A carbon free filter for collection of large volume samples of cellular biomass from oligotrophic waters

Isotopic analysis of cellular biomass has greatly improved our understanding of carbon cycling in the environment. Compound specific radiocarbon analysis (CSRA) of cellular biomass is being increasingly applied in a number of fields. However, it is often difficult to collect sufficient cellular biomass for analysis from oligotrophic waters because easy-to-use filtering methods that are free of carbon contaminants do not exist. The goal of this work was to develop a new column based filter to autonomously collect high volume samples of biomass from oligotrophic waters for CSRA using material that can be baked at 450°C to remove potential organic contaminants. A series of filter materials were tested, including uncoated sand, ferrihydrite-coated sand, goethite-coated sand, aluminum-coated sand, uncoated glass wool, ferrihydrite-coated glass wool, and aluminum-coated glass wool, in the lab with 0.1 and 1.0 μm microspheres and Escherichia coli. Results indicated that aluminum-coated glass wool was the most efficient filter and that the retention capacity of the filter far exceeded the biomass requirements for CSRA. Results from laboratory tests indicate that for oligotrophic waters with 1×10(5) cells ml(-1), 117l of water would need to be filtered to collect 100 μg of PLFA for bulk PLFA analysis and 2000 l for analysis of individual PLFAs. For field sampling, filtration tests on South African mine water indicated that after filtering 5955l, 450 μg of total PLFAs were present, ample biomass for radiocarbon analysis. In summary, we have developed a filter that is easy to use and deploy for collection of biomass for CSRA including total and individual PLFAs.     

Elaboration and application of mathematical model for estimation of mould contamination of some building materials based on ergosterol content determination

The study presents a mathematical function describing a correlation between the amount of ergosterol and the number of colony-forming units (CFU) of mould contaminating selected building materials such as: a block of cellular concrete, gypsum—carton board and gypsum—carton board covered with emulsion paint. The dependence obtained for a particular material as well as an average dependence for all the investigated materials has been described by means of an exponential equation. It has been found out that there is high, statistically significant correlation between ergosterol content and CFU number of mould in all of the building materials. The correlation coefficients have ranged from r=0.790 to 0.933. The elaborated equation describing the above dependence can be applied to estimate mould contamination by means of culture methods within the range 10³–10⁸ CFU/m² of the surface. In addition, the estimated level of ergosterol in these materials has been shown to be the criterion by which to evaluate the degree of filamentous fungal contamination. It has been assessed that an ergosterol content exceeding the level of 3.96 mg/m² indicates the active development of mould. This criterion has been applied to evaluate several building materials i.e.: concrete, gypsum board, emulsion coatings, brick, plaster, wallpaper, glass wool, mineral wool and wood. No statistically significant differences have been observed between CFU number of mould calculated from a model equation on the basis of the ergosterol content and CFU number of mould experimentally determined by traditional methods. The results presented in this paper show that the elaborated equation of correlation between the ergosterol content and CFU number of mould can be applied to estimate mould contamination of different building materials, based on the determination of ergosterol.     

Evaluation of Filters for Removal of Bacteriophages from Air

Glass wool, nonabsorbent cotton, fiberglass filter medium, and a commercial absolute filter were tested for effectiveness in removing aerosolized bacterial viruses under low flow rate (1 ft(3)/min) and high flow rate (10 to 25 ft(3)/min) air-flow conditions. Special equipment was designed for measurement of filter efficiencies under the two air-flow conditions. Under low air-flow rate test conditions, glass wool was only 98.543 to 99.83% efficient, whereas cotton (five layers), fiberglass medium (three layers), and the commercial absolute filter were at least 99.900, 99.999, and 99.999 efficient, respectively. Glass wool and cotton were not used under higher air-flow conditions because they were difficult to assemble in leak-tight filters. The commercial absolute filter and fiberglass medium (three layers) were at least 99.990 and 99.999% efficient, respectively, under the higher air flow conditions. A stainless-steel filter of simple design and fitted with three layers of fiberglass medium was found to be greater than 99.999% efficient in removing high concentrations (20,000 to 70,000 plaque-forming units per cubic foot) of aerosolized bacteriophages from air moving at a low flow rate (1 ft(3)/min). Use of this filter on pressure-vacuum tanks in the fermentation industry is suggested. Several other uses of such a filter are proposed.     

