This article has been updated with information from the Wadsworth Center of the New York State Department of Health.
NEW YORK – Labs and companies across the world are betting on alternative specimen types for COVID-19 testing – such as saliva, nasal swabs, or oral solution – to increase the speed and convenience of sample collection, aiming to overcome one of the challenges in rapidly scaling up testing efforts.
The ability to lift stay-at-home orders and reopen businesses and schools hinges on the availability of large-scale diagnostic testing. While testing numbers across the US have climbed to more than 200,000 tests per day on average over the past week, experts say this is not nearly enough to allow for sufficient surveillance.
Among the reasons for the testing crunch is the cumbersome sample collection. The overwhelming majority of molecular SARS-CoV-2 tests current uses a nasopharyngeal swab, considered the gold standard for many respiratory disease tests, which goes deep into the nasal cavity and needs to be administered by a healthcare professional. Both NP swabs and the viral or universal transport medium (VTM or UTM) to store the sample until testing have been in short supply. In addition, healthcare workers need to wear full protective gear to take the sample, a procedure that is unpleasant and frequently makes subjects cough or sneeze.
But alternatives have emerged over the past few weeks and are increasingly being implemented. For example, New York City Mayor Bill de Blasio said that NYC Health and Hospitals community testing sites will start letting patients collect their own nasal swabs and saliva samples this week, which he said is "simpler but also safer, especially for that healthcare worker." The city plans to use these types of samples "aggressively," he said.
"We need partnership from the private labs who do the processing work," he said, adding that the City has been "engaged in conversations" with labs already about adapting their testing processes.
So far, though, saliva samples and nasal swabs have been rare for COVID-19 testing. The Wadsworth Center of the New York State Department of Health, for example, has been validating saliva paired with nasal swabs as appropriate specimen types, according to a spokesperson, and expects to be able to use them in the field "very soon."
Also, on April 10, the US Food and Drug Administration granted the Rutgers Clinical Genomics Laboratory Emergency Use Authorization for a SARS-CoV-2 RT-PCR test, the first to include saliva as a specimen type. As of Wednesday, the lab, housed at RUCDR Infinite Biologics, remained the only one with FDA clearance for saliva-based coronavirus testing. Its EUA requires the use of a saliva collection device from Spectrum Solutions, which contains patented preservation reagents to stabilize RNA and DNA.
According to Andy Brooks, chief operating officer and director of technology development at RUCDR Infinite Biologics, being able to take saliva samples has increased sample collection volumes and reduced the need for personal protective equipment (PPE) among healthcare workers. "For a lot of the state and government collection facilities, we have tripled, almost quadrupled throughput," he said. "We've reduced PPE by 90 percent and we're letting our health professionals actually take care of patients instead of administer swab tests."
The lab has increased its testing capacity to 10,000 samples per day, he added, and is on track to double capacity by the end of next week, with a goal of getting to 30,000 to 40,000 daily samples in the near future. While the lab supports a variety of hospitals and healthcare networks, including those that use NP swabs, the ratio of swabs to saliva samples "has shifted tremendously" over the past few weeks, Brooks said.
For their EUA submission, the Rutgers scientists compared results from saliva samples with those from nasopharyngeal or oropharyngeal samples for 60 patients suspected of having COVID-19 and found them to be 100 percent concordant, and to have similar sensitivity. "We showed that they are equivalent in overall performance," Brooks said. "However, the robustness of the test is improved with saliva."
Others may follow Rutgers' example soon. Spectrum Solutions, on its website, said it is providing its saliva collection kits to "processing labs, university labs, and hospital labs that are working to validate the established protocols on their equipment to submit for FDA EUA approval for testing saliva samples detecting COVID-19 infection." In addition, Brooks said his team has been helping several commercial labs bring saliva testing online. "I think you're going to see more and more labs that will be doing saliva, for sure," he said.
Independently, groups at the University of Melbourne in Australia and at Yale University have been developing their own SARS-CoV-2 tests that use saliva as input. The Melbourne team, which published its results in the Journal of Clinical Microbiology last week, compared results from NP swabs and saliva in 522 patients who came to the Royal Melbourne Hospital for COVID-19 screening. They found that 85 percent of patients who tested positive with NP swabs also tested positive with saliva. Also, one of the patients testing negative with NP swabs was positive with saliva. The researchers also found that viral load seemed to be higher in NP swabs.
Meanwhile, in a study posted as a preprint on MedRxiv a week ago, the Yale researchers compared results from nasopharyngeal swabs and saliva samples from 44 COVID-19 patients with severe disease and found that virus titers were higher in the saliva samples. Also, in a fifth of 38 matched NP-saliva samples, saliva tested positive while NP swabs were negative, and in 8 percent of matching samples, it was the opposite. In addition, they tested longitudinal samples of each type in a number of hospitalized patients and found that saliva samples were less variable than NP swabs, which sometimes gave a negative result one day and a positive one the next day.
