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Coronavirus Disease 2019

COVID-19 Testing


Testing New/Noteworthy

  • SARS-CoV-2 Viral Mutations: Impact on COVID-19 Tests—(3-30-31) New FDA webpage provides information on the impact of viral mutations on COVID-19 tests, recommendations for clinical laboratory staff and health care providers, and information about certain tests for which the FDA has identified potential impacts on performance due to SARS-CoV-2 genetic mutations.

Background

This webpage provides an overview of SARS-CoV-2 tests with a focus on the use and interpretation of diagnostic (viral) tests. The LAC DPH Testing Guidelines and Resources page hosts local testing and laboratory information as well as testing recommendations.

Currently Available SARS-COV-2 Test Types

  • Diagnostic (viral) tests -- molecular and antigen tests
    • Direct viral detection methods with sensitive molecular tests (e.g. RT-PCR) are recommended for diagnosing current infection with SARS-CoV-2.
    • Antigen tests can also be used to diagnose acute infection but they are less accurate than PCR tests.
  • See SARS-CoV-2 Diagnostic (Viral) Tests Summary Grid and Discussion
  • Serologic tests -- these antibody tests can be used to help support or establish a diagnosis in limited circumstances. See Serology Tests.

Emergency Use Authorizations (EUAs)

Only tests with emergency use authorizations (EUAs) from the FDA should be used for patient care. A wide variety of molecular tests, antigen tests, and serologic tests now have EUAs.

It is important for clinicians to review the EUA for any SARS-CoV-2 test(s) they are using to understand the specific performance characteristics and the instructions for use. All SARS-CoV-2 EUAs are listed on the FDA COVID-19 Emergency Authorization website.

Accuracy of COVID-19 Tests

No tests give a 100% accurate result; tests need to be evaluated to determine their sensitivity and specificity, ideally by comparison with a gold standard.

For COVID-19, there is no clear-cut gold-standard which makes the evaluation of test accuracy challenging. No clinical performance data is required for an EUA and there has been limited information on how these different COVID-19 tests perform in real world settings and with different populations.

Test accuracy is dependent on many factors including the test technology, specimen collection and handling, pretest probability of disease, the timing of test, and the specific characteristics of the assay. Clinicians should review the EUA and supporting materials for any SARS-CoV-2 test(s) they are using to understand the specific performance characteristics, the sample collection, storage and transport instructions, and what negative and positive test results mean.

Point of Care Tests

Both molecular and antigen tests have been approved by the FDA for waived testing at the point-of-care setting.

For facilities performing only waived SARS-CoV-2 antigen tests, a temporary allowance has been made during the California emergency. Facilities may begin waived SARS-CoV-2 antigen testing as soon as they have submitted an application for a California Clinical Laboratory Registration and received acknowledgment of receipt. See CDPH Temporary Allowance for Waived SARS-CoV-2 Testing for more information.

All positive test results from POC devices must be reported to Public Health.

In addition to closely following the manufacturer's instructions for use,, it is critically important that testing sites observe best practices for handling and performing tests on infectious disease specimens and fully train all staff engaged in sampling and test operations. The CDPH LFS Branch website provides guidance on the operation of COVID-19 testing and other resources including Information for Physician Offices and Urgent Care Clinics. For additional information on POC testing see CDC Guidance for SARS-CoV-2 POC Testing.

SARS-CoV-2 Diagnostic (Viral) Tests

Summary Grid (Table 1)
DIAGNOSTIC (VIRAL) TESTS

Shows current infection

Method

Molecular Tests

Amplifies specific fragment of viral RNA using nucleic acid amplification

  • Reverse-transcription polymerase chain reaction (RT-PCR)-current laboratory standard
  • Isothermal amplification methods-(LAMP, NEAR, etc)-ultra fast NAA methods

Antigen Tests

Lateral flow immunoassays to detect viral surface proteins

Type of sample*
*Refer to the specific assay IFU for acceptable sample type(s)

Nasopharyngeal (NP), oropharyngeal, or nasal swab; saliva; lower respiratory tract specimens NP or nasal swab.