Selective isolation of bacteria for metagenomic analysis: Impact of membrane characteristics on bacterial filterability

For indirect DNA extraction for metagenomics studies, bacterial cells can be effectively separated from sample debris by using a simple size exclusion technique, such as filtration, and thereafter lysed. The requirement for the optimal recovery of cells in filtrates is critical to achieve sufficient DNA yield and a representative population. Particles smaller than the filter pore size are expected to be found in the filtrate, whereas particles larger than the filter pore sizes are excluded. However, this is not always the case. It is established that the membrane pore size influences filtration efficiency to some degree. In addition the physicochemical characteristics of the filter suspension and characteristics of the microbial cells being filtered influence the exclusion property of a membrane. This review provides an overview of membrane filtration techniques and the factors that affect filterability of bacteria cells through a filter membrane. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:853–866, 2015     

Filtration capture immunoassay for bacteria: optimization and potential for urinalysis.

A novel assay utilizing immuno-labeling, filtration, and electrochemistry for the rapid detection of bacteria has been optimized for the detection of Escherichia coli O157:H7. Bacteria were specifically labeled with alkaline phosphatase conjugated polyclonal antibodies and captured on a polycarbonate track-etched membrane filter (0.2 microm pore size). The filter was then placed directly against a glassy carbon electrode, incubated with enzyme substrate, and the product detected by square wave voltammetry. The high speed and capture efficiency of membrane filtration and inherent sensitivity of electrochemical detection produced a 25-min assay with a detection limit of 5 x 10(3) E. coli O157:H7 per ml using a filtration volume of 100 microl (i.e. 500 cells filtered). The labeling, filtration, and electrochemical steps were optimized, and the assay performance using electrochemical and colorimetric detection methods was compared. The assay was used to detect E. coli O157:H7 that was spiked into filter-sterilized urine at clinically relevant concentrations.     

Automated protein precipitation by filtration in the 96-well format

The use of automated protein precipitation by filtration in the 96-well format as a rapid sample preparation technique for high throughput bioanalysis using liquid chromatography tandem mass spectrometry is reported. A robotic sample processor is used to aspirate sequentially a plasma sample and acetonitrile separated by air gaps. These are then mixed by being dispensed into individual channels of a 96-well filter block. The resulting supernatant is separated from the precipitated plasma proteins by the application of gentle vacuum using a custom manifold. The filtered supernatants are collected into a deep well microtitre plate, evaporated to dryness using a heated 96-well dry down station and reconstituted in water prior to analysis. The efficiency of the extraction procedure is measured by the Lowry method for determining protein concentration. This method was used to optimise both the volume and the order of reagent addition, and to compare several prototype 96-well filter blocks. Using the optimised procedure a specific, precise and accurate method was developed for the β-agonist salbutamol in rabbit plasma with a calibration range of 1 to 100 ng/ml from 100 μl of sample.     

Effect of particle size distribution on filtration performance of cell culture medium

Cell culture media used in CHO-based biologic processes are typically sterile filtered to prevent microbial contamination prior to inoculation. In this study, the impact of common sterile filter throughput on a different, commercially available cell culture media was evaluated from the intermediate-adsorption fouling model of the filtration model. The key particle size range for optimum filter performance was discussed and identified by measuring the submicron order particle size distribution. It may be possible to predict the performance of filter capacity with size-exclusive separation by understanding the media particle counts and size distribution.     

Activation of polymeric materials towards enzymatic postgrafting and cross-linking

A methodology to activate inert polymeric materials to enzymatic functionalisation is described herein. Plasma irradiation can be used to graft compounds containing a moiety that is reactive towards an enzyme of interest. Subsequently, such enzyme can be used to either postgraft functional compounds or cross-link the polymeric materials. Argon plasma was utilised to graft 2-aminoethyl methacrylate onto cotton and wool fibres, introducing surface alkylamine groups to impart reactivity towards transglutaminase and tyrosinase. The efficiency of plasma grafting was verified by ATR-FTIR. Enzyme postgrafting of fluorescent peptides coupled with confocal microscopy was used to demonstrate transglutaminase activity towards cotton, a material typically inert to this enzyme. The grafting of alkylamines onto wool resulted in additional cross-linking by both enzymes, leading to significantly increased yarn breaking load and elongation at break. This technology permits the activation of inert materials towards enzymatic postgrafting, with applications in fields as diverse as textiles and biomaterials.     

Simultaneous filtration and immobilization of cells from a flowing suspension using a bioreactor containing polyethylenimine coated cotton threads: Application in the continuous inversion of concentrated sucrose syrups

Polyethylenimine(PEI)-coated cotton threads were shown to have potential for reducing microbial load from a flowing suspension. Turbid cell suspensions perfused through the PEI column appeared as totally clear in the effluent. The adhesion efficiency of the matrix was found to depend on the concentration of PEI used to treat the threads. Threads coated with 2.5% PEI were found to show optimal retention of cells. A considerable amount of binding was seen over a broad range of ionic concentration (0–0.3 M) and pH (3.6–10.3). Under similar conditions control threads did not show any filtration capacity. Saccharomyces cerevisiae, Saccharomyces fragilis, Escherichia coli and an Acetobacter species could be effectively filtered using PEI-coated threads. This technique can find potential for the simultaneous filtration and immobilization of cells in a bioreactor to be used in continuous bioprocessing as exemplified for the inversion of sucrose syrups using baker's yeast. The bioreactor could continuously hydrolyse 60% (w/v) sucrose syrups with a productivity of 2.25 kg/day for over a month without loss in efficiency.     

