ABSTRACT
The use of RNA sequencing from wastewater samples is proven to be a valuable way for estimating infection dynamics and circulating lineages of SARS-CoV-2. This approach has the advantage of being independent from patient population testing and symptomatic disease courses. However, it is equally important to develop easily accessible and scalable tools which can highlight critical changes in infection rates and dynamics over time across different locations given the sequencing data from the wastewater. Here we provide the first analysis of variant dynamics in Germany using wastewater sequencing and present PiGx SARS-CoV-2, a bit-by-bit reproducible end-to-end pipeline with comprehensive reports. To our knowledge, this is the first pipeline that includes all steps from raw-data to shareable reports, additional taxonomic analysis, deconvolution and geospatial time series analysis. Using our pipeline on a dataset of wastewater samples, from different locations across Berlin, over the time period from February 2021 to June 2021, we could reconstruct the dynamic of the Variant of Concern (VoC) B.1.1.7 (alpha). Additionally, we detected the unique signature mutation M:T26767C for the VoC B.1.617.2 (delta) and its raise in early June. We also show that SARS-CoV-2 mutation load measured from wastewater sequencing is correlated with actual case numbers and it has potential to be used in a predictive manner. All in all, our study provides additional evidence that systematic wastewater analysis using sequencing and computational methods can be used for modeling the infection dynamics of SARS-CoV-2. In addition, the results show that our tool can be used to tease out new mutations and to detect any emerging new lineages of concern before clinical detection. Our approach can support efforts for establishing continuous monitoring and early-warning projects for COVID-19 or any other infectious disease.
Subject(s)
Communicable Diseases , COVID-19ABSTRACT
The coronavirus disease 2019 (COVID-19) pandemic, caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an ongoing global health threat with more than two million infected people since its emergence in late 2019. Detailed knowledge of the molecular biology of the infection is indispensable for understanding of the viral replication, host responses, and disease progression. We provide gene expression profiles of SARS-CoV and SARS-CoV-2 infections in three human cell lines (H1299, Caco-2 and Calu-3 cells), using bulk and single-cell transcriptomics. Small RNA profiling showed strong expression of the immunity and inflammation-associated microRNA miRNA-155 upon infection with both viruses. SARS-CoV-2 elicited approximately two-fold higher stimulation of the interferon response compared to SARS-CoV in the permissive human epithelial cell line Calu-3, and induction of cytokines such as CXCL10 or IL6. Single cell RNA sequencing data showed that canonical interferon stimulated genes such as IFIT2 or OAS2 were broadly induced, whereas interferon beta (IFNB1) and lambda (IFNL1-4) were expressed only in a subset of infected cells. In addition, temporal resolution of transcriptional responses suggested interferon regulatory factors (IRFs) activities precede that of nuclear factor-{kappa}B (NF-{kappa}B). Lastly, we identified heat shock protein 90 (HSP90) as a protein relevant for the infection. Inhibition of the HSP90 charperone activity by Tanespimycin/17-N-allylamino-17-demethoxygeldanamycin (17-AAG) resulted in a reduction of viral replication, and of TNF and IL1B mRNA levels. In summary, our study established in vitro cell culture models to study SARS-CoV-2 infection and identified HSP90 protein as potential drug target for therapeutic intervention of SARS-CoV-2 infection.