Physicochemical properties of Shiga toxigenic Escherichia coli

AIMS: To investigate the physicochemical surface properties, such as cellular surface charge, hydrophobicity and electron donor/acceptor potential of a selection of Shiga toxigenic Escherichia coli (STEC) isolates grown in broth and agar culture. METHODS AND RESULTS: Cellular surface charge was determined using zeta potential measurements. Hydrophobicity of the isolates was determined using bacterial adhesion to hydrocarbons assay, hydrophobic interaction chromatography and contact angle measurements. Microbial adhesion to solvents was used to determine the electron donor/acceptor characteristics. No differences of surface charge measurements were found between broth and agar grown cultures. Isolates belonging to serogroup O157 and serotypes O26:H11 and O111:H- were significantly (P < 0.05) less negatively charged than other STEC serotypes tested. All strains were hydrophilic with most methods and demonstrated a lower hydrophobicity in agar culture compared with broth culture. All strains demonstrated a strong microbial adhesion to chloroform indicating that STEC possess an electron donor and basic character. A relationship between serogroup O157 and other STEC serotypes was apparent using principal-component analysis (PCA). CONCLUSIONS: Combining the results for physicochemical properties using PCA differentiated between strains belonging to the O157 serogroup and other STEC/non-STEC strains. PCA found similar results for broth and agar grown cultures. SIGNIFICANCE AND IMPACT OF THE STUDY: Particular serotypes of STEC possess similar physicochemical properties which may play a role in their pathogenicity or potential attachment to various surfaces.     

Effects of DNP on the cell surface properties of marine bacteria and its implication for adhesion to surfaces

Abstract

The effect of 2, 4-dinitrophenol (DNP) on the extracelluar polysaccharides (EPS), cell surface charge, and the hydrophobicity of six marine bacterial cultures was studied, and its influence on attachment of these bacteria to glass and polystyrene was evaluated. DNP treatment did not influence cell surface charge and EPS production, but had a significant effect on hydrophobicity of both hydrophilic (p = 0.05) and hydrophobic (p = 0.01) cultures. Significant reduction in the attachment of all the six cultures to glass (p = 0.02) and polystyrene (p = 0.03) was observed after DNP treatment. Moreover, hydrophobicity but not the cell surface charge or EPS production influenced bacterial cell attachment to glass and polystyrene. From this study, it was evident that DNP treatment influenced bacterial cell surface hydrophobicity, which in turn, reduced bacterial adhesion to surfaces.     

Investigating the effects of positive charge and hydrophobicity on the cell selectivity, mechanism of action and anti-inflammatory activity of a Trp-rich antimicrobial peptide indolicidin

To investigate the effects of positive charge and hydrophobicity on the cell selectivity, mechanism of action and anti-inflammatory activity of a Trp-rich antimicrobial peptide indolicidin (IN), a series of IN analogs with Trp→Lys substitution were synthesized. All IN analogs displayed an approximately 7- to 18-fold higher cell selectivity, compared with IN. IN, IN-1 and IN-2 depolarized (50−90%) the cytoplasmic membrane potential of Staphylococcus aureus close to minimal inhibitory concentration (5–10 μg mL⁻¹). However, other IN analogs (IN-3 and IN-4) displayed very low ability in membrane depolarization even at 40 μg mL⁻¹. Confocal laser-scanning microscopy revealed that IN-3 and IN-4 penetrated the Escherichia coli cell membrane, whereas IN, IN-1 and IN-2 did not enter the cell membrane. In the gel retardation assay, IN-3 and IN-4 bound more strongly to DNA compared with IN, IN-1 and IN-2. These findings suggest that the mechanism of antimicrobial action of IN-3 and IN-4 may be involved in the inhibition of intracellular functions via interference with DNA/RNA synthesis. Unlike IN, all IN analogs did not inhibit nitric oxide production or inducible nitric oxide synthase mRNA expression in lipopolysaccharide-stimulated mouse macrophage RAW264.7 cells, indicating that the hydrophobicity of IN is more important for anti-inflammatory activity in lipopolysaccharide-treated macrophage cells than the positive charge.     

