Antimicrobials and antimicrobial resistance genes in the shadow of COVID-19 pandemic: A wastewater-based epidemiology perspective.

Higher usage of antimicrobial agents in both healthcare facilities and the communities has resulted in an increased spread of resistant bacteria. However, the improved infection prevention and control practices may also contribute to decreasing antimicrobial resistance (AMR). In the present study, wastewater-based epidemiology (WBE) approach was applied to explore the link between COVID-19 and the community usage of antimicrobials, as well as the prevalence of resistance genes. Longitudinal study has been conducted to monitor the levels of 50 antimicrobial agents (AAs), 24 metabolites, 5 antibiotic resistance genes (ARGs) and class 1 integrons (intI 1) in wastewater influents in 4 towns/cities over two years (April 2020 - March 2022) in the South-West of England (a total of 1,180 samples collected with 87,320 individual AA measurements and 8,148 ARG measurements). Results suggested higher loads of AAs and ARGs in 2021-22 than 2020-21, with beta-lactams, quinolones, macrolides and most ARGs showing statistical differences. In particular, the intI 1 gene (a proxy of environmental ARG pollution) showed a significant increase after the ease of the third national lockdown in England. Positive correlations for all quantifiable parent AAs and metabolites were observed, and consumption vs direct disposal of unused AAs has been identified via WBE. This work can help establish baselines for AMR status in communities, providing community-wide surveillance and evidence for informing public health interventions. Overall, studies focused on AMR from the start of the pandemic to the present, especially in the context of environmental settings, are of great importance to further understand the long-term impact of the pandemic on AMR.

Understanding treatment of pain during SARS-CoV-2 pandemic in a two-year intercity longitudinal study using wastewater-based epidemiology.

SARS-CoV-2 pandemic had a significant impact on the society, economy, and health of people around the world with consequences that need to be better understood for future pandemic preparedness. This manuscript provides insights into the usage of pharmaceuticals for pain treatment management throughout SARS-CoV-2 pandemic. Four towns and cities with a total population of > 1 million people covering an area of 2000 km2 in South West England were monitored for twenty-four months. Results showed different patterns in pain pharma usage, with small towns having higher population normalised daily loads (PNDLs) than big cities for majority of pain killers studied. This is likely due to demographics of these cities with smaller cities having older population. Per capita consumption of non-steroidal anti-inflammatory drugs (NSAIDs) increased compared to pre-pandemic usage in line with SARS-CoV-2 infections (ibuprofen and acetaminophen), while body pain drugs (diclofenac and naproxen) decreased in line with restrictions and closure of sports facilities. Changes in population normalised daily intake (PNDI) of pain killers were particularly apparent during the 1st and 3rd national lockdown. Comparison of PNDIs with prescriptions highlighted differences related to medication availability (OTC drugs) and patients' nonadherence (prescribed drugs). In addition, several instances of direct disposal events across the catchments were observed which raises an issue of lack of pharma compliance and general understanding of potential environmental impacts from pharma usage.

Assessment of restriction measures implemented during COVID pandemics on community lifestyle choices via wastewater-based epidemiology.

SARS-CoV-2 pandemic affected lifestyle habits, and the mental health and wellbeing of people around the world. In this manuscript, two towns (Paulton and Radstock) and two cities (Bath and Bristol) in Southwest England (> 1 million people) were monitored for two years using Wastewater-Based Epidemiology to assess impacts of COVID (including management measures such as lockdowns and movement restrictions) on community lifestyle choices: illicit drugs, legal stimulants, abused pharmaceuticals and pain pharma usage. Results were triangulated with key dates captured during the pandemic (national lockdowns, restrictions and social distancing measures, etc.). This highlighted a reduction in cocaine intake (as benzoylecgonine) (community average: -36 %) during the first lockdown and an increase in illicit drugs usage after the 3rd national lockdown (community averages for amphetamine: +8 %, cocaine/benzoylecgonine: +39 %, ketamine:+70 %) when restrictions were removed, and social interaction and recreational activities increased. There was a reduction in the intake of caffeine (as 1,7-dimethylxanthine) (community average:-39 %) after 3rd national lockdown coinciding with pubs reopening while nicotine intake (via cotinine) remained stable indicating lack of impact of COVID on smoking habits. Pain pharma often used in pain management resulting from injuries linked with sport activities (naproxen and diclofenac) showed decrease in usage due to lockdown restrictions in physical exercise and access to gyms/sport facilities.

