title: Nested polymerase chain reaction (PCR) targeting 16S rDNA for bacterial identification in empyema.
abstract: BACKGROUND
  Empyema in children causes significant morbidity and mortality. However, identification of organisms is a major concern.
  
  
  OBJECTIVE
  To detect bacterial pathogens in pus specimens of children with empyema by 16S rDNA nested polymerase chain reaction (PCR) and correlate it with culture and sensitivity.
  
  
  METHODS
  Sixty-six children admitted to the paediatric ward with a diagnosis of empyema were enrolled prospectively. Aspirated pus was subjected to cytochemical examination, culture and sensitivity, and nested PCR targeting 16S rDNA using a universal eubacterial primer.
  
  
  RESULTS
  Mean (SD) age was 5·8 (1·8) years (range 1-13). Analysis of aspirated pus demonstrated total leucocyte count >1000×10(6)/L, elevated protein (≧20 g/L) and decreased glucose (≤2·2 mmol/L) in 80·3%, 98·5% and 100%, respectively. Gram-positive cocci were detected in 29 (43·9%) and Gram-negative bacilli in two patients. Nested PCR for the presence of bacterial pathogens was positive in 50·0%, compared with 36·3% for culture.
  
  
  CONCLUSIONS
  16S rDNA PCR improves rates of detection of bacteria in pleural fluid, and can detect bacterial species in a single assay as well as identifying unusual and unexpected causal agents.
authors.forename: Rajniti
authors.initials: R
authors.lastname: Prasad
authors.forename: Chhaya
authors.initials: C
authors.lastname: Kumari
authors.forename: B K
authors.initials: BK
authors.lastname: Das
authors.forename: Gopal
authors.initials: G
authors.lastname: Nath
mesh_terms: D000293:Adolescent; D001419:Bacteria / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D001424:Bacterial Infections / Q000175:diagnosis* / Q000382:microbiology; D001431:Bacteriological Techniques / Q000379:methods*; D002648:Child; D002675:Child, Preschool; D003430:Cross-Sectional Studies; D004275:DNA, Ribosomal / Q000235:genetics*; D004653:Empyema / Q000175:diagnosis* / Q000382:microbiology; D005260:Female; D006801:Humans; D007223:Infant; D008297:Male; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011446:Prospective Studies; D012336:RNA, Ribosomal, 16S / Q000235:genetics
chemical_list: D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S
keywords: Culture,; Empyema,; Nested PCR