Organocatalytic Copolymerization of Cyclic Lysine Derivative and ε-Caprolactam toward Antibacterial Nylon-6 Polymers.

Functional polymers of nylon-6, particularly those with sustained antibacterial functions, have many practical applications. However, the development of functional ε-caprolactam monomers for the subsequent ring-opening copolymerization (ROCOP) formation of these materials remains a challenge. Here we report a t-BuP4-mediated ROCOP of dimethyl-protected cyclic lysine with ε-caprolactam, followed by quaternization, affording antibacterial nylon-6 polymers bearing quaternary ammonium functionality with high molecular weight (up to 77.4 kDa). The antibacterial nylon-6 polymers exhibited good physical and mechanical properties and strong antimicrobial activities. At 25 mol % quaternary ammonium group incorporation, the nylon-6 polymer demonstrated complete killing of Staphylococcus aureus (Gram-positive) and Escherichia coli (Gram-negative). The results from this study may provide a strategy for the facile preparation of antibacterial nylon-6 polymers to addressing the public health and safety challenges.

Multifunctional PVCL nanogels with redox-responsiveness enable enhanced MR imaging and ultrasound-promoted tumor chemotherapy.

Development of versatile nanoplatforms that simultaneously integrate therapeutic and diagnostic features for stimuli-responsive delivery to tumors remains a great challenge. In this work, we report a novel intelligent redox-responsive hybrid nanosystem composed of MnO2 nanoparticles (NPs) and doxorubicin (DOX) co-loaded within poly(N-vinylcaprolactam) nanogels (PVCL NGs) for magnetic resonance (MR) imaging-guided and ultrasound-targeted microbubble destruction (UTMD)-promoted tumor chemotherapy. Methods: PVCL NGs were first synthesized via a precipitation polymerization method, decorated with amines using ethylenediamine, and loaded with MnO2 NPs through oxidation with permanganate and DOX via physical encapsulation and Mn-N coordination bonding. The as-prepared DOX/MnO2@PVCL NGs were well characterized. UTMD-promoted cellular uptake and therapeutic efficacy of the hybrid NGs were assessed in vitro, and a xenografted tumor model was used to test the NGs for MR imaging and UTMD-promoted tumor therapy in vivo.Results: The as-prepared DOX/MnO2@PVCL NGs with a size of 106.8 nm display excellent colloidal stability, favorable biocompatibility, and redox-responsiveness to the reductive intracellular environment and tumor tissues having a relatively high glutathione (GSH) concentration that can trigger the synchronous release of Mn2+ for enhanced T1-weighted MR imaging and DOX for enhanced cancer chemotherapy. Moreover, the DOX/MnO2@PVCL NGs upon the UTMD-promotion exhibit a significantly enhanced tumor growth inhibition effect toward subcutaneous B16 melanoma owing to the UTMD-improved cellular internalization and tumor penetration. Conclusion: Our work thereby proposes a promising theranostic nanoplatform for stimuli-responsive T1-weighted MR imaging-guided tumor chemotherapy.

Nylon-6/chitosan core/shell antimicrobial nanofibers for the prevention of mesh-associated surgical site infection.

The state-of-the-art hernia meshes, used in hospitals for hernia repair, are predominantly polymeric textile-based constructs that present high mechanical strength, but lack antimicrobial properties. Consequently, preventing bacterial colonization of implanted prosthetic meshes is of major clinical relevance for patients undergoing hernia repair. In this study, the co-axial electrospinning technique was investigated for the development of a novel mechanically stable structure incorporating dual drug release antimicrobial action. Core/shell structured nanofibers were developed, consisting of Nylon-6 in the core, to provide the appropriate mechanical stability, and Chitosan/Polyethylene oxide in the shell to provide bacteriostatic action. The core/shell structure consisted of a binary antimicrobial system incorporating 5-chloro-8-quinolinol in the chitosan shell, with the sustained release of Poly(hexanide) from the Nylon-6 core of the fibers. Homogeneous nanofibers with a "beads-in-fiber" architecture were observed by TEM, and validated by FTIR and XPS. The composite nanofibrous meshes significantly advance the stress-strain responses in comparison to the counterpart single-polymer electrospun meshes. The antimicrobial effectiveness was evaluated in vitro against two of the most commonly occurring pathogenic bacteria; S. aureus and P. aeruginosa, in surgical site infections. This study illustrates how the tailoring of core/shell nanofibers can be of interest for the development of active antimicrobial surfaces.

