No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Urban Wastewater Treatment Plants as Hotspots for Antibiotic Resistant Bacteria and Genes Spread into the Environment: A Review
Abstract
Urban wastewater treatment plants (UWTPs) are among the main sources of antibiotics' release into the environment. The occurrence of antibiotics may promote the selection of antibiotic resistance genes (ARGs) and antibiotic resistant bacteria (ARB), which shade health risks to humans and animals. In this paper the fate of ARB and ARGs in UWTPs, focusing on different processes/technologies (i.e., biological processes, advanced treatment technologies and disinfection), was critically reviewed. The mechanisms by which biological processes influence the development/selection of ARB and ARGs transfer are still poorly understood. Advanced treatment technologies and disinfection process are regarded as a major tool to control the spread of ARB into the environment. In spite of intense efforts made over the last years to bring solutions to control antibiotic resistance spread in the environment, there are still important gaps to fill in. In particular, it is important to: (i) improve risk assessment studies in order to allow accurate estimates about the maximal abundance of ARB in UWTPs effluents that would not pose risks for human and environmental health; (ii) understand the factors and mechanisms that drive antibiotic resistance maintenance and selection in wastewater habitats. The final objective is to implement wastewater treatment technologies capable of assuring the production of UWTPs effluents with an acceptable level of ARB.
... Marten Klatt [10] used a membrane bioreactor to achieve efficient removal of hospital pathogens, but the biodegradation of some recalcitrant substances was not sufficient. Rizzo [11] reported that UV radiation has a broad spectrum of applications in deep treatment; the radiation generated by UV light can damage cellular DNA and cause the water to produce strong oxidizing groups such as •OH and H 2 O 2 under high-energy radiation to achieve virus inactivation and organic matter degradation in the wastewater of the deep treatment process, and no byproducts are produced, which is a green and widely applicable method [12]. It is a green and widely applicable process. ...
... The absorbance of ciprofloxacin was measured at 277 nm with a total reaction duration of 120 min. The photocatalytic reaction was carried out in a homemade photocatalytic reactor by varying the AC loading of Fe 3+ -TiO 2 /AC (5%, 10%, 20%, and 30%, mass percentage) and the doping of Fe 3+ (0.1%, 0.2%, 0.3%, 0.4%, 1%, molar percentage) to investigate the optimal ratio of the materials; changing the solution pH (3,5,7,9,11), the initial concentration of pollutants (1-10 mg/L), and the intensity of light (40 W, 29 W) to investigate the variation in the pollutant removal rate under the influence of different factors; and finally, the photocatalytic stability of the composites. The degradation rate of pollutants can be calculated using Equation (1): ...
Low-concentration antibiotic wastewater is difficult to treat rapidly using conventional photocatalysts. For this reason, this paper simplified the traditional sol–gel method to prepare Fe3+-TiO2/AC composites and characterized the properties of the composites using FT-IR, XRD, SEM, BET, and TEM. The results demonstrated that iron was uniformly dispersed on the surface of the composites, and the activated carbon (AC) was successfully loaded with iron-doped titanium dioxide. Afterward, ciprofloxacin (CIP) was used as the target degradant, and the effects of different activated carbon loadings, iron-doping, pH, initial concentrations, and UV light intensities on the removal of ciprofloxacin were investigated. The repetitive photocatalytic stability of the composites was studied, and the reaction mechanism was explored by using free radical quenching experiments. The results demonstrated that while iron doping reduced the rate at which photogenerated electrons and holes could combine, loading AC increased the usage efficiency of the composites’ adsorption and catalytic active sites. According to the parameter tests, the circumstances that led to the highest CIP degradation efficiency (94.59%) were as follows: 10 mg/L CIP, 0.5 g/L 0.2%Fe3+-TiO2/20%AC, and solution pH of 7 under 40 min of UV light irradiation. In addition, the Fe3+-TiO2/AC composite material has excellent cyclic stability, the degradation rate of CIP can still reach 87.73% at 60 min after four repeated degradation tests under the same conditions. The applicability of this method could be expanded to the treatment of various industrial organic pollutants in water.
... Water, particularly drinking water, contaminated with animal or human faces containing antibiotic-resistant bacteria is considered a key source of the spread of antibiotic resistance, antibiotic residues, and extracellular mobile genetic elements associated with antibiotic-resistant organisms and a source of new antibiotic resistance genes (Larsson & Flach, 2022). Urban and livestock wastewater, circulating water, and effl uents from antimicrobial drug manufacturing facilities are identifi ed as hotspots for antibiotic-resistant bacteria and resistance genes that spread into the environment and require special monitoring (Rizzo et al., 2013;Burgmann et al., 2018;Symochko et al., 2019a;Symochko et al., 2023). ...
The excessive and uncontrolled use of antibiotics in healthcare and agriculture has become the main cause of antibiotic resistance and the spread of antibiotic-resistant microorganisms in the environment. Every year, antibiotic resistance continues to grow worldwide, posing a significant challenge for humanity. The World Health Organization recognizes antibiotic resistance as one of the top 10 global threats to public health, jeopardizing the achievement of the United Nations Sustainable Development Goal. The consequences of antimicrobial resistance extend beyond the effective treatment of infections; it is a public health issue with global consequences. Socioeconomic losses and environmental threats are closely linked to this problem. Moreover, antibiotic resistance has long-term and unpredictable consequences related to wars and military conflicts. The impact of the war and a clear understanding of the global scope of antibiotic resistance as a result of hostilities can be seen through the outcome of russia's invasion of Ukraine on February 24, 2022. This military conflict not only highlighted the importance of preserving human life and promoting peaceful development at both national and global levels but also revealed potential risks for various global environmental issues, particularly the increasing spread of antibiotic resistance. The disruption caused by the war has transformed the issue of antibiotic resistance in Ukraine from a national concern to a powerful factor that affects the global scale of this problem, transcending borders. During the war, resources and priorities have shifted toward military needs, relegating human health and environmental protection to secondary importance. Migration and loss of highly qualified medical specialists and biologists, deterioration of the environmental situation, sanitary, hygienic conditions, lack of food resources and medicines, and other factors directly and indirectly contribute to the emergence and spread of antibiotic resistance globally. This review demonstrates the relevance of the issue of antibiotic resistance, the main mechanisms of the development and spread of antibiotic resistance, the global scale and consequences of antibiotic resistance due to the war in Ukraine and aims to provide an overview of international initiatives to reduce antibiotic resistance, in particular the “One Health” concept and strategies, which are currently being implemented. Solving the issue of antibiotic resistance globally and at the level of the country affected by war requires a comprehensive approach, which includes mandatory systematic surveillance of antibiotic resistance, control of the use of antimicrobial drugs, informing all segments of society about the use of antimicrobial drugs and the consequences of antibiotic resistance, the introduction of educational programs to prevent an improper use of antibiotics, environmentally safe methods of waste disposal in medical, food, and other enterprises that produce or use antibiotics, safe disposal of antibiotics, etc.
... take up recombinant genes upon disposal of sludge from urban wastewater treatment and biomanufacturing waste151,152 . HGT was evident after the 1988 release of Pseudomonas putida engineered for phenol-degradation (Box 1). ...
... The questions pertaining to public utilities and infrastructure of the water supply, including the provision of both freshwater and sewage services, are fundamental to the urbanization phenomenon. The present problems for architects, planners and policymakers are the availability of jobs, health care, social services and access to facilities of basic human needs such as clean water and sanitation ( [6] and [7] ), The water quality is on the other hand compromised by the presence, in urban and rural water sources, of a significant number of pathogens [8] and anthropogenic chemicals [9] Municipal and municipal wastewater collection facilities asignificant element in aquatic contamination in the world has been recognized [10]. The majority of household and industrial waste water is dumped directly in water sources in many developed countries without any filtering or only after primary treatment. ...
Many water supplies are now contaminated by anthropogenic sources such as
domestic and agricultural waste, as well as manufacturing activities, the
public's concern about the environmental effects of wastewater contamination
has grown. Several traditional wastewater treatment methods, such as
chemical coagulation, adsorption, and activated sludge, have been used to
eliminate pollution; however, there are several drawbacks, most notably high
operating costs, because of its low operating and repair costs, the usage of
aerobic waste water treatment as a reductive medium is gaining popularity.
