Biofilm Formation Research Articles

The effects of anodization and instrumentation on titanium abutment surface characteristics and biofilm formation.

To assess the impact of anodization and instrumentation on titanium abutment surface characteristics (surface roughness and wettability) and biofilm formation (viability and mass). Titanium discs were obtained from pre-milled abutment blanks made of titanium-6aluminum-7niobium alloy. Polished samples were divided into three groups: un-anodized, gold-anodized, and pink-anodized. Instrumentation methods included no-instrumentation, air polishing, and titanium scaling treatment. Surface roughness was measured using an optical profilometer, and wettability was determined by measuring the contact angles using the sessile drop method with an optical tensiometer. Biofilm formation by Streptococcus sanguinis was evaluated based on the biofilm viability and mass. The biofilm viability was evaluated through colony-forming unit counting (CFU/mL), and biofilm mass was assessed with crystal violet staining (mean absorbance measured at 490nm, in optical density values). Sample surfaces before and after biofilm formation were also examined by scanning electron microscope (SEM). Two-way ANOVA was performed to determine the group differences, and Spearman's correlation (ρ) was used to analyze the correlation among surface roughness, wettability, and CFU/mL (α=0.05). Pink anodization significantly increased surface roughness (0.38±0.07µm, p<0.001) compared to un-anodized samples (0.25±0.01µm), while gold anodization did not (0.24±0.03µm, p=0.301). Among pink-anodized groups, air polishing resulted in significantly lower surface roughness (0.33±0.08µm) compared to titanium scaling (0.51±0.11µm, p<0.001) and no instrument treatment (0.38±0.07µm, p=0.050). Anodization significantly increased wettability (p<0.001), while instrumentation with a titanium scaling decreased it (p<0.001). The combination of un-anodized samples and titanium scaling treatment showed the lowest wettability with the highest contact angle (70.72±2.63°). The biofilm viability, measured by CFU/mL, was significantly inhibited by anodization (p<0.001) and air polishing (p<0.001) while promoted by titanium scaling (p<0.001). Gold-anodized titanium discs subjected to air polishing exhibited the lowest CFU/mL (279,420±16,300), while un-anodized samples instrumented with a titanium scaler had the highest CFU/mL (945,580±13,580). Biofilm mass, quantified by optical density values, was significantly inhibited by anodization (p<0.001) as well as air polishing (p=0.001). A moderate negative correlation was observed between CFU and wettability (ρ=-0.55, p<0.001). Gold- and pink-anodized titanium surfaces were more hydrophilic, leading to less biofilm formation than un-anodized ones. Biofilm formation was inhibited by air polishing while promoted by titanium scaling. Gold anodization combined with air polishing had the least biofilm formation and can be considered the preferred abutment anodization/instrumentation combination.

Defending Ti6Al4V against Biofilm Formation with Albumin Biofunctionalization.

Surface biofunctionalization with structurally perturbed albumin, as well as with other plasmatic proteins, inhibits the initial bacterial adhesion and biofilm formation, involved in numerous healthcare-associated infections. In fact, we have reported this protective effect with thermally treated plasmatic proteins, such as albumin and fibrinogen, adsorbed on flat silica surfaces. Here, we show that albumin biofunctionalization also works properly on flat Ti6Al4V substrates, which are widely used to fabricate medical devices. The protective effect is conserved even in biologically relevant fluids, containing other proteins that potentially adsorb onto and/or displace preadsorbed albumin from the biofunctionalized substrates. We further demonstrate that the presence of structurally perturbed albumin on the substrate does not trigger macrophage activation and the release of inflammatory mediators. Consequently, surface biofunctionalization with thermally perturbed albumin is a simple strategy to prepare antibacterial, nonimmunogenic medical devices.

Autoregulation of the Master Regulator Spo0A Controls Cell-Fate Decisions in Bacillus subtilis.