Antimicrobials and antimicrobial resistance genes in a one-year city metabolism longitudinal study using wastewater-based epidemiology.

This longitudinal study tests correlations between antimicrobial agents (AA) and corresponding antimicrobial resistance genes (ARGs) generated by a community of >100 k people inhabiting one city (Bath) over a 13 month randomised monitoring programme of community wastewater. Several AAs experienced seasonal fluctuations, such as the macrolides erythromycin and clarithromycin that were found in higher loads in winter, whilst other AA levels, including sulfamethoxazole and sulfapyridine, stayed consistent over the study period. Interestingly, and as opposed to AAs, ARGs prevalence was found to be less variable, which indicates that fluctuations in AA usage might either not directly affect ARG levels or this process spans beyond the 13-month monitoring period. However, it is important to note that weekly positive correlations between individual associated AAs and ARGs were observed where seasonal variability in AA use was reported: ermB and macrolides CLR-clarithromycin and dmCLR-N-desmethyl clarithromycin, aSPY- N-acetyl sulfapyridine and sul1, and OFX-ofloxacin and qnrS. Furthermore, ARG loads normalised to 16S rRNA (gene load per microorganism) were positively correlated to the ARG loads normalised to the human population (gene load per capita), which indicates that the abundance of microorganisms is proportional to the size of human population and that the community size, and not AA levels, is a major driver of ARG levels in wastewater. Comparison of hospital and community wastewater showed higher number of AAs and their metabolites, their frequency of occurrence and concentrations in hospital wastewater. Examples include: LZD-linezolid (used only in severe bacterial infections) and AMX-amoxicillin (widely used, also in community but with very low wastewater stability) that were found only in hospital wastewater. CIP-ciprofloxacin, SMX-sulfamethoxazole, TMP-trimethoprim, MTZ-metronidazole and macrolides were found at much higher concentrations in hospital wastewater while TET-tetracycline and OTC-oxytetracycline, as well as antiretrovirals, had an opposite trend. In contrast, comparable concentrations of resistant genes were observed in both community and hospital wastewater. This supports the hypothesis that AMR levels are more of an endemic nature, developing over time in individual communities. Both hospital and community wastewater had AAs that exceeded PNEC values (e.g. CLR-clarithromycin, CIP-ciprofloxacin). In general, though, hospital effluents had a greater number of quantifiable AAs exceeding PNECs (e.g. SMX-sulfamethoxazole, ERY-erythromycin, TMP-trimethoprim). Hospitals are therefore an important consideration in AMR surveillance as could be high risk areas for AMR.

Community infectious disease treatment with antimicrobial agents - A longitudinal one year study of antimicrobials in two cities via wastewater-based epidemiology.

Antimicrobial resistance (AMR) is one of the most significant global health threats. Inappropriate and over-usage of antimicrobial agents (AAs) is a major driver for AMR. Wastewater-based epidemiology (WBE) is a promising tool for monitoring AA usage in communities which is, for the first time, explored in this large scale, longitudinal study. Two contrasting urban catchment areas have been investigated: one city and one small town in the Southwest of the UK over a 13-month period in 2018-2019. Per capita daily intake of 17 AAs and metabolites has been estimated and obtained estimates were triangulated with catchment specific AA prescription data to understand AA usage patterns (both seasons driven prescription and AA prescription compliance). Results have demonstrated positive correlations for all quantifiable parent AAs and metabolites in wastewater, and spatial variability in AA usage was observed even in neighbouring urban areas. WBE and catchment specific prescription data showed similar seasonal trends but with low correlation in intake. The reasons might be variable prescribing patterns, prescription/intake outside the studied catchment, and/or lack of patient compliance. WBE proved useful in differentiating between consumption vs topical usage and/or direct disposal of unused AA. WBE is considered superior to prescription data as it provides information on AAs prescribed outside of the monitoring catchment, e.g. HIV antivirals and TB drugs. However, data triangulation, of both prescription data and wastewater data, provides the most comprehensive approach to understanding AA usage in communities.

