Wastewater-based epidemiology (WBE) is a great approach that enables us to comprehensively monitor the community to determine the scale and dynamics of infections in a city, particularly in metropolitan cities with a high population density. Therefore, we monitored the time course of the SARS-CoV-2 RNA concentration in raw sewage in the Frankfurt metropolitan area, the European financial center. To determine the SARS-CoV-2 RNA concentration in sewage, we continuously collected 24 h composite samples twice a week from two wastewater treatment plant (WWTP) influents (Niederrad and Sindlingen) serving the Frankfurt metropolitan area and performed RT-qPCR analysis targeting three genes (N gene, S gene, and ORF1ab gene). In August, a resurgence in the SARS-CoV-2 RNA load was observed, reaching 3 × 10¹³ copies/day, which represented similar levels compared to April with approx. 2 × 10¹⁴ copies/day. This corresponds to a continuous increase again in COVID-19 cases in Frankfurt since August, with an average of 28.6 incidences, compared to 28.7 incidences in April. Different temporal dynamics were observed between different sampling points, indicating local dynamics in COVID-19 cases within the Frankfurt metropolitan area. The SARS-CoV-2 RNA load to the WWTP Niederrad ranged from approx. 4 × 10¹¹ to 1 × 10¹⁵ copies/day, the load to the WWTP Sindlingen from approx. 1 × 10¹¹ to 2 × 10¹⁴ copies/day, which resulted in a preceding increase in these loading in July ahead of the weekly averaged incidences. The study shows that WBE has the potential as an early warning system for SARS-CoV-2 infections and a monitoring system to identify global hotspots of COVID-19.

Wastewater-based severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) surveillance of Frankfurt Airport by genome sequencing was used to detect SARS-CoV-2 variants entering the region. In November 2021, we found all characteristic mutations of Omicron in wastewater originating from Frankfurt Airport before the first confirmed clinical report from an arriving passenger on 26 November 2021.

In the present study, we investigated the impact of treated wastewater (TWW) irrigation on the prevalence of antibiotic resistance genes (ARGs) in subsoil pore-water, a so-far under-appreciated matrix. We hypothesized that TWW irrigation increases ARG prevalence in subsoil pore-water. This hypothesis was tested using a multiphase approach, which consisted of sampling percolated subsoil pore-water from lysimeter-wells of a real-scale TWW-irrigated field, operated for commercial farming practices, and controlled, laboratory microcosms irrigated with freshwater or TWW. We monitored the abundance of six selected ARGs (sul1, blaOXA-58, tetM, qnrS, blaCTX-M-32 and blaTEM), the intI1 gene associated with mobile genetic elements and an indicator for anthropogenic pollution and bacterial abundance (16S rRNA gene) by qPCR. The bacterial load of subsoil pore water was independent of both, irrigation intensity in the field study and irrigation water type in the microcosms. Among the tested genes in the field study, sul1 and intI1 exhibited constantly higher relative abundances. Their abundance was further positively correlated with increasing irrigation intensity. Controlled microcosm experiments verified the observed field study results: the relative abundance of several genes, including sul1 and intI1, increased significantly when irrigating with TWW compared to freshwater irrigation. Overall, TWW irrigation promoted the spread of ARGs and intI1 in the subsoil pore-water, while the bacterial load was maintained. The combined results from the real-scale agricultural field and the controlled lab microcosms indicate that the dissemination of ARGs in various subsurface environments needs to be taken into account during TWW irrigation scenarios.

PCR-based methods have caused a surge for integration of eco-physiological approaches into research on partial nitritation anammox (PNA). However, a lack of rigorous standards for molecular analyses resulted in widespread data misinterpretation and consequently lack of consensus. Data consistency and accuracy strongly depend on the primer selection and data interpretation. An in-silico evaluation of 16S rRNA gene eubacterial primers used in PNA studies from the last ten years unraveled the difficulty of comparing ecological data from different studies due to a variation in the coverage of these primers. Our 16S amplicon sequencing approach, which includes parallel sequencing of six 16S rRNA hypervariable regions, showed that there is no perfect hypervariable region for PNA microbial communities. Using qPCR analysis, we emphasize the significance of primer choice for quantification and caution with data interpretation. We also provide a framework for PCR based analyses that will improve and assist to objectively interpret and compare such results.

Implementation of partial nitritation anammox ( PNA ) in the mainstream (municipal wastewater treatment) is still under investigation. Microbial community structure and reactor type can influence the performance of PNA reactor; yet, little is known about the role of the community composition of the inoculum and the reactor configuration under mainstream conditions. Therefore, this study investigated the community structure of inocula of different origin and their consecutive community dynamics in four different lab‐scale PNA reactors with 16S r RNA gene amplicon sequencing. These reactors were operated for almost 1 year and subjected to realistic seasonal temperature fluctuations as in moderate climate regions, that is, from 20°C in summer to 10°C in winter. The sequencing analysis revealed that the bacterial community in the reactors comprised: (1) a nitrifying community (consisting of anaerobic ammonium‐oxidizing bacteria (An AOB ), ammonia‐oxidizing bacteria ( AOB ), and nitrite‐oxidizing bacteria ( NOB )); (2) different heterotrophic denitrifying bacteria and other putative heterotrophic bacteria ( HB ). The nitrifying community was the same in all four reactors at the genus level, although the biomasses were of different origin. Community dynamics revealed a stable community in the moving bed biofilm reactors ( MBBR ) in contrast to the sequencing batch reactors ( SBR ) at the genus level. Moreover, the reactor design seemed to influence the community dynamics, and reactor operation significantly influenced the overall community composition. The MBBR seems to be the reactor type of choice for mainstream wastewater treatment.