Direct comparison of RT-ddPCR and targeted amplicon sequencing for SARS-CoV-2 mutation monitoring in wastewater

Lou, Esther; Sapoval, Nicolae; McCall, Camille; Bauhs, Lauren; Carlson-Stadler, Russell; Kalvapalle, Prashant; Lai, Yanlai; Palmer, Kyle; Penn, Ryker; Rich, Whitney; Wolken, Madeline; Brown, Pamela J.; Ensor, Katherine B.; Hopkins, Loren; Treangen, Todd J.; Stadler, Lauren B.

doi:10.1016/j.scitotenv.2022.155059

Cited by 40 publications

(49 citation statements)

References 51 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We followed the same RNA extraction, library preparation, and sequencing protocols, as described in prior work 18 . Details on WWTP sample sites, and methods regarding sample collection procedures, and quantification of SARS-CoV-2 in wastewater samples can also be found in our previous publication 18 . Estimates of the viral load are provided by the Houston Health Department following the same methodology as outlined in the SARS-CoV-2 Wastewater Monitoring Dashboard 44 .…”

Section: Discussionmentioning

confidence: 99%

“…In comparison to one of the leading tools for analyzing SARS-CoV-2 wastewater sequencing data for VoC detection, QuaID demonstrated superior sensitivity. This is particularly important given that the underlying sample quality varies and the depth and breadth of coverage of amplicon sequencing data can vary widely across samples 18 . Furthermore, the ability to leverage indel information in the inference process makes QuaID overall more robust than approaches that rely solely on SNVs 9,15 .…”

Section: Discussionmentioning

confidence: 99%

“…Between February 23, 2021 and May 5th, 2022, we collected, processed, and analyzed 2,637 wastewater samples from the fifth-most populous metropolitan area in the US: Houston, Texas. Samples were collected weekly from 39 wastewater treatment plants (WWTPs, Supplementary Table 1, Supplementary Figure 1) distributed throughout the city of Houston and servicing more than 2 million Houston residents 13 . During the study period, the VoC detection signal clearly reflected the three major variants that affected Houston - Alpha, Delta, and Omicron (Figure 1B).…”

Section: Mainmentioning

confidence: 99%

“…Since wastewater represents a pooled sample of multiple hosts, it harbors a diversity of SARS-CoV-2 variants that are currently circulating in the population 1,2,10,13 , including potentially previously unreported genotypes 16 . Furthermore, variant detection and phasing is further complicated by uneven genome coverage 2,17,18 and environmental RNA degradation 5,19,20 which render phased assembly extremely difficult [21][22][23] . Despite these challenges, detection of VoCs in wastewater samples is important for monitoring the emergence and spread of variants and informing public health response 4,5,11,24,25 .…”

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

QuaID: Enabling Earlier Detection of Recently Emerged SARS-CoV-2 Variants of Concern in Wastewater

Sapoval¹,

Lou

Hopkins

et al. 2021

Preprint

Self Cite

View full text Add to dashboard Cite

SARS-CoV-2 RNA shedding in stool enabled wastewater surveillance for the genetic material of the virus. With the emergence of novel variants of concern and interest it becomes increasingly important to track arrival and spread of these variants. However, most current approaches rely on the manually curated lists of mutations phenotypically associated with the variants of concern. The resulting data has many overlaps between distinct variants leading to less specific characterization of complex sample mixtures that result from wastewater monitoring. In our work we propose a simple and specific method for characterization of wastewater samples by introducing the concept of quasi-unique mutations. Our approach is data driven and results in earlier detection and higher resolution of variants of concern emergence patterns in wastewater data.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Mainmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

QuaID: Enabling Earlier Detection of Recently Emerged SARS-CoV-2 Variants of Concern in Wastewater

