The Study On Values Of 16S RRNA Gene In Rapid Diagnosis Of Bacterial Bloodstream Infection In Neonates

Objective Bacterial bloodstream infection(BSI) is a major cause threatening the health even the life of newborn because of the high morbidity and mortality. Up to now, the clinical diagnosis of bacterial BSI is still depended on blood culture, this method has lower positive rate and get result slowly, so it is difficult to make diagnosis early. To find an accurate and rapid assay detecting bacteria directly from specimens is of great value. For an ideal target gene to identify bacteria, 16S rRNA gene has now been considered as an important marker. The aim of this research is to evaluate the diagnostic value of 16S rDNA in neonates with bacterial BSI , meanwhile compaired with blood culture and other non-specific diadynamic criteria.Methods Firstly , a universal primer(UP) was synthesized based on the highly conserved sequences of bacterial 16S rDNA. 37strains of common pathogenic bacteria representing for 19 species were amplified by polymerase chain reaction(PCR), and observed the result with agarose gel electrophoresis(AGE). Next, we tested the specificity and sensitivity of the PCR. Sencondly, blood specimens from 106 cases of suspected bacterial BSI were cultured and DNA isolated from these specimens was amplified by PCR and we also detected 20 cases of non-infected infants. All clinical data were analized using SPSS 11.5 for Windows software. Results The positive fragment with length of 371bp were amplified from all strains. The primer can only amplify bacteria and its sensitivity could be improved to 10CFU/ml(Escherichia coli ATCC25922). No signal was observed when human DNA and viruses were used as templates. Of 106 specimens with suspected bacterial BSI, 15 were positive(14.1%) on blood culture and 33 on PCR(31.1%). The positive rate of PCR was significantly higher than that of blood culture(p<0.01), while 20 specimens of non-infected infants were all negative on PCR. According to diagnostic criteria, 106 neonates suspected bacterial BSI were divided into 2 groups, bacterial BSI 41 cases(involved final andclinical diagnosis) and non bacterial BSI 65. This research revealed that the sensitivity, specificity, positive and negative predictive value ,diagnostic effection as well as diagnostic Index were 82.93%, 96.92%, 94.44% , 90.00%, 91.51% and 179.85 respectively, and the time of PCR spent only for 6~8h, improving the extracting technique of DNA, it can be 4~6h. Conclusions 16S rDNA amplified by PCR can be used to detect the pathogenic bacteria in specimens rapidly. It has higher sensitivity and specificity in diagnosis neonatal bacterial BSI as well as great value in separating bacterial BSI from localized bacterial infection and non-bacterial BSI.