Durability of the immune response to a third BNT162b2 dose; five months follow-up

BackgroundTwo doses of the BNT162b2 vaccine yielded high effectiveness that wanes within several months. The third dose was effective in mounting a significant humoral and cellular immune response.. MethodsWe followed BNT162b2-vaccinated health-care workers monthly for IgG and neutralizing antibody (NeutAb) titers. Avidity, T-cell activation and microneutralization of sera against different variants of concern (VOC) were assessed for a sub-cohort. Linear mixed models were used to compare the durability of the second and third doses, and to assess if Omicron breakthrough infections were associated with waning dynamics. ResultsOverall 3972 participants with a third dose were followed, the rate of waning of IgG and NeutAb was slower after the third (1.32%/day and 1.32%/day, respectively) compared to the second (2.26%/per day and 3.34%/day) dose. Live-neutralization of Omicron VOC was lower compared to previous strains and demonstrated similar waning from 111 (95%CI:75-166) to 26 (95%CI:16-42) within 4 months. Mean T cell activity decreased from 98{+/-}5.4 T cells/106 PBMC to 59{+/-}9.3, within 3-5 months. Omicron breakthrough infections were associated with lower IgG peak (ratio of means 0.86 95%CI 0.80-0.91), and among participants over 65y with faster waning of both IgG and NeutAb (ratio of mean rates 1.40 95% CI 1.13-1.68 and 3.58 95% CI 1.92-6.67). No waining in IgG avidity was obsereved during 112 days after the 3rd dose. ConclusionThe third dose is more durable than the second dose, yet Omicron is relatively resistant to direct neutralization. The level of humoral response may predict breakthrough infections.

NOVEL RT-qPCR ASSAYS ENABLE RAPID DETECTION AND DIFFERENTIATION BETWEEN SARS-COV-2 OMICRON (BA.1) AND BA.2 VARIANTS

In this report, we describe the development and initial validation of novel SARS-COV-2 Omicron-specific reactions that enable the identification of Omicron (BA.1) and BA.2 variants. Mutations that are either shared by both BA.1 and BA.2, or are exclusive for BA.1 or for BA.2 were identified by bioinformatic analysis, and corresponding probe-based quantitative PCR reactions were developed to identify them. We show that multiplex combinations of these reactions provide a single-reaction identification of the sample as BA.1, BA.2, or as non-Omicron SARS-COV-2. All four reactions described herein have a sensitivity of less than ten copies per reaction, and are amendable for multiplexing. The results of this study suggest that the new assays may be useful for testing both clinical and environmental samples to differentiate between these two variants.

Fourth Dose COVID mRNA Vaccines' Immunogenicity & Efficacy Against Omicron VOC

BACKGROUNDFollowing the emergence of the Omicron variant of concern, we investigated immunogenicity, efficacy and safety of BNT162b2 or mRNA1273 fourth dose in an open-label, clinical intervention trial. METHODSPrimary end-points were safety and immunogenicity and secondary end-points were vaccine efficacy in preventing SARS-CoV-2 infections and COVID-19 symptomatic disease. The two intervention arms were compared to a matched control group. Eligible participants were healthcare-workers (HCW) vaccinated with three BNT162b2 doses, and whose IgG antibody levels were [≤]700 BAU (40-percentile). IgG and neutralizing titers, direct neutralization of live VOCs, and T-cell activation were assessed. All participants were actively screened for SARS-CoV-2 infections on a weekly basis. RESULTSOf 1050 eligible HCW, 154 and 120 were enrolled to receive BNT162b2 and mRNA1273, respectively, and compared to 426 age-matched controls. Recipients of both vaccine types had a [~]9-10-fold increase in IgG and neutralizing titers within 2 weeks of vaccination and an 8-fold increase in live Omicron VOC neutralization, restoring titers to those measured after the third vaccine dose. Breakthrough infections were common, mostly very mild, yet, with high viral loads. Vaccine efficacy against infection was 30% (95%CI:-9% to 55%) and 11% (95%CI:-43% to +43%) for BNT162b2 and mRNA1273, respectively. Local and systemic adverse reactions were reported in 80% and 40%, respectively. CONCLUSIONSThe fourth COVID-19 mRNA dose restores antibody titers to peak post-third dose titers. Low efficacy in preventing mild or asymptomatic Omicron infections and the infectious potential of breakthrough cases raise the urgency of next generation vaccine development. Trial registration numberclicaltrials.gov: NCT05231005, NCT05230953