Influence of size, crosslinking degree and surface structure of poly(N-vinylcaprolactam)-based microgels on their penetration into multicellular tumor spheroids.

Current nanomedicine suffers from a big challenge due to the fact that most of the nanocarrier systems lack the desired tumor penetration depth, thereby limiting their clinical translation. Unlike the nanomaterials with a similar size or shape, microgels display excellent softness, fluidity and deformability, as well as stimuli-responsiveness in the tumor microenvironment. Herein, we report the synthesis of temperature-responsive poly(N-vinylcaprolactam)/oligo (ethylene glycol) acrylate/glycidyl methacrylate (PVCL/OEGA/GMA) microgels with different hydrodynamic radii (100-500 nm), crosslinking densities, 2-methoxyethyl acrylate (MEA) contents and OEGA chain lengths using a precipitation polymerization method and the investigation of the microgels in terms of their tumor penetration capability using a multicellular tumor spheroid (MCTS) model. The prepared microgels were well characterized with different techniques. We show that regardless of the size, crosslinking density, MEA content and OEGA chain length, all microgels display the desired cytocompatibility in the given concentration range. In vitro cellular uptake data reveal that similar to 2-dimensional (2-D) adherent cells, microgels with a smaller size display more enhanced cellular uptake than those having a larger size in the 3-D MCTS model. Likewise, 3-D MCTS penetration results indicate that the PVCL/OEGA/GMA microgels with the smallest radius of 100 nm exhibit the deepest penetration length. We then selected the microgels with a radius of 200 nm but with different physicochemical parameters to investigate their cellular uptake and tumor penetration behavior. Our data show that microgels with varying crosslinking densities, MEA contents and OEGA chain lengths do not have any appreciable changes in terms of their cellular uptake and penetration in the 3-D MCTS model. Our study provides new insights for the design of different microgel-based systems for further cancer theranostic applications.

Morphologically bioinspired hierarchical nylon 6,6 electrospun assembly recreating the structure and performance of tendons and ligaments.

Reconstructions of ruptured tendons and ligaments currently have dissatisfactory failure rate. Failures are mainly due to the mechanical mismatch of commercial implants with respect to the host tissue. In fact, it is crucial to replicate the morphology (hierarchical in nature) and mechanical response (highly-nonlinear) of natural tendons and ligaments. The aim of this study was to develop morphologically bioinspired hierarchical Nylon 6,6 electrospun assemblies recreating the structure and performance of tendons and ligaments. First, we built different electrospun bundles to find the optimal orientation of the nanofibers. A 2nd-level hierarchical assembly was fabricated with a dedicated process that allowed tightly joining the bundles one next to the other with an electrospun sheath, so as to improve the mechanical performance. Finally, a further hierarchical 3rd-level assembly was constructed by grouping several 2nd-level assemblies. The morphology of the different structures was assessed with scanning electron microscopy and high-resolution X-ray tomography, which allowed measuring the directionality of the nanofibers in the bundles and in the sheaths. The mechanical properties of the single bundles and of the 2nd-level assemblies were measured with tensile tests. The single bundles and the hierarchical assemblies showed morphology and directionality of the nanofibers similar to the tendons and ligaments. The strength and stiffness were comparable to that of tendons and ligaments. In conclusion, this work showed an innovative electrospinning production process to build nanofibrous Nylon 6,6 hierarchical assemblies which are suitable as future implantable devices and able to mimic the multiscale morphology and the biomechanical properties of tendons and ligaments.

Poly(N-vinylcaprolactam) Nanogels with Antiviral Behavior against HIV-1 Infection.

Stimuli-responsive nanogels offer promising perspectives for the development of next generation formulations for biomedical applications. In this work, poly(N-vinylcaprolactam) nanogels were synthesized varying the concentration of monomer and crosslinking agent. Thus, the inhibitory effect of poly(N-vinylcaprolactam) nanogels against HIV-1 infection is presented for the first time. In particular, we have demonstrated that one of the synthesized poly(N-vinylcaprolactam) nanogels with initial concentration of 80 mg of vinylcaprolactam and 4% of crosslinking agent shows antiviral behavior against HIV-1 infection since this nanogel inhibits the viral replication in TZM.bl target cells.

Soybean-modified polyamide-6 mats as a long-term cutaneous wound covering.

