Residual risk of Pseudomonas aeruginosa waterborne contamination in an intensive care unit despite the presence of filters at all water points-of-use.

OBJECTIVE: To assess the residual risk of waterborne contamination by Pseudomonas aeruginosa from a water network colonized by a single genotype [sequence type (ST) 299] despite the presence of antimicrobial filters in a medical intensive care unit (ICU). METHODS: During the first 19-month period since the ICU opened, contamination of the water network was assessed monthly by collecting water upstream of the filters. Downstream water was also sampled to assess the efficiency of the filters. P. aeruginosa isolates from patients were collected and compared with the waterborne ST299 P. aeruginosa by multiplex-rep polymerase chain reaction (PCR), pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing. Cross-transmission events by other genotypes of P. aeruginosa were also assessed. RESULTS: Overall, 1.3% of 449 samples of filtered water were positive for P. aeruginosa in inoculum, varying between 1 and 104 colony-forming units/100 mL according to the tap. All P. aeruginosa hydric isolates belonged to ST299 and displayed fewer than two single nucleotide polymorphisms (SNPs). Among 278 clinical isolates from 122 patients, 10 isolates in five patients showed identical profiles to the hydric ST299 clone on both multiplex-rep PCR and PFGE, and differed by an average of fewer than five SNPs, confirming the water network reservoir as the source of contamination by P. aeruginosa for 4.09% of patients. Cross-transmission events by other genotypes of P. aeruginosa were responsible for the contamination of 1.75% of patients. DISCUSSION/CONCLUSION: Antimicrobial filters are not sufficient to protect patients from waterborne pathogens when the water network is highly contaminated. A microbiological survey of filtered water may be needed in units hosting patients at risk of P. aeruginosa infections, even when all water points-of-use are fitted with filters.

Thermal, mechanical and biodegradation studies of biofiller based poly-3-hydroxybutyrate biocomposites.

Biodegradable poly-3-hydroxybutyrate [P(3HB)] and natural fillers - clay, peat, and birch wood flour - were used to prepare powdered composites to form pellets and granules. Pellets were produced by cold pressing of polymer and filler powder whereas granules were produced from the powders wetted with ethanol. Characterization techniques like IR spectroscopy, differential scanning calorimetry, X-ray analysis, mechanical analysis and electron microscopy were employed to study the properties of the initial P(3HB) and fillers and the composites. Analysis of the IR spectra of the composites showed the absence of chemical bonds between the components, i.e. the composites were physical mixtures. Young's moduli of the pellets prepared from initial materials varied considerably, and the highest value was obtained for P(3HB) pellets (350 MPa). Studies of biodegradation of composite pellets and granules in the soil for 35 days showed that the residual mass of the pellets had decreased to 68% for P(3HB); 56.4% for P(3HB)/peat; 67% for P(3HB)/wood flour, and 64% for P(3HB)/clay; granules exhibited a similar mass loss, residual mass of the granules of P(3HB) was 68.4%, P(3HB)/peat 46.4%; P(3HB)/wood flour 77%, and P(3HB)/clay 74%. This shows the significance of the material as an eco-friendly composite without sacrificing its mechanical properties.

Tracking the spread routes of opportunistic premise plumbing pathogens in a haematology unit with water points-of-use protected by antimicrobial filters.

BACKGROUND: Water networks in hospitals are frequently contaminated by opportunistic premise plumbing pathogens (OPPPs) leading to installation of antimicrobial filters on water points-of-use (POU) in order to limit patients' exposure. AIM: To assess the spread of OPPPs through secondary water routes (outside the plumbing system) in an adult haematology unit in which 52 out of 73 water POU were high risk for patients and protected by antimicrobial filters. METHODS: An observational audit identified six secondary water routes for which bacteria tracking and typing were performed in 315 surface samplings. Bacterial isolates were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and compared to the infra-species level by multiplex repetitive element sequence-based polymerase chain reaction and/or by restriction fragment length polymorphism in pulse-field gel electrophoresis. FINDINGS: Pseudomonas aeruginosa and Stenotrophomonas maltophilia, as well as non-pathogenic OPPP indicators, were detected in water collected upstream of antimicrobial filters. P. aeruginosa was the sole OPPP retrieved from tested surfaces (5.1%). The same clone of P. aeruginosa spread from water source to dry surfaces in the same room and cross-contaminated two sinks in different rooms. Three clones of non-pathogenic OPPP indicators spread more widely in different rooms. CONCLUSION: A strategy based on filtration of most (but not all) water POU in a haematology unit could be sufficient to limit the spread of OPPPs to the environment, provided a functional mapping of 'high-risk' POU has been undertaken. The residual spread of OPPPs and OPPP indicators linked to non-filtered water POU argues for careful monitoring of non-filtered water use.

Structural basis for the binding affinity of a homologous series of synthetic phenoxazone drugs with DNA: NMR and molecular mechanics analysis.