NATIONAL SCALE REAL-TIME SURVEILLANCE OF SARS-COV-2 VARIANTS DYNAMICS BY WASTEWATER MONITORING IN ISRAEL

In this report, we describe a national-scale monitoring of the SARS-COV-2 (SC-2) variant dynamics in Israel, using multiple-time sampling of twelve wastewater treatment plants. We used a combination of inclusive and selective quantitative PCR assays that specifically identify variants A19 or B.1.1.7 and tested each sample for the presence and relative viral RNA load of each variant. We show that between December-2020 and March-2021, a complete shift in the SC-2 variant circulation was observed, where the B.1.1.7 replaced the A19 in all examined test points. We further show that the normalized viral load (NVL) values and the average new cases per week reached a peak in January 2021, and then decreased gradually in almost all test points, in parallel with the progression of the national vaccination campaign, during February-March 2021. This study demonstrates the importance of monitoring SC-2 variant dynamics on a national scale through wastewater sampling. It also provides a proof-of-concept methodology for continuous surveillance by using a combination of inclusive and selective PCR tests, which is far more amendable for high throughput monitoring compared with sequencing. This approach may be useful for real-time dynamics surveillance of current and future variants, such as the Omicron (BA.1) variant. SynopsisThis study describes the continuous monitoring of the SARS CoV-2 variant B.1.1.7 circulation in wastewater in Israel using a positive/negative quantitative PCR assay.

Superior immunogenicity and effectiveness of the 3rd BNT162b2 vaccine dose

In a prospective cohort study involving 12,413 Health Care Workers (HCW), we assessed immunogenicity, vaccine-effectiveness (VE) and safety of the third BNT162b2 vaccine dose. One month after third dose, anti-RBD-IgG were induced 1.7-folds compared to one month after the second. A significant increase in avidity from 61.1% (95%CI:56.1-66.7) to 96.3% (95%CI:94.2-98.5) resulted in a 6.1-folds neutralizing antibodies induction. Linear mixed model demonstrated that the third dose elicited a greater response among HCW[≥]60 or those with [≥]two comorbidities who had a lower response following the second dose. VE of the third dose relative to two doses was 85.6% (95% CI, 79.2-90.1%). No serious adverse effects were reported. These results suggest that the third dose is superior to the second dose in both quantity and quality of IgG-antibodies and safely boosts protection from SARS-CoV-2 infection by generating high avidity antibodies to levels that are not significantly different between healthy and vulnerable populations.

Third BNT162b2 vaccination neutralization of SARS-CoV-2 Omicron infection

Using isolates of SARS-CoV-2 WT, Beta, Delta and most importantly Omicron we studied the capability of the BNT162b2 vaccine given in two or three doses to neutralize major SARS-CoV-2 variants of concern (VOC). We demonstrate low neutralization efficiency against delta and wild-type for vaccines with more than 5 months following the second BNT162b2 dose, with no neutralization efficiency against Omicron. We demonstrate the importance of a third dose, by showing a 100-fold increase in neutralization efficiency of Omicron following a third dose, with a 4-fold reduced neutralization compared to that against the Delta VOC. The durability of the effect of the third dose is yet to be determined.

SPECIFIC DETECTION OF SARS-COV-2 B.1.1.529 (OMICRON) VARIANT BY FOUR RT-qPCR DIFFERENTIAL ASSAYS

In this report, we describe four RT-qPCR assays that enable rapid identification of the newly emerging SARS-COV-2 Omicron (B.1.1.529) variant of concern. The assays target Omicron characteristic mutations in the nsp6 (Orf1a), spike and nucleocapsid genes. We demonstrate that the assays are straightforward to assemble and perform, are amendable for multiplexing, and may be used as a reliable first-line tool to identify B.1.1.529 suspected samples. Importantly, this is a preliminary development report. Further validation and optimization of the assays described herein will be published hereafter.

