Rolling Circle Amplification (RCA): an approach for quick detection and identification of fungal species

Conventional methods for fungal identification in the clinical laboratory rely on morphological and physiological tests. These tests often need several days or weeks to complete and are frequently unspecific. Molecular identification mostly implies sequencing, which is relatively expensive and time-consuming, as well as impractical for large numbers of isolates. The Rolling Circle Amplification approach, known as RCA, is a quick, critical and economic method for fungal species' identification. Despite its high speed, this method is highly sensitive, and it has been widely used for the detection of pathogenic fungi. The specific probes are designed based on the differences in the nucleotide regions of the target gene for the target species. The amplification product can be visualized by agarose gel electrophoresis, but can also be visualized in gel-free systems using fluorescence staining of the amplified product by SYBR Green in combination with a UV transilluminator. Thus, the simplicity, sensitivity, robustness and low costs make RCA an attractive technique for the reliable identification of sibling species and other closely related fungi.