Influent Samples Research Articles

Levels, consumption, and variations of eight artificial sweeteners in the wastewater treatment plants of Dalian city, China

Artificial sweeteners (ASs) are emerging contaminants in the environment, primarily derived from wastewater treatment plant (WWTP) effluents. In this study, the influents and effluents of three WWTPs in the Dalian urban area, China, were analyzed for the distribution of 8 typical ASs to investigate their seasonal fluctuations in the WWTPs. The results showed that acesulfame (ACE), sucralose (SUC), cyclamate (CYC), and saccharin (SAC) were both detected in the influent and effluent water samples of WWTPs, with concentrations ranging from not detected (ND) to 14.02 μg·L−1. In addition, SUC was the most abundant ASs type, accounting for 40 %–49 % and 78 %–96 % of the total ASs in the influent and effluent water, respectively. The WWTPs revealed high removal efficiencies of CYC, SAC, and ACE, while the SUC removal efficiency was poor (26 % ± 36 %). The ACE and SUC concentrations were higher in spring and summer, and all ASs showed lower levels in winter, which may be caused by the high consumption of ice-cream in warmer months. The per capita ASs loads in the WWTPs were determined in this study based on the wastewater analysis results. The calculated per capita daily mas loads for individual ASs ranged from 0.45 g·d−1·1000p−1 (ACE) to 2.04 g·d−1·1000p−1 (SUC). In addition, the relationship between per capita ASs consumption and socioeconomic status showed no significant correlation.

High resolution mass spectrometry (HRMS) determination of drugs in wastewater and wastewater based epidemiology in Cadiz Bay (Spain)

Multi-residue methods for the determination of the myriad of compounds of emerging concern (CECs) entering in the environment are key elements for further assessment on their distribution and fate. Here, we have developed an analytical protocol for the simultaneous analysis of 195 prescription, over-the-counter, and illicit drugs by using a combination of solid phase extraction (SPE) and determination by liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS). The method was applied to the analysis of influent sewage samples from 3 wastewater treatment plants (WWTPs) from Cadiz Bay (SW Spain), enabling the quantification of more than 100 pharmaceuticals, 19 of them at average concentrations higher than 1 μg L−1, including caffeine (92 μg L−1), paracetamol (72 μg L−1), and ibuprofen (56 μg L−1), as well as several illicit drugs (e.g., cocaine). Wastewater based epidemiology (WBE) was applied for 27 of the detected compounds to establish their consumption in the sampling area, which has been never attempted before. Caffeine, naproxen, and salicylic acid stood out because of their high consumption (638, 51, and 20 g d−1·1000pop−1, respectively). Regarding illicit drugs, cocaine showed the highest frequency of detection and we estimated an average consumption of 3683 mg d−1·1000pop−1 in Cadiz Bay. The combination of new HRMS methods, capable of discriminating thousands of chemicals, and WBE will allow for a more comprehensive characterization of chemical substances and their consumption in urban environments in the near future.

Removal of Manganese and Iron from Groundwater by using Chlorination and Rapid Sand Filtration: A Case Study

The average groundwater extraction from Murunkan well field, Mannar, is around 3.4 MCM/year, and it was triggered by extensive development projects and resettlements that took place in the last decade. Manganese and Iron variations in Murunkan groundwater reported during some periods created various operational and aesthetic issues in the potable water supply. This study is focused on treating iron and manganese in groundwater using chlorination and Rapid Sand Filtration (RSF) techniques. Pebble layers, height of 205 mm with the particle sizes ranging from 2-25 mm, were filled at the bottom of RSF model filter. Filter media, height of 700 mm with 0.45 mm effective size (D10) and a uniformity coefficient (Cu) of 1.33, was placed above the pebble layers. The filter was operated continuously for 15 days, and influent and effluent water samples were collected from 15 trials. Filter backwashing was performed after every two trials to maintain a consistent filtration rate. Raw water pH values ranged from 6.74 – 7.48. Chlorine solution (1.08-1.26 ppm) was added at the inlet point, and a 2.88 m water column was maintained, allowing adequate mixing and oxidation time. The results revealed that average removals of iron and manganese as 60% and 73%, respectively, while maximum removals of 93% and 91% were observed for iron and manganese, respectively.

