Multiplex characterization of human pathogens  including species and antibiotic-resistance gene  identification.
The efficient medical treatment of infections requires detailed information about the pathogens involved and potential antibiotic-resistance mechanisms. The dramatically increasing incidence of multidrug-resistant bacteria especially highlights the importance of sophisticated diagnostic tests enabling a fast patient-customized therapy. However, the current molecular detection methods are limited to either the detection of species or only a few antibiotic-resistance genes.In this work, we present a human pathogen characterization assay using a rRNA gene microarray identifying 75 species comprising bacteria and fungi. A statistical classifier was developed to facilitate the automated species identification. Additionally, the clinically most important β-lactamases were identified simultaneously in a 100-plex reaction using padlock probes and the same microarray. The specificity and sensitivity of the combined assay was determined using clinical isolates. The detection limit was 10(5) c.f.u. ml(-1), recovering 89 % of the detectable β-lactamase-encoding genes specifically. The total assay time was less than 7 hand the modular character of the antibiotic-resistance detection allows the easy integration of further genetic targets. In summary, we present a fast, highly specific and sensitive multiplex pathogen characterization assay.
Molecular Diagnostics, AIT Austrian Institute of Technology, 1190 Vienna, Austria.
Ivan
I
Barišić
Molecular Diagnostics, AIT Austrian Institute of Technology, 1190 Vienna, Austria.
Josefine
J
Petzka
Molecular Diagnostics, AIT Austrian Institute of Technology, 1190 Vienna, Austria.
Silvia
S
Schoenthaler
Molecular Diagnostics, AIT Austrian Institute of Technology, 1190 Vienna, Austria.
Klemens
K
Vierlinger
Molecular Diagnostics, AIT Austrian Institute of Technology, 1190 Vienna, Austria.
Christa
C
Noehammer
Molecular Diagnostics, AIT Austrian Institute of Technology, 1190 Vienna, Austria.
Herbert
H
Wiesinger-Mayr
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001419:Bacteria / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D006801:Humans; D046228:Microarray Analysis / Q000379:methods; D008826:Microbial Sensitivity Tests; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012680:Sensitivity and Specificity; D013045:Species Specificity; D001618:beta-Lactamases / Q000235:genetics
D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D001618:beta-Lactamases