Rechargeable antimicrobial surface modification of polyethylene.

Polyethylene films were surface modified, to incorporate amine and amide functionalities, and subsequently were evaluated for their ability to recharge the antimicrobial N-halamine structures after contact with sodium hypochlorite, a common food-approved sanitizer. Surfaces were tested for chlorine retention and release, as well as antimicrobial activity against microorganisms relevant to food quality and food safety, including Escherichia coli K-12, Pseudomonas fluorescens, Bacillus cereus, and Listeria monocytogenes. N-Halamine functionalized polyethylene exhibited chlorine rechargeability, maintaining 5 to 7 nmol/cm2 N-halamine structures for six successive charges. The N-halamine functionalized films achieved a 4-log reduction for all organisms tested and maintained a greater than 3-log reduction for four successive uses, suggesting that the modified polyethylene films are capable of providing rechargeable antimicrobial activity. The modified films exhibited antimicrobial activity in aqueous suspensions (P < 0.05) and reduced microbial growth in diluted broth (P < 0.05), suggesting the potential for biocidal action even in the presence of organic matter. Such a rechargeable antimicrobial surface could supplement existing cleaning and sanitation programs in food processing environments to reduce the adhesion, growth, and subsequent cross-contamination of food pathogens, as well as food spoilage organisms.

Expression of a cDNA sequence encoding human purine nucleoside phosphorylase in rodent and human cells.

A cDNA sequence which contains the entire coding region for human purine nucleoside phosphorylase (PNP) was recombined for selection and expression in mammalian cells. Plasmids containing either the simian virus 40 early promoter or the mouse metallothionein promoter positioned just upstream of the PNP coding sequence were constructed. These plasmids also contained the gene for a methotrexate-resistant dihydrofolate reductase, allowing for selection and amplification of positive transferrents after transfection of cells by the DNA-calcium phosphate coprecipitation technique. Expression of human PNP activity was readily detected in both mouse (L) and CHO cells by isoelectric focusing of cell extracts followed by histochemical staining for PNP activity. The simian virus 40 early promoter directed considerable expression of human PNP activity in CHO cells but only scant activity in mouse cells. The mouse metallothionein promoter was not successful in effecting human PNP expression in CHO cells but provided substantial human PNP activity in mouse cells and was inducible by incubation with zinc. HeLa cell transferrents were isolated and screened for the presence of transferred PNP cDNA sequences by Southern hybridization analysis. RNA transcripts derived from the transferred PNP cDNA were identified in one of these cell lines.

Preparation of Biocatalytic Microparticles by Interfacial Self-Assembly of Enzyme-Nanoparticle Conjugates Around a Cross-Linkable Core.

Rational design of hierarchical interfacial assembly of reusable biocatalytic microparticles is described in this chapter. Specifically, purified enzymes and functionalized nanoparticles are electrostatically assembled at the interface of cross-linked microparticles which are formed through ring opening metathesis polymerization. The diameters of microparticle assemblies average 10µm, and they show enhanced kinetic efficiency as well as improved stability against heat, pH, and solvent denaturation when compared to stabilities of the corresponding native enzymes.

Nucleotide sequence of Rattus norvegicus mitochondrial DNA that includes the genes for tRNAile, tRNAgln and tRNAf-met.

The nucleotide sequence of a segment of mtDNA from Rattus norvegicus (rat) which contains the genes for tRNAile, tRNAgln and tRNAf-met has been determined. A detailed comparison has been made between this sequence and the corresponding sequences of mouse, human and bovine mtDNAs with regard to the primary and secondary structure of the tRNA genes, the regions connecting the tRNA genes, and the regions flanking the tRNA genes which code for the carboxyl terminus of URF-1 and the amino terminus of URF-2. No differences were found in the nucleotide sequences of the genes for tRNAile, tRNAgln and tRNAf-met in mtDNAs from three different female lines of rats (SASCO-1, SASCO-2 and Wild-UT) that differ by substitutions of 0.8% to 1.8% of their total nucleotides.

Surface components of Onchocerca volvulus.

