Some pathogens are excreted in sufficient quantities by infected individuals through feces, urine, saliva, and other bodily fluids, and can therefore be detected in wastewater. This allows for a test strategy-independent monitoring of the circulation of pathogens, as it examines the excretions of individuals who may not undergo pathogen diagnostics due to mild symptoms, for example. Wastewater analysis provides the opportunity to determine the trend of pathogen load in wastewater and circulating variants specific to a particular geographic region and its population. The results of wastewater surveillance should be evaluated in conjunction with other data.

Date: 13.07.2023

In principle, it is possible to sequence the nucleic acids of pathogens in wastewater samples. For example, virus variants of SARS-CoV-2 in wastewater can be determined to learn more about their circulation in the population.

Date: 13.07.2023

Wastewater-based surveillance should be understood as a supplementary tool for assessing the epidemiological situation. Especially when differences to other monitoring parameters exist, surveillance systems should be compared to identify the causes (e.g., utilization of testing services, reporting delays, altered virus shedding rate due to different virus variants, or changed immune status). Based on the insights gained about infection dynamics, adjustments to the testing strategy can be made.

Date: 13.07.2023

The early detection of the increasing or decreasing infection dynamics of SARS-CoV-2 compared to other surveillance systems depends, among other factors, on the prevailing virus variant, the testing strategy, and reporting delays. There are different estimates of the actual lead time in the current literature. While some studies have observed a lead time of up to three weeks, others have found no lead time in other studies.

Date: 13.07.2023

Wastewater samples are taken regularly from the inlets of the respective wastewater treatment plants. The samples are processed by concentrating the contained pathogens and isolating their nucleic acids in suitable laboratories. Most pathogens can then be detected using Polymerase-Chain-Reaction (PCR) methods if they are present in sufficiently high concentrations in the samples. For further semi-quantitative assessment, a correction is necessary, based on, for example, existing viral fecal indicators or precipitation levels at the time of sampling.

Date: 13.07.2023

In the ESI-CorA project in cooperation with other projects, 48 wastewater treatment plants were sampled from November 2021-April 2023, resulting in a coverage of approx. 16 %. As part of AMELAG, 148 wastewater treatment plants are currently being sampled, which corresponds to a coverage of approx. 32 %.

Date: 18.01.2024

Wastewater is highly heterogeneous over the course of the day and week. Most of the human wastewater load occurs during the morning on weekdays, while on weekends, it is more evenly distributed throughout the day. To obtain a wastewater sample which is representative for the day, it is necessary to take so-called composite samples. This means, for example, continuously and automatically taking samples of 0.5 liters every hour over the course of 24 hours and then combining and homogenizing them to prepare a 1-liter analysis sample for the laboratory. Wastewater samples are currently taken from raw wastewater, immediately after the primary treatment of wastewater at the treatment plant.

Date: 13.07.2023

SARS-CoV-2 is easily detectable in wastewater samples. Wastewater samples used for SARS-CoV-2 analysis can be stored for approximately 4-7 days when refrigerated at 4°C, following appropriate enrichment and extraction methods. Longer storage durations, storage at temperatures above 4°C, or freezing of raw wastewater can lead to a significant reduction in the analysis results.

Date: 13.07.2023

To definitively detect pathogens in wastewater, the Polymerase Chain Reaction (PCR) method is used. PCR amplifies the genetic fingerprint, i.e., the deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) of a given pathogen. The genes or gene fragments of the agents are enriched, amplified, and subsequently detected. PCR is one of the most important methods in molecular biology and is used on a regular basis today.

Date: 13.07.2023

A change in laboratory methods (e.g. in sample preparation or PCR analysis) can lead to a change in sensitivity. This can be reflected in noticeable jumps in the curve. To indicate this, changes in methods are indicated by a vertical dashed line in the weekly report.

Date: 10.07.2024

In the context of PCR analysis, controls are conducted in accordance with common quality standards to ensure the quality and accuracy of the analysis. Additionally, not just one, but two to three genes are amplified in a PCR. This so-called redundant system compensates for analysis errors in the majority of cases.

Date: 14.11.2023

At each participating wastewater treatment plant, influent samples are taken, and relevant sampling parameters (e.g., weather data, flow rate, pH, temperature) are transmitted. The samples are then sent to the laboratories and analyzed for various biomarkers. The data is checked for plausibility and quality at the Federal Environment Agency (UBA) and subsequently normalized. Normalization helps to eliminate effects of factors like heavy rainfall.