Spo0A in Bacillus subtilis is activated by phosphorylation (Spo0A~P) upon starvation and differentially controls a set of genes involved in biofilm formation and sporulation. The spo0A gene is transcribed by two distinct promoters, a σA-recognized upstream promoter Pv during growth, and a σH-recognized downstream promoter Ps during starvation, and appears to be autoregulated by four Spo0A~P binding sites (0A1-4 boxes) localized between two promoters. However, the autoregulatory mechanisms and their impact on differentiation remain elusive. Here, we determined the relative affinity of Spo0A~P for each 0A box and dissected each promoter in combination with the systematic 0A box mutations. The data revealed that (1) the Pv and Ps promoters are on and off, respectively, under nutrient-rich conditions without Spo0A~P, (2) the Ps promoter is activated by first 0A3 and then 0A1 during early starvation with low Spo0A~P, (3) during later starvation with high Spo0A~P, the Pv promoter is repressed by first 0A1 and then 0A2 and 0A4, and (4) during prolonged starvation, both promoters are silenced by all 0A boxes with very high Spo0A~P. Our results indicate that the autoregulation of spo0A is one of the key determinants to achieve a developmental increase in Spo0A~P, leading to a temporal window for entry into biofilm formation or sporulation.

Quorum sensing regulates virulence factors in the coral pathogen Vibrio coralliilyticus.

The bacterial pathogen Vibrio coralliilyticus causes disease in coral species worldwide. The mechanisms of V. coralliilyticus coral colonization, coral microbiome interactions, and virulence factor production are understudied. In other model Vibrio species, virulence factors like biofilm formation, toxin secretion, and protease production are controlled through a density-dependent communication system called quorum sensing (QS). Comparative genomics indicated that V. coralliilyticus genomes share high sequence identity for most of the QS signaling and regulatory components identified in other Vibrio species. Here, we identify an active QS signaling pathway in two V. coralliilyticus strains with distinct infection etiologies: type strain BAA-450 and coral isolate OCN008. In V. coralliilyticus, the inter-species AI-2 autoinducer signaling pathway in both strains controls expression of the master QS transcription factor and LuxR/HapR homolog VcpR to regulate >300 genes, including protease production, biofilm formation, and two conserved type VI secretion systems (T6SSs). Activation of T6SS1 by QS results in the secretion of effectors and enables interbacterial competition and killing of prey bacteria. We conclude that the QS system in V. coralliilyticus is functional and controls the expression of genes involved in relevant bacterial behaviors typically associated with host infection.IMPORTANCEVibrio coralliilyticus infects many marine organisms, including multiple species of corals, and is a primary causative agent of tissue loss diseases and bacterial-induced bleaching. Here, we investigated a common cell-cell communication mechanism called quorum sensing, which is known to be intimately connected to virulence in other Vibrio species. Our genetic and chemical studies of V. coralliilyticus quorum sensing uncovered an active pathway that directly regulates the following key virulence factors: proteases, biofilms, and secretion systems. These findings connect bacterial signaling in communities to the infection of corals, which may lead to novel treatments and earlier diagnoses of coral diseases in reefs.

Evaluation of the antibacterial properties of four bioactive biomaterials for chronic wound management.

Chronic wound infections present a prevalent medical issue and a multifaceted problem that significantly impacts healthcare systems worldwide. Biofilms formed by pathogenic bacteria are fundamental virulence factors implicated in the complexity and persistence of bacterial-associated wound infections, leading to prolonged recovery times and increased risk of infection. This study aims to investigate the antibacterial effectiveness of commonly employed bioactive wound healing compositions with a particular emphasis on their effectiveness against common bacterial pathogens encountered in chronic wounds - Staphylococcus epidermidis, Escherichia coli, and Pseudomonas aeruginosa to identify optimal wound product composition for managing chronic wound infections. This study tested the antibacterial and antibiofilm effectiveness of four bioactive wound healing materials by performing in vitro antibacterial assays and measuring ion release profiles. The anti-biofilm effectiveness differed extensively among the biomaterials tested and slightly among the bacterial species. Particularly, copper and zinc-doped borate bioactive glass wound healing compositions inhibited the three clinically relevant bacteria in both planktonic and biofilm forms, which were found to be ascribed to the copper and zinc gradual release. The findings suggest that copper and zinc-doped bioactive glasses hold great promise for improving chronic wound management by providing strong antibacterial action and promoting faster healing.