Vapour–phase activities of essential oils against antibiotic sensitive and resistant bacteria including MRSA

Aims:  To determine whether essential oil (EO) vapours could reduce surface and airborne levels of bacteria including methicillin-resistant Staphylococcus aureus (MRSA).
Methods and Results:  The antibacterial activity of geranium and lemongrass EO individually and blended were evaluated over a range of concentrations by direct contact and vapour diffusion. The EO were tested in vitro against a selection of antibiotic-sensitive and -resistant bacteria, including MRSA, vancomycin-resistant Enterococci (VRE), Acinetobacter baumanii and Clostridium difficile . An EO blend containing lemongrass and geranium was used to formulate BioScent^TM that was dispersed into the environment using the ST Pro^TM machine. The effects were variable depending on the methods used. In a sealed box environment, MRSA growth on seeded plates was reduced by 38% after 20 h exposure to BioScent^TM vapour. In an office environment, the ST Pro^TM machine dispersing BioScent^TM effected an 89% reduction of airborne bacteria in 15 h, when operated at a constant output of 100%.
Conclusions:  EO vapours inhibited growth of antibiotic-sensitive and -resistant bacteria in vitro and reduced surface and airborne levels of bacteria.
Significance and Impact of the Study:  Results suggest that EO vapours, particularly Bioscent^TM, could be used as a method of air disinfection.     

Individual and Coupled Effects of Echinoderm Extracts and Surface Hydrophobicity on Spore Settlement and Germination in the Brown Alga Hincksia irregularis

Marine substrata possess cues that influence the behavior of fouling organisms. Initial adhesion of fouling algal zoospores to surfaces is also theorized to depend primarily upon interactions between substrata and spore cell bodies and flagellar membranes. In an effort to identify cues and surface characteristics that influence spore settlement and early development, the effects of bioactive echinoderm extracts, surface charge, and surface hydrophobicity were examined individually and in tandem on zoospore settlement and germination in Hincksia irregularis. Experiments utilizing 96-well plastic culture plates confirmed that spore settlement and germination were significantly affected by surface charge and hydrophobicity as well as by echinoderm metabolites, both individually and in tandem. Spore settlement rates in the dark over 30?min were >?400% higher on hydrophobic surfaces than on positively and negatively charged surfaces. Spore germling numbers were >?300% higher on hydrophobic surfaces than on positively and negatively charged surfaces when spores were allowed to settle in the light for 30?min and the settled spores allowed to subsequently germinate for 24?h. Spore germling numbers were consistently >?25% higher on hydrophobic surfaces than on positively and negatively charged surfaces when equal numbers of spores were allowed to completely settle in the light and subsequently germinate for 24?h. H. irregularis germ tube lengths were also significantly longer on positively charged plates than on negatively charged plates. All echinoderm extracts tested had significant effects on germination and settlement at levels below those of estimated ecological concentrations. Short-term (30?min) exposure and subsequent germination experiments indicated that higher concentrations of extracts had rapid toxic effects on algal spores. Synchronous effects of echinoderm extracts and plate charge upon spore settlement varied considerably and did not show a strong dose response relationship. Long-term (24?h) exposure of spores to echinoderm extracts had dosage dependent effects on germination and spore survival. The results of this study indicate that H. irregularis spores possess the capacity for complex responses to their environment, utilizing combined cues of surface charge, surface energy and biochemistry to determine where to settle and germinate. These responses may aid spores in the detection of suitable substrata and conditions for settlement in the marine environment.     

Production of class II bacteriocins by lactic acid bacteria; an example of biological warfare and communication

Lactic acid bacteria (LAB) fight competing Gram-positive microorganisms by secreting anti-microbial peptides called bacteriocins. Peptide bacteriocins are usually divided into lantibiotics (class I) and non-lantibiotics (class II), the latter being the main topic of this review. During the past decade many of these bacteriocins have been isolated and characterized, and elements of the genetic mechanisms behind bacteriocin production have been unravelled. Bacteriocins often have a narrow inhibitory spectrum, and are normally most active towards closely related bacteria likely to occur in the same ecological niche. Lactic acid bacteria seem to compensate for these narrow inhibitory spectra by producing several bacteriocins belonging to different classes and having different inhibitory spectra. The latter may also help in counteracting the possible development of resistance mechanisms in target organisms. In many strains, bacteriocin production is controlled in a cell-density dependent manner, using a secreted peptide-pheromone for quorum-sensing. The sensing of its own growth, which is likely to be comparable to that of related species, enables the producing organism to switch on bacteriocin production at times when competition for nutrients is likely to become more severe. Although today a lot is known about LAB bacteriocins and the regulation of their production, several fundamental questions remain to be solved. These include questions regarding mechanisms of immunity and resistance, as well as the molecular basis of target-cell specificity. This revised version was published online in August 2006 with corrections to the Cover Date.     

