title: Specific detection of fungal pathogens by 18S rRNA gene PCR in microbial keratitis.
abstract: BACKGROUND
  The sensitivity and specificity of 18S rRNA polymerase chain reaction (PCR) in the detection of fungal aetiology of microbial keratitis was determined in thirty patients with clinical diagnosis of microbial keratitis.
  
  
  METHODS
  Corneal scrapings from patients were used for Gram stain, culture and PCR analysis. PCR was performed with primer pairs targeted to the 18S rRNA gene. The result of the PCR was compared with conventional culture and Gram staining method. The PCR positive samples were identified by DNA sequencing of the internal transcribed spacer (ITS) region of the rRNA gene. Main outcome measures were sensitivity and specificity of PCR in the detection of fungus in corneal keratitis.
  
  
  RESULTS
  Combination of microscopy and culture gave a positive result in 11 of 30 samples of microbial keratitis. PCR detected 10 of 11 samples that were positive by conventional method. One of the 19 samples that was negative by conventional method was positive by PCR. Statistical analysis revealed that the PCR to have a sensitivity of 90.9% and specificity of 94.7% in the detection of a fungal aetiology in microbial keratitis.
  
  
  CONCLUSIONS
  PCR is a rapid, sensitive and useful method to detect fungal aetiology in microbial keratitis.
authors.affiliation: Department of Ophthalmology, School of Medical Sciences, Universiti Sains Malaysia, Malaysia. zunaina@yahoo.com
authors.forename: Zunaina
authors.initials: Z
authors.lastname: Embong
authors.forename: Wan Hazabbah
authors.initials: WH
authors.lastname: Wan Hitam
authors.forename: Chan Yean
authors.initials: CY
authors.lastname: Yean
authors.forename: Nur Haslindawaty Abdul
authors.initials: NH
authors.lastname: Rashid
authors.forename: Balqis
authors.initials: B
authors.lastname: Kamarudin
authors.forename: Siti Khaironi Zainal
authors.initials: SK
authors.lastname: Abidin
authors.forename: Sabariah
authors.initials: S
authors.lastname: Osman
authors.forename: Zainul F
authors.initials: ZF
authors.lastname: Zainuddin
authors.forename: Manickam
authors.initials: M
authors.lastname: Ravichandran
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D003315:Cornea / Q000382:microbiology*; D003430:Cross-Sectional Studies; D004271:DNA, Fungal; D015821:Eye Infections, Fungal / Q000175:diagnosis*; D005260:Female; D005658:Fungi / Q000235:genetics*; D006801:Humans; D007634:Keratitis / Q000382:microbiology* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D012337:RNA, Ribosomal, 18S / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D017421:Sequence Analysis; D013997:Time Factors
chemical_list: D004271:DNA, Fungal; D012337:RNA, Ribosomal, 18S