Das Autoselect-system – ein Automatensystm zu Selektion von Antibiotikaproduzenten. II. Agarabfüllautomat

An agardeliverer, one of the machines of the Autoselect-system is described. This machine allows to pour melted agar automatically into a row of 8 microculture cups of a cassette with 64 cups. By changing the agarcontainer provided for one medium against another container with 8 chambers the machine offers the possibility to deliver 1 to 8 media simultaneously. In this respect the machine gets more and more interest not only for the selection of antibiotic producers, but also for taxonomic, genetic and other studies. Sterile conditions are ensured by sterilizing the head of the machine before starting the work.     

Das Autoselect-System – ein Automatensystem zur Selektion von Antibiotikaproduzenten. VI. Achtfach-Stanzautomat

An eightfold punch belonging to the Aùtoselect-system is described. It is provided for the preparation of bioassay plates. From the agar of large quadratic test plates moving automatically on a carriage of the machine, 64 holes are made by eight punches. The punches arranged in a row over the test plate are lowered, if the carriage stops, and so they cut out and exhaust agar discs in a pattern of 8 × 8.     

Increase in Sensitivity for the Assay of Neomycin in Milk

A study was conducted to design a more sensitive and flexible technique for the assay of neomycin in milk. Sterile Antibiotic Assay medium (Difco; 15 ml) was poured into glass petri dishes with aluminum tops equipped with absorbent discs. Seed agar (4 ml; containing 1% sodium chloride) inoculated with standardized amounts of the test organism (Staphylococcus epidermidis ATCC 12228) was laid over the hardened base agar, and stainless-steel cylinders were placed equal distances apart in the agar. The plates were refrigerated for 30 min and the cups were removed in an atmosphere of minimal contamination. The resultant holes were sealed with melted agar. After preparation, the holes were used for assay procedures. Samples to be assayed were pipetted into the holes and the plates were refrigerated for 90 min at 4 C. Plates were incubated for 18 to 20 hr at 32 C. The zones of inhibition were recorded and compared with a standard curve. The approximate sensitivity of this method was 0.05 mug/ml.     

Das Autoselect-System – ein Automatensystem zur Selektion von Antibiotikaproduzenten. III. Ein Impfautomat zum Isolieren von Einzelkolonien

An inoculator enabling the isolation of colonies from petridishes onto agar-filled microcultur cups, arranged in a pattern of 8 × 8 in cassettes, is described. The transfer of the colonies takes place by turning of an inoculation cross with 4 loops. While the cross stops and lowers, one loop is sterilized, another, which has been sterilized shortly before, is being cooled in sterile water, the next one is taking off some material from a colony and the last is spreading the material on the agar of a cup. The capacity of the machine accounts about 600 colonies per hour.     

Advances in rearing ofLixophaga diatraeae [Dipt.: Tachinidae]

Larvae ofDiatraea saccharalis (F) were reared on artificial diet in 30-ml cups (1–3 larvae/cup). Adults of the larviparous tachinid parasite,Lixophaga diatraeae (Townsend), were removed from emergence and holding cages with a modified vacuum sweeper. Maggots were extracted from 10 to 14-day-old female flies, that had been disinfected with 1% NaOCl by one of two methods. In method I, fly uteri were removed and placed in a 10-ml vial containing a 0.15% agar-water solution with 3–4 glass beads; rapid vibration of the vial ruptured the uteri and distributed the maggots in the agar solution. In method 2, whole flies were blended with 50-ml water in a blender, and maggots were separated from fly particles by screening; then they were suspended in the agar solution. A procedure was devised for determining the number of maggots obtained by each method. The maggot-agar solution was injected into cups containing host larvae and maggots sought out and parasitized the host larvae; however, the percentage producing puparia generally decreased with increases of host larva and/or maggot density. Puparia were harvested from cups by hand and washed in 1% NaOCl to disinfect and destroy host larval webbing.     