First quantitative detection of tomato brown rugose fruit virus in wastewater in Louisiana

We investigated the occurrence of tomato brown rugose fruit virus (ToBRFV) at a conventional wastewater treatment plant in Louisiana over a 13-month period, from March 2017 to March 2018. Influent, secondary effluent, and final effluent wastewater samples were collected monthly, and viruses were concentrated by the adsorption-elution method using an electronegative filter, followed by the detection using quantitative polymerase chain reaction. ToBRFV was detected in 10 (77 %) of 13 influent samples, 9 (69 %) of 13 in secondary effluent, and 6 (50 %) of 12 final effluents. The concentrations of ToBRFV in the influent samples ranged from 3.5 to 6.1 log10 copies/L and it was always higher than those in secondary or final effluents. Wastewater samples showed a high positive ratio of ToBRFV during fall and winter months. The findings highlight that routine monitoring of new viral indicator such as ToBRFV is necessary to understand its environmental distribution and correlation with pathogenic viruses. This is the first study providing quantitative data on the occurrence of ToBRFV in wastewater.

Phenotypic and genotypic characteristics of beta-lactamase dominant with CARBA, AmpC, and ESBL-producing bacteria in municipal wastewater influent in Helsinki, Finland

Analysing samples of municipal wastewater influent (before treatment) can help to map the status of antibiotic-resistant bacteria (ARB) at the population level in sewershed communities and may also help in predicting the public health risks of ARB in surface water because of the outfall of wastewater. In this study, we investigated the bacterial isolates carrying beta-lactamase genes in wastewater and compared their genotypic and phenotypic characteristics. A total of 399 bacterial isolates grown on CHROMagarESBL (n=207) and CHROMagarKPC (n=192) from composite wastewater influent samples (n=7) from the Viikinmäki wastewater treatment plant (Helsinki) were subcultured, nucleic acid was extracted, and the prevalence of different beta-lactamase genes was screened with multiplex polymerase chain reaction (PCR). All PCR-positive isolates were identified with MALDI-TOF. A total of 32.6% of isolates (130 of 399) were PCR positive for at least one resistance gene, and 13% of these positive isolates out of 130 had at least three resistance genes. Among the 22 detected genes, blaGES group was the most prevalent, at 25.8% (n=198; many isolates carried multiple genes), followed by blaMOX (13.1%) and blaTEM (10.1%) as most frequently detected. Furthermore, out of 18 different bacterial species/genera detected as carrying beta-lactamase genes, A. hydrophila/caviae (28.5%), Enterobacter spp. (16.9%), and E. coli (14.6%) were the most prevalent. Enterobacter spp., Aeromonas spp., and K. cryocescens potentially carried AmpC genes, and E. coli carried ESBL genes. We recorded a huge variety of beta-lactamases (blaAmpC, blaESBL, and blaCARBA) genes in many potential pathogens that probably originated from both enteric and environmental sources.

High-resolution characterization of short-term temporal variability in the taxonomic and resistome composition of wastewater influent.

