|
Abstract
|
Loop mediated isothermal amplification (LAMP) technique is a good novel candidate for the fast molecular diagnosis of C. albicans infection. The aim of this study was to evaluate an enhanced LAMP-based method for rapid diagnosis of Candida albicans. We used 35 and 89 yeast isolates from oral cavities of healthy people and patients with OPC, respectively. The results of gold standard and conventional identification experiments including physiochemical, microscopic examination, culture-based and PCR methods were compared to conventional LAMP and a new enhanced LAMP method. We have introduced an enhanced protocol of LAMP employing uracil DNA glycosylase (UDG) treatment, a closed-tube system, non-denatured DNA and the known DNA amplification enhancers; DMSO and tetramethyl ammonium (TMA) oxalate, to decrease false positive results and increase specificity and sensitivity of LAMP assays. Old-school phenotypical, physiochemical and culture-based examinations were failed by wrong identification of 12 isolates. Eight of them were C. dubliniensis isolates wrongly identified as C. albicans. Our established enhanced closed-tube UDG- LAMP (ECT UDG-LAMP) method could successfully distinguish among these genetically close species. Moreover, 5 LAMP products were sequenced. Sequence analysis showed the high similarity between our native ITS sequences and other fungal ITS sequences. In conclusion, the results revealed promising potential of using ECT UDG-LAMP technique as a high specificity, cost-effective and user-friendly laboratory diagnosis method which could be done directly on non-purified specimens containing mixed fungal populations such as oral washes.
|