Decreased Sensitivity of Rapid Antigen Test Is Associated with a Lower Viral Load of Omicron than Delta SARS-CoV-2 Variant

ABSTRACT Large-scale screening for SARS-CoV-2 infection is an important tool for epidemic prevention and control. The appearance of new variants associated with specific mutations can call into question the effectiveness of rapid diagnostic tests (RDTs) deployed massively at national and international levels. We compared the clinical and virological characteristics of individuals infected by Delta or Omicron variants to assess which factors were associated with a reduced performance of RDT. A commercially available RDT as well as the evaluation of the viral load (VL) and the detection of replicate intermediates (RIs) were carried out retrospectively on positive SARS-CoV-2 nasopharyngeal specimens from health care workers of the Pitié-Salpêtrière Hospital infected by the Delta or Omicron variant between July 2021 and January 2022. Of the 205 samples analyzed (104 from individuals infected with Delta and 101 with Omicron), 176 were analyzed by RDT and 200 by RT-PCR for VL and RIs. The sensitivity of the TDR for Omicron was significantly lower than that observed for Delta (53.8% versus 74.7%, respectively, P < 0.01). Moreover, the Delta VL was significantly higher than that measured for Omicron (median Ct 21.2 versus 24.1, respectively, P < 0.01) and associated with the positivity of the RDT in multivariate analysis. We demonstrate a lower RDT sensitivity associated with a lower VL at the time of diagnosis on Omicron-infected individuals in comparison to those infected with the Delta variant. This RDT lower sensitivity should be taken into account in the large-scale screening strategy and in particular in case of strong suspicion of infection where testing should be repeated. IMPORTANCE Previous reports have shown a variability in the diagnostic performance of RDTs. In the era of SARS-CoV-2 variants and the use of RDT, mutation associated with these variants could affect the test performance. We evaluate the sensitivity of the RDT Panbio COVID-19 Ag (Abbott) with two variants of concern (VOC), the Delta and Omicron variants. In order to investigate whether clinical characteristics or virological characteristics can affect this sensitivity, we collected clinical information and performed a specific RT-PCR that detected the RIs as a marker of the viral replication and viral cycle stage. Our results showed that Omicron was less detected than the Delta variant. A lower viral load of Omicron variant in comparison to Delta variant explained this decreased sensitivity, even if they are at the same stage of the disease and the viral cycle and should be taken into account with the use of RDT as diagnostic tool.

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The ASM Journals program strives for constant improvement in our submission and publication process. Please tell us how we can improve your experience by taking this quick Author Survey. The authors assessed the factors that affect the reduced performance of RDT among individuals infected with the Delta and Omicron variants. The paper is well written, and I must commend the researchers for a good job done.
I have therefore attached minor comments for your attention METHODS Line 105: Replaced categorical variables with discrete variables How was positive RDT defined in your study. Was it a binary or continuous variable and how was it coded? Also, the researchers should list the number of factors that were included in the logistic regression model here in the methods section and indicate how they were measured and coded.
Reviewer #2 (Comments for the Author): This is a interesting manuscript where a reduction in sensitivity of rapid antigen test for SARS-CoV-2 Omicron variant compared to delta variant is reported. Moreover, a lower viral load associated to omicron variant would explain the reduction in sensitivity.
However, I have some comments to be address prior to publication: 1. The authors should make a comment in the discussion about a potential bias on the sampling. With a 100 samples of each variant, it could be possible that the lower viral load in omicron would be associated to this sampling and not to the variant itself.
2. Mean Ct values are really low either in delta or in omicron samples group, so the viral loads are high in general. So, the authors should consider in the discussion other reasons for the sensitivity reduction other than the viral load. I do not believe that a sensitivity of 54% for omicron variants is only associated to low viral loads when the avarage Ct is 24.
3. There is a clear difference in the number of vaccine shots between delta an omicron individuals. 46% on omicron individuals got 3 shots, and only 11% for delta. The authors should analyze and discuss how this fact may explain the results reported. A stronger immunization could either cause a lower viral load or direct scavenge viral antigens.
4. If possible, the authors should also include previous infection as a variable. Probably, there are reinfected individuals in the omicron group and that could explain either the reduced viral load or the reduced sensitivity of the test for the same reasons discussed in point 3.