Sapoval¹,

Lou

Hopkins

et al. 2021

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…Droplet digital PCR (ddPCR) is a modified PCR method where amplification is performed in submicroliter droplets, and the number of positive droplets is enumerated for absolute quantification of the targets or the genomic variations. ddPCR provides the absolute quantification of targets and is also reported to be more sensitive for the detection of known SARS-CoV-2 mutations [ 16 ].…”

Section: Introductionmentioning

confidence: 99%

Confirming Multiplex RT-qPCR Use in COVID-19 with Next-Generation Sequencing: Strategies for Epidemiological Advantage

Carpenter

Tamrakar

Chahar³

et al. 2022

Global Health

View full text Add to dashboard Cite

Rapid identification and tracking of emerging SARS-CoV-2 variants are critical for understanding the transmission dynamics and developing strategies for interrupting the transmission chain. Next-Generation Sequencing (NGS) is an exceptional tool for whole-genome analysis and deciphering new mutations. The technique has been instrumental in identifying the variants of concern (VOC) and tracking this pandemic. However, NGS is complex and expensive for large-scale adoption, and epidemiological monitoring with NGS alone could be unattainable in limited-resource settings. In this study, we explored the application of RT-qPCR-based detection of the variant identified by NGS. We analyzed a total of 78 deidentified samples that screened positive for SARS-CoV-2 from two timeframes, August 2020 and July 2021. All 78 samples were classified into WHO lineages by whole-genome sequencing and then compared with two commercially available RT-qPCR assays for spike protein mutation(s). The data showed good concordance between RT-qPCR and NGS analysis for specific SARS-CoV-2 lineages and characteristic mutations. RT-qPCR assays are quick and cost-effective and thus can be implemented in synergy with NGS for screening NGS-identified mutations of SARS-CoV-2 for clinical and epidemiological interest. Strategic use of NGS and RT-qPCR can offer several COVID-19 epidemiological advantages.

show abstract

SARS‐CoV‐2 in digital era: Diagnostic techniques and importance of nucleic acid quantification with digital PCRs

Aravind Kumar,

Aradhana,

Harleen

et al. 2023

Reviews in Medical Virology

View full text Add to dashboard Cite

Studies related to clinical diagnosis and research of SARS‐CoV‐2 are important in the current pandemic era. Although molecular biology has emphasised the importance of qualitative analysis, quantitative analysis with nucleic acids in relation to SARS‐CoV‐2 needs to be clearly emphasised, which can provide perspective for viral dynamic studies of SARS‐CoV‐2. In this regard, the requirement and utilization of digital PCR in COVID‐19 research has substantially increased during the pandemic, necessitating the aggregation of its cardinal applications and future scopes. Hence, this meta‐review comprehensively addresses and emphasises the importance of nucleic acid quantification of SARS‐CoV‐2 RNA with digital PCR (dPCR). Various quantitative techniques of clinical significance like immunological, proteomic and nucleic acid‐based diagnosis and quantification, have been comparatively discussed. Furthermore, the core part of the article focusses on the working principle and advantages of digital PCR, along with its applications in COVID‐19 research. Several important applications like viral load quantitation, environmental surveillance and assay validation have been extensively investigated and discussed. Certain key future scopes of clinical importance, like mortality prediction, viral/variant‐symbiosis, and antiviral studies were also identified, suggesting several possible digital PCR applications in COVID‐19 research.

show abstract

Direct comparison of RT-ddPCR and targeted amplicon sequencing for SARS-CoV-2 mutation monitoring in wastewater

Cited by 40 publications

References 51 publications

QuaID: Enabling Earlier Detection of Recently Emerged SARS-CoV-2 Variants of Concern in Wastewater

QuaID: Enabling Earlier Detection of Recently Emerged SARS-CoV-2 Variants of Concern in Wastewater

Confirming Multiplex RT-qPCR Use in COVID-19 with Next-Generation Sequencing: Strategies for Epidemiological Advantage

SARS‐CoV‐2 in digital era: Diagnostic techniques and importance of nucleic acid quantification with digital PCRs

Contact Info

Product

Resources

About