SPECIFIC DETECTION OF SARS-COV-2 VARIANTS B.1.1.7 (ALPHA) AND B.1.617.2 (DELTA) USING A ONE-STEP QUANTITATIVE PCR ASSAY

In this report, we describe the development of an RT-qPCR assay, termed Alpha Delta assay, which can detect SARS-COV-2 (SC-2) and distinguish between the Alpha (B.1.1.7) and Delta (B.1.617.2) variants. The Alpha- and Delta-specific reactions in the assay target mutations that are strongly linked to the target variant. The Alpha reaction targets the D3L substitution in N gene, and the Delta reaction targets the spike gene 156-158 mutations. Additionally, we developed a second Delta-specific assay, used as a confirmatory test for the Alpha Delta assay that targets the 119-120 deletion in the Orf8 gene. Both reactions have similar sensitivities of 15-25 copies per reaction, similar to the sensitivity of commercial SC-2 detection tests. The Alpha Delta assay and the Orf8-119del assay were successfully used to classify clinical samples that were subsequently analyzed by whole genome sequencing. Lastly, we show that the Alpha Delta and Orf8-119del assays correctly identified the presence of Alpha and Delta lineages RNA in wastewater samples. This study provides a rapid, sensitive and cost-effective tool for detecting and classifying two worldwide dominant SC-2 variants. It also highlights the importance of a timely diagnostic response to the emergence of new SC-2 variants with significant consequences on global health.

The unique evolutionary dynamics of the SARS-CoV-2 Delta variant

The SARS-Coronavirus-2 (SARS-CoV-2) driven pandemic was first recognized in late 2019, and the first few months of its evolution were relatively clock-like, dominated mostly by neutral substitutions. In contrast, the second year of the pandemic was punctuated by the emergence of several variants that bore evidence of dramatic evolution. Here, we compare and contrast evolutionary patterns of various variants, with a focus on the recent Delta variant. Most variants are characterized by long branches leading to their emergence, with an excess of non-synonymous substitutions occurring particularly in the Spike and Nucleocapsid proteins. In contrast, the Delta variant that is now becoming globally dominant, lacks the signature long branch, and is characterized by a step-wise evolutionary process that is ongoing. Contrary to the "star-like" topologies of other variants, we note the formation of several distinct clades within Delta that we denote as clades A-E. We find that sequences from the Delta D clade are dramatically increasing in frequency across different regions of the globe. Delta D is characterized by an excess of non-synonymous mutations, mostly occurring in ORF1a/b, some of which occurred in parallel in other notable variants. We conclude that the Delta surge these days is composed almost exclusively of Delta D, and discuss whether selection or random genetic drift has driven the emergence of Delta D.

BNT162b2 Vaccination efficacy is marginally affected by the SARS-CoV-2 B.1.351 variant in fully vaccinated individuals.

BackgroundIsraeli has vaccinated over 80% of its adult population, with two doses of the Pfizer BNT162b2 vaccine. This intervention has been highly successful in curtailing the coronavirus 2 outbreak. One major concern is the ability of the virus to mutate which potentially can cause SARS-CoV-2 to partially escape from the immune system. Here we evaluate the efficacy of the Pfizer vaccine against the B.1.351 variant. MethodsThe Ministry of Health, initiated sequencing of selected positive swab samples identified as being of interest. We used logistic regression, with variant type as the dependent variable, vaccination status as the main explanatory variable, controlling for age, sex, subpopulation, place of residence and time of sample, to estimate the odds ratio for a vaccinated case to have the B. 1.351 versus the B.1.1.7 variant, within vaccinated and unvaccinated persons who tested positive. FindingsThere were 19 cases of B.1.351 variant (3.2%) among those vaccinated more than 14 days before the positive sample and 88 (3.5%) among the unvaccinated. The estimated odds ratio was 1.29 [95% CI: 0.66-2.50]. From this result, assuming the efficacy against the B.1.1.7 variant to be 95%, the estimated efficacy against the B.1.351 variant was 94% [95% CI: 87-97%]. InterpretationDespite the concerns caused by the B.1.351 variant, the BNT162b2 vaccine seems to provide substantial immunity against both that variant and the B.1.1.7. Our results suggest that from 14 days following the second vaccine dose the efficacy of BNT162b2 vaccine is at most marginally affected by the B.1.351 variant. FundingNo funding

The rise and fall of an emerging SARS-CoV-2 variant with the spike protein mutation L452R

Emerging SARS-CoV-2 variants may threaten global vaccination efforts and awaited reduction in outbreak burden. In this study, we report a novel variant carrying the L452R mutation that emerged from a local B.1.362 lineage, B.1.362+L452R. The L452R mutation is associated with the Delta and Epsilon variants and was shown to cause increased infection and reduction in neutralization in pseudoviruses. Indeed, the B.1.362+L452R variant demonstrated a X4-fold reduction in neutralization capacity of sera from BNT162b2-vaccinated individuals compared to a wild-type strain. The variant infected 270 individuals in Israel between December 2020 and March 2021, until diminishing due to the gain in dominance of the Alpha variant in February 2021. This study demonstrates an independent, local emergence of a variant carrying a critical mutation, L452R, which may have the potential of becoming a variant of concern and emphasizes the importance of routine surveillance and detection of novel variants among efforts undertaken to prevent further disease spread.

