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    Microarray-based genotyping of Salmonella:
    inter-laboratory evaluation of reproducibility and standardization potential (2011)

    Art
    Zeitschriftenartikel / wissenschaftlicher Beitrag
    Autoren
    Grønlund, Hugo
    Riber, Leise
    Vigre, Håkan
    Löfström, Charlotta
    Folling, Liselotte
    Huehn, Stephan
    Malorny, Burkhard
    Rådström, Peter
    Rudi, Knut
    Hoorfar, Jeffrey
    Quelle
    International Journal of Food Microbiology; 145(Suppl. 1) — S. 79–85
    ISSN: 0168-1605
    Sprache
    Englisch
    Verweise
    DOI: 10.1016/j.ijfoodmicro.2010.08.007
    Pubmed: 20817296
    Kontakt
    Institut für Lebensmittelsicherheit und -hygiene

    Königsweg 69
    14163 Berlin
    Tel.+49 30 838 62550 Fax.+49 30 838 46029
    email:lebensmittelhygiene@vetmed.fu-berlin.de

    Abstract / Zusammenfassung

    Bacterial food-borne infections in humans caused by Salmonella spp. are considered a crucial food safety issue. Therefore, it is important for the risk assessments of Salmonella to consider the genomic variation among different isolates in order to control pathogen-induced infections. Microarray technology is a promising diagnostic tool that provides genomic information on many genes simultaneously. However, standardization of DNA microarray analysis is needed before it can be used as a routine method for characterizing Salmonella isolates across borders and laboratories. A comparative study was designed in which the agreement of data from a DNA microarray assay used for typing Salmonella spp. between two different labs was assessed. The study was expected to reveal the possibility of obtaining the same results in different labs using different equipment in order to evaluate the reproducibility of the microarray technique as a first step towards standardization. The low-density array contains 281 57-60-mer oligonucleotide probes for detecting a wide range of specific genomic marker genes associated with antibiotic resistance, cell envelope structures, mobile genetic elements and pathogenicity. Several critical methodology parameters that differed between the two labs were identified. These related to printing facilities, choice of hybridization buffer, wash buffers used following the hybridization and choice of procedure for purifying genomic DNA. Critical parameters were randomized in a four-factorial experiment and statistical measures of inter-lab consistency and agreement were performed based on the kappa coefficient. A high level of agreement (kappa=0.7-1.0) in microarray results was obtained even when employing different printing and hybridization facilities, different procedures for purifying genomic DNA and different wash buffers. However, less agreement (Kappa=0.2-0.6) between microarray results were observed when using different hybridization buffers, indicating this parameter as being highly critical when transferring a standard microarray assay between laboratories. In conclusion, this study indicates that DNA microarray assays can be reproduced in at least two different facilities, which is a pre-requisite for the development of standard guidelines.