Various physicochemical and surface properties controlling the bioactivity of cerium oxide nanoparticles

Amidst numerous emerging nanoparticles, cerium oxide nanoparticles (CNPs) possess fascinating pharmacological potential as they can be used as a therapeutic for various oxidative stress-associated chronic diseases such as cancer, inflammation and neurodegeneration due to unique redox cycling between Ce³⁺ and Ce⁴⁺ oxidation states on their surface. Lattice defects generated by the formation of Ce³⁺ ions and compensation by oxygen vacancies on CNPs surface has led to switching between CeO₂ and CeO_2–x during redox reactions making CNPs a lucrative catalytic nanoparticle capable of mimicking key natural antioxidant enzymes such as superoxide dismutase and catalase. Eventually, most of the reactive oxygen species and nitrogen species in biological system are scavenged by CNPs via an auto-regenerative mechanism in which a minimum dose can exhibit catalytic activity for a longer duration. Due to the controversial outcomes on CNPs toxicity, considerable attention has recently been drawn towards establishing relationships between the physicochemical properties of CNPs obtained by different synthesis methods and biological effects ranging from toxicity to therapeutics. Unlike non-redox active nanoparticles, variations in physicochemical properties and the surface properties of CNPs obtained from different synthesis methods can significantly affect their biological activity (inactive, antioxidant, or pro-oxidant). Moreover, these properties can influence the biological identity, cellular interactions, cellular uptake, biodistribution, and therapeutic efficiency. This review aims to highlight the critical role of various physicochemical and the surface properties of CNPs controlling their biological activity based on 165 cited references.     

Growth phase-dependent surface properties of Legionella pneumophila and their role in adhesion to stainless steel coated QCM-D sensors

Legionella pneumophila cell surface hydrophobicity and charge are important determinants of their mobility and persistence in engineered water systems (EWS). These surface properties may differ depending on the growth phase of L. pneumophila resulting in variable adhesion and persistence within EWS. We describe the growth-dependent variations in L. pneumophila cell surface hydrophobicity and surface charge using the microbial adhesion to hydrocarbon assay and microelectrophoresis, respectively, and their role in cell adhesion to stainless steel using a quartz crystal microbalance with dissipation (QCM-D) monitoring instrument. We observed a steady increase in L. pneumophila hydrophobicity during their lifecycle in culture media. Cell surfaces of stationary phase L. pneumophila were significantly more hydrophobic than their lag and midexponential counterparts. No significant changes in L. pneumophila cell surface charge were noted. Morphology of L. pneumophila remained relatively constant throughout their lifecycle. In the QCM-D study, lag and exponential phase L. pneumophila weakly adhered to stainless steel surfaces resulting in viscoelastic layers. In contrast, stationary phase bacteria were tightly and irreversibly bound to the surfaces, forming rigid layers. Our results suggest that the stationary phase of L. pneumophila would highly favour their adhesion to plumbing surfaces and persistence in EWS.     

Effective antimicrobial activity of Cbf‐K16 and Cbf‐A7A13 against NDM‐1‐carrying Escherichia coli by DNA binding after penetrating the cytoplasmic membrane in vitro