They also enrolled almost 100 asymptomatic healthcare workers in a study that collected one or both sample types every few days. In two cases, saliva tested positive earlier than NP swabs, suggesting that it is more sensitive for detecting mild or subclinical infections.
According to Anne Wyllie, an associate research scientist at the Yale School of Public Health and the lead author of the preprint, which has been submitted to a journal for publication, the team has now expanded its study population to 85 patients and 61 matched samples and "saliva is still performing nicely," with a much lower false negative rate than NP swabs.
She said the reason her group started assessing saliva was a shortage of NP swabs, which they eventually sourced from a company in Taiwan and of viral transport media, which their lab started making in house based on a CDC protocol.
In contrast to the Rutgers test, their protocol does not require a special saliva collection device. The Yale group has been asking participants to spit into standard hospital urine collection cups, and for previous projects that involved saliva, Wyllie said she used standard lab reagent tubes of various sizes.
The response her group has received to its preprint has been enormous. "The number of emails I have received asking for protocols lately suggests a lot of people are looking into [saliva testing] and a lot of people are interested in it" she said, and "shows how desperate people are to have options, especially to test a lot of people in a short amount of time."
However, labs and companies bringing testing online for the first time may be reluctant to move away from the gold standard of NP swabs. "The growing data coming out showing that saliva is a viable option will really help different companies and institutions make these decisions," Wyllie said.
For now, her team continues to build out its dataset comparing NP swabs and saliva. "For research purposes, it's working great for us. We'll continue using it for screening healthcare workers," she said. Likewise, the diagnostic labs at Yale are comparing pairs of NP swabs and saliva in order to establish how they can potentially establish saliva testing. Right now, when a healthcare worker tests positive in the Yale study, she said, that result still needs to be confirmed in a CLIA laboratory using an NP swab.
In addition, her team is looking into other ways of streamlining the testing process, for example, by multiplexing qPCR assays. Right now, three separate qPCR assays – one for each target – need to be run for each sample.
Commercial labs, for the most part, have not ventured into COVID-19 testing with saliva yet. "Saliva samples have not been validated for use with the SARS-CoV-2 RNA tests offered through Quest Diagnostics at this time," said a spokesperson for Quest.
KorvaLabs, an anti-doping testing lab, and Curative, meanwhile, received FDA Emergency Use Authorization for a COVID-19 test last week that includes "oral fluid" as a specimen, a kind of modified saliva sample where patients are asked to cough a few times and then swab the inside of their mouth, which they can do themselves in the presence of a healthcare professional.
LabCorp also is not using saliva at the moment but "is evaluating all options to scale and expand the range of testing as needed," according to a spokesperson, basing its decisions on "quality, validity, availability, and what is required to scale quickly."
Last week, the company received an FDA Emergency Use Authorization for its Pixel by LabCorp COVID-19 Test home collection kit, which will become available in the next few weeks in most US states, with exceptions that include New York and New Jersey. The kit contains a nasal swab that patients can use to collect their own sample from the front of their nose.
Others have been exploring the use of nasal swabs, as well. Earlier this month, for example, a group from France published a paper in the Journal of Clinical Microbiology in which they compared nasal and nasopharyngeal swabs for COVID-19 testing. They found that out of 37 patients that were positive by NP swab, 89 percent also tested positive with nasal swabbing, and all negative NP samples were also negative with nasal swabs.
Senior author David Veyer, a virologist at Georges Pompidou European Hospital in Paris, said that his institution, the Assistance Publique Hôpitaux de Paris, which is part of the largest university hospital group in Europe, had experienced a shortage of NP swabs, prompting his group to test traditional swabs, for which there is no risk of supply disruption. Based on the results of the study, "several laboratories or hospitals of our institutions have adopted nasal swabbing as a convenient alternative in [the] time of [a] nasopharyngeal swab shortage," he said.
Yet another group, at the University of Washington, has tested the use of self-collected dry mid-turbinate swabs without the use of universal transport media. They combined this with a direct RT-PCR testing protocol that does not require RNA extraction, an approach others have also explored. In a BioRxiv preprint posted last week, the UW team reported that their simplified protocol appears to be at least as sensitive as conventional testing, based on results from 11 COVID patients.
Co-senior author Lea Starita, a research assistant professor at UW, said the team developed the protocol in response to "an acute shortage of UTM and RNA extraction reagents." Her group continues to evaluate its performance with more COVID patients and healthy volunteers. It also is studying the stability of viral RNA over time and under different temperatures to mimic sample shipping and the viability of virus in the sample.
While the swabs her team has been using are the regular ones that are still in short supply, she said she hopes that cutting the UTM and RNA extraction reagents from the testing pipeline will be helpful. Also, changes to the upstream logistics side of testing will be needed for volumes to ramp up significantly, she said. "In addition, changes to the regulatory structure of lab testing will be necessary to allow more widespread community testing outside of the healthcare system."