Furthermore, it is simple to produce and has a high efficacy and potential to
degrade pollutants.
... Among various ECs, elimination of antibiotics from wastewater poses a significant challenge to conventional WWTPs. Compounding this issue, these WWTPs serve as incubators for the growth of antibiotic-resistant bacteria (ARBs) and genes (ARGs) and thus are considered as hotspots for the dissemination of ARBs and ARGs [124,125]. Recently, a full-scale AOP based on UV-C/PMS (UV wavelength range 100À280 nm) was set up for the treatment of antibiotics and ARGs for the first time. The Estiviel WWTP in Toledo, Spain, treats an average flow of 12,000 m 3 /day and employs a tertiary treatment, which consists of coagulation/flocculation/decantation, rotofilter, and UV-C reactor [126]. ...
Recently, a new class of organic pollutants has been widely reported in aquatic environments, including the treated effluents. These pollutants, referred to as emerging contaminants (ECs) in the scientific literature, are not commonly monitored or regulated in wastewater but have the potential to cause adverse effects both on the environment and human health. Due to the inability of conventional wastewater treatment plants to efficiently remove ECs, physicochemical treatment technologies have gained tremendous popularity as powerful tools for tackling this critical issue. These advanced treatment methods exploit various physical and chemical processes to eliminate ECs, which can significantly improve the quality of treated effluents and reduce the environmental impact. This chapter provides a comprehensive overview of these physicochemical treatment technologies, including high retention separation membranes and electrified catalytic membranes, Fenton, photo-Fenton and electro-Fenton advanced oxidation processes (AOPs), UV photolysis, ozonation, sulfate radical-based AOPs, and photocatalytic advanced oxidation/reduction processes. We discussed the underlying principles, mechanisms, and performance of these methods for the removal of ECs. A significant aspect of the chapter is the scale-up of the physicochemical treatment technologies. We explored case studies of pilot-scale applications, highlighting the setup, operational conditions, and outcomes of full-scale implementations.
... However, during the last century, environmental microbial communities have been constantly subjected to invasion events by antibiotic resistant bacteria (ARB), and their associated ARGs, which have been enriched or released through anthropogenic activities. For example, release and reuse of wastewater effluents or the application of manure to soils are known to be primary conduits of AMR spread in the aquatic and terrestrial environments [9][10][11][12] . But even habitats with no direct anthropogenic impact are regularly exposed to lower frequencies of such invasion events, for example, through wildlife and aerial depositions. ...
When antimicrobial resistant bacteria (ARB) and genes (ARGs) reach novel habitats, they can become part of the habitat’s microbiome in the long term if they are able to overcome the habitat’s biotic resilience towards immigration. This process should become more difficult with increasing biodiversity, as exploitable niches in a given habitat are reduced for immigrants when more diverse competitors are present. Consequently, microbial diversity could provide a natural barrier towards antimicrobial resistance by reducing the persistence time of immigrating ARB and ARG. To test this hypothesis, a pan-European sampling campaign was performed for structured forest soil and dynamic riverbed environments of low anthropogenic impact. In soils, higher diversity, evenness and richness were significantly negatively correlated with relative abundance of >85% of ARGs. Furthermore, the number of detected ARGs per sample were inversely correlated with diversity. However, no such effects were present in the more dynamic riverbeds. Hence, microbiome diversity can serve as a barrier towards antimicrobial resistance dissemination in stationary, structured environments, where long-term, diversity-based resilience against immigration can evolve.
... The alternative lies in developing tailored solutions for each case using new technology that enables high-quality purification, facilitating the reuse of treated water in the effluent production area and avoiding the extensive and costly sewer networks that require huge initial and maintenance investments, despite which they continue to cause ongoing contamination of aquifers, continental surface waters, and coasts. 2 The alternative solution of applying tailored purification to each area, based on the specific characteristics of the effluent, has faced criticism that lacks consistent arguments, driven solely by the desire to maintain the immense business of wastewater management. [5][6][7][8][9][10][11][12][13][14][15][16][17][18] This article unequivocally demonstrates the benefits of applying contemporary technology to local problems, enabling the complete reuse of water and sludge without posing any health or environmental risks. Additionally, it highlights the flexibility of the system, which can also provide solutions for small populations, something that is impossible with current standard technology. ...
The only solution to avoid pollution due to sewage is to treat the effluent and reuse water and sludge at the wastewater production site. This makes it necessary to install treatment plants distributed along the sewer system to prevent the movement of effluents with the consequent losses to the aquifer. This new management is called decentralized wastewater treatment. To make this change possible, it is necessary to develop treatment plants that do not represent a health hazard for citizens, eliminating the nuisance caused by traditional plants. These plants need to be robust, operate autonomously, have reduced maintenance and exhaustive control of each process, where each cubic meter of effluent is perfectly controlled, avoiding losses to the aquifer or discharges that may occur due to deficiencies in their purification capacity; constantly producing water and sludge, which are directly reusable with all the health and environmental guarantees regardless of variations in inlet effluent or environmental conditions. This article presents a paradigmatic case of a decentralized solution with conditions of maximum complexity that has been solved efficiently, saving around 98% of the initial investment, completely avoiding wastewater discharges into the environment and reusing 100% of the treated water, in 14 years of operation.
... Waste water tr eatment plants (WWTPs) ar e of particular inter est for the study of AMR because they collect w astew ater from multiple sources, including human and non-human sources, and provide a suitable environment for close bacterial interaction and exchange of antibiotic-resistance genes (ARGs). In WWTPs, high nutrient concentration, high bacterial density, and the presence of sub-inhibitory concentrations of antibiotics and other selective a gents suc h as heavy metals and disinfectants favour horizontal gene transfer (HGT) in bacteria (Rizzo et al. 2013, Karkman et al. 2018, Manaia et al. 2018, Pazda et al. 2019. Of particular interest is to know which bacteria are the hosts of clinically significant ARGs and how the composition of the hosts is altered b y the w astew ater tr eatment pr ocess. ...
Wastewater treatment plants (WWTPs) provide a suitable environment for the interaction of antibiotic resistant bacteria and antibiotic-resistance genes (ARGs) from human, animal, and environmental sources. The aim was to study the influent and effluent of two WWTPs in Croatia to identify bacterial hosts of clinically important beta-lactamase genes (blaTEM, blaVIM, blaOXA-48-like) and observe how their composition changes during the treatment process. A culture-independent epicPCR was used to identify the ARG hosts, and 16S rRNA amplicon sequencing to study the entire bacterial community. Different wastewater sources contributed to the significant differences in bacterial composition of the wastewater between the two WWTPs studied. A total of 167 genera were detected by epicPCR, with the Arcobacter genus, in which all ARGs studied were present, dominating in both WWTPs. In addition, the clinically important genera Acinetobacter and Aeromonas contained all ARGs examined. The blaOXA-48-like gene had the highest number of hosts, followed by blaVIM, while blaTEM had the narrowest host range. Based on 16S rRNA gene sequencing, ARG hosts were detected in both abundant and rare taxa. The number of hosts carrying investigated ARGs was reduced by wastewater treatment. EpicPCR provided valuable insights into the bacterial hosts of horizontally transmissible beta-lactamase genes in Croatian wastewater.
... Since wastewater treatment plants function as a confluence point for bacteria and sewage from various sources, they can be hotspots for selecting and disseminating antibiotic resistance genes [25]. In addition, the high bacterial density in raw sewage provides a favorable environment for the horizontal transfer of antibiotic-resistant genes between bacteria via mobile genetic elements, such as plasmids or transposons [23,26]. ...