Das Autoselect-System – ein Automatensystem zur Selektion von Antibiotikaproduzenten. V. Achtfach-Pipettierautomat

An automatic eightfold-pipetter has been developed with the same basic unit as the diluter of the Autoselect-system. The pipetter is fitted with eight pipettes on a bridge over the basic unit. By means of the pipettes a volume of 0.05 ml from eight tubes is exhauseted and delivered in the holes of a test plate. Both the test plate and a cassette with 64 tubes are located on a moving carriage. the test plate on the first floor and the cassette on the ground floor. If the pipettes transfer the samples from the tubes to the holes the distance between the pipettes must be changed by a special device from 20 to 30 mm, because the distance between the holes on the test plate differs from that of the tubes. For the transfer of 64 samples 2 minutes only are needed.     

A simple agar plate assay for screening siderophore producer yeasts.

Yeasts produce hydroxamate-type siderophores (iron-binding compounds) in response to Fe-stress conditions. Because these siderophores are important to the biocontrol of postharvest diseases of apple and pears, a method for screening siderophore producer yeast was developed.The screening method was carried out in special Petri dishes with eight or nine wells (25-mm diameter). These wells were filled with siderophore production medium and seeded with yeasts isolated from epiphytic apple microflora. After yeasts grew (24-48 h), holes (2-mm diameter) were made in the agar of each well. Holes were filled with an acid solution of ferric perchlorate. After 10-15 min, reddish halos appeared in the bottom of the plate and their intensities were compared with standards. Standards were prepared in the same special dish with rhodotorulic acid solutions (concentrations between 0.05 and 1 g/l) plus 2% agar. When agar solidified into wells, holes were made and filled with ferric perchlorate solution. Color intensities of reddish halos were proportional to siderophore concentration and the detection limit was 0.1 g/l. It was possible to correlate the production of siderophore in solid medium with the results obtained in liquid medium. The methodology was also a useful tool for making a preliminary assessment of the influence of different factors on the siderophore production.     

Performance of polymyxin B agar-based tests among carbapenem-resistant Enterobacterales

Therapeutic options for infections caused by Carbapenem-resistant Enterobacterales (CRE) are restricted and include polymyxins-centred schemes. Evaluation of in vitro susceptibility is difficult and time consuming. Agar-based methodologies are an alternative to broth microdilution (BMD) and we aimed to evaluate the accuracy of those methods among Enterobacterales. A total of 137 non-duplicated CRE were subjected to polymyxin B BMD, agar screening test (Mueller Hinton plates containing 3 µg ml⁻¹ of polymyxin B) and agar dilution (antibiotic serially diluted 0·25–64 µg ml⁻¹). CRE of 42·3% were resistant to polymyxin B (MICs range: 0·25–>64 µg ml⁻¹) and 16·8% presented borderline MICs. Sensitivity, specificity, PPV and NPV were 86·2, 98·7, 98 and 90·7% for screening test and 86·2, 97·5, 96·1 and 90·6% for agar dilution. ME was 0·73 and 1·5% for screening and agar dilution respectively; VME was 5·8% for both techniques. In general, agar-based methods had a good performance. As far as we know, this is the first study to propose an agar screening test using polymyxin B instead of colistin.     

Life in Zero Magnetic Field. V. E. coli Resistance to Antibiotics

Twenty‐six E. coli strains, isolated from human subjects, were tested for antibiotic drug resistance using the dilution of antibiotic solutions in agar culture medium. The bacterial strains were then exposed to zero magnetic field in a well‐controlled laboratory area, where a Helmholtz coil compensated the local geomagnetic field. The exposure time to the zero magnetic field was 6 days. The antibiotic drugs with antimicrobial large action spectra used to evaluate bacteria resistance were ampicillin, ceftazidime, tetracycline, ofloxacin, and kanamycin. The aqueous solutions of drug had dilutions of 0.25, 0.50, 4, 8, 16, 32, 64, and 128 µm/mL, respectively. Two types of microorganisms were detected: strains sensitive and strains nonsensitive to geomagnetic field compensation. We found that the magnetic‐sensitive strains represent about one‐third of the analyzed samples, statistical analysis emphasizing the general tendency of diminishing resistance against antibiotics.     

