Establishment and application of loop-mediated isothermal amplification for Candida albicans / 临床检验杂志

ABSTRACT

Objective@#To establish a simple and rapid loop-mediated isothermal amplification (LAMP) to detect Candida albicans (CA) and explore its clinical value.@*Methods@#The Primer Explorer 5.0 software was used to design 4 primers for amplification of CA by LAMP. The system and conditions of LAMP reaction were optimized to evaluate its specificity and the minimum limit in the detection. The vaginal swabs were collected from 123 vulvovaginal candidiasis (VVC) patients and 42 healthy individuals. Fungal culture, LAMP test, PCR test and 1.79 mol/L KOH microscopy were conducted in parallel. Fungal culture was used as the reference method for VVC diagnosis. The positive rates between two groups were compared by Pearson χ² test. The consistency of the results from LAMP, PCR, microscopy and culture were analyzed by Kappa test and P＜0.05 was considered as statistically significant difference. ROC curve analysis was used to evaluate LAMP and PCR test for diagnosis of VVC.@*Results@#The optimum reaction temperature of LAMP was 61 ℃ with high specificity. No cross reaction with other strains was found. The minimum detection limit was 10³ copies/ul. The positive rates of LAMP, PCR and microscopy between VVC and healthy group showed statistically significant difference（LAMP χ²=68.576；PCR χ²=64.918；microscopy χ²=50.076，P＜0.01）. LAMP detection and PCR showed good consistency (κ=0.744, 0.720), but microscopy examination showed poor consistency (κ=0.533). LAMP showed diagnostic sensitivity of 87.62%, specificity of 88.33%, positive predictive value of 92.93% and negative predictive value of 80.30%. The area under the curves of LAMP and PCR were 0.873 and 0.888, respectively. No difference in efficacies between LAMP and PCR was found (Z=0.849, P=0.395 6), but the lowest detection time of LAMP was shorter than 1 hour.@*Conclusion@#A rapid, reliable, sensitive and specific LAMP technique for detecting CA was established. The comprehensive screening performance should be superior to the routine method in laboratories, so LAMP could be used for the supplementary diagnosis and monitoring therapeutic effects in CA infection.