The relative contribution of factors responsible for the environmental exposure of active pharmaceutical ingredients (APIs) is of interest for appropriate remedial measures. This study was carried out to evaluate the post-lockdown levels of APIs in water resources, in comparison to our previously published study from 2016. The environmental levels of 28 drugs from different classes were analyzed in surface water (Yamuna River), aquifers, and leachate samples collected from 26 locations in Delhi-NCR using the previously validated liquid chromatography-mass spectrometry (LC-MS/MS) methods. In addition, the prevalence of antimicrobial resistance in coliforms isolated from targeted surface water samples was also studied. This study revealed that more than 90% of APIs, including antibiotics, decreased drastically in both surface water and aquifers compared to our previous data. Selected samples subjected to antimicrobial resistance (AMR) analysis revealed the presence of cephalosporin-resistant coliform bacteria. Tracing cephalosporins in the surface and drain water samples revealed the presence of ceftriaxone in the drain and water samples from Yamuna River. Higher levels of ceftriaxone in landfill leachate were also found, which were found to be associated with coliform resistance and indicate the un-segregated disposal of medical waste into landfills. Social restrictions enforced due to COVID-19 resulted in a drastic decrease in antimicrobials and other APIs in aquatic water resources. Increased ceftriaxone and cephalosporin resistance was seen in coliform from surface water and drain, indicating the possibility of hospital waste and treatment-related drugs entering Yamuna River. Enforcement of the regulations for the safe disposal of antibiotics at hospitals and preliminary disinfection of hospital sewage before its inflow into common drains might help minimize the spread of antibiotic resistance in the environment.
... These differences may be due to some factors such as population density, industrial zones, agricultural practices, and wastewater treatment plants, which can significantly impact the prevalence of antibiotic-resistant bacteria in aquatic environments. 41,42 In a nutshell, the main goal achieved in this study was setting a reproducible sampling method by using APSs, as an efficient and easy-tohandle tool for monitoring the presence of MDR bacteria. These results hold significant importance from the One Health perspective, as they ...
... Municipal wastewater treatment facilities are major reservoirs of antibiotics and their metabolites (Jia et al., 2012). These compounds are often not entirely removed during the wastewater treatment process (Hijosa-Valsero et al., 2011;Rizzo et al., 2013;Rowan, 2011); therefore, treated municipal wastewater (TMW) may contain these compounds (Giger et al., 2003). As TMW is discharged to the environment (e.g., surface water, groundwater, agricultural soil, etc.), the exposure of bacteria to sublethal concentrations of antibiotics in these environments can create a selective pres-sure that drives the development and proliferation of antibioticresistant strains. ...
The spread of antimicrobial resistance (AMR) in agricultural systems via irrigation water is a serious public health issue as it can be transmitted to humans through the food chain. Therefore, understanding the dissemination routes of antibiotic resistance genes (ARGs) in agricultural systems is crucial for the assessment of health risks associated with eating fresh vegetables such as spinach and radish irrigated with treated municipal wastewater (TMW). In this study, we investigated the bacterial community structure and resistome in the soil–plant–earthworm continuum after irrigation of spinach and radish with TMW containing the antibiotics trimethoprim (TMP), sulfamethoxazole (SMZ), and sulfapyridine (SPD) using 16S rRNA gene sequencing and high throughput quantitative PCR (HT-qPCR). The study was conducted in two phases: Phase I involved eight weeks of spinach and radish production using TMW for irrigation, whereas Phase II entailed three weeks of earthworm exposure to contaminated plant material obtained in Phase I. The 16S data indicated that the rhizosphere bacterial community composition and structure were more resilient to antibiotic residuals in the irrigated water, with radish showing less susceptibility than spinach than those of bulk soils. The HT-qPCR analysis revealed that a total of 271 ARGs (out of 285) and 9 mobile genetic elements (MGEs) (out of 10) were detected in all samples. Higher diversity and abundance of ARGs were observed for samples irrigated with higher concentrations of antibiotics in both spinach and radish treatments. However, compared to spinach, radish ARG dynamics in the soil biome were more stable due to the change of antibiotic introduction to the soil. At the class level, multi-drug resistance (MDR) class was altered significantly by the presence of antibiotics in irrigation water. Compared to earthworm fecal samples, their corresponding soil environments showed a higher number of detected ARGs, suggesting that earthworms could play a role in reducing ARG dissemination in the soil environments. These findings will not only provide insight into the dissemination of ARGs in agricultural environments due to antibiotic residuals in irrigated water but could help understand the potential human health risks associated with ARGs.
... Human and animal waste is the primary source of antibiotic dissemination into the environment, containing significant amounts of unmetabolized antimicrobials [37]. The widespread use of antibiotics in households results in their presence in wastewater, making municipal wastewater treatment plants (WWTPs) major contributors to ARGs and antibiotic-resistant bacteria (ARB) in the environment [38]. ...
This review aims to examine the existence of Pseudomonas aeruginosa (P. aeruginosa) and their antibiotic resistance genes (ARGs) in aquatic settings and the alternative treatment ways. P. aeruginosa in a various aquatic environment have been identified as contaminants with impacts on human health and the environment. P. aeruginosa resistance to multiple antibiotics, such as sulfamethoxazole, ciprofloxacin, quinolone, trimethoprim, tetracycline, vancomycin, as well as specific antibiotic resistance genes including sul1, qnrs, blaVIM, blaTEM, blaCTX, blaAIM-1, tetA, ampC, blaVIM. The development of resistance can occur naturally, through mutations, or via horizontal gene transfer facilitated by sterilizing agents. In addition, an overview of the current knowledge on inactivation of Pseudomonas aeruginosa and ARG and the mechanisms of action of various disinfection processes in water and wastewater (UV chlorine processes, catalytic oxidation , Fenton reaction, and ozonation) is given. An overview of the effects of nanotechnology and the resulting wetlands is also given.
... The dissemination of AMR is not solely affected by the antibiotics; there are other additional factors, including pharmaceuticals, pesticides, and heavy metals in water, which can induce selective pressure stress in bacteria, contributing to the selection of antimicrobialresistant bacteria [18]. Research indicates that high levels of ARGs can be transferred under heavy metal stress in bacterial communities within sludge [19]. ...
Valley surface water is considered a focal public health concern owing to the presence of multi-drug-resistant bacteria. The distribution of antimicrobial resistance (AMR) bacteria in the surface water is affected by the presence of multiple factors, including antibiotics coming from wastewater discharge or other contaminant sources such as pharmaceuticals, biocides, and heavy metals. Furthermore, there is evidence suggesting that high levels of antibiotic resistance genes (ARGs) can be transferred within bacterial communities under the influence of heavy metal stress. Hence, the primary aim of this study is to investigate the presence of heavy metals and bacterial ARGs in upstream as well as downstream locations of Wadi Hanifah Valley in Riyadh, Saudi Arabia. Sample collection was conducted at eighteen surface water sites within the valley in total. The selection of ARGs was associated with the most common antibiotics, including β-lactam, tetracycline, erythromycin, gentamicin, sulphonamide, chloramphenicol, vancomycin, trimethoprim, and colistin antibiotics, which were detected qualitatively using polymerase chain reaction (PCR) technology. The tested antibiotic resistance genes (ARGs) included (blaNDM-1 (for the antibiotic class Beta-lactamases), mecA (methicillin-resistant Staphylococcus aureus), tet(M) and tet(B) (for the antibiotic class Tetracycline), ampC (for the antibiotic class Beta-lactamases), vanA (for the antibiotic class vancomycin), mcr-1 (for the antibiotic class colistin), erm(B) (for the antibiotic class erythromycin), aac6′-Ie-aph2-Ia (for the antibiotic class Gentamicin), sulII (for the antibiotic class sulphonamide), catII (for the antibiotic class Chlorophincol), and dfrA1 (for the antibiotic class trimethoprim). Moreover, an assessment of the levels of heavy metals such as lithium (Li), beryllium (Be), chromium (Cr), cobalt (Co), arsenic (As), cadmium (Cd), tin (Sn), mercury (Hg), and lead (Pb) was conducted by using inductively coupled plasma mass spectrometry (ICPMS). According to our findings, the concentrations of sulphonamide, erythromycin, and chloramphenicol ARGs (erm(B), sulII, and catII) were observed to be the most elevated. Conversely, two ARGs, namely mecA and mcr-1, were not detected in the samples. Moreover, our data illustrated a significant rise in ARGs in the bacteria of water samples from the upstream sites as compared with the water samples from the downstream sites of Wadi Hanifah Valley. The mean concentration of Li, Be, Cr, Co, As, Cd, Sn, Hg, and Pb in the water samples was estimated to be 37.25 µg/L, 0.02 µg/L, 0.56 µg/L,0.32 µg/L, 0.93 µg/L, 0.01 µg/L, 200.4 µg/L, 0.027 µg/L, and 0.26 µg/L, respectively, for the selected 18 sites. Furthermore, it was revealed that the concentrations of the screened heavy metals in the water samples collected from various sites did not surpass the maximum limits set by the World Health Organization (WHO). In conclusion, this study offers a concise overview of the presence of heavy metals and ARGs in water samples obtained from the Wadi Hanifah Valley in Riyadh, KSA. Such findings will contribute to the ongoing monitoring and future risk assessment of ARGs spread in surface water.