Authorized for use at point of care (POC)

Most are not, but some are. Most are, at least one is not.

Test turn-around time

Standard RT-PCR: < 48 hours

POC tests range from 15-45 min
Range from 15-30 min

Interpretation of positive tests

RNA from SARS-CoV-2 was detected. The patient is infected with the virus and considered to be contagious.

Note: A positive test in an asymptomatic person within 3 months of a prior lab-confirmed COVID-19 infection likely represents detection of non-viable virus.

Antigens from SARS-CoV-2 were detected. The patient is infected with SARS-CoV-2 virus and considered to be contagious.

Note: While false positive results may occur when the test instructions for use are not followed or when screening asymptomatic low risk persons, patients with positive antigen tests must be treated as a case and must isolate. See Antigen Tests: Confirmatory Testing Recommendations and FAQs for more information.

Interpretation of negative tests

Negative tests should always be interpreted in the context of the exposure history and clinical presentation of person being tested.

A negative result in a patient with a high pre-test probability of infection (e.g. there is strong clinical suspicion for COVID-19 and/or recent exposure) has an increased likelihood of being a false negative.

A negative result in a patient with low pre-test probability of infection is more likely to be a true negative.
Negative tests should always be interpreted in the context of the exposure history and clinical presentation of person being tested.

In most cases, negative antigen tests are considered presumptive and should be confirmed with a RT-PCR test. Persons with symptoms consistent with COVID-19 should be instructed to isolate themselves pending confirmation with a RT-PCR test.

A negative result in a patient with a low pre-test probability of infection is more likely to be a true negative. Confirmatory testing is not needed unless important for clinical management or infection control. 

Considerations for Use

RT-PCR is recommended for the following:

  • Persons with symptoms of COVID-19
  • Persons who were close contacts to COVID-19*
  • For people without symptoms living/working in a high-risk setting (such as a skilled nursing facility) or who are identified as part of an outbreak detection and response

*A close contact cannot be released early from quarantine on the basis of negative molecular test.

Due to their rapid turn-around time antigen tests are useful to identify infected persons in the following situations:

  • Persons with symptoms of COVID-19*
  • Persons who were close contacts** to COVID-19
  • Persons identified as part of an outbreak investigation/response.

Antigen tests can have a role in screening when used to rapidly detect and monitor introduction of the virus into high risk congregate settings such as skilled nursing facilities.

Antigen tests are not recommended for screening in the general population in the absence of a recent known exposure.

*Refer to assay specific IFU for symptomatic window of detection

**A close contact cannot be released early from quarantine on the basis of negative antigen test.

Discussion of SARS-COV-2 Diagnostic (Viral) Tests

Molecular

Molecular tests are the most accurate tests for diagnosing current COVID-19. Molecular tests detect the unique genetic sequence of the SARS-CoV-2 virus by using nucleic acid amplification (NAA) technology such as reverse transcription polymerase chain reaction (RT-PCR) or other amplification methods (e.g. LAMP). RT-PCR is highly sensitive and specific for detecting SARS-CoV-2 virus and is the recommended “gold standard” method for diagnosing current infection. Most molecular tests are moderate or high complexity and must be run in a laboratory, however some are also authorized to be conducted in the point-of-care (POC) setting under a Clinical Laboratory Improvement Amendments (CLIA) Certificate of Waiver.

Not all molecular tests have equivalent performance, for instance rapid isothermal NAA tests have significantly lower sensitivity than standard laboratory based NAA tests and rapid RT-PCR tests.. The FDA is now publishing comparative performance data of molecular tests against a standard reference panel. The sensitivity of molecular tests is also dependent on the timing of testing, sampling technique, and sample type. Molecular tests are considered to be very specific.