Wastewater-based epidemiology (WBE) for population-level surveillance of antimicrobial resistance (AMR) is gaining significant traction, but the impact of wastewater sampling methods on results is unclear. In this study, we characterized taxonomic and resistome differences between single-timepoint-grab and 24 h composites of wastewater influent from a large UK-based wastewater treatment work [WWTW (population equivalent: 223 435)]. We autosampled hourly influent grab samples (n=72) over three consecutive weekdays, and prepared additional 24 h composites (n=3) from respective grabs. For taxonomic profiling, metagenomic DNA was extracted from all samples and 16S rRNA gene sequencing was performed. One composite and six grabs from day 1 underwent metagenomic sequencing for metagenomic dissimilarity estimation and resistome profiling. Taxonomic abundances of phyla varied significantly across hourly grab samples but followed a repeating diurnal pattern for all 3 days. Hierarchical clustering grouped grab samples into four time periods dissimilar in both 16S rRNA gene-based profiles and metagenomic distances. 24H-composites resembled mean daily phyla abundances and showed low variability of taxonomic profiles. Of the 122 AMR gene families (AGFs) identified across all day 1 samples, single grab samples identified a median of six (IQR: 5-8) AGFs not seen in the composite. However, 36/36 of these hits were at lateral coverage <0.5 (median: 0.19; interquartile range: 0.16-0.22) and potential false positives. Conversely, the 24H-composite identified three AGFs not seen in any grab with higher lateral coverage (0.82; 0.55-0.84). Additionally, several clinically significant human AGFs (bla VIM, bla IMP, bla KPC) were intermittently or completely missed by grab sampling but captured by the 24 h composite. Wastewater influent undergoes significant taxonomic and resistome changes on short timescales potentially affecting interpretation of results based on sampling strategy. Grab samples are more convenient and potentially capture low-prevalence/transient targets but are less comprehensive and temporally variable. Therefore, we recommend 24H-composite sampling where feasible. Further validation and optimization of WBE methods is vital for its development into a robust AMR surveillance approach.

Vitamin D receptor antagonist activity in wastewater effluents—potential for endocrine disruption

BackgroundEndocrine disrupting chemicals have been identified for a number of human endocrine systems, but there are no reports on vitamin D-antagonistic activities in environmental samples.ObjectivesWe have investigated if there are compounds present in the environment that can act as Vitamin D receptor (VDR) antagonists.MethodsWater samples were collected of the influent and effluent water from five Swedish wastewater treatment facilities and concentrated with solid phase extraction. VDR antagonistic properties of the samples were tested with a cell-based in vitro assay responsive to vitamin D signaling. Cytotoxicity was monitored by three different assays.ResultsWe observed a dose-dependent decrease in the VDR signaling in most studied samples, although the effect was overlapping with cytotoxicity for the influent samples. For effluent samples, we observed clear VDR antagonistic effects also in non-cytotoxic concentrations. The observed effects could not be explained by presence of natural organic matter or cadmium in the water.DiscussionThe vitamin D endocrine system regulates a broad range of physiological processes, and disruption of this system could be associated with serious health consequences. In this study, we report environmental presence of compounds with VDR antagonistic properties, compounds which constitute a new group of potential endocrine disruptors. The VDR antagonism was observed in wastewater treatment facility effluent waters, which are discharged into water systems used as raw water for drinking water production. The findings reported in this study may indicate a potential hazard to human health and aquatic life. Future research is needed to investigate the presence of VDR antagonists in the environment, identification of the causative compounds, and studies of exposure of humans and aquatic organisms to these compounds.

The fate of SARS-CoV-2 viral RNA in coastal New England wastewater treatment plants

Municipal sewage carries SARS-CoV-2 viruses shed in the human stool by infected individuals to wastewater treatment plants (WWTPs). It is well-established that increasing prevalence of COVID-19 in a community increases the viral load in its WWTPs. Despite the fact that wastewater treatment facilities serve a critical role in protecting downstream human and environmental health through removal or inactivation of the virus, little is known about the fate of the virus along the treatment train. To assess the efficacy of differing WWTP size and treatment processes in viral RNA removal we quantified two SARS-CoV-2 nucleocapsid (N) biomarkers (N1 and N2) in both liquid and solids phases for multiple treatment train locations from seven coastal New England WWTPs. SARS-CoV-2 biomarkers were commonly detected in the influent, primary treated, and sludge samples (returned activated sludge, waste activated sludge, and digested sludge), and not detected after secondary clarification processes or disinfection. Solid fractions had 470 to 3,700-fold higher concentrations of viral biomarkers than liquid fractions, suggesting considerably higher affinity of the virus for the solid phase. Our findings indicate that a variety of wastewater treatment designs are efficient at achieving high removal of SARS CoV-2 from effluent; however, quantifiable viral RNA was commonly detected in wastewater solids at various points in the facility. This study supports the important role municipal wastewater treatment facilities serve in reducing the discharge of SARS-CoV-2 viral fragments to the environment and highlights the need to better understand the fate of this virus in wastewater solids.

