Wastewater surveillance monitoring of SARS-CoV-2 variants of concern and dynamics of transmission and community burden of COVID-19.

Wastewater-based surveillance is a valuable approach for monitoring COVID-19 at community level. Monitoring SARS-CoV-2 variants of concern (VOC) in wastewater has become increasingly relevant when clinical testing capacity and case-based surveillance are limited. In this study, we ascertained the turnover of six VOC in Alberta wastewater from May 2020 to May 2022. Wastewater samples from nine wastewater treatment plants across Alberta were analysed using VOC-specific RT-qPCR assays. The performance of the RT-qPCR assays in identifying VOC in wastewater was evaluated against next generation sequencing. The relative abundance of each VOC in wastewater was compared to positivity rate in COVID-19 testing. VOC-specific RT-qPCR assays performed comparatively well against next generation sequencing; concordance rates ranged from 89% to 98% for detection of Alpha, Beta, Gamma, Omicron BA.1 and Omicron BA.2, with a slightly lower rate of 85% for Delta (p < 0.01). Elevated relative abundance of Alpha, Delta, Omicron BA.1 and BA.2 were each associated with increased COVID-19 positivity rate. Alpha, Delta and Omicron BA.2 reached 90% relative abundance in wastewater within 80, 111 and 62 days after their initial detection, respectively. Omicron BA.1 increased more rapidly, reaching a 90% relative abundance in wastewater after 35 days. Our results from VOC surveillance in wastewater correspond with clinical observations that Omicron is the VOC with highest disease burden over the shortest period in Alberta to date. The findings suggest that changes in relative abundance of a VOC in wastewater can be used as a supplementary indicator to track and perhaps predict COVID-19 burden in a population.

Rapid transition between SARS-CoV-2 variants of concern Delta and Omicron detected by monitoring municipal wastewater from three Canadian cities.

Monitoring the communal incidence of COVID-19 is important for both government and residents of an area to make informed decisions. However, continuous reliance on one means of monitoring might not be accurate because of biases introduced by government policies or behaviours of residents. Wastewater surveillance was employed to monitor concentrations of SARS-CoV-2 RNA in raw influent wastewater from wastewater treatment plants serving three Canadian Prairie cities with different population sizes. Data obtained from wastewater are not directly influenced by government regulations or behaviours of individuals. The means of three weekly samples collected using 24 h composite auto-samplers were determined. Viral loads were determined by RT-qPCR, and whole-genome sequencing was used to charaterize variants of concern (VOC). The dominant VOCs in the three cities were the same but with different proportions of sub-lineages. Sub-lineages of Delta were AY.12, AY.25, AY.27 and AY.93 in 2021, while the major sub-lineage of Omicron was BA.1 in January 2022, and BA.2 subsequently became a trace-level sub-variant then the predominant VOC. When each VOC was first detected varied among cities; However, Saskatoon, with the largest population, was always the first to present new VOCs. Viral loads varied among cities, but there was no direct correlation with population size, possibly because of differences in flow regimes. Population is one of the factors that affects trends in onset and development of local outbreaks during the pandemic. This might be due to demography or the fact that larger populations had greater potential for inter- and intra-country migration. Hence, wastewater surveillance data from larger cities can typically be used to indicate what to expect in smaller communities.

RNA in Municipal Wastewater Reveals Magnitudes of COVID-19 Outbreaks across Four Waves Driven by SARS-CoV-2 Variants of Concern.

There are no standardized protocols for quantifying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater to date, especially for population normalization. Here, a pipeline was developed, applied, and assessed to quantify SARS-CoV-2 and key variants of concern (VOCs) RNA in wastewater at Saskatoon, Canada. Normalization approaches using recovery ratio and extraction efficiency, wastewater parameters, or population indicators were assessed by comparing to daily numbers of new cases. Viral load was positively correlated with daily new cases reported in the sewershed. Wastewater surveillance (WS) had a lead time of approximately 7 days, which indicated surges in the number of new cases. WS revealed the variant α and δ driving the third and fourth wave, respectively. The adjustment with the recovery ratio and extraction efficiency improved the correlation between viral load and daily new cases. Normalization of viral concentration to concentrations of the artificial sweetener acesulfame K improved the trend of viral load during the Christmas and New Year holidays when populations were dynamic and variable. Acesulfame K performed better than pepper mild mottle virus, creatinine, and ammonia for population normalization. Hence, quality controls to characterize recovery ratios and extraction efficiencies and population normalization with acesulfame are promising for precise WS programs supporting decision-making in public health.

Distribution of heavy metal resistance elements in Canadian Salmonella 4,[5],12:i:- populations and association with the monophasic genotypes and phenotype.

Salmonella 4,[5],12:i:- are monophasic S. Typhimurium variants incapable of producing the second-phase flagellar antigen. They have emerged since the mid-1990s to become one of the most prevalent Salmonella serotypes causing human disease world-wide. Multiple genetic events associated with different genetic elements can result in the monophasic phenotype. Several jurisdictions have reported the emergence of a Salmonella 4,[5],12:i:- clone with SGI-4 and a genetic element (MREL) encoding a mercury resistance operon and antibiotic resistance loci that disrupts the second phase antigen region near the iroB locus in the Salmonella genome. We have sequenced 810 human and animal Canadian Salmonella 4,[5],12:i:- isolates and determined that isolates with SGI-4 and the mercury resistance element (MREL; also known as RR1&RR2) constitute several global clades containing various proportions of Canadian, US, and European isolates. Detailed analysis of the data provides a clearer picture of how these heavy metal elements interact with bacteria within the Salmonella population to produce the monophasic phenotype. Insertion of the MREL near iroB is associated with several deletions and rearrangements of the adjacent flaAB hin region, which may be useful for defining human case clusters that could represent outbreaks. Plasmids carrying genes encoding silver, copper, mercury, and antimicrobial resistance appear to be derived from IS26 mediated acquisition of these genes from genomes carrying SGI-4 and the MREL. Animal isolates with the mercury and As/Cu/Ag resistance elements are strongly associated with porcine sources in Canada as has been shown previously for other jurisdictions. The data acquired in these investigations, as well as from the extensive literature on the subject, may aid source attribution in outbreaks of the organism and interventions to decrease the prevalence of this clone and reduce its impact on human disease.

Complete Genome Sequences of Six Salmonella enterica Serovar 4,[5],12:i:- Isolates from Canada.

Salmonella enterica with antigenic formula 4,[5],12:i:- is a monophasic variant of S. enterica serovar Typhimurium that has emerged globally as a human pathogen over the last 3 decades. We describe the closed genomes and plasmids from six S. enterica 4,[5],12:i:- isolates recovered from stool samples obtained during investigation of human food poisoning cases reported to PulseNet Canada that are suitable for use as reference strains.