  • A positive molecular test result generally confirms a SARS-CoV-2 infection. Note: RT-PCR tests may remain persistently positive for prolonged periods (up to 12 weeks) after a patient has recovered due to prolonged detection of non-viable RNA (see CDC Decision Memo).
  • Negative molecular tests must be interpreted in the context of the exposure history and clinical presentation. False-negative tests have been well documented, especially early in the course of infection (see Variation in False-Negative Rate of RT-PCR–Based SARS-CoV-2 Tests by Time Since Exposure).
    Note: a negative test result from lower sensitivity molecular tests in symptomatic persons should be considered presumptive and confirmation with a more sensitive RT-PCR is recommended

Laboratory turn-around time (TAT) for PCR test results should be less than 48 hours, however results can be delayed if laboratories are experiencing high volumes. Currently available POC molecular tests results have a TAT ranging from 15-45 minutes.

See FDA list of EUAs for Molecular Tests for more information on the performance and use of specific authorized tests.

Antigen

Antigen tests detect the presence of SARS-CoV-2 viral surface proteins. The main advantages of antigen tests are that results are available within 15-30 minutes, they are relatively simple to perform, and they are less expensive than RT-PCR tests. Most currently available EUA authorized SARS-CoV-2 antigen tests are all approved as POC tests for settings with a CLIA waiver. The intended for use of antigen tests is to evaluate symptomatic persons within 5-12 days of symptom onset (the number of days varies by manufacturer). See FDA’s list of EUAs for Antigen Tests.

In practice, antigen tests are being used more widely for diagnostic testing and for screening purposes outside their intended use. It is important that providers are aware of the limitations of these tests and that they use and interpret antigen test results based on the probability the patient has COVID-19 (pre-test probability). A summary of when confirmatory testing of antigen results is recommended is included below.

Limitations of Antigen Tests

False negative results are a limitation of these tests. Recently published reports as well as local experience has found that antigen tests in symptomatic persons are less sensitive than initially reported to the FDA. In addition, these tests have a much lower sensitivity when used to screen asymptomatic persons (~35-40%).

False positives are another limitation of antigen tests. While the specificity of antigen tests is generally high false positive antigen tests are known to occur when the manufacturer’s instructions for use are not followed correctly or if there are inadequate quality assurance procedures. False positives are also more likely to occur when testing persons with a low pre-test probability of infection (e.g., screening asymptomatic persons without risk factors in a low-prevalence setting).The FDA has issued an alert to healthcare providers regarding the potential for false-positive antigen results and steps to mitigate this risk.

Visit FAQs below for more on false positive antigen results.

Antigen test considerations for use

When used correctly, rapid antigen tests can help quickly identify patients early in the course of SARS-CoV-2 infection when viral load is highest and who pose the greatest risk of SARS-CoV-2 transmission to others. They perform best when there is a high pre-test probability of infection (e.g., symptoms consistent with COVID-19, recent exposure to a known case, and living/working in a setting where a high proportion of persons are infected).

Diagnostic testing

Antigen tests are useful as part of the evaluation of individuals with symptoms consistent with COVID-19 and/or those with a recent exposure to SARS-CoV-2. Positive antigen results are generally considered diagnostic of infection whereas negative results are presumptive.

Facilities using antigen tests for diagnostic testing should have the ability to collect same day specimens for confirmatory RT-PCR testing of negative antigen test results.

Screening

Antigen tests are not recommended for screening in the general population in the absence of a recent known exposure. 

When used for screening purposes (testing asymptomatic persons with no recent exposure), confirmation with RT-PCR is recommended for persons who have a positive antigen test result, see Confirmatory testing below.

Antigen tests can have a role in screening when used to rapidly detect and monitor introduction of the virus into high risk congregate settings such as skilled nursing facilities. Timely results can help inform decisions about patient management and infection control.

Providers or facilities using antigen tests for screening should refer to their setting-specific protocols and be aware of the performance characteristics of these tests. Results should be interpreted in the context of symptomatology, risk of exposure to SARS-CoV-2, and prevalence of COVID-19 in the setting.