Primary stability of threaded cups in THR--an experimental study.

Four threaded cups were tested up to their lever-out moments, torque-in moments and their resistance to failure. A was a parabolic-shaped, B was a spherical, C was a spherical-shaped too, and D was a conical shaped cup. Cup A and D represent cups which have proven themselves in clinical applications, but not cup B. The threads were determined and showed different constructive features. The cups were torqued into precise cavities in PVC foam cubes, after that they were levered out in a testing machine. The lever-out moments of all the cups showed significant differences; the results were: A: 78.4 Nm, B: 88.7 Nm, C: 117.5 Nm, D; 136.6 Nm. In the case of the torque-in moments there were no significant differences between A and B, neither between C and D. The differences in stiffness between B and C were not significant, but they were between the others. The primary stability against lever-out and the torque-in moment of threaded cups for artificial hip replacement can be basically influenced by different constructive features. Hence lever-out moment and torque-in moment should be understood and tested as independent variables.     

Antimicrobial Residues in Milk. Comparison of Different Agar Diffusion Methods

Different agar diffusion methods were compared in order to find a sensitive method for the detection of various antimicrobial residues in milk. A total of 588 producer milk samples were analyzed using subsets of the most sensitive methods. With the IDF method, 2 positive cases (0.34 %) appeared among the producer milk samples, with the Thermocult method 13 positive cases (2.21 %) and with the Test agar pH 8 method with trimethoprim and glucose 4 positive cases (0.68 %). A combination of the IDF method and the Test agar pH 8 method resulted in 6 positive cases (1.02 %) and a combination of the Thermocult method and the Test agar pH 8 method in 17 positive cases (2.89 %). With penicillinase 41 % of the positive cases were identified as β-lactam antibiotics and with p-aminobenzoic acid 18 % of the positive cases were identified as sulphonamides. 41 % of the positive cases remained unexplained. The best combination for the detection of antimicrobial agents in milk seems to be that of the Thermocult method and the Test agar pH 8 method with trimethoprim and glucose.     

Biochemical investigation of lens induction in vitro. I. Induction properties of the eye cup and ectodermal response

1. Optic cups of 48, 72 and 96 hours old chick embryos were prepared, cultured and recombined with ectoderm. With the optic cups of 48 hours old embryos, lens formation occurred in 16% of the cases. With the optic cups of 72 hours old embryos, lens formation occurred in 28% of the cases. Optic cups of 96 hours old embryos were not able to induce a lens. 2. The optic cup proved to be able to induce a lens more than once. 3. Ectoderm of the head of 72 hours old embryos was still able to form a lens. 4. Using homogenized eye cups of 72 hours old embryos, lens induction occurred only in a few cases. When the optic cups were cut into small pieces, lens induction occurred in 30% of the cases. This suggests that intact cells are necessary to obtain lens induction.     

THE NUMBERS AND GROWTH RATES OF THE BACTERIA IN HEVEA LATEX, AMMONIATED FIELD LATEX AND AMMONIATED LATEX CONCENTRATE

SUMMARY: The rates of growth of bacteria on Hevea latex systems in the presence and absence of ammonia have been derived from colony count data obtained on a modified Kligler's iron agar medium. For fresh latex, ammoniated field latex, and ammoniated latex concentrate, variations in the bacterial populations encountered are given for a period of about one year's testing, with some data on commercial samples of latex concentrate. Field latex at routine collection contained about 8 × 10⁶ bacteria/ml. This could be reduced by 62% by the use of sterile tapping cups. In the routine latex, the mixed population grew logarithmically after a lag phase of 2·1/2–3 hr. In clean latex the lag phase was extended to 4 hr. On ammoniation to 0.3% (w/w), the population was reduced over about 2 hr, but subsequently logarithmic growth recurred, without a lag phase. On concentration and further ammoniation to 0·7% (w/w), the count dropped still further, levels of 10/ml or less being achieved in clean conditions, but very slow logarithmic growth again occurred with a lag phase preceding it.     