Rapid And high throughput RT-qPCR assay for identification and differentiation between SARS-CoV-2 variants B.1.1.7 and B.1.351

Emerging SARS-CoV-2 (SC-2) variants with increased infectivity and vaccine resistance are of major concern. Rapid identification of such variants is important for the public health activities and provide valuable data for epidemiological and policy decision making. We developed a multiplex quantitative RT-qPCR (qPCR) assay that can specifically identify and differentiate between the emerging B.1.1.7 and B.1.351 SC-2 variants. In a single assay, we combined four reactions: one that detects SC-2 RNA independently of the strain, one that detects the D3L mutation, which is specific to variant B.1.1.7, and one that detects the 242-244 deletion, which is specific to variant B.1.351. The fourth reaction identifies human RNAseP gene, serving as an endogenous control for RNA extraction integrity. We show that the strain-specific reactions target mutations that are strongly associated with the target variants, and not with other major known variants. The assays specificity was tested against a panel of respiratory pathogens (n=16), showing high specificity towards SC-2 RNA. The assays sensitivity was assessed using both In-vitro transcribed RNA and clinical samples, and was determined to be between 20 and 40 viral RNA copies per reaction. The assay performance was corroborated with Sanger and whole genome sequencing, showing complete agreement with the sequencing results. The new assay is currently implemented in the routine diagnostic work at the Central Virology Laboratory, and may be used in other laboratories to facilitate the diagnosis of these major worldwide circulating SC-2 variants.

The neutralization potency of anti-SARS-CoV-2 therapeutic human monoclonal antibodies is retained against novel viral variants

A wide range of SARS-CoV-2 neutralizing monoclonal antibodies (mAbs) were reported to date, most of which target the spike glycoprotein and in particular its receptor binding domain (RBD) and N-terminal domain (NTD) of the S1 subunit. The therapeutic implementation of these antibodies has been recently challenged by emerging SARS-CoV-2 variants that harbor extensively mutated spike versions. Consequently, the re-assessment of mAbs, previously reported to neutralize the original early-version of the virus, is of high priority. Four previously selected mAbs targeting non-overlapping epitopes, were evaluated for their binding potency to RBD versions harboring individual mutations at spike positions 417, 439, 453, 477, 484 and 501. Mutations at these positions represent the prevailing worldwide distributed modifications of the RBD, previously reported to mediate escape from antibody neutralization. Additionally, the in vitro neutralization potencies of the four RBD-specific mAbs, as well as two NTD-specific mAbs, were evaluated against two frequent SARS-CoV-2 variants of concern (VOCs): (i) the B.1.1.7 variant, emerged in the UK and (ii) the B.1.351 variant, emerged in South Africa. Variant B.1.351 was previously suggested to escape many therapeutic mAbs, including those authorized for clinical use. The possible impact of RBD mutations on recognition by mAbs is addressed by comparative structural modelling. Finally, we demonstrate the therapeutic potential of three selected mAbs by treatment of K18-hACE2 transgenic mice two days post infection with each of the virus strains. Our results clearly indicate that despite the accumulation of spike mutations, some neutralizing mAbs preserve their potency against SARS-CoV-2. In particular, the highly potent MD65 and BL6 mAbs are shown to retain their ability to bind the prevalent novel viral mutations and to effectively protect against B.1.1.7 and B.1.351 variants of high clinical concern.

A unique SARS-CoV-2 spike protein P681H strain detected in Israel

Routine detection, surveillance and reporting of SARS-CoV-2 novel variants is important, as these threaten to hinder vaccination efforts. Herein we report a local novel strain that includes a non-synonymous mutation in the spike (S) protein - P681H and additional synonymous mutations. The P681H Israeli strain has not been associated with higher infection rates and was neutralized by sera from vaccinated individuals in comparable levels to the B.1.1.7 strain and a non-P681H strain from Israel.