Benefits of Immobilized Bacteria in Bioremediation of Sites Contaminated with Toxic Organic Compounds

Although bioremediation is considered the most environmentally friendly and sustainable technique for remediating contaminated soil and water, it is most effective when combined with physicochemical methods, which allow for the preliminary removal of large quantities of pollutants. This allows microorganisms to efficiently eliminate the remaining contaminants. In addition to requiring the necessary genes and degradation pathways for specific substrates, as well as tolerance to adverse environmental conditions, microorganisms may perform below expectations. One typical reason for this is the high toxicity of xenobiotics present in large concentrations, stemming from the vulnerability of bacteria introduced to a contaminated site. This is especially true for planktonic bacteria, whereas bacteria within biofilms or microcolonies have significant advantages over their planktonic counterparts. A physical matrix is essential for the formation, maintenance, and survival of bacterial biofilms. By providing such a matrix for bacterial immobilization, the formation of biofilms can be facilitated and accelerated. Therefore, bioremediation combined with bacterial immobilization offers a comprehensive solution for environmental cleanup by harnessing the specialized metabolic activities of microorganisms while ensuring their retention and efficacy at target sites. In many cases, such bioremediation can also eliminate the need for physicochemical methods that are otherwise required to initially reduce contaminant concentrations. Then, it will be possible to use microorganisms for the remediation of higher concentrations of xenobiotics, significantly reducing costs while maintaining a rapid rate of remediation processes. This review explores the benefits of bacterial immobilization, highlighting materials and processes for developing an optimal immobilization matrix. It focuses on the following four key areas: (i) the types of organic pollutants impacting environmental and human health, (ii) the bacterial strains used in bioremediation processes, (iii) the types and benefits of immobilization, and (iv) the immobilization of bacterial cells on various carriers for targeted pollutant degradation.

Calcium Phosphate (CaP) Composite Nanostructures on Polycaprolactone (PCL): Synergistic Effects on Antibacterial Activity and Osteoblast Behavior

Bone tissue engineering aims to develop biomaterials that are capable of effectively repairing and regenerating damaged bone tissue. Among the various polymers used in this field, polycaprolactone (PCL) is one of the most widely utilized. As a biocompatible polymer, PCL is easy to fabricate, cost-effective, and offers consistent quality control, making it a popular choice for biomedical applications. However, PCL lacks inherent antibacterial properties, making it susceptible to bacterial adhesion and biofilm formation, which can lead to implant failure. To address this issue, this study aims to enhance the antibacterial properties of PCL by incorporating calcium phosphate composite (PCL_CaP) nanostructures onto its surface via hydrothermal synthesis. The resulting “PCL_CaP” nanostructured surfaces exhibited improved wettability and demonstrated mechano-bactericidal potential against Escherichia coli and Bacillus subtilis. The flake-like morphology of the fabricated CaP nanostructures effectively disrupted bacteria membranes, inhibiting bacterial growth. Furthermore, the “PCL_CaP” surfaces supported the adhesion, proliferation, and differentiation of pre-osteoblasts, indicating their potential for bone tissue engineering applications. This study demonstrates the promise of calcium phosphate composite nanostructures as an effective antibacterial coating for implants and medical devices, with further research required to evaluate their long-term stability and in vivo performance.

Redox-Modulating Capacity and Effect of Ethyl Acetate Roots and Aerial Parts Extracts from Geum urbanum L. on the Phenotype Inhibition of the Pseudomonas aeruginosa Las/RhI Quorum Sensing System

Pseudomonas aeruginosa is an opportunistic pathogen that causes nosocomial infections of the urinary tract, upper respiratory tract, gastrointestinal tract, central nervous system, etc. It is possible to develop bacteremia and sepsis in immunocompromised patients. A major problem in treatment is the development of antibiotic resistance. Therefore, new preparations of natural origin are sought, such as plant extracts, which are phytocomplexes and to which it is practically impossible to develop resistance. Geum urbanum L. (Rosacea) is a perennial herb known for many biological properties. This study aimed to investigate the redox-modulating capacity and effect of ethyl acetate (EtOAc) extracts from roots (EtOAcR) and aerial parts (EtOAcAP) of the Bulgarian plant on the phenotype inhibition of the P. aeruginosa Las/RhI quorum sensing (QS) system, which primarily determines drug resistance in pathogenic bacteria, including biofilm formation, motility, and pigment production. We performed QS assays to account for the effects of the two EtOAc extracts. At sub-minimal inhibitory concentrations (sub-MICs) ranging from 1.56 to 6.25 mg/mL, the biofilm formation was inhibited 85% and 84% by EtOAcR and 62% and 39% by EtOAcAP extracts, respectively. At the same sub-MICs, the pyocyanin synthesis was inhibited by 17–27% after treatment with EtOAcAP and 26–30% with EtOAcR extracts. The motility was fully inhibited at 3.12 mg/mL and 6.25 mg/mL (sub-MICs). We investigated the inhibitory potential of lasI, lasR, rhiI, and rhiR gene expression in biofilm and pyocyanin probes with the PCR method. Interestingly, the genes were inhibited by two extracts at 3.12 mg/mL and 6.25 mg/mL. Antiradical studies, assessed by DPPH, CUPRAC, and ABTS radical scavenging methods and superoxide anion inhibition showed that EtOAcAP extract has effective antioxidant capacity. These results could help in the development of new phytocomplexes that could be applied as biocontrol agents to inhibit the phenotype of the P. aeruginosa QS system and other antibiotic-resistant pathogens.

