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 × 10
copies/day, which represented similar levels compared to April with approx. 2 × 10
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 × 10
to 1 × 10
copies/day, the load to the WWTP Sindlingen from approx. 1 × 10
to 2 × 10
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.
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.
Partial Nitritation/Anammox (PN/A) is a well-established technology for side-stream nitrogen removal from highly concentrated, warm wastewaters. The focus has now shifted to weakly concentrated ...municipal wastewaters with much lower concentrations and temperatures. The major challenge is the temperature, which ranges from moderate 20 °C in summer to cold 10 °C in winter. For this study, the most frequently used configurations for side-stream applications were exposed to a slow temperature reduction from 20 °C to 10 °C to simulate a realistic temperature gradient. To evaluate the behavior of the different biomasses based on their properties, four lab reactors were operated in two different configurations. Synthetic wastewater was used to avoid side effects of heterotrophic growth. Differences in the response of the different reactor systems to this temperature gradient clearly indicated, that the geometry of the biomass has a major impact on the overall PN/A performance at low temperatures: While anammox activity in suspended biomass suffered already at 15 °C, it persevered in granular biomass as well as in biofilms on carriers for temperatures down to <13 °C. Further, anammox activity in thicker biofilms was less affected than in thinner biofilms and even adaption to low temperatures was observed.
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•Operation of 4 different PN/A reactors over a temperature gradient of 20° to 10 °C.•Thicker biofilms show better performance and less nitrite accumulation.•Biofilms (MBBR) retain more anammox biomass.•Microbial community in MBBR more stable than in SBR.
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•Current design and operational strategies for mainstream PN/A are summarized.•Combined ON/OFF and IN/OUT strategies are necessary for successful operation.•A mechanistic framework ...linking engineering, microbiome, and modeling is proposed.•Knowledge readiness levels for key process indicators are defined.•Success relies on integrated research within the framework to boost predictability.
Twenty years ago, mainstream partial nitritation/anammox (PN/A) was conceptually proposed as pivotal for a more sustainable treatment of municipal wastewater. Its economic potential spurred research, yet practice awaits a comprehensive recipe for microbial resource management. Implementing mainstream PN/A requires transferable and operable ways to steer microbial competition as to meet discharge requirements on a year-round basis at satisfactory conversion rates. In essence, the competition for nitrogen, organic carbon and oxygen is grouped into ‘ON/OFF’ (suppression/promotion) and ‘IN/OUT’ (wash-out/retention and seeding) strategies, selecting for desirable conversions and microbes. Some insights need mechanistic understanding, while empirical observations suffice elsewhere. The provided methodological R&D framework integrates insights in engineering, microbiome and modeling. Such synergism should catalyze the implementation of energy-positive sewage treatment.
Municipal wastewater collected in areas with moderate climate is subjected to a gradual temperature decrease from around 20 °C in summer to about 10 °C in winter. A lab-scale moving bed biofilm ...reactor (MBBR) with carrier material (K3 from AnoxKaldnes) was used to test the tolerance of the overall partial nitritation/anammox process to this temperature gradient. A synthetic influent, containing only ammonium and no organic carbon was used to minimize denitrification effects. After stable reactor operation at 20 °C, the temperature was slowly reduced by 2 °C per month and afterward held constant at 10 °C. Along the temperature decrease, the ammonium conversion dropped from an average of 40 gN m–3 d–1 (0.2 gN kgTSS h–1) at 20 °C to about 15 gN m–3 d–1 (0.07 gN kgTSS h–1) at 10 °C, while the effluent concentration was kept <8 mgNH4‑N l–1 during the whole operation. This also resulted in doubling of the hydraulic retention time over the temperature ramp. The MBBR with its biofilm on 10 mm thick carriers proved to sufficiently sustain enough biomass to allow anammox activity even at 10 °C. Even though there was a minor nitrite-build up when the temperature dropped below 12.5 °C, reactor performance recovered as the temperature decrease continued. Microbial community analysis by 16S rRNA amplicon analysis revealed a relatively stable community composition over the entire experimental period.
The Publisher regrets that this article is an accidental duplication of a published article, http://dx.doi.org/10.1016/j.wroa.2020.100066. The duplicate article has therefore been withdrawn. The ...full Elsevier Policy on Article Withdrawal can be found at https://www.elsevier.com/about/our-business/policies/article-withdrawal.The Publisher regrets that this article is an accidental duplication of a published article, http://dx.doi.org/10.1016/j.wroa.2020.100066. The duplicate article has therefore been withdrawn. The full Elsevier Policy on Article Withdrawal can be found at https://www.elsevier.com/about/our-business/policies/article-withdrawal.
