Search Results (158)

Dental caries is a major global health issue associated with biofilm formation by Streptococcus mutans (S. mutans). Conventional antimicrobials often fail to eliminate biofilms due to their structural resistance, highlighting the need for new strategies. This study investigated the antibacterial and antibiofilm effects of protamine peptides (PPs), which are cell-penetrating antimicrobial peptides derived from salmon protamine, alone and in combination with antimicrobial agents. Antimicrobial susceptibility was evaluated using alamarBlue^® and colony count assays, while biofilm formation was analyzed using crystal violet staining, confocal microscopy, and extracellular polysaccharide (EPS) quantification. PP exhibited moderate antibacterial activity but strongly suppressed EPS accumulation and biofilm development, leading to a flattened biofilm structure. Cotreatment with ε-poly-L-lysine (PL) significantly enhanced antibacterial and antibiofilm effects compared with either agent alone, whereas this effect was not observed with other cationic polymers. Fluorescence imaging revealed that PL promoted the intracellular localization of PP without increasing membrane damage, indicating a cooperative mechanism by which PL enhances membrane permeability and PP targets intracellular sites. These findings demonstrate that combining a cell-penetrating peptide with a membrane-active agent is a novel approach to overcome bacterial tolerance. The PP–PL combination effectively suppressed S. mutans growth and biofilm formation through dual action on membranes and EPS metabolism, offering a promising basis for the development of peptide-based preventive agents and biofilm-resistant dental materials. Full article

Biofilm formation by non-C. albicans Candida (NAC) species is a major factor in device-associated infections, yet few studies have examined their development under physiologically relevant conditions. This study evaluated the biofilm-forming capacity of Candida tropicalis, Candida parapsilosis sensu stricto and Candida albicans on stainless steel surfaces in the presence of artificial saliva, simulating the respiratory tract environment of tracheostomized patients. Standardized inocula were incubated for 24 h, and biofilms were assessed through quantification of viable cells, biomass, biofilm matrix production and structural characterization by scanning electron microscopy (SEM). C. tropicalis produced the most robust biofilms compared to C. albicans and C. parapsilosis stricto sensu isolates, with significantly higher biomass and biofilm matrix (p < 0.001). C. parapsilosis sensu stricto developed less dense yet structurally defined biofilm networks. SEM confirmed mature and compact biofilm architecture, especially in C. tropicalis. These results demonstrate the strong intrinsic biofilm-forming ability of NAC species on stainless steel under host-like conditions, reinforcing their capacity to persist on medical surfaces and their relevance as independent contributors to biofilm-related contamination and infection. Full article

Adherent-invasive E. coli (AIEC) is closely related to inflammatory bowel disease (IBD). However, its pathogenic mechanism has not yet been fully elucidated. Using a BLASTP search, we discovered that the amino acid sequence of a putative protein (UFP37798.1) in the AIEC LF82 strain is highly homologous to some regulators in the SlyA family. We named it EruA. We displayed the secondary structures of EruA using bioinformatics, overexpressed the His₆-tagged EruA protein using SDS-PAGE, and dissected the genetic organization of the eruA chromosomal region using 5′RACE. We constructed an eruA deletion mutant (ΔeruA) and a complementary strain (CΔeruA) of the LF82 strain. The transcriptomes of wild-type (WT) and ΔeruA bacteria were compared using RNA sequencing and qRT-PCR, thereby identifying 32 differentially expressed genes (DEGs). Based on YASARA software and EMSA analysis, EruA directly binds to the consensus sequences (P_fimA and P_tnaB) in the promoter region of the fimA and tnaB genes from these DEGs. By using a super-resolution confocal microscope (SCM), counting CFUs of colonies on plates, indole quantification, and crystal violet staining of biofilms adhered to tubes or 96-well plates, we found that EruA activates the fimA to promote bacterial adhesion to intestinal epithelial cells and activates the tnaB to enhance bacterial indole production and biofilm formation. Moreover, EruA helps AIEC resist environmental stress and enhances bacterial survival within macrophages as well as loading in mouse tissues. Notably, EruA promotes AIEC colonization in the colons of mice and exacerbates intestinal inflammation caused by bacterial infection in mice with DSS-induced inflammatory colitis, manifested by weight loss, colon length shortening, and pathological changes in colon tissues. Therefore, EruA plays a key role in the pathogenicity of AIEC. Full article

