Bacterial adhesion measurements on soft contact lenses using a Modified Vortex Device and a Modified Robbins Device

S. marcescens 8100 andP. aeruginosa 15442 were used to study bacterial adhesion to hydrogel contact lenses which had not been worn. Bacterial removal from unworn lens materials was assessed with a calibrated vortex device modified with a digital rpm readout and fitted with a test tube attachment (MVD). The MVD, which relies on a whirlpool-like force to remove the bacteria, showed that bacteria adhered to the same degree to etafilcon A, vifilcon A and polymacon lenses under standardized conditions. Tracking the isoenzyme patterns of these bacterial species over time showed instability ofS. marcescens upon repeated passage. This instability was not evident withP. aeruginosa. Bacterial adhesion ofP. aeruginosa 15442, to human worn and unworn etafilcon A materials was determined with a Modified Robbins Device. The MRD was closed off at both ends stopping medium and bacterial movement after 1 h of fluid flow over the lens surface. The results show that immediately following this 1-h period more bacteria adhere to unworn contact lenses than to worn lenses. However, bacterial counts were equivalent on worn and unworn lenses following 5 h of static incubation.     

Adhesion of Pseudomonas aeruginosa,Achromobacter xylosoxidans,Delftia acidovorans,Stenotrophomonas maltophilia to contact lenses under the influence of an artificial tear solution

Abstract

Corneal infection is a devastating sight-threatening complication that is associated with contact lens (CL) wear, commonly caused by Pseudomonas aeruginosa. Lately, Achromobacter xylosoxidans, Delftia acidovorans, and Stenotrophomonas maltophilia have been associated with corneal infection. This study investigated the adhesion of these emerging pathogens to CLs, under the influence of an artificial tear solution (ATS) containing a variety of components commonly found in human tears. Two different CL materials, etafilcon A and senofilcon A, either soaked in an ATS or phosphate buffered saline, were exposed to the bacteria. Bacterial adhesion was investigated using a radio-labeling technique (total counts) and plate count method (viable counts). The findings from this study revealed that in addition to P. aeruginosa, among the emerging pathogens evaluated, A. xylosoxidans showed an increased propensity for adherence to both CL materials and S. maltophilia showed lower viability. ATS influenced the viable counts more than the total counts on CLs.     

Bacterial colonization of rigid gas permeable and hydrogel contact lenses by Staphylococcus aureus

Staphylococcus aureus ATCC 6538 and a clinical isolate of S. aureus from a bacterial keratitis patient were examined for their ability to adhere to etafilcon A, polymacon, silafocon, and pauflufocon A, B and C contact lenses. Both isolates adhered more to the rigid gas permeable (RGP) materials than to the hydrogel lenses tested (P < 0.05). S. aureus ATCC 6538 adhered to the etafilcon A material to a greater extent than did the clinical isolate (P < 0.05). there were no statistically significant differences in the recovery of staphylococci from unworn lens materials when surface area, composition and ionicity were evaluated for either the hydrogel or rgp lenses tested against lenses of a similar type. however, differences were observed when hydrogel lenses were evaluated against rgp lenses (P < 0.05). these differences may be related to water content. Journal of Industrial Microbiology & Biotechnology (2000) 24, 113–115. Received 12 August 1999/ Accepted in revised form 02 November 1999     

Microbiome precision editing: Using PEG as a selective fermentation initiator against methicillin‐resistant Staphylococcus aureus

Recent creation of a Unified Microbiome Initiative (UMI) has the aim of understanding how microbes interact with each other and with us. When pathogenic Staphylococcus aureus infects the skin, the interplay between S. aureus and skin commensal bacteria occurs. Our previous data revealed that skin commensal bacteria can mediate fermentation against the growth of USA300, a community‐acquired methicillin‐resistant S. aureus MRSA. By using a fermentation process with solid media on a small scale, we define poly(ethylene glycol) dimethacrylate (PEG‐DMA) as a selective fermentation initiator which can specifically intensify the probiotic ability of skin commensal Staphylococcus epidermidis bacteria. At least five short‐chain fatty acids including acetic, butyric and propionic acids with anti‐USA300 activities are produced by PEG‐DMA fermentation of S. epidermidis. Furthermore, the S. epidermidis‐laden PEG‐DMA hydrogels effectively decolonized USA300 in skin wounds in mice. The PEG‐DMA and its derivatives may become novel biomaterials to specifically tailor the human skin microbiome against invading pathogens.     

