[Rapid diagnosis of neonatal sepsis by 16SrRNA genes PCR amplification and genechip hybridization].
OBJECTIVE
To explore a method for rapid diagnosis of sepsis in newborn infants.


METHODS
(1) The primers and oligonucleotide probes were designed and synthesized based on the sequences of bacterial 16SrRNA gene. The gene chip was prepared through the probes printed onto special glass slides. The gene chip included 18 special probes: universal probe 1, universal probe 2, Gram positive bacterial probe, Gram negative bacterial probe 1, Gram negative bacterial probe 2, Staphylococcus aureus, coagulase negative staphylococcus (CoNS) 1, CoNS 2, Escherichia coli, Hemophilus influenzae, Listeria monocytogenes, Streptococcus pneumoniae, Streptococcus agalactiae, Bacteroides fragilis, Bacillus, Meningococcus, Corynebacterium, Propionibacterium; (2) Blood specimens from 285 cases of suspected septicemia were cultured and bacterial 16S rRNA gene was detected separately; DNA isolated from blood specimens and cerebrospinal fluid was amplified by PCR, and PCR products were hybridized with the probes on the gene chips. Hybridization results were scanned and read by laser-scanner.


RESULTS
(1) Of the 285 cases, 17 were positive by PCR and the positive rate (5.96%) was significantly higher than that of blood culture (2.81%) (P < 0.01). When blood culture was taken as control, the sensitivity of PCR was 100% and Specificity was 96.75%, the index of accurate diagnosis was 0.968. (2) The 17 specimens which showed positive results by PCR were further hybridized on the gene chip. All were positive by universal probes. Among all of them, 5 were positive by E. coli probe; 4 were positive by Staphylococcus epidermidis; two were positive by Bacillus and Propionibacterium probes, separately; 4 were positive by CoNS. The 8 specimens which showed positive results by both PCR and blood culture, the result of gene chip hybridization coincided with the result of blood culture.


CONCLUSIONS
Detection of the bacterial 16SrRNA genes in clinical specimens by gene chip hybridization technology can diagnose neonatal septicemia rapidly. This method has higher sensitivity and specificity than blood culture or other methods and can provide a rapid way for the etiological diagnosis of neonatal septicemia. Therefore the genechip method may be valuable and practical in early diagnosis of neonatal septicemia.
Department of Internal Medicine, Children's Hospital Affiliated to Medical College, Zhejiang University, Hangzhou 310003, China.
Mei-qin
MQ
Tong
Shi-qiang
SQ
Shang
Yi-dong
YD
Wu
Zheng-yan
ZY
Zhao
D020459:Genes, rRNA; D006801:Humans; D007231:Infant, Newborn; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D018805:Sepsis / Q000175:diagnosis*; D013997:Time Factors