Respiratory Testing: Preparing for an Uncertain Future

The global emergence of COVID-19 has resulted in a radical shift in the management of patients presenting with respiratory and/or flu-like symptoms of fever, malaise, and myalgias. In the past, such symptoms could have been interpreted as consistent with infection with influenza or respiratory syncytial viruses, but under current conditions, SARS-CoV-2 is often considered in the differential diagnosis. Currently, a patient with these symptoms is likely to simply take a rapid antigen test at home, or present to their local clinic or hospital for testing. However, a negative result for COVID-19 could be giving the patient a false sense of security, with their symptoms actually being caused by something serious such as influenza.

SARS-CoV-2 has shifted the way the healthcare sector approaches respiratory patients, as well as how the general population reacts to this set of symptoms. This article delves into this changing clinical landscape and explores how the lessons learned from previous viral outbreaks have been applied to create fast and accurate molecular tests that are helping to not only diagnose COVID-19 now, but also to provide “variant-ready” solutions for the detection of SARS-CoV-2, influenza A and B, and other viruses that may cause similar symptoms.

The Global Burden of Respiratory Diseases

Respiratory viruses have had a long history of impact on human health, but rarely have they disrupted our way of life like SARS-CoV-2. Humans are not strangers to the coronavirus family, and most strains usually just cause mild upper-respiratory issues, like the common cold. However, occasionally, new strains emerge that are more deadly — such as MERS-CoV or SARS-CoV — or more transmissible, as is the case with SARS-CoV-2.¹ Fortunately, the general nature of these viruses is to adapt to become less pathogenic. New variants ranging in transmissibility and severity will continue to emerge but, over time, it is likely that the predominant strains will become more like the other endemic coronaviruses in circulation, resulting in milder symptoms. As this natural progression occurs, the current programme of vaccinations and boosters, as well as natural immunity resulting from infections, should provide ever-increasing levels of protection from severe disease. 

Influenza has followed a similar evolution, but still causes significant mortality and morbidity globally. Seasonal strains affect millions every year, and are responsible for hundreds of thousands of deaths, with one study reporting 290.000 to 650.000 seasonal influenza-associated respiratory deaths each year between 1999 and 2015.² Like all viruses, influenza continuously evolves in order to survive and better infect its hosts. Occasionally, strains appear that create a major public health emergency, such as influenza A subtype H1N1, which was responsible for the 1918 and 2009 pandemics, the former resulting in an estimated 50 million deaths worldwide.³

The rapid evolution of these viruses means that diagnostic tools must also continually adapt and develop to help guide clinicians in the management of their patients. The consequences of a misdiagnosis can be serious, including ineffective therapies potentially leading to more severe symptoms and disease progression as the causative pathogen escapes treatment.

The need for rapid, sensitive, and definitive results is now even more pressing with the advent of antiviral therapies, which are more effective if administered early in the course of infection.

Designing Robust Diagnostic Tests

Molecular methods remain the gold standard for diagnosing respiratory infections in a clinical setting. For COVID-19,⁴ rapid antigen tests are adequate in symptomatic patients who have high viral loads but lose accuracy when a patient is further from peak replication.⁵ Many of these patients show up to a hospital with inflammatory complications a week or so after symptoms first appear, usually with lower viral loads, and this may return a negative result with a rapid antigen test.

Designing and building molecular tests is an intricate, multistep process that usually begins with an unmet clinical need. Diagnostic companies assess where they can make the most significant impact, looking at what test is needed to help physicians, microbiologists, or infection control specialists quickly and accurately diagnose patients. However, it’s not just the opinion of a company that determines the design of a test. Advice from various groups — including physicians, nurses, pharmacists, virologists, public health specialists, and regulatory personnel — help to ensure the resulting test addresses the relevant clinical need. It usually takes years to develop a product and launch it into the market, beginning with the initial assay design and optimisation, before undergoing validation to prove its accuracy. The data is then submitted to a regulatory body — such as the FDA — and, if approved, it can be manufactured under strict quality control conditions and made available to laboratories and other health facilities to start responding to clinical need.

This intensive process makes it crucial to design a product that can withstand the test of time which, in the case of many viruses, means one that takes into consideration the continuous change and evolution of the target pathogen. To stay one step ahead of infectious diseases, diagnostic tests need to meet the clinical needs of today and, crucially, the evolutionary pressures of tomorrow. If we don’t have sufficient data about a pathogen, or the knowledge of a disease, then this may not be possible. Fortunately, in most cases, the lessons learned from previous outbreaks or other organisms can be used to help develop the tools of the future.