Engineered skin coverings have been adopted clinically to support extensive and deep wounds that result in fewer healthy skin remaining and therefore take longer to heal. Nonetheless, these biomaterials demand intensive labor and an expensive final cost. In comparison to conventional bandages, which do not meet all the requirements of wound care, electrospun fiber mats could potentially provide an excellent environment for healing. In this work, we developed two nanostructured scaffolds based on polyamide-6 (PA-6) to be tested as a wound covering in a rat model of full-thickness incisional wound healing. The central idea was to create a bioconstruct that is simple to implement and biologically safe, with a high survival rate, which provides physical support and biological recognition for new functional tissues. An unmodified PA-6 and a soybean-modified PA-6 were employed as nanofibrillar matrices in this study. The biomaterials showed a dimensional homology to natural extracellular matrix components and neither in vitro toxicity nor in vivo side effects. Both polymeric scaffolds were resistant to the sterilization process and could promote the attachment of 3T3 fibroblast cells, besides successfully incorporating the growth factor PDGF-BB, which had its bioactivity extended for up to 12 h under simulated conditions. The modification of PA-6 chains with a fatty acid derivative increased the scaffold's surface free energy, favoring cell proliferation, collagen formation, and ECM secretion. These results confirm the potential of these materials as a topical dermal covering for skin regeneration.

Time-dependent deformation of artificial muscles based on Nylon 6.

Considering the potential applications of the Nylon 6 with thermal-induced deformation, we studied the creep deformation of non-twisted Nylon 6 wires and Nylon 6 artificial muscles as functions of annealing temperature. For comparison, we also studied the creep deformation of chicken muscle fibers in a temperature range of 20 to 35 °C. The experimental results showed that we could use the standard linear viscoelastic model to describe the creep deformation of the chicken muscle fibers, the non-twisted Nylon 6 wires, and the Nylon 6 artificial muscles. A simple method was developed to calculate the mechanical (elastic) constants and viscous resistance coefficient (viscosity) of the three different materials. The activation energy for the creep deformation of the chicken muscle fibers in the temperature of 20 to 35 °C was 18.79 kJ/mol. For the non-twisted Nylon 6 wires, the activation energy for the creep deformation was generally larger than that of the chicken muscle fibers, and was dependent on the annealing temperature. For the Nylon 6 artificial muscles, the activation energy for the creep deformation was smaller than that of the chicken muscle fibers.

Design of antimicrobial polycaprolactam nanocomposite by immobilizing subtilisin conjugated Au/Ag core-shell nanoparticles for biomedical applications.

Preparation of the gold core and silver shell NP (AuAgNP) is challenging because of the facile oxidation of silver. Here such a NP is carefully synthesized and conjugated with subtilisin to arrive at a stable spherical material of 120-130 nm in diameter (AuAgSNP). A biomaterial prepared by immobilizing AuAgSNP on polycaprolactam (PCL) exhibits antibiofilm properties against S. aureus and E. coli, but with lesser potency than the one prepared with bare AuAgNP. Subtilisin degrades the adhesive surface proteins of the bacteria thereby preventing the biofilm formation. Subtilisin conjugated AuAgSNP is not cytotoxic to 3T3 cells at its MIC, in contrast to AuAgNP. The presence of subtilisin promotes the fibroblast proliferation. This study indicates that AuAgSNP has antibacterial/antibiofilm activities as well as biocompatibility unlike NPs which are very cytotoxic to cells. Hence AuAgSNP can be used in medical implants and devices.

Inhibition of Candida albicans biofilm and hyphae formation by biocompatible oligomers.

Candida albicans is a yeast pathogen known for its virulence and high morbidity rate, and it easily colonizes host tissues and implant devices and forms mature biofilms, which play an important role in pathogenesis and drug resistance. In this study, we investigated the abilities of thermoresponsive oligomers of N-vinylcaprolactam (OVCLs) to inhibit biofilm formation by C. albicans. One synthetic and four commercial OVCLs (≤MW 240 000) at a concentration of 5 µg ml-1 were found to decrease C. albicans biofilm formation by more than 90% at 37°C, but to be less effective at 25°C. Microscopic observations showed that OVCLs clearly inhibited hyphal formation and cell aggregation by C. albicans, and this appeared to be responsible for their antibiofilm effects. In addition, effective antibiofouling coatings of OVCL/poly(lactic-co-glycolic acid) polymer blends were prepared on glass surfaces. SIGNIFICANCE AND IMPACT OF THE STUDY: The emergence of multidrug-resistant Candida strains has prompted searches for new antifungals. The antibiofilm and antihyphae properties of OVCLs and OVCL coating against a fluconazole-resistant Candida albicans strain are present in this study. These findings suggest that OVCL and OVCL-coated biomaterials are potentially useful for controlling fungal biofilm formation by and the virulence of antifungal-resistant C. albicans.

