Identification of bacteria in paraffin-embedded tissues using 16S rDNA sequencing from a neonate with necrotizing enterocolitis.
The conventional approach to bacterial identification in paraffin-embedded tissue relies mainly on morphology, with the aid of Gram stain. This approach is only able to provide some clues; it does not offer the capability of accurate identification. Bacterial identification based on sequencing of 16S ribosomal DNA (rDNA) is able to identify bacteria to the species level. Here we demonstrate the application of this technique in the postmortem examination, for which we had obtained different results from premortem and postmortem blood cultures. In the postmortem examination, abundant gram-negative rods were present in gastrointestinal tract lumen and lymphatic space. DNA was extracted from paraffin-embedded tissue of the above sections and subjected to 16S rDNA polymerase chain reaction for the first 500 base pairs, followed by sequencing. The results of sequencing correlated well with the postmortem culture result. In this review, the newly emerging field of bacterial identification with molecular techniques employing both broad-range and targeted approaches and their clinical applications and limitations are discussed.
He
Mai
M
Department of Pathology, Women & Infants Hospital of Rhode Island, Providence, RI, USA. mhe@wihri.org
Hong
Tao
T
Lauro
Patricia
P
Pinar
Halit
H
D001344:Autopsy; D004269:DNA, Bacterial / Q000302:isolation & purification*; D004275:DNA, Ribosomal / Q000302:isolation & purification*; D020345:Enterocolitis, Necrotizing / Q000175:diagnosis* / Q000382:microbiology*; D004926:Escherichia coli; D004927:Escherichia coli Infections / Q000150:complications / Q000175:diagnosis*; D006801:Humans; D007231:Infant, Newborn; D008297:Male; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000302:isolation & purification
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S