New Delhi metallo‐beta‐lactamase‐1(NDM‐1)‐carrying isolates, which are resistant to most clinical used antibiotics except for tigecycline and colistin, have been found worldwide. Cathelicidin‐BF (BF‐30) is found in the venom of the snake Bungarus fasciatus and exhibits broad antimicrobial activity. Cbf‐K₁₆ and Cbf‐A₇A₁₃ were obtained by mutating Lys¹⁶, Ala⁷, and Ala¹³ of BF‐30, respectively. To investigate their antimicrobial activities against NDM‐1 carrying bacteria, recombinant Escherichia coli BL21 (DE3)‐NDM‐1 with high NDM‐1 activity was constructed by inserting the Klebsiella pneumoniae NDM‐1 gene (GenBank accession no. HQ328085) into a pET28a vector and transforming it into E. coli BL21 (DE3). The peptides showed effective antimicrobial activities against NDM‐1‐carrying E. coli, and the minimum inhibitory concentrations of Cbf‐K₁₆ and Cbf‐A₇A₁₃ were only 4 and 8 µg/ml, whereas those of minimum bactericidal concentrations were 8 and 16 µg/ml, respectively. A time course experiment showed that colony forming unit counts rapidly decreased, and bacteria were thoroughly eliminated within 3 and 6 h by the Cbf‐K₁₆ and Cbf‐A₇A₁₃ treatments, respectively. The peptides penetrated the bacterial cell membrane and enabled β‐galactosidase leakage, and caused the cytoplasmic membrane to become permeable, and finally bound to the DNA. The genomic DNA of E. coli was completely unable to migrate on an agarose gel after Cbf‐K₁₆ treatment (8 µg/ml). These data demonstrated that Cbf‐K₁₆ and Cbf‐A₇A₁₃ possess effective antimicrobial activity against drug‐resistant strains, including NDM‐1 carrying E. coli BL21 (DE3)‐NDM‐1, by binding to DNA after penetrating the cytoplasmic membrane in vitro, which may have potential therapeutic value for the treatment of NDM‐1‐carrying bacterial infections. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Synthesis,conformation, and membrane modifying properties of the trikoningin KB lipopeptaibols: Effect of hydrophobicity and chirality in position 1

Summary We synthesized by solution-phase methods the naturally occurring, 10-amino acid residue lipopeptaibol antibiotics trikoningins KBI and KBII, and the [l-Iva¹] KB analogue, in which the amino acid in position 1 is different, with the aim at investigating the effect of hydrophobicity and chirality in that position. A solution conformational analysis, using FT-IR absorption and CD techniques, indicated that all of the three decapeptides are predominantly helical in a membrane-mimetic environment. Permeability measurements showed an increase of the activity from the [Aib¹] peptide to the more hydrophobic [Iva¹] peptides. Conversely, the effect of a change in chirality, obtained by replacingd-Iva¹ withl-Iva, turned out to be of minor significance.     

Measuring the Binding Stoichiometry of HIV-1 Gag to Very-Low-Density Oligonucleotide Surfaces Using Surface Plasmon Resonance Spectroscopy

The interaction of the HIV Gag polyprotein with nucleic acid is a critical step in the assembly of viral particles. The Gag polyprotein is composed of the matrix (MA), capsid (CA), and nucleocapsid (NC) domains. The NC domain is required for nucleic acid interactions, and the CA domain is required for Gag-Gag interactions. Previously, we have investigated the binding of the NC protein to d(TG)(n) oligonucleotides using surface plasmon resonance (SPR) spectroscopy. We found a single NC protein is able to bind to more than one immobilized oligonucleotide, provided that the oligonucleotides are close enough together. As NC is believed to be the nucleic acid binding domain of Gag, we might expect Gag to show the same complex behavior. We wished to analyze the stoichiometry of Gag binding to oligonucleotides without this complication due to tertiary complex formation. We have therefore analyzed Gag binding to extremely low oligonucleotide density on SPR chips. Such low densities of oligonucleotides are difficult to accurately quantitate. We have determined by Fourier transform ion cyclotron (FTICR) mass spectrometry that four molecules of NC bind to d(TG)(10) (a 20-base oligonucleotide). We developed a method of calibrating low-density surfaces using NC calibration injections. Knowing the maximal response and the stoichiometry of binding, we can precisely determine the amount of oligonucleotide immobilized at these very-low-density surfaces (<1 Response Unit). Using this approach, we have measured the binding of Gag to d(TG)(10). Gag binds to a 20-mer with a stoichiometry of greater than 4. This suggests that once Gag is bound to the immobilized oligonucleotide, additional Gag molecules can bind to this complex.     

