Enterovirus, Molecular Detection, PCR, Plasma
NY State Approved Indicates the status of NY State approval and if the test is orderable for NY State clients.
Aids in diagnosing enterovirus infections
Real-Time Polymerase Chain Reaction (PCR)/RNA Probe Hybridization
(PCR is utilized pursuant to a license agreement with Roche Molecular Systems, Inc.)
Reporting Name A shorter/abbreviated version of the Published Name for a test; an abbreviated test name
Enterovirus PCR, P
Enterovirus (Coxsackievirus and Echovirus)
Hand, Foot, and Mouth Disease
Enterovirus (Coxsackievirus and Echovirus)
Hand, Foot, and Mouth Disease
Specimen Type Describes the specimen type needed for testing
Specimen Required Defines the optimal specimen. This field describes the type of specimen required to perform the test and the preferred volume to complete testing. The volume allows automated processing, fastest throughput and, when indicated, repeat or reflex testing.
Container/Tube: Lavender top (EDTA)
Specimen Volume: 3 mL
Forms: If not ordering electronically, submit a Microbiology Request Form (Supply T244) with the specimen.
Specimen Minimum Volume Defines the amount of specimen required to perform an assay once, including instrument and container dead space. Submitting the minimum specimen volume makes it impossible to repeat the test or perform confirmatory or perform reflex testing. In some situations, a minimum specimen volume may result in a QNS (quantity not sufficient) result, requiring a second specimen to be collected.
Mild OK; Gross reject
Specimen Stability Information Provides a description of the temperatures required to transport a specimen to the laboratory. Alternate acceptable temperature(s) are also included.
|Plasma EDTA||Refrigerated (preferred)||7 days|
Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test
Enteroviruses are positive-sense RNA viruses in the Picornaviridae family. These viruses were initially classified by serotype as polioviruses (3 types), echoviruses (31 types, including types 22 and 23 which are now classified as parechoviruses), coxsackievirus A (23 types) and coxsackievirus B (6 types). However, genomic studies have demonstrated that there is significant overlap in the biological characteristics of different serotypes and more recently isolated enteroviruses are now named with consecutive numbers (eg, EV68, EV69).
The normal site of enterovirus replication is the gastrointestinal tract where the infection is typically subclinical. However, in a proportion of cases, the virus spreads to other organs, causing systemic manifestations, including mild respiratory disease (eg, common cold); conjunctivitis; hand, foot, and mouth disease; aseptic meningitis; myocarditis; and acute flaccid paralysis. Collectively, enteroviruses are the most common cause of upper respiratory tract disease in children. In addition, the enteroviruses are the most common cause of central nervous system (CNS) disease; they account for almost all viruses recovered in culture from spinal fluid. Differentiation of enteroviruses from other viruses and bacteria that cause CNS disease is important for the appropriate medical management of these patients.
Traditional cell culture methods require 6 days, on average, for enterovirus detection. In comparison, real-time PCR allows same-day detection. Detection of enterovirus nucleic acid by PCR is also the most sensitive diagnostic method for the diagnosis of CNS infection caused by these viruses.
Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.
A positive result indicates the presence of enterovirus RNA in the specimen.
Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances
A negative result does not rule out the possibility of enterovirus infection. This assay may detect virus from a variety of specimen types in asymptomatic individuals. This assay should only be used for patients with a clinical history and symptoms consistent with enterovirus infection, and must be interpreted in the context of the clinical picture. This test should not be used to screen asymptomatic patients.
Accuracy/Diagnostic Sensitivity and Specificity:
We compared the generic detection of enteroviruses from spinal fluid by conventional tube cell culture (MCR-5) and by LightCycler PCR. Of 715 specimens tested, enteroviruses were detected in 65 (9%) by conventional cell culture and 82 (11%) by LightCycler PCR. Twenty-two of 82 (27%) were exclusively positive by PCR; whereas, only 5 of 65 (8%) were exclusively positive by conventional cell cultures.
Supplemental Data (Spiking Studies):
To supplement the data above, 30 or more negative specimens of each specimen type (cerebrospinal fluid/sterile body fluid, dermal/ocular/rectal swabs, plasma and upper and lower respiratory specimens) were spiked with enterovirus culture control at approximately 10 to 50 targets/mcL (the approximate limit of detection). The spiked specimens were run in a blinded manner along with negative (non spiked) specimens of each specimen type. Of the spiked specimens, 97% to 100% were positive, and 100% of the non spiked specimens were negative. A total of 489 spiked and non spiked specimens were tested.
The assay detected all 64 members of an enterovirus panel, consisting of coxsackieviruses, polio viruses, echoviruses, and other enteroviruses.
Analytical Specificity/Limit of Detection (LoD):
The lower LoD of this assay is approximately 10 to 50 RNA target copies/mcL. This was confirmed in all specimen types accepted for this assay.