Background/Objectives: Bacterial biofilms formed by Escherichia coli pose a significant challenge in veterinary medicine due to their intrinsic resistance to antibiotics. Antimicrobial peptides (AMPs) represent a promising alternative. AMPs exert their bactericidal activity by binding to negatively charged phospholipids in bacterial membranes via electrostatic interactions, leading to membrane disruption and rapid cell lysis. Methods: In vitro assays including MIC determination, biofilm eradication testing (crystal violet, colony counts, and CLSM), swimming motility, and EPS quantification were performed. CRISPR/Cas9 was used to construct and complement a kduD mutant. A transposon mutagenesis library was screened for biofilm-defective mutants. In an in vivo murine excisional wound infection model treated with the mouse cathelicidin-related antimicrobial peptide (CRAMP-34), wound closure and bacterial burden were monitored. Gene expression changes were analyzed via RT-qPCR. Results: CRAMP-34 effectively eradicated pre-formed biofilms of a clinically relevant, porcine-origin E. coli strain and promoted wound healing in the murine infection model. We conducted a genome-wide transposon mutagenesis screen, which identified kduD as a critical gene for robust biofilm formation. Functional characterization revealed that kduD deletion drastically impairs flagellar motility and alters exopolysaccharide production, leading to defective biofilm architecture without affecting growth. Notably, the anti-biofilm activity of CRAMP-34 phenocopied aspects of the kduD deletion, including motility inhibition and transcriptional repression of a common set of biofilm-related genes. Conclusions: This research highlights CRAMP-34 as a potent anti-biofilm agent and unveils kduD as a previously unrecognized regulator of E. coli biofilm development, which is also targeted by CRAMP-34. Full article

Background: Tracheostomy procedures are associated with increased risk of nosocomial infections due to microbial colonization and biofilm formation on tube surfaces. These biofilms contribute to persistent infections and hinder clinical recovery. Plant-derived products have gained interest as alternative strategies for preventing device-associated infections. Methods: This study evaluated the phytochemical composition and the antimicrobial, anti-adherent, antibiofilm, anti-inflammatory, antioxidant, and toxicity properties of Tapirira obtusa bark extract (TOBE). Antimicrobial activity was determined by minimum inhibitory concentrations (MICs). Biofilm formation and microbial viability were assessed in mono- and mixed-species biofilms. Anti-inflammatory effects were evaluated by NF-κB inhibition and TNF-α quantification. Antioxidant activity was measured using the DPPH assay. Phytochemical analysis identified major bioactive groups, and toxicity was tested in the Galleria mellonella model. Results: TOBE exhibited notable antimicrobial activity, with MIC values between 3.9 and 31.25 µg/mL. At 78 µg/mL, the extract significantly reduced biofilm biomass and microbial viability (p < 0.05). TOBE also downregulated NF-κB activation and decreased TNF-α levels. Antioxidant assays confirmed radical-scavenging capacity. Phytochemical screening revealed phenolics, flavonoids, and tannins, and toxicity results indicated a safe profile. Conclusion: TOBE effectively inhibits microbial growth and biofilm development on orotracheal tube surfaces while exhibiting anti-inflammatory and antioxidant properties without detectable toxicity. These findings support its potential as a plant-based adjunct for preventing tracheostomy-related infections and improving patient outcomes. Full article