Polyphenolic Polymersomes of Temperature-Sensitive Poly(N-vinylcaprolactam)-block-Poly(N-vinylpyrrolidone) for Anticancer Therapy.

We report a versatile synthesis for polyphenolic polymersomes of controlled submicron (<500 nm) size for intracellular delivery of high and low molecular weight compounds. The nanoparticles are synthesized by stabilizing the vesicular morphology of thermally responsive poly(N-vinylcaprolactam)n-b-poly(N-vinylpyrrolidone)m (PVCLn-PVPONm) diblock copolymers with tannic acid (TA), a hydrolyzable polyphenol, via hydrogen bonding at a temperature above the copolymer's lower critical solution temperature (LCST). The PVCL179-PVPONm diblock copolymers are produced by controlled reversible addition-fragmentation chain transfer (RAFT) polymerization of PVPON using PVCL as a macro-chain transfer agent. The size of the TA-locked (PVCL179-PVPONm) polymersomes at room temperature and upon temperature variations are controlled by the PVPON chain length and TA:PVPON molar unit ratio. The particle diameter decreases from 1000 to 950, 770, and 250 nm with increasing PVPON chain length (m = 107, 166, 205, 234), and it further decreases to 710, 460, 290, and 190 nm, respectively, upon hydrogen bonding with TA at 50 °C. Lowering the solution temperature to 25 °C results in a slight size increase for vesicles with longer PVPON. We also show that TA-locked polymersomes can encapsulate and store the anticancer drug doxorubicin (DOX) and higher molecular weight fluorescein isothiocyanate (FITC)-dextran in a physiologically relevant pH and temperature range. Encapsulated DOX is released in the nuclei of human alveolar adenocarcinoma tumor cells after 6 h incubation via biodegradation of the TA shell with the cytotoxicity of DOX-loaded polymersomes being concentration-dependent. Our approach offers biocompatible and intracellular degradable nanovesicles of controllable size for delivery of a variety of encapsulated materials. Considering the particle monodispersity, high loading capacity, and a facile two-step aqueous assembly based on the reversible temperature-responsiveness of PVCL, these polymeric vesicles have significant potential as novel drug nanocarriers and provide a new perspective for fundamental studies on thermo-triggered polymer assemblies in solutions.

* Thermosensitive Poly(N-vinylcaprolactam) Injectable Hydrogels for Cartilage Tissue Engineering.

Injectable hydrogels have gained prominence in the field of tissue engineering for minimally invasive delivery of cells for tissue repair and in the filling of irregular defects. However, many injectable hydrogels exhibit long gelation times or are not stable for long periods after injection. To address these concerns, we used thermosensitive poly(N-vinylcaprolactam) (PNVCL) hydrogels due to their cytocompatibility and fast response to temperature stimuli. Changes in the PNVCL molecular weight and concentration enabled the development of hydrogels with tunable mechanical properties and fast gelation times (<60 s when the temperature was raised from room temperature to physiologic temperature). Chondrocytes (CHs) and mesenchymal stem cells were encapsulated in PNVCL hydrogels and exhibited high viability (∼90%), as monitored by Live/Dead staining and Alamar Blue assays. Three-dimensional constructs of CH-laden PNVCL hydrogels supported cartilage-specific extracellular matrix production both in vitro and after subcutaneous injection in nude rats for up to 8 weeks. Moreover, biochemical analyses of constructs demonstrated a time-dependent increase in glycosaminoglycans (GAGs) and collagen, which were significantly augmented in the implants cultured in vivo. Histological analyses also demonstrated regular distribution of synthesized cartilage components, including abundant GAGs and type II collagen. The findings from this study demonstrate thermosensitive PNVCL as a candidate injectable biomaterial to deliver cells for cartilage tissue engineering.

Antibacterial and antifungal screening of natural products sourced from Australian fungi and characterisation of pestalactams D-F.

