Impact of reduced vancomycin susceptibility on the therapeutic outcome of MRSA bloodstream infections

Background

Resistance to anti-tuberculosis drugs is a serious public health problem. Multi-drug resistant tuberculosis (MDR-TB), defined as resistance to at least rifampicin and isoniazid, has been reported in all regions of the world. Current phenotypic methods of assessing drug susceptibility of M. tuberculosis are slow. Rapid molecular methods to detect resistance to rifampicin have been developed but they are not affordable in some high prevalence countries such as those in sub Saharan Africa. A simple multi-well plate assay using mycobacteriophage D29 has been developed to test M. tuberculosis isolates for resistance to rifampicin. The purpose of this study was to investigate the performance of this technology in Kampala, Uganda.

Methods

In a blinded study 149 M. tuberculosis isolates were tested for resistance to rifampicin by the phage assay and results compared to those from routine phenotypic testing in BACTEC 460. Three concentrations of drug were used 2, 4 and 10 μg/ml. Isolates found resistant by either assay were subjected to sequence analysis of a 81 bp fragment of the rpoB gene to identify mutations predictive of resistance. Four isolates with discrepant phage and BACTEC results were tested in a second phenotypic assay to determine minimal inhibitory concentrations.

Results

Initial analysis suggested a sensitivity and specificity of 100% and 96.5% respectively for the phage assay used at 4 and 10 μg/ml when compared to the BACTEC 460. However, further analysis revealed 4 false negative results from the BACTEC 460 and the phage assay proved the more sensitive and specific of the two tests. Of the 39 isolates found resistant by the phage assay 38 (97.4%) were found to have mutations predictive of resistance in the 81 bp region of the rpoB gene. When used at 2 μg/ml false resistant results were observed from the phage assay. The cost of reagents for testing each isolate was estimated to be 1.3US$ when testing a batch of 20 isolates on a single 96 well plate. Results were obtained in 48 hours.

Conclusion

The phage assay can be used for screening of isolates for resistance to rifampicin, with high sensitivity and specificity in Uganda. The test may be useful in poorly resourced laboratories as a rapid screen to differentiate between rifampicin susceptible and potential MDR-TB cases.     

Immobilization of galactose ligands on acrylic acid graft-copolymerized poly(ethylene terephthalate) film and its application to hepatocyte culture

Surface modification of argon-plasma-pretreated poly(ethylene terephthalate) (PET) films via UV-induced graft copolymerization with acrylic acid (AAc) was carried out. Galactosylated surfaces were then obtained by coupling a galactose derivative (1-O-(6'-aminohexyl)-D-galactopyranoside) to the AAc graft chains with the aid of a water-soluble carbodiimide (WSC) and N-hydroxysulfosuccinimide (sulfo-NHS). The modified PET films were characterized by X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), and water contact-angle measurements. The galactosylated PET films were used as substrates for hepatocyte culture. The effects of surface carboxyl group concentration on the extent of galactose ligand immobilization, the extent of hepatocyte attachment, and the surface morphology were investigated. The amount of the galactose ligands immobilized on the PET surface increased with the AAc polymer graft concentration. AFM images revealed that the surface roughness of the PET film increased after graft copolymerization with AAc, but did not change appreciably with the subsequent immobilization of the galactose ligands. At the surface carboxyl group concentration of about 0.56 micromol/cm(2) or galactose ligand concentration of about 0.51 micromol/cm(2), the hepatocyte culture on the galactosylated surface exhibited the optimum concentration and physiological functions and formed aggregates or spheroids after just 1 day of culture. The albumin and urea synthesis functions of these hepatocytes were comparable to or higher than those of the hepatocytes cultured on the collagen-modified PET substrates.     