The burden of city's pain treatment - A longitudinal one year study of two cities via wastewater-based epidemiology.

This paper explores Wastewater-Based Epidemiology (WBE) as a tool enabling understanding of city's pain treatment in an intercity longitudinal study. An intensive 13-month monitoring programme was undertaken in two adjacent urban areas in South-West England: a small commuter town Keynsham and the city of Bath (>180 samples collected). The study has shown a great potential of using triangulated WBE and National health Service (NHS) prescription data in understanding pain treatment in two contrasting communities with strong apparent seasonal patterns of short pain medications vs chronic pain treatment as well as the type of treatment used (e.g. oral vs topical). Community-wide usage of Non-Steroidal Anti-inflammatory Drugs (NSAIDs) and paracetamol in the intercity study is population size and season driven with the highest usage recorded in winter months. This contrasts with other pain pharmaceuticals, especially those used for chronic pain, where no/limited seasonal usage was recorded. Unmetabolized NSAIDs are, to a large extent, directly disposed of into the sewerage system bypassing metabolism due to their topical application. This is particularly apparent in winter months with naproxen showing the highest seasonal variability. Pharma/met (ratio of pharmaceutical and its metabolite concentration) analysis allows for tracking topical (non-metabolic) application/down-the-drain disposal of pharmaceuticals with frequent instances of direct disposal of NSAIDs into the sewerage system observed. Normalisation of pharma markers to population size shows comparable estimates of pharma usage in the two cities confirming population as the main driver of pharma loads in wastewater. Variable application patterns of pain pharmaceuticals make back-calculation of intake more convoluted. Intake calculated using percentage excretion of parent NSAIDs will likely lead to overestimation, as it is assumed that NSAIDs are subject to extensive metabolism (this is not the case for topical applications). Intake calculated using percentage excretion of metabolites (or parent compound) as consumption markers leads to underestimation of NSAIDs usage due to contributions from topical application not being accounted for. Prescription data indicates cumulative internal and topical usage, but the data ignores large proportion of over-the-counter usage. Therefore, we have proposed a combined approach allowing for estimation of total usage including, and differentiating between, topical application and oral administration.

Quantifying community-wide antimicrobials usage via wastewater-based epidemiology.

Increasing usage of antimicrobials is a significant contributor to the emergence and dissemination of antimicrobial resistance. Wastewater-based epidemiology is a useful tool for evaluating public health, via the monitoring of chemical and biological markers in wastewater influent, such as antibiotics. Sixteen antimicrobials and their metabolites were studied: sulfonamides, trimethoprim, metronidazole, quinolones, nitrofurantoin, cyclines, and antiretrovirals. Correction factors (CFs) for human drug excretion, for various drug forms, were determined via a systematic literature review of pharmacokinetic research. Analyte stability was examined over a 24 h study. The estimation of community-wide drug intake was evaluated using the corresponding catchment prescription data. Overall, antimicrobials excreted in an unchanged form were often observed to over-estimate daily intake. This could be attributed to biotransformation, e.g., via glucuronide cleavage, or direct disposal of unused drugs. Acetyl-sulfonamides, trimethoprim, hydroxy-metronidazole, clarithromycin, ciprofloxacin, ofloxacin, tetracycline, and oxytetracycline generally performed well in the estimation of drug intake, relative to prescription records. The low prevalence of quinolone and trimethoprim metabolites, and the low stability of nitrofurantoin, limited the ability to evaluate these metabolites and their respective CFs.

Human population as a key driver of biochemical burden in an inter-city system: Implications for One Health concept.

