INTRODUCTION<\/h2>\n
Almost 25 years have passed since the hypothesis that wastewater can be treated as a highly diluted urine sample [1, 2]. Gradually, a new interdisciplinary field of science emerged \u2013 Wastewater Based Epidemiology (WBE), which was initially focused primarily on licit and illicit drugs [3] and gradually expanded to\u00a0include other groups of substances and markers excreted by the population into wastewater. Municipal wastewater therefore contains a complex mixture of\u00a0chemical substances, including human metabolites \u2013 biomarkers. Quantitative measurement of these specific substances will provide information, for example, about the way of eating, health status of the population, occurrence of diseases, alcohol and drug consumption, and the population\u2019s exposure to environmental contaminants [4].<\/span><\/p>\n Given that SARS-CoV-2 viral particles were excreted by the infected population in urine and feces into municipal wastewater even before the onset of disease symptoms, the Covid-19 pandemic contributed significantly to increasing the importance of the epidemiological approach to wastewater [5, 6], which can also be used as an early warning tool before the onset of the disease.<\/span><\/p>\n The determination of selected drugs in municipal wastewater has long been used at the European and global level for monitoring drug consumption in monitored urban agglomerations; monitoring results are regularly published on the website of the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) [7, 8].<\/span><\/p>\n The TGM WRI hydrochemical laboratory has been dealing with the determination of illegal substances in wastewater for more than ten years. The range of\u00a0determined licit and illicit drugs is gradually expanding, and new substances are included in the analyses according to the current situation on the drug scene. The analytical method was developed according to the procedure published by Postigo et al. [9]. It is a fully automated on-line SPE and LC-MS\/MS (ESI) drug determination method. The analytical methods in ESI+ or ESI-mode are accredited, and the laboratory annually participates in an international interlaboratory comparison of tests. This interlaboratory comparison takes place as part of global monitoring of the drug situation organized by the SCORE-network [14]. A weekly monitoring sampling campaign is implemented every year in spring, usually from April to May, and was not interrupted even during the Covid-19 pandemic. We used the results obtained as part of these sampling campaigns to monitor the impact of the pandemic on drug use from the point of view of the epidemiological approach to wastewater, as we had the measurement results available for both \u201cnormal\u201d situation before the pandemic and during the period affected by Covid-19 and related measures. These measures also affected WWTP operation, which operated in a special regime.<\/span><\/p>\n From the beginning of March 2020 up to and including 2022, various regulations and measures affected by the current pandemic situation were gradually announced, expanded and cancelled, mainly in connection with the number of infected and hospitalized persons. The following overview briefly shows the announcement dates of\u00a0selected measures, especially those that were in force at the time of\u00a0the\u00a0weekly monitoring campaigns for determination of illicit substances. An\u00a0overall detailed overview of government resolutions related to the fight against coronavirus is available on\u00a0the\u00a0Government of the Czech Republic website [10].<\/span><\/p>\n Brief overview of selected measures related to Covid-19 pandemic:<\/span><\/span><\/em><\/p>\n 2019 \u2013 in December, Covid-19 was first identified in Wuhan, China<\/p>\n 2020<\/p>\n 2021<\/p>\n 2022<\/p>\n For comparison, sampling and analyses of wastewater from the Prague Central Wastewater Treatment Plant (CWWTP), at the inflow to the old (OWL) and new\u00a0(NWL) water line were chosen. At OWL, water is treated from main sewers B, D, E and F, at NWL from sewers A, C and K. The sewer network of the\u00a0capital city of Prague is shown in Fig. 1<\/span><\/em>. The sampling campaign always started on\u00a0Tuesday or Wednesday and ended a week later. 24-hour composite samples were collected, and sampling was carried out every 15 minutes at the inflow to\u00a0the\u00a0WWTP, behind the screens (after rough pre-treatment). Samples were cooled to 4\u20138 \u00b0C; if they could not be processed within 48 hours or, in the most urgent cases, within 72 hours, they were kept frozen at -20 \u00b0C until analysis. The\u00a0sampling dates in individual years are not exactly the same, but they were always carried out in April and May. In the years affected by the pandemic, the\u00a0sampling was also related to the capacity of the CWWTP, which worked in a\u00a0special regime. Sampling took place from 2nd to 8th April 2019, from 20th\u00a0to\u00a026th May 2020, from 20th to 26th April 2021, and from 6th to 12th April 2022.<\/span><\/p>\n \u00a0<\/span>Results were compared for MDMA (ecstasy, 3,4-methylenedioxy-N-methamphetamine), methamphetamine and its metabolite amphetamine, cocaine and its main metabolite benzoylecgonine, and the tetrahydrocannabinol (THC) metabolite 11-nor-9-carboxy-delta- 9-THC (nor-THC), i.e., for the most common drugs. Amphetamine is used as a drug in many countries rather than methamphetamine; in the Czech Republic, it is considered a metabolite of methamphetamine in wastewater. Ecstasy and cocaine are common \u201cparty\u201d drugs, i.e.,\u00a0drugs often used at parties and dancing events.