Similar viral loads in Omicron infections regardless of vaccination status

BackgroundAlthough SARS-CoV-2 booster vaccinations are underway, breakthrough infections with Omicron variants are occurring. This study analyzed associations between Omicron sublineage (BA.1.1 and BA.2) viral load and vaccination history. MethodsViral loads in nasopharyngeal swabs were evaluated by quantitative real-time PCR, and the virus strain was evaluated by whole-genome analysis or TaqMan assay. ResultsA total of 611 patients positive for an Omicron SARS-CoV-2 variant were included; 199 were unvaccinated, 370 had received two vaccine doses, and 42 had received three doses. Similar viral loads and Ct values of BA.1.1 and BA.2 were detected regardless of vaccination history. No correlations between age and BA.1.1 and BA.2 viral load were observed. ConclusionOmicron-infected patients who had received a third vaccine dose had viral loads similar to patients with two doses or who were unvaccinated.

SARS-CoV-2 Omicron sublineage BA.2 replaces BA.1.1: genomic surveillance in Japan from September 2021 to March 2022

ObjectiveThe new emerging Omicron strain of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently spreading worldwide. We aimed to analyze the genomic evolution of the shifting Omicron virus subtypes. MethodsThe study included 1,297 individuals diagnosed as SARS-CoV-2 positive by PCR test or antigen quantification test from September 2021 to March 2022. Samples were analyzed by whole genome sequencing analysis (n=489) or TaqMan assay (n=808). ResultsAfter the outbreak of the SARS-CoV-2 Delta strain, the Omicron strain spread rapidly in Yamanashi, Japan. BA.1.1 was the predominant sublineage of the Omicron strain from January to mid-February 2022, but the number of cases of sublineage BA.2 began to increase after mid-February, and this sublineage was shown to have replaced BA.1.1 by the end of March 2022. We observed higher viral and antigen levels of sublineage BA.2 than of sublineage BA.1.1 in nasopharyngeal swab samples. However, no difference in viral load by patient age was apparent between sublineages BA.1.1 and BA.2. ConclusionsA transition from sublineage BA.1.1 to sublineage BA.2 was clearly observed over approximately one month. Omicron sublineage BA.2 was found to be more transmissible owing to its higher viral load regardless of patient age.

Classification of Omicron BA.1, BA.1.1 and BA.2 sublineages by TaqMan assay consistent with whole genome analysis data

ObjectiveRecently, the Omicron strain of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread and replaced the previously dominant Delta strain. Several Omicron sublineages (BA.1, BA.1.1 and BA.2) have been identified, with in vitro and preclinical reports showing that the pathogenicity and therapeutic efficacy differs between BA.1 and BA.2. We sought to develop a TaqMan assay to identify these subvariants. MethodsA TaqMan assay was constructed for rapid identification and genotyping of Omicron sublineages. We analyzed three characteristic mutations of the spike gene, {Delta}69-70, G339D and Q493R, by TaqMan assay. The accuracy of the TaqMan assay was examined by comparing its results with the results of whole genome sequencing (WGS) analysis. ResultsA total of 169 SARS-CoV-2 positive samples were analyzed by WGS and TaqMan assay. The 127 samples determined as BA.1/BA.1.1 by WGS were all positive for {Delta}69-70, G339D and Q493R by TaqMan assay. Forty-two samples determined as BA.2 by WGS were negative for {Delta}69-70 but positive for G339D and Q493R by TaqMan. The concordance rate between WGS and the TaqMan assay was 100% (169/169). ConclusionTaqMan assays targeting characteristic mutations are useful for identification and discrimination of Omicron sublineages.

Pooling RT-PCR test of SARS-CoV-2 for large cohort of 'healthy' and infection-suspected patients: A prospective and consecutive study on 1,000 individuals

BackgroundSARS-CoV-2 testing reagents are expected to become in short supply worldwide. However, little is unknown whether the pooling strategy detects SARS-CoV-2 with accuracy. MethodTo validate the feasibility of pooling samples, serial dilution analysis and spike-in experiment were conducted using synthetic DNA and nucleic acids extracted from SARS-CoV-2 positive and negative patients. Furthermore, we studied a total of 1,000 individuals, who were 667 "healthy" (195 healthcare workers and 472 hospitalized patients with other disorders than COVID-19 infection) individuals and 333 infection-suspected patients with cough and fever, were tested. ResultsSerial dilution analysis showed the limit of detection of around 10-100 copies according to National Institute of Infectious Diseases, Japan. Spike-in experiment demonstrated RT-qPCR detect positive signal in pooling samples of SARS-CoV-2 negative and positive patient at the 5-, 10-, 20-fold dilution. By screening with pooling strategy by the end of April, 2020, there are 12 COVID-19 patients in 333 infection suspected patients (3.6%) and zero in 667 "healthy". We obtained these results with total running 538 times (instead of 1,000 times) by pooling strategy. ConclusionPooling samples is feasible for saving test reagents and detecting SARS-CoV-2 in clinical setting to prevent the spread of the virus and nosocomial transmission.