Background: Bacterial keratitis, a major cause of corneal blindness, is frequently associated with biofilm-forming pathogens such as Klebsiella pneumoniae. Cyclic-di-GMP (c-di-GMP) controls biofilm development, which increases antibiotic resistance and makes treatment more difficult, highlighting the need for innovative therapeutic approaches. Methods: This study investigated cinnamaldehyde as a potential ocular therapeutic using combined computational and experimental approaches. Molecular docking and in vitro assays (XTT, resazurin reduction, crystal violet staining, qRT-PCR, and fluorescence microscopy) were used to evaluate the anti-biofilm and immunomodulatory activities of cinnamaldehyde (CA) against Klebsiella pneumoniae. Results: CA inhibited biofilm formation in a dose-dependent manner (≈89% at 1000 µM; >50% at 250 µM), reduced bacterial attachment to contact lenses, and downregulated key biofilm genes (mrkA, mrkC, ybtS, bolA). Docking analysis revealed strong binding affinity to the mrkH regulator (−5.46 kcal/mol. CA maintained more than 80% corneal cell viability by increasing IL-10, suppressing inflammatory mediators (IL-1β, IL-6, and TNF-α), and improving bacterial clearance. Conclusions: This study combines computational docking, biofilm quantification, immune cell assays, and functional gene expression analyses to reveal the ability of cinnamaldehyde not only to suppress biofilm formation but also to enhance macrophage-mediated clearance and modulate corneal immune responses, a multi-target approach not previously described in the context of bacterial keratitis. Such effects highlight its potential as a novel ocular drug candidate for protecting corneal integrity in infectious keratitis. Full article

The leading cause of post-surgical hospital readmission is the emergence of hospital-acquired infections (HAIs), where surgical site infections (SSIs) constitute a substantial negative impact on patient outcome and contribute annual direct costs estimated to range from $28.4 billion to $45 billion in the U.S. To address the need for novel antimicrobial coating strategies, previous research has demonstrated that certain microbes can degrade poly(aspartic acid) (PAA)-based coatings, suggesting potential limitations of single-compound approaches that must be considered when designing antimicrobial surfaces. In this proof-of-concept study, we investigated whether ordered sequential coatings combining thermally synthesized PAA (tPAA) and oleic acid (OleA) might produce enhanced antimicrobial effects compared to individual compounds. Despite concerns regarding PAA biodegradability, the benefits of using PAA include low cytotoxicity and an ability to chelate metals such as calcium and facilitate bone mineralization and growth post-surgery. Using simple yet effective methods of surface coating applications which utilize tPAA and OleA, we investigated the potential of these ordered coatings to attenuate planktonic and sessile (biofilm) growth and development in Pseudomonas aeruginosa and Escherichia coli in vitro. Application of these ordered coatings resulted in up to 62% reduction in bacterial carrying capacity for P. aeruginosa and up to 43% reduction in biofilm mass relative to untreated controls. Further, confocal imaging via immunohistochemical labeling revealed methods for evaluating the impact of treatments targeting biofilm development through extracellular DNA quantification. Additionally, these coatings show dose-dependent cytotoxic effects against 3T3 mouse fibroblast cells. These preliminary findings, along with results derived from cytotoxicity assessment and physicochemical characterization via dynamic light scattering, suggest that ordered tPAA-OleA coating systems warrant further investigation as potential antimicrobial strategies, though additional validation, including testing against diverse clinical isolates, mechanistic studies, and in vivo evaluation, would be required before clinical application. Full article

