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Background: Resistance to carbapenem antibiotics is emerging worldwide among Enterobacteriaceae. To prevent hospital transmission due to unnoticed carriage of carbapenemase producing micro-organisms in newly admitted patients, or follow-up of patients in an outbreak setting, a molecular screening method was developed for detection of the most prevalent carbapenemase genes; bla(OXA-48), bla(VIM), bla(IMP), bla(NDM), and bla(KPC). Methods: A real-time multiplex PCR assay was evaluated using a collection of 86 Gram negative isolates, including 62 carbapenemase producers. Seven different laboratories carried out this method and used the assay for detection of the carbapenemase genes on a selection of 20 isolates. Results: Both sensitivity and specificity of the multiplex PCR assay was 100%, as established by results on the strain collection and the inter-laboratory comparisons. Conclusions: In this study, we present a multiplex real-time PCR that is a robust, reliable and rapid method for the detection of the most prevalent carbapenemases bla(OXA-48), bla(VIM), bla(IMP), bla(NDM), and bla(KPC), and is suitable for screening of broth cultured rectal swabs and for identification of carbapenemase genes in cultures.

Item does not contain fulltext, BACKGROUND: Late April 2009, human infection with variant influenza virus A(H1N1)v emerged in the Northern Americas posing a threat that this virus may become the next pandemic influenza virus. OBJECTIVES: To prepare laboratories for surge capacity for molecular diagnosis of patients suspected for A(H1N1)v infection in the Netherlands. STUDY DESIGN: A panel of 10 blinded specimens containing seasonal A(H1N1) or A(H3N2), or A/Netherlands/602/2009(H1N1)v influenza virus, or negative control was distributed to the outbreak assistance laboratories (OAL) together with influenza virus A (M-gene), swine influenza virus A (NP-gene) and influenza virus A(H1N1)v (H1v-gene) specific primers and probes and protocol (CDC Atlanta, USA). Laboratories were asked to implement and test this protocol. RESULTS: All OAL were able to detect A(H1N1)v using the CDC M-gene reagents, the majority with similar sensitivity as the in-house M-gene based assays. RT-PCRs used in routine diagnostic setting in the OAL specifically designed to detect H1, H3, or NS1 from seasonal influenza A viruses, did not or at very low level cross-react with A(H1N1)v. The CDC swine NP-gene and H1v-gene RT-PCRs showed somewhat reduced sensitivity compared to the CDC and in-house M-gene RT-PCRs. In contrast, in-house developed A(H1N1)v specific H1v-gene and N1v-gene RT-PCRs showed equal sensitivity to CDC and in-house M-gene RT-PCRs. CONCLUSIONS: The Dutch OAL are prepared for detection and specific identification of A(H1N1)v, although some level of cross-reactivity was observed with seasonal influenza viruses. Additionally, M-gene based generic influenza A virus detection is recommended to be able to detect emerging influenza A viruses in routine settings.

Contains fulltext : 81491.pdf (publisher's version ) (Closed access), BACKGROUND: Experience with a highly pathogenic avian influenza outbreak in the Netherlands (2003) illustrated that the diagnostic demand for respiratory viruses at different biosafety levels (including BSL3), can increase unexpectedly and dramatically. OBJECTIVES: We describe the measures taken since, aimed at strengthening national laboratory surge capacity and improving preparedness for dealing with diagnostic demand during outbreaks of (emerging) respiratory virus infections, including pandemic influenza virus. STUDY DESIGN: Academic and peripheral medical-microbiological laboratories collaborated to determine minimal laboratory requirements for the identification of viruses in the early stages of a pandemic or a large outbreak of avian influenza virus. Next, an enhanced collaborative national network of outbreak assistance laboratories (OAL) was set up. An inventory was made of the maximum diagnostic throughput that this network can deliver in a period of intensified demand. For an estimate of the potential magnitude of this surge demand, historical counts were calculated from hospital- and physician-based registries of patients presenting with respiratory symptoms. RESULTS: Number of respiratory physician-visits ranged from 140,000 to 615,000 per month and hospitalizations ranged from 3000 to 11,500 per month. The established OAL-network provides rapid diagnostic response with agreed quality requirements and a maximum throughput capacity of 1275 samples/day (38,000 per month), assuming other routine diagnostic work needs to be maintained. CONCLUSIONS: Thus surge demand for diagnostics for hospitalized cases (if not distinguishable from other respiratory illness) could be handled by the OAL network. Assessing etiology of community acquired acute respiratory infection however, may rapidly exceed the capacity of the network. Therefore algorithms are needed for triaging for laboratory diagnostics; currently this is not addressed in pandemic preparedness plans.