... The rise in antibiotic-resistant bacteria poses the most significant challenge, as even small concentrations of antibiotics have the potential to foster their development (Nazaret & Aminov, 2014). These bacteria can generate genes that provide them with protection against antibiotics (Rizzo et al., 2013). According to the World Bank's projections, by the year 2050, these antibiotic-resistant bacteria have the potential to cause the deaths of 10 million people annually and drive approximately 28 million people into poverty (Bloom et al., 2017). ...
A multistage effluent treatment plant (ETP) was installed in a pharmaceutical factory in Comilla District, Bangladesh, to treat the discharged effluent from the factory. This study evaluated the treatment performance of the ETP through physical, chemical, and biological treatment processes. The treatment plant consisted of equalization, neutralization, anaerobic followed by aerobic biodegradation, two-stage clarification, reed bed bio-filtration, and post-aeration units. Water quality parameters including pH, turbidity, TDS, TSS, TS, COD and BOD5 were analyzed at the IUBAT-Environmental Engineering Laboratory. The ultimate removal efficiencies were 100% for turbidity, 97% for TSS, 95% for COD, and 97% for BOD5. The average reduction ranges for, turbidity, TSS, TS, COD and BOD5 were 201 FTU to 0 FTU, 512 mg/L to 13 mg/L, 912 mg/L to 530 mg/L, 2034 mg/L to 99 mg/L, and 1126 mg/L to 29 mg/L respectively. The anaerobic treatment unit achieved 1%, 17%, and 37% removal efficiencies for turbidity, COD, and BOD5, while the aerobic unit achieved 76%, 86%, and 86% removal efficiencies. The chemical treatment process was found effective for TSS and turbidity removal. The Reed Bed System performed well in removing BOD5, COD, and Turbidity of the effluent. The ETP's final discharge complied with water quality standards set by the Department of Environment, Bangladesh.
... In some countries, biosolids are used as fertilizer in agriculture and forestry or for landscaping and land remediation (Kang et al., 2022;LeBlanc et al., 2009;Marchuk et al., 2023). However, land applications of biosolids have been found to enhance the abundance of ARGs in soils (Bondarczuk et al., 2016;Kang et al., 2022;Munir et al., 2011;Murray et al., 2019;Rizzo et al., 2013;Ross and Topp, 2015;Wang et al., 2023;Yang et al., 2018). ...
... Several of the cultivated environmental samples from our study contained few CFUs, it is however noteworthy, that from these few, MDR bacteria were found that are associated with disease in humans (Brinas et al., 2002;Paterson, 2006;Aiello et al., 2008;Kim et al., 2009;Rizzo et al., 2013;Akbari et al., 2016;Prokesch et al., 2016;DavinRegli et al., 2019;CDC, 2023). ...
Several studies have investigated the effects of swimming in sewage-polluted recreational beach water, highlighting the associated health hazards. To mitigate potential pathogen transmission, it is imperative that the polluted water is released away from recreational waters and foreshores, where children tend to play. At present, domestic sewage in the Faroe Islands solely undergoes primary wastewater treatment within primary settling tanks before being discharged into the ocean. Effluents are a major anthropogenic source of antibiotic resistance genes and antibiotic resistant bacteria, which are released into the environment. The aim of this study was to investigate antibiotic resistant Gram-negative bacteria and antibiotic resistance genes in influents and effluents of wastewater subjected solely to primary treatment, along with their release into the environment during both summer and winter. Water samples were collected from influents and effluents as well as with increasing distance away from the wastewater outlet and from nearby tidepools. Samples were cultured on MacConkey agar with four different antibiotics for detection of antibiotic-resistant bacteria and antibiotic resistance genes were quantified by droplet digital PCR. All multi-drug resistant bacteria were identified using the API 20E kit. We observed an overall decrease of the abundance of Gram-negative bacteria from the effluents compared to influents, however, we observed the opposite trend in the antibiotic resistance genes. Antibiotic resistant bacteria and antibiotic resistance genes in addition to multi-drug resistant bacteria were found in the surrounding oceanic and several terrestrial tidepool samples. Of the multi-drug resistant bacteria, we found, e.g., Escherichia coli, P. aeruginosa, and A. hydrophila species, which can be pathogenic, potentially causing an infection if encountering a host. These results indicate a relatively wide pollution range of the effluents from the septic tank and treated sewage released into the environment, posing a potential hazard for both humans and wildlife.
... The release of antibiotics and resistance genes into the environment has inspired intensive research efforts to trace their distribution pattern and mode of degradation. (Martinez, 2009;Pruden et al., 2006;Rizzo et al., 2013) This makes the search for exact mechanism causing the occurrence of multi-drug resistant microbes becoming uncertain. ...
The eutrophication of open water bodies, especially in coastal water area, has received intensive public focuses in recent years. Biological treatment of organic pollutants in industrial and domestic wastewater requires microbial flora to participate. The environmental impact of these microbes, particularly in regard to the spread of antibiotic resistance genes and pathogenicity in coastal water, remains largely unknown. We initiate studies to examine antibiotic resistance genes (ARGs) in microbial flora and study the mode of antibiotic spread in the coastal waters of Bohai Bay, North China by using high performance liquid chromatography-mass spectrum analysis and quantitative polymerase chain reaction. The metabolic features of bacteria related to the presence of ARGs in the coastal area showed the enrichment of Pseudomonadale species with triple resistance properties (ampicillin, kanamycin and gentamycin) at the estuary site locating adjacent to a coastal sewage treatment plant where both type I and type II polyketide synthase genes and b-lactam type antibiotics are detected. Pseudomonadale order is phylogenetically belonged to Proteobacteria Phylum, Gamma-proteobacteria Class and contains Pseudomonadaceae family and Pseudomonas genus. QPCR analysis suggests that the percent occupation of Pseudomonadale species to total Proteobacteria in the examined stations is close to several sampling stations close to sewage treatment plants, where consistent release of organic wastes occurred.
... Indeed, WWTP processes seem to be favorable places for Horizontal Gene Transfer (HGT) and the selection of antibiotic resistant bacteria (ARB) because of high microbial density, high nutrient content and presence of ECs and other xenobiotics (Qiao et al., 2018). After treatment, a certain quantity of ARGs and/or ARB and/or Mobile Genetic Elements (MGEs) remains in water (Laht et al., 2014;Lekunberri et al., 2017;Maganha de Almeida Kumlien et al., 2021;Munir et al., 2011;Rizzo et al., 2013). Fig. 3 summarizes concentrations of ARGs groups in freshwater ecosystems and in effluent wastewater. ...
... Mexico, which is the second-largest economy in LA, is reported to be one of the top consumers of antibiotics in the region (SRE, 2023). In LA, over 50 % of the wastewater is released into the environment untreated (Rizzo et al., 2013;Souza et al., 2022). Unfortunately, wastewater treatment WWTPs in Mexico and many other countries lack proper regulations and technology to effectively remove many of the antibiotics, resulting in the discharge of antibiotics into water and sediments (Martínez-Orgániz et al., 2021). ...
