Anti-biofilm Mechanism Research Articles

Deciphering the killing mechanisms of potassium iodide in combination with antimicrobial photodynamic therapy against cross-kingdom biofilm

IntroductionThe co-existence of S. mutans and C. albicans is frequently detected in root caries and early child caries and is reported to be associated with recurrent caries. The aim of this study was to investigate the effects of potassium iodide (KI) in combination with toluidine blue O-mediated antimicrobial photodynamic therapy (aPDT) on S. mutans and C. albicans mixed-species biofilm, as well as the antibiofilm mechanisms involved.MethodsMixed-species biofilm was constructed of S. mutans and C. albicans on dentin blocks. The antibiofilm efficacy, cytotoxicity and antibiofilm mechanism of KI in combination with aPDT were determined and evaluated.ResultsKI+TBO-aPDT treatment caused reduction in microorganism counts, metabolic activity, and biofilm biomass of mixed-species biofilm without inducing cytotoxicity to hDPCs (human dental pulp cells). Observations such increased ROS (reactive oxygen species) levels, impaired cell membrane function, cell apoptosis and reduced expression in several genes seem to be artifacts of reduced growth and general killing by KI+TBO-aPDT treatment.DiscussionThese data suggested that KI in combination with aPDT as an innovative approach to combat S. mutans and C. albicans biofilm, and thus as an optional treatment for caries.

Antibiofilm mechanism of 2R,3R-dihydromyricetin by targeting sortase A and its application against Staphylococcus aureus adhesion on eggshell

Biofilm formation of Staphylococcus aureus in food processing environments raises significant safety concerns, necessitating the development of new antibiofilm approaches for controlling S. aureus contamination. This study aimed to elucidate the antibiofilm mechanism of 2R,3R-dihydromyricetin (DMY), a natural flavonoid, against S. aureus and evaluate its efficacy in reducing bacterial adhesion to eggshell. The results revealed that DMY was a potent inhibitor of S. aureus sortase A (SrtA) with an IC50 of 73.43 μM, preventing bacterial adhesion to fibrinogen and subsequent biofilm formation. Fluorescence quenching assay and surface plasmon resonance analysis confirmed that DMY could directly bind to S. aureus SrtA. Notably, circular dichroism spectra demonstrated a conformational change in SrtA from α-helical to β-sheet structure upon DMY binding. Molecular dynamics simulation suggested that DMY bound to the catalytic pocket of S. aureus SrtA via hydrophobic interactions and hydrogen bonds. Furthermore, fluorescence microscopic observations further revealed that DMY attenuated the biofilm-related phenotype of SrtA by decreasing the anchoring of S. aureus protein A (SpA) onto cell wall. Importantly, pretreatment with 125 μg/mL DMY significantly reduced 1.14–1.75 log CFU/cm2 of S. aureus adhered on eggshells. Overall, these findings highlight how specific targeting of SrtA by DMY inhibits the attachment stages of biofilm development in S. aureus, making it a promising candidate for a novel disinfectant against this pathogen in the food industry.

Inhibitory Mechanism of the Oregano Essential Oil on the Growth and Biofilm Formation of Streptococcus mutans

Objective: The objective was to determine the in vitro antibacterial effect of the Oregano essential oil (OEO) on Streptococcus mutans ( S mutans) and to explore the anti-biofilm effect and mechanism of OEO. Methods: The in vitro antibacterial susceptibility of OEO was detected by the growth curve and colony-forming unit (CFU) counting method. Confocal laser scanning microscope (CLSM) and crystal violet assay were used to explore the anti-biofilm activity of OEO, and the anti-biofilm mechanism was explored from the aspects of extracellular polysaccharides (EPS) synthesis, acidoid, and extracellular DNA changes. OEO was characterized using a gas chromatography-mass spectrometer and cytotoxicity was tested using a cell counting Kit-8 assay. Results: The minimum inhibitory concentration of OEO was 0.25 mg/mL, and the CFU showed the ability of OEO to kill planktonic bacteria. Scanning electron microscopy and CLSM showed that OEO could hinder the development of biofilms, harm bacteria, and eliminate mature biofilms by changing EPS synthesis and pH value. Carvacrol in OEO played a major role in biological antibacterial activity. OEO had low cytotoxicity and good biocompatibility. Conclusions: OEO had excellent antibacterial and anti-biofilm effects on the key virulence traits of S mutans, indicating potential prospects in preventing dental caries.