Maintaining cultures of ectomycorrhizal and plant pathogenic fungi in sterile water cold storage.

Mycelial cultures of 64 isolates of 14 species of ectomycorrhizal fungi and 27 isolates of 15 species of plant pathogenic fungi were grown on agar medium in Petri dishes. Mycelial discs, 8 mm in diameter, were removed from the cultures and stored in sterile distilled water in test tubes at 5 degrees C. Sixty-four, 61, and 41 isolates of the symbiotic fungi were viable after 1, 2, and 3 years storage respectively. Only 19, 10, and 8 isolates of the pathogenic fungi were viable after 1, 2, and 3 years storage, respectively. Time in pure culture before water storage did not affect viability of any fungal species following water storage. After 3 years storage, four fungi (three symbionts and one pathogen) were tested and found to have retained their original growth rates and root-infecting abilities on pine seedlings. The same four isolates, however, maintained on agar slants at 5 degrees C and subcultured every 4 to 6 months, grew slower and did not infect as many feeder roots of pine as the water-stored isolates.     

A rapid method for culturing guinea pig gastric mucous cell monolayers

Summary A method has been developed for growing confluent primary cultured monolayers of guinea pig gastric mucous cells suitable for in vitro electrophysiological, transport, and pharmacological studies. Isolated mucous cells were enriched on a one-step Percoll density gradient and plated on fibronectin-coated plastic dishes or in small cups with holes containing glutaraldehyde-fixed Vitrogen gels. These cups were designed to fit in Ussing chambers. Mucous cells attached, proliferated, and formed confluent monolayers in 3 d. The low cuboidal cells contained periodic acid Schiff-positive mucous granules that were negative by Bowie and indirect immunofluorescent staining for pepsinogen. Electron microscopy revealed polarized mucous cells with microvilli, mucous granules, microfilaments, small mitochondria, some vacuoles, and junctional complexes that excluded wheat germ agglutinin-peroxidase. No basal lamina was present. Monolayers could be maintained for over 2 wk but subcultures were not made. The cultures were virtually free of fibroblasts. Epithelial sheets produced by this simple and rapid method can be used for electrophysiological, ion transport, and pharmacological studies. This research was supported in part by National Institutes of Health grants GM7806, AM31158, AM 15681, and AM 30303.     

Injured coliforms in drinking water.

Coliforms were enumerated by using m-Endo agar LES and m-T7 agar in 102 routine samples of drinking water from three New England community water systems to investigate the occurrence and significance of injured coliforms. Samples included water collected immediately after conventional treatment, during the backwash cycle, at various points in the distribution system, and 1 week after the break and subsequent repair of a distribution main. Injured coliforms in these samples averaged greater than 95%. m-T7 agar yielded 8- to 38-fold more coliforms than did m-Endo agar LES. The geometric mean of coliforms recovered by m-Endo agar LES was less than 1 confirmed coliform per 100 ml, although m-T7 agar yielded 5.7 to 67.5 confirmed coliforms per 100 ml. In addition, the majority of these samples giving positive results on m-T7 agar produced no detectable counts on m-Endo agar LES. These findings indicated that coliforms were injured and largely undetected by use of accepted analytical media in the systems examined.     

Design, construction and modularity of pressure-fit acetabular cups]

To enable a comparison of different pressfit acetabular cups objective criteria are essential. The aim of this study is to describe the design features of this type of cup and to analyse currently available cups. 30 implants were systematically measured and analysed. The mean surface roughness (Ra) was determined and configurations established with the light section technique. For further evaluation the cups were transversely sectioned. The cups are made of pure titanium, titanium alloy or polyethylene coated with titanium. Five implants take the form of monoblocks. The configuration is predominately (n = 25) flattened spherical. The size of eight cups corresponds to the outer diameter, 19 cups have a larger outer diameter (overdimensioning), 3 cups have a smaller outer diameter (underdimensioning). The magnitude of overdimensioning is, on average, 1.9%. 9 cups are provided with plugs, hollow cylinders, fins or rings as outer stabilizers. Surface roughness achieved with corundum blasting is 6.8 microns. Titanium porous-coated implants have a surface roughness of 21-32 microns. 24 cups have polyethylene inserts, most of which are snap-fixed with equatorial lips. For 16 cups, full-ceramic inserts are available. 4 cups have a metal insert. Titanium implants with structured or HAC-coated surfaces have become the accepted standard for cementless acetabular cup implantation. Together with ceramic, metal, or modified polyethylene inserts they meet the requirement for permanent osteo-integrative stability.     