HiSpike: A high-throughput cost effective sequencing method for the SARS-CoV-2 spike gene

The changing nature of the corona virus of the SARS-CoV-2 pandemic poses unprecedented challenges to the worlds health systems. New and virulent emerging spike gene variants, such as the UK 20I/501Y.V1 and South African 20H/501Y.V2, could jeopardize global efforts to produce immunity and reduce mortality. These challenges require effective real-time genomic surveillance solutions that the medical community can quickly adopt. The SARS-CoV-2 spike protein mediates host receptor recognition and entry into the cell and therefore, it is most susceptible to generation of variants with increased transmissibility and pathogenicity. The spike protein is also the primary target of neutralizing antibodies in COVID-19 patients and the most common antigen for induction of effective vaccine immunity. Therefore, tight monitoring of the spike protein gene variants is key to mitigating COVID-19 spread and vaccine escape mutants. Currently, the ARTIC method for SARS-CoV-2 whole genome sequencing is applied worldwide. However, this method commonly requires more than 96 hours (4-5 days) from start to finish and at present high sample sequence demands, sequencing resources are quickly exhausted. In this work, we present HiSpike, a method for high-throughput targeted next generation sequencing (NGS) of the spike gene. This simple three-step method can be completed in less than 30 hours and can sequence 10-fold more samples compared to the conventional ARTIC method and at a fraction of the cost. HiSpike was proven valid, and has identified, at high quality, multiple spike variants from real-time field samples, such as the UK and the South African variants. This method will certainly be effective in discovering future spike mutations. Therefore, running HiSpike for full sequencing of the spike gene of all positive SARS-CoV-2 samples could be considered for near real-time detection of known and emerging spike mutations as they evolve. HiSpike provides affordable sequencing options to help laboratories conserve resources, hence it provides a tool for widespread monitoring, that can support critical knowledge-based decisions.

City-level SARS-CoV-2 sewage surveillance

The COVID-19 pandemic created a global crisis impacting not only healthcare systems, but also world economies and society. Recent data have indicated that fecal shedding of SARS-CoV-2 is common, and that viral RNA can be detected in wastewater. This suggests that wastewater monitoring is a potentially efficient tool for both epidemiological surveillance, and early warning for SARS-CoV-2 circulation at the population level. In this study we sampled an urban wastewater infrastructure in the city of Ashkelon, Israel, during the end of the first COVID-19 wave in May 2020 when the number of infections seemed to be waning. We were able to show varying presence of SARS-CoV-2 RNA in wastewater from several locations in the city during two sampling periods. This was expressed as a new index, Normalized Viral Load (NVL), which can be used in different area scales to define levels of virus activity such as red (high) or green (no), and to follow morbidity in the population at tested area. Our index showed the rise in viral load between the two sampling periods (one week apart) and indicated an increase in morbidity that was evident a month later in the population. Thus, this methodology may provide an early indication for SARS-CoV-2 infection outbreak in a population before an outbreak is clinically apparent. HIGHLIGHTSO_LIDetecting the presence of SARS-CoV-2 virus RNA in urban wastewater C_LIO_LIThe city sewer system may provide an early indication for SARS-CoV-2 infection and may be used as early warning for SARS-CoV-2 outbreaks C_LIO_LINVL index defines various infected urban zones from red (high) to green (low) C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/20215244v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@360a84org.highwire.dtl.DTLVardef@1ec8004org.highwire.dtl.DTLVardef@1c8ae93org.highwire.dtl.DTLVardef@3d670c_HPS_FORMAT_FIGEXP M_FIG C_FIG

Improved Sensitivity, Safety, and Rapidity of COVID-19 Tests by Replacing Viral Storage Solution with Lysis Buffer

Conducting numerous, rapid, and reliable PCR tests for SARS-CoV-2 is essential for our ability to monitor and control the current COVID-19 pandemic. Here, we tested the sensitivity and efficiency of SARS-CoV-2 detection in clinical samples collected directly into a mix of lysis buffer and RNA preservative, thus inactivating the virus immediately after sampling. We tested 79 COVID-19 patients and 20 healthy controls. We collected two samples (nasopharyngeal swabs) from each participant: one swab was inserted into a test tube with Viral Transport Medium (VTM), following the standard guideline used as the recommended method for sample collection; the other swab was inserted into a lysis buffer supplemented with nucleic acid stabilization mix (coined NSLB). We found that RT-qPCR tests of patients were significantly more sensitive with NSLB sampling, reaching detection threshold 2.1{+/-}0.6 (Mean{+/-}SE) PCR cycles earlier then VTM samples from the same patient. We show that this improvement is most likely since NSLB samples are not diluted in lysis buffer before RNA extraction. Re-extracting RNA from NSLB samples after 72 hours at room temperature did not affect the sensitivity of detection, demonstrating that NSLB allows for long periods of sample preservation without special cooling equipment. We also show that swirling the swab in NSLB and discarding it did not reduce sensitivity compared to retaining the swab in the tube, thus allowing improved automation of COVID-19 tests. Overall, we show that using NSLB instead of VTM can improve the sensitivity, safety, and rapidity of COVID-19 tests at a time most needed.