Waste Water Treatment Plants (WWTP) aim to reduce contamination in effluent water; however, studies indicate antimicrobial resistance genes (ARGs) persist post-treatment, potentially leading to their spread from human populated areas into the environment. This study evaluated the impact of a large WWTP serving 125,000 people on the Iskar River in Bulgaria, by characterizing the spatial and short-term temporal dynamics in bacterial community dynamics and resistance profiles of the surface water. Pairs of samples were collected biweekly on four dates from two different locations, one about 800 m after the WWTP effluents and the other 10 km downstream. Taxonomic classification revealed the dominance of Pseudomonodota and Bacteriodota, notably the genera Flavobacterium, Aquirufa, Acidovorax, Polynucleobacter, and Limnohabitans. The taxonomic structure corresponded with both lentic and lotic freshwater habitats, with Flavobacterium exhibiting а significant decrease over the study period. Principal Coordinate Analysis revealed statistically significant differences in bacterial community composition between samples collected on different dates. Differential abundance analysis identified notable enrichment of Polynucleobacter and Limnohabitans. There were shifts within the enriched or depleted bacterial taxa between early and late sampling dates. High relative abundance of the genes erm(B), erm(F), mph(E), msr(E) (macrolides); tet(C), tet(O), tet(W), tet(Q) and tet(X) (tetracyclines); sul1 and sul2 (sulphonamides); and cfxA3, cfxA6 (beta-lactams) were detected, with trends of increased presence in the latest sampling dates and in the location closer to the WWTP. Of note, genes conferring resistance to carbapenems blaOXA-58 and blaIMP-33-like were identified. Co-occurrence analysis of ARGs and mobile genetic elements on putative plasmids showed few instances, and the estimated human health risk score (0.19) according to MetaCompare2.0 was low. In total, 29 metagenome-assembled genomes were recovered, with only a few harbouring ARGs. This study enhances our understanding of freshwater microbial community dynamics and antibiotic resistance profiles, highlighting the need for continued ARGs monitoring.
This study investigated the occurrence, removal rate, and potential risks of 43 organic micropollutants (OMPs) in four municipal wastewater treatment plants (WWTPs) in Korea. Results from two-year intensive monitoring confirmed the presence of various OMPs in the influents, including pharmaceuticals such as acetaminophen (pain relief), caffeine (stimulants), cimetidine (H2-blockers), ibuprofen (non-steroidal anti-inflammatory drugs- NSAIDs), metformin (antidiabetics), and naproxen (NSAIDs) with median concentrations of >1 μg/L. Some pharmaceuticals (carbamazepine-anticonvulsants, diclofenac-NSAIDs, propranolol-β-blockers), corrosion inhibitors (1H-benzotriazole-BTR, 4-methyl-1H-benzotriazole-4-TTR), and perfluorinated compounds (PFCs) were negligibly removed during WWTP treatment. The OMP concentrations in the influents and effluents were mostly lower in August than those of other months (p-value < 0.05) possibly due to wastewater dilution by high precipitation or enhanced biodegradation under high-temperature conditions. The anaerobic-anoxic-oxic process (A2O) with a membrane bioreactor exhibited higher OMP removal than other processes, such as A2O with sedimentation or the conventional activated sludge process (p-value < 0.05). Pesticides (DEET and atrazine), corrosion inhibitors (4-TTR and BTR), and metformin were selected as priority OMPs in toxicity-driven prioritization, whereas PFCs were determined as priority OMPs given their persistence and bioaccumulation properties. Overall, our results contribute to an important database on the occurrence, removal, and potential risks of OMPs in Korean WWTPs.
Landfilling is the most popular, economical and simple method for disposing a wide range of waste, which on the other hand beyond doubt serves as a reservoir of toxic substances. The leachate that are released from the contaminants affects the surrounding soil, groundwater and human health. Apart from harbouring contaminants from different sources like pharmaceuticals, medical, livestock, domestic wastes, heavy metals etc., landfill leachates (LFLs) facilitate the survival and dissemination of antibiotic resistance genes (ARGs). Due to the increased population which is directly proportional to the amount of waste produced requires more stringent protocol for the control and treatment of waste that is being dumped. The current review illustrates a panoramic view on the ARGs in the LFLs that includes the occurrence, source, treatment technologies and the scientometrics to analyse the research hotspots. The literature survey confirmed that only a limited number of studies have been reported so far for removal of ARGs in LFLs and still search for a cost effective, streamlined and ubiquitous method of treatment has never ceased. This could be due to the increase as well as the detrimental nature of ARGs, hence a few suitable methods already proposed for removal of ARGs from various sources and their challenges have also been discussed, to provide an insight of different methods to be investigated in LFLs.
Graphical Abstract
In activated sludge, the antibiotic resistance genes (ARGs) can be present either in the intracellular (iDNA) or extracellular DNA fraction (exDNA). Recent advances in the exDNA extraction methodology allow a better profiling of the pool of ARGs.
There is a growing concern about how microplastics could harm human health and ecosystems. The ability of microplastics to transport hydrophobic organic contaminants is well recognized. The potential for hydrophilic compounds, such as pharmaceuticals, to be adsorbed onto plastic substrates has been demonstrated by a recent study on these chemicals. Due to their extensive use, these substances are now pervasively in the environment and coexist with microplastics. Particular matrices’ physical and chemical characteristics control how plastics and pharmaceuticals are distributed and what happens when vectors transport them. This chapter’s objectives were to summarize and assess the various microplastic-pharmaceutical interactions and factors that affect the fate and mobility of hydrophilic chemicals, such as medications and personal care products (PPCPs), as well as their adsorption on microplastic surfaces. Kinetic microplastic investigations that have mostly focused on antibiotics have drawn some attention to a number of PPCP compounds, including steroidal hormones, antibacterial treatments, and nonsteroidal anti-inflammatory drugs (NSAIDs). The chapter also examines ecological factors that impact medicines, such as the sorption of PPCPs onto microplastics, such as pH, salinity, and dissolved organics. The ecotoxicological impacts of microplastics absorbed by PPCP on ecosystems and human health are also being investigated.
The article presents an analysis of the data of modern literature devoted to the study of early diagnosis of wound infection. It is well known that wound healing is a very complex and dynamic mechanism of wound re-epithelialization. At the same time, the normal microflora of the skin plays an important function for maintaining homeostasis and the formation of the skin. There are about 1000 species of microorganisms belonging to the normal flora of human skin and do not cause any harm to healthy people. At the same time, there are microorganisms that, when they enter the wound, lead to the development of infectious complications of wounds as a result of a violation of the integrity of the skin. They include both gram-positive (Staphylococcus aureus, Staphylococcus epidermidis) and gram-negative bacteria (Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa, Enterobacter spp., Morganella spp., etc.). Early detection of these microorganisms will contribute to timely and high-quality treatment of wound infection. Currently, there are certain conditions that limit the use of microbiological research methods used to establish a clinical diagnosis of wound infection (long duration, labor intensity, required level of qualification of specialists, etc.). This dictates the need to develop new, fast and easy-to-use methods for diagnosing wound infection. To this end, a group of researchers from Russia (Skolkovo Institute of Science and Technology) and the USA (University of Texas at Austin) have recently developed wearable sensors for the diagnosis of wound infection. These sensors can be embedded in wound dressings and are able to detect certain biomarkers indicating the presence of wound infection. Among these biomarkers, pH and uric acid are the most commonly used, but there are many others (lactic acid, oxygenation, inflammatory mediators, bacterial metabolites or the bacteria themselves). Currently, the development of microelectronics, the emergence of biochemical sensors, active microfluidics and painless microneedles have led to the creation of new generations of wearable biosensors that provide completely new opportunities in the fight against wound infection.