Anti-Biofilm Effects of Z102-E of Lactiplantibacillus plantarum against Listeria monocytogenes and the Mechanism Revealed by Transcriptomic Analysis.

Lactic acid bacteria (LAB) are the most common probiotics, and they present excellent inhibitory effects on pathogenic bacteria. This study aimed to explore the anti-biofilm potential of the purified active substance of Lactiplantibacillus plantarum, named Z102-E. The effects of Z102-E on Listeria monocytogenes were investigated in detail, and a transcriptomic analysis was conducted to reveal the anti-biofilm mechanism. The results indicated that the sub-MIC of Z102-E (3.2, 1.6, and 0.8 mg/mL) decreased the bacterial growth and effectively reduced the self-aggregation, surface hydrophobicity, sugar utilization, motility, biofilm formation, AI-2 signal molecule, contents of extracellular polysaccharides, and extracellular protein of L. monocytogenes. Moreover, the inverted fluorescence microscopy observation confirmed the anti-biofilm effect of Z102-E. The transcriptomic analysis indicated that 117 genes were up-regulated and 214 were down-regulated. Z102-E regulated the expressions of genes related to L. monocytogenes quorum sensing, biofilm formation, etc. These findings suggested that Z102-E has great application potential as a natural bacteriostatic agent.

Food byproduct-derived zinc-binding peptides: Dual suppression of planktonic and biofilm Staphylococcus aureus

With the increasing globalization of the food supply, foodborne infection and food poisoning brought by Staphylococcus aureus (S. aureus) have progressively become intractable issues. This work aimed to research the dual inhibitory effects of zinc-binding peptides (termed CSSP-Zn) on both planktonic and biofilm of S. aureus 6538, with respect to its utilization on pork models. Further research showed CSSP-Zn could disrupt the cytomembrane of permeability and integrity, triggering cytoplasm overflow, thereby causing proton motive force (PMF) collapse and metabolic disorder in planktonic strain 6538. The antibiofilm mechanism of CSSP-Zn primarily involved disrupting the expression of polysaccharide intercellular adhesion (PIA) biosynthesis-related genes (e.g. icaA, icaB, icaC and icaD), causing an enervated intercellular adhesion and lessened biofilm viability. When employed to a pork model, CSSP-Zn could obviously attenuate strain 6538 colonization and enterotoxin secretion. These results not only demonstrated the practical antibacterial efficacy of CSSP-Zn but also offered a novel strategy for mitigating the risk of S. aureus contamination in pork.

New insights into the antibiofilm activity and mechanism of Mannosylerythritol Lipid-A against Listeria monocytogenes EGD-e

Listeria monocytogenes is one of the leading causative agents of foodborne disease outbreaks worldwide. Herein, the antibiofilm effect and mechanism of Mannosylerythritol Lipid-A against L. monocytogenes EGD-e is reported for the first time. MEL-A effectively attenuated biofilm formation while reducing the viability and motility of bacteria within the biofilm in the early stage, and influenced bacterial adhesion by affecting the secretion of extracellular polysaccharides and eDNA. RT-qPCR revealed that MEL-A significantly suppressed the expression of genes involved in flagellar movement and virulence. Untargeted LC-MS metabolomics indicated that MEL-A affected the fluidity and permeability of cell membranes by significantly upregulating unsaturated fatty acids, lipids and glycoside metabolites, and affected protein biosynthesis, nucleotide metabolism and DNA synthesis and repair by significantly downregulating amino acid metabolism and nucleic acid metabolism. These pathways may constitute the key targets of biofilm formation inhibition by MEL-A. Furthermore, MEL-A showed good removal effects on mature biofilms under different temperatures, different materials and milk. Our data indicated that MEL-A could be used as a novel antibiofilm agent to improve food safety. Our study provides new insights into the possible inhibitory mechanism of MEL-A and the response of L. monocytogenes EGD-e to MEL-A.

