Discrimination of Candida nivariensis and Candida bracarensis among Candida glabrata sensu lato isolates from clinical isolates in Tunisia

Background: Candida nivariensis and Candida bracarensis are new species of Candida that are phenotypically similar to Candida glabrata sensu stricto, causing significant problems for their identification by traditional laboratory methods. This study used a singleplex PCR method for the rapid identification of members of the Candida glabrata species complex ( Candida glabrata sensu stricto, Candida nivariensis and Candida bracarensis ). Furthermore, we tried to choose an appropriate extraction method, which is an important factor for the success of the PCR approach. Methods A total of 163 clinical isolates cultured from urine samples, vaginal swabs, placenta, intrauterine device, and urinary catheter in patients from the Maternity and Neonatology Center of Monastir were screened. A singleplex PCR was used targeting the RPL31 gene for the discrimination between species of the Candida glabrata complex. Four different DNA extraction methods, two commercial kits (GF-1Tissue/Blood), the phenol–chloroform isoamylic method, and chelating resin, were applied to obtain and determine the most effective DNA extraction method. The DNA quantity and quality were determined using Nanodrop and PCR. Results The Singleplex PCR assay amplified a 1.061 bp amplicon from all 163 Candida glabrata sensu stricto isolates, thus identifying all clinical isolates in Tunisia as Candida glabrata sensu stricto. Low DNA concentrations were measured for all methods, and the results showed that with one method, PCR success was 100%. The results of DNA purity and quantity measurements show variant results. Conclusion Our results obtained from a collection of clinical Candida glabrata sensu lato isolates show that Candida nivariensis and Candida bracarensis are not clinically important or prevalent in Tunisia. For the extraction method, Chelex (chelating resin) turned out to be a rapid, low-cost method that can provide high-quality DNA.