Biodegradable magnesium (Mg) alloys are promising materials for temporary orthopaedic implants, combining favourable mechanical properties and superplastic behaviour with in vivo resorption. This enables (i) prolonged implant duration, (ii) fabrication of complex-shaped prostheses via superplastic forming (SPF), (iii) elimination of removal surgery, and (iv) reduced risk of long-term complications. However, rapid corrosion under physiological conditions remains a major limitation, highlighting the need for surface treatments that slow degradation while preserving implant integrity. This study investigates the effects of hydrothermal surface treatment on MgAZ31-SPF alloys, focusing on immunomodulatory responses, antibacterial potential, and degradation behaviour. Hydrothermally treated MgAZ31-SPF (MgAZ31-SPF-HT) extracts released lower Mg²⁺ concentrations (29.2 mg/dL) compared to untreated MgAZ31-SPF (47.5 mg/dL) while maintaining slightly alkaline pH (7–8.7), indicating improved control of early degradation. In vitro assays with human peripheral blood mononuclear cells (hPBMCs) and normal human dermal cells (NHDCs) showed that MgAZ31-SPF-HT extracts maintained higher cell viability over 24–72 h. Gene expression analysis revealed significant downregulation of pro-inflammatory markers CTSE and TNF-α, while protein quantification via ELISA and BioPlex confirmed reduced secretion of TNF-α, TGF-β1, TGF-β2, IL-6, and IL-8, suggesting mitigation of early immune activation. Antibacterial assays demonstrated limited Staphylococcus aureus colonisation on both MgAZ31-SPF and MgAZ31-SPF-HT scaffolds, with CFU counts (~10⁵–10⁶) well below the threshold for mature biofilm formation (~10⁸), and SEM analysis confirmed sparse bacterial distribution without dense EPS-rich layers. Overall, hydrothermal treatment improves Mg alloy biocompatibility by controlling Mg²⁺ release, modulating early immune responses, and limiting bacterial adhesion, highlighting its potential to enhance clinical performance of Mg-based implants. Full article

This study aimed to explore the phytochemical variability, nutrient composition, and bioactive profile of the edible mushroom species Craterellus cornucopioides with specimens originating from Romanian flora. Its nutritional profile, including its proximate composition and energy value, was determined using standardized analytical methods. The mean contents of total polyphenols and caffeic acid derivatives, quantified by spectrophotometric assay, were established. HPLC–DAD–ESI⁺ analysis enabled the identification and quantification of individual phenolic constituents. Its antioxidant potential was systematically assessed using the following in vitro complementary assays: FRAP, ABTS, CUPRAC, DPPH, and ORAC. Antimicrobial activity was evaluated in vitro against MSSA, MRSA, Bacillus cereus, Enterococcus faecalis, Listeria monocytogenes, Escherichia coli, Pseudomonas aeruginosa, and Candida albicans using well diffusion, broth microdilution, and anti-biofilm assays. The high amounts of polyphenols, including gentisic acid and protocatechuic acid, underlined a biologically relevant phytochemical composition. In fact, all tested extracts and, in particular, CE3 extract consistently displayed strong antioxidant properties, as indicated by five complementary in vitro tests (FRAP, ABTS, CUPRAC, DPPH, and ORAC). In addition, CE1-4 extracts expressed in vitro antimicrobial potential towards all tested organisms except for Pseudomonas aeruginosa. Our results underscore C. cornucopioides’ nutritional, antioxidant, and antimicrobial potential, thus supporting its classification as an edible but under-explored mushroom species with promising applications in both the food and pharmaceutical industries. Full article

Background: Carbapenem-resistant Acinetobacter baumannii (CRAB) is a critical public health threat, particularly in hospital environments where treatment options are limited. Drug repurposing offers a rapid and cost-effective strategy to address antimicrobial resistance. This study evaluates KP46 (tris(8-quinolinolato)gallium(III)), an orally bioavailable gallium-based anticancer agent, for its antimicrobial potential against CRAB. Methods: KP46 was synthesized and characterized using spectroscopic and crystallographic techniques. Its antibacterial activity was assessed against planktonic and biofilm-associated CRAB strains, including multidrug-resistant clinical isolates. Mechanistic studies included intracellular reactive oxygen species (ROS) quantification, membrane integrity assays, and resistance development profiling. Results: KP46 exhibited potent antibacterial activity against both susceptible and carbapenem-resistant A. baumannii strains. It inhibited planktonic growth and disrupted early biofilm formation. KP46 induced intracellular oxidative stress, leading to membrane damage and cell death. Resistance development was significantly slower compared to meropenem, and KP46 retained efficacy against meropenem-resistant isolates. Conclusions: KP46 demonstrates dual-action antimicrobial activity and a low propensity for resistance development, positioning it as a promising candidate for repurposing against CRAB infections. These findings support further preclinical evaluation of KP46 as an orally active therapeutic agent targeting both planktonic and biofilm-associated bacterial populations. resistance development and retained efficacy against meropenem-resistant strains. Full article

