Contribution, Composition, and Structure of EPS by In Vivo Exposure to Elucidate the Mechanisms of Nanoparticle-Enhanced Bioremediation to Metals.

Bacterial extracellular polymeric substances (EPS) have been recently found to contribute most for metal removal in nanoenhanced bioremediation. However, the mechanism by which NPs affect EPS-metal interactions is not fully known. Here, Halomonas sp. was employed to explore the role of EPS after in vivo exposure to Cd/Pb and polyvinylpyrrolidone (PVP) coated iron oxide nanoparticles (IONPs, 20 mg L-1) for 72 h. Cd-IONPs produced the highest concentrations of EPS proteins (136.3 mg L-1), while Cd induced the most production of polysaccharides (241.0 mg L-1). IONPs increased protein/polysaccharides ratio from 0.2 (Cd) to 1.2 (Cd-IONPs). The increased protein favors the formation of protein coronas on IONPs surface, which would promote Cd adsorption during NP-metal-EPS interaction. FTIR analysis indicated that the coexistence of Cd and IONPs interacted with proteins more strongly than with polysaccharides. Glycosyl monomer analyses suggested mannose and glucose as target sugars for EPS complexation with metals, and IONPs reduced metal-induced changes in monosaccharide profiles. Protein secondary structures changed in all treatments, but we could not distinguish stresses induced by metals from those by IONPs. These findings provide greater understanding of the role of EPS in NP-metal-EPS interaction, providing a better underpinning knowledge for the application of NP-enhanced bioremediation.

Nanoparticles as antibiotic-delivery vehicles (ADVs) overcome resistance by MRSA and other MDR bacterial pathogens: The grenade hypothesis.

OBJECTIVES: The aim of this study was to examine how the concentrated delivery of less effective antibiotics, such as the ß-lactam penicillin G, by linkage to nanoparticles (NPs), could influence the killing efficiency against various pathogenic bacteria, including methicillin-resistant Staphylococcus aureus (MRSA) and other multidrug resistant (MDR) strains. METHODS: The ß-lactam antibiotic penicillin G (PenG) was passively sorbed to fluorescent polystyrene NPs (20nm) that were surface-functionalized with carboxylic acid (COO--NPs) or sulfate groups (SO4--NPs) to form a PenG-NP complex. Antimicrobial activities of PenG-NPs were evaluated against Gram-negative and Gram-positive bacteria, including antibiotic resistant strains. Disc diffusion, microdilution assays and live/dead staining were performed for antibacterial assessments. RESULTS: The results showed that bactericidal activities of PenG-NP complexes were statistically significantly (P<0.05) enhanced against Gram-negative and Gram-positive strains, including MRSA and MDR strains. Fluorescence imaging verified that NPs comigrated with antibiotics throughout clear zones of MIC agar plate assays. The increased bactericidal abilities of NP-linked antibiotics are hypothesized to result from the greatly increased densities of antibiotic delivered by each NP to a given bacterial cell (compared with solution concentrations of antibiotic), which overwhelms the bacterial resistance mechanism(s). CONCLUSIONS: As a whole, PenG-NP complexation demonstrated a remarkable activity against different pathogenic bacteria, including MRSA and MDR strains. We term this the 'grenade hypothesis'. Further testing and development of this approach will provide validation of its potential usefulness for controlling antibiotic-resistant bacterial infections.

Improved metal remediation using a combined bacterial and nanoscience approach.

Polyvinylpyrrolidone (PVP) coated iron oxide nanoparticles (NPs) were used to explore the potential for improved bioremediation of metals by interaction with the Gram-negative bacterium Halomonas sp. The combined approach improved metal removal and shortened metal remediation times (approx. 100% removal of Pb after 24 h, of Cd after 48 h) compared with bacteria- or NP-only controls. NPs also demonstrated the ability to reduce metal toxicity to bacteria and enhance bacterial growth efficiencies in an additive manner. Cd, Pb, and Fe (from NPs) were analyzed in the following operationally-defined components: EPS, cell-wall, cell membrane, and cytoplasmic fractions; EPS was most important in metal removal. There was a significant promotion of Cd intracellular transportation, but not Pb, by NPs. Reduced Pb internalization may have resulted from EPS acting as an uptake barrier coupled with an effective efflux system of Halomonas sp. as a resistance mechanism. In addition, the majority of Fe was present in bacterial membranes, compared with Cd or Pb, suggesting that bacteria may take up iron oxide NPs as a potential nutrient while recognizing Cd or Pb as toxicants.

Gold Nanoparticles with Antibiotic-Metallopolymers toward Broad-Spectrum Antibacterial Effects.