Seven-year performance of biosand filters in rural Rwanda

Abstract Waterborne diseases remain a significant public health problem, and biosand filters (BSFs) are a common household water filtration method for improving drinking water quality to prevent diseases. The purpose of this study was to evaluate the effectiveness of BSFs over time in a rural Rwandan community. Sixteen BSFs were installed – eight in 2015 and eight in 2018. Influent and effluent samples were tested with Colilert Presence/Absence kits in 2015, 2017, 2018, 2019, and 2022. About 92.9% of filters were still in use in July 2022, but only 64.3% were functioning well, and 50% of effluent samples tested positive for fecal coliforms. A Kruskal–Wallis H test showed no statistically significant difference in effluent percent positive for fecal coliforms by filter age [χ2 (1) = 4.00, p = 0.41]. Recorded observations about each filter, such as rusted diffuser plates, may be useful for understanding the need for maintenance over time.

Exploring resistomes and microbiomes in pilot-scale microalgae-bacteria wastewater treatment systems for use in low-resource settings

Antibiotic resistance genes (ARGs) released into the environment are an emerging human and environmental health concern, including ARGs spread in wastewater treatment effluents. In low-to-middle income countries (LMICs), an alternate wastewater treatment option instead of conventional systems are low-energy, high-rate algal ponds (HRAP) that use microalgae-bacteria aggregates (MABA) for waste degradation. Here we studied the robustness of ARG removal in MABA-based pilot-scale outdoor systems for 140 days of continuous operation. The HRAP system successfully removed 73 to 88 % chemical oxygen demand and up to 97.4 % ammonia, with aggregate size increasing over operating time. Fourteen ARG classes were identified in the HRAP influent, MABA, and effluent using metagenomics, with the HRAP process reducing total ARG abundances by up to 5-fold from influent to effluent. Parallel qPCR analyses showed the HRAP system significantly reduced exemplar ARGs (p < 0.05), with 1.2 to 4.9, 2.7 to 6.3, 0 to 1.5, and 1.2 to 4.8 log-removals for sul1, tetQ, blaKPC, and intl1 genes, respectively. Sequencing of influent, effluent and MABAs samples showed associated microbial communities differed significantly, with influent communities by Enterobacteriales (clinically relevant ARGs carrying bacteria), which were less evident in MABA and effluent. In this sense, such bacteria might be excluded from MABA due to their good settling properties and the presence of antimicrobial peptides. Microalgae-bacteria treatment systems steadily reduced ARGs from wastewater during operation time, using sunlight as the energetic driver, making them ideal for use in LMIC wastewater treatment applications.

Quantification of microplastics in wastewater systems of German industrial parks and their wastewater treatment plants

Microplastics (MP) enter the aquatic environment via several pathways. Many research groups have focused on municipal discharge, while research on industrial sources is rare. This study provides one of the first insights into MP occurrence and distribution in the wastewater systems of industrial parks (IPs) and their wastewater treatment plants (IPWWTPs). The effluents from production plants as well as influent, effluent, and internal samples from the IPWWTPs were assessed. Sampling methods for parallel MP mass and number analyses were developed for varying conditions. The total item emissions of MP (≥10 μm) into the environment were analyzed using μ-Raman spectroscopy and ranged from 3 · 102 to 8 · 104 MP m−3, with a median of 6 · 103 MP m−3 per IPWWTP. Masses analyzed using differential scanning calorimetry showed an MP mass discharge into the environment of 0.2 to 11 mg m−3 with a median of 3.7 mg m−3 per IPWWTP. MP item concentrations within an IPWWTP varied by two to three log levels over several days. Fibers were rare in all samples. Polymer types varied depending on the types of industrial sites and the production plants discharging into the IPWWTP. Within an IP, MP could be allocated to its dischargers, which could be useful for future regulatory requirements. Further research is needed to include different types of IPs producing various polymers and additional processing plants to expand this data set.

