@article {44717,
	title = {Novel Nested-Seq Approach for SARS-CoV-2 Real-Time Epidemiology and In-Depth Mutational Profiling in Wastewater},
	journal = {Int J Mol Sci},
	volume = {22},
	number = {16},
	year = {2021},
	note = {Avgeris, MargaritisAdamopoulos, Panagiotis GGalani, AikateriniXagorari, MarietaGourgiotis, DimitriosTrougakos, Ioannis PVoulgaris, NikolaosDimopoulos, Meletios-AthanasiosThomaidis, Nikolaos SScorilas, Andreaseng16689/Special Account for Research Grants of the National and Kapodistrian University of Athens/Switzerland2021/08/28 06:00Int J Mol Sci. 2021 Aug 7;22(16):8498. doi: 10.3390/ijms22168498.},
	month = {Aug 7},
	abstract = {Considering the lack of effective treatments against COVID-19, wastewater-based epidemiology (WBE) is emerging as a cost-effective approach for real-time population-wide SARS-CoV-2 monitoring. Here, we report novel molecular assays for sensitive detection and mutational/variant analysis of SARS-CoV-2 in wastewater. Highly stable regions of SARS-CoV-2 RNA were identified by RNA stability analysis and targeted for the development of novel nested PCR assays. Targeted DNA sequencing (DNA-seq) was applied for the analysis and quantification of SARS-CoV-2 mutations/variants, following hexamers-based reverse transcription and nested PCR-based amplification of targeted regions. Three-dimensional (3D) structure models were generated to examine the predicted structural modification caused by genomic variants. WBE of SARS-CoV-2 revealed to be assay dependent, and significantly improved sensitivity achieved by assay combination (94\%) vs. single-assay screening (30\%-60\%). Targeted DNA-seq allowed the quantification of SARS-CoV-2 mutations/variants in wastewater, which agreed with COVID-19 patients{\textquoteright} sequencing data. A mutational analysis indicated the prevalence of D614G (S) and P323L (RdRP) variants, as well as of the Beta.1.1.7/alpha variant of concern, in agreement with the frequency of Beta.1.1.7/alpha variant in clinical samples of the same period of the third pandemic wave at the national level. Our assays provide an innovative cost-effective platform for real-time monitoring and early-identification of SARS-CoV-2 variants at community/population levels.},
	keywords = {*COVID-19/epidemiology/virology, *Pandemics, 2019-nCov, 231-0004089064) and on 22 March 2021 to the Patent Cooperation Treaty (PCT, application number PCT/EP2021/057316). All other authors declare no competing, Covid-19, D614g, Environmental Monitoring/methods, February 2021 to the Hellenic Industrial Property Organisation (case number, Humans, interests., nested PCR, RNA virus in an environmental sample, RNA, Viral/*isolation \& purification, SARS-CoV-2 mutations, SARS-CoV-2 NGS, SARS-CoV-2 sequencing, SARS-CoV-2/*isolation \& purification, Sewage, spike protein, University of Athens, and the authors M.A., P.G.A., N.V., N.S.T. and A.S. on 11, wastewater-based epidemiology, Wastewater/*virology},
	isbn = {1422-0067 (Electronic)1422-0067 (Linking)},
	author = {Avgeris, M. and Adamopoulos, P.G. and Galani, A. and Xagorari, M. and Gourgiotis, D. and Trougakos, I.P. and Voulgaris, N. and Dimopoulos, M.A. and Thomaidis, N.S. and Scorilas, A.}
}