Preparing Molecular Tests for the Future

RNA viruses are prone to random genetic drift, which forms part of their adaptive ability to survive. This has been witnessed with the emergence of multiple variants of SARS-CoV-2, each arising from selective pressures in certain populations, or from variations arising in regions of the virus that are not subject to immune selection. These broad evolutionary changes give the virus a survival advantage, and can make new variants harder to detect if tests are not designed to accommodate this variation.

This issue was encountered during the 2009 influenza pandemic when tests were developed for a single antigenic target that subsequently drifted. Many diagnostic companies struggled to keep up with this outbreak, and the sensitivity of rapid antigen tests plummeted due to variations within the protein target, resulting in countless undiagnosed patients.⁷ Indeed, in 2009 and 2010, the poor performance of diagnostic tests for detection of influenza prompted the Centers for Disease Control and Prevention in the US to recommend avoiding use of these tests.⁸ There will always be a small percentage of variation within a single target, and therefore a number of cases will manage to avoid detection. Unfortunately, it’s difficult to predict when and where genetic variation will occur, and how fast changes will be, and so diagnostic methods need to account for this.

Learning from the H1N1 outbreak, Cepheid developed a test to cover multiple independent and conserved influenza A targets, reducing the chance that future influenza strains will escape detection. This was developed into an assay that detects not just seasonal influenza but is also predicted to detect future influenza strains of pandemic concern such as H5N1, H7N9 and avian influenza, and also respiratory syncytial virus (RSV), providing a more thorough screening solution to improve patient management.

This technological know-how and experience proved crucial when SARS-CoV-2 emerged, and accelerated the development of the Xpert® Xpress CoV-2/Flu/RSVplus test, which provides a combined screening option for SARS-CoV-2, RSV, influenza A and influenza B in a single self-contained cartridge for the GeneXpert® system. This solution helps to rapidly diagnose patients presenting with non-specific upper respiratory symptoms, or where physicians suspect one of these pathogens is present, providing a clearer course of action for rapid patient management.

To minimise the number of variants that could potentially evade detection, this test comprises multiple targets for each virus.

This method provides accuracy that is comparable to reference labs, yet can be performed close to the patient, away from a central location. It helps to avoid false-negative results now and is designed to continue to be effective well into the future, ensuring affected patients get the medical attention or advice that they need. High sensitivity not only helps to improve individual patient care, it also helps to avoid individuals unknowingly spreading their infection, potentially putting others at risk and further exacerbating the pressures on public health services.

Building Enduring Molecular Tests

We can’t afford to be unprepared for the next pandemic, trying to reactively develop a new test from scratch when we are confronted with an emerging pathogen. This robust technology and approach to molecular diagnostics will be the foundation for building the tools to fight a potential SARS-CoV-3, or whatever the next pathogen will be. Diagnostic companies aim to design molecular tests that offer the broadest possible range of coverage, helping to ensure that we have the capability to recognise both current and future strains. This will help to make our healthcare services as ‘pandemic ready’ as possible to meet future needs.

If the tests are ready — or at least close — for the next pathogen, widespread testing could be integrated earlier to help with infection control procedures. A study of German healthcare services reported that around 20 percent of SARS-CoV-2 cases up to the 21st of September 2021 were associated with outbreaks, with one per cent in hospitals and four per cent in long-term care facilities.⁹ Healthcare services were already under immense strain from the increased number of inpatients, with these outbreaks further exacerbating the issue. Moreover, infections in these environments generally spread disproportionately among the elderly and the vulnerable, often resulting in inferior patient outcomes.⁹

Interestingly, outbreaks in healthcare facilities were more strongly associated with the first and second waves of the pandemic, before widespread vaccinations.⁹ As we have witnessed, once pharmaceutical interventions started to make an impact, morbidity and mortality decreased. If the same is true for future pathogens, infection control procedures — such as accurate and fast molecular testing — could prove critical in managing outbreaks until vaccine and drug development can make an impact. This could involve testing both patients and staff en masse to identify cases earlier, potentially helping to improve patient outcomes, as well as protecting everyone in the hospital environment.

Conclusion

This concept of preparing a test for an uncertain future is an integral part of the product development pipeline at Cepheid. The company has taken the same principles learned from other emerging diseases — such as multidrug-resistant tuberculosis, influenza, Ebola, mpox, and healthcare-associated infections — and is applying them across the board, while also developing innovative technologies focusing on future pandemic preparedness.

Originally published 9/2022.