ABSTRACT
Background: Reports showed presence of SARS-CoV-2 genetic material in wastewater. Wastewater concentration methods are optimized for detection of non-enveloped viruses so need to be adopted for enveloped viruses and their genetic material. Methods: Conventional (cRT-PCR) and quantitative real time RT-PCR (qRT-PCR) were used as readouts to compare 4 water concentration methods namely, (A) filtration on negatively charged membrane followed by extracting RNA from it, (B) adsorbtion-elution method, (C) flocculation with skimmed milk and (D) polyethylene glycol precipitation, to detect SARS-CoV-2 RNA and 229E human coronavirus (229E-HCoV) as a model for spike-containing enveloped virus from fresh and wastewater. Results: On using cRT-PCR: recovery rate of SARS-CoV-2 RNA was better using method A then B for fresh water and method B then D for wastewater. 229E-HCoV recovery from fresh water was better using method C then A and methods B then D for wastewater. On using qRT-PCR, both methods A and B were better for SARS-CoV-2 RNA recovery from both fresh and wastewater. For the 229E-HCoV methods A was the most efficient for fresh water and method B for wastewater. Conclusion: Method B is recommended for SARS-CoV-2 RNA or whole 229E-HCoV recovery from wastewater.
ABSTRACT
Wastewater-based epidemiology has been described as a valuable tool for monitoring the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a community. However, there is no consensus on the best concentration method to allow reliable detection of SARS-CoV-2 in this matrix, considering different laboratory facilities. This study compares two viral concentration methods, ultracentrifugation (ULT) and skimmed-milk flocculation (SMF), for detecting SARS-CoV-2 in wastewater samples. The analytical sensitivity (limits of detection and quantification [LoD/LoQ]) of both methods was evaluated using a bovine respiratory syncytial virus (BRSV) as a surrogate. Three different approaches were conducted to establish LoD of each method based on the assays on the standard curve (ALoDsc), on the dilution of internal control (ALoDiC), and the processing steps (PLoD). For PLoD, ULT method had the lowest value (1.86 × 103 genome copy/microliter [GC/µL]) when compared to the SMF method (1.26 × 107 GC/µL). The LoQ determination showed a mean value of 1.55 × 105 GC/µL and 3.56 × 108 GC/µL to ULT and SMF, respectively. The detection of SARSCoV-2 in naturally contaminated wastewater revealed 100% (12/12) and 25% (3/12) of detection using ULT and SMF with quantification ranging from 5.2 to 7.2 log10 genome copy/liter (GC/L) and 5.06 to 5.46 log10 GC/L, respectively. The detection success rate of BRSV used as an internal control process was 100% (12/12) for ULT and 67% (8/12) for SMF, with an efficiency recovery rate ranging from 12 to 38% and 0.1 to 5%, respectively. Our data consolidates the importance of assessing the methods used; however, further analysis should be carried out to improve low-cost concentration methodologies, essential for use in low-income and developing countries.
Subject(s)
COVID-19 , Viruses , Animals , Cattle , SARS-CoV-2/genetics , COVID-19/diagnosis , Wastewater , Limit of Detection , RNA, ViralABSTRACT
[Display omitted] • P(DAC-NIPAM) significantly improved the removal of levofloxacin and tetracycline. • P(DAC-NIPAM) had strong interaction with antibiotics for its multiple functional groups. • The hydrophobic groups on P(DAC-NIPAM) tightly bridged micelles of antibiotics and SDS. • Compact flocs were formed for shrinkage of P(DAC-NIPAM) molecule at the LCST. • Flocculation simulation further confirmed application feasibility of thermosensitive flocculants. Antibiotics were detected in worldwide natural water especially in COVID-19 period. The common flocculants rarely removed the dissolved antibiotics from natural water and wastewater. The flocculation improvement of organic polymer flocculants might solve the issue of antibiotic pollution or promote the removal efficiencies of antibiotics in water/wastewater treatment plants. Herein, a thermosensitive flocculant, P(DAC-NIPAM), was prepared via one-step method. It was investigated that the relationship between the various functional groups of P(DAC-NIPAM) and its flocculation performances in the treatment of simulated water containing levofloxacin, tetracycline, colloidal particles and natural organic matters. The removal mechanisms were discussed. The results indicated that the rich cationic, hydrophilic and hydrophobic groups of P(DAC-NIPAM) enhanced the interaction between flocculants and pollutants. The bridging of P(DAC-NIPAM) among micelles, charge neutralization, hydrogen bond between P(DAC-NIPAM) and two antibiotics, the shrinkage of P(DAC-NIPAM) molecule and enhancement of hydrophobicity when water temperature was above low critical solution temperature (LCST), co-flocculation and co-settlement of multiple pollutants all contributed to the efficient removal of levofloxacin and tetracycline from water. Flocculation simulation further confirmed that thermosensitive flocculant combined with heating plates was a potential candidate for antibiotic treatment in actual water treatment plants. [ FROM AUTHOR]