Heparin-coupled poly(poly(ethylene glycol) monomethacrylate)-Si(111) hybrids and their blood compatible surfaces

Well-defined (nearly monodispersed) poly(poly(ethylene glycol)monomethacrylate)-Si hybrids were prepared via surface-initiated atom transfer radical polymerization (ATRP) of the poly(ethylene glycol)monomethacrylate (PEGMA) macromonomer on the hydrogen-terminated Si(111) surface (Si-H surface). Both the active chloride groups at the chain ends (from the ATRP process) and the chloride groups converted from some ( approximately 32%) of the -OH groups of the Si-C bonded PEGMA polymer, or P(PEGMA), brushes were used as leaving groups for the covalent coupling of heparin. For the heparinized P(PEGMA)-Si hybrid surfaces, protein adsorption and platelet adhesion were significantly suppressed. The well-defined and dense P(PEGMA) brushes, prepared from surface-initiated ATRP, had allowed the immobilization of a relatively high concentration of heparin (about 14 mug/cm(2)). The resulting silicon surface exhibited significantly improved antithrombogenecity with a plasma recalcification time (PRT) of about 150 min. The persistence of high bioactivity for the immobilized heparin on the hybrid surfaces can be attributed to the biocompatibility of the PEGMA units, as well as their role as spacers in providing the immobilized heparin with a higher degree of conformational freedom in a more hydrophilic environment. Thus, the heparin-coupled P(PEGMA)-Si hybrids with anti-fouling and antithrombogenic surfaces are potentially useful in silicon-based implantable devices and tissue engineering.     

Surface-active and stimuli-responsive polymer--Si(100) hybrids from surface-initiated atom transfer radical polymerization for control of cell adhesion

A simple two-step method was developed for the covalent immobilization of atom-transfer radical polymerization (ATRP) initiators on the hydrogen-terminated Si(100) (Si-H) surface. Well-defined functional polymer-Si hybrids, consisting of covalently tethered brushes of poly(ethylene glycol) monomethacrylate (PEGMA) polymer, N-isopropylacrylamide (NIPAAm) polymer, and NIPAAm-PEGMA copolymers and block copolymers on Si-H surfaces, were prepared via surface-initiated ATRP. Kinetics study revealed that the chain growth from the silicon surface was consistent with a "controlled" process. Surface cultures of the cell line 3T3-Swiss albino on the hybrids were evaluated. The PEGMA graft-polymerized silicon [Si-g-P(PEGMA)] surface is very effective in preventing cell attachment and growth. At 37 degrees C [above the lower critical solution temperature (LCST, approximately 32 degrees C) of NIPAAm], the seeded cells adhered, spread, and proliferated on the NIPAAm graft polymerized silicon [Si-g-P(NIPAAm)] surface. Below the LCST, the cells detached from the Si-g-P(NIPAAm) surface spontaneously. Incorporation of PEGMA units into the NIPAAm chains of the Si-g-P(NIPAAm) surface via copolymerization resulted in more rapid cell detachment during the temperature transition. The "active" chain ends on the Si-g-P(PEGMA) and Si-g-P(NIPAAm) hybrids were also used as the macroinitiators for the synthesis of diblock copolymer brushes. Thus, not only are the hybrids potentially useful as stimuli-responsive adhesion modifiers for cells in silicon-based biomedical microdevices but also the active chain ends on the hybrid surfaces offer opportunities for further surface functionalization and molecular design.     

Assessment of in vitro bioactivity of hyaluronic acid and sulfated hyaluronic acid functionalized electroactive polymer

Electrically conductive polypyrrole (PPY) was surface functionalized with hyaluronic acid (HA) and sulfated hyaluronic acid (SHA) to improve its surface biocompatibility. The immobilization of HA on the PPY film was facilitated by the use of a cross-linker having the appropriate functional groups. The biological activity of the HA functionalized PPY film was assessed by means of an in vitro PC12 cell culture. The cell attachment on different substrates was studied and determined by bicinchoninic acid protein analysis. Cell attachment on the HA functionalized PPY film surface was significantly enhanced in the presence of nerve growth factor. The SHA functionalized PPY film was obtained by the sulfonation of the immobilized HA using pyridinesulfonate. The retention of the biological activity of the immobilized HA after sulfonation was evaluated by the in vitro assessment of the plasma recalcification time (PRT) and platelet adhesion on the substrate. The PRT observed from the SHA functionalized PPY film was significantly prolonged compared with the HA functionalized PPY. Some reduction of platelet adhesion was observed for the SHA functionalized PPY film, compared with that of the HA functionalized PPY film.     

