title: 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.
abstract: 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.
  
  
  OBJECTIVE
  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.
  
  
  METHODS
  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' 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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Jiazhen
authors.initials: J
authors.lastname: Chen
authors.affiliation: Department of Medical Microbiology and Parasitology, Shanghai Medical College, Fudan University, Shanghai 200032, China.
authors.forename: Yongfeng
authors.initials: Y
authors.lastname: Fu
authors.affiliation: The MOE Key Laboratory of Public Health Safety, School of Public Health, Fudan University, Shanghai 200032, China.
authors.forename: Liwen
authors.initials: L
authors.lastname: Ju
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Xinyu
authors.initials: X
authors.lastname: Miao
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Yaojie
authors.initials: Y
authors.lastname: Shen
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Lei
authors.initials: L
authors.lastname: He
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Wenjie
authors.initials: W
authors.lastname: Wang
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Jialin
authors.initials: J
authors.lastname: Jin
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Lingyun
authors.initials: L
authors.lastname: Shao
authors.affiliation: Ibis Biosciences, Inc., An Abbott Company, Carlsbad, CA, USA.
authors.forename: Rangarajan
authors.initials: R
authors.lastname: Sampath
authors.affiliation: Ibis Biosciences, Inc., An Abbott Company, Carlsbad, CA, USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Ecker
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Min
authors.initials: M
authors.lastname: Li
authors.affiliation: Department of Medical Microbiology and Parasitology, Shanghai Medical College, Fudan University, Shanghai 200032, China. Electronic address: xjcheng@shmu.edu.cn.
authors.forename: Xunjia
authors.initials: X
authors.lastname: Cheng
authors.affiliation: Department of Infectious Diseases, Huashan Hospital, Fudan University, Shanghai 200040, China. Electronic address: zhangwenhong@fudan.edu.cn.
authors.forename: Wenhong
authors.initials: W
authors.lastname: Zhang
mesh_terms: D002675:Child, Preschool; D004770:Enterovirus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D005260:Female; D006232:Hand, Foot and Mouth Disease / Q000175:diagnosis* / Q000821:virology; D006801:Humans; D007223:Infant; D008297:Male; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014773:Virology / Q000379:methods*
keywords: CMV; CSF; CVA; EBV; ECHO; EV; Enterovirus; Epstein–Barr virus; HFMD; HHV; HSV; RT-PCR/ESI-MS; VZV; cerebrospinal fluid; coxsackievirus A; cytomegalovirus; echovirus; enterovirus; hand, foot and mouth disease; herpes simplex virus; human herpesvirus; reverse transcription-PCR coupled with electrospray ionization mass spectrometry; varicella zoster virus