Comparison of aerosol and bioaerosol collection on air filters

Air filters efficiency is usually determined by non-biological test aerosols, such as potassium chloride particles, Arizona dust or di-ethyl-hexyl-sebacate (DEHS) oily liquid. This research was undertaken to asses, if application of non-biological aerosols reflects air filters capacity to collect particles of biological origin. The collection efficiency for non-biological aerosol was tested with the PALAS set and ISO Fine Test Dust. Flow rate during the filtration process was 720 l/h, and particles size ranged 0.246–17.165 μm. The upstream and downstream concentration of the aerosol was measured with a laser particle counter PCS-2010. Tested bioaerosol contained 4 bacterial strains of different shape and size: Micrococcus luteus, Micrococcus varians, Pseudomonas putida and Bacillus subtilis. Number of the biological particles was estimated with a culture-based method. Results obtained with bioaerosol did not confirmed 100% filters efficiency noted for the mineral test dust of the same aerodynamic diameter. Maximum efficiency tested with bacterial cells was 99.8%. Additionally, cells reemission from filters into air was also studied. Bioaerosol contained 3 bacterial strains: Micrococcus varians, Pseudomonas putida and Bacillus subtilis. It was proved that the highest intensity of the reemission process was during the first 5 min. and reached maximum 0.63% of total number of bacteria retained in filters. Spherical cells adhered stronger to the filter fibres than cylindrical ones. It was concluded that non-biological aerosol containing particles of the same shape and surface characteristics (like DEHS spherical particles) can not give representative results for all particles present in the filtered air.     

An aerobiological study of pollen grains and fungal spores of Barcelona (Spain)

Summary A study of concentration of airborne pollen grains and fungal spores has been carried out in Barcelona (Spain) during 1989–90. The volumetric method of filtration, previously described for airborne pollen analysis (Suarez-Cervera and Seoane-Camba, 1983) has been used. In this case, the filters have also been cultivated in Czapecdox-agar, Sabouraud-agar and Sabouraud-agar with streptomycin for the identification of the fungal colonies. Analysis of the number of fungal spores growing on the filter shows that the maxima of colonies of spores developed in culture per m³ of air filtered, correspond to September–December. Pollen and spore concentrations start from November–December, reach a maximum in March–April and decline progressively until September–October. Therefore, in the city of Barcelona, the greatest concentration occurs in spring and the lowest in autumn.     

Proceedings: A method for preservation of bacteria and bacteriophages by drying in vacuo

An efficient and practical method was established to preserve bacterial strains and bacteriophages. The method is characterized by drying without freezing and by use of a cotton wool plug (nonabsorbent) to prevent contamination. Drying conditions were examined by measuring temperature, vacuum, and residual moisture of the samples. From the measurement, it was found that the cotton wool plug acts as a buffer and a desiccant. Thus, the specimens reached optimal conditions during storage. Another point of advantage is that the temperature of the specimen during the drying procedure was 2–5 °C; therefore, the evaporation of the water is rapid and the time of completion is shorter than that during lyophilization.     

The effect of heparin, caffeine and calcium ionophore A23187 on in vitro induction of the acrosome reaction in frozen-thawed bovine and caprine spermatozoa

The effect of heparin (5 IU), caffeine (5 mM) and calcium-ionophore A23187 (0.1 mM) on motility and in vitro induction of the acrosome reaction in glass wool filtered frozen-thawed bull and goat semen was studied. The motile spermatozoa fraction was obtained after glass wool filtration of frozen-thawed semen. The seminal plasma was removed from filtered semen by centrifugation, and the sperm pellet was resuspended in Sperm-TALP medium. Samples of treated and untreated control semen of both species were incubated at 37 degrees C. At 1, 15 and 30 min of incubation the proportions of progressively motile and acrosome-reacted spermatozoa were assessed. Trypan blue and Giemsa stain was used to differentiate live and dead spermatozoa having undergone acrosome reaction. Glass wool filtration enhanced the proportion of motile spermatozoa from 43% to 62% in the bovine and from 41% to 60% in the caprine. Whereas the effect of incubation with caffeine, heparin and calcium-ionophore on spermatozoan motility was negligible, the treatment of semen with calcium-ionophore resulted in a significantly improved percentage of live spermatozoa with true acrosome reaction at all stages of incubation, both in the bovine and the caprine.     

