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. 2018 Sep 25;18(1):480.
doi: 10.1186/s12879-018-3381-5.

Identification of nine cryptic species of Candida albicans, C. glabrata, and C. parapsilosis complexes using one-step multiplex PCR

Amir Arastehfar  1 , Wenjie Fang  1   2 , Weihua Pan  3 , Wanqing Liao  2 , Liang Yan  2 , Teun Boekhout  4   5   6
Affiliations

Identification of nine cryptic species of Candida albicans, C. glabrata, and C. parapsilosis complexes using one-step multiplex PCR

Amir Arastehfar et al. BMC Infect Dis. .

Abstract

Background: Candida albicans, Candida glabrata, and Candida parapsilosis are three prevalent causes of candidiasis, worldwide. These species are considered as nine medically important complex species. Limited knowledge about these newly recognized species prompted us to develop a one-step, multiplex PCR to detect and identify them in clinical settings.

Methods: Primers targeting Hyphal Wall Protein I gene for the C. albicans, C. dubliniensis, C. africana, Intergenic Spacer for the C. glabrata, C. nivariensis, C. bracarensis, and Intein and ITS rDNA for the C. parapsilosis, C. orthopsilosis, and C. metapsilosis were designed. Using 168 CBS reference strains and 280 clinical isolates, the specificity and reproducibility of the developed assay were evaluated.

Results: Our developed assay successfully identified and distinguished all the nine species. No cross-reaction with closely- and distantly-related yeast species, Aspergillus species and human DNA was observed, resulting in 100% specificity. The ambiguous results obtained by MALDI-TOF for C. albicans and C. africana were corrected by our 9-plex PCR assay. This assay identified all the cryptic complex species from two test sets from Iran and China, correctly.

Conclusions: Our developed multiplex assay is accurate, specific, cost/time-saving, and works without the tedious DNA extraction steps. It could be integrated into routine clinical laboratories and as a reliable identification tool and has the potential to be implemented into epidemiological studies to broaden the limited knowledge of cryptic species complexes.

Keywords: Candida; Cryptic species; Molecular diagnosis; Multiplex PCR.

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Conflict of interest statement

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

Weihua Pan is the editor of BMC Infectious Diseases and we declare that her co-authorship has not influenced the editorial processes.

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Figures

Fig. 1
Fig. 1
Primer information. List of covered species, their PCR product sizes, and their locations are depicted
Fig. 2
Fig. 2
Amplified PCR products for nine target cryptic species complexes are shown. M. 100 bps Molecular ladder (SM 0324), 1–2 C. albicans (1090 bps), 3–4 C. africana (901 bps), 5–6 C. dubliniensis (721 bps), 7–8 C. nivariensis (586 bps), 9–10 C. orthopsilosis (476 bps), 11–12 C. glabrata (386 bps), 13–14 C. metapsilosis (308 bps), 15–16 C. bracarensis (214 bps), 17–18 C. parapsilosis (112 bps)
See this image and copyright information in PMC

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