ABSTRACT
Antibiotics were detected in worldwide natural water especially in COVID-19 period. The common flocculants rarely removed the dissolved antibiotics from natural water and wastewater. The flocculation improvement of organic polymer flocculants might solve the issue of antibiotic pollution or promote the removal efficiencies of antibiotics in water/wastewater treatment plants. Herein, a thermosensitive flocculant, P(DAC-NIPAM), was prepared via one-step method. It was investigated that the relationship between the various functional groups of P(DAC-NIPAM) and its flocculation performances in the treatment of simulated water containing levofloxacin, tetracycline, colloidal particles and natural organic matters. The removal mechanisms were discussed. The results indicated that the rich cationic, hydrophilic and hydrophobic groups of P(DAC-NIPAM) enhanced the interaction between flocculants and pollutants. The bridging of P(DAC-NIPAM) among micelles, charge neutralization, hydrogen bond between P(DAC-NIPAM) and two antibiotics, the shrinkage of P(DAC-NIPAM) molecule and enhancement of hydrophobicity when water temperature was above low critical solution temperature (LCST), co-flocculation and co-settlement of multiple pollutants all contributed to the efficient removal of levofloxacin and tetracycline from water. Flocculation simulation further confirmed that thermosensitive flocculant combined with heating plates was a potential candidate for antibiotic treatment in actual water treatment plants.
ABSTRACT
Wastewater-based epidemiology is a widely used tool to detect prevalence of viruses in the population. In the current COVID-19 pandemic, many countries began to analyze the novel coronavirus in sewage samples, and it was found a reliable method to monitor the tendencies of COVID-19 infections in different areas. The viral titer was observed to increase 4-10 days earlier in wastewater than the number of clinical cases. Therefore, the method could be used for early prediction. The method development started in April 2020 at National Public Health Centre (NPHC). Various concentration (flocculation, ultrafiltration) and RNA isolation methods (commercial kits and classic precipitation methods) were compared. The flocculation method showed low recovery rate, while the quality of the ultrafiltration method depended strongly on the type of filter unit. For the national survey, a specially manufactured membrane was chosen, due to its good recovery and reliable availability. The results of nucleic acid isolation were similar with the different methods, a commercial kit (Zymo Research) recommended to feces and soil was chosen due to its higher inhibitor-removal ability. RNA concentration is quantified by quantitative RT-PCR (designed for the nucleocapsid protein 1 gene), similar to the method used for clinical diagnostics. Systematic wastewater sampling started in end of May in Budapest;the survey was extended to all county capitals by the beginning of July. The operators of the wastewater treatment plants (WWTP) from the raw sewage carry the sampling out weekly after the grid filter, and the samples are shipped to the laboratory within 24 hours. Most WWTP does not have composite auto-sampler;therefore, sampling is carried out in the peak-load in most places. The results are available in 36-72 hours and published to NPHC website within a week. A decreasing trend was observable in the data from the end of May to the beginning of June, in parallel with the decline of the first wave of the epidemic. After that, the concentration of SARS-CoV-2 stagnated at a low level until beginning of August. The increasing trend in the wastewater was followed by an increase of the confirmed COVID-19 cases approximately 2 weeks later. Data processing is still ongoing for better modeling of the correlation between clinical data and SARS-CoV-2 concentration in wastewater.
ABSTRACT
In the pandemic of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) many strategies have been performed in order to control viral spread in the population and known the real-time situation about the number of infected persons. In this sense, Wastewater Based Epidemiology (WBE) has been applied as an excellent tool to evaluate the virus circulation in a population. In order to obtain reliable results, three low-cost viral concentration methods were evaluated in this study, polyethylene glycol (PEG) precipitation, skimmed milk flocculation (SM) and Aluminum polychloride flocculation, for Pseudomonas aeruginosa bacteriophage PP7 as a surrogate for non-enveloped viruses and Bovine Coronavirus (BCoV) as a surrogate for enveloped virus, with emphasis for SARS- CoV-2. Our results suggest that PEG precipitation for viral concentration, for both enveloped and non-enveloped virus from wastewater is an appropriate approach since it was more sensitive compared to SM flocculation and Aluminum polychloride flocculation. This methodology can be used for WBE studies in order to follow the epidemiology of the SARS-CoV-2 pandemic, mainly in developing countries where the economic resources are frequently limited.