Detection of representative enteropathogenic bacteria, Vibrio spp., pathogenic Escherichia coli, Salmonella spp., Shigella spp., and Yersinia enterocolitica, using a virulence factor gene-based oligonucleotide microarray.
Rapid identification of enteropathogenic bacteria in stool samples is critical for clinical diagnosis and antimicrobial therapy. In this study, we describe the development of an approach that couples multiplex PCR with hybridization to a DNA microarray, to allow the simultaneous detection of the 10 pathogens. The microarray was synthesized with 20-mer oligonucleotide probes that were designed to be specific for virulence-factor genes of each strain. The detection limit for genomic DNA from a single strain was approximately 10 fg. In the presence of heterogeneous non-target DNA, the detection sensitivity of the array decreased to approximately 100 fg. We did not observe any non-specific hybridization. In addition, we successfully used this oligonucleotide-based DNA microarray to identify the causative agents in clinical stool samples from patients with food-borne enteritis.
Department of Microbiology, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Dong-Hun
DH
Kim
Bok-Kwon
BK
Lee
Yong-Dae
YD
Kim
Sung-Keun
SK
Rhee
Young-Chang
YC
Kim
D001431:Bacteriological Techniques / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics* / Q000302:isolation & purification; D004755:Enterobacteriaceae / Q000235:genetics / Q000472:pathogenicity*; D004756:Enterobacteriaceae Infections / Q000175:diagnosis* / Q000382:microbiology; D005243:Feces / Q000382:microbiology*; D005517:Foodborne Diseases / Q000175:diagnosis / Q000382:microbiology; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D037521:Virulence Factors / Q000235:genetics* / Q000302:isolation & purification
D004269:DNA, Bacterial; D037521:Virulence Factors