This paper tests the hypothesis that human population and city function are key drivers of biochemical burden in an inter-city system, which can be used to inform One Health actions as it enables a holistic understanding of city's metabolism encompassing all of the activities of a city in a single model: from lifestyle choices, through to health status and exposure to harmful chemicals as well as effectiveness of implemented management strategies. Chemical mining of wastewater for biophysico-chemical indicators (BCIs) was undertaken to understand speciation of BCIs in the context of geographical as well as community-wide socioeconomic factors. Spatiotemporal variabilities in chemical and biological target groups in the studied inter-city system were observed. A linear relationship (R2 > 0.99) and a strong positive correlation between most BCIs and population size (r > 0.998, p < 0.001) were observed which provides a strong evidence for the population size as a driver of BCI burden. BCI groups that are strongly correlated with population size and are intrinsic to humans' function include mostly high usage pharmaceuticals that are linked with long term non-communicable conditions (NSAIDs, analgesics, cardiovascular, mental health and antiepileptics) and lifestyle chemicals. These BCIs can be used as population size markers. BCIs groups that are produced as a result of a specific city's function (e.g. industry presence and occupational exposure or agriculture) and as such are not correlated with population size include: pesticides, PCPs and industrial chemicals. These BCIs can be used to assess city's function, such as occupational exposure, environmental or food exposure, and as a proxy of community-wide health. This study confirmed a strong positive correlation between antibiotics (ABs), population size and antibiotic resistance genes (ARGs). This confirms the population size and AB usage as the main driver of AB and ARG levels and provides an opportunity for interventions aimed at the reduction of AB usage to reduce AMR. Holistic evaluation of biophysicochemical fingerprints (BCI burden) of the environment and data triangulation with socioeconomic fingerprints (indices) of tested communities are required to fully embrace One Health concept.

Diagnosing Down-the-Drain Disposal of Unused Pharmaceuticals at a River Catchment Level: Unrecognized Sources of Environmental Contamination That Require Nontechnological Solutions.

Down-the-drain disposal of pharmaceuticals remains an overlooked and unrecognized source of environmental contamination that requires nontechnological "at-source" solutions. Monitoring of 31 pharmaceuticals over 7 days in five wastewater treatment plants (WWTPs) serving five cities in South-West UK revealed down-the-drain codisposal of six pharmaceuticals to three WWTPs (carbamazepine and propranolol in city A, sildenafil in city B, and diltiazem, capecitabine, and sertraline in city D), with a one-off record codisposal of estimated 253 pills = 40 g of carbamazepine and estimated 96 pills = 4 g of propranolol in city A accounting for their 10- and 3-fold respective increases in wastewater daily loads. Direct disposal of pharmaceuticals was found to affect the efficiency of wastewater treatment with much higher pharmaceutical removal (decrease in daily load) during "down-the-drain disposal" days. This is due to lack of conjugated glucuronide metabolites that are cleaved during "consumption-only" days, with the release of a parent pharmaceutical counterbalancing its removal. Higher removal of pharmaceuticals during down-the-drain disposal days reduced pharmaceutical loads reaching receiving environment, albeit with significant levels remaining. The estimated daily loads in receiving water downstream from a discharge point accounted for 13.8 ± 3.4 and 2.1 ± 0.2 g day-1 of carbamazepine and propranolol, respectively, during consumption-only days and peaked at 20.9 g day-1 (carbamazepine) and 4.6 g day-1 (propranolol) during down-the-drain disposal days. Actions are needed to reduce down-the-drain disposal of pharmaceuticals. Our recent work indicated that down-the-drain disposal of pharmaceuticals doubled since the last study in 2005, which may be due to the lack of information and messaging that informs people to dispose of unused medicines at pharmacies. Media campaigns that inform the public of how to safely dispose of medicines are key to improving rates of return and reducing pharmaceutical waste in the environment. The environment is a key motivator for returning unused medicines to a pharmacy and so messaging should highlight environmental risks associated with improper disposal.

A high prevalence of blaOXA-48 in Klebsiella (Raoultella) ornithinolytica and related species in hospital wastewater in South West England.