A technique employing Sephadex G25 gel filtration has been developed for the rapid isolation and purification of live microfilariae of Onchocerca volvulus from subcutaneous nodules and skin samples. Microfilariae, adult worms and L3 larvae have been surface radiolabelled using the Iodogen technique. Two proteins have been characterised on the surface of uterine microfilariae: these have apparent molecular weights of 14,800 and 15,000. A MW 15,000 protein was the only molecule labelled on the surface of skin microfilariae. Ten proteins were labelled on adult male worms: these have molecular weights of 15,000, 17,500, 20,000, 22,000, 24,000, 29,000, 32,000, 37,000, 42,000, and 50,000. Some, if not all, of these proteins were also identified on female worms. Seven proteins were labelled on the surface of L3 larvae: these have molecular weights of 17,500, 48,000, 50,000, 52,000, 54,000, 57,000, and 105,000. Three of the adult surface proteins were precipitated by selected human infection serum: these are the MW 17,500, 32,000 and 42,000 molecules. The microfilarial surface proteins were not precipitated by human infection serum. The antiserum used in these experiments was shown by Western blot analysis to contain high levels of antibody with specificity for microfilarial and adult antigens. Indirect immunofluorescent assays showed these sera to contain antibody which bound to the surface of adult worms and eggs but not microfilariae. The possibility that skin microfilariae absorb host serum albumin was investigated: Western blot analysis and surface immunofluorescence assays using a specific anti-human albumin serum gave negative results. Fluorescent lectin binding studies revealed the presence of stage-specific carbohydrate moieties exposed on the surface of adult worms and eggs. Microfilariae do not have surface carbohydrate determinants.

Onchocerciasis in Sierra Leone. I. Studies on the prevalence and transmission at Gbaiima village.

A longitudinal survey--parasitological, clinical, immunological and entomological--of onchocerciasis is being conducted in Gbaiima village in Sierra Leone. The estimated Annual Transmission Potential (ATP) is 5863. More than 80% of the annual transmission occurs between October and December. Four species of the Simulium damnosum complex are known to breed in a nearby river. The relative role of these species as vectors has yet to be determined. The total population (598 persons) aged one year and over were examined. Based on microfilarial and nodular rates the prevalence of onchocerciasis was 68.6%. In persons above 15 years of age this prevalence was 88.9%. Microfilarial and nodular rates were related to age. Severe skin lesions occurred in 1.0% of persons. 24 adults (7.5%) were blind (but the cause of the blindness was not determined).

Concentrations of oxygen delivered by air entrainment oxygen masks.

The air entrainment devices from oxygen masks of four manufacturers (Henleys Medical Supplies Ltd, Vickers Medical, Intersurgical Ltd, C R Bard International Ltd) were studied. All were found to deliver concentrations of oxygen close to those specified. The literature is reviewed and it is suggested that provision of a total flow in excess of 60 litres/minute is most likely to provide a constant inspired oxygen concentration, whichever nominal concentration or design of mask is selected and provided distal obstruction is avoided.

Covalent immobilization of lysozyme on ethylene vinyl alcohol films for nonmigrating antimicrobial packaging applications.

The objective of this study was to develop a new antimicrobial film, in which lysozyme was covalently attached onto two different ethylene vinyl alcohol copolymers (EVOH 29 and EVOH 44). The EVOH surface was modified with UV irradiation treatment to generate carboxylic acid groups, and lysozyme was covalently attached to the functionalized polymer surface. Surface characterization of control and modified films was performed using attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and dye assay. The value of protein loading after attachment on the surface was 8.49 µg protein/cm(2) and 5.74 µg protein/cm(2) for EVOH 29 and EVOH 44, respectively, after 10 min UV irradiation and bioconjugation. The efficacy of the EVOH-lysozyme films was assessed using Micrococcus lysodeikticus. The antimicrobial activity of the films was tested against Listeria monocytogenes and was similar to an equivalent amount of free enzyme. The reduction was 1.08 log for EVOH 29-lysozyme, 0.95 log for EVOH 44-lysozyme, and 1.34 log for free lysozyme. This work confirmed the successful use of lysozyme immobilization on the EVOH surface for antimicrobial packaging.

Covalent attachment of lactase to low-density polyethylene films.

