A Dual Loci Quality Assurance and Control Framework for Real-Time Evaluation of Signal Accuracy in Wastewater Surveillance of Pathogens with High Rates of Mutation DOI Creative Commons
Ocean Thakali, Élisabeth Mercier,

Walaa Eid

et al.

Research Square (Research Square), Journal Year: 2023, Volume and Issue: unknown

Published: Aug. 15, 2023

Abstract Wastewater surveillance of coronavirus disease 2019 (COVID-19) commonly applies reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to quantify severe acute respiratory syndrome 2 (SARS-CoV-2) RNA concentrations in wastewater over time. In most applications worldwide, maximal sensitivity and specificity RT-qPCR has been achieved, part, by monitoring two or more genomic loci SARS-CoV-2. Ontario, Canada, the provincial Surveillance Initiative reports average copies CDC N1 N2 normalized fecal biomarker pepper mild mottle virus. November 2021, emergence Omicron variant concern, harboring a C28311T mutation within probe region, challenged accuracy consensus between measurements this study, we developed applied novel real-time dual quality assurance control framework based on relative difference City Ottawa dataset identify loss assay period from July 10, 2022 January 31, 2023. Further analysis via sequencing allele-specific revealed high proportion mutations C28312T A28330G during study period, both across province. It is hypothesized that nucleotide especially A28330G, led inefficient annealing, resulting reduction assay. This highlights importance implementing criteria continually evaluate, near real-time, signal produced rely detection pathogens whose genomes undergo rates mutation.

Language: Английский

Real-time evaluation of signal accuracy in wastewater surveillance of pathogens with high rates of mutation DOI Creative Commons
Ocean Thakali, Élisabeth Mercier,

Walaa Eid

et al.

Scientific Reports, Journal Year: 2024, Volume and Issue: 14(1)

Published: Feb. 14, 2024

Abstract Wastewater surveillance of coronavirus disease 2019 (COVID-19) commonly applies reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to quantify severe acute respiratory syndrome 2 (SARS-CoV-2) RNA concentrations in wastewater over time. In most applications worldwide, maximal sensitivity and specificity RT-qPCR has been achieved, part, by monitoring two or more genomic loci SARS-CoV-2. Ontario, Canada, the provincial Surveillance Initiative reports average copies CDC N1 N2 normalized fecal biomarker pepper mild mottle virus. November 2021, emergence Omicron variant concern, harboring a C28311T mutation within probe region, challenged accuracy consensus between measurements this study, we developed applied novel real-time dual quality assurance control framework based on relative difference City Ottawa dataset identify loss assay period from July 10, 2022 January 31, 2023. Further analysis via sequencing allele-specific revealed high proportion mutations C28312T A28330G during study period, both across province. It is hypothesized that nucleotide especially A28330G, led inefficient annealing, resulting reduction assay. This highlights importance implementing criteria continually evaluate, near real-time, signal produced rely detection pathogens whose genomes undergo rates mutation.

Language: Английский

Citations

8
Wastewater surveillance monitoring of SARS-CoV-2 variants of concern and dynamics of transmission and community burden of COVID-19 DOI Creative Commons
Maria E. Hasing,

Bonita E. Lee,

Tiejun Gao

et al.

Emerging Microbes & Infections, Journal Year: 2023, Volume and Issue: 12(2)

Published: July 6, 2023

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

Language: Английский

Citations

15
Digital microfluidics with distance-based detection – a new approach for nucleic acid diagnostics DOI
Man Ho, N. Sathishkumar, Alexandros A. Sklavounos

et al.

Lab on a Chip, Journal Year: 2023, Volume and Issue: 24(1), P. 63 - 73

Published: Nov. 21, 2023

There is great enthusiasm for using loop-mediated isothermal amplification (LAMP) in point-of-care nucleic acid tests (POC NAATs), as an alternative to PCR. While techniques like LAMP eliminate the need rapid temperature cycling a portable format, these systems are still plagued by requirements dedicated optical detection apparatus analysis and manual off-chip sample processing. Here, we developed new microfluidic system LAMP-based POC NAATs address limitations. The combines digital microfluidics (DMF) with distance-based (DBD) direct signal readout. This first report of use (i) or (ii) DMF DBD - thus, describe number characterization steps taken determine optimal combinations reagents, materials, processes reliable operation. For example, was found be quite sensitive background signals from low molecular weight products; Capto™ adhere bead-based clean-up procedure isolate desirable high-molecular-weight products analysis. method validated application SARS-CoV-2 saliva. able distinguish between saliva containing no virus, viral load (10

Language: Английский

Citations

15
Emergence and spread of SARS-CoV-2 variants of concern in Canada: a retrospective analysis from clinical and wastewater data DOI Creative Commons
David Champredon, Devan Becker, Shelley Peterson

et al.