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•TWW irrigation intensity and sul1 &intI1 abundance correlate in a real-scale field.•ARGs &intI1 increase in subsoil pore-water during TWW irrigation in mesocosms.•No increase of ARGs ...&intI1 in freshwater irrigated mesocosms.•TWW irrigation does not affect the bacterial load of subsoil pore-water.
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.
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•PICRUSt as tool to predict functional potential from 16S rRNA sequencing data.•Accuracy of PICRUSt tested for microbial communities employed in nitrogen removal.•Two main points were ...discovered from comparing PICRUSt with qPCR.•Higher abundance of target genes results in better correlations.•The more genome information available, the stronger the correlation.
The diversity and dynamics of microorganisms in engineered ecosystems have a high impact on performance and operational stability. Nitrogen removal from wastewater is one example of such complex and dynamic ecosystems. Following the microbial community composition and its functional potential is highly valuable information for optimizing performance. Molecular methods and data analysis tools have become more and more popular in recent years. PICRUSt, a bioinformatics tool to predict the functional potential of a sample from 16S rRNA amplicon sequencing, was tested in the context of autotrophic nitrogen removal for its accuracy. Two experimental studies were extended by qPCR to demonstrate how qPCR can be used to deliver information about the accuracy of PICRUSt predictions. Two main points were discovered: (1) the correlation between qPCR data and PICRUSt predictions depends on the relative abundance of the target gene. With higher abundance, better correlations are achievable; (2) the more genome information available, the stronger the correlation.
This paper highlights the current challenges with quantitative estimation of ammonia‐oxidizing bacteria (AOB) employing quantitative real‐time PCR (qPCR) in diverse natural‐ and engineered‐ ...environments. In this work we concentrates on: (i) identifying the coverage and specificity of the available primers and TaqMan probes for amoA gene based on current databases; (ii) the development of new primers and TaqMan probes for differential quantification of N. europaea and N. eutropha.
Summary
The choice of primer and TaqMan probes to quantify ammonia‐oxidizing bacteria (AOB) in environmental samples is of crucial importance. The re‐evaluation of primer pairs based on current genomic sequences used for quantification of the amoA gene revealed (1) significant misrepresentations of the AOB population in environmental samples, (2) and a lack of perfect match primer pairs for Nitrosomonas europaea and Nitrosomonas eutropha. We designed two new amoA cluster 7‐specific primer pairs and TaqMan probes to quantify N. europaea (nerF/nerR/nerTaq) and N. eutropha (netF/netR/netTaq). Specificity and quantification biases of the newly designed primer sets were compared with the most popular primer pair (amoA1f/amoA2r) using DNA from various AOB cultures as individual templates as well as DNA mixtures and environmental samples. Based on the qPCR results, we found that the newly designed primer pairs and the most popular one performed similarly for individual templates but differed for the DNA mixtures and environmental samples. Using the popular primer pair introduced a high underestimation of AOB in environmental samples, especially for N. eutropha. Thus, there is a strong need for more specific primers and probes to understand the occurrence and competition between N. europaea and N. eutropha in different environments.
Abstract
Waste stabilization ponds (WSP) exist worldwide to treat wastewater, especially in warm climates. They are characterized by simple operation and maintenance and over 50 years many WSP were ...built in urban communities in Namibia. This study characterized and evaluated nine of these WSP systems in terms of their influent and effluent water quality and compared them with the requirements for water reuse in agriculture. In their current state none of them adhered with the Namibian or the new European reuse standards, especially due to tCOD concentrations above 100 mg/L caused by high algal fractions in the pCOD. The algae related chlorophyll-a concentrations correlated linearly with the pCOD and this correlation can therefore be used to fractionate the tCOD for further judgement. Additionally, microbial community analyses determined the composition of pathogens in the WSP influent and effluent, this helped to assess potential risks and distinguish between potentially toxic and non-toxic cyanobacteria. The EU requirement of less than 1,000 E. coli per 100 mL for fodder crop irrigation was only achieved with one WSP system which was enhanced with additional pre- and post-treatment. This research delivers a first overview of the current situation and can be used as basis to establish possible enhancement measures for existing WSP as well as to investigate possible effluent application in agricultural irrigation.
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