Estimating the prevalence of depression using wastewater-based epidemiology: A case study in Qinghai Province, West China

Wastewater-based epidemiology (WBE) is considered a cost-effective alternative approach capable of determining the consumption and prevalence of drug use in communities, however, the application of WBE for estimating the prevalence of depression has seldom been reported. In this study, the prevalence of antidepressants was estimated in five cities in Qinghai Province, west China to examine the feasibility of using WBE to estimate the depression prevalence. Residual concentrations of the drugs varied from different wastewater treatment plants (WWTPs) in five cities. Venlafaxine (0.06–720 ng/L), O-desmethylvenlafaxine (1.31–1659 ng/L), paroxetine (<LOD-69.9 ng/L), sertraline (0.20–3.36 ng/L), and nortriptyline (<LOD-50.8 ng/L) were in the top concentrations quartile in the influent samples of 17 WWTPs. Back-calculation estimated venlafaxine (538 mg/1000 inh/d) as the most consumed antidepressant in Qinghai Province, followed by paroxetine (159 mg/1000 inh/d), sertraline (150 mg/1000 inh/d) and amitriptyline (97.2 mg/1000 inh/d). The prevalence of depression was 17.8 % based on antidepressant usage (2.50 %), which was consistent with the data conducted by traditional survey (16.7 %). Risk assessment showed that fluoxetine, citalopram and venlafaxine in effluents might cause ecological risks to aquatic environment. This study provided a promising way for monitoring antidepressant usage and depression prevalence through WBE, which would improve the understanding of depression disease and provide guidance for public health care.

Three years of wastewater surveillance for new psychoactive substances from 16 countries

The proliferation of new psychoactive substances (NPS) over recent years has made their surveillance complex. The analysis of raw municipal influent wastewater can allow a broader insight into community consumption patterns of NPS. This study examines data from an international wastewater surveillance program that collected and analysed influent wastewater samples from up to 47 sites in 16 countries between 2019 and 2022. Influent wastewater samples were collected over the New Year period and analysed using validated liquid chromatography – mass spectrometry methods. Over the three years, a total of 18 NPS were found in at least one site. Synthetic cathinones were the most found class followed by phenethylamines and designer benzodiazepines. Furthermore, two ketamine analogues, one plant based NPS (mitragynine) and methiopropamine were also quantified across the three years. This work demonstrates that NPS are used across different continents and countries with the use of some more evident in particular regions. For example, mitragynine has highest mass loads in sites in the United States, while eutylone and 3-methylmethcathinone increased considerably in New Zealand and in several European countries, respectively. Moreover, 2F-deschloroketamine, an analogue of ketamine, has emerged more recently and could be quantified in several sites, including one in China, where it is considered as one of the drugs of most concern. Finally, some NPS were detected in specific regions during the initial sampling campaigns and spread to additional sites by the third campaign. Hence, wastewater surveillance can provide an insight into temporal and spatial trends of NPS use.