Confirmatory testing

To account for reduced antigen test accuracy, confirmatory testing with a more sensitive test (e.g., RT-PCR) is recommended after negative antigen test results in symptomatic persons and after positive antigen test results in asymptomatic persons without exposure, see below. Clinical judgment should be used when deciding whether confirmatory RT-PCR testing should be performed. See Testing FAQs; What if I am concerned about an antigen result being a false positive and What if I am concerned about a result being a false negative for more detailed information.

Note: Skilled nursing facilities and other congregate settings using antigen tests should follow their setting specific guidelines.

If indicated, confirmatory testing should be done using RT-PCR on specimen collected as soon as possible after the antigen test and not longer than 48 hours after the initial specimen collection. Tests performed >2 days apart should be considered separate results and discordant results may be due to changes in viral dynamics.

Antigen Tests: Confirmatory Testing Recommendations

Person with symptoms consistent with COVID-19

  • If Positive – Confirmatory testing is not recommended. The patient must isolate and their close contacts must quarantine.

Asymptomatic person with known exposure in the past 14days (e.g., a close contact to a known case or living/working in an outbreak setting)

  • If Positive – Confirmatory testing is generally not recommended*. The patient must isolate and their close contacts must quarantine.
    *Consider confirmatory testing if there is lower pre-test probability of infection (e.g., non-household exposure) and/or to minimize unnecessary quarantine of their close contacts.
  • If Negative – Presumptive negative. Confirmatory testing may be needed if clinical judgment deems it important for patient management or infection control. The patient should continue to quarantine as instructed. If confirmatory testing is performed, a negative RT-PCR test does not release them from the required quarantine.

Asymptomatic person with no recent known exposure (e.g. screening)

Antigen tests perform poorly in asymptomatic persons with low risk of infection and are not recommended for screening the general population. If antigen tests were used for screening:

  • If Positive – Confirmatory testing is recommended. The patient must be treated as a case and instructed to isolate while awaiting the RT-PCR test result.
    Note: if confirmatory testing is not performed, then the result is considered diagnostic and the patient must isolate their isolation and their close contacts must quarantine.
  • If Negative – Confirmatory testing is not needed. It is important that the patient understand that the test could be falsely negative. The patient should be advised to continue all physical distancing and mask requirements. See When testing asymptomatic persons without a known exposure.

Related References

Interpreting Viral Test Results-Key Points
  • Interpreting the result of a diagnostic test for COVID-19 depends on two things: the accuracy of the test and the pre-test probability
  • Factors that increase the pre-test probability of infection include:
    • Symptoms consistent with COVID-19
    • Recent exposure to a person with COVID-19
    • Living or working in a setting where a high proportion of persons are infected (e.g. nursing home or workplace with an outbreak)
    • Prevalence of infection among the group being tested
  • A positive molecular test generally rules in infection with COVID-19 but is less good for ruling out COVID-19 if negative. This is because molecular tests have a high specificity but lower sensitivity.
  • A positive antigen test in a person with high pretest probability of infection rules in infection. To account for reduced antigen test accuracy, confirmatory testing with a more sensitive test (e.g., RT-PCR) is recommended after negative antigen test results in symptomatic persons and positive antigen test results in asymptomatic persons.
  • Negative viral tests for COVID-19 should always be interpreted in the context of the exposure history and clinical presentation.
    • A single negative viral test should not be used to rule-out infection when there is a high probability of infection nor can it be used to release a confirmed case from isolation.
    • A close contact cannot be released early from quarantine on the basis of a negative viral test.

Serology

Serology Tests detect waning or past SARS-CoV-2 virus infection indirectly, by measuring the antibody response to the virus. Serology assays should not replace direct viral detection methods for diagnosing an active SARS-CoV-2 infection. At this point in time, until the presence, durability, and duration of immunity is established, serology tests they should not be used to determine immune status.

See FDA list of EUA authorized serology tests and performance information.