Acylase-Based Coatings on Sandblasted Polydimethylsiloxane-Based Materials for Antimicrobial Applications

Indwelling medical devices, such as urinary catheters, often experience bacterial colonization, forming biofilms that resist antibiotics and the host’s immune defenses through quorum sensing (QS), a chemical communication system. This study explores the development of antimicrobial coatings by immobilizing acylase, a quorum-quenching enzyme, on sandblasted polydimethylsiloxane (PDMS) surfaces. PDMS, commonly used in medical devices, was sandblasted to increase its surface roughness, enhancing acylase attachment. FTIR analysis confirmed that acylase retained its three-dimensional structure upon immobilization, preserving its enzymatic activity. The antibacterial efficacy of the coatings was tested against Pseudomonas aeruginosa (P. aeruginosa) (a common biofilm-forming pathogen), Staphylococcus aureus (S. aureus), and Escherichia coli (E. coli). The results showed that sandblasted PDMS surfaces had improved bacterial adhesion due to increased focal adhesion points, but acylase-functionalized surfaces had significantly reduced bacterial attachment and biofilm formation. Notably, the coatings inhibited P. aeruginosa growth by 40% under static conditions, demonstrating the potential of acylase-functionalized PDMS for medical applications. This approach offers a promising strategy for creating antimicrobial surfaces that prevent biofilm-related infections in urinary catheters and other medical devices. The findings highlight the dual role of surface roughness in enhancing enzyme attachment while reducing bacterial adhesion through effective QS inhibition.

Limosilactobacillus reuteri AJCR4: A Potential Probiotic in the Fight Against Oral Candida spp. Biofilms

Oral candidiasis is one of the most common infections in the immunocompromised. Biofilms of Candida species can make treatments difficult, leading to oral infection recurrence. This research aimed to isolate a Lactobacillus with anti-Candida effects from the oral cavity. An oral Lactobacillus was isolated in caries-free individuals. The best isolate was evaluated against Candida spp. planktonic and biofilm forms. The bacterial impacts on Candida biofilms’ adhesion to acrylic discs were analyzed through an in vitro test. L. reuteri AJCR4 had the best anti-Candida activity in the preliminary screening. Results were promising in both planktonic and biofilms, particularly with C. albicans SC5314 and C. tropicalis ATCC750, where no viable cells were detected when using the cell-free supernatant (undiluted). In C. glabrata ATCC2001 and C. parapsilosis ATCC22019 biofilms, reductions of 3 Log10 and more than 2 Log10, respectively, were noted when using a cell suspension of L. reuteri ACJR4 (108 CFU/mL). On polymethyl methacrylate acrylic discs, the cell-free supernatant reduced Candida adhesion, resulting in no viable cell detection on the surface. In conclusion, L. reuteri AJCR4 demonstrated notable antifungal activity against Candida biofilms. This oral isolate and its postbiotic can be a potential alternative strategy to oral candidiasis, especially to treat recalcitrant infections.

Virulent and multidrug-resistant Aeromonas in aquatic environments of Kerala, India: potential risks to fish and humans.