Sewage analysis as an alternative tool for assessing drug of abuse and new psychoactive substances in Tunisia

Many studies attest to the pollution of wastewaters by organic molecules including drug of abuse (DAs) residues and new psychoactive substances (NPS) at trace levels. The occurrence of these emerging micropollutants in influent wastewaters (IWW) from three Tunisian Wastewater Treatment Plants (WWTPs) was assessed. Influent wastewater composite samples (24 h) were collected over 7 consecutive days in November 2019. The determination and quantification of 11 drug of abuse or their metabolites was performed by the application of an optimized multi-residue method liquid chromatography tandem mass spectrometry (LC–MS/MS). MDMA, THC and the cocaine metabolite benzoyl ecgonine were the most detected substances across the three investigated sewage plants. A new wastewater-based epidemiology (WBE) approach was applied in this study to estimate illicit drug consumption. This innovative approach was used to calculate and to assess collective drug consumption of illicit drug at a community level, based on the concentration of selected illicit substances and their major metabolites in influent wastewater. The average MDMA consumption found in the selected cities ranged between 35,8–1531,1 mg day− 1/1000 inhabitants and increased during the weekends. Cocaine consumption varied from 24.5 to 179.8 mg day− 1/1000 inhabitants.Complementary qualitative investigation of new psychoactive substances was monitored for the first time for an African country, examining the occurrence of 33 NPS in wastewaters samples. Out of 33 totals screened NPS across all sampling sites, 16 were tentatively identified with this approach. The 16 detected NPS covered most of the representative and used molecules of different NPS classes; including synthetic opioids, synthetic cathinones, amphetamines derivatives and synthetic cannabinoids.

Sub-sewershed Monitoring to Elucidate Down-the-Drain Pesticide Sources.

Pesticides have been reported in treated wastewater effluent at concentrations that exceed aquatic toxicity thresholds, indicating that treatment may be insufficient to adequately address potential pesticide impacts on aquatic life. Gaining a better understanding of the relative contribution from specific use patterns, transport pathways, and flow characteristics is an essential first step to informing source control measures. The results of this study are the first of their kind, reporting pesticide concentrations at sub-sewershed sites within a single sewer catchment to provide information on the relative contribution from various urban sources. Seven monitoring events were collected from influent, effluent, and seven sub-sewershed sites to capture seasonal variability. In addition, samples were collected from sites with the potential for relatively large mass fluxes of pesticides (pet grooming operations, pest control operators, and laundromats). Fipronil and imidacloprid were detected in most samples (>70%). Pyrethroids were detected in >50% of all influent and lateral samples. There were significant removals of pyrethroids from the aqueous process stream within the facility to below reporting limits. Imidacloprid and fiproles were the only pesticides that were detected above reporting limits in effluent, highlighting the importance of source identification and control for the more hydrophilic compounds. Single source monitoring revealed large contributions of fipronil, imidacloprid, and permethrin originating from a pet groomer, with elevated levels of cypermethrin at a commercial laundry location. The results provide important information needed to prioritize future monitoring efforts, calibrate down-the-drain models, and identify potential mitigation strategies at the site of pesticide use to prevent introduction to sewersheds.

Dispersive Liquid - Liquid Micro extraction Technique Coupled with High Performance Liquid Chromatography Diode Array Detector for the Determination of Sulfonamides in Water Samples

A dispersive liquid-liquid microextraction combined with high-performance liquid chromatography with diode array detection (DLLME-HPLC-DAD) has been developed for the determination of 15 sulfonamides in water samples. The effects of experimental parameters on the performance of the method such as the type and volume of extraction and disperser solvents, pH of the solution, effect of salt, and centrifugation time were evaluated, and optimum conditions were established. Under these optimum conditions, linearity was found in the range of 2 - 1 000 μg L-1 with regression coefficients better than 0.9990. The limit of detection (LOD) and limit of quantification (LOQ) values were in the range of 0.6 - 7.8 μg L-1 and 1.5 - 8.3 µg L-1, respectively. Intra-day and inter-day precision ranged from 1.5 - 9.7% and from 3.1 - 9.9%, respectively. The accuracy of the method were also evaluated in samples from three different sources of water (river water, groundwater, and tap water) were found to range from 70.5 to 103.9% with %RSD values ranging from 0.5 to 9.8%, with the exception of sulfaguanidine in river water and groundwater where the percentage recovery ranged from 54.3 - 84.9% at the two higher concentration levels (100 and 400 µg L-1).Furthermore, the effect of sample matrix on the extraction of target compounds was evaluated using the relative recovery (%RR) of the samples from three different water sources relative to the ultrahigh purity (UHP) water sample spiked with target analytes, and found in the range of 70.3 - 106.1%.The proposed method was successfully applied to wastewater treatment plant samples and Sulfisoxazole (SSO) was detected in the influent samples in the range of 14.8 - 17.8 µg L-1 with corresponding %RSD, (n = 5) value of 1.6% to 1.9%.The results indicated that the proposed method is effective for the extraction and determination of the sulfonamides in water samples.