<\/span><\/p>\n Methods for determining the above-mentioned analytes are described in\u00a0detail in Posp\u00edchalov\u00e1 et al. [11]. Before the actual analysis, the samples were conditioned to laboratory temperature, filtered through membrane filters to\u00a0remove solid impurities, and a mixed solution of internal standards, including isotopically labelled ones, was added to the filtrate. This was followed by online SPE-LC-MS\/MS determination, according to the analyte in ESI+ (MDMA, methamphetamine, amphetamine, cocaine, benzoylecgonine) or ESI- mode (nor-THC), and evaluation of the measured results.<\/span><\/p>\n After the borders were closed due to the Covid-19 pandemic, precursors for the production of meth became more difficult to find in the Czech Republic because these substances are largely smuggled from abroad, where their sale is not strictly limited by legislation. Closing of borders also affected the availability of cocaine and heroin, which also led to an increase in the prices of individual drugs. The situation on the drug scene gradually \u201cnormalized\u201d during 2020 and most probably adapted well to the existing state in 2021 and 2022. More detailed information about the situation on the drug scene during the state of emergency is available on the National Monitoring Centre for Drugs and Addiction website [12].<\/span><\/p>\n In this paper, we tried to evaluate the drug situation in the Czech Republic from the point of view of municipal wastewater analysis, i.e., use wastewater based epidemiology which, based on our experience, has a great informative value in terms of behaviour of the population connected to the sewage system in the monitored area.<\/span><\/p>\n Fig. 2<\/span><\/span><\/em> compares the average concentration values of individual drugs in\u00a0the\u00a0compared years. All values for a given analyte obtained from analyses of\u00a0samples taken at both water lines are averaged.<\/span><\/p>\n Porovn\u00e1v\u00e1ny byly v\u00fdsledky pro MD<\/p>\n It is obvious that in the first pandemic year (i.e., 2020), there was a partial reduction in the consumption of methamphetamine and ecstasy (MDMA) from the point of view of wastewater analysis. In the case of methamphetamine, this reduction was probably caused by the poorer availability of precursors for its production; in 2021, the situation on the methamphetamine drug market apparently normalized, and in the last pandemic year, at least during the sampling campaign, consumption even increased. For ecstasy, a typical \u201cparty\u201d drug, this was most probably the impact of the ban on organizing various events where ecstasy is used, and this trend continued in 2021. In 2022,\u00a0<\/span>during the sampling period, most of the nationwide measures were cancelled, and the drug scene reacted very quickly to this fact.<\/span><\/p>\n Marijuana, or its metabolite nor-THC, which is determined in wastewater, and cocaine\u2019s main metabolite benzoylecgonine, which is crucial for monitoring cocaine consumption, saw an increase in incidence over the years compared. Very high values for benzoylecgonine in 2022 are probably again related to the lifting of restrictive measures and the possibility to organize dance and other parties again. In the following figures, the measured results for individual drugs and their metabolites in the compared years are presented in the form of graphs. They always show the analyte concentration, date of sampling, and the water line where the sampling for the given substance was carried out. Weekends are marked with red or yellow columns.<\/span><\/p>\n The changes during the weekly monitoring are clearly visible here. It should be emphasized that the concentrations of monitored substances in wastewater are compared. Daily wastewater flows through a treatment plant do not vary much from day to day. Higher flows are mainly caused by rainfall, when the wastewater is diluted, and thus the concentrations of the monitored substances can be reduced.<\/span><\/p>\n Fig. 3 <\/span><\/span><\/em>to 6<\/span><\/em> show the determined concentrations of methamphetamine and amphetamine which, as mentioned above, is mainly a metabolite of methamphetamine in the Czech Republic.<\/span><\/p>\n Methamphetamine consumption does not vary much between the days of\u00a0the week, as can be seen in Figs. 3<\/span><\/em> and 5<\/span><\/em>. In Fig. 6<\/span><\/em>, the lower concentration on\u00a09th\u00a0April was probably caused by heavy rainfall and thus dilution of a water sample. In the following days, there was also a higher consumption of this drug, which could have been caused by the cancellation of emergency measures on 11th April. This increase is also clearly visible in Fig. 2<\/span><\/em>.<\/span><\/p>\n <\/p>\n Marijuana consumption is monitored by the rate of occurrence of its stable metabolite nor-THC (11-nor-9-carboxy-delta-9-THC). With this drug, as\u00a0with methamphetamine, consumption does not depend on specific days of\u00a0the\u00a0week. Figs. 7<\/span><\/em> to 10<\/span><\/em> document its consumption.<\/span><\/p>\n According to the wastewater analysis, marijuana consumption gradually increased between 2019 and 2022; practically the same applies to this drug as\u00a0in\u00a0the case of methamphetamine.<\/span><\/p>\n The determined concentration of ecstasy in municipal wastewater is presented in Figs. 11<\/span><\/em> to 14<\/span><\/em>. Due to its nature, consumption of this drug was reduced during Covid-19 pandemic.