Structural insights on anti-biofilm mechanism of heated slightly acidic electrolyzed water technology against multi-resistant Staphylococcus aureus biofilm on food contact surface

Slightly acidic electrolyzed water (SAEW) has proven to be an efficient and novel sanitizer in food and agriculture field. This study assessed the efficacy of SAEW (30 mg/L) at 40 °C on the inactivation of foodborne pathogens and detachment of multi-resistant Staphylococcus aureus (MRSA) biofilm. Furthermore, the underlying mechanism of MRSA biofilm under heated SAEW at 40 °C treatment on metabolic profiles was investigated. The results showed that the heated SAEW at 40 °C significantly effectively against foodborne pathogens of 1.96–7.56 (lg (CFU/g)) reduction in pork, chicken, spinach, and lettuce. The heated SAEW at 40 °C treatment significantly reduced MRSA biofilm cells by 2.41 (lg (CFU/cm2)). The synergistic effect of SAEW treatment showed intense anti-biofilm activity in decreasing cell density and impairing biofilm cell membranes. Global metabolic response of MRSA biofilms, treated by SAEW at 40 °C, revealed the alterations of intracellular metabolites, including amino acids, organic acid, fatty acid, and lipid. Moreover, signaling pathways involved in amino acid metabolism, energy metabolism, nucleotide synthesis, carbohydrate metabolites, and lipid biosynthesis were functionally disrupted by the SAEW at 40 °C treatment. As per our knowledge, this is the first research to uncover the potential mechanism of heated SAEW treatment against MRSA biofilm on food contact surface.

The Anti-Microbial Peptide Citrocin Controls Pseudomonas aeruginosa Biofilms by Breaking Down Extracellular Polysaccharide.

Citrocin is an anti-microbial peptide that holds great potential in animal feed. This study evaluates the anti-microbial and anti-biofilm properties of Citrocin and explores the mechanism of action of Citrocin on the biofilm of P. aeruginosa. The results showed that Citrocin had a significant inhibitory effect on the growth of P. aeruginosa with a minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of 0.3 mg/mL. All five concentrations (1/4MIC, 1/2MIC, MIC, 2MIC, and 4MIC) of Citrocin inhibited P. aeruginosa biofilm formation. Citrocin at the MIC, 2MIC and 4MIC removed 42.7%, 76.0% and 83.2% of mature biofilms, respectively, and suppressed the swarming motility, biofilm metabolic activity and extracellular polysaccharide production of P. aeruginosa. Metabolomics analysis indicated that 0.3 mg/mL of Citrocin up- regulated 26 and down-regulated 83 metabolites, mainly comprising amino acids, fatty acids, organic acids and sugars. Glucose and amino acid metabolic pathways, including starch and sucrose metabolism as well as arginine and proline metabolism, were highly enriched by Citrocin. In summary, our research reveals the anti-biofilm mechanism of Citrocin at the metabolic level, which provides theoretical support for the development of novel anti-biofilm strategies for combatting P. aeruginosa.

Ultrasonic-assisted extraction, anti-biofilm activity, and mechanism of action of Ku Shen (Sophorae Flavescentis Radix) extracts against Vibrio parahaemolyticus.