Background Many different techniques have been designed for the quantification of JAK2V617F allelic burden, sometimes producing discrepant results. Design and Methods JAK2V617F quantification techniques were compared among 16 centers using 11 assays based on quantitative polymerase chain reaction (with mutation-specific primers or probes, or fluorescent resonance energy transfer/melting curve analysis), allele-specific polymerase chain reaction, conventional sequencing or pyrosequencing. Results A first series of blinded samples (granulocyte DNA, n=29) was analyzed. Seven assays (12 centers) reported values inside the mean±2SD; the mean coefficient of variation was 31%. Sequencing techniques lacked sensitivity, and strong discrepancies were observed with four techniques, which could be attributed to inadequate standards or to different modes of expression of results. Indeed, quantification of JAK2V617F in relation to another control gene produced higher than expected values, suggesting the possibility of more than two JAK2 copies/cell. After calibration of assays with common 1% to 100% JAK2V617F standards (dilutions of UKE-1 cells in normal leukocytes), 14 centers tested ten new samples. JAK2V617F allelic burdens greater or equal than 1% were then reliably quantified by five techniques – one allele specific-polymerase chain reaction and four TaqMan allele-specific quantitative polymerase chain reaction assays, including one previously giving results outside the mean±2SD – with a lower mean coefficient of variation (21%). Of these, only the two TaqMan allele-specific quantitative polymerase chain reaction assays with primer-based specificity could detect 0.2% JAK2V617F. Conclusions Techniques expressing the allelic burden as JAK2V617F/total JAK2 and using a common set of standards produced similar quantification results but with variable sensitivity. Calibration to a reference standard improved reproducibility.

Microstructural development in gas tungsten arc (GTA) welded silicon- and aluminum-based transformation-induced plasticity (TRIP) steels was studied by optical and electron microscopy. The fusion zone (FZ) of both welds contained complex inclusions. Energy-dispersive spectroscopic (EDS) analysis on these inclusions showed that the center of the inclusions contained oxides of silicon and aluminum in silicon- and aluminum-based steels, respectively. Epitaxial enrichment of manganese, sulfur, and phosphorous was found on the oxides in the inclusions, resulting in depletion of solutes in solid solution and thereby reducing the stability of austenite in the fusion zone. The fusion line of aluminum-based steel weldments contained higher amounts of allotriomorphic ferrite than silicon-based steel weldments due the partitioning of aluminum at the fusion line during solidification. The retained austenite (RA) contents in the heat-affected zones (HAZs) and FZs of both TRIP steels were found to be about 8 and 4 pct by volume, respectively., Materials Science and Engineering, Mechanical, Maritime and Materials Engineering

Op drinkwaterproductiebedrijf Spannenburg doseert Vitens zuivere zuurstof aan het beluchte ruwe water om een volledige ammoniumomzetting in de voorfilters te bereiken en nagroei in het distributienet te voorkomen. Deze werkwijze is uniek in Nederland en waarschijnlijk ook uniek in de wereld. Zuurstof in water oplossen lijkt op het eerste gezicht niet moeilijk. Dit is het wel als men zuurstof doseert in een open goot waar het water invalt en men eisen stelt aan het rendement van de zuurstofinbreng. In dit artikel wordt onder meer ingegaan op de manier waarop het uiteindelijk gelukt is om zuurstof in het water op te lossen met een rendement boven de 80 procent

Er bestaat tot nog toe geen enkel hard bewijs dat trekvogels het hoogpathogene H5N1 hebben verspreid. Verschillende meningen van verschillende organisaties en personenen. In dit artikel aandacht voor de vogeltrek en ophokplicht, het besmettingsrisico en aanwijzingen voor preventieve zorg voor het welzijn van hobbypluimvee

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Pompstation Spannenburg had na de invoering van diepere ontharding tot 1,2 mmol/l te kampen met verkorte looptijden van de nafilters en kalkafzetting in leidingen, op meetapparatuur en kleppen. Ook de consument ondervond meer hinder van kalkafzetting dan voorheen. De oorzaak hiervan bleek de relatief hoge pH-waarde van het ontharde water. Om de problemen te verhelpen is kooldioxidedosering geïntroduceerd. Na ongeveer 1,5 jaar bedrijfservaring blijkt dit een enorme kostenbesparing op te leveren: de looptijden van de nafilters zijn verdubbeld van de oorspronkelijke 45 uur naar 90 uur. Daarnaast blijven kleppen, leidingen en meetapparatuur vrij van kalkafzetting. Ook de consument kan het verschil merken door de veel lagere kalkafzetting op de huishoudelijke apparatuur