Use of Serologic Tests

CDC Recommendations for Use of Serologic Tests include:

Diagnostic uses:

  • To help establish a diagnosis when a patient presents with complications of COVID-19 illness, such as multisystem inflammatory syndrome in children (MIS-C), in adults (MIS-A), and other post-acute sequelae of COVID-19.
  • To support diagnosis of acute COVID-19 illness for persons who present late. For persons who present 9-14 days after illness onset, serologic testing can be offered in addition to recommended direct viral detection methods such as PCR. This will improve diagnostic accuracy at a time when the sensitivity of PCR detection is decreasing, and serologic testing is increasing.

Non-diagnostic uses of serological tests:

When SARS-CoV-2 serology tests should not be used:

  • As sole basis to diagnose or rule out infection with SARS-CoV-2.
  • To screen for asymptomatic shedders.
  • To make decisions about grouping persons residing in or being admitted to congregate settings, such as schools, dormitories, or correctional facilities.
  • To release a person from isolation or quarantine or to clear a person to return to work.
  • To determine if a patient is immune or protected from re-infection.
  • To guide PPE use or infection control measures.

To learn more, see current serology guidance:

Considerations for Testing Asymptomatic Persons

Routine viral testing of asymptomatic persons in the general population in the absence of an exposure has limited value. Due to uncertainties in test performance in asymptomatic individuals, particularly with those at lower likelihood of infection, the meaning of a positive or negative test result becomes less clear. Antigen tests perform particularly poorly in asymptomatic persons and should be generally not be used unless in conjunction with RT-PCR tests.

There are situations where testing asymptomatic people is strongly recommended, such as recently exposed close contacts of a known case, persons who are part of contact investigations and/or outbreaks, and routine testing of staff and residents of nursing homes. In the absence of clear clusters of cases or known exposures, the utility of routinely testing asymptomatic persons in other situations and settings such as schools, hospitals, and workplaces is not clear. See Smart Testing for COVID-19 Virus and Antibodies, Center for Infectious Disease Research and Policy for more discussion.

Serial antigen testing of asymptomatic persons within a closed congregate setting, such as a long-term care facility or a correctional facility, has been suggested by the CDC as a potential strategy to rapidly identify cases of SARS-CoV-2 infection to prevent further transmission in the facility. CDC cites modeling evidence shows that outbreak control depends largely on the frequency of testing and the speed of reporting and is only marginally improved by high test sensitivity.

When testing asymptomatic persons without a known exposure

It is important to emphasize that negative test results only indicate that the test did not detect the virus at the time the test was taken. With widespread community transmission, any person who interacts with other people runs a daily risk of acquiring COVID-19. This daily risk increases in crowded places, in confined spaces (especially indoors), with close contact, and when protective actions, such as maintaining physical distancing, correctly using face coverings when around others, frequent hand hygiene, and other job-specific protective measures, are not followed.

A false negative result can happen when an infected individual is still incubating the infection (i.e., that the test was taken too early in the course of the infection) and/or that the test simply failed to detect the SARS-CoV-2 virus.

A negative test does NOT mean the person can safely ignore physical distancing and mask requirements. Refer the patient to Information about Testing.


Definitions

  • Pretest probability: Probability of a patient having an infection before the test result is known; based on the proportion of people in a community with the disease at a given time (prevalence) and the clinical presentation of the patient.
  • Negative predictive value: Probability that a patient who has a negative test result truly does not have the infection.
  • Positive predictive Value: Probability that a patient who has a positive test result truly does have the infection.
Frequently Asked Questions

Negative SARS-CoV-2 viral test results should always be interpreted in the context of the patient’s exposure history and the clinical presentation of person being tested.

A negative viral test result in a patient with a high pre-test probability of infection has an increased likelihood of being a false negative. If there is strong clinical suspicion for COVID-19 (compatible symptoms and/or exposure), and a patient has a negative viral test result, the patient should continue isolation and be managed as a presumed positive.

