Detection of common candidemia pathogens based on PCR combined with MALDI-TOF MS / 中华检验医学杂志

ABSTRACT

Objective:A high-throughput assay for the detection of five common clinical Candidaemia pathogens was established by combining polymerase chain reaction (PCR) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS).Method:Establishment of methodology. We selected Candida albicans, Candida parapsilosis, Candida glabrata, Candida krusei, Candida tropicalis to be the target pathogens and the internal transcribed spacer (ITS) region as the target gene. Specific single base extension primers were designed to perform single base extension reaction in the same reaction system. MALDI-TOF MS was used to detect the characteristic peaks of each target pathogen. The sensitivity and specificity of the detection system were verified by using spiked blood samples. Totally 108 blood samples from proven or suspected candidaemia patients were collected from October 2021 to September 2022 in a hospital in Beijing. The results of nucleic acid mass spectrometry were compared with those of clinical blood culture. Results:The established nucleic acid mass spectrometry detection system can simultaneously detect five common clinical Candida species. Each strain can produce specific product peaks and there is no mutual interference between the strains. The detection limit of Candida albicans was 100 CFU/ml. The detection limit of Candida parapsilosis, Candida glabrata, Candida krusei and Candida tropicalis was 10 CFU/ml. For the 108 blood samples, the sensitivity, specificity, positive predictive value and negative predictive value of nucleic acid mass spectrometry were 94.74% (36/38), 97.14% (68/70), 92.31% (36/39) and 98.55% (68/69), respectively. The McNemar χ 2 test showed no significant difference between the two methods ( P>0.05), and the Kappa consistency test showed good consistency between the two methods ( Kappa=0.9, P<0.05). Conclusion:A nucleic acid mass spectrometry detection system suitable for clinical candida detection was successfully constructed, and the method validation results were consistent with the clinical blood culture.