Background and objective:
Three SARS-CoV-2 Variants of Concern (VoCs) are circulating in the European Union/European Economic Area (EU/EEA) countries as of December 2022: Omicron BA.2, BA.4, and BA.5. ECDC also currently recognizes three Variants of Interest (VOIs): Omicron BA.2.75, BQ.1, and XBB.1. This work analyses the occurrence and spread of VoCs and VoIs in Italy in October and November 2022, by monitoring untreated wastewater.

Methods:
During October and November 2022, 332 wastewater samples were collected from 167 Wastewater Treatment Plants throughout Italy. Variant characterization was performed by amplifying a ~1600 bp fragment of the spike protein, followed by Sanger (individual samples) and long-read nanopore sequencing (pooled samples by Regions/Autonomous Provinces).

Results:
In October 2022, mutations characteristic of Omicron BA.4/BA.5 were detected in the vast majority of the samples. Amino acid substitutions characteristic of sub-lineages BQ.1 (K444T and N460K) or BQ.1.1 (BQ.1 + R346T) were detected for the first time in wastewater in four Regions/A.P. despite the low prevalence documented in clinical cases at the time of sampling. In November 2022, the positivity rate for BQ.1/BQ.1.1 almost quadrupled, and the number of Regions/A.P. positive for the new Omicron subvariant tripled compared to October. Moreover, in November there was the detection of two other VoIs never observed before in sewage in Italy: BA.2.75 and XXB.1 (the latter found in a Region where no clinical cases associated with this variant had been documented).

Conclusions:
Results suggest that BQ.1/BQ.1.1 is becoming dominant in late 2022, as predicted by the ECDC, and other minor subvariants are emerging, confirming environmental surveillance as a powerful tool for tracking the spread of SARS-CoV-2 variants/subvariants in the population.