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Michael Loeffelholz
Vice President, Scientific Affairs
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September 04, 2024
RESPIRATORY HEALTH
Expert Perspective
Emerging & Evolving Respiratory Viruses: Test Design Matters
Flu viruses and SARS-CoV-2 pose an ongoing public health challenge, largely because they evolve quickly and often. To help clinicians limit the spread of these viruses, diagnostic technology must account for genetic evolution. However, tests differ in their ability to detect new variants. Let’s review recent epidemiological data and explore diagnostic challenges and solutions.
Spread of Seasonal Respiratory Illnesses
World Health Organization (WHO) data show COVID-19 cases peaked when new variants appeared, especially the Omicron variant, causing surges in early 2022 and early 2023. From mid-2023 to 2024, weekly COVID-19 cases decreased, but the WHO still receives reports of hundreds of thousands of cases each week.1
In the United States, COVID-19 hospital admissions followed these global trends, according to data from the Centers for Disease Control and Prevention (CDC). Major peaks happened in early 2021 and early 2022. But the numbers declined in 2023 and early 2024, thanks to vaccinations and other health measures.2
During the 2023/2024 season in Europe, influenza was the most common respiratory virus, followed by respiratory syncytial virus (RSV) and SARS-CoV-2. Flu cases peaked in late 2023, with influenza A (H3) and influenza A (H1N1)pdm09 being the main types.3 This trend also happened globally, where influenza A subtypes were detected the most.4
In the United States, the 2023/2024 flu season has been moderate, and most positive flu tests detected influenza A (H3).5 Australia had similar trends, with high flu activity in early 2020, mid-2022, early 2023, and early 2024. Influenza A (H3) and influenza A (H1N1)pdm09 were most common.6
Virus Genetic Drift & Its Impact on Testing
Recent studies in the Journal of Clinical Microbiology and the Journal of Pathology, Microbiology, and Immunology have shown that new strains of influenza A with matrix gene mutations can lead to false-negative results in tests that rely on a single gene target. For example, a study by Landry and Owen (2022) found that an influenza A H1N1 strain escaped detection by tests targeting only the matrix gene.9 This shows the need for multiple gene targets in molecular tests.
Genetic Evolution of SARS-CoV-2
SARS-CoV-2 has evolved rapidly, especially with the Omicron variant and its subtypes, challenging diagnostic accuracy. Data from the CDC show significant genetic changes from the original strain.12 This drift can affect the sensitivity of diagnostic tests, requiring ongoing updates and test validation.
Analysis of Diagnostic Technology
In a study by Jorgensen et al.,10 some commercially available tests showed false negative results due to changes in the influenza A H3N2 matrix gene sequence. The study highlighted that new circulating viral strains can cause a drop in test performance when tests are not designed to be robust against mutations.10
Cepheid's diagnostic tests handle viral genetic drift well because they use primers and probes targeting multiple influenza genes, including matrix gene targets from both human and bird influenza A strains. This target redundancy enabled the Cepheid multiplex test for influenza A, influenza B, RSV, and SARS-CoV-2 to withstand the mutations reported by Jorgensen, yielding no false negatives in the comparative study.10
This multi-target test design strategy provides broad strain coverage and reduces the impact of genetic mutations. These attributes are essential for future pandemic preparedness. A multi-target test design can enable accurate detection of various viral subtypes. Recent data from in silico analyses using the Global Initiative on Sharing All Influenza Data (GSAID) sequence database show Cepheid tests maintained over 96% sequence similarity with all tested human and bird influenza A strains, effectively covering recent genetic drifts.11
Importance of Rapid Testing
A study published in the Journal of Clinical Virology in 2018 by Pedersen et al. showed the benefits of rapid testing for flu and RSV. Point-of-care testing (POCT) for these viruses can reduce the time it takes to get results from 5.2 hours with lab tests to just 20 minutes. This quick turnaround is crucial. Without POCT, 70% of virus-positive patients ended up sharing rooms with virus-negative patients, and 27% of virus-positive patients got unnecessary antibiotics.7 Rapid testing helps ensure patients receive the right treatment quickly, reducing the spread of infections.
