Sample collection and transport strategies to enhance yield, accessibility, and biosafety of COVID-19 RT-PCR testing

Abstract

Introduction. Non-invasive sample collection and viral sterilizing buffers have independently enabled workflows for more widespread COVID-19 testing by reverse-transcriptase polymerase chain reaction (RT-PCR).Gap statement. The combined use of sterilizing buffers across non-invasive sample types to optimize sensitive, accessible, and biosafe sampling methods has not been directly and systematically compared.Aim. We aimed to evaluate diagnostic yield across different non-invasive samples with standard viral transport media (VTM) versus a sterilizing buffer eNAT- (Copan diagnostics Murrieta, CA) in a point-of-care diagnostic assay system.Methods. We prospectively collected 84 sets of nasal swabs, oral swabs, and saliva, from 52 COVID-19 RT-PCR-confirmed patients, and nasopharyngeal (NP) swabs from 37 patients. Nasal swabs, oral swabs, and saliva were placed in either VTM or eNAT, prior to testing with the Xpert Xpress SARS-CoV-2 (Xpert). The sensitivity of each sampling strategy was compared using a composite positive standard.Results. Swab specimens collected in eNAT showed an overall superior sensitivity compared to swabs in VTM (70 % vs 57 %, P=0.0022). Direct saliva 90.5 %, (95 % CI: 82 %, 95 %), followed by NP swabs in VTM and saliva in eNAT, was significantly more sensitive than nasal swabs in VTM (50 %, P<0.001) or eNAT (67.8 %, P=0.0012) and oral swabs in VTM (50 %, P<0.0001) or eNAT (58 %, P<0.0001). Saliva and use of eNAT buffer each increased detection of SARS-CoV-2 with the Xpert; however, no single sample matrix identified all positive cases.Conclusion. Saliva and eNAT sterilizing buffer can enhance safe and sensitive detection of COVID-19 using point-of-care GeneXpert instruments.

Keywords: Inactivation; Nasal; Oral; Saliva; eNAT.

Fig. 1.

Study flowchart.

Fig. 2.

Comparative testing of different respiratory specimens using the Xpert Xpress SARS-CoV-2 test. (A) Percent positive rate and (B) N2 gene cycle threshold (Ct) values of samples from all participants with at least one SARS-CoV-2 positive sample (N=84 for all samples and N=37 for NP swab). NP=Nasopharyngeal: VTM=Viral transport medium; eNAT=eNAT transport media, Copan diagnostics. ns=not statistically different. ****P<0.0001; ***P<0.001, **P=0.02.

Fig. 3.

eNAT as a transport media for saliva. Saliva diluted with eNAT at 1 : 1, 1 : 2, and 1 : 4 ratio showing (A) Percent positive rate and (B) N2-Ct values from patient saliva samples tested directly (N=44), as a swab in eNAT (N=44), diluted 1 : 1 (N=44), 1 : 2 (N=42), and 1 : 4 (N=42) in eNAT transport media. ns=not statistically significant.