Microorganisms which are resistant to antibiotics are a global threat to the health of humans and animals. Wastewater treatment plants are known hotspots for the dissemination of antibiotic resistances. Therefore, novel methods for the inactivation of pathogens, and in particular antibiotic‐resistant microorganisms (ARM), are of increasing interest. An especially promising method could be a water treatment by physical plasma which provides charged particles, electric fields, UV‐radiation, and reactive species. The latter are foremost responsible for the antimicrobial properties of plasma. Thus, with plasma it might be possible to reduce the amount of ARM and to establish this technology as additional treatment stage for wastewater remediation. However, the impact of plasma on microorganisms beyond a mere inactivation was analyzed in more detail by a proteomic approach. Therefore, Escherichia coli GW‐AmxH19, isolated from hospital wastewater in Germany, was used. The bacterial solution was treated by a plasma discharge ignited between each of four pins and the liquid surface. The growth of E. coli and the pH‐value decreased during plasma treatment in comparison with the untreated control. Proteome and antibiotic resistance profile were analyzed. Concentrations of nitrite and nitrate were determined as long‐lived indicative products of a transient chemistry associated with reactive nitrogen species (RNS). Conversely, hydrogen peroxide served as indicator for reactive oxygen species (ROS). Proteome analyses revealed an oxidative stress response as a result of plasma‐generated RNS and ROS as well as a pH‐balancing reaction as key responses to plasma treatment. Both, the generation of reactive species and a decreased pH‐value is characteristic for plasma‐treated solutions. The plasma‐mediated changes of the proteome are discussed also in comparison with the Gram‐positive bacterium Bacillus subtilis . Furthermore, no effect of the plasma treatment, on the antibiotic resistance of E. coli , was determined under the chosen conditions. The knowledge about the physiological changes of ARM in response to plasma is of fundamental interest to understand the molecular basis for the inactivation. This will be important for the further development and implementation of plasma in wastewater remediation.
Pharmaceuticals are one of the emerging pollutants (EPs) in river waters across the world. Due to their toxic effects on aquatic organisms, they have drawn the global attention of the scientific community concerned with river ecosystems. This paper reviews the existing occurrence data for pharmaceutical pollutants (PPs) reported in river waters in some part of the world. Using algae, macroinvertebrates, and fish as biotic indicator groups in water to reflect river health condition (RHC), an attempt has been made to assess the ecological risk due to the presence of PPs.
After ascertaining the predicted no-effect concentration (PNEC) of PPs for selected groups of aquatic organisms, the risk quotient (RQ) is estimated based on their measured environmental concentration (MEC). When MEC> PNEC and RQ >1 for any of the biotic indicator, ecologically it is ‘high risk’ condition.
For river health assessment, physico-chemical characteristics of river water quality has been divided into three groups (dissolved oxygen related parameters, nutrients, and EPs, PPs represent EPs) in this study, based on their ease of measurement and frequency of monitoring. A framework for calculating separate indicator group score and the overall river health index has been developed. Colour-coded hexagonal pictorial forms representing indicator group condition and RHC provide a direct visible perception of the existing aquatic environment and a scientific basis for prioritization of corrective measures. The analyses indicate that many rivers across the world are under ‘high risk’ conditions due to PPs having MEC >PNEC and RQ >1. Algae is the most frequently affected group of biotic indicators, followed by MI and fish.
A review for selection of appropriate technology to reduce PPs from the wastewater streams has been summarised. It appears that constructed wetlands are at present the most suitable nature-based solutions, particularly for the developing economies of the world.
Surveillance methods of circulating antibiotic resistance genes (ARGs) are of utmost importance in order to tackle what has been described as one of the greatest threats to humanity in the 21 st century. In order to be effective, these methods have to be accurate, quickly deployable, and scalable. In this study, we compare metagenomic shotgun sequencing (TruSeq DNA sequencing) of wastewater samples with a state-of-the-art PCR-based method (Resistomap HT-qPCR) on four wastewater samples that were taken from hospital, industrial, urban and rural areas. ARGs that confer resistance to 11 antibiotic classes have been identified in these wastewater samples using both methods, with the most abundant observed classes of ARGs conferring resistance to aminoglycoside, multidrug-resistance (MDR), macrolide-lincosamide-streptogramin B (MLSB), tetracycline and beta-lactams. In comparing the methods, we observed a strong correlation of relative abundance of ARGs obtained by the two tested methods for the majority of antibiotic classes. Finally, we investigated the source of discrepancies in the results obtained by the two methods. This analysis revealed that false negatives were more likely to occur in qPCR due to mutated primer target sites, whereas ARGs with incomplete or low coverage were not detected by the sequencing method due to the parameters set in the bioinformatics pipeline. Indeed, despite the good correlation between the methods, each has its advantages and disadvantages which are also discussed here. By using both methods together, a more robust ARG surveillance program can be established. Overall, the work described here can aid wastewater treatment plants that plan on implementing an ARG surveillance program.
The widespread use of antibiotics since the 50s, generates a significant release of these molecules in the environment (excretion via urine and feces) which can be found at concentrations ranging from 1-100 ng/L in wastewater. Due to the high microbial biomass and the abundance of nutrients, wastewater treatment plants (WWTP) represent a suitable habitat for horizontal gene transfer. Because they occupy a key position between human activities and the environment, WWTP may play a major role in limiting the dissemination of antibiotic resistance genes, therefore contributing to the preservation The parameters which influence these transfers in wastewater treatment plants are still poorly known, especially because of methodological limitations. Therefore the aim of our study was to identify environmental factors affecting the stability and transfer of a mobile genetic element model, the plasmid pB10 in bacterial communities (biomass from wastewater treatment plants and river sediments) maintained in microcosms. So far, the transfer of resistance genes have been studied mainly with methods based on the cultivation of microorganisms on selective media that we know now they underestimate the observed phenomena. Also, an approach based on quantitative PCR was developed for detecting the release of a mobile DNA template from the host bacterium E. coli DH5α. Couples of designed primers/probes were very specific and have been developed by taking advantage of the mosaic structure of the bacterial genome. The proposed approach is based on the over time measurements of the number of plasmids pB10 and its bacterial host DH5α, where an increased ratio (pB10/DH5α) implies a release of the plasmid to the indigenous bacteria. This method was used to assess the impact of some environmental parameters on the release of DNA in complex microbial communities. Two groups of factors could be distinguished according to whether they influence the persistence of plasmid pB10 in communities in microcosms (oxygenation / mixing, addition of antibiotics at sub-inhibitory concentrations as amoxicillin and sulfamethoxazole frequently found in treatment plant) and / or they favor his release in bacterial communities (biofilms, sediments). Without inducing genes transfers, the antibiotics tested, even at sub-lethal concentrations, could participate in the dissemination of resistance genes by facilitating their persistence.
Keywords: Antibiotics, resistance genes, horizontal transfers, wastewater treatment plant, river sediments, quantitative PCR, pB10 plasmid.
To evaluate the antibiogram and antibiotic resistance genes of some Vibrio strains isolated from wastewater final effluents in a rural community of South Africa. V. vulnificus (18), V. metschnikovii (3), V. fluvialis (19) and V. parahaemolyticus (12) strains were isolated from final effluents of a wastewater treatment plant (WWTP) located in a rural community of South Africa. The disk diffusion method was used for the characterization of the antibiogram of the isolates. Polymerase chain reaction (PCR) was employed to evaluate the presence of established antibiotic resistance genes using specific primer sets.
The Vibrio strains showed the typical multidrug-resistance phenotype of an SXT element. They were resistant to sulfamethoxazole (Sul), trimethoprim (Tmp), cotrimoxazole (Cot), chloramphenicol (Chl), streptomycin (Str), ampicillin (Amp), tetracycline (Tet) nalidixic acid (Nal), and gentamicin (Gen). The antibiotic resistance genes detected includes dfr18 and dfrA1 for trimethoprim; floR, tetA, strB, sul2 for chloramphenicol, tetracycline, streptomycin and sulfamethoxazole respectively. Some of these genes were only recently described from clinical isolates, demonstrating genetic exchange between clinical and environmental Vibrio species.
These results demonstrate that final effluents from wastewater treatment plants are potential reservoirs of various antibiotics resistance genes. Moreover, detection of resistance genes in Vibrio strains obtained from the wastewater final effluents suggests that these resistance determinants might be further disseminated in habitats downstream of the sewage plant, thus constituting a serious health risk to the communities reliant on the receiving waterbodies.
