Detection and identification of viral pathogens in patients with hand, foot, and mouth disease by multilocus PCR, reverse-transcription PCR and electrospray ionization mass spectrometry

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• 作者：Jiazhen Chen ; Yongfeng Fu ; Liwen Ju ; Xinyu Miao ; Yaojie Shen ; Lei He ; Wenjie Wang ; Jialin Jin ; Lingyun Shao ; Rangarajan Sampath ; David J. Ecker ; Ying Zhang ; Min Li ; Xunjia Cheng ; Wenhong Zhang
• 关键词：RT-PCR/ESI-MS ; reverse transcription-PCR coupled with electrospray ionization mass spectrometry ; HFMD ; hand ; foot and mouth disease ; EV ; enterovirus ; CVA ; coxsackievirus A ; ECHO ; echovirus ; CSF ; cerebrospinal fluid ; HSV ; herpes simplex virus ; VZV ; varicella zoster virus ; EBV ; Epstein&ndash ; Barr virus ; CMV ; cytomegalovirus ; HHV ; human herpesvirus
• 刊名：Journal of Clinical Virology
• 出版年：February, 2014
• 年：2014
• 卷：59
• 期：2
• 页码：115-119
• 全文大小：631 K

文摘

Background

Rapid detection and identification of viruses are important for early diagnosis and effective surveillance of hand, foot, and mouth disease (HFMD). We described a novel assay using multilocus PCR and reverse transcription-PCR coupled with electrospray ionization mass spectrometry (RT-PCR/ESI-MS) to simultaneously detect and identify human enterovirus A-D, adenovirus A-F, human herpesvirus 1-8, parvovirus B19 and polyomavirus.

Objectives

To evaluate the accuracy and efficacy of the RT-PCR/ESI-MS method, to detect and type enterovirus from specimens of clinical diagnosed HFMD patients.

Study design

In this study, 152 specimens of clinically diagnosed HFMD patients were studied by the assay using RT-PCR/ESI-MS method. The detection and typing of enterovirus by RT-PCR/ESI-MS were compared with results from reference molecular methods.

Results

The assay detected enteroviruses in 97 (63.8%) specimens, resulting in a sensitivity of 93.8% (95% CI: 91.8-95.7%) and a specificity of 87.5% (95% CI: 84.8-90.2%) compared with a reference clinical diagnostic test. Most enterovirus genotypes (65/84; 77%) determined by the platform were consistent with 5鈥?UTR sequence analysis, and most misidentifications resulted from the virus library, which could be further improved by updating the enterovirus database. In addition to enteroviruses, herpesviruses, polyomaviruses, adenoviruses and human rhinoviruses were detected and identified in 55 (36%) HFMD specimens by RT-PCR/ESI-MS.

Conclusion

With the capability of high throughput and detection and typing of multiple clinically relevant viruses simultaneously, RT-PCR/ESI-MS can be a rapid and robust laboratory tool for identifying viral pathogens.