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Thank you for submitting your paper to Microbiology Spectrum.
The authors assessed the factors that affect the reduced performance of RDT among individuals infected with the Delta and Omicron variants. The paper is well written, and I must commend the researchers for a good job done.
I have therefore attached minor comments for your attention

Line 105: Replaced categorical variables with discrete variables
How was positive RDT defined in your study. Was it a binary or continuous variable and how was it coded? Also, the researchers should list the number of factors that were included in the logistic regression model here in the methods section and indicate how they were measured and coded.

Reviewer #1
The authors assessed the factors that affect the reduced performance of RDT among individuals infected with the Delta and Omicron variants. The paper is well written, and I must commend the researchers for a good job done.

I have therefore attached minor comments for your attention
We would like to thank the reviewer for his kind review and comments.

Line 105: Replaced categorical variables with discrete variables
As suggested by the reviewer we have modified the line 105.

How was positive RDT defined in your study? Was it a binary or continuous variable and how was it coded?
In this study, positive RDT was defined according to the manufacturer's recommendations, with the appearance of bands, even a simple trace, at two specifics heights on the RDT: one marking SARS-CoV-2 antigen positivity and the other marking successful completion of the technique. It was a binary variable coded "negative" or "positive".
3. Also, the researchers should list the number of factors that were included in the logistic regression model here in the methods section and indicate how they were measured and coded.
We fully agree with the reviewer that adding informations of the variables included in the logistic regression model could improve our paper. Variables included in the multivariate analysis appeared in the Table 2 and we have added in the methods section how these variables were measured and coded (lines 110 to 115).

Reviewer #2 (Comments for the Author):
This is a interesting manuscript where a reduction in sensitivity of rapid antigen test for SARS-CoV-2 Omicron variant compared to delta variant is reported. Moreover, a lower viral load associated to omicron variant would explain the reduction in sensitivity.
However, I have some comments to be address prior to publication: We would like to thank the reviewer for his kind review and comments.
1. The authors should make a comment in the discussion about a potential bias on the sampling. With a 100 samples of each variant, it could be possible that the lower viral load in omicron would be associated to this sampling and not to the variant itself.
We agree with the reviewer about this potential bias. As suggested, we have added a comment about this limitation in the discussion section (line 177).

Mean
Ct values are really low either in delta or in omicron samples group, so the viral loads are high in general. So, the authors should consider in the discussion other reasons for the sensitivity reduction other than the viral load. I do not believe that a sensitivity of 54% for omicron variants is only associated to low viral loads when the average Ct is 24.
We also agree about the questioning reduce sensitivity for Omicron variant even with an estimated high viral load. Nevertheless, we observed in our study, in absence of difference on the delay between the symptoms onset and sampling, a shift of 3 Ct (median Ct of 21.2 for Delta variant and 24.1 for Omicron variant), which lead to a potential 10-fold decrease of the viral load for Omicron variant in comparison to Delta variant, which could explain partially this reduce sensitivity. We also discuss this point in the other reviewer's remarks.
3. There is a clear difference in the number of vaccine shots between delta an omicron individuals.
46% on omicron individuals got 3 shots and only 11% for delta. The authors should analyze and discuss how this fact may explain the results reported. A stronger immunization could either cause a lower viral load or direct scavenge viral antigens.
We thank the reviewer about this particular point. Indeed 4. If possible, the authors should also include previous infection as a variable. Probably, there are reinfected individuals in the omicron group and that could explain either the reduced viral load or the reduced sensitivity of the test for the same reasons discussed in point 3.
Unfortunately, we do not have this information for the majority of the participants and to avoid any declarative bias (as remembering or asymptomatic infection), we have decided to not include this variable. However, we have added this point as a limit of the study (line 179).