The objective of this study is to optimize the ultrasonic-assisted extraction process of Ku Shen (Sophorae Flavescentis Radix) extracts (KSE) against Vibrio parahaemolyticus and explore their anti-biofilm activity and mechanism of action. The ultrasonic-assisted extraction process of KSE optimized by single factor experiment, Box-Behnken design and response surface methodology was as follows: 93% ethanol as solvent, liquid/material ratio of 30 mL/g, ultrasonic power of 500 W, extraction temperature of 80°C and time of 30 min. Under these conditions, the diameter of inhibition circle of KSE was 15.60 ± 0.17 mm, which had no significant difference with the predicted value. The yield of dried KSE is 32.32 ± 0.57% and the content of total flavonoids in KSE was 57.02 ± 5.54%. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of KSE against V. parahaemolyticus were 0.25 and 0.5 mg/mL, respectively. Crystal violet staining, Congo red plate, spectrophotometry, CCK-8 and scanning electron microscopy were used to investigate the activity and mechanism of action of KSE against V. parahaemolyticus biofilm. The results showed that the sub-MIC of KSE could significantly inhibit biofilm formation, reduce the synthesis of polysaccharide intercellular adhesin (PIA) and the secretion of extracellular DNA. In addition, the inhibition rate of biofilm formation and clearance rate of mature biofilm of 1.0 mg/mL KSE were 85.32 and 74.04%, and the reduction rate of metabolic activity of developing and mature biofilm were 77.98 and 74.46%, respectively. These results were confirmed by visual images obtained by scanning electron microscopy. Therefore, KSE has the potential to further isolate the anti-biofilm agent and evaluate it for the preservation process of aquatic products.

Anti-Biofilm Activity of Assamsaponin A, Theasaponin E1, and Theasaponin E2 against Candida albicans.

Biofilm formation plays a crucial role in the pathogenesis of Candida albicans and is significantly associated with resistance to antifungal agents. Tea seed saponins, a class of non-ionic triterpenes, have been proven to have fungicidal effects on planktonic C. albicans. However, their anti-biofilm activity and mechanism of action against C. albicans remain unclear. In this study, the effects of three Camellia sinensis seed saponin monomers, namely, theasaponin E1 (TE1), theasaponin E2 (TE2), and assamsaponin A (ASA), on the metabolism, biofilm development, and expression of the virulence genes of C. albicans were evaluated. The results of the XTT reduction assay and crystal violet (CV) staining assay demonstrated that tea seed saponin monomers concentration-dependently suppressed the adhesion and biofilm formation of C. albicans and were able to eradicate mature biofilms. The compounds were in the following order in terms of their inhibitory effects: ASA > TE1 > TE2. The mechanisms were associated with reductions in multiple crucial virulence factors, including cell surface hydrophobicity (CSH), adhesion ability, hyphal morphology conversion, and phospholipase activity. It was further demonstrated through qRT-PCR analysis that the anti-biofilm activity of ASA and TE1 against C. albicans was attributed to the inhibition of RAS1 activation, which consequently suppressed the cAMP-PKA and MAPK signaling pathways. Conversely, TE2 appeared to regulate the morphological turnover and hyphal growth of C. albicans via a pathway that was independent of RAS1. These findings suggest that tea seed saponin monomers are promising innovative agents against C. albicans.

Structural dynamics and anti-biofilm screening of novel imidazole derivative to explore their anti-biofilm inhibition mechanism against Pseudomonas Aeruginosa

The biofilm formation is still prevalent mechanism of developing the drug resistance in the Pseudomonas aeruginosa, gram-negative bacteria, known for its major role in nosocomial, ventilator-associated pneumonia (VAP), lung infections and catheter-associated urinary tract infections. As best of our knowledge, current study first time reports the most potent inhibitors of LasR, a transcriptional activator of biofilm and virulence regulating genes in, Pseudomonas aeruginosa LasR, utilizing newly functionalized imidazoles (5a-d), synthesized via 1,3-dipolar cycloaddition using click approach. The synthesized ligands were characterized through Mass Spectrometry and 1H NMR. The binding potency and mode of biding of ligands. Quantum Mechanical(QM) methods were utilized to investigate the electronic basis, HOMO/LUMO and dipole moment of the geometry of the ligands for their binding potency. Dynamics cross correlation matrix (DCCMs) and protein surface analysis were further utilized to explore the structural dynamics of the protein. Free energy of binding of ligands and protein were further estimated using Molecular Mechanical Energies with the Poisson–Boltzmann surface area (MMPBSA) method. Molecular Docking studies revealed significant negative binding energies (5a − 10.33, 5b −10.09, 5c − 10.11, and 5d −8.33 KJ/mol). HOMO/LUMO and potential energy surface map estimation showed the ligands(5a) with lower energy gaps and larger dipole moments had relatively larger binding potency. The significant change in the structural dynamics of LasR protein due to complex formation with newlyfunctionalized imidazoles ligands. Hydrogen bond surface analysis followed by MMPBSA calculations of free energy of binding further complemented the Molecular docking revelations showing the specifically ligand (5a) having the relatively higher energy of binding(-65.22kj/mol). Communicated by Ramaswamy H. Sarma