Changing Performance of SARS-CoV-2 Rapid Antigen Tests
Studies in the Journal of Clinical Microbiology have looked at the sensitivity of rapid antigen tests against SARS-CoV-2 variants, including Omicron and Delta. These studies found that while some rapid antigen tests remain effective, others have reduced sensitivity.14
Value of Multiplexed Testing
RSV, influenza, and SARS-CoV-2 co-circulate and have many similar signs and symptoms, making multiplex tests essential. These tests can detect multiple viruses simultaneously, making fast, definitive diagnoses possible. These tests also aid clinicians in grouping hospitalized patients appropriately and prescribing the right antiviral treatments.
Cepheid’s SARS-CoV-2 Surveillance
Cepheid’s approach to SARS-CoV-2 testing targets multiple genes, including the RdRP, E, and N2 genes. This strategy ensures high inclusivity across viral variants, with in silico analyses predicting no false-negative results for new or circulating variants.13
Quality Test Design Where Patients Receive Care
Cepheid’s GeneXpert® platform offers on-demand, PCR testing in hospital labs and near-patient settings with performance equal to batch-based laboratory systems wherever testing is performed. This allows for timely and accurate diagnosis of respiratory viruses, including influenza A, influenza B, RSV, and SARS-CoV-2.15, 16, 17
Test design that focuses on multiple genetic targets and broad strain coverage reduces the impact of genetic drift and increases accuracy. As respiratory viruses continue to evolve, the quality of molecular test design will likely continue to emerge as a driver of clinical decision-making and patient impact.
IVD. In Vitro Diagnostic Medical Device. May not be available in all countries.
References:
1. WHO. SARS-CoV-2 Status—Global. Accessed May 2024. https://data.who.int/dashboards/covid19/cases
2. CDC. SARS-CoV-2 Status—U.S. Accessed May 2024. https://nextstrain.org/ncov/gisaid/global/6m
3. WHO. Respiratory Virus Status—Europe 2023/4 Season. Accessed May 2024. https://erviss.org
4. WHO. Influenza—Global. Accessed May 2024. https://app.powerbi.com/view
5. CDC. Influenza—U.S. 2023/4. Accessed May 2024. https://www.cdc.gov/flu/weekly/index.htm
6. WHO. Influenza—Australia. Accessed May 2024. https://app.powerbi.com/view
7. Pedersen, C, et al. Using a novel rapid viral test to improve triage of emergency department patients with acute respiratory illness during flu season. J Clin Virol. 2018 Sep 15(108):72-76.
8. Fenstermacher KZJ, et al. Pre- and Post-implementation Comparison of the Impact of Emergency Department (ED)-Based COVID-19 Point-of-Care Testing on ED Patient Metrics. Open Forum Infect Dis. 2023 Nov 27;10(Suppl 2):ofad500.521.
9. Landry ML, Owen M. Failure to Detect Influenza A H1N1 Highlights the Need for Multiple Gene Targets in Influenza Molecular Tests. J Clin Microbiol. 2023 Jul 20;61(7):e0044823.
10. Jørgensen, RL, et al. Emergence of circulating influenza A H3N2 viruses with genetic drift in the matrix gene: be alert of false-negative test results. APMIS. 2022 Oct;130(10):612-617.
11. Cepheid. Multi-target Design for Broad Coverage of Influenza Strains: Mitigating Impact of Recent Genetic Drift. Medical/Scientific Affairs Bulletin.
12. Centers for Disease Control and Prevention. Genetic Evolution of SARS-CoV-2. COVID Data Tracker. Accessed May 2024. https://covid.cdc.gov/covid-data-tracker/#variant-proportions.
13. Cepheid. Xpert Xpress SARS-CoV-2/Flu/RSV plus. Medical/Scientific Affairs Bulletin.
14. Rao, A, et al. Sensitivity of Rapid Antigen Tests Against SARS-CoV-2 Omicron and Delta Variants. medRxiv [Preprint]. 2023 Feb 10.
15. Moran, A, et al. Detection of SARS-CoV-2 by Use of the Cepheid Xpert Xpress SARS-CoV-2 and Roche cobas SARS-CoV-2 Assays. J Clin Microbiol. 2020 Jul 23;58(8):e00772-20.
16. Banerjee, D, et al. Comparison of Six Sample-to-Answer Influenza A/B and Respiratory Syncytial Virus Nucleic Acid Amplification Assays Using Respiratory Specimens from Children. J Clin Microbiol. 2018 Oct 25;56(11):e00930-18.
17. Liu, YL, et al. Diagnostic Accuracy of Xpert Xpress Flu/RSV for the Detection of Influenza and Respiratory Syncytial Viruses. Jpn J Infect Dis. 2022 Mar 24;75(2):183-191.
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