Bacterial infection has evolved into one of the most dangerous global health crises. Designing potent antimicrobial agents that can combat drug-resistant bacteria is essential for treating bacterial infections. In this paper, a strategy to graft metallopolymer-antibiotic bioconjugates on gold nanoparticles is developed as an antibacterial agent to fight against different bacterial strains. Thus, these nanoparticle conjugates combine various components in one system to display enhanced bactericidal efficacy, in which small sized nanoparticles provide high surface area for bacteria to contact, cationic metallopolymers interact with the negatively charged bacterial membranes, and the ß-lactam antibiotics' sterilzation capabilities are improved via evading intracellular enzymolysis by ß-lactamase. This nanoparticle-based antibiotic-metallopolymer system exhibits an excellent broad-spectrum antibacterial effect, particularly for Gram-negative bacteria, due to the synergistic effect of multicomponents on the interaction with bacteria.

Recyclable magnetic nanoparticles grafted with antimicrobial metallopolymer-antibiotic bioconjugates.

Over-prescription and improper use of antibiotics has led to the emergence of bacterial resistance, posing a major threat to public health. There has been significant interest in the development of alternative therapies and agents to combat antibiotic resistance. We report the preparation of recyclable magnetic iron oxide nanoparticles grafted with charged cobaltocenium-containing metallopolymers by surface-initiated reversible addition-fragmentation chain transfer (RAFT) polymerization. ß-Lactam antibiotics were then conjugated with metallopolymers to enhance their vitality against both Gram-positive and Gram-negative bacteria. The enhanced antibacterial activity was a result of synergy of antimicrobial segments that facilitate the inhibition of hydrolysis of antibiotics and local enhancement of antibiotic concentration on a nanoparticle surface. These magnetic nanoparticles can be recycled numerous times without losing the initial antimicrobial potency. Studies suggested negligible toxicity of metallopolymer-grafted nanoparticles to red blood cells and minimal tendency to induce resistance in bacteria.

A novel method for the synergistic remediation of oil-water mixtures using nanoparticles and oil-degrading bacteria.

Releases of crude oil and other types of oil from numerous sources can impose catastrophic physical, chemical, and biological effects on aquatic ecosystems. While currently-used oil removal techniques possess many advantages, they have inherent limitations, including low removal efficiencies and waste disposal challenges. The present study quantified the synergistic interactions of polyvinylpyrrolidone (PVP) coated magnetite nanoparticles (NP) and oil-degrading bacteria for enhanced oil removal at the laboratory scale. The results showed that at relatively high oil concentrations (375 mg L-1), NP alone could remove approximately 70% of lower-chain alkanes (C9-C22) and 65% of higher-chain (C23-C26), after only 1 h, when magnetic separation of NP was used. Removal efficiency did not increase significantly after that, which was likely due to saturation of the NP with oil. Microbial bioremediation, using strains of oil-degrading bacteria, removed almost zero oil immediately but 80-90% removal after 24-48 h. The combination of NPs and oil-degrading bacterial strains worked effectively to remove essentially 100% of oil within 48 h or less. This was likely due to the sorption of oil components to NPs and their subsequent utilization by bacteria as a joint Fe and C source, although the mechanisms of removal require further testing. Furthermore, results showed that the emission of selected volatile organic compounds (VOCs) and semi volatile organic compounds (SVOCs) were reduced after addition of NPs and bacteria separately. When combined, VOC and SVOC emissions were reduced by up to 80%.

Charged Metallopolymer-Grafted Silica Nanoparticles for Antimicrobial Applications.

Inappropriate and frequent use of antibiotics has led to the development of antibiotic-resistant bacteria, which cause infectious diseases that are difficult to treat. With the rising threat of antibiotic resistance, the need to develop effective new antimicrobial agents is prominent. We report antimicrobial metallopolymer nanoparticles, which were prepared by surface-initiated reversible addition-fragmentation chain transfer polymerization of a cobaltocenium-containing methacrylate monomer from silica nanoparticles. These particles are capable of forming a complex with ß-lactam antibiotics, such as penicillin, rejuvenating the bactericidal activity of the antibiotic. Disk diffusion assays showed significantly increased antibacterial activities against both Gram-positive and Gram-negative bacteria. The improved efficiencies were attributed to the inhibition of hydrolysis of the ß-lactam antibiotics and enhancement of local antibiotics concentration on a nanoparticle surface. In addition, hemolysis evaluations demonstrated minimal toxicity to red blood cells.

Trio Act of Boronolectin with Antibiotic-Metal Complexed Macromolecules toward Broad-Spectrum Antimicrobial Efficacy.

