On June 16, the U.S. Food and Drug Administration (FDA) posted updates to their Molecular Diagnostic Templates for developers that intend their assay to be used for pooled patient samples or for screening asymptomatic individuals not suspected of having COVID-19. FDA has previously authorized molecular diagnostic tests for individuals suspected of having COVID-19 by their health care provider, where patients may be symptomatic, pre-symptomatic, or asymptomatic. Importantly, it has always been at the discretion of health care providers whether or not to test asymptomatic individuals. There is currently no SARS-CoV-2 diagnostic test for the broad screening of asymptomatic individuals for COVID-19.

Recognizing that organizations may want to conduct screening of asymptomatic individuals as part of a broader strategy to help ensure the safety of their employees, patients, students and others, FDA has updated their Emergency Use Authorization (EUA) templates and available FAQs to assist developers who wish to develop tests for screening asymptomatic individuals. Given that testing asymptomatic individuals means an increased number of tests and an expected lower level of disease prevalence, particularly if the population is at low risk for exposure to or contracting COVID-19, developers may be interested in using pooling techniques to preserve testing resources.

The technique of “pooling” samples allows a lab to mix several samples together in a “batch” and then test the pooled sample with a single diagnostic test. Therefore, fewer tests are run overall, fewer testing supplies are used and results can be returned more quickly. This makes pooling samples for testing large populations of asymptomatic patients an attractive option for developers, but as samples are diluted there is a higher risk of false negative results.

Broad screening using a highly sensitive test, especially given the asymptomatic testing pool, leads to the most accurate results, which is why the FDA has provided validation recommendations designed to establish high sensitivity for tests intended for this purpose in the updated templates. FDA has noted that in general, point-of-care tests tend to be less sensitive than laboratory-based tests. Additionally, although current data suggests that asymptomatic and symptomatic individuals with COVID-19 may have similar levels of viral genetic material in their anterior nares and other upper respiratory sites, there is limited data on the viral loads at different time-points, in asymptomatic and pre-symptomatic individuals.

With these considerations in mind FDA has provided the following updates to the molecular diagnostic test templates:

• Screening asymptomatic individuals with a previously unauthorized test

  • FDA recommends a clinical study in the intended asymptomatic population where the test results are compared to comparator assay for each patient enrolled.

  • At least 20 positive samples should be prospectively collected in the intended use population and be sufficient to demonstrate the following minimum performance:

    • Positive Percent Agreement (PPA) ≥ 95% (Lower Bound of the two-sided 95% confidence interval >76%);

    • Negative Percent Agreement (NPA) ≥ 98% (Lower Bound of the two-sided 95% confidence interval >95%).

  • Samples should be collected at a minimum of 3 geographically diverse sites.

• Adding asymptomatic population to an authorized EUA test

  • If the assay is highly sensitive as determined by testing with the FDA SARS-CoV-2 Reference Panel or a recognized international standard, a post-authorization study may be appropriate.

  • FDA recommends testing a minimum of 20 consecutively collected asymptomatic positive specimens and at least 100 consecutively collected negative specimens based on the results of the candidate test. All specimens should then be tested with another EUA authorized molecular assay.

  • PPA should be ≥95% (Lower Bound of the two-sided 95% confidence interval >76%) and NPA should be ≥ 98% (Lower bound of the two-sided 95% confidence interval >95%).

• Sample pooling with a new test (not previously authorized)

  • The number of samples to be combined in one pool for testing should be established for each specimen type in the pooling claim.

  • FDA recommends a clinical validation study:

    • The number of enrolled patient specimens should be sufficient to ensure at least 30 comparator method positive samples are collected

    • Those with positive results by the candidate assay should each be pooled with n-1 (e.g., where n=4, n-1=3) comparator method negative samples and tested by the candidate assay.

    • All samples that were identified individually as positive by the test should still be positive when tested in pools

    • The clinical validation study should demonstrate that the positive samples with viral load close to the assay’s limit of detection (LoD) (i.e., weak positives) are accurately detected by the candidate test in a pool of negative samples.

  • As described above, samples should be collected from a minimum of 3 geographically diverse sites.

• Adding sample pooling to an authorized EUA test

  • FDA recommends a clinical study comparing the performance of the candidate assay when testing single specimens and specimen pools.

  • A minimum of 20 individual specimens which are positive by the candidate assay should be evaluated using similar methods described above.

Finally, FDA has stated that they generally do not regulate the use of a test for surveillance purposes, such as determining the prevalence of acute infections in a population. However, it is understood that results have been returned to individuals tested for COVID-19 surveillance purposes. If surveillance testing is performed by a non-CLIA certified laboratory, an individual who tests positive for SARS-CoV-2 should have a confirmatory test performed by a CLIA-certified laboratory. Surveillance with return of results and surveillance with pooled or batched testing should be validated on a test platform and test of high sensitivity and positive tests should have a confirmatory test.

References:

1. Coronavirus (COVID-19) Update: Facilitating Diagnostic Test Availability for Asymptomatic Testing and Sample Pooling, available at: https://www.fda.gov/news-events/press-announcements/coronavirus-covid-19-update-facilitating-diagnostic-test-availability-asymptomatic-testing-and
2. FAQs on Testing for SARS-CoV-2, https://www.fda.gov/medical-devices/emergency-situations-medical-devices/faqs-testing-sars-cov-2

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