Date: 13.07.2023

The normalized SARS-CoV-2 gene concentrations are initially transformed using a base-10 logarithm. The resulting values or aggregations of these values (e.g., weekly averages or averages across different locations) are smoothed using a locally weighted regression (LOESS method). Confidence intervals for the smoothed values are calculated pointwise based on the corresponding quantile of the t-distribution. For the trend calculation, the smoothed values are transformed back to the original scale, and the percentage difference to the previous week is calculated. The LOESS method uses a locally weighted regression function, so a certain proportion of all observed viral loads is included in predicting a viral load on the LOESS curve, with the influence decreasing as the distance from the time of the predicted viral load increases. The curve on which the predicted virus loads lie is estimated separately for each analysis (on a site-specific or aggregated level). The proportion of included virus loads is determined for each location using the generalized cross-validation method, optimizing the predictive quality of the curve. The idea of the cross-validation method is to remove an observed virus load from the data, predict it based on a proportion of all other observed virus loads around the measured value using the LOESS method, and calculate the resulting deviation between observed and predicted viral load. This procedure is performed for all measurement points, and the calculated deviations are summed. The optimal proportion of included viral loads according to the cross-validation criterion is the one that minimizes this sum. This results in a smooth curve and a predicted viral load for each time point (including between measurement points). Since all measurement points are considered for the described estimation of the LOESS curve, it is possible for newly added values to retrospectively alter the LOESS curves of past weeks.

Date: 12.10.2023

Yes, wastewater samples are regularly tested for SARS-CoV-2 and other pathogens in other countries. Wastewater-based surveillance is gaining global importance and is being expanded in many countries. In some countries like Austria or the Netherlands, wastewater-based surveillance for SARS-CoV-2 is used on a widespread basis and covers a large part of the population. In some countries like Canada or the USA, decisions on public health measures are also supported by wastewater analysis.

Date: 13.07.2023

The normalization of data is done based on the dry weather inflow of the respective wastewater treatment plant. This compensates for dilution effects from factors like rainfall and thawing.

In the project AMELAG, the ratio of flow rate during the sampling period (Q_{KA, aktuell}) to the dry weather inflow (Q_{KA, median}) is calculated, and then multiplied by the geometric mean of the existing PCR values ("Gene gemittelt"). The determination of the dry weather inflow involves using the median of all previously reported flow rate values for the wastewater treatment plant.

Genes normalized = (Q_{KA, aktuell} / Q_{KA, median}) * Gene gemittelt

Date: 14.11.2023

In principle, trend analyses of infection dynamics are possible. However, determining the exact incidence (new cases) or prevalence (frequency in the population) and potential underreporting is currently not possible for a municipality or city with a travelling or commuting population, as too many parameters are unknown (e.g., shedding load and shedding duration of an infected person; shedding rates are also dependent on virus variants and immune status). Different site conditions and virus mutations will require ongoing adjustments to the analysis methods and modeling.

Date: 13.07.2023

Wastewater-based surveillance is independent of the testing strategy and the population's testing behavior. Different sampling strategies and analysis methods can also answer different questions. While individual testing allows for concrete measures for the individual, this is not possible with wastewater analysis. However, wastewater-based surveillance may describe the trend of SARS-CoV-2 infection dynamics. The results of wastewater-based surveillance should be considered in the context of other surveillance systems to generate added value for situation management.

Date: 13.07.2023

Wastewater testing serves as a supplement to existing surveillance systems as it cannot address a number of aspects of public health. The following aspects cannot be addressed: virulence and clinical relevance of the detected agents or their variants, disease severity, healthcare burden, affected population groups (gender or age distribution), risk factors, protective factors, and vaccine effectiveness. Advice on individual infection control measures is also not possible.

Date: 13.07.2023

Various aspects need to be considered, such as relevance for public health, potential measures, or the risk of reintroduction of an agent. For example, wastewater-based surveillance could offer added value for public health for the following pathogens, in addition to SARS-CoV-2: influenza viruses, multidrug-resistant agents, Respiratory Syncytial Viruses (RSV), and generally for newly emerging pathogens. The benefit of testing for influenza and antibiotic-resistant bacteria is being evaluated in the AMELAG project.

Date: 13.07.2023

The wastewater surveillance data can be found at various locations on the internet. A detailed epidemiological report, including an assessment of the situation, can be found in the weekly report of wastewater surveillance (http://www.rki.de/abwassersurveillance). Parts of the data are also included in the ARE Weekly Report (https://influenza.rki.de/Wochenberichte.aspx) and in the Infection Radar at the Federal Ministry of Health (https://infektionsradar.gesund.bund.de/de).

Date: 21.02.2024

Infectious SARS-CoV-2 has not been detected in raw wastewater so far. Infectious SARS-CoV-2 have been detected in the stool of infected individuals in a few cases, which could enter wastewater treatment plants through wastewater. In raw wastewater, before treatment in the treatment plants, only the genetic material, i.e., the RNA of the virus, has been detected thus far. During the SARS epidemic of 2002/2003, only a few cases of infectious viruses of the closely related SARS-CoV-1 were isolated from hospital wastewater.

Date: 13.07.2023