Comparing Molecular Methods for Early Detection and Serotyping of Enteroviruses in Throat Swabs of Pediatric Patients

Pai-Shan Chiang,Min-Shi Lee,Shu-Ting Luo,Tzou-Yien Lin,Kuo-Chien Tsao,Mei-Liang Huang

doi:10.1371/journal.pone.0048269

Abstract

BackgroundEnteroviruses include over 100 serotypes and usually cause self-limited infections with non-specific symptoms in children, with the exceptions of polioviruses and enterovirus 71 which frequently cause neurologic complications. Therefore, early detection and serotyping of enteroviruses are critical in clinical management and disease surveillance. Traditional methods for detection and serotyping of enteroviruses are virus isolation and immunofluorescence assay, which are time-consuming. In this study, we compare virus isolation and two molecular tests for detection and serotyping of enteroviruses in clinical samples.MethodsOne hundred and ten throat swabs were collected from pediatric outpatients with enterovirus-like illnesses (hand-foot-mouth disease, herpangina, and non-specific febrile illness). Virus isolation was conducted using multiple cell lines and isolated viruses were serotyped using immunofluorescent assay. In the molecular tests, a semi-nested RT-PCR and a novel CODEHOP platform were used to detect the 5′UTR and VP1 genes of enteroviruses, respectively. Amplified nucleotides were sequenced and genotyped.ResultsAmong the 110 cases, 39(35%), 52(47%), and 46(42%) were tested positive with these three tests, respectively. Using the consensus results of these three tests as the gold standard, agreement of the VP1 CODEHOP test was 96%, which is higher than those of the virus isolation (89%) and the 5′-UTR test (88%). The VP1 CODEHOP test also has the best performance on serotyping confirmed with serum neutralization tests.ConclusionsThe VP1 CODEHOP test performed well for detection and serotyping of enteroviruses in clinical specimens and could reduce unnecessary hospitalization cares during enterovirus seasons.

Highlights

Human enteroviruses include over 100 serotypes and can be divided into four species using molecular typing[1]
Early detection and serotyping of enteroviruses infections are critical in clinical management and disease surveillance
Several clinical studies have documented that molecular diagnosis based on polymerase chain reaction (PCR) is time-saving and more sensitive than virus isolation for detection of enterovirus infections in hospitalized patients [6,7] [8,9,10] but few studies have been conducted in outpatients

Summary

Introduction

Human enteroviruses include over 100 serotypes and can be divided into four species using molecular typing[1]. The traditional standard methods for detection and serotyping of enterovirus infections are virus isolation and immunofluorescence assay (IFA), which are time-consuming and labor-intensive [4,5]. No study has compared molecular tests and virus isolation/IFA for serotyping of human enteroviruses using clinical specimens These methods had been used to detect enteroviruses in clinical specimens including throat swabs, stool samples and cerebrospinal fluid (CSF) [5,6,7,9,10,11,12], reports elucidating the comparison among the diagnostic approaches are limited. Enteroviruses include over 100 serotypes and usually cause self-limited infections with non-specific symptoms in children, with the exceptions of polioviruses and enterovirus 71 which frequently cause neurologic complications. We compare virus isolation and two molecular tests for detection and serotyping of enteroviruses in clinical samples

Methods

Results

Conclusion