Analysis of Inteins in the Candida parapsilosis Complex for Simple and Accurate Species Identification

Abstract

cInteins are coding sequences that are transcribed and translated with flanking sequences and then are excised by an autocatalytic process. There are two types of inteins in fungi, mini-inteins and full-length inteins, both of which present a splicing domain containing well-conserved amino acid sequences. Full-length inteins also present a homing endonuclease domain that makes the intein a mobile genetic element. These parasitic genetic elements are located in highly conserved genes and may allow for the differentiation of closely related species of the Candida parapsilosis (psilosis) complex. The correct identification of the three psilosis complex species C. parapsilosis, Candida metapsilosis, and Candida orthopsilosis is very important in the clinical setting for improving antifungal therapy and patient care. In this work, we analyzed inteins that are present in the vacuolar ATPase gene VMA and in the threonyl-tRNA synthetase gene ThrRS in 85 strains of the Candida psilosis complex (46 C. parapsilosis, 17 C. metapsilosis, and 22 C. orthopsilosis). Here, we describe an accessible and accurate technique based on a single PCR that is able to differentiate the psilosis complex based on the VMA intein. Although the ThrRS intein does not distinguish the three species of the psilosis complex by PCR product size, it can differentiate them by sequencing and phylogenetic analysis. Furthermore, this intein is unusually present as both mini-and full-length forms in C. orthopsilosis. Additional population studies should be performed to address whether this represents a common intraspecific variability or the presence of subspecies within C. orthopsilosis.

Changing epidemiology of candidaemia: Increase in fluconazole‐resistant Candida parapsilosis

“…Differentiation of C parapsilosis into C parapsilosis sensu stricto, C orthopsilosis and C metapsilosis was performed using the polymerase chain reaction (PCR) method. Gene research has been studied using primers and conditions described by Prandini et al…”

Species-specific detection of Candida tropicalis using evolutionary conserved intein DNA sequences

“…Previously, the use of intein sequence for molecular identification of C. parapsilosis complex using threonyl‐tRNA synthetase ( ThrRS ) gene was reported (Prandini et al . ). We have also shown the specificity of VMA intein sequence for specific detection of C. glabrata (Kumar and Ramesh ).…”

“…Vacuolar membrane ATPase are family of proteins present in all domains of life and are one of the highly conserved protein (Prandini et al . ; Swithers et al . ).…”

Identification and Differentiation of Candida parapsilosis Complex Species by Use of Exon-Primed Intron-Crossing PCR

“…Thus, a four-species complex has been suggested to replace the three-species complex (19). Thus, it is clinically important to distinguish among the members of this species complex.So far, various molecular approaches, such as PCR-restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA, real-time PCR, PCR with specific primers, matrixassisted laser desorption ionization-time of flight mass spectrometry, sequencing analysis, and a simple PCR developed recently, were established to differentiate the former three-species complex (6,(21)(22)(23)(24)(25)(26)(27). Although turquoise blue colonies on BBL CHROMagar Candida medium (Becton, Dickinson and Company, Sparks, MD) or sequence analysis could be used to distinguish L. elongisporus from the three-species complex (19), no other molecular tool has been well established.…”