Comparison of media and methods for detecting and enumerating Listeria monocytogenes in refrigerated cabbage.

Direct plating, selective enrichment, and cold enrichment followed by secondary selective enrichment procedures were compared for detecting and enumerating Listeria monocytogenes in chopped cabbage stored at 5 degrees C for up to 64 days. Addition of Fe3+ to solid media enhanced detection of the organism. Cold enrichment (5 degrees C) in nutrient broth and brain heart infusion broth followed by secondary enrichment (48 h, 30 degrees C) in Trypticase soy-yeast extract-antibiotic broth and thiocyanate-nalidixic acid broth and plating on selective agar media (Doyle and Schoeni selective enrichment agar [minus acriflavin hydrochloride, supplemented with 5 micrograms of Fe3+/ml] and McBride Listeria agar) resulted in the detection of highest populations.     

Reusable vs. disposable cups revisited: guidance in life cycle comparisons addressing scenario,model, and parameter uncertainties for the US consumer

Purpose

Despite interest in an environmentally conscious decision between disposable and reusable cups, a comprehensive and current study for US consumers is not yet available. Guidance in favor of single-use cups rely on outdated or non-ISO-compliant results with limited uncertainty information. Such claims are insufficiently generalizable. This article delivers an updated comparative life cycle impact assessment of reusable ceramic cups and single-use expanded polystyrene cups.

Methods

The ReCiPe midpoint model was selected. Scenario uncertainties are addressed by evaluating compliant standard dishwashing appliance models from 2004 to 2013 used in 26 US subregional utility grids. A utility snapshot from 2009 is applied with extension to recent shifts in generation from increased penetration of natural gas and renewable energy. Parameter uncertainty is quantified through statistical methods.

Results

Where there is statistical difference, results almost entirely favor reusable cups in the USA. For climate change, 16 % of users have higher impact for ceramic cups washed in 2013 by minimally compliant dishwashers. Higher climate change impacts for 32 % of reusable cup users is indicated with 2004 average dishwashers, though using a cup twice between washes shifts the impact in favor of the reusable cup.

Conclusions

Disposable cup scenarios do not account for film sleeves, lids, printing, and less conservative shipping weights and distances and therefore reflect a best case scenario. Impact for reusable cups is expected to decrease further as the electricity mix becomes less CO₂-intensive with replacement of coal-fired generators by natural gas, wind, and solar and as less efficient dishwashers are replaced with new units compliant to current laws.     

Performance of a semi-automated antibiotic susceptibility testing system (ABAC)

The ABAC system for antibiotic susceptibility testing was compared with an agar diffusion method in 14960 tests, including 23 antibacterial agents. Identical breakpoints were used. Only 3% major discrepancies (M.d.; sensitive vs resistant) and 19% minor discrepancies (m.d.; intermediate vs sensitive or resistant) were noted. Major discrepancies were mainly found for methicillin ( Staphylococcus aureus ), netilmicin ( Pseudomonas aeruginosa ), chloramphenicol, sulphamethoxazole and tri-methoprim ( Proteus sp.) and were checked by quantitative susceptibility tests. These showed ABAC to be at fault in 41–47% of discrepancies, the diffusion test in 21–32% and 21–37% were intermediate. Half of the m.d. involved beta-lactams, which is explained by too low breakpoints. Except for methicillin and netilmicin the overall results showed ABAC to be equal to the agar diffusion method. Technical faults, like leakage and incorrect filling of cups in the plastic rotors of ABAC, occurred in 14% of the rotors.