Antibiofilm activity of polyethylene glycol-quercetin nanoparticles-loaded gelatin-N,O-carboxymethyl chitosan composite nanogels against Staphylococcus epidermidis.

Biofilms, such as those from Staphylococcus epidermidis, are generally insensitive to traditional antimicrobial agents, making it difficult to inhibit their formation. Although quercetin has excellent antibiofilm effects, its clinical applications are limited by the lack of sustained and targeted release at the site of S. epidermidis infection. Polyethylene glycol-quercetin nanoparticles (PQ-NPs)-loaded gelatin-N,O-carboxymethyl chitosan (N,O-CMCS) composite nanogels were prepared and assessed for the on-demand release potential for reducing S. epidermidis biofilm formation. The formation mechanism, physicochemical characterization, and antibiofilm activity of PQ-nanogels against S. epidermidis were studied. Physicochemical characterization confirmed that PQ-nanogels had been prepared by the electrostatic interactions between gelatin and N,O-CMCS with sodium tripolyphosphate. The PQ-nanogels exhibited obvious pH and gelatinase-responsive to achieve on-demand release in the micro-environment (pH 5.5 and gelatinase) of S. epidermidis. In addition, PQ-nanogels had excellent antibiofilm activity, and the potential antibiofilm mechanism may enhance its antibiofilm activity by reducing its relative biofilm formation, surface hydrophobicity, exopolysaccharides production, and eDNA production. This study will guide the development of the dual responsiveness (pH and gelatinase) of nanogels to achieve on-demand release for reducing S. epidermidis biofilm formation.

Anti-biofilm mechanism of a synthetical low molecular weight poly-d-mannose on Salmonella Typhimurium

In this study, a low molecular weight poly-d-mannose (LMWM) was separated from a mixed polysaccharide synthesized previously. Monosaccharide composition, Fourier-Transform infrared spectroscopy (FT-IR), periodate oxidation and smith degradation were determined. After safety evaluation, the inhibition of LMWM on the different biofilm formation stages of Salmonella enterica serovar Typhimurium (S. Typhimurium) was tested in vitro. Furthermore, the effect of LMWM on the adhesion of S. Typhimurium to Caco-2 cells and cell surface hydrophobicity (CSH) were observed. Results indicated that LMWM was a homopolysaccharide without cytotoxicity and hemolysis, containing both α-mannose and β-mannose. It showed obvious anti-biofilm activity on S. Typhimurium and mainly activated on the initial adhesion and formation stage, even better than the commercial S. cerevisiae mannan (CM). LMWM inhibited the adhesion of S. Typhimurium on Caco-2 cells with the inhibition rate of 61.04 % at 2 mg/ml. Meanwhile, LMWM decreased the hydrophobicity of S. Typhimurium cell surface. In conclusion, the inhibitory effect on S. Typhimurium biofilm was not caused by bacteriostatic or bactericidal activity of LMWM. The specific anti-adhesion and the decrease of bacterial CSH by LMWM may closely relate to anti-biofilm mechanism. This study provides some supports for the application of LMWM as antibiotics alternative on S. Typhimurium in the future.

