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Advancing molecular testing

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January 2000

Molecular testing historically has been implemented when traditional testing hasn’t worked well or in the absence of a traditional assay, noted David Hillyard, MD, director of molecular pathology and associate medical director for infectious disease testing, ARUP Laboratories, Salt Lake City. "There is no better example than HIV," Dr. Hillyard told CAP TODAY.

RT-PCR and bDNA tests for HIV viral load "compete for the majority of the U.S. market. Each has undergone fairly rapid generational improvement," Dr. Hillyard said. RT-PCR has the disadvantage of limited dynamic range but is highly specific and is the only FDA-approved assay. New versions of continuously monitored PCR tests with extended dynamic range eventually will be available.

Dr. Hillyard agreed that bDNA has some false positives, perhaps three to five percent, he said, but "In our experience, those false positives have been less than 200 copies." In clinical settings, this should not be a problem provided physicians understand these limitations at the very low end. Finally, bDNA has higher throughput.

Dr. Hillyard is using tests for cytomegalovirus largely to assist bone marrow transplant surgeons. "There is controversy about whether one should be using antigenemia or quantitative PCR," Dr. Hillyard said. Almost everyone gets infected with CMV, so a too-sensitive test would pick up CMV in so many people that it wouldn’t be useful. He uses antigenemia and a qualitative assay that has been adjusted so it is not too sensitive. "We will move toward antigenemia pending the releases of improved quantitative PCR assays," he stated.

"It is very important to do molecular testing on CSF for HSV and varicella-zoster in the clinical setting of encephalitis," he explained. "This has been a real success story for molecular testing," which can detect more than 95 percent of HSV infections, compared to about five percent for culture. Unfortunately, surveys document tremendous variability in home-brew assays for HSV.

"This is a general theme for all PCR tests," Dr. Hillyard said. "One of our greatest challenges is to bring more uniform quality to PCR tests so they are accurate and resistant to artifacts of contamination."

For HCV, qualitative molecular testing helps confirm the diagnosis and document successful endpoints in therapy. And because data link HCV viral load to prognosis and response rates, many clinicians order a quantitative test and follow viral load during therapy. Others get a pretherapy viral load and order a qualitative test at the end of therapy to see whether virus is undetectable. "There is nowhere near the consensus about how HCV tests should be used as there is for HIV," commented Dr. Hillyard.

William Check, PhD