Direct detection of Corynebacterium striatum, Corynebacterium propinquum, and Corynebacterium simulans in sputum samples by high-resolution melt curve analysis

• Published in: BMC infectious diseases Vol. 21; no. 1; p. 21
• Main Authors: Xu, Shuai, Qiu, Xiaotong, Hou, Xuexin, Zhou, Haijian, Chen, Dongke, Wang, Xuebing, Han, Lichao, Li, Dan, Sun, Lina, Ji, Xingzhao, Li, Minghui, Zhang, Jingshan, Li, Mengtong, Li, Zhenjun
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 07.01.2021; BioMed Central; BMC
• Subjects: Assaying; Bacterial identification; Bacterial Proteins - genetics; Clinical isolates; Confidence intervals; Corynebacteria; Corynebacterium; Corynebacterium - genetics; Corynebacterium - isolation & purification; Corynebacterium Infections - diagnosis; Corynebacterium Infections - microbiology; Corynebacterium species; Deoxyribonucleic acid; Diagnostic software; Diagnostic systems; Diagnostics; DNA; DNA Primers - genetics; Epidemiology; Genetic diversity; Genotyping Techniques - methods; High resolution; High-resolution melt curve analysis; Humans; Identification and classification; Laboratories; Limit of Detection; Mass spectrometry; Melting; Melting curve; Methods; Microbiological assay; Microorganisms; Nosocomial infections; Pathogens; Polymerase chain reaction; Real-Time Polymerase Chain Reaction - methods; Scientific imaging; Software; Species; Sputum; Sputum - microbiology; Testing; Varieties; China; France; United States > US; Germany; Diagnostics; High-resolution melt curve analysis; Bacterial identification; Corynebacterium species
• ISSN: 1471-2334; 1471-2334
• DOI: 10.1186/s12879-020-05633-z

Pulmonary infections caused by non-diphtheriae corynebacteria are increasing. However, rapid identification of Corynebacterium species poses a challenge due to the low genetic variation within the genus. Three reference strains and 99 clinical isolates were used in this study. A qPCR followed by high-resolution melting (HRM) targeting ssrA was performed to simultaneously identify C. striatum, C. propinquum and C. simulans. To further evaluate this assay's performance, 88 clinical sputum samples were tested by HRM and the detection results were compared with those of the traditional culture method and multiple cross-displacement amplification (MCDA) assay. The melting curve produced by a pair of universal primers generated species-specific HRM curve profiles and could distinguish the three target species from other related bacteria. The limit of detection of HRM assay for DNA from the three purified Corynebacterium species was 100 fg. Compared with the culture method, HRM detected 22 additional positive specimens, representing a 23.9% relative increase in detection rate. The HRM assay had 98.4% (95% confidence interval [CI], 90.5-99.9%) sensitivity and 100% (95% CI, 82.8-100%) specificity. Additionally, 95.5% concordance between HRM and MCDA (κ = 0.89 [95% CI, 0.79-0.99]) was noted. The HRM assay was a simple, rapid, sensitive, and specific diagnostic tool for detecting C. striatum, C. propinquum, and C. simulans, with the potential to contribute to early diagnosis, epidemiological surveillance, and rapid response to outbreak.