Wastewater monitoring using a novel, cost-effective PCR-based method that rapidly captures the transition patterns of SARS-CoV-2 variant prevalence (from Delta to Omicron) in the absence of conventional surveillance evidence

Abstract

Background: The magnitude of Omicron SARS-CoV-2 variant of concern (VOC) global spread necessitates reinforcement of surveillance implementation. Conventional VOC surveillance based on genotyping of clinical samples is characterized by certain challenges related to available sequencing capacity, population sampling methodologies, and demands in terms of time, labor, and resources. Wastewater-based SARS-CoV-2 VOC surveillance constitutes a valuable supplementary practice, since it does not require extensive sampling, and provides information on the prevalence of the disease in a timely and cost-effective manner. Methods: A highly sensitive real-time RT-PCR assay was developed, for targeted Omicron VOC detection and quantification in wastewater samples. The assay exclusively amplifies sequences with the S:Δ69/70 deletion, thus its performance is not hampered by the presence of the Delta VOC. The method was incorporated in the analysis of composite daily samples taken from the main Wastewater Treatment Plant (WWTP) of Thessaloniki, Greece from 1 December 2021 to 9 January 2022. Results: The Omicron VOC was detected for the first time in samples from the Thessaloniki WWTP on 19 December 2021. In the following 10-day period, a rapid increase in Omicron sewage viral load was observed with an estimated early doubling time of 1.86 days. The proportion of the total SARS-CoV-2 load attributed to Omicron reached 91.09% on 7 January, revealing a fast Delta-to-Omicron VOC transition pattern. The detection of Omicron in wastewater preceded the outburst of reported (presumable) Omicron cases in the city by approximately 7 days. Conclusions: The proposed wastewater-based surveillance approach enables rapid, real-time data acquisition on the Omicron VOC prevalence and transmission dynamics. Timely provision of these results to State authorities can readily influence the decision-making process for targeted public health interventions, including control measures, awareness, and preparedness.

Monitoring of SARS-CoV-2 variant dynamics in wastewater by digital RT-PCR : from Alpha to Omicron BA.2 VOC

“…These approaches have been developed and applied to the emergence of Alpha and Delta VOC. To our knowledge, few data are available regarding the emergence and the dynamics of the Omicron BA.1 and BA.2 VOC (Agrawal et al, 2022b; Ahmed et al, 2022; Chassalevris et al, 2022; Kirby et al, 2022; Smyth et al, 2022).…”

A qualitative RT-PCR assay for the specific identification of the SARS-CoV-2 B.1.1.529 (Omicron) Variant of Concern

“…To the best of our knowledge, when this paper was published as a preprint [ 19 , 28 ], OmMet was the first single one-step RT-PCR assay reported for the specific detection of Omicron. Since then, two other PCR-based assays have been developed for the detection of Omicron in clinical and environmental samples [ 29 , 30 ]. These methods take advantage of other unique features present in the S gene of Omicron (e.g.…”

“…typical deletions or insertions) and have been described to be highly specific and sensitive for Omicron. However, according to our in silico amplification, the del69-70 based assays as described in [ 29 , 30 ], should not be able to detect the lineage Omicron BA.2, which is currently (end of April 2022) prevalent, as it is missing that particular deletion.…”

A RT-qPCR system using a degenerate probe for specific identification and differentiation of SARS-CoV-2 Omicron (B.1.1.529) variants of concern

“…Recently, several single one-step RT-qPCRs reported for the specific detection of Omicron in clinical and environmental samples have been developed [19][20][21]. As Omicron continues to spread and evolve into several lineages and sub-lineages, these methods suffer, as they are not able to differentiate between multiple Omicron sub-lineages, although they have been described as highly specific and sensitive for Omicron.…”