Role of molecular diagnostics in ocular microbiology

Abstract

Although microbial culture remains the gold standard for diagnosis of many ocular infections, the technique is limited by low yield, inability to detect certain organisms, and potentially long delays to results. DNA-based molecular diagnostic techniques use detection of specific nucleic acid sequences as evidence for presence of suspected pathogens. The polymerase chain reaction (PCR) is a powerful molecular biology technique that allows for detection of fewer than 10 copies of pathogen genome. Recent technical advances in PCR have permitted quantitation of pathogen load using quantitative PCR (qPCR), and have permitted multiplexing of primer sets. Use of pan-bacterial and pan-fungal primers for ribosomal DNA sequences has allowed diagnosis of bacterial and fungal infections using molecular techniques. In this review, we highlight recent advances in the application of PCR to the diagnosis of anterior segment and posterior segment ocular infectious diseases.

Keywords: Acanthamoeba; Ocular infection; Toxoplasmosis; glaucomatocyclitic crisis; keratitis; polymerase chain reaction; retinitis; uveitis.

Figure 1

Schematic of the polymerase chain reaction. DNA to be detected (black) is incubated with oligonucleotide primers (red and blue) complementary to sequences in the suspected pathogen. Hybridization of primers allows DNA synthesis by a thermostable polymerase. PCR is an exponential process; after N rounds, 2^N copies of the starting material will be generated in theory.