Antibiofilm Activity of Probiotic Strains against Aggregatibacter actinomycetemcomitans

Aggregatibacter actinomycetemcomitans is a significant agent for periodontal disease. It develops biofilms on gingival plaque and promotes chronic pro-inflammatory response to periodontal tissues. As bacterial cells within the biofilm, this pathogen successfully eluded the human immune system by preventing the uptake of biofilm cells by phagocytic cells. Antibiotic-resistant biofilms might harm the environment, increase mortality, and add to the cost of healthcare. Anti-biofilms are necessary to prevent the growth of biofilms. It has been demonstrated that probiotics change how immune system cells produce cytokines. Therefore, the efficacy of five probiotic lactobacillus strains to prevent A. actinomycetemcomitans from growing bacteria and forming biofilms was evaluated. The organisms were incubated in anaerobic conditions at 37°C for a certain period. The antimicrobial test was done using the lawn method and measuring the size of the inhibition zone produced by the probiotic spots for each probiotic strain. Tetracycline antibiotic disc was used as a positive control to compare the size of the inhibition zone. The antibiofilm activity was performed using probiotic strains at their exponential and stationary phases against the exponential phase of A. actinomycetemcomitans. Our finding showed the probiotic lactobacillus could inhibit the growth of A. actinomycetemcomitans. The highest inhibition was demonstrated by L. johnsonii NBRC 13952. Besides, all probiotic lactobacillus used for this study significantly (p<0.05) inhibited the A. actinomycetemcomitans biofilm using both stationary and exponential phases. In conclusion, probiotic lactobacillus might be a potential agent that can be used to counteract the periodontal pathogen biofilm. Further study should be done to elucidate the antibiofilm mechanism, and several trials of the probiotic lactobacilli against multiple species of biofilms might be helpful in evaluating their effectiveness.

In silico design and mechanistic study of niosome-encapsulated curcumin against multidrug-resistant Staphylococcus aureus biofilms

Curcumin, an important natural component of turmeric, has been known for a long time for its antimicrobial properties. This study aimed to investigate the anti-biofilm action of the niosome-encapsulated curcumin and explore the involved anti-biofilm mechanism. In silico investigations of ADME-Tox (absorption, distribution, metabolism, excretion, and toxicity) were first performed to predict the suitability of curcumin for pharmaceutical application. Curcumin showed low toxicity but at the same time, low solubility and low stability, which, in turn, might reduce its antimicrobial activity. To overcome these intrinsic limitations, curcumin was encapsulated using a biocompatible niosome system, and an encapsulation efficiency of 97% was achieved. The synthesized curcumin-containing niosomes had a spherical morphology with an average diameter of 178 nm. The niosomal curcumin was capable of reducing multi-drug resistant (MDR) Staphylococcus aureus biofilm 2–4-fold compared with the free curcumin. The encapsulated curcumin also demonstrated no significant cytotoxicity on the human foreskin fibroblasts. To understand the interaction between curcumin and S. aureus biofilm, several biofilm-related genes were analyzed for their expression. N-acetylglucosaminyl transferase (IcaD), a protein involved in the production of polysaccharide intercellular adhesion and known to play a function in biofilm development, was found to be downregulated by niosomal curcumin and showed high binding affinity (-8.3 kcal/mol) with curcumin based on molecular docking analysis. Our study suggests that the niosome-encapsulated curcumin is a promising approach for the treatment of MDR S. aureus biofilm and can be extended to biofilms caused by other pathogens.

A nanocomposite hydrogel for co-delivery of multiple anti-biofilm therapeutics to enhance the treatment of bacterial biofilm-related infections

