ARTICLE Design and validation of a low density array (Nosochip) for the detection and identification of the main pathogenic bacteria and fungi responsible for nosocomial pneumonia S. Burteau & P. Bogaerts & R. de Mendonça & L. Irenge & C. Berhin & J. Hiffe& N. de San & P. Beyne & S. Hamels & Y. Glupczynski & M. Struelens & J.-L. Gala& J. Remacle Published online: 29 September 2007 # Springer-Verlag 2007 AbstractThe aim of this study was to be able to amplify and to detect on one array 27 different etiologic agents found in nosocomial pneumonia, some being phylogenet- ically closely related and others very distant. The assay is based on the use of consensus primers combined with the identification of the resulting amplicons by hybridization on specific capture probes present on an array. Three genes were necessary in order to coverthe different pathogens. We took a redundancy of at leasttwo positive spots to confirm theidentity ofeach species. Each probewas present in triplicate on the array. The detection limit was between 10 and 1,000 DNA copies in the assay depending on the bacteria and the probe. The assay was also specific when tested both on reference collection strains correspon to the 27 species of interest and on 57 otherbacterial species of the normal human flora. Accuracy of the assay was assessed on 200 clinical isolatesand somepoly- morphisms were indeed observed for 5 species. Effective- nessof theassaywas preliminarilyvalidatedon 25 endotracheal aspirates and sputum samples, and the results were in accordance either with the cell culture or with the sequencing. Polybacterial infections were well detected in threesamples. The resultsshow thata combination of appropriate polymerase chain reaction (PCR) and redunda cy of signalson the array allowspecific screening of bacteria belonging to different species and genus and eve fungi.The results open the way for a possible molecular detection of bacteria in the clinical diagnostic setting. Introduction In recent years, culture-independent molecular methods b on the polymerase chain reaction (PCR) have been propos and used for microbiological diagnostics with the aim of speeding up the diagnostics [ 1]. Several PCR assays have developed to detect species-specific genes: for instance, the identification of Streptococcus pneumoniae by the amplifi tion of the pneumolysin gene [ 5], detection of mecA and f genes in the identification of methicillin-resistant Staphylo- coccus aureus [13], identification of mycobacterial species amplification of the specificMycobacterium tuberculosis Eur J Clin Microbiol Infect Dis (2008) 27:17–27 DOI 10.1007/s10096-007-0394-1 S. Burteau : J. Hiffe : J. Remacle (*) URBC, FUNDP, 61 Rue de Bruxelles, 5000,Namur, Belgium e-mail: Jose.remacle@fundp.ac.be P. Bogaerts : C. Berhin : Y. Glupczynski Department of Microbiology, Cliniques Universitaires, UCL, Mont-Godinne, 5530 Yvoir, Belgium R. de Mendonça : N. de San : M. Struelens Department of Microbiology, Hopital Erasme, ULB, 1070 Brussels, Belgium L. Irenge : J.-L. Gala Laboratory of Applied Molecular Technologies, Center for Human Genetics, UCL, Av Mounier 52, 1200 Brussels, Belgium J.-L. Gala Defence Laboratories Department, Belgian Armed Forces, Brussels, Belgium P. Beyne CHR, A. Albert 1er, 5000 Namur, Belgium S. Hamels Eppendorf Array Technologies, SA, Rue du seminaire 20, 5000 Namur, Belgium