HHV-6 and Chronic Fatigue Syndrome (ME/CFS): The Difficulties of Diagnosis by Cort Johnson (8/08)

"There is good reason that it has taken so long to build a case for this virus playing a role in chronic fatigue syndrome – its very difficult to find…..Its quite possible to have a significant infection in brain tissues but no virus in the serum by DNA testing’.

Daram Ablashi – co-discover of HHV-6A/B

Several features complicate efforts to determine HHV-6A’s prevalence, abundance and effects.

    • Many of the cells HHV-6A replicates in are found in the central nervous system making it difficult to study them
    • Unlike most viruses it is largely spread through cell to cell transmission and thus is largely absent from the medium (blood, serum, plasma) usually used to detect pathogens
    • HHV-6’s latency in immune cells means serum or plasma (not whole blood) are the appropriate mediums to assess an active infection.
    • The currently available commercial tests cannot satisfactorily measure HHV-6A activity
At least six methods are used by to detect HHV-6. The HHV-6 Foundation asserts that the following commercial tests can be used, “albeit imperfectly’, to characterize active infections; PCR DNA tests on serum or plasma, IgM early antigen, IgG or IgM antibodies, primary cell culture and lymphocyte response.

HHV-6A Tests – An Overview

Antibody Tests – Antibodies or immunoglobulins are proteins produced by B-cells. They block a pathogens ability to infect other cells.

IgM Antibodies - IgM antibodies are the first antibodies produced by B cells in response to a primary infection. Because IgM antibodies usually peak and decline earlier than IgG antibodies they are generally believed to be better markers of active infection in children. Since IgM antibodies are typically not produced in response to HHV-6 reactivation they are not a suitable test for adults. (HHV-6 Foundation)

IgG Antibodies – IgG antibodies are the most abundant antibodies in the circulation. Because they can still be present after an infection has passed they are generally not considered good markers of active infection. The IgG antibody test is routinely positive for about 90% of adults (HHV-6 Foundation)

Wallace states that at least with regard to ME/CFS patients even high antibody titers are unreliable and culture studies are needed to verify them (Wallace et. al. 1999). Dr. Montoya argues, however, that very high levels of IgG antibodies (1:320, 1:640) in adults with the appropriate symptoms are sufficient to support a clinical diagnosis of an active HHV-6 infection. I

Low IgG antibody titers may not mean that an active infection is not present as people with weakened immune systems may not be able to mount a strong antibody response.

Polymerase Chain Reaction (PCR) - PCR tests for specific amino acid sequences found in viral DNA. Unfortunately the lack of a standard primer for HHV-6 makes comparing results across studies more difficult (Opsahl and Kennedy 2005).

The use of some PCR techniques to search for an active infection has been criticized on the following grounds (HHV-6 Foundation):
    • Because white blood cells typically contain latent HHV-6 viruses PCR must be done on serum.
    • HHV-6’s low replication rate and cell-cell transmission means few virions are typically present in the serum even in active infections; PCR tests must be extremely sensitive to pick up signs of active infection.
    • Since HHV-6 is ubiquitous in the population quantitative PCR tests are needed to establish between active and latent infections.
Quantitative PCR Tests – Sensitive quantitative PCR tests on serum or plasma (but not blood) can detect active HHV-6 infection.

Qualitative PCR Tests - Qualitative PCR tests on serum or plasma (but not blood) can detect active HHV-6 infection.

Nested PCR Tests – Nested PCR tests amplify traditional PCR results and can detect an active infection on serum or plasma (but not whole blood). Repeat testing is recommended to rule out the possibility of contamination and a false positive reading. Nested PCR can be used to differentiate HHV-6A from B using whole blood.

PCR Passe? – Dr. Montoya’s Valcyte trials with patients containing high antibody titers to both EBV and HHV-6 rebut the idea that PCR is effective in diagnosing treatable infections. While it was not a hard and fast rule Montoya found Valcyte responders tended to have high and the non-responders low antibody levels. Montoya reported that none of the Valcyte responders tested positive for HHV-6/EBV infection using PCR tests; PCR is not an acceptable test for determining which patients will respond to Valcyte. Montoya is continuing to attempt to define the Valcyte responsive subset in ME/CFS.

Rapid Culture – In culture tests a sample is cultured to see if HHV-6 is present. At the completion of the culture period samples are tested for the presence of specific proteins produced by actively replicating HHV-6. Using cultures to grow out the virus to determine if it is present is time-consuming, difficult and expensive. Only small labs provide culture tests. Culture tests can determine if the virus is present but cannot determine its degree of activity. Many people carry the viruses in their cells that can grow if stimulated properly.

A Missing Test - While many commercial labs in the U.S. are proficient at picking up acute HHV-6 infections, the HHV-6 Foundation asserts none are capable at identifying the kind of chronic, low-level infection usually found in ME/CFS patients. According to the Foundation "the most sensitive test for active infection was available briefly to researchers in the late 90’s and has never made it into commercial production; it was based on antibodies to the early antigen (EA) or one of the first proteins produced by the virus in the early stages of infection." (HHV-6 Foundation)

After entering the cell the virus uses the transcription and translation equipment of the cell to produce three kinds of viral proteins; IE, early (E) and late (L). Synthesized almost immediately upon infection, IE proteins regulate the expression needed to build more virus particles (virions). The E proteins are mainly involved in DNA metabolism and replication and the late proteins build such components of the mature virus particle as the virus wall, etc. Many antibody tests test for antigen produced on the outside of the viral particle; i.e. found on the viral wall. It is possible, however, for a pathogen to be active and produce detrimental proteins without replicating. It is these proteins that the early antigen tests measure.

Testing Facilities: The HHV-6 Foundation provides a list of testing facilities, and costs and recommendations for CFS and other patients with possible HHV-6 infection. The development of a sensitive test for HHV-6 infection is the Foundations top priority.

The HHV-6 Foundation Website

 Part I: CFS and HHV-6 - An Overview

Part III: HHV-6 and Disease