Transcriptomic profiling of human corona virus (HCoV)-229E -infected human cells and genomic mutational analysis of HCoV-229E and SARS-CoV-2

Human coronaviruses (HCoVs) cause mild to severe respiratory infection. Most of the common cold illnesses are caused by one of four HCoVs, namely HCoV-229E, HCoV-NL63, HCoV-HKU1 and HCoV-OC43. Several studies have applied global transcriptomic methods to understand host responses to HCoV infection, with most studies focusing on the pandemic severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome CoV (MERS-CoV) and the newly emerging SARS-CoV-2. In this study, Next Generation Sequencing was used to gain new insights into cellular transcriptomic changes elicited by alphacoronavirus HCoV-229E. HCoV-229E-infected MRC5 cells showed marked downregulation of superpathway of cholesterol biosynthesis and eIF2 signaling pathways. Moreover, upregulation of cyclins, cell cycle control of chromosomal replication, and the role of BRCA1 in DNA damage response, alongside downregulation of the cell cycle G1/S checkpoint, suggest that HCoV-229E favors S phase for viral infection. Intriguingly, a significant portion of key factors of cell innate immunity, interferon-stimulated genes (ISGs) and other transcripts of early antiviral response genes were downregulated early in HCoV-229E infection. On the other hand, early upregulation of the antiviral response factor Apolipoprotein B mRNA editing enzyme catalytic subunit 3B (APOBEC3B) was observed. APOBEC3B cytidine deaminase signature (C-to-T) was previously observed in genomic analysis of SARS-CoV-2 but not HCoV-229E. Higher levels of C-to-T mutations were found in countries with high mortality rates caused by SARS-CoV-2. APOBEC activity could be a marker for new emerging CoVs. This study will enhance our understanding of commonly circulating HCoVs and hopefully provide critical information about still-emerging coronaviruses. Author summaryHuman coronaviruses (HCoVs) generate respiratory tract infections. HCoV-229E is one of four known HCoV strains that circulate annually in the population for several decades. Beside these, three pandemic CoV emerged since year 2002, the Severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV) and SARS-CoV-2. These three strains attracted most attention for extensive research and less consideration has been given to the commonly infecting HCoVs. In this study we use Next generation sequencing analysis to understand global transcriptomic changes in human host cells following HCoV-229E infection. We found several cellular pathways that change during infection that involve cholesterol biosynthesis, cell cycle control, DNA replication, DNA repair, innate immune response and an interesting RNA editing enzyme which could be involve in CoVs pathogenesis.

Regressing SARS-CoV-2 sewage measurements onto COVID-19 burden in the population: a proof-of-concept for quantitative environmental surveillance

SARS-CoV-2 is an RNA virus, a member of the coronavirus family of respiratory viruses that includes SARS-CoV-1 and MERS. COVID-19, the clinical syndrome caused by SARSCoV-2, has evolved into a global pandemic with more than 2,900,000 people infected. It has had an acute and dramatic impact on health care systems, economies, and societies of affected countries within these few months. Widespread testing and tracing efforts are employed in many countries in order to contain and mitigate this pandemic. Recent data has indicated that fecal shedding of SARS-CoV-2 is common, and that the virus can be detected in wastewater. This indicates that wastewater monitoring is a potentially efficient tool for epidemiological surveillance of SARS-CoV-2 infection in large populations at relevant scales. Collecting raw sewage data, representing specific districts, and crosslinking this data with the number of infected people from each location, will enable us to derive and provide quantitative surveillance tools. In particular, this will provide important means to (i) estimate the extent of outbreaks and their spatial distributions, based primarily on in-sewer measurements (ii) manage the early-warning system quantitatively and efficiently (and similarly, verify disease elimination). Here we report the development of a virus concentration method using PEG or alum, providing an important a tool for detection of SARS-CoV-2 RNA in sewage and relating it to the local populations and geographic information. This will provide a proof of concept for the use of sewage associated virus data as a reliable epidemiological tool.