BMC Infectious Diseases, Journal Year: 2024, Volume and Issue: 24(1)

Published: Jan. 29, 2024

Abstract Background The spread of SARS-CoV-2 has been studied at unprecedented levels worldwide. In jurisdictions where molecular analysis was performed on large scales, the emergence and competition numerous SARS-CoV-2lineages have observed in near real-time. Lineage identification, traditionally from clinical samples, can also be determined by sampling wastewater sewersheds serving populations interest. Variants concern (VOCs) lineages associated with increased transmissibility and/or severity are particular Method Here, we consider data sources to assess VOCs Canada retrospectively. Results We show that, overall, wastewater-based VOC identification provides similar insights surveillance based samples. Based data, synchrony introduction as well speeds across most Canadian provinces despite geographical size country differences provincial public health measures. Conclusion particular, it took approximately four months for Alpha Delta contribute half incidence. contrast, Omicron achieved same contribution less than one month. This study significant benchmarks enhance planning future VOCs, some extent pandemics caused other pathogens, quantifying rate invasion Canada.

Language: Английский

Citations

5
Hyperplex PCR enables highly multiplexed analysis of point mutations in wastewater: long-term SARS-CoV-2 variant surveillance in Sweden as a case study DOI
Ruben R. G. Soares,

Javier Edo Varg,

Attila Szabó

et al.

Water Research, Journal Year: 2025, Volume and Issue: 274, P. 123154 - 123154

Published: Jan. 16, 2025

Language: Английский

Citations

0
Underestimation of SARS-CoV-2 in wastewater due to single or double mutations in the N1 qPCR probe binding region DOI Creative Commons
Jianxian Sun, Minqing Ivy Yang, Jiaxi Peng

et al.

Water Research X, Journal Year: 2024, Volume and Issue: 22, P. 100221 - 100221

Published: Jan. 1, 2024

Wastewater surveillance using RT-qPCR has now been widely adopted to track circulating levels of SARS-CoV-2 virus in many sewersheds. The CDC qPCR assays targeting two regions (N1 and N2) within the N gene are commonly used, but a discrepancy between biomarkers noticed by independent studies these methods since late 2021. reason is presumed be due mutations targeted N1 probe. In this study, we systematically investigated unequivocally confirmed that underlying for was probe target, single mutation could cause significant drop signal. We first proportion related wastewater samples (Jan 2021-Dec 2022) nested PCR LC-MS. Based on relative proportions alleles, separated data into four time periods corresponding different variant waves: Period I (Alpha Delta waves with 0 mutation), II (BA.1/BA.2 found all Omicron strains), III (BA.5.2* wave mutations), IV (BQ.1* mutations). Significantly lower copies N2 from Periods II-IV compared those observed wastewater. To further pinpoint extent which each impacted quantification, response among synthetic oligomers mutations. This study highlighted impact even just one or qPCR-based SARS-CoV-2.

Language: Английский

Citations

2
Quantification and Differentiation of SARS-CoV-2 Variants in Wastewater for Surveillance DOI Creative Commons
Teresa Kumblathan, Yanming Liu, Xiaoli Pang

et al.

Environment & Health, Journal Year: 2023, Volume and Issue: 1(3), P. 203 - 213

Published: Aug. 9, 2023

Wastewater surveillance plays an important role in the monitoring of infections SARS-CoV-2 at community level. We report here determination and differentiation its variants concern 294 wastewater samples collected from two major Canadian cities May 2021 to March 2023. The overall method analysis involved extraction virus viral components using electronegative membranes, situ stabilization concentration RNA onto magnetic beads, direct on beads. Multiplex reverse transcription quantitative polymerase chain reaction (RT-qPCR) assays, targeting specific naturally selected mutations SARS-CoV-2, enabled detection Alpha, Beta, Gamma, Delta, Omicron variants. An triplex RT-qPCR assay three mutations, HV 69-70 deletion, K417N, L452R, was able detect differentiate BA.1/BA.3, BA.2/XBB, BA.4/5. This had efficiencies 90-104% for all mutation targets a limit 28 copies per reaction. Analyses over two-year span showed concentrations trends as they emerge participating program. were consistent with clinical reports same period. At beginning each wave, corresponding detectable wastewater. For example, BA.2 variant high 104 100 mL January 2022, when approximately only 50-60 cases infection reported Canada. These results show that strategy highly sensitive assays are potentially useful newly emerging other viruses future biomonitoring.

