Development and evaluation of loop-mediated isothermal amplification (LAMP) for the rapid diagnosis of Candida parapsilosis

The Candida parapsilosis family has emerged as a major opportunistic and nosocomial pathogen. It causes multifaceted pathology in immuno-compromised and normal hosts, notably low birth weight neonates. In the present study, a novel method, known as loop-mediated isothermal amplification (LAMP), was described for the rapid and specific detection of the species, using primer sets derived from the 5.8 S ribosomal RNA gene of C. parapsilosis (internal transcribed spacer 2, ITS2). Amplification products can be detected macroscopically by visual inspection in vials using SYBRGreen I as well as by electrophoresis on agarose gel. The LAMP assay resulted in specific amplification of the ITS2 of C. parapsilosis using pure cultures after a 45-min reaction at 65°C; no cross-reactivity with other fungi including other Candida species was observed. The detectable DNA limit was 0.01 pg fungal DNA per reaction, equivalent to 3.74 × 10-3cfu/ml. In addition, specific amplification was achieved using 30 proven C. parapsilosis strains from patients samples. The method provides a powerful tool for rapid diagnostics in the clinical laboratory, and has potential for use in ecological studies.