The characteristics of biofilms have exacerbated the issue of clinical antibiotic resistance, rendering it a pressing challenge in need of resolution. The combination of biofilm-dispersing agents and antibiotics can eliminate biofilms and promote healing synergistically in infected wounds. In this study, we developed a novel nanocomposite hydrogel (NC gel) comprised of the poly(lactic acid)-hyperbranched polyglycerol (PLA-HPG) based bioadhesive nanoparticles (BNPs) and a hydrophilic carboxymethyl chitosan (CS) network. The NC gel was designed to co-deliver two biofilm-dispersing agents (an NO-donor SNO, and an α-amylase Am) and an antibiotic, cefepime (Cef), utilizing a synergistic anti-biofilm mechanism in which Am loosens the matrix structure and NO promotes the release of biofilm bacteria via quorum sensing, and Cef kills bacteria. The drug-loaded NC gel (SNO/BNP/CS@Am-Cef) demonstrated sustained drug release, minimal cytotoxicity, and increased drug-bacterial interactions at the site of infection. When applied to mice infected with methicillin-resistant Staphylococcus aureus (MRSA) biofilms in vivo, SNO/BNP/CS@Am-Cef enhanced biofilm elimination and promoted wound healing compared to traditional antibiotic treatments. Our work demonstrates the feasibility of the co-delivery of biofilm-dispersing agents and antibiotics using the NC gel and presents a promising approach for the polytherapy of bacterial biofilm-related infections.

Mode of action of antimicrobial agents albofungins in eradicating penicillin- and cephalosporin-resistant Vibrio parahaemolyticus biofilm.

Albofungin is a promising broad-spectrum antimicrobial compound against multidrug-resistant bacteria. In the present study, we further investigated albofungin's biofilm eradication activity and its potential mode of action against drug-resistant Vibrio parahaemolyticus. Among all derivatives, albofungin exhibited the best antibiofilm and antibacterial activity with rapid killing effects at 0.12 µg mL-1. Confocal microscopy observation exhibited that albofungin disrupted V. parahaemolyticus biofilms by killing or dispersing biofilm cells. Meanwhile, scanning electron microscope and fluorescent staining experiments demonstrated that albofungin rapidly destroyed the integrity and permeability of the bacterial cell membrane. Moreover, this study revealed an antibiofilm mechanism of albofungin involving inhibition of peptidoglycan biosynthesis, flagella assembly pathways, and secretion system proteins in V. parahaemolyticus by quantitative proteomics and validation experiments. Our results highlighted albofungin's mechanism of action in planktonic cells and biofilms and suggested further development and potential applications of albofungin for treating infections caused by penicillins-and-cephalosporins-resistant V. parahaemolyticus. IMPORTANCE Infections caused by multidrug-resistant bacteria, as well as a scarcity of new antibiotics, have become a major health threat worldwide. To tackle the demand for new and effective treatments, we investigated the mechanism of action of albofungin, a natural product derived from Streptomyces, which exhibits potent antimicrobial activity against multidrug-resistant bacteria. Albofungin showed potent biofilm eradication activity against penicillins-and-cephalosporins-resistant Vibrio parahaemolyticus, which expresses a novel metallo-β-lactamase and, thus, reduces their sensitivity to various antibiotics. We observed membrane disruption and permeation mechanisms in planktonic cells and biofilms after albofungin treatment, while albofungin had a weak interaction with bacterial DNA. Moreover, the antibiofilm mechanism of albofungin included inhibition of peptidoglycan biosynthesis, flagellar assembly pathways, and secretion system proteins. Our finding suggested potential applications of albofungin as an antibacterial and antibiofilm therapeutic agent.

Antibiofilm activity of bromelain from pineapple against Staphylococcus aureus

Bromelain is a set of proteolytic enzymes usually obtained from pineapple (Ananas comosus). Although bromelain has distinguished therapeutic properties, little is known about its proteolytic potential against opportunistic pathogens related to wound healing complications, such as Staphylococcus aureus. This study aimed to investigate the antibiofilm and antibacterial activity of bromelain in 43 clinical strains of S. aureus isolated from chronic wounds and blood cultures. Bromelain’s activity against S. aureus biofilm in vitro was assessed by analyzing biofilm formation in cultures grown in the presence of 1% bromelain and biofilm destruction after the addition of 1% bromelain to mature biofilms. Proteinase K and sodium metaperiodate were also added to mature biofilms in parallel to compare their activity with that of bromelain and, together with exopolysaccharide and protein production rate assays, to determine the chemical composition of the biofilm extracellular matrix of selected strains of S. aureus. Bromelain was also evaluated for its DNase activity and impact on cellular hydrophobicity and auto-aggregation. Mueller-Hinton agar dilution was used to determine bromelain minimal inhibitory concentration (MIC). Biofilm assays showed that 1% bromelain significantly inhibits S. aureus biofilm formation (p = 0.0157) by up to 4-fold and destroys its mature biofilms (p < 0.0001) by up to 6.4-fold, both compared to the control grown without bromelain. Biofilms of methicillin-resistant S. aureus strains isolated from chronic wounds were the most affected by bromelain treatment. No antibacterial activity was detected with bromelain MIC assays and the proteolytic activity of bromelain was identified as the main antibiofilm mechanism of the enzyme, though its DNase activity may also contribute. The epithelial therapeutic properties of bromelain combined with its antibiofilm activity against S. aureus make it a promising alternative to compose the therapeutic arsenal for the control of S. aureus biofilms in the context of wound care.