The assay did not crossreact with a specificity panel containing other RNA-containing viruses (rhinovirus; reovirus; influenza virus, types A and B; respiratory syncytial virus; and parainfluenza virus) and DNA-containing viruses (herpes simplex, Epstein-Barr virus, varicella-zoster virus, and cytomegalovirus.
This is a qualitative assay and results are reported a either negative or positive for targeted enterovirus RNA.
Clinical Reference Provides recommendations for further in-depth reading of a clinical nature
1. Enterovirus surveillance-United States, 1970-2005. MMWR Morbidity Mortality Weekly Report Sept 15 2006;55(SS08);1-20
2. Foray S, Pailloud F, Thouvenot D, et al: Evaluation of combining upper respiratory tract swab samples with cerebrospinal fluid examination for the diagnosis of enteroviral meningitis in children. J Med Virology 1999;57(2):193-197
3. Furione M, Zavattoni M, Gatti M, et al: Rapid detection of enteroviral RNA in cerebrospinal fluid (CSF) from patients with aseptic meningitis by reverse transcription-nested polymerase chain reaction. New Microbiol 1998;21(4):343-351
Method Description Describes how the test is performed and provides a method-specific reference
For this real-time reverse-transcription laboratory-developed PCR assay, viral nucleic acid is extracted by the MagNA Pure automated instrument (Roche Applied Science) from specimens, followed by amplification and detection on the Roche LightCycler 2.0 instrument. This PCR assay has been optimized to detect a target sequence in the polyprotein region. Primers amplify a 193-bp product.
Enterovirus genomic RNA is first transcribed to cDNA by reverse transcriptase, followed by amplification of the cDNA product. The LightCycler instrument can rapidly (30-40 minutes) detect amplicon development through stringent air-controlled temperature cycling in capillary cuvettes. The detection of amplified products is based on the fluorescence resonance energy transfer (FRET) principle. For FRET product detection, a hybridization probe with a donor fluorophore, fluorescein, on the 3'-end is excited by an external light source and emits light that is absorbed by a second hybridization probe with an acceptor fluorophore, LC-Red 640, at the 5'-end. The acceptor fluorophore then emits a light of a different wavelength that can be measured with a signal that is proportional to the amount of specific PCR product. FRET (with subsequent production of a detectable fluorescent signal) only occurs when the probes have specifically annealed to the target sequence of the amplicon.
Melting curve analysis is performed following PCR amplification and is the detection phase of the assay, since it offers greater sensitivity than the amplification phase and maintains high specificity.
The melting phase of the assay occurs as follows:
Starting at 45 degrees Celsius, which allows the probes to bind to the amplified product, the temperature in the thermal chamber is then slowly raised to 80 degrees Celsius and the fluorescence is measured at frequent intervals to determine the point where half of the fluorescence is lost as the probes are denatured (ie, "melt") off of the target. This is called the melting temperature (Tm) of that virus. Analysis of the PCR amplification and probe melting curves is accomplished through the use of LightCycler software. (Cockerill FR III, Uhl JR: Applications and challenges of real-time PCR for the clinical microbiology laboratory. In Rapid Cycle Real-Time PCR Methods and Applications. Edited by U Reischl, et al. Germany, Springer, 2002, pp 3-30)
Day(s) and Time(s) Test Performed Outlines the days and times the test is performed. This field reflects the day and time the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time required before the test is performed. Some tests are listed as continuously performed, which means assays are performed several times during the day.
Monday through Sunday; varies
Analytic Time Defines the amount of time it takes the laboratory to setup and perform the test. This is defined in number of days. The shortest interval of time expressed is "same day/1 day," which means the results may be available the same day that the sample is received in the testing laboratory. One day means results are available 1 day after the sample is received in the laboratory.
Same day/1 day
Maximum Laboratory Time Defines the maximum time from specimen receipt at Mayo Medical Laboratories until the release of the test result
Specimen Retention Time Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded
Performing Laboratory Location The location of the laboratory that performs the test
Test Classification Provides information regarding the medical device classification for laboratory test kits and reagents. Tests may be classified as cleared or approved by the US Food and Drug Administration (FDA) and used per manufacturer's instructions, or as products that do not undergo full FDA review and approval, and are then labeled as an Analyte Specific Reagent (ASR), Investigation Use Only (IUO) product, or a Research Use Only (RUO) product.
This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the U.S. Food and Drug Administration.
CPT Code Information Provides guidance in determining the appropriate Current Procedural Terminology (CPT) code(s) information for each test or profile. The listed CPT codes reflect Mayo Medical Laboratories interpretation of CPT coding requirements. It is the responsibility of each laboratory to determine correct CPT codes to use for billing.
LOINC® Code Information Provides guidance in determining the Logical Observation Identifiers Names and Codes (LOINC) values for the result codes returned for this test or profile.
|Result ID||Reporting Name||LOINC Code|
|56070||Report Status||In Process|