Development of a prototype wound dressing technology which can detect and report colonization by pathogenic bacteria

A new methodology for detecting the microbiological state of a wound dressing in terms of its colonization with pathogenic bacteria such as Staphylococcus aureus or Pseudomonas aeruginosa has been developed. Here we report how stabilized lipid vesicles containing self-quenched carboxyfluorescein dye are sensitive to lysis only by toxins/virulence factors from P. aeruginosa and S. aureus but not by a non-toxic Escherichia coli species. The development of the stabilized vesicles is discussed and their response to detergent (triton), bacterial toxin (α-hemolysin) and lipases (phospholipase A₂). Finally, fabrics with stabilized vesicles attached via plasma deposited maleic anhydride coupling are shown visibly responding to S. aureus (MSSA 476) and P. aeruginosa (PAO1) but not E. coli DH5α in a prototype dressing.     

Bacterial compounds,as biocontrol agent against early blight (Alternaria solani) and tobacco cut worm (Spodoptera litura Fab.) of tomato (Lycopersicon esculentum Mill.)

Abstract

Tomato, the important vegetable crop faces yield loses by foliar disease, Early Blight caused by Alternaria solani and infestation by tobacco cut worm, Spodoptera litura. Synthetic pesticides used for disease and pest control resulted in environmental pollution as well as development of resistance. By way of a need to develop a new biopesticide, bacteria were tested for their anti-fungal and insecticidal activity. Volatile compounds and peptides from the bacteria, Bacillus subtilis, Staphylococcus aureus and Pseudomonas aeruginosa were able to inhibit the hyphal growth and melanin production of A. solani. S. aureus showed the highest inhibitory effect as well as reduced the disease severity. B. subtilis exhibited the highest mortality rate of 87% and 83% against I and II instar of S. litura, respectively. The bacteria offered effective biocontrol at 35?°C. Bioactive substances from the bacteria can be used as potential biocontrol agents against the pest and disease of tomato.     

Intelligent Bayes Classifier (IBC) for ENT infection classification in hospital environment

Electronic Nose based ENT bacteria identification in hospital environment is a classical and challenging problem of classification. In this paper an electronic nose (e-nose), comprising a hybrid array of 12 tin oxide sensors (SnO₂) and 6 conducting polymer sensors has been used to identify three species of bacteria, Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), and Pseudomonas aeruginosa (P. aeruginosa) responsible for ear nose and throat (ENT) infections when collected as swab sample from infected patients and kept in ISO agar solution in the hospital environment. In the next stage a sub-classification technique has been developed for the classification of two different species of S. aureus, namely Methicillin-Resistant S. aureus (MRSA) and Methicillin Susceptible S. aureus (MSSA). An innovative Intelligent Bayes Classifier (IBC) based on "Baye's theorem" and "maximum probability rule" was developed and investigated for these three main groups of ENT bacteria. Along with the IBC three other supervised classifiers (namely, Multilayer Perceptron (MLP), Probabilistic neural network (PNN), and Radial Basis Function Network (RBFN)) were used to classify the three main bacteria classes. A comparative evaluation of the classifiers was conducted for this application. IBC outperformed MLP, PNN and RBFN. The best results suggest that we are able to identify and classify three bacteria main classes with up to 100% accuracy rate using IBC. We have also achieved 100% classification accuracy for the classification of MRSA and MSSA samples with IBC. We can conclude that this study proves that IBC based e-nose can provide very strong and rapid solution for the identification of ENT infections in hospital environment.     

Modelling antisepsis using defined populations of facultative and anaerobic wound pathogens grown in a basally perfused biofilm model

An in vitro model was developed to assess the effects of topical antimicrobials on taxonomically defined wound biofilms. Biofilms were exposed over seven days to povidone-iodine, silver acetate or polyhexamethylene biguanide (PHMB) at concentrations used in wound dressings. The rank order of tolerance in multi-species biofilms, based on an analysis of the average bacterial counts over time was P. aeruginosa > methicillin-resistant Staphylococcus aureus (MRSA) > B. fragilis > S. pyogenes. The rank order of effectiveness for the antimicrobials in the biofilm model was povidone-iodine > PHMB > silver acetate. None of the test compounds eradicated P. aeruginosa or MRSA from the biofilms although all compounds except silver acetate eliminated S. pyogenes. Antimicrobial effectiveness against bacteria grown in multi-species biofilms did not correlate with planktonic susceptibility. Defined biofilm populations of mixed-species wound pathogens could be maintained in the basal perfusion model, facilitating the efficacy testing of treatments regimens and potential dressings against multi-species biofilms composed of wound isolates.     

