Challenges to global control of SARS-CoV-2 have underscored the need for accurate, safe, and rapidly scalable laboratory testing to inform rapid diagnosis and implementation of appropriate clinical and public health measures. Furthermore, early detection of emerging infectious disease threats is critical to limit spread. Over the last 4 decades, 1-3 newly emerging pathogens have been identified annually [1]. The risk of emerging infections is increasing, in part due to closer proximity of humans to wildlife and expanding geographic ranges of insect and arthropod vectors [2,3]. Outbreaks of diseases caused by respiratory tract infections, such as COVID-19, severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), pandemic H1N1 influenza, and avian influenza, have specifically illustrated the global health burden posed by respiratory viruses with RNA genomes, prompting the implementation of public health policies and efforts to monitor and contain outbreaks [4,5]. Due to the genetic diversity of respiratory viral pathogens and the overlap between syndromes associated with distinct viruses, contemporary disease control efforts generally rely on a combination of methods, including rapid identification of emerging viral strains through targeted polymerase chain reaction (PCR)–based methods coupled with sequencing-based methods to enable molecular epidemiologic tracking of transmission. These activities can be streamlined by new methods of viral detection, such as metagenomic next-generation sequencing (NGS), to simultaneously detect and sequence respiratory viruses from human specimens [6].

Lower per-sample sequencing costs, accessible results through user-friendly analysis tools, and rapid turnaround of sequencing data have led to a growing number of diagnostic and public health laboratories applying NGS-based pathogen detection and profiling, which has enhanced and accelerated the detection and surveillance of emerging viruses [7-9]. Reliable detection of respiratory viruses requires robust methods for sample collection and preservation of viral RNA during transport because poor sample quality can lead to missed identification of significant pathogenic viruses [10]. Traditionally, laboratory diagnosis of respiratory viral infections has relied on collection of nasopharyngeal swabs by health care providers and uninterrupted cold chain transport to testing laboratories. In a previous study, evaluation of nasopharyngeal swabs from individuals with a known respiratory infection by metagenomic RNA sequencing showed high diagnostic agreement with a commercially available reverse transcription–PCR (RT-PCR) viral respiratory panel in a clinical laboratory setting, generated partial or full-length sequences in 86% of known positive samples, and detected additional viruses compared to the commercial panel [11].

However, community- and household-based sampling strategies offer potential advantages over sampling by health care workers in preparation for and during an outbreak. Specifically, anterior nasal swabs represent a noninvasive means for the detection and profiling of respiratory viruses via molecular diagnostic methods, provided appropriate measures to maintain sample integrity are taken [12-14]. Furthermore, nasal swabs can be collected at home by patients or their family members, limiting opportunities for further transmission from exposed individuals within health care facilities. Recent work has demonstrated high percent positive agreement in the detection of SARS-CoV-2 by targeted RT-PCR from self-collected nasal swabs compared to health care provider–collected nasopharyngeal swabs [15-17]. The feasibility of self-collected nasal swabs for detection of pathogen carriage and infection has also been previously demonstrated in an outpatient clinic setting for PCR-based detection of influenza from symptomatic patients [12,14]. Studies to date have focused on PCR-based detection of specific pathogens, and relatively few studies have directly compared the yield from participants to trained researchers or clinical workers.

In this feasibility study, we assessed detection of respiratory viruses by shotgun RNA sequencing using a room temperature–stable at-home nasal swab sampling kit, comparing swabs collected by participants to swabs collected by trained research staff. We also determined the acceptability to participants of collecting nasal swabs from their children and potential barriers to collection at home.

In this pilot, proof-of-concept study, we found that respiratory samples (anterior nasal swabs) self-collected from adults and children who were trained via online video and written instructions had comparable detection of respiratory viruses to samples concurrently collected by trained research personnel, with transport at ambient temperatures, as assessed by unbiased detection of respiratory viruses by NGS. All participants found the home collection procedures acceptable, and 85% reported minimal or no difficulty collecting the samples.

Our findings are consistent with prior studies that have found home collection of anterior nasal swabs to be acceptable in general populations, with high levels of good-quality samples [13,19-24]. Our results are also consistent with a growing body of studies indicating high agreement between self-collected swabs in adults and children (by parents) and swabs collected by health care workers or research personnel; κ ≥0.85 has been reported for the detection of several respiratory viruses, including influenza viruses, SARS-CoV-2, and metapneumovirus [12,13,17,25-30]. To our knowledge, this study is the first to evaluate agreement in self-collected versus researcher-collected nasal swabs specifically for shotgun RNA sequencing.

We considered any risk of nasal injury by self-collection to be minimal and outweighed by the potential benefit of understanding the feasibility and acceptability of self-collection of nasal samples for future research and public health surveillance. In one large, community-based study of over 4000 individuals given home collection kits with nasal swabs to be used on themselves or family members, there was one mild adverse event that did not require any medical attention and had no long-term impacts [31].

This method could be applied to test exposed or symptomatic individuals without the need for visits to a health care facility during outbreaks of emerging respiratory viruses [24,30,32]. In addition, it could streamline surveillance for known or emerging infectious disease threats by application to sentinels. Prior studies have supported the contributions of self-collected swab submission for molecular testing as a supplement to passive and syndromic surveillance for acute respiratory infections [11,14,19,20,32,33]. Guidance from the United States Food and Drug Administration (FDA) to permit the submission of self-collected swabs for SARS-CoV-2 testing illustrates the advantages of this strategy to reduce opportunities for transmission from infected individuals to health care workers and vulnerable populations and to mitigate bottlenecks associated with sample collection by providers [34]. The nasal swab collection kits used in this study have a shelf life of 3 years and stabilize samples at ambient temperature for up to 30 days. This allows for economical stocking and transport of collection kits during an outbreak or in support of surveillance efforts.

This study had several limitations. The sample size was small in this pilot study. Recent respiratory illness was self-reported and no information was collected about the duration of symptoms or the time between onset of symptoms and sample collection. It is therefore possible that individuals may no longer have been shedding virus at the time of sampling. Diagnostic performance of metagenomic sequencing was not compared to a reference method (eg, FDA-cleared PCR panels for the detection of respiratory pathogens), although high agreement has been previously reported [11].

We detected a diverse range of respiratory viruses, determined viral genome sequences in the majority of cases, and showed that viral genomes were identical in staff- and self-collected swabs. Our identification of a divergent strain of HRV-C with only 74% nucleotide sequence identity with the most similar known strain demonstrates the power of unbiased pathogen detection by metagenomics in the context of the well-described diversity of rhinoviruses [35,36]. This suggests an advantage of sequencing-based surveillance for generating viral genomes for high-resolution molecular typing. Studies in hospitalized patients have shown that this genotypic information can assist in efforts to contain transmission [37-39].

In summary, our findings support a growing body of evidence that at-home nasal swab kits for viral detection can be reliably used, are well-tolerated, and provide a scalable solution for the detection and profiling of respiratory viruses. In addition to prior work showing the reliability of self-collected nasal swabs with PCR-based diagnostic tests, our findings suggest that self-collected nasal swabs can be reliable for shotgun RNA sequencing. As metagenomics is integrated into routine diagnostic workflows in a rapidly growing number of laboratories, pathogen genotypic information generated as part of diagnostic testing can be more easily shared and is likely to prove valuable in support of surveillance research and public health applications.