When using lower sensitivity assays (e.g. antigen tests and some molecular tests), negative results are considered presumptive and confirmation with RT-PCR is recommended, especially if important for clinical management or infection control. If the initial negative test was a RT-PCR, consider repeating the test to confirm infection.

Reminder:

  • A close contact cannot be released early from quarantine on the basis of negative viral test(s).

Any patient with a positive antigen test for COVID-19 should be treated as a case and isolated. 

False positives antigen tests are known to occur when instructions for use are not followed correctly or if there are inadequate quality assurance procedures. False positives are also more likely to occur when testing asymptomatic persons with low probability of infection (e.g. screening asymptomatic persons in a low-prevalence setting).

If a false positive antigen test result is suspected, then a confirmatory RT-PCR test is recommended. Note: confirmatory RT-PCR testing is not recommended in situations where there a high pre-test probability of infection (e.g. COVID-19 is clinically suspected, they are a household close contact, they live or work in a setting with an outbreak).

If confirmatory testing is pursued, the person must remain isolated until the result of the RT-PCR test is available.

  • A single negative RT-PCR test from a specimen collected within 2 days of the initial collection can be used to rule out infection. Tests performed >2 days apart should be considered separate results and discordant results may be due to changes in viral dynamics.
  • To optimize sensitivity, CDC recommends collecting a high-quality sample (NP swab) using a sensitive RT-PCR test. NP swabs used correctly have a higher sensitivity than other specimen sources.

Any new* positive PCR test for COVID-19 should be considered true, treated as a case, and isolated per guidelines.

*A repeat positive PCR test within 3 months of recovery from a confirmed SARS-CoV-2 infection in the absence of symptoms likely represents persistent shedding of non-viable viral RNA. See Patients with a History of Recent Recovery from COVID-19.

RT-PCR tests are highly specific and are the current gold-standard method for diagnosing current infection with SARS-CoV-2. Despite their high specificity, false positives are possible in the setting of low pre-test probability (e.g., screening asymptomatic persons with no known exposure in low prevalence settings).

If a patient is asymptomatic with no known exposures to COVID and there are concerns that a test is falsely positive, confirmatory laboratory based RT-PCR tests can be performed. In these situations, the person must remain isolated until all of the results of the RT-PCR are available.

  • Two negative laboratory performed RT-PCR tests collected > 24 hours apart can be used to rule out infection (unless the person becomes symptomatic)

When false-positive tests are suspected, CDC recommends that facilities should review and gather the following information:

  • Review the FDA EUA and instructions for use of the point-of-care test to ensure correct use. For instance, the use of viral transport medium has been shown to produce false-positive results and should not be used for some antigen platforms (including BD Veritor and Quidel Sofia platforms).
  • Review recent control results to ensure accuracy of the POC test machine. If able, perform procedural quality control tests to ensure correct operation of the machine.
  • Compare the percent positivity of the samples that were run that day (or week) to their previous percent positivity (e.g., the rolling 7-day average percent positive). Identifying many positives in one day might indicate an issue with the test results (either due to operator error or test supplies).
  • Consider the pre-test probability of disease. Testing in low-prevalence populations with antigen or molecular tests might produce false positives.

A low pretest probability has a high negative predictive value but a lower positive predictive value. This means there is an increased likelihood of true negatives and false positives.

A high pretest probability has high positive predictive value and a low negative predictive value. This means there is an increased likelihood of true positives and false negatives.

For instance, if testing a population with a COVID-19 prevalence of <1% (e.g., screening asymptomatic persons in non-outbreak settings) with a single test with 99% specificity, the positive predictive value (probability that a positive test is a true-positive) could be <40%. If the prevalence is >10% (e.g., testing asymptomatic persons as part of an outbreak response) with that same test with 99% specificity, the positive predictive value may be >90%.

See CDC interpreting test results.



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  • Public Health has made reasonable efforts to provide accurate translation. However, no computerized translation is perfect and is not intended to replace traditional translation methods. If questions arise concerning the accuracy of the information, please refer to the English edition of the website, which is the official version.

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