Surface properties from the S-layer of Clostridium thermosaccharolyticum D120-70 and Clostridium thermohydrosulfuricum L111-69

Labelling experiments using a positively charged topographical marker for electron microscopy, polycationized ferritin, showed that the S-layers of two closely related clostridia Clostridium thermohydrosulfuricum L111-69 and C. thermosaccharolyticum D120-70 do not exhibit a net negative charge, as usually observed for bacterial cell surfaces. Chemical modification of reactive sites confirmed that amino and carboxyl groups are exposed on the S-layer surface of both strains. Amino-specific, bifunctional agents crosslinked both S-layer lattices. Studies with carbodiimides revealed that only the S-layer surface of C. thermohydrosulfuricum L111-69 had amino and carboxyl groups closely enough aligned to permit electrostatic interactions between the constituent protomers. The regular structure of this S-layer lattice was lost upon converting the carboxyl groups into neutral groups by amidation. Disintegration of both S-layer lattices occurred upon N-acetylation or N-succinylation of the free amino groups. Adhesion experiments showed that in neutral and weakly alkaline environment whole cells of C. thermosaccharolyticum D120-70 exhibited a stronger tendency to bind to charged surfaces than whole cells of C. thermohydrosulfuricum L111-69, but showed a lower tendency to bind to hydrophobic materials.     

The extension peptide of plant ferritin from sea lettuce contributes to shell stability and surface hydrophobicity

Plant ferritins have some unique structural and functional features. Most of these features can be related to the plant-specific "extension peptide" (EP), which exists in the N-terminus of the mature region of a plant ferritin. Recent crystallographic analysis of a plant ferritin revealed the structure of the EP, however, two points remain unclear: (i) whether the structures of well-conserved EP of plant ferritins are common in all plants, and (ii) whether the EP truly contributes to the shell stability of the plant ferritin oligomer. To clarify these matters, we have cloned a green-plant-type ferritin cDNA from a green alga, Ulva pertusa, and investigated its crystal structure. Ulva pertusa ferritin (UpFER) has a plant-ferritin-specific extension peptide composed of 28 amino acid residues. In the crystal structure of UpFER, the EP lay on and interacted with the neighboring threefold symmetry-related subunit. The amino acid residues involved in the interaction were very highly conserved among plant ferritins. The EPs masked the hydrophobic pockets on the ferritin shell surface by lying on them, and this made the ferritin oligomer more hydrophilic. Furthermore, differential scanning calorimetric analysis of the native and its EP-deletion mutant suggested that the EP contributed to the thermal stability of the plant ferritin shell. Thus, the shell stability and surface hydrophobicity of plant ferritin were controlled by the presence or absence of the plant-ferritin-specific EP. This regulation can account for those processes such as shell stability, degradation, and association of plant ferritin, which are significantly related to iron utilization in plants.     

Living organisms and the quantum-mechanical wave properties of matter

Heat capacity data for the solid chemical elements and for 10 amino acids from proteins, and Kopp's Law heat capacity values for the chemical elements, show that proteins have extremely low heat capacities compared to other substances. The type of quantum-mechanical interaction that is highly predominant in low-heat-capacity substances reveals the quantum wave properties of matter. Therefore, the quantum-mechanical wave properties of matter are highly predominant in proteins. Some speculations about the meaning of this result are examined and assessed.     

Surface hydrophobicity and water transport of the toad urinary bladder: Effects of vasopressin

Summary The present study investigated whether the hydrophobic properties (wettability) of the luminal surface of the toad urinary bladder might play a role in modulating water transport across this epithelium. In the absence of vasopressin (ADH), water transport across the tissue was low, while luminal surface hydrophobicity (water contact angle) was relatively high. Following stimulation by ADH, water transport increased and surface hydrophobicity decreased. The addition of indomethacin to inhibit ADH-induced prostaglandin synthesis did not reduce these actions of ADH. In an attempt to alter water transport in this tissue, a liposomal suspension of surface-active phospholipids was administered to the luminal surface. This addition had no detectable influence on the low basal rates of water transport, but blocked the ADH-induced stimulation of water transport. We suggest that surface-active phospholipids on the toad bladder luminal membrane may contribute to the hydrophobic characteristics of this tissue. ADH may act to decrease surface hydrophobicity, facilitating the movement of water molecules across an otherwise impermeable epithelium. This surface alteration may be associated with the appearance of water channels in the apical membrane.     