Aeromonas inhabit diverse aquatic habitats and are recognized as both opportunistic and primary pathogens of fish and humans. This study delineates the biochemical and gyrB sequence-based molecular identification of 14 Aeromonas strains isolated from aquatic environments in Kerala, India, identifying them as A. dhakensis (50%), A. hydrophila (28.6%), and A. jandaei (21.4%). These strains exhibit a high prevalence of virulence genes (act, flaA, ser, gcat, lip, and ela) implicated in pathogenesis in both fish and humans. These findings underline the emergence of A. dhakensis, often misidentified as A. hydrophila, as a potential pathogen, highlighting the necessity for comprehensive identification methods. Significantly, all strains demonstrated beta-hemolysis and moderate to strong biofilm formation, enhancing their infectivity potential. Moreover, all isolates exhibited multidrug resistance, with a multiple antimicrobial resistance (MAR) index ranging from 0.39 to 0.56, and a significant presence of class 1 (500-1100bp) and class 2 (250-700bp) integrons, indicating their potential risk to both fish and human populations. Our results underscore the role of aquatic environment as a repository for virulent and multidrug-resistant Aeromonas spp., emphasizing the imperative for prudent antimicrobial usage and regular monitoring of antimicrobial resistance (AMR) in these environments.

Characterization and Probiotic Potential of Levilactobacillus brevis DPL5: A Novel Strain Isolated from Human Breast Milk with Antimicrobial Properties Against Biofilm-Forming Staphylococcus aureus

Lactobacillus is a key genus of probiotics commonly utilized for the treatment of oral infections The primary aim of our research was to investigate the probiotic potential of the newly isolated Levilactobacillus brevis DPL5 strain from human breast milk, focusing on its ability to combat biofilm-forming pathogens such as Staphylococcus aureus. Employing in vitro approaches, we demonstrate L. brevis DPL5′s ability to endure at pH 3 with survival rates above 30%, and withstand the osmotic stress often found during industrial processes like fermentation and freeze drying, retaining over 90% viability. The lyophilized cell-free supernatant of L. brevis DPL5 had a significant antagonistic effect against biofilm-producing nasal strains of Staphylococcus aureus, and it completely eradicated biofilms at subinhibitory concentrations of 20 mg·mL−1. Higher concentrations of 69 mg·mL−1 were found to have a 99% bactericidal effect, based on the conducted probability analysis, indicating the production of bactericidal bioactive extracellular compounds capable of disrupting the biofilm formation of pathogens like S. aureus. Furthermore, genome-wide sequencing and analysis of L. brevis DPL5 with cutting-edge Nanopore technology has uncovered over 50 genes linked to probiotic activity, supporting its ability to adapt and thrive in the harsh gut environment. The genome also contains multiple biosynthetic gene clusters such as lanthipeptide class IV, Type III polyketide synthase (T3PKS), and ribosomally synthesized, and post-translationally modified peptides (RiPP-like compounds), all of which are associated with antibacterial properties. Our study paves the way for the further exploration of DPL5, setting the stage for innovative, nature-inspired solutions to combat stubborn bacterial infections.

A strategy for controlling Hypervirulent Klebsiella pneumoniae: inhibition of ClpV expression

The emergence and prevalence of hypervirulent Klebsiella pneumoniae (hvKP) have proposed a great challenge to control this infection. Therefore, exploring some new drugs or strategies for treating hvKP infection is an urgent issue for scientific researchers. In the present study, the clpV gene deletion strain of hvKP (ΔclpV-hvKP) was constructed using CRISPR-Cas9 technology, and the biological characteristics of ΔclpV-hvKP were investigated to explore the new targets for controlling this pathogen. The results showed that clpV gene deletion did not affect the growth ability of hvKP. However, knocking out the clpV gene markedly decreased the mucoid phenotype and the biofilm formation ability of hvKP. It reduced the interspecific competition of hvKP with Escherichia coli, Salmonella, Pseudomonas aeruginosa, and Staphylococcus aureus. The clpV deletion significantly changed the transcriptome profile of hvKP, inhibited the expression of virulence factors, and decreased the lethality of hvKP against Galleria mellonella larvae. In vitro experiments showed that lithocholic acid could inhibit the expression of the clpV gene and reduce the virulence of hvKP. Our data suggested that the clpV gene may be a potential target for decreasing hvKP infection risk.

