Mutations Can Affect Accuracy of Some COVID-19 False Negative Test Results

Pharmacists and other healthcare professionals providing COVID-19 testing should be aware that SARS-CoV-2 genetic variants can affect the accuracy of testing. The FDA recently sent out an alert pointing out that mutations could cause false negatives in patients who are tested and offered some advice on dealing with the issue. Here is more information.

WASHINGTON, DC – Mutations in the SARS-CoV-2 virus not only can make it more or less transmissible or likely to cause more or less serious cases, they can also make it more difficult to detect.

The U.S. Food and Drug Administration (FDA) sent out a letter to healthcare professionals and clinical laboratory staff, alerting them that false negative results can occur with any molecular test for the detection of COVID-19 if a mutation occurs in the part of the virus’ genome assessed by that test.

Thus far, the FDA has identified three molecular tests that have received emergency use authorization whose accuracy could be affected by genetic variations. Those tests include:

  • Accula SARS-Cov-2 Test: The FDA’s analysis, including additional information provided by the manufacturer, Mesa Biotech Inc., indicates that the Accula SARS-Cov-2 Test performance might be affected when a SARS-CoV-2 virus patient sample having a genetic variant at position 28881 (GGG to AAC) is tested. “While the impact does not appear to be significant, the FDA is providing this alert to health care providers out of an abundance of caution while we continue to gather additional data and work with the manufacturer,” according to the alert.
  • TaqPath COVID-19 Combo Kit: The FDA’s analysis, including additional information provided by the manufacturer, Thermo Fisher Scientific, Inc., and multiple reports from clinical laboratories, indicates that one of three targets of the TaqPath COVID-19 Combo Kit (which may also be labeled as the TaqPath COVID-19 Combo Kit Advanced) has significantly reduced sensitivity due to certain mutations, including one of the mutations in the recently identified B.1.1.7 variant (UK VOC-202012/01). The test is designed to detect multiple genetic targets, so the FDA suggests that overall test sensitivity should not be impacted and that the pattern of detection when certain mutations are present may help with early identification of new variants in patients to reduce further spread of infection.
  • Linea COVID-19 Assay Kit: The FDA’s analysis, including additional information provided by the manufacturer, Applied DNA Sciences, Inc., indicates that one of the two targets of the Linea COVID-19 Assay Kit has significantly reduced sensitivity due to certain mutations, including one of the mutations in the recently identified B.1.1.7 variant. Again, the FDA points out that, since the test is designed to detect multiple genetic targets, the overall test sensitivity should not be impacted.

“The presence of SARS-CoV-2 genetic variants in a patient sample can potentially change the performance of the SARS-CoV-2 test,” the regulators explain. “Molecular tests designed to detect multiple SARS-CoV-2 genetic targets are less susceptible to the effects of genetic variation than tests designed to detect a single genetic target. The FDA reminds clinical laboratory staff and health care providers about the risk of false negative results with all laboratory tests, including molecular tests. No test is perfect. Laboratories should expect some false results to occur even when very accurate SARS-CoV-2 tests are used. The prevalence of genetic variants may vary in any population and may lead to more false negative results than otherwise expected.”

Based on that information, the FDA is recommending that those who use molecular tests for the detection of SARS-CoV-2 to:

  • Understand that genetic variants of SARS-CoV-2 arise regularly, and false negative test results can occur.
  • Be aware that tests that use multiple genetic targets to determine a final result are less likely to be impacted by increased prevalence of genetic variants.
  • Consider negative results in combination with clinical observations, patient history, and epidemiological information.
  • Use repeat testing with a different test (with different genetic targets) as an option if COVID-19 is still suspected after receiving a negative test result.

“Most molecular tests for SARS-CoV-2 are designed to detect specific RNA sequences found in the viral genome,” according to the alert. “However, the SARS-CoV-2 virus mutates regularly, resulting in several genetically unique variants, each with different RNA sequences, that may be found in infected individuals as the pandemic progresses. The FDA has been monitoring the SARS-CoV-2 virus mutations, and potential impact on testing, throughout the pandemic. 

“Since molecular tests look for a specific sequence, it is possible for certain virus mutations to affect the performance of certain tests if the mutation is in a region of the genome targeted by the test. The FDA monitors these potential performance changes by comparing the sequence of genetic variants of the SARS-CoV-2 virus with the RNA sequences targeted by authorized SARS-CoV-2 tests. The clinical impact of genetic variants on test sensitivity is influenced by the sequence of the variant, the design of test and the prevalence of the variant in the patient population. Tests that rely on the detection of multiple regions of the genome may be less impacted by genetic variation in the SARS-CoV2 genome than tests that rely on detection of only a single region.”

The mutation in the news – the recently identified B.1.1.7 variant -- has been associated with an increased risk of transmission. The FDA points out the importance of early identification of the variant in patients to help reduce further spread of infection. The B.1.1.7 variant carries many mutations, including a double deletion at positions 69 and 70 on the spike protein gene (S-gene), which is the mutation that appears to impact the pattern of detection when using the TaqPath COVID-19 Combo Kit and the Linea COVID-19 Assay Kit, the letter explains.

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