A group of antibiotic resistance genes (ARGs) (blaTEM, blaCTX-M-1, mecA, armA, qnrA, and qnrS) were analyzed by real-time quantitative PCR (qPCR) in bacteriophage DNA isolated from feces from 80 healthy humans. Seventy-seven
percent of the samples were positive in phage DNA for one or more ARGs. blaTEM, qnrA, and, blaCTX-M-1 were the most abundant, and armA, qnrS, and mecA were less prevalent. Free bacteriophages carrying ARGs may contribute to the mobilization of ARGs in intra- and extraintestinal
environments.
Effectiveness of conventional chlorine and ozone disinfection on reduction of antibiotic resistance was compared with less commonly applied heterogeneous photocatalytic process. For this purpose plasmid DNA isolated from a multi-resistant Escherichia coli (E. coli) HB101 was treated in two different concentrations with the three oxidation processes. Oxidative damage on the plasmid DNA was analyzed with gel electrophoresis by comparing the extent of conformational changes in the DNA structure. The effectiveness of the applied oxidant in reducing the risk of resistance transfer was also evaluated by comparing the ability of treated plasmid DNA to transform competent cells. Chlorine did not affect plasmid DNA structure at the studied doses, while ozone and photocatalytic treatment resulted in conformational changes and the damage increased with increasing oxidant doses. Transformation experiments confirmed a similar trend. Chlorine did not affect the transformability and the cell counts of competent cells transformed with chlorine treated plasmid DNA were similar to those transformed by non-treated plasmid DNA in the control experiments.
The evolutionary events in organisms can be tracked to the transfer of genetic material. The inheritance of genetic material among closely related organisms is a slow evolutionary process. On the other hand, the movement of genes among distantly related species can account for rapid evolution. The later process has been quite evident in the appearance of antibiotic resistance genes among human and animal pathogens. Phylogenetic trees based on such genes and those involved in metabolic activities reflect the incongruencies in comparison to the 16S rDNA gene, generally used for taxonomic relationships. Such discrepancies in gene inheritance have been termed as horizontal gene transfer (HGT) events. In the post-genomic era, the explosion of known sequences through large-scale sequencing projects has unraveled the weakness of traditional 16S rDNA gene tree based evolutionary model. Various methods to scrutinize HGT events include atypical composition, abnormal sequence similarity, anomalous phylogenetic distribution, unusual phyletic patterns, etc. Since HGT generates greater genetic diversity, it is likely to increase resource use and ecosystem resilience.
The assembling of bacterial communities in conventional activated sludge (CAS) bioreactors was thought, until recently, to be chaotic and mostly unpredictable. Studies done over the last decade have shown that specific, and often, predictable random and non-random factors could be responsible for that process. These studies have also motivated a "structure-function" paradigm that is yet to be resolved. Thus, elucidating the factors that affect community assembly in the bioreactors is necessary for predicting fluctuations in community structure and function. For this study activated sludge samples were collected during a one-year period from two geographically distant CAS bioreactors of different size. Combining community fingerprinting analysis and operational parameters data with a robust statistical analysis, we aimed to identify relevant links between system performance and bacterial community diversity and dynamics. In addition to revealing a significant β-diversity between the bioreactors' communities, results showed that the largest bioreactor had a less dynamic but more efficient and diverse bacterial community throughout the study. The statistical analysis also suggests that deterministic factors, as opposed to stochastic factors, may have a bigger impact on the community structure in the largest bioreactor. Furthermore, the community seems to rely mainly on mechanisms of resistance and functional redundancy to maintain functional stability. We suggest that the ecological theories behind the Island Biogeography model and the species-area relationship were appropriate to predict the assembly of bacterial communities in these CAS bioreactors. These results are of great importance for engineers and ecologists as they reveal critical aspects of CAS systems that could be applied towards improving bioreactor design and operation.
The activated sludge process is one of the biological treatment methods used in many countries to reduce the high levels of organic and mineral pollutants and pathogenic micro-organisms present in wastewater. The present work was undertaken to study the dynamic and antibiotic-resistance of faecal coliforms (FC) in the activated sludge system of Beni Mellal. This work has also as objective the study of the survival of FC, protozoan cysts, helminth eggs and FC antibiotic resistance in the sludge dehydrated in drying beds in order to know if the agricultural usage of sludge presents any problems to public health. The activated sludge treatment of Beni Mellal resulted in an average reduction of FC and faecal streptococci of 90.75 and 91.06%, respectively. The overall resistance (resistance to at least one antibiotic) of 111 FC strains isolated from the system was 72.07%. This treatment system did not increase the incidence of FC antibiotic resistance in treated wastewaters. The antibiotic resistance of FC was found to be similar in both raw (71.05%) and treated sewage (77.77%). High levels of antibiotic resistance were towards streptomycin (54.05%), ampicillin (42.34%), amoxicillin (42.34%) and amoxicillin–clavulanic acid (31.53%). The treatment of sludge in drying beds appeared to be efficient in eliminating pathogenic micro-organisms: FC, protozoan cysts and helminth eggs. Moreover, the FC antibiotic resistance did not change over time in sludge-drying bed. According to the standard norms, agricultural utilization of this sludge cannot be excluded. However, it is important to study in the receptor environment the survival and the behaviour of antibiotic-resistant FC present in sludge and water.
The spread of antibiotic-resistant bacteria is a growing problem and a public health issue. In recent decades, various genetic mechanisms involved in the spread of resistance genes among bacteria have been identified. Integrons – genetic elements that acquire, exchange, and express genes embedded within gene cassettes (GC) – are one of these mechanisms. Integrons are widely distributed, especially in Gram-negative bacteria; they are carried by mobile genetic elements, plasmids, and transposons, which promote their spread within bacterial communities. Initially studied mainly in the clinical setting for their involvement in antibiotic resistance, their role in the environment is now an increasing focus of attention. The aim of this review is to provide an in-depth analysis of recent studies of antibiotic-resistance integrons in the environment, highlighting their potential involvement in antibiotic-resistance outside the clinical context. We will focus particularly on the impact of human activities (agriculture, industries, wastewater treatment, etc.).
At present, very little is known about the fate and persistence of multiresistant bacteria (MRB) and their resistance genes in natural aquatic environments. Treated, but partly also untreated sewage of the city of Lausanne, Switzerland is discharged into Vidy Bay (Lake Geneva) resulting in high levels of contamination in this part of the lake. In the present work we have studied the prevalence of MRB and resistance genes in the wastewater stream of Lausanne. Samples from hospital and municipal raw sewage, treated effluent from Lausanne’s wastewater treatment plant (WTP) as well as lake water and sediment samples obtained close to the WTP outlet pipe and a remote site close to a drinking water pump were evaluated for the prevalence of MRB. Selected isolates were identified (16S rRNA gene fragment sequencing) and characterized with regards to further resistances, resistance genes, and plasmids. Mostly, studies investigating this issue have relied on cultivation-based approaches. However, the limitations of these tools are well known, in particular for environmental microbial communities, and cultivation-independent molecular tools should be applied in parallel in order to take non-culturable organisms into account. Here we directly quantified the sulfonamide resistance genes sul1 and sul2 from environmental DNA extracts using TaqMan real-time quantitative PCR. Hospital sewage contained the highest load of MRB and antibiotic resistance genes (ARGs). Wastewater treatment reduced the total bacterial load up to 78% but evidence for selection of extremely multiresistant strains and accumulation of resistance genes was observed. Our data clearly indicated pollution of sediments with ARGs in the vicinity of the WTP outlet. The potential of lakes as reservoirs of MRB and potential risks are discussed.