New easy lab methods for the extraction of phycobiliproteins and pigments from cyanobacteria

Abstract Cyanobacteria are a source of a variety of valuable substances. These include extracellular polymeric substances (EPS) and natural dyes. EPS are produced by cyanobacteria to protect themselves against environmental stresses, serve as a water reservoir, and play an essential role in the formation of biofilms. They can be used, for example, due to their antimicrobial properties. Cyanobacteria are phototrophic organisms and synthesize the pigments chlorophyll a and carotenoids for photosynthesis. However, they also possess other light-harvesting complexes in the form of phycobilisomes, which combine with proteins to form so-called phycobiliproteins (PBP). These enable cyanobacteria to utilize a larger light spectrum than plants. Both the pigments and the PBP can be used as natural colorants and offer antioxidant properties, among other things, which enables applications in medicine. Methods for the combined extraction of EPS, PBP, and pigments have already been described in the literature. Still, in this work, the methods were simplified and optimized to obtain the best possible method for use in the laboratory. The optimized downstream process is based on an extraction of the EPS with deionized water followed by an extraction of the PBP with potassium phosphate buffer and finally the extraction of the pigments with methanol. The PBP and pigments are extracted by adding fixed ratios of solvents followed by a single extraction for a certain period. The universal applicability was demonstrated over a broad biomass spectrum from 4 mg to 80 mg dry mass and the method was transferred to several cyanobacterial strains.

Potential Use of Selected Natural Compounds with Anti-Biofilm Activity

Antibiotic resistance in microorganisms is an escalating global concern, exacerbated by their formation of biofilms, which provide protection through an extracellular matrix and communication via quorum sensing, enhancing their resistance to treatment. This situation has driven the search for alternative approaches, particularly those using natural compounds. This study explores the potential of phytochemicals, such as quercetin, apigenin, arbutin, gallic acid, proanthocyanidins, and rutin, known for their antibacterial properties and ability to inhibit biofilm formation and disrupt mature biofilms. The methods used in this study included a comprehensive review of current literature assessing the bioavailability, distribution, and effective concentrations of these compounds in treating biofilm-associated infections. The results indicate that these phytochemicals exhibit significant antibacterial effects, reduce biofilm’s structural integrity, and inhibit bacterial communication pathways. Moreover, their potential use in combination with existing antibiotics may enhance therapeutic outcomes. The findings support the conclusion that phytochemicals offer promising additions to anti-biofilm strategies and are capable of complementing or replacing conventional treatments, with appropriate therapeutic levels and delivery mechanisms being key to their effectiveness. This insight underscores the need for further research into their clinical applications for treating infections complicated by biofilms.

Characterization and enhanced antibiofilm activity of Annona muricata extract in combination with fluconazole against Candida albicans

Introduction: Candida albicans biofilm formation contributes significantly to antifungal resistance, necessitating new treatment strategies. Annona muricata Lin., a traditional herbal remedy, has shown promise in combating microbial infections. This study evaluated the antibiofilm activity of the methanol extract of A. muricata leaves, alone or combined with fluconazole, against C. albicans. Methods: Phytochemicals in the methanol extract were analyzed using LC-MS, biofilm metabolic activity was assessed with the XTT assay, and morphological changes were examined via scanning electron microscopy (SEM). Molecular docking evaluated the binding of compounds in the A. muricata extract to the Sap3 receptor (PDB: 2H6T). Results: LC-MS identified 17 phytochemicals in the methanol extract. The extract showed dose-dependent inhibition of biofilm formation, reaching ~60% inhibition at 240 μg/ml. Fluconazole inhibition increased from 32% to 76% as concentrations rose from 15 to 240 μg/ml. The combination of A. muricata and fluconazole enhanced inhibition, ranging from 74% to 78% at 15–240 μg/ml. SEM analysis revealed altered biofilm morphology and loss of cell integrity with the combination treatment. Phytochemicals showed high binding affinity (−9.7 to −8.0 kcal/mol) for the Sap3 enzyme, suggesting therapeutic potential. Conclusion: These findings suggest that compounds in the A. muricata methanol extract synergize with fluconazole at sub-inhibitory concentrations to suppress C. albicans biofilm formation. This paves the way for antifungal treatments that may mitigate fluconazole resistance using this plant extract.