Bacterial infections, particularly by Gram-negative pathogens, have become a serious threat to global healthcare due to the diminishing effectiveness of existing antibiotics. We report a nontraditional therapy to combine three components in one macromolecular system, in which boronic acid adheres to peptidoglycan or lipopolysaccharide via boron-polyol based boronolectin chemistry, cationic metal polymer frameworks interact with negatively charged cell membranes, and ß-lactam antibiotics are reinstated with enhanced vitality to attack bacteria via evading the detrimental enzyme-catalyzed hydrolysis. These macromolecular systems exhibited high efficacy in combating pathogenic bacteria, especially Gram-negative strains, due to synergistic effects of multicomponents on interactions with bacterial cells. In vitro and in vivo cytotoxicity and hemolysis evaluation indicated that these multifunctional copolymers did not induce cell death by apoptosis, as well as did not alter the phenotypes of immune cells and did not show observable toxic effect on red blood cells.

Complete Genome Sequence of Vibrio gazogenes ATCC 43942.

Vibrio gazogenes ATCC 43942 has the potential to synthesize a plethora of metabolites which are of clinical and agricultural significance in response to environmental triggers. The complete genomic sequence of Vibrio gazogenes ATCC 43942 is reported herein, contributing to the knowledge base of strains in the Vibrio genus.

Aflatoxin-Exposure of Vibrio gazogenes as a Novel System for the Generation of Aflatoxin Synthesis Inhibitors.

Aflatoxin is a mycotoxin and a secondary metabolite, and the most potent known liver carcinogen that contaminates several important crops, and represents a significant threat to public health and the economy. Available approaches reported thus far have been insufficient to eliminate this threat, and therefore provide the rational to explore novel methods for preventing aflatoxin accumulation in the environment. Many terrestrial plants and microbes that share ecological niches and encounter the aflatoxin producers have the ability to synthesize compounds that inhibit aflatoxin synthesis. However, reports of natural aflatoxin inhibitors from marine ecosystem components that do not share ecological niches with the aflatoxin producers are rare. Here, we show that a non-pathogenic marine bacterium, Vibrio gazogenes, when exposed to low non-toxic doses of aflatoxin B1, demonstrates a shift in its metabolic output and synthesizes a metabolite fraction that inhibits aflatoxin synthesis without affecting hyphal growth in the model aflatoxin producer, Aspergillus parasiticus. The molecular mass of the predominant metabolite in this fraction was also different from the known prodigiosins, which are the known antifungal secondary metabolites synthesized by this Vibrio. Gene expression analyses using RT-PCR demonstrate that this metabolite fraction inhibits aflatoxin synthesis by down-regulating the expression of early-, middle-, and late- growth stage aflatoxin genes, the aflatoxin pathway regulator, aflR and one global regulator of secondary metabolism, laeA. Our study establishes a novel system for generation of aflatoxin synthesis inhibitors, and emphasizes the potential of the under-explored Vibrio's silent genome for generating new modulators of fungal secondary metabolism.

Long-term antihypertensive effects of far-infrared ray irradiated from wooden board in spontaneously hypertensive rats.

BACKGROUND: Far-infrared ray (FIR) has been widely used in promoting health and has been shown to exert beneficial effects in vascular function. The non-thermal effect of FIR has been found to play a significant role in the protective effect on some vascular-related diseases, but its protective effects and use against hypertension have not been clearly presented. METHODS: In the present study, by using a wooden board coated with FIR-irradiated materials, we evaluated the long-term antihypertensive effect on spontaneously hypertensive rats (SHRs) in the environment in contact with the FIR-irradiated wooden board. SHRs were placed on the wooden board with or without FIR radiation for 4 weeks. RESULTS: The systolic blood pressure (BP) of SHRs undergoing different treatments was measured weekly using a tail-cuff method. FIR radiation significantly reduced the systolic BP of the SHRs along with a decreasing plasma level of angiotensin II and an increasing plasma level of bradykinin. In addition, long-term contact of FIR did not significantly affect the BP in normotensive Wistar Kyoto rats (WKYs). CONCLUSIONS: Our results provided the evidence based on which FIR radiation could be considered an effective non-pharmacological choice for preventing hypertension.

Engineering nanoparticles to silence bacterial communication.

