The study tracks the rapid shift from Omicron BA.1 to BA.2 subvariant dominance in Sweden
In a recently published study to medRxiv* preprint server, researchers tracked the rapid shift from the new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant of anxiety (VOC) Omicron BA.1 subvariant dominance to BA.2 subvariant dominance in Sweden between January and March 2022
Estimates revealed that during the Omicron wave in January 2022, the daily number of new cases of coronavirus disease 2019 (COVID-19) increased to almost 50 million worldwide, far exceeding the daily peak of 14 million COVID-19 cases due to Delta VOC in April 2021, which represents an unprecedented transfer of Omicron.
An analysis of reverse transcription-polymerase chain reaction (RT-PCR) that can genotype Omicron BA.1 cases directly in primary SARS-CoV-2 RT-PCR testing enabled the detection of BA.1-positive on a large scale in Sweden daily, as helped to source data for the current study.
About the study
In the current study, the researchers used an improved, more sensitive multiplex version of ribonucleic acid (RNA) extraction-free protocol from the Centers for Disease Control and Prevention (CDC) SARS-CoV-2 RT-PCR assay. This assay used an increased sample and reaction volume to enable simultaneous detection of general SARS-CoV-2 infection status, ribonuclease P (RNase P) sample integrity, and Omicron BA.1 variant status.
The analysis tracked Omicron BA.1 to BA.2 subvariant transition by analyzing 174,933 nasopharyngeal samples collected from several districts in Central Sweden between January 26 and March 8, 2022, on a daily basis. The nasopharyngeal swabs were anonymized and subjected to heat inactivation prior to RT-PCR testing.
The Public Health Agency gave in vitro expanded Omicron BA.1 reference test, used for nail (S)BA1 evaluation of primer-probe sets.
The researchers performed RT-PCR in eight replicates per concentration of serially diluted (1:10) Omicron BA.1-positive clinical trials to determine the log-linear cycle threshold (C)T) range for the nucleocapsid (N1) and SBA1 primer probe sets under assay conditions. Furthermore, they performed the analysis in the presence and absence of SBA1 the primer probe set for 185 clinical samples to evaluate the probe’s effect on N1 CT values.
Finally, they used whole genome sequencing (WGS) to sequence the entire SARS-CoV-2 genome.
Study result
The Omicron BA.1 subvariant was the dominant variant among covid-19-positive cases in January 2022; However, the BA.1 fraction steadily decreased to 11% in March 2022, and the Omicron BA.2 sub-variant eventually competed with the BA.1 sub-variant across all healthcare districts in Central Sweden. In addition, Omicron BA.2-infected samples had almost twice as much viral RNA as BA.1-infected samples, which partly explains how the BA.2 sub-variant currently competes with the BA.1 variant globally.
The increased amount of virus in the upper respiratory tract (pharynx) of Omicron BA.2 case samples indicated its higher infectivity compared to BA.1 subvariant, but this was a surprising revelation. The researchers thus exposed 3,392 samples to N1 / RP / SBA1 analysis and an extraction-based assay directed against N- and RNA-dependent RNA polymerase (RdRp), located 5 ‘inside the open reading frame (ORF) 1ab. RT-PCR analysis for the N gene showed linear correlation for both N and RdRp in the extraction-based assay, confirming the increased number of virus copies in BA.1-negative COVID-19 samples in both the assays and the genes probed. In addition, WGS confirmed that all 118 BA.1-negative samples were of BA.2 lineage in this assay.
All 698 samples that tested SARS-CoV-2-positive in RT-PCR analysis were genotyped Omicron BA.1-negative by whole genome sequencing, which shows that Omicron BA.2 was the variant that competed with BA.1 in the Swedish population.
The authors also observed the BA.2-specific ORF3a: H78Y mutation in approximately 40% of BA.2 cases classified by WGS. These case samples belonged almost exclusively to the southernmost Swedish region, where they accounted for 72% of covid-19 cases (103/144). In striking contrast to previous reports from southern Europe, WGS showed no cases of covid-19 due to Delta / Omicron recombination or concomitant infection.
Conclusions
The RT-PCR assay used in the current study strongly differentiated general COVID-19 status and Omicron BA.1 variant status in a single RT-PCR reaction, enabling real-time monitoring of the Omicron BA.1 / BA.2 transition in Sweden on a mass scale.
Furthermore, this assay can monitor the Omicron subvariant transition where needed, saving time and challenges associated with genotyping by WGS sequencing due to its compatibility with the well-established CDC N1 / RNaseP probe sets.
*Important message
medRxiv publishes preliminary scientific reports that are not peer-reviewed and therefore should not be considered as crucial, guide clinical practice / health-related behavior or be treated as established information.