Eighteen natural products sourced from Australian micro- or macro-fungi were screened for antibacterial and antifungal activity. This focused library was comprised of caprolactams, polyamines, quinones, and polyketides, with additional large-scale isolation studies undertaken in order to resupply previously identified compounds. Chemical investigations of the re-fermented culture from the endophytic fungus Pestalotiopsis sp. yielded three caprolactam analogues, pestalactams D-F, along with larger quantities of the known metabolite pestalactam A, which was methylated using diazomethane to yield 4-O-methylpestalactam A. The chemical structures of the previously undescribed fungal metabolites were determined by analysis of 1D/2D NMR and MS data. The structure of 4-O-methylpestalactam A was confirmed following single crystal X-ray diffraction analysis. The antibacterial and antifungal activity of all compounds was assessed, which identified three compounds, (1S,3R)-austrocortirubin, (1S,3S)-austrocortirubin, and 1-deoxyaustrocortirubin with mild activity (100 µM) against Gram-positive isolates and one compound, 2-hydroxy-6-methyl-8-methoxy-9-oxo-9H-xanthene-1-carboxylic acid, with activity against Cryptococcus neoformans and Cryptococcus gattii at 50 µM.

[Selective regulation of laccase isoform production by the Lentinus strigosus 1566 fungus].

The effects of a number of culture medium components, such as peptone, yeast extract, mono- and disaccharides, copper ions, 2,6-dimethylphenol, and polycaproamide fiber, on the laccase activity dynamics in the culture liquid and laccase isoform production by the Lentinus strigosus 1566 fungus were studied. It was demonstrated that some saccharides selectively induced or inhibited the synthesis of different laccase isoforms. Similar action was exerted by copper ions, 2,6-dimethylphenol, and polycaproamide fiber, as well as by their combination. Selective in vivo regulation of the production of certain laccase isoforms by basidial fungi by means of altering the culturing medium composition can be utilised for various biotechnological purposes.

Design of a papain immobilized antimicrobial food package with curcumin as a crosslinker.

Contamination of food products by spoilage and pathogenic microorganisms during post process handling is one of the major causes for food spoilage and food borne illnesses. The present green sustainable approach describes the covalent immobilization of papain to LDPE (low density polyethylene), HDPE (high density polyethylene), LLDPE (linear low density polyethylene) and PCL (polycaprolactam) with curcumin as the photocrosslinker. About 50% of curcumin and 82-92% of papain were successfully immobilized on these polymers. After 30 days, the free enzyme retained 87% of its original activity, while the immobilized enzyme retained more than 90% of its activity on these polymers. Papain crosslinked to LLDPE exhibited the best antibiofilm properties against Acinetobacter sp. KC119137.1 and Staphylococcus aureus NCIM 5021 when compared to the other three polymers, because of the highest amount of enzyme immobilized on this surface. Papain acts by damaging the cell membrane. The enzyme is able to reduce the amount of carbohydrate and protein contents in the biofilms formed by these organisms. Meat wrapped with the modified LDPE and stored at 4°C showed 9 log reduction of these organisms at the end of the seventh day when compared to samples wrapped with the bare polymer. This method of crosslinking can be used on polymers with or without functional groups and can be adopted to bind any type of antimicrobial agent.

Roles of hydroxyapatite allocation and microgroove dimension in promoting preosteoblastic cell functions on photocured polymer nanocomposites through nuclear distribution and alignment.

This study clarifies how hydroxyapatite (HA) allocation and microgroove dimension affect mouse preosteoblastic MC3T3-E1 cell functions on microgrooved substrates of polymer nanocomposites. Using replica molding from micromachined silicon wafer templates, we fabricated photocured poly(ε-caprolactone) triacrylate (PCLTA)/HA nanocomposite substrates with parallel microgrooves (two groove widths of 5 and 15 µm and one groove depth of 5 µm). Four types of microgrooved substrates were made: "homogeneous" ones of PCLTA and PCLTA/HA with uniform distribution and two "heterogeneous" laminated microgrooved substrates with HA only in the PCLTA matrix in the ridges or bottom. These substrates were used to regulate MC3T3-E1 cell attachment, proliferation, alignment, nuclear circularity and distribution, and mineralization. MC3T3-E1 cell attachment and proliferation were much higher on the microgrooved substrates of PCLTA/HA than on those of PCLTA, in particular, on the 5 µm wide microgrooved substrate with PCLTA/HA ridges and PCLTA bottom. The shape and distribution of MC3T3-E1 cytoskeleton and nuclei were altered by the substrate topography and HA allocation. For 5 µm wide heterogeneous microgrooved substrates with HA only in the ridges, MC3T3-E1 cells exhibited better spreading perpendicular to the microgrooves but tended to extend along the microgrooves containing HA in the bottom. The widest cells and the roundest/largest cell nuclei were observed on the heterogeneous substrate with PCLTA/HA ridges, while the narrowest cells with the best elongation were found on the homogeneous PCLTA/HA substrate. The trend in MC3T3-E1 cell mineralization on the substrates was consistent with that in cell/nuclear elongation. Osteocalcin mRNA expression was significantly higher on the PCLTA/HA substrates than on the PCLTA ones and also on the microgrooved substrates of PCLTA/HA than on the flat ones, regardless of the groove width of 5 or 15 µm.