Polymer films to which bioactive compounds such as enzymes are covalently attached offer potential for in-package processing of food. Beta-galactosidase (lactase) was covalently attached to surface-functionalized low-density polyethylene films. A two-step wet chemical functionalization introduced 15.7 nmol/cm2 primary amines to the film surface. Contact angle, dye assays, X-ray photoelectron spectroscopy, and appropriate protein assays were used to characterize changes in film surface chemistry after each step in the process of attachment. Glutaraldehyde was used to covalently attach lactase to the surface at a density of 6.0 microg protein per cm2 via reductive amination. The bond between the covalently attached lactase and the functionalized polyethylene withstood heat treatment in the presence of an ionic denaturant with 74% enzyme retention, suggesting that migration of the enzyme into the food product would be unlikely. The resulting polyethylene had an enzyme activity of 0.020 lactase units (LU)/cm2 (approximately 4500 LU/g). These data suggest that enzymes that may have applications in foods can be covalently attached to inert polymer surfaces, retain significant activity, and thus have potential as a nonmigratory active packaging materials.

Postoperative nitrous oxide analgesia. Administration from air entrainment oxygen masks with a primary flow of Entonox.

Nitrous oxide is occasionally used as an analgesic agent in the postoperative period. The feasibility of administering a known concentration from an air entrainment oxygen mask, using a primary flow of Entonox, (50% oxygen/50% nitrous oxide) was investigated. Accurox (C.R. Bard Canada Inc.) blenders, disconnected from their facemasks, were studied using a primary flow of Entonox. An increase in air entrainment of approximately 13% was demonstrated. Concentrations of about 20% nitrous oxide in oxygen enriched air, appropriate for postoperative analgesia, can be produced, but the methods are extravagant in the use of Entonox and are likely to be slightly unreliable.

Pediatric risk of mortality scoring overestimates severity of illness in infants.

OBJECTIVE: To validate Pediatric Risk of Mortality (PRISM) scoring in infants and children admitted for intensive care. DESIGN: Validation cohort. SETTING: A five-bed pediatric ICU and three cots providing intensive care for surgical neonates, within a 159-bed tertiary care children's hospital. PATIENTS: All patients admitted for intensive care during an 18-month period, January 1990 to July 1991. METHODS: Admission (first 24 hrs) PRISM scoring was introduced as a routine procedure. Discretion was allowed in requesting arterial blood gas measurements and clotting studies. All other parameters were intended to be measured on all patients. MEASUREMENTS AND MAIN RESULTS: PRISM scores were obtained on 380 (88%) of 433 patients. Median age was 15 months. A complete PRISM score was obtained in 24% of cases and a score as intended (i.e., allowing discretionary omissions) was obtained in 56% of patients. Comparison of observed and predicted mortality rates using chi square goodness-of-fit tests showed a significantly better observed outcome for all patients (chi 2(5) = 12.04, p < .05). In-depth analysis indicates that the model works well for children (chi 2(5) = 1.80, p > .75), but that observed outcome is significantly better than predicted for infants (chi 2(5) = 17.46, p < .01). Underscoring of children is not the cause of this finding. CONCLUSIONS: In our center, PRISM scoring overestimates severity of illness in infants. PRISM scoring is not institutionally independent and therefore, at present, a comparison between units may not be justified. A reappraisal of the parameter ranges for infants is suggested.

Origin and direction of replication in mitochondrial DNA molecules from the genus Drosophila.

Mitochondrial DNA (mtDNA) obtained from ovaries of Drosophila simulans, D. mauritiana, D. takahashii, D. yakuba and D. virilis was examined by electron microscopy. From a consideration of the structural properties of replicative intermediates, it was concluded that in mtDNA molecules of each species, synthesis on one strand can be up to 97% complete before synthesis on the complementary strand is initiated. MtDNA molecules of each species contain a single A+T-rich region which shows species-specific size variation from 1.0 kb (D. virilis) to 4.8 kb (D. simulans), and maps at the same position in all molecules relative to three common EcoRI sites. The structural properties of complex forms, interpreted as having originated from replicative intermediates, and produced by either partial denaturation or EcoRI digestion, are consistent with the hypothesis that replication is initiated within the A+T-rich region and proceeds unidirectionally around the molecule towards the nearest common EcoRI site. The replication origin is located near the center of the A+T-rich region in D. simulans and D. mauritiana, but lies closer to that end of the A+T-rich region which is distal to the nearest common EcoRI site in D. takahashii, D. yakuba and D. virilis.