<\/span><\/p>\n The graphical representations above in particular confirm the weekend consumption of ecstasy. Fig. 11<\/span><\/em>, where the results are from a year unaffected by\u00a0the\u00a0pandemic situation, shows an up to fourfold increase in consumption at the weekend compared to a normal working day. In 2020, during the monitoring event, restrictive measures were still in force, including a state of emergency. However, there was already a reduction in the number of infected persons, and thus also less tension in the population. The state of emergency was lifted in mid-May 2020. The situation was similar in 2021: the state of emergency ended on 11th April, and measures according to the pandemic law remained in force. On 23rd April, there was another relaxation, to which the drug scene immediately reacted, as it was again possible to organize social events. Fig.\u00a014<\/span><\/em>\u00a0shows the situation most clearly for 2022. The reaction to the information that most of\u00a0the\u00a0emergency measures will be lifted from 11th April was immediate; the\u00a0consumption of ecstasy increased more than three times.<\/span><\/p>\n Cocaine is monitored through the determination of its most important metabolite, benzoylecgonine. It is an expensive drug used mainly at parties of\u00a0the\u00a0population group with higher incomes. In this case, we specifically selected the\u00a0inflow to the old water line (OWL) for monitoring, where the water from main sewer B is also discharged. Here, the cocaine concentration, or its metabolite benzoylecgonine, is the highest in the whole of Prague and prevails over methamphetamine, which is a typical dominant drug in the Czech Republic [13].<\/span><\/p>\n The results obtained for cocaine can be seen in Figs. 15<\/span><\/em> to 18<\/span><\/em>.<\/span><\/p>\n Basically, the same applies to cocaine as it does to ecstasy. In the graph in\u00a0Fig.\u00a015<\/span><\/em> (2019), there is again a typical increase at the weekend in the concentration of the monitored drug, especially its metabolite. In 2020 (Fig. 16<\/span><\/em>), the concentration is approximately the same throughout the week. As a\u00a0result of\u00a0the\u00a0restrictions imposed by the state of emergency, it was forbidden to\u00a0organize social events; the expectation of a certain relaxation did not show in the case of cocaine. In 2021 (Fig. 17<\/span><\/em>), the impact of the relaxation of emergency measures has already partially manifested itself. However, the most significant change is in METHODOLOGY<\/h2>\n
History of emergency measures in the Czech Republic<\/h3>\n
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Wastewater sampling and analysis<\/h3>\n
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<\/a><\/h2>\nFig. 1. Sewer network of the capital city of Prague<\/h6>\n
RESULTS AND DISCUSSION<\/h2>\n
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Fig. 2. Average concentration values of individual drugs and their metabolites in\u00a0the\u00a0compared years<\/h6>\n
Methamphetamine<\/h3>\n
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Fig. 3. Concentration of methamphetamine and amphetamine in wastewater sampled at the inflow to NWL in 2019 from April 3 to 9<\/h6>\n
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Fig. 4. Concentration of methamphetamine and amphetamine in wastewater sampled at the inflow to NWL in 2020 from May 20 to 26<\/h6>\n
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Fig. 5. Concentration of methamphetamine and amphetamine in wastewater sampled at the inflow to NWL in 2021 from April 20 to 26<\/h6>\n
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Fig. 6. Concentration of methamphetamine and amphetamine in wastewater sampled at the inflow to NWL in 2022 from April 6 to 12<\/h6>\n
Marijuana<\/h3>\n
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\u00a0<\/span>Fig. 7. Concentration of nor-THC (11-nor-9-carboxy-delta-9-THC) in wastewater sampled at the inflow to the OWL in 2019 from April 3 to 9<\/h6>\n
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Fig. 8. Concentration of nor-THC (11-nor-9-carboxy-delta-9-THC) in wastewater sampled at the inflow to the OWL in 2020 from May 20 to 26<\/h6>\n
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Fig. 9. Concentration of nor-THC (11-nor-9-carboxy-delta-9-THC) in wastewater sampled at the inflow to the OWL in 2021 from April 20 to 26<\/h6>\n
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Fig. 10. Concentration of nor-THC (11-nor-9-carboxy-delta-9-THC) in wastewater sampled at the inflow to the OWL in 2022 from April 6 to 12<\/h6>\n
Ecstasy (MDMA,\u00a03,4-methylenedioxy-N-methamphetamine)<\/span><\/h3>\n
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Fig. 11. Concentration of ecstasy (MDMA, 3,4-methylendioxy-N-methamphetamine) in\u00a0wastewater sampled at the inflow to OWL in 2019 from April 3 to 9<\/h6>\n
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Fig. 12. Concentration of ecstasy (MDMA, 3,4-methylene-dioxy-methamphetamine) in\u00a0wastewater sampled at the inflow to OWL in 2020 from May 20 to 26<\/h6>\n
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Fig. 13. Concentration of ecstasy (MDMA, 3,4-methylendioxy-N-methamphetamine) in\u00a0wastewater sampled at the inflow to OWL in 2021 from April 20 to 26<\/h6>\n
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Fig. 14. Concentration of ecstasy (MDMA, 3,4-methylene-dioxy-methamphetamine) in\u00a0wastewater sampled at the inflow to OWL in 2022 from April 6 to 12<\/h6>\n
Cocaine<\/h3>\n