenuated in mutants that reduce overall dispersal or that increase dispersal at the cost of homogenizing local mechanical properties. Collectively, our findings provide fundamental insights into the mechanisms of biofilm dispersal, advancing our understanding of how pathogens disseminate.

D-Methionine-induced DNases disperse established Burkholderia pseudomallei biofilms and promotes ceftazidime susceptibility

Burkholderia pseudomallei biofilm is correlated with pathogenesis, antibiotic resistance, and relapsing cases of melioidosis, leading to challenges in clinical management. There is increasing interest in employing biofilm dispersal agents as adjunctive treatments for biofilm-associated infections. Methionine (Met) has shown promise as an anti-biofilm agent by inducing bacterial DNase production, resulting in the degradation of extracellular DNA (eDNA) and dispersion of bacterial biofilm. In this study, we investigated the impact of 0.05–50 μM D-Met and L-Met on the 24-h established biofilm of a clinical isolate, B. pseudomallei H777. Our findings revealed the ability of D-Met and L-Met to disperse the established biofilm in a non-dose-dependent manner accompanied by eDNA depletion. Real-time PCR analysis further identified an up-regulation of bacterial nuclease genes, including recJ, eddB, nth, xth, and recD, in the presence of 0.05 μM D-Met. Similarly, recJ and eddB in B. pseudomallei were up-regulated in response to the presence of 0.05 μM L-Met. Notably, D-Met enhanced the susceptibility of B. pseudomallei H777 biofilm cells to ceftazidime. Our findings indicate a correlation between methionine supplementation and the up-regulation of nuclease genes, leading to eDNA depletion and the dispersal of preformed B. pseudomallei H777 biofilm. This enhances the susceptibility of biofilm cells to ceftazidime, showing promise in combating biofilm-associated B. pseudomallei infections.

Tyrosol blocks E. coli anaerobic biofilm formation via YbfA and FNR to increase antibiotic susceptibility

Bacteria within mature biofilms are highly resistant to antibiotics than planktonic cells. Oxygen limitation contributes to antibiotic resistance in mature biofilms. Nitric oxide (NO) induces biofilm dispersal; however, low NO levels stimulate biofilm formation, an underexplored process. Here, we introduce a mechanism of anaerobic biofilm formation by investigating the antibiofilm activity of tyrosol, a component in wine. Tyrosol inhibits E. coli and Pseudomonas aeruginosa biofilm formation by enhancing NO production. YbfA is identified as a target of tyrosol and its downstream targets are sequentially determined. YbfA activates YfeR, which then suppresses the anaerobic regulator FNR. This suppression leads to decreased NO production, elevated bis-(3’−5’)-cyclic dimeric GMP levels, and finally stimulates anaerobic biofilm formation in the mature stage. Blocking YbfA with tyrosol treatment renders biofilm cells as susceptible to antibiotics as planktonic cells. Thus, this study presents YbfA as a promising antibiofilm target to address antibiotic resistance posed by biofilm-forming bacteria, with tyrosol acting as an inhibitor.