The molecular basis of the marked structure-activity relationship for a homologous series of DNA-binding phenoxazone drugs (ActII-ActIV) has been investigated by NMR spectroscopy and molecular mechanics. The spatial structures of the complexes between the drugs and a model deoxytetranucleotide, 5'-d(TpGpCpA), have been determined by molecular mechanics methods using homonuclear (1)H-(1)H 2D-NOESY and heteronuclear (1)H-(31)P (HMBC) NMR spectroscopic data. Observed intermolecular NOE contacts and equilibrium binding studies confirm that the binding affinity of the synthetic phenoxazone derivatives with d(TGCA) decreases with an increase in the number of CH(2) groups in the dimethylaminoalkyl side chains, i.e., ActII > ActIII > ActIV, in agreement with the observed biological activity of these compounds. Molecular mechanics calculations of the spatial structures of the intercalated complexes of ActII-ActIV with d(TGCA) indicate that the different binding constants of the phenoxazone derivatives with the DNA oligomer are due to the different degrees of intercalation of the chromophore and the different steric arrangements of aminoalkyl side chains in the minor groove of the tetramer duplex; this results in different distances between the negatively-charged phosphates of the DNA duplex and the terminal positively-charged N(CH(3))(2) groups of the side chains.

Spectrophotometric investigation of the hetero-association of caffeine and thiazine dye in aqueous solution.

The self-association of thiazine dye, Methylene Blue (MB), and its hetero-association with Caffeine (CAF), were studied in aqueous solution by means of spectrophotometry in the visible range of spectrum. Concentration and temperature dependences of molar absorption of the interacting molecules were used to analyse dynamic equilibrium in solution in terms of two-component model of molecular hetero-association. The magnitudes of equilibrium dimerization and hetero-association constants as well as thermodynamic parameters, enthalpy and entropy, were determined. The calculation of the fraction of different types of associates in the mixed solution, containing Methylene Blue and Caffeine, was done. It was concluded that the hetero-association of Methylene Blue and Caffeine molecules results in lower effective concentration of the dye in solution, which may account for the alteration of its biological activity.

NMR investigation of the complexation of daunomycin with deoxytetranucleotides of different base sequence in aqueous solution.

500 MHz NMR spectroscopy has been used to investigate the complexation of the anthracycline antibiotic daunomycin (DAU) with self-complementary deoxytetranucleotides, 5'-d(CGCG), 5'-d(GCGC), 5'-d(TGCA), 5'-d(ACGT) and 5'-d(AGCT), of different base sequence in aqueous salt solution. 2D homonuclear 1H NMR spectroscopy (TOCSY and NOESY) and heteronuclear 1H - 31P NMR spectroscopy (HMBC) have been used for complete assignment of the non-exchangeable protons and the phosphorus resonance signals, respectively, and for a qualitative determination of the preferred binding sites of the drug. Analysis shows that DAU intercalates preferentially into the terminal sites of each of the tetranucleotides and that the aminosugar of the antibiotic is situated in the minor groove of the tetramer duplex, partly eclipsing the third base pair. A quantitative determination of the complexation of DAU with the deoxytetranucleotides has been made using the experimental concentration and temperature dependences of the drug proton chemical shifts; these have been analysed in terms of the equilibrium reaction constants, limiting proton chemical shifts and thermodynamical parameters (enthalpies deltaH, entropies deltaS) of different drug-DNA complexes (1:1, 1:2, 2:1, 2:2) in aqueous solution. It is found that DAU interacts with sites containing three adjacent base pairs but does not show any significant sequence specificity of binding with either single or double-stranded tetranucleotides, in contrast with other intercalating drugs such as proflavine, ethidium bromide and actinomycin D. The most favourable structures of the 1:2 complexes have been derived from the induced limiting proton chemical shifts of the drug in the intercalated complexes with the tetranucleotide duplex, in conjunction with 2D NOE data. It has been found that the conformational parameters of the double helix and the orientation of the DAU chromophore in the intercalated complexes depend on base sequence at the binding site of the tetramer duplexes in aqueous solution.

1H-NMR determination of the thermodynamics of drug complexation with single-stranded and double-stranded oligonucleotides in solution: ethidium bromide complexation with the deoxytetranucleotides 5'-d(ApCpGpT), 5'-d(ApGpCpT), and 5'-d(TpGpCpA).

The thermodynamical parameters (free energy, enthalpy, and entropy) of complex formation between ethidium bromide and single-stranded and double-stranded tetranucleotides of different base sequence [5-d(TpGpCpA), 5-d(ApCpGpT), and 5-d(ApGpCpT) have been determined from the temperature dependencies of 500 MHz proton nmr chemical shifts. The analysis enables the contributions to be differentiated for the formation of different types of complexes (1:1, 2:1, 1.2 and 2:2) in aqueous solution. The results have been interpreted in terms of the main types of intermolecular interactions responsible for formation of the different complexes; van der Waals and electrostatic interactions are important for formation of complexes of ethidium bromide with single-stranded tetranucleotides, whereas van der Waals and hydrophobic interactions play a significant role in the binding of the dye to the tetramer duplexes.