Residual Disinfection of Wool Blankets Treated with Formaldehyde

SUMMARY: Treatment with formaldehyde during laundering conferred a persistent residual disinfectant action on wool blankets but not on cotton ones. The former was shown by a slower rate of bacterial contamination of blankets during use or by the more rapid disappearance of nonsporing bacteria from contaminated blankets during storage. The formaldehyde treated blankets had no perceptible odour during use, and did not cause irritation of the skin or mucous membranes. It is suggested that the formaldehyde treatment of wool blankets might be worthy of trial as a means of reducing bacterial contamination in rooms occupied by patients with a high risk of infection.     

Fluorometric Method for Determining the Efficiency of Spun-Glass Air Filtration Media

The procedures and equipment needed to measure filtration efficiency by means of fluorescent aerosols are described. The filtration efficiency of individual lots of spun-glass air filtration medium or of entire air filtration systems employing such media was determined. Data relating to the comparative evaluation of spun-glass filter media by means of the fluorometric method described, as well as by conventional biological procedures, are presented.     

Factors Affecting the Persistence of Staphylococcus aureus on Fabrics

The persistence of Staphylococcus aureus (Smith) on wool blanket, wool gabardine, cotton sheeting, cotton knit jersey, cotton terry cloth, and cotton wash-and-wear fabrics was studied. The fabrics were exposed to bacterial populations by three methods: direct contact, aerosol, and a lyophilized mixture of bacteria and dust having a high content of textile fibers. The contaminated fabrics were held in 35 or 78% relative humidities at 25 C. In general, the persistence time of S. aureus populations on fabrics held in 35% relative humidity was substantially longer when the fabrics were contaminated by exposure to aerosolized cultures or to dust containing bacteria than when contaminated by direct contact. In a 78% relative humidity, bacterial populations on the fabrics persisted for substantially shorter periods of time regardless of the mode of contamination or fabric type. Cotton wash-and-wear fabric (treated with a modified triazone resin) was the material on which populations of S. aureus persisted for the shortest time. This organism retained its virulence for Swiss mice after being recovered from wool gabardine swatches held 4 weeks in 35% relative humidity and 6 weeks in 78% relative humidity.     

Extremozymes for improving wool properties

The project ‘EXTRETEX’ funded by the German Federal Foundation Environment (DBU, Osnabrück, Germany) aims at the improvement of wool properties dyeability, handle, felting behaviour and degree of whiteness by means of enzymes derived from extremophilic micro-organisms. In this paper the effects of a commercial thermo- and alkalistable protease on wool with regard to the degree of whiteness, the dyeability and the felting behaviour are presented. A method to treat wool top and wool fabric was developed on a laboratory scale in which the protease was integrated into the pre-washing step of a dyeing process. This treatment method was than scaled up and tested on an industrial winch beck for fabric. With this method—the addition of enzyme in the pre-washing step—the degree of whiteness is generally enhanced. Dyeing untreated and the enzyme-treated wool with Lanasol Blue 8G leads to an improved dyestuff uptake and a distinctive difference in the colour shade for the latter. Microscopy pictures of fibre cross-sections of these samples display a more even distribution of the dyestuff and a better penetration in the enzyme-treated wool fibres but the colour fastness of the enzyme-treated wool is decreased. Though the felting behaviour of the protease treated wool is significantly improved the felting tendency is still too high for an antifelting finish. An increased damage of the enzyme-treated wool in comparison with the untreated one was not observed.     

Feeding behaviour,midgut distension and ovarian development in Phlebotomus papatasi (Diptera: Psychodidae)

Phlebotomus papatasi females were fed through membranes or from cotton wool soaked in blood, water, sucrose or sodium chloride solutions. In membrane-fed flies, all diets were routed to the midgut and not to the crop. Following the meals that went to the midgut, females showed ovarian development at least 3 times greater than in sucrose-fed, autogenous control flies. Neither small quantities of water arriving in the midgut following drinking from soaked cotton wool, nor piercing of a membrane without feeding, stimulated ovarian development. Flies exhibited different feeding behaviour namely, blood feeding, sugar feeding, and water drinking. The blood-feeding behaviour was typical of flies ingesting any of the experimental diets through membranes, or blood or saline from cotton wool. The other two types of behaviour were observed in flies which fed from soaked cotton wool. The type of behaviour was characterized by the depth of penetration of the mouthparts into the substrate, the deployment of the palps and the degree of contact between the palps and the surface. It is suggested that the stimuli which control the routing of meals to the crop or to the midgut are derived from these types of behaviour.