Antimicrobial Efficacy of Trifluoro-Anilines Against Vibrio Species

Vibrios are naturally present in marine ecosystems and are commonly allied with live seafood. Vibrio species frequently cause foodborne infections, with Vibrio parahaemolyticus recently becoming a significant contributor to foodborne illness outbreaks. In response, aniline and 68 of its aniline derivatives were studied due to their antibacterial effects targeting V. parahaemolyticus and Vibrio harveyi. Among these, 4-amino-3-chloro-5-nitrobenzotrifluoride (ACNBF) and 2-iodo-4-trifluoromethylaniline (ITFMA) demonstrated both antibacterial and antibiofilm properties. The minimum inhibitory concentrations (MIC) for ACNBF and ITFMA were 100 µg/mL and 50 µg/mL, respectively, against planktonic cells. The active compounds effectively suppressed biofilm formation in a manner dependent on the dosage. Additionally, these trifluoro-anilines significantly reduced virulence factors such as motility, protease activity, hemolysis, and indole production. Both trifluoro-anilines caused noticeable destruction to the membrane of bacterial cells and, at 100 µg/mL, exhibited bactericidal activity against V. parahaemolyticus within 30 min. Toxicity assays using the Caenorhabditis elegans and seed germination models showed that the compounds displayed mild toxicity. As a result, ACNBF and ITFMA inhibited the growth of both planktonic cells and biofilm formation. Furthermore, these active compounds effectively prevented the formation of biofilm on the surfaces of shrimp and squid models, highlighting their potential use in controlling seafood contamination.

Hypermutability bypasses genetic constraints in SCV phenotypic switching in Pseudomonas aeruginosa biofilms

Biofilms are critical in the persistence of Pseudomonas aeruginosa infections, particularly in cystic fibrosis patients. This study explores the adaptive mechanisms behind the phenotypic switching between Small Colony Variants (SCVs) and revertant states in P. aeruginosa biofilms, emphasizing hypermutability due to Mismatch Repair System (MRS) deficiencies. Through experimental evolution and whole-genome sequencing, we show that both wild-type and mutator strains undergo parallel evolution by accumulating compensatory mutations in factors regulating intracellular c-di-GMP levels, particularly in the Wsp and Yfi systems. While wild-type strains face genetic constraints, mutator strains bypass these by accessing alternative genetic pathways regulating c-di-GMP and biofilm formation. This increased genetic accessibility, driven by higher mutation rates and specific mutational biases, supports sustained cycles of SCV conversion and reversion. Our findings underscore the crucial role of hypermutability in P. aeruginosa adaptation, with significant implications for managing persistent infections in clinical settings.

Two-component system UhpAB facilitates the pathogenicity of avian pathogenic Escherichia coli through biofilm formation and stress responses.

UhpAB increases the pathogenicity of APEC.UhpAB activates the expression of virulence genes fepG, ldrD, ycgV, and ydeI.UhpAB promotes biofilm formation and enhances stress tolerance.UhpAB contributes to APEC evading attack by the host immune system.

Characterization and genomic insights into bacteriophages Kpph1 and Kpph9 against hypervirulent carbapenem-resistant Klebsiella pneumoniae

ABSTRACT The increasing incidence of infections attributed to hypervirulent carbapenem-resistant Klebsiella pneumoniae (Hv-CRKp) is of considerable concern. Bacteriophages, also known as phages, are viruses that specifically infect bacteria; thus, phage-based therapies offer promising alternatives to antibiotic treatments targeting Hv-CRKp infections. In this study, two isolated bacteriophages, Kpph1 and Kpph9, were characterized for their specificity against the Hv-CRKp K. pneumoniae NUHL30457 strain that possesses a K2 capsule serotype. Both phages exhibit remarkable environmental tolerance, displaying stability over a range of pH values (4–11) and temperatures (up to 50°C). The phages demonstrate potent antibacterial and antibiofilm efficacy, as indicated by their capacity to inhibit biofilm formation and to disrupt established biofilms of Hv-CRKp. Through phylogenetic analysis, it has been revealed that Kpph1 belongs to the new species of Webervirus genus, and Kpph9 to the Drulisvirus genus. Comparative genomic analysis suggests that the tail fiber protein region exhibits the greatest diversity in the genomes of phages within the same genus, which implies distinct co-evolution histories between phages and their corresponding hosts. Interestingly, both phages have been found to contain two tail fiber proteins that may exhibit potential depolymerase activities. However, the exact role of depolymerase in the interaction between phages and their hosts warrants further investigation. In summary, our findings emphasize the therapeutic promise of phages Kpph1 and Kpph9, as well as their encoded proteins, in the context of research on phage therapy targeting hypervirulent carbapenem-resistant Klebsiella pneumoniae.