Monitoring organic micropollutants in stormwater runoff with the method of fingerprinting

The ecological state of receiving water bodies can be significantly influenced by organic micropollutants that are emitted via stormwater runoff. Reported efforts to quantify the emission of micropollutants mainly focus on sampling at combined sewer overflows and storm sewer outfalls, which can be challenging. An alternative method, called fingerprinting, was developed and tested in this study. The fingerprinting method utilizes wastewater treatment plant (WWTP) influent samples and derives the proportion of stormwater in a sample. This is achieved by comparing the wet weather vs dry weather concentrations of substances-tracers which are present only in wastewater. It is then possible to estimate the concentration of organic micropollutants in stormwater runoff from measurements in the influent of a WWTP based on a mass balance. In this research, the fingerprinting method was applied in influent samples obtained in five WWTPs in the Netherlands. In total, 28 DWF and 22 WWF samples were used. The chosen tracers were ibuprofen, 2-hydroxyibuprofen, naproxen and diclofenac. Subsequently, the concentration in stormwater runoff of 403 organic micropollutants was estimated via the WWF samples. The substances that were present and analyzed included glyphosate and AMPA, 24 out of 254 pesticides, 6 out of 28 organochlorine pesticides, 45 out of 63 pharmaceuticals, 15 out of 15 PAHs, 2 of the 7 PCBs, and 20 of 33 other substances (e.g. bisphenol-A). A comparison with findings from other studies suggested that the fingerprinting method yields trustworthy results. It was also noted that a representative and stable dry weather flow reference concentration is a strict requirement for the successful application of the proposed method.

First reported detection of the mobile colistin resistance genes, mcr-8 and mcr-9, in the Irish environment

The emergence and dissemination of mobile colistin resistance (mcr) genes across the globe poses a significant threat to public health, as colistin remains one of the last line treatment options for multi-drug resistant infections. Environmental samples (157 water and 157 wastewater) were collected in Ireland between 2018 and 2020. Samples collected were assessed for the presence of antimicrobial resistant bacteria using Brilliance ESBL, Brilliance CRE, mSuperCARBA and McConkey agar containing a ciprofloxacin disc. All water and integrated constructed wetland influent and effluent samples were filtered and enriched in buffered peptone water prior to culture, while wastewater samples were cultured directly. Isolates collected were identified via MALDI-TOF, were tested for susceptibility to 16 antimicrobials, including colistin, and subsequently underwent whole genome sequencing. Overall, eight mcr positive Enterobacterales (one mcr-8 and seven mcr-9) were recovered from six samples (freshwater (n = 2), healthcare facility wastewater (n = 2), wastewater treatment plant influent (n = 1) and integrated constructed wetland influent (piggery farm waste) (n = 1)). While the mcr-8 positive K. pneumoniae displayed resistance to colistin, all seven mcr-9 harbouring Enterobacterales remained susceptible. All isolates demonstrated multi-drug resistance and through whole genome sequencing analysis, were found to harbour a wide variety of antimicrobial resistance genes i.e., 30 ± 4.1 (10–61), including the carbapenemases, blaOXA-48 (n = 2) and blaNDM-1 (n = 1), which were harboured by three of the isolates. The mcr genes were located on IncHI2, IncFIIK and IncI1-like plasmids. The findings of this study highlight potential sources and reservoirs of mcr genes in the environment and illustrate the need for further research to gain a better understanding of the role the environment plays in the persistence and dissemination of antimicrobial resistance.