Biomolecules of microbial origin are gaining attention for their use in various industries, including cosmetics, due to their broad bioactivities, peculiar properties, and sustainability. This study aimed to develop a novel, eco-friendly nanoemulsion from fungal chitosan hydrochloride (ChC), Pseudomonas aeruginosa biosurfactant (PaB), and grape seed oil (GSO), and to assess its antimicrobial action, biofilm control, and biocompatibility. High-energy emulsification was performed to produce the nanoemulsion (CCh-PaB-GSO), which was characterized by FTIR. Its stability was monitored for 30 days via DLS, zeta potential (ZP), and PDI. The minimum inhibitory concentration (MIC) for cariogenic Streptococcus species, inhibitory fraction concentration (FIC), influence on exopolysaccharide (EPS) quantification produced by bacteria, bacteria’s cell wall hydrophobicity, and biofilm control were determined. Biocompatibility was assessed using the HET-CAM technique by determining the irritation potential. FTIR analysis confirmed the formation the interaction between the substances that compound the nanoemulsion. The CCh-PaB-GSO had nanometric micelles (169.5–203.4 nm), PDI (0.241–0.271), and a positive ZP (+20.25 to +31.94 mV). It showed a consistent MIC (2.0 mg/mL CCh, 0.1 mg/mL PaB, and 3.2 mg/mL GSO) for all tested Streptococcus species and an indifferent interaction effect, FIC (1.32). At sub-MIC, the CCh-PaB-GSO effectively reduced EPS and microbial cell wall hydrophobicity, inhibiting biofilm adhesion. The CCh-PaB-GSO demonstrated biocompatibility with no signs of irritation. In conclusion, the ChC-PaB-GSO system forms an effective and stable nanoemulsion with potential for application as an eco-sustainable and biocompatible product for caries control. Full article

This study aimed to characterize Mycoplasmosis bovis strain 16M—a highly virulent isolate from one Chinese outbreak—as a candidate for challenge models and inactivated vaccine development. We assessed strain 16M through morphological observation, PCR identification, drug susceptibility testing, growth titer and biofilm quantification, immunological profiling, and calf challenge experiments. We used genomic resequencing to evaluate the genetic stability across 150 passages. Classified as the prevalent ST52 lineage in China, strain 16M showed phylogenetic proximity to strain 08M and exhibited multidrug resistance (notably to macrolides). It achieved higher titers and stronger biofilm formation than other isolates and the reference strain PG45. In calves, intratracheal inoculation with 16M induced universal infection, severe pulmonary consolidation, and peribronchial cuffing, with significantly higher disease scores (p < 0.01). The inactivated 16M vaccine elicited elevated antigen-specific IgG titers, PBMC proliferation, and IFN-γ production versus PG45. Post challenge, immunized calves showed reduced pathological lesions, shorter bacterial shedding, and lower disease scores than the infected controls (p < 0.05). Genetic stability was confirmed for virulence-associated genes (e.g., adhesion proteins), with stable titers and biofilm production within 50 generations. Strain 16M combines high virulence for challenge modeling and industrial-scale vaccine suitability, owing to its robust growth, stable immunogenicity, and genetic consistency. Full article