Comparison of Macromolecular Oxidation by Reactive Oxygen Species in Three Bacterial Genera Exposed to Different Antibiotics

Proteins and lipids maybe important targets of oxidation and this may alter their functions. We evaluated whether ceftazidima (CAZ), piperacillin (PIP), chloramphenicol (CMP), and ciprofloxacin (CIP) could oxidize the macromolecules in the three bacterial genera Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. There was an increase in lipid peroxidation observed in these three species. However, this was lower in the Gram negative bacteria than in S. aureus. A reduction of the carbonyl residue in S. aureus with ciprofloxacin was observed whereas in Gram negative bacteria the antibiotics increased the carbonyl residue with respect to the control. Although the strains suffered a rise in advanced oxidation protein products (AOPP) in the presence of ciprofloxacin, the S. aureus strain had a smaller increase of AOPP than the other strains. The results described in this article provide data about the susceptibility of the three bacterial genera to the oxidative stress induced by the antibiotics studied.     

Bronchopneumonia and oral health in hospitalized older patients. A pilot study

Aims: To correlate microbial findings obtained by bronchoalveolar lavage in pneumonia patients with the clinical situation of the oral cavity. Method: Quantitative aerobic and anaerobic cultures were carried out in 150 ml samples of bronchoalveolar lavage (BAL) obtained by means of an endoscope (Video Endoscope Pentax®) inserted per as in the infected bronchus. Material: Twenty consecutive patients with a tentative clinical diagnosis of bronchopneumonia in whom BAL was carried out for diagnostic purposes. A clinical evaluation of the oral health status (oral hygiene, caries, periodontal diseases) was subsequently carried out. Results: In seven edentulous subjects wearing complete dentures the culture of anaerobic microorganisms was negative or yielding less than 100 cfu/ml BAL. Two patients yielded high counts of S. aureus and one high counts of P. aeruginosa. In the 13 subjects with natural teeth left one showed high counts of Veillonella spp. (anaerobic)+P. aeruginosa, one high counts of Veillonella spp. +S. aureus, one high counts of P. aeruginosa + S. aureus and one high counts of E. coli. These four subjects showed poor oral hygiene, periodontal pockets and a BAL microflora consistent with periodontal pathology. Conclusion: The results of this pilot study suggest that microorganisms of denture plaque or associated with periodontal diseases may give rise to aspiration pneumonia in susceptible individuals.     

A novel cationic‐peptide coating for the prevention of microbial colonization on contact lenses

Aims: To develop an antimicrobial peptide with broad spectrum activity against bacteria implicated in biomaterial infection of low toxicity to mammalian cells and retaining its antimicrobial activity when covalently bound to a biomaterial surface. Methods and Results: A synthetic peptide (melimine) was produced by combining portions of the antimicrobial cationic peptides mellitin and protamine. In contrast to the parent peptide melittin which lysed sheep red blood cells at >10 μg ml^?1, melimine lysed sheep red blood cells only at concentrations >2500 μg ml^?1, well above bactericidal concentrations. Additionally, melimine was found to be stable to heat sterilization. Evaluation by electron microscopy showed that exposure of both Pseudomonas aeruginosa and Staphylococcus aureus to melimine at the minimal inhibitory concentration (MIC) produced changes in the structure of the bacterial membranes. Further, repeated passage of these bacteria in sub‐MIC concentrations of melimine did not result in an increase in the MIC. Melimine was tested for its ability to reduce bacterial adhesion to contact lenses when adsorbed or covalently attached. Approximately 80% reduction in viable bacteria was seen against both P. aeruginosa and S. aureus for 500 μg per lens adsorbed melimine. Covalently linked melimine (18 ± 4 μg per lens) showed >70% reduction of these bacteria to the lens. Conclusions: We have designed and tested a synthetic peptide melimine incorporating active regions of protamine and mellitin which may represent a good candidate for development as an antimicrobial coating for biomaterials. Significance and Impact of the Study: Infection associated with the use of biomaterials remains a major barrier to the long‐term use of medical devices. The antimicrobial peptide melimine is an excellent candidate for development as an antimicrobial coating for such devices.     

Synthesis of a novel class of some 1,3,4-oxadiazole derivatives as antimicrobial agents

In an effort to discover new candidates with improved antimicrobial activities we report here the synthesis and in vitro biological evaluation of various series of 2-{(3,4,5-trimethoxy phenyl-1,3,4-oxadiazolyl)-5-thio}-4-(morpholino)-6-(phenyl ureido)-s-triazine (7a-i) and 2-{(3,4,5-trimethoxy phenyl-1,3,4-oxadiazolyl)-5-thio}-4-(morpholino)-6-(phenyl thioureido)-s-triazine (8a-g). Antimicrobial properties of the title compounds were investigated against two Gram ( + ve) bacteria (S. aureus, B. subtilis), two Gram ( ? ve) bacteria (P. aeruginosa, E. coli) and yeast-like fungi (C. albicans) using the broth microdilution method.     

