Specific detection of fungal pathogens by 18S rRNA gene PCR in microbial keratitis

Zunaina Embong¹ , Wan Hazabbah Wan Hitam¹ , Chan Yean Yean² , Nur Haslindawaty Abdul Rashid³ , Balqis Kamarudin⁴ , Siti Khaironi Zainal Abidin⁵ , Sabariah Osman² , Zainul F Zainuddin³ and Manickam Ravichandran²

¹Department of Ophthalmology, School of Medical Sciences, Universiti Sains Malaysia, Malaysia

²Department of Medical Microbiology and Parasitology, School of Medical Sciences, Universiti Sains Malaysia, Malaysia

³School of Health Sciences, Universiti Sains Malaysia, Malaysia

⁴Universiti Malaya, Malaysia

⁵Universiti Teknologi Malaysia, Malaysia

author email corresponding author email

BMC Ophthalmology 2008, 8:7doi:10.1186/1471-2415-8-7


Published:	29 April 2008

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.

Conclusion

PCR is a rapid, sensitive and useful method to detect fungal aetiology in microbial keratitis.