Abstract
BackgroundEnterovirus (EV) infections are commonly associated with encephalitis and meningitis. Detection of enteroviral RNA in clinical specimens has been demonstrated to improve the management of patients, by ruling out other causes of disease.MethodTo develop a sensitive and reliable assay for routine laboratory diagnosis, we developed a real-time one step reverse transcription polymerase chain reaction (RT-PCR) assay with minor groove binder probes and primers modified with complementary locked primer technology (TMC-PCR). We checked the sensitivity of the developed assay by comparing it to a previously published TaqMan probe real-time one-step RT-PCR (TTN-PCR) procedure using enteroviral isolates, Enterovirus Proficiency panels from Quality Control on Molecular Diagnostics (QCMD-2007), and clinical specimens from patients with suspected EV infections.ResultsOne hundred clinical specimens from 158 suspected viral meningitis cases were determined to be positive by the TMC-PCR assay (63.29%), whereas only 60 were found to be positive by the TTN-PCR assay (37.97%). The positive and negative agreements between the TMC-PCR and TTN-PCR assays were 100% and 59.2%, respectively.ConclusionThis data suggest that the TMC-PCR assay may be suitable for routine diagnostic screening from patient suspected EV infection.
Highlights
Enterovirus (EV) infections are commonly associated with encephalitis and meningitis
The positive and negative agreements between the TMC-polymerase chain reaction (PCR) and time one-step RT-PCR (TTN-PCR) assays were 100% and 59.2%, respectively. This data suggest that the TMC-PCR assay may be suitable for routine diagnostic screening from patient suspected EV infection
Viruses and Controls Five reference strains belonging to distinct genogroups [enterovirus 71 (EV71), coxsackievirus B2 (CVB2), echo 30 (E30), coxsackievirus A24 (CVA24), and poliovirus type 1 (P1)] were obtained from the American Type Culture Collection (ATCC) and were used to optimize TTN-PCR conditions and to evaluate analytical performance
Summary
Enterovirus (EV) infections are commonly associated with encephalitis and meningitis. Detection of enteroviral RNA in clinical specimens has been demonstrated to improve the management of patients, by ruling out other causes of disease. EVs are associated with diverse clinical syndromes, ranging from mild febrile illness to severe central nervous system diseases, such as aseptic meningitis and encephalitis, potentially leading to paralysis [1,2]. Serotypes of EVs have traditionally been classified into echoviruses, coxsackieviruses, the identification of EVs in suspected patients, molecular methods such as reverse transcription PCR (RT-PCR), real-time RT-PCR, and nucleic acid sequence-based amplification offer more sensitive, specific, and rapid results [15,16,17]. Several groups have described real-time RT-PCR methods for the detection of EVs in cerebrospinal fluid (CSF) [18,19,20,21,22,23]
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