The alarming spread of bacterial resistance to traditional antibiotics has warranted the study of alternative antimicrobial agents. Quorum sensing (QS) is a chemical cell-to-cell communication mechanism utilized by bacteria to coordinate group behaviors and establish infections. QS is integral to bacterial survival, and therefore provides a unique target for antimicrobial therapy. In this study, silicon dioxide nanoparticles (Si-NP) were engineered to target the signaling molecules [i.e., acylhomoserine lactones (HSLs)] used for QS in order to halt bacterial communication. Specifically, when Si-NP were surface functionalized with ß-cyclodextrin (ß-CD), then added to cultures of bacteria (Vibrio fischeri), whose luminous output depends upon HSL-mediated QS, the cell-to-cell communication was dramatically reduced. Reductions in luminescence were further verified by quantitative polymerase chain reaction (qPCR) analyses of luminescence genes. Binding of HSLs to Si-NPs was examined using nuclear magnetic resonance (NMR) spectroscopy. The results indicated that by delivering high concentrations of engineered NPs with associated quenching compounds, the chemical signals were removed from the immediate bacterial environment. In actively-metabolizing cultures, this treatment blocked the ability of bacteria to communicate and regulate QS, effectively silencing and isolating the cells. Si-NPs provide a scaffold and critical stepping-stone for more pointed developments in antimicrobial therapy, especially with regard to QS-a target that will reduce resistance pressures imposed by traditional antibiotics.

Functionalised nanoparticles complexed with antibiotic efficiently kill MRSA and other bacteria.

Antibiotic-resistant bacterial infections are a vexing global health problem and have rendered ineffective many previously-used antibiotics. Here we demonstrate that antibiotic-linkage to surface-functionalized silica nanoparticles (sNP) significantly enhances their effectiveness against Escherichia coli, and Staphylococcus aureus, and even methicillin-resistant S. aureus (MRSA) strains that are resistant to most antibiotics. The commonly-used antibiotic penicillin-G (PenG) was complexed to dye-labeled sNPs (15 nm diameter) containing carboxyl groups located as either surface-functional groups, or on polymer-chains extending from surfaces. Both sNPs configurations efficiently killed bacteria, including MRSA strains. This suggests that activities of currently-ineffective antibiotics can be restored by nanoparticle-complexation and used to avert certain forms of antibiotic-resistance.

Antimicrobial metallopolymers and their bioconjugates with conventional antibiotics against multidrug-resistant bacteria.

Bacteria are now becoming more resistant to most conventional antibiotics. Methicillin-resistant Staphylococcus aureus (MRSA), a complex of multidrug-resistant Gram-positive bacterial strains, has proven especially problematic in both hospital and community settings by deactivating conventional ß-lactam antibiotics, including penicillins, cephalosporins, and carbapenems, through various mechanisms, resulting in increased mortality rates and hospitalization costs. Here we introduce a class of charged metallopolymers that exhibit synergistic effects against MRSA by efficiently inhibiting activity of ß-lactamase and effectively lysing bacterial cells. Various conventional ß-lactam antibiotics, including penicillin-G, amoxicillin, ampicillin, and cefazolin, are protected from ß-lactamase hydrolysis via the formation of unique ion-pairs between their carboxylate anions and cationic cobaltocenium moieties. These discoveries could provide a new pathway for designing macromolecular scaffolds to regenerate vitality of conventional antibiotics to kill multidrug-resistant bacteria and superbugs.

Surface-functionalization effects on uptake of fluorescent polystyrene nanoparticles by model biofilms.

A study was conducted to investigate the role of nanoparticle (NP) surface functionalization/charge on their uptake by biofilms. Biofilms, bacterial colonies attached to surfaces via extracellular polymers, are effective at removing suspended nanomaterials from the aqueous phase. However, the mechanisms regulating particle uptake are unknown. Here, it was shown that the mechanism was strongly dependent on the nanoparticle surface ionization, and not the core composition of the NP. Uptake experiments were conducted using laboratory-cultured biofilms. The biofilms were incubated in the presence of fluorescent polystyrene NPs with either negatively-charged surfaces (i.e. functionalized with sulfated (SO(4) (-)-NP) or carboxylated (COO(-)-NP) groups) or positively-charged surfaces (functionalized with primary amines, Amine-P). Particles with negatively-charged sulfated surfaces associated most strongly to biofilms across all experimental conditions. Associations of positively-charged amine particles with biofilms were greatest at high ionic conditions resembling those of seawater, but were sensitive to changes in ionic strength. Sorption of COO(-)-NPs was lowest, relative to other particle types, and was not sensitive to ionic strength. The results of this study support an emerging precedent that biofilms may be an effective player in the binding and sequestration of nanoparticles in aqueous systems.

Robust antimicrobial compounds and polymers derived from natural resin acids.