Antibiofilm mechanism of peppermint essential oil to avert biofilm developed by foodborne and food spoilage pathogens on food contact surfaces.

Establishing efficient methods to combat bacterial biofilms is a major concern. Natural compounds, such as essential oils derived from plants, are among the favored and recommended strategies for combatting bacteria and their biofilm. Therefore, we evaluated the antibiofilm properties of peppermint oil as well as the activities by which it kills bacteria generally and particularly their biofilms. Peppermint oil antagonistic activities were investigated against Vibrio parahaemolyticus, Listeria monocytogenes, Pseudomonas aeruginosa, Escherichia coli O157:H7, and Salmonella Typhimurium on four food contact surfaces (stainless steel, rubber, high-density polyethylene, and polyethylene terephthalate). Biofilm formation on each studied surface, hydrophobicity, autoaggregation, metabolic activity, and adenosine triphosphate quantification were evaluated for each bacterium in the presence and absence (control) of peppermint oil. Real-time polymerase chain reaction, confocal laser scanning microscopy, and field-emission scanning electron microscopy were utilized to analyze the effects of peppermint oil treatment on the bacteria and their biofilm. Results showed that peppermint oil (1/2× minimum inhibitory concentration [MIC], MIC, and 2× MIC) substantially lessened biofilm formation, with high bactericidal properties. A minimum of 2.5-log to a maximum of around 5-log reduction was attained, with the highest sensitivity shown by V. parahaemolyticus. Morphological experiments revealed degradation of the biofilm structure, followed by some dead cells with broken membranes. Thus, this study established the possibility of using peppermint oil to combat key foodborne and food spoilage pathogens in the food processing environment.

Antibiofilm Activity and Mechanism of Linalool against Food Spoilage Bacillus amyloliquefaciens.

Pellicle biofilm-forming bacteria Bacillus amyloliquefaciens are the major spoilage microorganisms of soy products. Due to their inherent resistance to antibiotics and disinfectants, pellicle biofilms formed are difficult to eliminate and represent a threat to food safety. Here, we assessed linalool's ability to prevent the pellicle of two spoilage B. amyloliquefaciens strains. The minimum biofilm inhibitory concentration (MBIC) of linalool against B. amyloliquefaciens DY1a and DY1b was 4 μL/mL and 8 μL/mL, respectively. The MBIC of linalool had a considerable eradication rate of 77.15% and 83.21% on the biofilm of the two strains, respectively. Scanning electron microscopy observations revealed that less wrinkly and thinner pellicle biofilms formed on a medium supplemented with 1/2 MBIC and 1/4 MBIC linalool. Also, linalool inhibited cell motility and the production of extracellular polysaccharides and proteins of the biofilm matrix. Furthermore, linalool exposure reduced the cell surface hydrophobicity, zeta potential, and cell auto-aggregation of B. amyloliquefaciens. Molecular docking analysis demonstrated that linalool interacted strongly with quorum-sensing ComP receptor and biofilm matrix assembly TasA through intermolecular hydrogen bonds, hydrophobic contacts, and van der Waals forces interacting with site residues. Overall, our findings suggest that linalool may be employed as a potential antibiofilm agent to control food spoilage B. amyloliquefaciens.