[Detection and identification of seven clinical common pathogenic bacteria by oligonucleotide microarray].
OBJECTIVE
Using 16S rDNA and 23S rDNA genes as the target sequences to develop a system based on oligonucleotide microarray and to detect the seven clinical pathogenic bacteria, commonly seen.


METHODS
Double polymerase chain reaction (PCR) was applied to amplify the segments of 16S rDNA and 23S rDNA genes of the target bacteria. An oligonucleotide microarray was constructed to simultaneously detect EHEC O157:H7, Vibrio parahaemolyticus, Salmonella sp., Vibrio cholerae, Listeria monocytogenes, Campylobacter jejuni and Shigella sp. Specificity, sensitivity and reproducibility of the microarray during detection were checked. And then microarray was used to detect the microbes in stool specimens of 81 patients with diarrhea and vomiting.


RESULTS
The double PCR method could simultaneously amplify the target sequences of 16S rDNA and 23S rDNA genes of the seven pathogens. The sensitivity of the developed oligonucleotide microarray could reach 10(3) cfu/ml but no positive results were presented for non-targeted bacteria. The coefficients of differentiation in one lot or among different lots of the microarray slices were 3.89% - 5.81%. The positive detection rate of the stool specimens by oligonucleotide microarray was 39.5% (32/81), with a coincidence of 96.3% (78/81) for the patients and another coincidence of 96.8% (31/32) for bacterial genus or species identification, when comparing to the results by routine bacteriological examinations.


CONCLUSIONS
The established assay in this study based on oligonucleotide microarray to detect the seven pathogenic bacteria has many advantages such as convenient, rapid, accurate, stable and high flux, which is suitable for clinical specimen examination and epidemiological field investigation.
Xing
Jian-ming
JM
Department of Laboratory Medicine, Huzhou Maternity and Child Care Hospital, Huzhou 313000, China.
Zhang
Su
S
Zhang
Hong-he
HH
Shen
Cui-fen
CF
Bi
Dan
D
Li
Gang
G
Yao
Li-hui
LH
D001419:Bacteria / Q000302:isolation & purification*; D016123:Campylobacter jejuni / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics*; D019453:Escherichia coli O157 / Q000302:isolation & purification; D006801:Humans; D008089:Listeria monocytogenes / Q000302:isolation & purification; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012338:RNA, Ribosomal, 23S / Q000235:genetics; D015203:Reproducibility of Results; D012475:Salmonella / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D012760:Shigella / Q000302:isolation & purification; D014734:Vibrio cholerae / Q000302:isolation & purification; D014736:Vibrio parahaemolyticus / Q000302:isolation & purification
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D012338:RNA, Ribosomal, 23S