Microsporidial Keratitis in India: 16S rRNA Gene-Based PCR Assay for Diagnosis and Species Identification of Microsporidia in Clinical Samples

Abstract

This study is the first to validate PCR-based assays for detection of microsporidial DNA in corneal scrapings. Pan microsporidian PCR can be a useful adjunct with smear examination in the diagnosis of microsporidial keratitis.

Outbreak of microsporidial keratoconjunctivitis with rugby sport due to soil exposure

“…Nonetheless, PCR allowed the identification of microsporidia in the subspecies level. 19,26 Recently a study showed that 11.4% of patients with microsporidial keratoconjunctivitis have concomitant adenoviral infection. 27 Specific PCR for adenovirus and enterovirus is negative for all of our specimens.…”

“…In addition, PCR is a sensitive way for organism detection, and subsequent sequencing of PCR products is used to identify particular microsporidial species. 19,20 The use of drugs namely albendazole, fumagillin, itraconazole, and propamidine isethionate has been associated with remission of microsporidial keratoconjunctivitis. However, it has been demonstrated to be less effective in stromal keratitis, in which corneal transplantation is usually required.…”

Microsporidial Keratitis

“…Results are excellent on resin embedded sections stained with toluidine blue [31]. Sensitivity of detection by Gram's stain was 90.3%; by Giemsa stain was 64.52% and by modified Ziehl Neelsen stain was 87% as observed by Joseph et al, in their study [32].…”

“…Though this is a rapid and sensitive method it is not specific and many other structures like fungal spores will also fluoresce and may give false positive results [31]. Studies show sensitivity of potassium hydroxide with calcoflour white to be 93.5% [32].…”

“…Apart from small-subunit rRNA, large subunit rRNA and the intergenic spacer region can also be used as a target sequence [32]. Panmicrosporidial primers (V1 and PMP2) and species specific primers (V1-EB450 andV1-SI500) can be used for detecting microsporidia in clinical specimen [35].…”