Tracking SARS-CoV-2 in Sewage: Evidence of Changes in Virus Variant Predominance during COVID-19 Pandemic
Abstract
:1. Introduction
2. Materials and Methods
2.1. Wastewater Sample Collection and Processing
2.2. Quantification of SARS-CoV-2 RNA in Wastewater Concentrates by RT-qPCR
2.3. SARS-CoV-2 Whole-Genome Sequences Used for Nucleotide Sequence Analyses
2.4. Nested RT-PCR (nPCR) Amplification
2.5. Sanger Sequencing Analysis of RT-PCR Products
2.6. NGS Analysis of RT-PCR Products
2.7. Generation of SARS-CoV-2 Sequence Contigs and Identification of SNPs
3. Results
3.1. Detection of SARS-CoV-2 RNA in Wastewater Samples
3.2. Analysis of Nucleotide Sequence Variation among SARS-CoV-2 RNA Sequences from Clinical Samples
3.3. Generation of nPCR Products for Nucleotide Sequence Analyses
3.4. Nucleotide Sequence Analysis of nPCR Products from Wastewater Concentrates
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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SARS-CoV-2 RTqPCR (No. of Replicates) 1 | Nested RT-PCR Gene Target (PCR Product Size) 2 | Genome Sequenced (No. of Nucleotides) | ||||||
---|---|---|---|---|---|---|---|---|
Sampling Date | RdRP Gene (log10 gc/L Sewage) | E Gene (log10 gc/L Sewage) | nsp2-PLPro Gene nPCR1 (714 nt) | RdRP Gene nPCR2 (523 nt) | RdRP Gene nPCR3 (527 nt) | RdRP Gene nPCR4 (235 nt) | ORF8b-N Gene nPCR5 (612 nt) | |
14-Jan-20 | - | - | - | - | - | - | - | - |
11-Feb-20 | - | - | - | - | - | + | + | 847 |
11-Mar-20 | 4.84 ± 0.45 [4.18–5.52] (n = 10) | 4.98 ± 0.40 [4.63–5.41] (n = 3) | + | + | + | + | + | 2376 |
14-Apr-20 | 5.27 ± 0.30 [4.77–5.91] (n = 11) | 5.78 ± 0.07 [5.71–5.84] (n = 3) | + | + | + | + | + | 2376 |
12-May-20 | <3.5 (n = 11) 3 | - | + | + | + | + | + | 2376 |
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Martin, J.; Klapsa, D.; Wilton, T.; Zambon, M.; Bentley, E.; Bujaki, E.; Fritzsche, M.; Mate, R.; Majumdar, M. Tracking SARS-CoV-2 in Sewage: Evidence of Changes in Virus Variant Predominance during COVID-19 Pandemic. Viruses 2020, 12, 1144. https://doi.org/10.3390/v12101144
Martin J, Klapsa D, Wilton T, Zambon M, Bentley E, Bujaki E, Fritzsche M, Mate R, Majumdar M. Tracking SARS-CoV-2 in Sewage: Evidence of Changes in Virus Variant Predominance during COVID-19 Pandemic. Viruses. 2020; 12(10):1144. https://doi.org/10.3390/v12101144
Chicago/Turabian StyleMartin, Javier, Dimitra Klapsa, Thomas Wilton, Maria Zambon, Emma Bentley, Erika Bujaki, Martin Fritzsche, Ryan Mate, and Manasi Majumdar. 2020. "Tracking SARS-CoV-2 in Sewage: Evidence of Changes in Virus Variant Predominance during COVID-19 Pandemic" Viruses 12, no. 10: 1144. https://doi.org/10.3390/v12101144
APA StyleMartin, J., Klapsa, D., Wilton, T., Zambon, M., Bentley, E., Bujaki, E., Fritzsche, M., Mate, R., & Majumdar, M. (2020). Tracking SARS-CoV-2 in Sewage: Evidence of Changes in Virus Variant Predominance during COVID-19 Pandemic. Viruses, 12(10), 1144. https://doi.org/10.3390/v12101144