Discovery of a capuramycin analog that kills nonreplicating Mycobacterium tuberculosis and its synergistic effects with translocase I inhibitors.

Capuramycin (1) and its analogs are strong translocase I (MurX/MraY) inhibitors. In our structure-activity relationship studies of capuramycin analogs against Mycobacterium tuberculosis (Mtb), we observed for the first time that a capuramycin analog, UT-01320 (3) killed nonreplicating (dormant) Mtb at low concentrations under low oxygen conditions, whereas selective MurX inhibitors killed only replicating Mtb under aerobic conditions. Interestingly, 3 did not exhibit MurX enzyme inhibitory activity even at high concentrations, however, 3 inhibited bacterial RNA polymerases with the IC50 values of 100-150 nM range. A new RNA polymerase inhibitor 3 displayed strong synergistic effects with a MurX inhibitor SQ 641 (2), a promising preclinical tuberculosis drug.

Antibiofilm properties of interfacially active lipase immobilized porous polycaprolactam prepared by LB technique.

Porous biomaterial is the preferred implant due to the interconnectivity of the pores. Chances of infection due to biofilm are also high in these biomaterials because of the presence of pores. Although biofilm in implants contributes to 80% of human infections, there are no commercially available natural therapeutics against it. In the current study, glutaraldehyde cross linked lipase was transferred onto a activated porous polycaprolactam surface using Langmuir-Blodgett deposition technique, and its thermostability, slimicidal, antibacterial, biocompatibility and surface properties were studied. There was a 20% increase in the activity of the covalently crosslinked lipase when compared to its free form. This immobilized surface was thermostable and retained activity and stability until 100°C. There was a 2 and 7 times reduction in carbohydrate and 9 and 5 times reduction in biofilm protein of Staphylococcus aureus and Escherichia coli respectively on lipase immobilized polycaprolactam (LIP) when compared to uncoated polycaprolactam (UP). The number of live bacterial colonies on LIP was four times less than on UP. Lipase acted on the cell wall of the bacteria leading to its death, which was confirmed from AFM, fluorescence microscopic images and amount of lactate dehydrogenase released. LIP allowed proliferation of more than 90% of 3T3 cells indicating that it was biocompatible. The fact that LIP exhibits antimicrobial property at the air-water interface to hydrophobic as well as hydrophilic bacteria along with lack of cytotoxicity makes it an ideal biomaterial for biofilm prevention in implants.

Thermoresponsive oligomers reduce Escherichia coli O157:H7 biofouling and virulence.

Thermoresponsive polymers have potential biomedical applications for drug delivery and tissue engineering. Here, two thermoresponsive oligomers were synthesized, viz. oligo(N-isopropylacrylamide) (ONIPAM) and oligo(N-vinylcaprolactam) (OVCL), and their anti-biofouling abilities investigated against enterohemorrhagic E. coli O157:H7, which produces Shiga-like toxins and forms biofilms. Biofilm formation (biofouling) is closely related to E. coli O157:H7 infection and constitutes a major mechanism of antimicrobial resistance. The synthetic OVCL (MW 679) and three commercial OVCLs (up to MW 54,000) at 30 µg ml(-1) were found to inhibit biofouling by E. coli O157:H7 at 37 °C by more than 80% without adversely affecting bacterial growth. The anti-biofouling activity of ONIPAM was weaker than that of OVCL. However, at 25 °C, ONIPAM and OVCL did not affect E. coli O157:H7 biofouling. Transcriptional analysis showed that OVCL temperature-dependently downregulated curli genes in E. coli O157:H7, and this finding was in line with observed reductions in fimbriae production and biofouling. In addition, OVCL downregulated the Shiga-like toxin genes stx1 and stx2 in E. coli O157:H7 and attenuated its in vivo virulence in the nematode Caenorhabditis elegans. These results suggest that OVCL has potential use in antivirulence strategies against persistent E. coli O157:H7 infection.