Rapid detection of <i>Candida albicans</i> in oral exfoliative cytology samples by loop-mediated isothermal amplification

Abstract

Loop-mediated isothermal amplification(LAMP) rapidly amplifies DNA under isothermal conditions. The aim of this study was to detect Candida albicans and compare the positivity rate in the LAMP reaction with that of conventional methods for oral exfoliative cytology (EC) samples. Sixty-eight EC samples from 53 patients were subjected to LAMP analysis. These patients had been clinically diagnosed with leukoplakia, squamous cell carcinoma, oral lichen planus (OLP), stomatitis, oral candidiasis, and other malignancies. LAMP reactions were defined as positive when the sample turbidity exceeded 0.1 (arbitrary unit). Periodic acid-Schiff (PAS) staining and microbial culture were also performed to detect Candida species in EC samples. The LAMP reaction detected C. albicans in 42.6% of EC samples. Candida species were detected in 32.4% of the same samples by culturing and in 29.4% of samples by PAS staining. C. albicans DNA was detected most frequently in samples from OLP patients. We conclude that, in comparison to conventional methods for detection of C. albicans, the LAMP method is highly sensitive and timesaving, and does not require expensive equipment or diagnostic technology. It may therefore be useful for on-site screening of C. albicans at dental clinics.

Identification of Mycoses in Developing Countries

“…In next-generation LAMP, which employs loop primers, the original LAMP amplification time was significantly reduced (up to 30 min) [71]. Rapidity (less than 1 h), high sensitivity (detects up to six target copies), resistance to inhibitors, and easy visualization provided by simple amplicon detection systems [72] make LAMP one of the most popular isothermal assays for identification of a wide range of clinically important fungi from pure culture [73] as well as from environmental [74] and clinical [75][76][77][78][79] samples. The technique has been successfully optimized and validated for detection of C. auris from simulated environmental samples and clinical specimens and in some cases, it demonstrated superiority over quantitative real-time PCR (qRT-PCR) for direct identification of A. fumigatus in clinical samples [75,78].…”

Establishment and Application of Multiple Cross Displacement Amplification Coupled With Lateral Flow Biosensor (MCDA-LFB) for Visual and Rapid Detection of Candida albicans in Clinical Samples

“…To compare the sensitivity of MCDA with that of LAMP and qPCR assays for pure cultures, template DNA from the reference strains was diluted from 2 ng/μl to 2 fg/μl. For LAMP assays, the Loopamp Specified Microorganism “Fungi Candida albicans” Detection Kit (Eiken Chemical Co., Tokyo, Japan) was used in accordance with the manufacturer's instructions (Noguchi et al, 2017). In brief, the primer sequence for LAMP assay is targeting ITS region, which has obtained the patent NO.…”

“…Thus, it is imperative to develop and validate a rapid and accurate method for C. albicans identification. Several non-culture methods have been developed for C. albicans detection, including mass spectrometry (Zehm et al, 2012), immunoassay (Gunasekera et al, 2015), polymerase chain reaction (PCR) (Vahidnia et al, 2015), real-time PCR (Maaroufi et al, 2003; Kasai et al, 2006), polymerase spiral reaction (PSR) (Jiang et al, 2016), and loop-mediated isothermal amplification (LAMP) (Noguchi et al, 2017). Compared with the gold-standard culture method, these tests for C. albicans identification can save considerable time.…”

Molecular diagnostics in medical mycology

“…LAMP also uses four primers instead of the usual two, which provides a greater specificity than the typical pair of primers, although primer design is more challenging. LAMP has been applied to many of the major fungal pathogens 51 – 53 . A second type of isothermal amplification method used for fungal identification is called Nucleic Acid Sequence-Based Amplification (NASBA), which uses RNA as the template.…”