The ionic liquid 1-alkyl-3-methylimidazolium demonstrates comparable antimicrobial and antibiofilm behavior to a cationic surfactant

Biofilms are problematic in health and industry because they are resistant to various antimicrobial treatments. Ionic liquids are a novel class of low temperature liquid salts consisting of discrete anions and cations, and have attracted considerable interest as safer alternatives to organic solvents. Ionic liquids have interesting antimicrobial properties and some could find use in the development of novel antiseptics, biocides and antifouling agents. The antimicrobial and antibiofilm activity of 1-dodecyl-3-methylimiazolium iodide ([C₁₂MIM]I) was studied using the clinically important bacterial pathogens, Staphylococcus aureus SAV329 and Pseudomonas aeruginosa PAO1. The ionic liquid increased cell membrane permeability in both S. aureus and P. aeruginosa cells and impaired their growth, attachment and biofilm development. The ionic liquid exhibited superior antimicrobial and antibiofilm activity against the Gram-positive S. aureus compared to the Gram-negative P. aeruginosa cells. BacLight? staining and confocal microscope imaging confirmed that the ionic liquid treatment increased the cell membrane permeability of both the Gram-positive and Gram-negative bacteria. In addition, the antimicrobial and antibiofilm properties of [C₁₂MIM]I were similar or superior to those of cetyltrimethylammonium bromide (CTAB), a well-known cationic surfactant. It is concluded that the ionic liquid induced damage to bacterial cells by disrupting cell membrane, leading to inhibition of growth and biofilm formation. Overall, the results indicate that the ionic liquid 1-dodecyl-3-methylimiazolium iodide was effective in preventing S. aureus and P. aeruginosa biofilms and could have applications in the control of bacterial biofilms.     

Synthesis,molecular docking,binding free energy calculation and molecular dynamics simulation studies of benzothiazol-2-ylcarbamodithioates as Staphylococcus aureus MurD inhibitors

Abstract

A new series of benzothiazol-2-ylcarbamodithioate functional compounds 5a-f has been designed, synthesized and characterized by spectral data. These compounds were screened for their in vitro antibacterial activity against strains of Staphylococcus aureus (NCIM 5021, NCIM 5022 and methicillin-resistant isolate 43300), Bacillus subtilis (NCIM 2545), Escherichia coli (NCIM 2567), Klebsiella pneumoniae (NCIM 2706) and Psudomonas aeruginosa (NCIM 2036). Compounds 5a and 5d exhibited significant activity against all the tested bacterial strains. Specifically, compounds 5a and 5d showed potent activity against K. pneumoniae (NCIM 2706), while compound 5a also displayed potent activity against S. aureus (NCIM 5021). Compound 5d showed minimum IC₅₀ value of 13.37?μM against S. aureus MurD enzyme. Further, the binding interactions of compounds 5a-f in the catalytic pocket have been investigated using the extra-precision molecular docking and binding free energy calculation by MM-GBSA approach. A 30?ns molecular dynamics simulation of 5d/modeled S. aureus MurD enzyme was performed to determine the stability of the predicted binding conformation.     

Resistance to cadmium salts and metal absorption by different microbial species

The present study evaluates the inhibitory activity and the absorption of cadmium (Cd) salts by different microbial species, including Gram-positive (Staphylococcus aureus andS. epidermidis), Gram-negative (Pseudomonas aeruginosa, Escherichia coli, Salmonella typhimurium, andProteus mirabilis) bacteria and one yeast (Candida albicans). The metal absorption by growing cells was considered both in liquid and in solid medium. For one strain ofP. aeruginosa the presence of Cd deposits inside the cell was investigated by transmission electron microscopy. Generally, the Gram-negative species tested proved to be highly resistant to Cd ions and accumulated great amounts of Cd during growth. Two strains ofP. aeruginosa showed a high degree of resistance to Cd and were particularly efficient in removing the metal from solutions. The Gram-positive bacteria showed a heterogeneous behavior: anS. aureus strain susceptible to Cd absorbed, at low metal concentrations, higher amounts of metal than a Cd-resistant one. The metal absorption for Gram-negative species was dose dependent, while for the Cd-resistant staphylococci it reached a plateau. Our results suggest that microorganisms can represent a good model to study the interactions between heavy metals and living organisms.     

