A Study of Analytical and Clinical Sensitivity of Aptima SARS-CoV-2 Assay (Hologic) and Proposals of Complementary Tests for SARS-CoV-2 Detection in Low Viral Load Specimens

Abstract

Early and accurate detection of SARS-CoV-2 is important for diagnosis and transmission control. The use of high-throughput and automated testing allows laboratories to better deliver diagnostic testing given manpower and resource limitations. We validated the clinical and analytical performance of the Hologic Panther Aptima SARS-CoV-2 assay with an emphasis on detection of specimens with low viral loads. The clinical performance was evaluated using 245 clinical specimens, against a comparator PCR-based laboratory developed test (LDT). The analytical performance was determined by replicate testing of contrived samples in a ten-fold dilution series ( C T values 32–42, based on LDT). The Aptima assay had 96.7% overall percent agreement, 100% negative percent agreement and 88.1% positive percent agreement. It was able to consistently detect SARS-CoV-2 in contrived samples with C T = 32 by LDT (calculated 2354 copies/mL). The 95% limit of detection of the Aptima assay was estimated to be at LDT C T = 33 (equivalent to 870 copies/mL). The relative light units (RLU) × 1000 for 52 true positive clinical specimens was 962.2 ± 181.5, and that for the 186 true negative specimens was 264.6 ± 14.3. The Aptima assay was a reliable method with a high overall percent agreement against our comparator LDT. We propose that samples reported as negative by the Aptima assay with RLU > 350 be tested by a secondary method, in order to improve detection of samples with very low viral loads. Supplementary Information The online version contains supplementary material available at 10.1007/s00284-021-02730-3.

Role of Hologic® Panther AptimaTM SARS-CoV-2 assay in the detection of SARS-CoV-2: screening or diagnostic technique?

“…Other comparator studies showed similar values of sensitivity but higher PPA values [7,8,9]. The differences are probably due to the dissimilar way of sample selection.…”

“…Several studies have previously evaluated Hologic Panther Aptima assay ability for SARS-CoV-2 detection in comparison to different qPCR commercial kits [ 5 , 6 , 7 , 8 , 9 ]. The efficacy of the technique has already been demonstrated, although it is true that a previous study by our group [ 10 ] showed that the sensitivity of Hologic Panther Aptima assay is lower than TaqPath COVID-19 RT-PCR.…”

Evaluation of SARS-CoV-2 Detection Systems Using Clinical Samples and Standard Material: A Comparative Study

“…This TMA method targets two unique regions of the SARS-CoV-2 ORF-1ab gene and proceeds in a one-step process under isothermal conditions. Previous studies reported the low LoD of the Aptima SARS-CoV-2 assay, ranging from 62.5 to 870 copies/mL [ 3 , 5 , 6 ]. Based on this low LoD, Kimberly et al used the assay as a COVID-19 screening tool using 10/1 pooled samples [ 9 ].…”

“…The Aptima SARS-CoV-2 assay (Hologic, Bedford, MA, USA) is a transcription-mediated amplification (TMA) assay that uses an automated system with a turnaround time of 3.5 h, and can provide random access for reporting results of up to 60 samples per hour continuously thereafter [ 3 ]. Clinical comparative data have been obtained for the Aptima SARS-CoV-2 assay [ 3 , 4 , 5 , 6 , 7 , 8 ]), but to our knowledge, there have been no studies comparing the Aptima SARS-CoV-2 assay and the Real-Q Direct SARS-CoV-2 detection kit (Biosewoom, Seoul, Republic of Korea), which is currently used in our institution, with the same clinical samples. There have also been no studies of the analytical performance of the Real-Q Direct SARS-CoV-2 detection kit using reference materials.…”

Evolution and Impact of Nucleic Acid Amplification Test (NAAT) for Diagnosis of Coronavirus Disease

“…112 Hologic’s Aptima SARS-CoV-2 Assay employs TMA, while direct RNA amplification requires reverse transcriptase with the T7 RNA polymerase enzyme. Several authorized NATs based on TMA use the Aptima SARS-CoV-2 Assay, which is the Poplar SARS-CoV-2 TMA Pooling Assay, 113 the Quest Diagnostics HA SARS-CoV-2 Assay, 114 the Let us Get Checked Coronavirus Assay (COVID-19) TEST, 115 and the Procleix SARS-CoV-2 Assay, 116 which detects ORF1ab genes in two regions and an IC, which is intended to be carried out using the Panther System. The target RNA molecule is extracted with the help of a capture oligomer with a sequence complementary to the target protein’s specific region and a chain of deoxyadenosine residues.…”