Differential Metabolite Profiles during Fruit Development in High-Yielding Oil Palm Mesocarp

To better understand lipid biosynthesis in oil palm mesocarp, in particular the differences in gene regulation leading to and including de novo fatty acid biosynthesis, a multi-platform metabolomics technology was used to profile mesocarp metabolites during six critical stages of fruit development in comparatively high- and low-yielding oil palm populations. Significantly higher amino acid levels preceding lipid biosynthesis and nucleosides during lipid biosynthesis were observed in a higher yielding commercial palm population. Levels of metabolites involved in glycolysis revealed interesting divergence of flux towards glycerol-3-phosphate, while carbon utilization differences in the TCA cycle were proven by an increase in malic acid/citric acid ratio. Apart from insights into the regulation of enhanced lipid production in oil palm, these results provide potentially useful metabolite yield markers and genes of interest for use in breeding programmes.     

Analyzing the Secretome of Gut Microbiota as the Next Strategy For Early Detection of Colorectal Cancer

Dysbiosis of gut microbiome can contribute to inflammation, and subsequently initiation and progression of colorectal cancer (CRC). Throughout these stages, various proteins and metabolites are secreted to the external environment by microorganisms or the hosts themselves. Studying these proteins may help enhance our understanding of the host–microorganism relationship or they may even serve as useful biomarkers for CRC. However, secretomic studies of gut microbiome of CRC patients, until now, are scarcely performed. In this review article, the focus is on the roles of gut microbiome in CRC, the current findings on CRC secretome are highlighted, and the emerging challenges and strategies to drive forward this area of research are addressed.     

A High Resolution Computer Tomography Scoring System to Predict Culture-Positive Pulmonary Tuberculosis in the Emergency Department

This study evaluated the use of high-resolution computed tomography (HRCT) to predict the presence of culture-positive pulmonary tuberculosis (PTB) in adult patients with pulmonary lesions in the emergency department (ED). The study included a derivation phase and validation phase with a total of 8,245 patients with pulmonary disease. There were 132 patients with culture-positive PTB in the derivation phase and 147 patients with culture-positive PTB in the validation phase. Imaging evaluation of pulmonary lesions included morphology and segmental distribution. The post-test probability ratios between both phases in three prevalence areas were analyzed. In the derivation phase, a multivariate analysis model identified cavitation, consolidation, and clusters/nodules in right or left upper lobe (except anterior segment) and consolidation of the superior segment of the right or left lower lobe as independent positive factors for culture-positive PTB, while consolidation of the right or left lower lobe (except superior segment) were independent negative factors. An ideal cutoff point based on the receiver operating characteristic (ROC) curve analysis was obtained at a score of 1. The sensitivity, specificity, positivity predictive value, and negative predictive value from derivation phase were 98.5% (130/132), 99.7% (3997/4008), 92.2% (130/141), and 99.9% (3997/3999). Based on the predicted positive likelihood ratio value of 328.33 in derivation phase, the post-test probability was observed to be 91.5% in the derivation phase, 92.5% in the validation phase, 94.5% in a high TB prevalence area, 91.0% in a moderate prevalence area, and 76.8% in moderate-to-low prevalence area. Our model using HRCT, which is feasible to perform in the ED, can promptly diagnose culture-positive PTB in moderate and moderate-to-low prevalence areas.     

Layer-by-Layer Click Deposition of Functional Polymer Coatings for Combating Marine Biofouling

"Click" chemistry-enabled layer-by-layer (LBL) deposition of multilayer functional polymer coatings provides an alternative approach to combating biofouling. Fouling-resistant azido-functionalized poly(ethylene glycol) methyl ether methacrylate-based polymer chains (azido-poly(PEGMA)) and antimicrobial alkynyl-functionalized 2-(methacryloyloxy)ethyl trimethyl ammonium chloride-based polymer chains (alkynyl-poly(META)) were click-assembled layer-by-layer via alkyne-azide 1,3-dipolar cycloaddition. The polymer multilayer coatings are resistant to bacterial adhesion and are bactericidal to marine Gram-negative Pseudomonas sp. NCIMB 2021 bacteria. Settlement of barnacle ( Amphibalanus (= Balanus ) amphitrite ) cyprids is greatly reduced on the multilayer polymer-functionalized substrates. As the number of the polymer layers increases, efficacy against bacterial fouling and settlement of barnacle cyprids increases. The LBL-functionalized surfaces exhibit low toxicity toward the barnacle cyprids and are stable upon prolonged exposure to seawater. LBL click deposition is thus an effective and potentially environmentally benign way to prepare antifouling coatings.