Interpreting Laboratory Tests

Laboratory Testing

Laboratory testing is recommended to:

• Confirm suspected cases of varicella
• Confirm varicella as the cause of outbreaks
• Confirm varicella in severe cases (hospitalizations or deaths) or unusual cases
• Determine susceptibility to varicella
• Determine if suspected vaccine-related adverse events were caused by vaccine-strain VZV

 

Laboratory Confirmation of Suspected Varicella

The most sensitive method for confirming a diagnosis of varicella is the use of polymerase chain reaction (PCR) to detect VZV in skin lesions (vesicles, scabs, maculopapular lesions). Vesicular lesions or scabs, if present, are the best for sampling. Adequate collection of specimens from maculopapular lesions in vaccinated people can be challenging. However, one study (Evaluation of Laboratory Methods for Diagnosis of Varicella) comparing a variety of specimens from the same patients vaccinated with one dose suggests that maculopapular lesions collected with proper technique can be highly reliable specimen types for detecting VZV. Other sources such as nasopharyngeal secretions, saliva, urine, bronchial washings, and cerebrospinal fluid are less likely to provide an adequate sample and can often lead to false negative results.

Other viral isolation techniques for confirming varicella are direct fluorescent antibody assay (DFA) and viral culture. However, these techniques are generally not recommended because they are less sensitive than PCR and, in the case of viral culture, will take longer to generate results.

IgM serologic testing is considerably less sensitive than PCR testing of skin lesions. IgM serology can provide evidence for a recent active VZV infection, but cannot discriminate between a primary infection and reinfection or reactivation from latency since specific IgM antibodies are transiently produced on each exposure to VZV. IgM tests are also inherently prone to poor specificity.

Paired acute and convalescent sera showing a four-fold rise in IgG antibodies have excellent specificity for varicella, but are not as sensitive as PCR of skin lesions for diagnosing varicella. People with prior history of vaccination or disease history may have very high baseline titers, and may not achieve a four-fold increase in the convalescent sera. The usefulness of this method for diagnosing varicella is further limited as it requires two office visits. A single positive IgG ELISA result cannot be used to confirm a varicella case.

For more information, see Collecting Specimens for Varicella Zoster Virus (Chickenpox and Shingles) Testing.

 

Laboratory Criteria for Diagnosis

• Demonstration of VZV DNA by PCR tests from a clinical specimen, ideally scabs, vesicular fluid, or cells from the base of a lesion is the preferred method for varicella diagnosis. PCR is also useful for confirming breakthrough varicella. Other methods, such as DFA and culture, are available for diagnosis but are less sensitive and specific than PCR.
• Positive serologic test for varicella-zoster immunoglobulin M (IgM) antibody when a varicella-like rash is present.
• Four-fold or greater rise in serum varicella immunoglobulin G (IgG) antibody titer by any standard serologic assay between acute and convalescent sera.

For both unvaccinated and vaccinated people, PCR is the most reliable method for confirming infection.

See the Laboratory Testing section of the Varicella Surveillance Manual

 

Virus Strain Identification

PCR genotyping of VZV identified in specimens collected from skin lesions can be used to distinguish rashes due to wild-type VZV from rashes caused by vaccine type (Oka/Merck)-strain VZV. Samples of blood, cerebrospinal fluid, biopsy or autopsy specimens may also be tested for vaccine-strain/wild-type VZV discrimination to confirm etiology and to identify a vaccine-adverse event, though these specimens may be less likely to provide an adequate sample and can often lead to false negative results. The National VZV Laboratory at CDC and the American Public Health Laboratory Association Vaccine Preventable Diseases Reference Centers (VPD-RCs[2 pages]) have the capacity to distinguish wild-type VZV from Oka strain using both strain differential real-time PCR or PCR combined with restriction fragment length polymorphism (RFLP) analysis. VPD-RCs are located in the state laboratories of Wisconsin, California, New York, and Minnesota, and each VPD-RC receives specimens from a designated group of states.

Examples of possible varicella vaccine-adverse events for which PCR genotyping for VZV can be useful include varicella or a varicella-related complication in a vaccinated person 7 to 42 days after vaccination, herpes zoster in a vaccinated person, and suspected secondary vaccine-strain VZV transmission.

 

Evaluating VZV Susceptibility

IgG ELISA

A single serologic IgG test can be used to determine if a person has antibodies to VZV from past varicella disease or who may be candidates for varicella-zoster immune globulin (VZIG). (The product available in the U.S. is VariZIG). Commercial enzyme-linked immunosorbent assays (ELISAs) are recommended for the purpose of screening. Whole infected cell (wc) ELISA is the most commonly used test to determine if a person has antibodies to VZV from past varicella disease. Wc ELISA is done on blood samples. It can readily detect seroconversion to natural infection with VZV. Routine testing for varicella immunity following vaccination is not recommended because commercially available VZV IgG assays are not sensitive enough to detect all seroconversions after vaccination. The more sensitive purified glycoprotein ELISA (gpELISA) has been used in research settings to detect seroconversion after vaccination. However, testing with gpELISA is not available commercially.

IgG avidity

IgG avidity has been used in research settings to determine if a person who is IgG positive for VZV was infected with the virus in the past or more recently. The laboratory at CDC has developed an IgG avidity assay which can be used to determine if the most recent VZV rash was due to primary infection (varicella) or reactivation (herpes zoster). People infected in the past tend to have antibodies with high affinity for binding to the antigen compared with people with a more recent infection who have a low affinity. As a result, low avidity is an indicator for VZV primary infection. Vaccinated people are expected to undergo antibody affinity maturation following vaccination, leading to moderate to high IgG avidity VZV antibody. As such, measurements of VZV avidity in vaccinated people are unlikely to distinguish remote infection (or vaccination) from recent (breakthrough) infection with VZV. This test is not available commercially.

 

Establishing Laboratory Evidence of Immunity to Varicella

A positive IgG ELISA result indicates that a person has antibodies to VZV either from past varicella disease or vaccination. This test cannot distinguish whether the antibodies were from a past episode of varicella or vaccination. While many commercial VZV IgG ELISAs perform well enough to reliably detect seroconversion for infection by wild type virus, the performance specifications (specificity and sensitivity) of these methods vary widely. Some commercially available VZV IgG assays are not reliable, even for the detection of people with a history of natural disease. There are currently no commercially available VZV IgG methods sensitive and specific enough to reliably detect seroconversion to vaccine.

 

References

• Leung J, Harpaz R, Baughman AL, et al. Evaluation of laboratory methods for diagnosis of varicella. Clin Infect Dis. 2010 Jul 1;51(1):23-32.
• Schmid DS, Jumaan AO. Impact of Varicella Vaccine on Varicella-Zoster Virus Dynamics. Clin Microbiol Rev. 2010 Jan;23(1):202-17.