Klebsiella species occupy a wide range of environmental and animal niches, and occasionally cause opportunistic infections that are resistant to multiple antibiotics. In particular, Klebsiella pneumoniae (Kpne) has gained notoriety as a major nosocomial pathogen, due principally to the rise in non-susceptibility to carbapenems and other beta-lactam antibiotics. Whilst it has been proposed that the urban water cycle facilitates transmission of pathogens between clinical settings and the environment, the level of risk posed by resistant Klebsiella strains in hospital wastewater remains unclear. We used whole genome sequencing (WGS) to compare Klebsiella species in contemporaneous samples of wastewater from an English hospital and influent to the associated wastewater treatment plant (WWTP). As we aimed to characterize representative samples of Klebsiella communities, we did not actively select for antibiotic resistance (other than for ampicillin), nor for specific Klebsiella species. Two species, Kpne and K. (Raoultella) ornithinolytica (Korn), were of equal dominance in the hospital wastewater, and four other Klebsiella species were present in low abundance in this sample. In contrast, despite being the species most closely associated with healthcare settings, Kpne was the dominant species within the WWTP influent. In total, 29 % of all isolates harboured the blaOXA-48 gene on a pOXA-48-like plasmid, and these isolates were almost exclusively recovered from the hospital wastewater. This gene was far more common in Korn (68 % of isolates) than in Kpne (3.4 % of isolates). In general plasmid-borne, but not chromosomal, resistance genes were significantly enriched in the hospital wastewater sample. These data implicate hospital wastewater as an important reservoir for antibiotic-resistant Klebsiella, and point to an unsuspected role of species within the Raoultella group in the maintenance and dissemination of plasmid-borne blaOXA-48. This article contains data hosted by Microreact.

Estimation of community-wide multi-chemical exposure via water-based chemical mining: Key research gaps drawn from a comprehensive multi-biomarker multi-city dataset.

This paper explores the strong potential of chemical mining of wastewater for markers of community-wide intake of wide-ranging harmful chemicals belonging to several usage groups: industrial chemicals, personal care products, pesticides, illicit drugs, lifestyle chemicals and prescription pharmaceuticals as a proxy for multi-chemical community-wide exposure. An estimation of chemical intake in five contrasting town/cities based in the Avon River catchment in the South-West UK was undertaken. High-resolution spatiotemporal pharmaceutical prescription databases were used for system calibration, both in terms of biomarker selection and its correction factor, as well as for the overall system performance evaluation, both spatially and temporality. Only metabolism data accounting for phase two metabolism provided correct estimates of pharma intake. Using parent compounds as XCRs (xenobiotic compounds residue) was found to overestimate exposure due to an inclusion of directly disposed (unused) drugs. Spatiotemporal trends in XC intake were observed as a result of occupational exposure (higher bisphenol A (BPA) intake during weekday), and lifestyle choices (higher cocaine and pyrethroid pesticides intake during weekend). WBE is not intended to estimate individual exposure to chemicals. It can however provide estimates at a community level, and as a result, it has the potential to be developed into an early warning system, a powerful tool for large scale screening studies identifying communities at risk and in need of high resolution individual testing at a localised scale.

Orthodontic treatment considerations for a patient with erythropoietic protoporphyria.

Erythropoietic protoporphyria (EPP) is an inherited blood disorder in which formation of the heme group of hemoglobin is defective. Specifically, a deficiency of the enzyme ferrochelatase leads to the accumulation of protoporphyrin, resulting in often painful photosensitivity of the skin and tissues. The prevalence of EPP is estimated at 1:75,000 to 1:200,000. Photosensitivity is exhibited upon exposure to light with specific wavelengths through the creation of reactive oxygen products (oxidants), activation of the complement system, and mast cell degranulation. The aim of this article is to report the orthodontic treatment of an 11-year-old boy with EPP, a Class III skeletal relationship, and an anterior crossbite. Orthodontic treatment established normal overbite and overjet. Short-term periodontal and dental tissue responses to treatment were noted. Extra care was needed when collecting photographic and radiographic records for this patient and during some treatment procedures to avoid causing a photosensitive reaction of the skin or oral mucosa.

The aponeurotic tension model of craniofacial growth in man.