We report novel robust resin acid-derived antimicrobial agents that exhibit excellent antimicrobial activities against a broad spectrum of bacteria (6 Gram-positive and 7 Gram-negative) with selective lysis of microbial membranes over mammalian membranes. Our results indicate that hydrophobicity and unique structures of resin acids can be determining factors in dictating the antimicrobial activity.

Fabrication and characterization of multi-scale microlens arrays with anti-reflection and diffusion properties.

In this paper, an effective method for fabricating artificial compound-eye structures is demonstrated. The fabrication of high fill factor microlens arrays (MLAs) with sub-wavelength structures (SWSs) on a polycarbonate (PC) substrate involves nanoimprint and thermo-extrusion techniques by using two different scales of nano/micromolds. In addition, the MLAs with SWSs on the PC substrate would be replicated on a polymethylmethacrylate (PMMA) millimeter concave surface by hot-embossing, forming three-level compound-eye structures. The optical properties of these samples are characterized. The transmittances of two-level PC and three-level PMMA compound structures are increased by 2.5% and 2%, and the uniformities are enhanced by 18% and 24%, respectively.

A permanent optical storage medium exhibiting ultrahigh contrast, superior stability, and a broad working wavelength regime.

In this paper we demonstrate an optical storage medium having advantages of ultrahigh contrast, superior stability, and broadband working wavelengths. Combining a single shot of deep-ultraviolet (UV) laser illumination with a Au particle-assisted etching process, we formed broadband antireflective, one-dimensional silicon nanowire arrays (SiNWs) with selectively at specific positions. Optical measurements and three-dimensional finite-difference time domain (3D-FDTD) simulations revealed ultrahigh reflection contrast between the Au and the SiNWs for both far- and near-field regimes. Relative to typical organic-based storage media, Au films and SiNWs are more stable, both chemically and thermally; therefore, we suspect that this new storage medium would exhibit high stability toward moisture, sunshine, and elevated temperatures.

One-shot deep-UV pulsed-laser-induced photomodification of hollow metal nanoparticles for high-density data storage on flexible substrates.

In this paper, we report a new optical data storage method: photomodification of hollow gold nanoparticle (HGN) monolayers induced by one-shot deep-ultraviolet (DUV) KrF laser recording. As far as we are aware, this study is the first to apply HGNs in optical data storage and also the first to use a recording light source for the metal nanoparticles (NPs) that is not a surface plasmon resonance (SPR) wavelength. The short wavelength of the recording DUV laser improved the optical resolution dramatically. We prepared HGNs exhibiting two absorbance regions: an SPR peak in the near-infrared (NIR) region and an intrinsic material extinction in the DUV region. A single pulse from a KrF laser heated the HGNs and transformed them from hollow structures to smaller solid spheres. This change in morphology for the HGNs was accompanied by a significant blue shift of the SPR peak. Employing this approach, we demonstrated its patterning ability with a resolving power of a half-micrometer (using a phase mask) and developed a readout method (using a blue-ray laser microscope). Moreover, we prepared large-area, uniform patterns of monolayer HGNs on various substrates (glass slides, silicon wafers, flexible plates). If this spectral recording technique could be applied onto thin flexible tapes, the recorded data density would increase significantly relative to that of current rigid discs (e.g., compact discs).

Rapid screening of quorum-sensing signal N-acyl homoserine lactones by an in vitro cell-free assay.

A simple, sensitive, and rapid cell-free assay system was developed for detection of N-acyl homoserine lactone (AHL) autoinducers involved in bacterial quorum sensing (QS). The present approach improves upon previous whole-cell biosensor-based approaches in its utilization of a cell-free assay approach to conduct bioassays. The cell-free assay was derived from the AHL biosensor bacterium Agrobacterium tumefaciens NTL4(pCF218)(pCF372), allowing the expression of beta-galactosidase upon addition of exogenous AHLs. We have shown that beta-galactosidase expression is possible in cell-free solution [lysate from Agrobacterium tumefaciens NTL4(pCF218)(pCF372) culture]. Assay detection limits with the use of chromogenic substrate X-Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside) ranged from approximately 100 nM to 300 nM depending on the specific AHL. Replacement (of X-Gal) with the luminescent substrate Beta-Glo increased sensitivity to AHLs by 10-fold. A major advantage of the cell-free assay system is elimination of time-consuming steps for biosensor cell culture conditioning, which are required prior to whole-cell bioassays. This significantly reduced assay times from greater than 24 h to less than 3 h, while maintaining high sensitivity. Assay lysate may be prepared in bulk and stored (-80 degrees C) over 6 months for future use. Finally, the present protocol may be adapted for use with other biosensor strains and be used in high-throughput AHL screening of bacteria or metagenomic libraries.