Co-incidence of BA.1 and BA.2 at the start of Singapore's Omicron wave revealed by Community and University Campus wastewater surveillance

Wastewater surveillance (WWS) has been globally recognised to be a useful tool in quantifying SARS-CoV-2 RNA at the community and residential levels without biases associated with case-reporting. The emergence of variants of concern (VOCs) have given rise to an unprecedented number of infections even though populations are increasingly vaccinated. This is because VOCs have been reported to possess higher transmissibility and can evade host immune responses. The B.1.1.529 lineage (Omicron) has severely disrupted global plans to return to normalcy. In this study, we developed an allele-specific (AS) RT-qPCR assay which simultaneously targets the stretch of deletions and mutations in the spike protein from position 24–27 for quantitative detection of Omicron BA.2. Together with previous assays that detect mutations associated with Omicron BA.1 (deletion at position 69 and 70) and all Omicron (mutation at position 493 and 498), we report the validation and time series of these assays from September 2021 to May 2022 using influent samples from two wastewater treatment plants and across four University campus sites in Singapore. Viral RNA concentrations at the treatment plants corroborate with locally reported clinical cases, AS RT-qPCR assays revealed co-incidence of Omicron BA.1 and BA.2 on 12 January 2022, almost two months after initial BA.1 detection in South Africa and Botswana. BA.2 became the dominant variant by the end of January 2022 and completely displaced BA.1 by mid-March 2022. University campus sites were similarly positive for BA.1 and/or BA.2 in the same week as first detection at the treatment plants, where BA.2 became rapidly established as the dominant lineage within three weeks. These results corroborate clinical incidence of the Omicron lineages in Singapore and indicate minimal silent circulation prior to January 2022. The subsequent simultaneous spread of both variant lineages followed strategic relaxation of safe management measures upon meeting nationwide vaccination goals.

Detection of Carbamazepine, Diclofenac, and Ketoprofen via a SPE-HPLC-PDA Method in a Municipal Wastewater Treatment Plant in East Texas, USA

Increased demand for water reuse, recycling and reclamation heightens the need for improvements in the design of wastewater treatment plants that minimize or remove toxic pollutants or their metabolites. In this study, solid-phase extraction (SPE) technique utilizing C-18 cartridge was used for separation of three commonly used pharmaceuticals, namely, carbamazepine (CBZ), diclofenac (DF), and ketoprofen (KT) in wastewater samples. High performance liquid chromatography-photodiode array detector (HPLC-PDA) method was used for detection of CBZ, DF, and KT along the treatment stages (aeration, clarifier, raw activated sludge (RAS), chlorine contact chamber (CCC), and sulfur dioxide chamber) from Nacogdoches wastewater treatment plant (NWWTP), in East Texas, USA. Highest concentrations were found in the RAS followed by influent aeration stage. Pharmaceutical concentrations detected from the influent or RAS samples were 2.115 × 10−3 mg/L, 8.95 × 10−4 mg/L and 8.55 × 10−4 mg/L for CBZ, KT and DF, respectively. Highest detected total concentrations of pharmaceuticals were: carbamazepine (1.285 – 2.117 μg/L), diclofenac (0.600 – 0.865 μg/L), and ketoprofen (0.815 – 0.885 μg/L). The calculated % removal efficiencies along the treatment stages follow the order; CBZ: RAS > SO2 > clarifier ≈ aeration > CCC; KT: RAS > SO2 > aeration > CCC > clarifier; DF: RAS ≈ aeration ≈ CCC > clarifier > SO2. Determined removal efficiencies follow the order carbamazepine (∼ 39.2%) > diclofenac (∼ 30.6%) > ketoprofen (∼8.8%). Removal efficiencies of pharmaceuticals in the NWWTP may be affected by the human body, transition through activated wastewater treatment processes, and their metabolism and/or usage by humans.