Direct Nasal Swab for Rapid Test and Saliva as an Alternative Biological Sample for RT-PCR in COVID-19 Diagnosis

Abstract

Accurate and early diagnoses are prerequisites for prompt treatment. For coronavirus disease 2019 (COVID-19), it is even more crucial. Currently, choice of methods include rapid diagnostic tests and reverse transcription polymerase chain reaction (RT-PCR) using samples mostly of respiratory origin and sometimes saliva. We evaluated two rapid diagnostic tests with three specimen types using viral transport medium (VTM) containing naso-oropharyngeal (NOP) swabs, direct nasal and direct nasopharyngeal (NP) samples from 428 prospective patients. We also performed RT-PCR for 428 NOP VTM and 316 saliva samples to compare results. The sensitivity of the SD Biosensor Standard Q COVID-19 antigen (Ag) test kit drastically raised from an average of 65.55% (NOP VTM) to 85.25% (direct nasal samples), while RT-PCR was the gold standard. For the CareStart kit, the sensitivity was almost similar for direct NP swabs; the average was 84.57%. The specificities were ≥95% for both SD Biosensor Standard Q and CareStart COVID-19 Ag tests in all platforms. The kits were also able to detect patients with different variants as well. Alternatively, RT-PCR results from saliva and NOP VTM samples showed high sensitivities of 96.45% and 95.48% with respect to each other as standard. The overall results demonstrated high performance of the rapid tests, indicating the suitability for regular surveillance at clinical facilities when using direct nasal or direct NP samples rather than NOP VTM. Additionally, the analysis also signifies not showed that RT-PCR of saliva can be used as an choice of method to RT-PCR of NOP VTM, providing an easier, non-invasive sample collection method. IMPORTANCE There are several methods for the diagnosis of coronavirus disease 2019 (COVID-19), and the choice of methods depends mostly on the resources and level of sensitivity required by the user and health care providers. Still, reverse transcription polymerase chain reaction (RT-PCR) has been chosen as the best method using direct naso-oropharyngeal swabs. There are also other methods of fast detection, such as rapid diagnostic tests (RDTs), which offer result within 15 to 20 min and have become quite popular for self-testing and in the clinical setting. The major drawback of the currently used RT-PCR method is compliance, as it may cause irritation, and patients often refuse to test in such a way. RDTs, although inexpensive, suffer from low sensitivity due to technical issues. In this article, we propose saliva as a noninvasive source for RT-PCR samples and evaluate various specimen types at different times after infection for the best possible output from COVID-19 rapid tests.

Keywords: COVID-19; RT-PCR; SARS-CoV-2; rapid diagnostic tests; saliva; salivary specimen.

FIG 1

Flowchart showing the sample size and the steps of rapid testing and RT-PCR of VTM and saliva in two COVID-19 waves.

FIG 2

A violin curve depicting the distribution of age among the symptomatic male and female patients coming for COVID-19 RT-PCR testing.

FIG 3

A line graph comparing the C_T values for each target (N1 and N2) between nasal VTM and salivary sources.

FIG 4

ROC curve for the SD Biosensor kit presenting the cut points where highest sensitivity and specificity can be attained for N1 and N2 targets with NOP VTM RT-PCR as standard (a) and saliva RT-PCR as standard (b); Std, standard.

FIG 5

(a and b) ROC curve for SD Biosensor test kits using direct nasal samples (a) and ROC curve for CareStart test kits using direct NP samples (b) with NOP VTM RT-PCR as standard. (c and d) ROC curve for SD Biosensor test kits using direct nasal samples (c) and ROC curve for CareStart test kits using direct NP samples (d) with saliva RT-PCR as standard.

FIG 6

Scatter diagram showing the distribution of the C_T values of the infected patients during the first and second waves of COVID-19. The RT-PCR-positive samples that were missed by RDT are shown by the red triangles. In the first wave, only the SD Biosensor kit was used, and both SD Biosensor and CareStart kits were used during the second wave.

FIG 7

Heat map showing 9 major symptoms observed among the symptomatic patients. Each heat map contains day 1 to day 6 and later after onset of symptoms, with the right-sided bar representing the number of patients. (a and b) The total number of patients coming for testing (a) and the patients who were positive by rapid test only (b) are indicated.

FIG 8

Pie chart of the circulating variants during the two COVID-19 waves.