Occurrence of the genes encoding carbapenemases,ESBLs and class 1 integron-integrase among fermenting and non-fermenting bacteria from retail goat meat

The present study was planned to detect the genes encoding carbapenemases, ESBLs and class 1 integron-integrase among bacteria obtained from retail goat meat. Fermenting and non-fermenting bacterial isolates (n = 57), recovered from 61 goat meat samples, were identified by 16S rRNA gene sequencing. Antimicrobial susceptibility of isolates was tested by the broth dilution method using ceftazidime, cefotaxime, meropenem and imipenem. Plasmids were isolated and tested for their physical characters. Plasmids were subjected to screening of carbapenemase, ESBL and intI1 gene. Conjugation assay was performed using bla_NDM-positive isolates as the donor, and Escherichia coli HB101 as the recipient. Isolates showed the high rates of resistance to ceftazidime (77·2%), cefotaxime (70·2%), meropenem (22·8%) and imipenem (17·5%). They showed variability in number and size (~1 to >20 kb) of plasmids. Among all, 1, 4, 13 and 31 isolates showed the bla_KPC, bla_NDM, bla_SHV and bla_TEM genes, respectively. The bla_KPC-2 gene was observed in one E. coli isolate. The bla_NDM-1 gene was detected in Stenotrophomonas maltophilia (n = 2), Acinetobacter baumannii (n = 1) and Ochrobactrum anthropi (n = 1) isolates. These isolates co-harboured the bla_TEM and bla_SHV genes. The intI1 gene was detected in 22 (38·6%) isolates, and 16 of these isolates showed the carbapenemase and/or ESBL genes. The conjugative movement of bla_NDM gene could not be proved after three repetitive mating experiments. The presence of genes encoding carbapenemases and ESBLs in bacteria from goat meat poses public health risks.     

Substrate specificity and some physicochemical properties of autolytic enzymes of the bacterium <Emphasis Type="Italic">Lysobacter</Emphasis> sp. XL 1

The substrate specificity of autolytic enzymes of the bacterium Lysobacter sp. XL 1 has been established. The periplasmic enzyme A8, the cytosolic enzyme A1, and the enzyme A10 solubilized from the cell walls and membranes with Triton X-100 exhibit glucosaminidase activity; the cytosolic enzyme A4 and the enzyme A9 solubilized from the cell walls and membranes with LiCl exhibit the muramidase activity. The cytosolic enzymes A3 and A6 have N-acetylmuramoyl-L-alanine amidase activity, and the enzyme A5 exhibits the diaminopimelinoyl-alanine endopeptidase activity. Some physicochemical properties of the most active autolytic cytosolic enzymes of Lysobacter sp. XL 1 (endopeptidases A5 and A7 and N-acetylmuramoyl-L-alanine amidase A6) were studied. The enzymes exhibit maximal activity over a wide range of buffer concentrations in weakly alkaline medium and moderate temperatures. The investigated enzymes are comparatively thermolabile proteins.     

Enhanced antibacterial properties and the cellular response of stainless steel through friction stir processing

Biofilm related bacterial infection is one of the primary causes of implant failure. Limiting bacterial adhesion and colonization of pathogenic bacteria is a challenging task in health care. Here, a highly simplistic processing technique for imparting antibacterial properties on a biomedical grade stainless steel is demonstrated. Low-temperature high strain-rate deformation achieved using submerged friction stir processing resulted in a nearly single phase ultra-fine grain structure. The processed stainless steel demonstrated improved antibacterial properties for both Gram-positive and Gram-negative bacteria, significantly impeding biofilm formation during the in vitro study. Also, the processed stainless steel showed better compatibility with human fibroblasts manifested through apparent cell spreading and proliferation. The substantial antibacterial properties of the processed steel are explained in terms of the favorable electronic characteristics of the metal-oxide and by using classical Derjaguin–Landau–Verwey–Overbeek (DLVO) and the extended DLVO (XDLVO) approach at the cell–substrate interface.     

Surface electrical properties of <Emphasis Type="Italic">bacillus subtilis</Emphasis> cells and the effect of interaction with silicon dioxide particles

Depending on the amount of phosphate in the growth medium, the surface layer of the Bacillus subtilis cell wall may be composed mainly of either teichoic or teichuronic acids. Only in the former case the negative charge of the cell surface increases upon addition of silicon dioxide particles into the cell suspension.