Language: Английский

Citations

3
Underestimation of SARS-CoV-2 in wastewater due to single or double mutations in the N1 qPCR probe binding region DOI Creative Commons
Jianxian Sun, Minqing Ivy Yang, Jiaxi Peng

et al.

medRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Feb. 4, 2024

Abstract Wastewater surveillance using RT-qPCR has now been widely adopted to track circulating levels of SARS-CoV-2 virus in many sewer sheds. The CDC qPCR assays targeting two regions (N1 and N2) within the N gene are commonly used, but a discrepancy between biomarkers noticed by groups this method since late 2021. reason is presumed be due mutations targeted probe. In study, we systematically investigated unequivocally confirmed that underlying for was N1 probe target, single mutation could cause significant drop signal. We first proportion related wastewater samples (Jan 2021-Dec 2022) nested PCR LC-MS. Based on relative alleles, separated data into four time periods corresponding different variant waves: Period I (Alpha Delta waves with 0 mutation), II (BA.1/BA.2 wave found all Omicron strains), III (BA. 5.2* mutations), IV (BQ.1* mutations). Significantly lower copies N2 from Periods II-IV compared those observed wastewater. To further pinpoint extent which each impacted quantification, response among synthetic oligomers mutations. This study highlighted impact even just one or qPCR-based SARS-CoV-2.

Language: Английский

Citations

0
Hyperplex PCR enables the next-generation of wastewater-based surveillance systems: long-term SARS-CoV-2 variant surveillance in Sweden as a case study DOI Open Access
Ruben R. G. Soares,

Javier Edo Varg,

Attila Szabó

et al.

medRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: June 11, 2024

Abstract Wastewater-based epidemiology aims at measuring pathogens in wastewater as a means of deriving unbiased epidemiological information population scale, ranging from buildings and aircrafts to entire cities or countries. After gaining significant mainstream attention during the SARS-CoV-2 pandemic, field holds promise continuous monitoring early warning system tracking emerging viral variants new with pandemic potential. To expand current toolbox analytical techniques for analysis, we explored use Hyperplex PCR (hpPCR) analyse mutations samples collected weekly up 22 sites across Sweden between October 2022 December 2023. Approximately 900 were tested using dynamic probe panel multiplexity 10-to 18-plex, continuously adapted within 1-2 weeks quantify relevant concern over time. The simultaneously covered deletions, single nucleotide substitutions, well variable regions resorting degeneracy. By analysing all parallel gold standard methods including qPCR two different NGS technologies, performance hpPCR is herein shown bridge gap these by providing (1) systematic sensitivity simple design, (2) high without re-optimization requirements (3) 4-5-week earlier mutation detection compared excellent quantitative linearity good correlation frequency (r=0.88). Based on demonstrated performance, authors propose combined routine discovery high-frequency key pathogens/variants potential alternative analysis paradigm.

Language: Английский

Citations

0
A Dual Loci Quality Assurance and Control Framework for Real-Time Evaluation of Signal Accuracy in Wastewater Surveillance of Pathogens with High Rates of Mutation DOI Creative Commons
Ocean Thakali, Élisabeth Mercier,

Walaa Eid

et al.

Research Square (Research Square), Journal Year: 2023, Volume and Issue: unknown

Published: Aug. 15, 2023

Abstract Wastewater surveillance of coronavirus disease 2019 (COVID-19) commonly applies reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to quantify severe acute respiratory syndrome 2 (SARS-CoV-2) RNA concentrations in wastewater over time. In most applications worldwide, maximal sensitivity and specificity RT-qPCR has been achieved, part, by monitoring two or more genomic loci SARS-CoV-2. Ontario, Canada, the provincial Surveillance Initiative reports average copies CDC N1 N2 normalized fecal biomarker pepper mild mottle virus. November 2021, emergence Omicron variant concern, harboring a C28311T mutation within probe region, challenged accuracy consensus between measurements this study, we developed applied novel real-time dual quality assurance control framework based on relative difference City Ottawa dataset identify loss assay period from July 10, 2022 January 31, 2023. Further analysis via sequencing allele-specific revealed high proportion mutations C28312T A28330G during study period, both across province. It is hypothesized that nucleotide especially A28330G, led inefficient annealing, resulting reduction assay. This highlights importance implementing criteria continually evaluate, near real-time, signal produced rely detection pathogens whose genomes undergo rates mutation.

Language: Английский

Citations

1