Biofilm formation is a key factor in microbial survival and persistence, often contributing to reduced antimicrobial susceptibility. This systematic literature review investigates whether increased biofilm formation correlates with decreased antibiotic susceptibility. The literature search was conducted in the Pubmed database and we identified and screened 328 studies, with 35 ultimately meeting the inclusion criteria for detailed analysis. Findings reveal that the relationship between biofilm size and antimicrobial susceptibility is highly variable and influenced by multiple factors, including microbial species, strain-specific traits, antibiotic type, and experimental methodologies. While some studies report a positive correlation between biofilm biomass and reduced susceptibility, others show weak or no such relationships, and statistical support for a correlation is often lacking (also due to small sample sizes). The lack of standardized biofilm quantification methods and susceptibility metrics further complicates cross-study comparisons. These findings underscore the need for standardized protocols and more comprehensive datasets to clarify the complex interplay between biofilm formation and antibiotic susceptibility. Regardless of these difficulties, the available data clearly indicate that ‘bigger’ biofilms are not by definition less susceptible. Future research should prioritize diverse and sufficiently large strain collections and consistent methodologies to better understand and address biofilm-associated antimicrobial tolerance. Full article

Objectives: This study aimed to evaluate whether the addition of pressurized carbon dioxide (PCD) influences the antimicrobial efficacy of 2.5% sodium hypochlorite (NaOCl) against Enterococcus faecalis biofilm in root canals and dentinal tubules. Methods: Forty extracted human mandibular premolars with single canals were contaminated with E. faecalis for 10 days and randomly assigned to four groups (n = 10): 2.5% NaOCl, 2.5% NaOCl + CO₂, sterile saline, and sterile saline + CO₂. The pH and temperature of the NaOCl solution were measured before and after CO₂ incorporation. Microbial load was assessed by CFU counts before and after irrigation, and in dentin samples from the cervical, middle, and apical thirds. Oxidative stress was evaluated via lipid peroxidation (TBARS), protein carbonyl content, and total protein quantification. Biofilm metabolic activity was analyzed using the XTT reduction assay. Data were analyzed using one-way ANOVA on ranks and two-way repeated measures ANOVA (α = 0.05), a very large effect size (Cohen’s d) ≈ 1.756 was assumed. Results: All irrigation protocols significantly reduced bacterial load (p < 0.05). Both NaOCl groups outperformed the saline controls (p = 0.009). The addition of CO₂ to NaOCl slightly enhanced disinfection in the main canal but did not improve antimicrobial action in dentinal tubules. CO₂ incorporation reduced the pH of NaOCl from ~13.4 to 7.4 and slightly increased the temperature, making the solution more chemically reactive. However, both oxidative stress markers and the XTT assay showed that the combination with CO₂ impaired the antimicrobial effectiveness of NaOCl. Conclusions: Despite the improvement in bacterial reduction in the root canal lumen, the combination of PCD with NaOCl failed to enhance intratubular disinfection and reduced the oxidative damage and metabolic inactivation of the biofilm. CO₂ pressurization appears to limit the antimicrobial action of NaOCl. Full article

In the present study, the biofilm formation and ultraviolet-C (UVC) resistance characteristics of Escherichia coli isolated from an occluded biliary stent were compared with those of four E. coli O157:H7 strains (ATCC 35150, 43889, 43890, and 43895). To evaluate biofilm formation, the E. coli isolated from a stent and four E. coli O157:H7 strains were incubated at 37, 25, and 15 °C for 7 days, revealing that peak biofilm formation occurred at 37 °C (day 1), 25 °C (day 3), and 15 °C (day 5), with the stent-isolated strain consistently exhibiting significantly higher biofilm cell counts than the others (p < 0.05). The UVC treatment was less effective at reducing viable biofilm cells as the formation temperature decreased, with the stent-isolated E. coli biofilm formed at 15 °C showing the lowest reduction levels. Exopolysaccharide quantification revealed that all E. coli strains produced more extracellular polymeric substances (EPSs) at lower temperatures, with the stent-isolated E. coli biofilm formed at 15 °C showing significantly higher EPS levels than the other strains (p < 0.05), potentially explaining its greater UVC resistance. Based on these results, it was confirmed that the biofilm formed by the E. coli isolated from the stent at 15 °C exhibited the highest resistance to UVC, which can be attributed to its elevated exopolysaccharide production. This study demonstrates that both temperature and maturation period significantly influence E. coli biofilm characteristics and provides valuable insights into E. coli isolated from the stent, which may pose a risk of cross-contamination in food-related environments. Full article