Craniofacial growth is a scientific crossroad for the fundamental mechanisms of musculoskeletal physiology. Better understanding of growth and development will provide new insights into repair, regeneration and adaptation to applied loads. Traditional craniofacial growth concepts are insufficient to explain the dynamics of airway/vocal tract development, cranial rotation, basicranial flexion and the role of the cranial base in expression of facial proportions. A testable hypothesis is needed to explore the physiological pressure propelling midface growth and the role of neural factors in expression of musculoskeletal adaptation after the cessation of anterior cranial base growth. A novel model for craniofacial growth is proposed for: 1. brain growth and craniofacial adaptation up to the age of 20; 2. explaining growth force vectors; 3. defining the role of muscle plasticity as a conduit for craniofacial growth forces; and 4. describing the effect of cranial rotation in the expression of facial form.Growth of the viscerocranium is believed to be influenced by the superficial musculoaponeurotic systems (SMAS) of the head through residual tension in the occipitofrontalis muscle as a result of cephalad brain growth and cranial rotation. The coordinated effects of the regional SMAS develop a craniofacial musculoaponeurotic system (CFMAS), which is believed to affect maxillary and mandibular development.

Comparison of the Bolton Standards to longitudinal cephalograms superimposed on the occipital condyle (I-point).

OBJECTIVE: To compare traditional superimposition on sella turcica and the anterior cranial base (SACB) to superimposition referenced at the occipital condyle (I-point) for demonstrating craniofacial growth and development. MATERIALS AND METHODS: Tracings for ages 8, 10, 12, 15 and 18 were chosen from the Bolton Standards of Dentofacial Developmental Growth to compare superimposition with the traditional longitudinal reference at SACB (anterior curvature of sella turcica and anterior cranial base) to reference at I-point on the antero-inferior contour of the occipital condyles in norma lateralis. The serial tracings were superimposed using both the sagittal and postero-anterior (PA) tracings. Incremental growth of landmarks was measured in relation to Cartesian coordinates and compared between the superimposition methods. RESULTS: Sagittal and PA tracing superimpositions displayed an average 7 mm greater cephalad movement of landmarks, an average 2.4 mm greater ventral movement, and comparable transverse dimension with superimposition referenced at I-point as compared to the SACB reference. CONCLUSION: I-point superimposition demonstrates physiologic growth patterns concealed by traditional registration at sella turcica. The evolution of superimposition on SACB was based on convenience and reproducibility. Fundamental principles of bone development are consistent with the occipital condyles as a more biologic reference for relative craniofacial growth. Actual vertical growth is believed to be greater than displayed in this study, due to the cross-sectional and blended nature of the sample.

Cephalometric superimposition on the occipital condyles as a longitudinal growth assessment reference: I-point and I-curve.

This retrospective study tests the hypothesis that superimposition referenced at the occipital condyles (defined as I-point, I-curve) and oriented to the anterior cranial base (ACB) will display a growth pattern that is more consistent with independent evaluations, such as the Melsen necropsy specimens and the Bjork implant studies, when compared with traditional superimpositions referenced at sella turcica. Twenty-eight sets of serial lateral cephalometric radiographs were selected from an archived growth study. The apparent facial growth was compared using polar coordinate analysis from superimposition tracings of the serial films for each subject. The two superimposition methods were compared. The traditional method, ACB registered on the anterior curvature of sella turcica, versus registration on I-point while maintaining ACB parallel. I-point registered superimpositions consistently displayed a facial growth pattern that was more consistent with the classic necropsy specimens of children and the cephalometric studies superimposing on implant markers. Traditional ACB superimposition suggests that airway is restricted by normal growth. This apparent physiologic artifact does not occur when superimpositions are registered on I-point. Sella turcica displays vertical movement that is consistent with brain growth. These data indicate that registration on I-point is a more accurate physiologic representation of facial growth than the traditional ACB superimpositions. When compared with the traditional registration at sella turcica, I-point superimposition better elucidates physiologic growth patterns. As cephalometrics evolve from a two to a three dimensional science, it is important to use a more biologically valid registration for evaluating therapeutics and facial growth patterns.