Inactivation and injury of Escherichia coli O157:H7 and Staphylococcus aureus by pulsed electric fields

Two pathogenic microorganisms Escherichia coli O157:H7 and Staphylococcus aureus, suspended in peptone solution (0.1% w/v) were treated with 12, 14, 16 and 20 kV/cm electric field strengths with different pulse numbers up to 60 pulses. Pulsed electric field (PEF) treatment at 20 kV/cm with 60 pulses provided nearly 2 log reduction in viable cell counts of E. coli O157:H7 and S. aureus. S. aureus cells were slightly more resistant than E.coli O157:H7 cells. The results related to the effect of initial cell concentration of E. coli O157:H7 on the PEF inactivation showed that more inactivation was obtained by decreasing initial cell concentration. Any possible injury by PEF was also investigated after applying 20 kV/cm electric field to the microorganisms. As a result, it was determined that there was 35.92 to 43.36% injury in E. coli O157:H7 cells, and 17.26 to 30.86% injury in S. aureus cells depending on pulse number. The inactivation results were also described by a kinetic model.     

Alteration of protein homeostasis mediates the interaction of Pseudomonas aeruginosa with Staphylococcus aureus

Intracellular protein degradation is essential for the survival of all organisms, but its role in interspecies interaction is unknown. Here, we show that the ClpXP protease of Pseudomonas aeruginosa suppresses its antimicrobial activity against Staphylococcus aureus, a common pathogen co-isolated with P. aeruginosa from polymicrobial human infections. Using proteomic, biochemical, and molecular genetic approaches, we found that this effect is due to the inhibitory effects of ClpXP on the quorum sensing (QS) of P. aeruginosa, mainly by degrading proteins (e.g., PhnA, PhnB, PqsR, and RhlI) which are critical for the production of QS signal molecules PQS and C4-HSL. We provide evidence that co-culturing with S. aureus induces a decrease in the activity of ClpXP in P. aeruginosa, an effect which was also achieved by the treatment of P. aeruginosa with N-acetylglucosamine (GlcNAc), a widespread chemical present on the surface of diverse cell types from bacteria to humans. These findings extend the range of biological events governed by proteolytic machinery to microbial community structure, thus also suggesting that a chemical-induced alteration of protein homeostasis is a mechanism for interspecies interactions.     

<Emphasis Type="Italic">Staphylococcus aureus</Emphasis> sigma B-dependent emergence of small-colony variants and biofilm production following exposure to <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> 4-hydroxy-2-heptylquinoline-<Emphasis Type="Italic">N-</Emphasis> oxide

Background  

Staphylococcus aureus and Pseudomonas aeruginosa are often found together in the airways of cystic fibrosis (CF) patients. It was previously shown that the P. aeruginosa exoproduct 4-hydroxy-2-heptylquinoline-N-oxide (HQNO) suppresses the growth of S. aureus and provokes the emergence of small-colony variants (SCVs). The presence of S. aureus SCVs as well as biofilms have both been associated with chronic infections in CF.     

Influence of Heterologously Expressed azurin from Pseudomonas aeruginosa on the Adhesion and Invasion of Pathogenic Bacteria to the Caco-2 Cell Line

This study proposed to investigate the effect of azurin on the major stages of pathogenesis (adhesion and invasion) of intestinal bacterial pathogens (Salmonella spp. and Escherichia coli) and epithelial pathogens (Staphylococcus aureus and Pseudomonas aeruginosa) on the human colorectal adenocarcinoma (Caco-2) cell line. Azurin protein was produced by cloning the azurin gene into pET21a and heterologous expression in E. coli BL21. The protein was purified using affinity chromatography and confirmed by Western blotting. The purified protein was evaluated by three experiments of adhesion and invasion assays, including exclusion, competition, and replacement. Azurin was observed to significantly inhibit the attachment and invasion of S. aureus, Salmonella spp., and E. coli, while no such inhibitory effects were observed on P. aeruginosa. In fact, the protein increased the adhesion of P. aeruginosa to the cell. In conclusion, our study proposes that azurin is a potential prophylactic or preventive helper candidate to inhibit the attachment and invasion of pathogenic bacteria to host cells and reduce the progression of the infection process. Our study also reveals the involvement of azurin in bacteria-host cell interactions, providing novel and important insights toward the elucidation of its biological function in this field. Thus, this study provides new opportunities to use azurin as an adjunct therapy against critical stages of infection by a wide range of pathogenic bacteria.