RESUMEN
In 2013, the Thermo Scientific™ SureTect™ Listeria monocytogenes Real-Time PCR Assay was certified by the AOAC Research Institute (RI) Performance Tested Methods(SM) program as a rapid method for the detection of L. monocytogenes from a wide range of food matrixes and surface samples. This report details the method modification studies undertaken to extend the analysis of this PCR assay to the Applied Biosystems™ 7500 Fast PCR Instrument and Applied Biosystems RapidFinder™ Express 2.0 software allowing the use of the SureTect assay on a 96 well format PCR cycler in addition to the current workflow, which uses the 24 well Thermo Scientific PikoReal™ PCR Instrument and Thermo Scientific SureTect software. Because this study was deemed by AOAC-RI to be a level 2 method modification study, a representative range of food matrixes covering raw ground turkey, 2% fat pasteurized milk, and bagged lettuce as well as stainless steel surface samples were analyzed with the Applied Biosystems 7500 Fast PCR Instrument and RapidFinder Express 2.0 software. All testing was conducted in comparison to the reference method detailed in International Organization for Standardization (ISO) 6579:2002. No significant difference by probability of detection statistical analysis was found between the SureTect Listeria monocytogenes PCR Assay or the ISO reference method methods for any of the matrixes analyzed during the study.
Asunto(s)
Técnicas Bacteriológicas/instrumentación , Listeria monocytogenes/genética , Listeria monocytogenes/aislamiento & purificación , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
The Thermo Scientific™ SureTect™ Listeria species Real-Time PCR Assay was certified during 2013 by the AOAC Research Institute (RI) Performance Tested Methods(SM) program as a rapid method for the detection of Listeria species from a wide range of food matrixes and surface samples. A method modification study was conducted in 2015 to extend the matrix claims of the product to a wider range of food matrixes. This report details the method modification study undertaken to extend the use of this PCR kit to the Applied Biosystems™ 7500 Fast PCR Instrument and Applied Biosystems RapidFinder™ Express 2.0 software allowing use of the assay on a 96-well format PCR cycler in addition to the current workflow, using the 24-well Thermo Scientific PikoReal™ PCR Instrument and Thermo Scientific SureTect software. The method modification study presented in this report was assessed by the AOAC-RI as being a level 2 method modification study, necessitating a method developer study on a representative range of food matrixes covering raw ground turkey, 2% fat pasteurized milk, and bagged lettuce as well as stainless steel surface samples. All testing was conducted in comparison to the reference method detailed in International Organization for Standardization (ISO) 6579:2002. No significant difference by probability of detection statistical analysis was found between the SureTect Listeria species PCR Assay or the ISO reference method methods for any of the three food matrixes and the surface samples analyzed during the study.
Asunto(s)
Análisis de los Alimentos , Microbiología de Alimentos , Listeria/genética , Listeria/aislamiento & purificación , Reacción en Cadena en Tiempo Real de la Polimerasa , Humanos , Listeria/clasificación , Reacción en Cadena en Tiempo Real de la Polimerasa/normas , Estándares de ReferenciaRESUMEN
Multidrug-resistant Acinetobacter baumannii (MDRAB) is an increasing problem worldwide. Prevalence of carbapenem resistance in Acinetobacter spp. due to acquired carbapenemase genes is not known in Finland. The purpose of this study was to examine prevalence and clonal spread of multiresistant A. baumannii group species, and their carbapenemase genes. A total of 55 Acinetobacter isolates were evaluated with repetitive PCR (DiversiLab) to analyse clonality of isolates, in conjunction with antimicrobial susceptibility profile for ampicillin/sulbactam, colistin, imipenem, meropenem, rifampicin and tigecycline. In addition, a new real-time PCR assay, detecting most clinically important carbapenemase genes just in two multiplex reactions, was developed. The assay detects genes for KPC, VIM, IMP, GES-1/-10, OXA-48, NDM, GIM-1, SPM-1, IMI/NMC-A, SME, CMY-10, SFC-1, SIM-1, OXA-23-like, OXA-24/40-like, OXA-58 and ISAbaI-OXA-51-like junction, and allows confident detection of isolates harbouring acquired carbapenemase genes. There was a time-dependent, clonal spread of multiresistant A. baumannii strongly correlating with carbapenamase gene profile, at least in this geographically restricted study material. The new carbapenemase screening assay was able to detect all the genes correctly suggesting it might be suitable for epidemiologic screening purposes in clinical laboratories.
Asunto(s)
Acinetobacter baumannii/enzimología , Acinetobacter baumannii/genética , Proteínas Bacterianas/genética , Genes Bacterianos/genética , Hospitales , Tipificación Molecular/métodos , Reacción en Cadena de la Polimerasa Multiplex/métodos , beta-Lactamasas/genética , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/aislamiento & purificación , Antibacterianos/farmacología , Células Clonales , Finlandia , Pruebas de Sensibilidad Microbiana , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducibilidad de los Resultados , Análisis de Secuencia de ADN , Factores de TiempoRESUMEN
The addition of potassium fluoride 'switches on' the fluorescence of sensors and while potassium chloride and bromide cause no fluorescence change; the fluorescence can be 'switched off' by removing the potassium cation from the benzocrown ether receptors of sensors and through the addition of [2.2.2]-cryptand and restored by the addition of the potassium cation as potassium chloride.