The prospects for using bacterial DNA as an intrinsic probe for HOCl and secondary oxidants/chlorinating agents associated with it has been evaluated using both in vitro and in vivo studies. Single-strand and double-strand breaks occurred in bare plasmid DNA that had been exposed to high levels of HOCl, although these reactions were very inefficient compared to polynucleotide chain cleavage caused by the OH-generating reagent, peroxynitrite. Plasmid nicking was not increased when intact Escherichia coli were exposed to HOCl; rather, the amount of recoverable plasmid diminished in a dose-dependent manner. At concentration levels of HOCl exceeding lethal doses, genomic bacterial DNA underwent extensive fragmentation and the amount of precipitable DNA–protein complexes increased several-fold. The 5-chlorocytosine content of plasmid and genomic DNA isolated from HOCl-exposed E. coli was also slightly elevated above controls, as measured by mass spectrometry of the deaminated product, 5-chlorouracil. However, the yields were not dose-dependent over the bactericidal concentration range. Genomic DNA recovered from E. coli that had been subjected to phagocytosis by human neutrophils occasionally showed small increases in 5-chlorocytosine content when compared to analogous cellular reactions where myeloperoxidase activity was inhibited by azide ion. Overall, the amount of isolable 5-chlorouracil from the HOCl-exposed bacterial cells was far less than the damage manifested in polynucleotide bond cleavage and cross-linking.
The environment, and especially freshwater, constitutes a reactor where the evolution and the rise of new resistances occur. In water bodies such as waste water effluents, lakes, and rivers or streams, bacteria from different sources, e.g., urban, industrial, and agricultural waste, probably selected by intensive antibiotic usage, are collected and mixed with environmental species. This may cause two effects on the development of antibiotic resistances: first, the contamination of water by antibiotics or other pollutants lead to the rise of resistances due to selection processes, for instance, of strains over-expressing broad range defensive mechanisms, such as efflux pumps. Second, since environmental species are provided with intrinsic antibiotic resistance mechanisms, the mixture with allochthonous species is likely to cause genetic exchange. In this context, the role of phages and integrons for the spread of resistance mechanisms appears significant. Allochthonous species could acquire new resistances from environmental donors and introduce the newly acquired resistance mechanisms into the clinics. This is illustrated by clinically relevant resistance mechanisms, such as the fluoroquinolones resistance genes qnr. Freshwater appears to play an important role in the emergence and in the spread of antibiotic resistances, highlighting the necessity for strategies of water quality improvement. We assume that further knowledge is needed to better understand the role of the environment as reservoir of antibiotic resistances and to elucidate the link between environmental pollution by anthropogenic pressures and emergence of antibiotic resistances. Only an integrated vision of these two aspects can provide elements to assess the risk of spread of antibiotic resistances via water bodies and suggest, in this context, solutions for this urgent health issue.
Activated sludge (AS) contains highly complex microbial communities. In this study, PCR-based 454 pyrosequencing was applied to investigate the bacterial communities of AS samples from 14 sewage treatment plants of Asia (mainland China, Hong Kong, and Singapore), and North America (Canada and the United States). A total of 259 K effective sequences of 16S rRNA gene V4 region were obtained from these AS samples. These sequences revealed huge amount of operational taxonomic units (OTUs) in AS, that is, 1183-3567 OTUs in a sludge sample, at 3% cutoff level and sequencing depth of 16,489 sequences. Clear geographical differences among the AS samples from Asia and North America were revealed by (1) cluster analyses based on abundances of OTUs or the genus/family/order assigned by Ribosomal Database Project (RDP) and (2) the principal coordinate analyses based on OTUs abundances, RDP taxa abundances and UniFrac of OTUs and their distances. In addition to certain unique bacterial populations in each AS sample, some genera were dominant, and core populations shared by multiple samples, including two commonly reported genera of Zoogloea and Dechloromonas, three genera not frequently reported (i.e., Prosthecobacter, Caldilinea and Tricoccus) and three genera not well described so far (i.e., Gp4 and Gp6 in Acidobacteria and Subdivision3 genera incertae sedis of Verrucomicrobia). Pyrosequencing analyses of multiple AS samples in this study also revealed the minority populations that are hard to be explored by traditional molecular methods and showed that a large proportion of sequences could not be assigned to taxonomic affiliations even at the phylum/class levels.
The overuse or misuse of antibiotics has accelerated antibiotic resistance, creating a major challenge for the public health in the world. Sewage treatment plants (STPs) are considered as important reservoirs for antibiotic resistance genes (ARGs) and activated sludge characterized with high microbial density and diversity facilitates ARG horizontal gene transfer (HGT) via mobile genetic elements (MGEs). However, little is known regarding the pool of ARGs and MGEs in sludge microbiome. In this study, the transposon aided capture (TRACA) system was employed to isolate novel plasmids from activated sludge of one STP in Hong Kong, China. We also used Illumina Hiseq 2000 high-throughput sequencing and metagenomics analysis to investigate the plasmid metagenome. Two novel plasmids were acquired from the sludge microbiome by using TRACA system and one novel plasmid was identified through metagenomics analysis. Our results revealed high levels of various ARGs as well as MGEs for HGT, including integrons, transposons and plasmids. The application of the TRACA system to isolate novel plasmids from the environmental metagenome, coupled with subsequent high-throughput sequencing and metagenomic analysis, highlighted the prevalence of ARGs and MGEs in microbial community of STPs.
Antibiotics were one of the great discoveries of the 20th century. However, resistance appeared even in the earliest years of the antibiotic era. Antibiotic resistance continues to become worse, despite the ever-increasing resources devoted to combat the problem. One of the most important factors in the development of resistance to antibiotics is the remarkable ability of bacteria to share genetic resources via Lateral Gene Transfer (LGT). LGT occurs on a global scale, such that in theory, any gene in any organism anywhere in the microbial biosphere might be mobilized and spread. With sufficiently strong selection, any gene may spread to a point where it establishes a global presence. From an antibiotic resistance perspective, this means that a resistance phenotype can appear in a diverse range of infections around the globe nearly simultaneously. We discuss the forces and agents that make this LGT possible and argue that the problem of resistance can ultimately only be managed by understanding the problem from a broad ecological and evolutionary perspective. We also argue that human activities are exacerbating the problem by increasing the tempo of LGT and bacterial evolution for many traits that are important to humans.
Plasmid-mediated dissemination of antibiotic resistance genes is widely recognized to take place in many environmental compartments but remains difficult to study in a global perspective because of the complexity of the environmental matrices considered and the lack of exhaustive tools. In this report, we used a molecular approach based on quantitative PCR to monitor the fate of the antibiotic resistance plasmid pB10 and its donor host in microbial communities collected from various wastewater treatment plant (WWTP) sludges and maintained in microcosms under different conditions. In aerated activated sludge microcosms, pB10 did not persist because of an apparent loss of the donor bacteria. The persistence of the donor bacteria noticeably increased in non-aerated activated sludge microcosms or after amending antibiotics (sulfamethoxazole or amoxicillin) at sub-inhibitory concentrations, but the persistence of the donor bacteria did not stimulate the dissemination of pB10. The dissemination of the plasmid appeared as an increasing plasmid to donor ratio in microcosm setups with microbial communities collected in anaerobic digesters or the spatially organized communities from fixed biofilm reactors. As a whole, the data collected suggest that some WWTP processes, more than others, may sustain microbial communities that efficiently support the dissemination of the multiple-antibiotic-resistance plasmid pB10.
The utilization of antibiotics to control infectious diseases is one of the biggest advances in human and veterinary health care. However, the generalized use of antibiotics has been accompanied by a worrisome increase in the prevalence of antibiotic-resistant bacteria. This evidence motivated numerous studies on the diversity and distribution of antibiotic-resistant bacteria and resistance genetic determinants not only in clinic but also in different environmental compartments. Given the particular importance that the anthropic water cycle (waste water/surface water/drinking water) may have in the development and dissemination of antibiotic-resistant organisms, this chapter aims at summarizing the recent advances in this area. Sections 1 and 2 are an Introduction to antibiotic resistance, summarizing some mechanisms and modes of resistance acquisition. In Sect. 3, the contribution of the environmental pollution and other anthropic pressures for antibiotic resistance evolution is discussed. The use of different methodologies and the limitations to achieve general conclusions on the characterization and quantification of antibiotic resistance in aquatic environments are examined in Sects. 4 and 7. Sections 5–7 summarize recent evidences on the widespread distribution of antibiotic resistance in different compartments of the anthropic water cycle. The scarcity of studies giving evidences on the direct effect of anthropic pressures on antibiotic resistance acquisition and maintenance in treated waste/drinking waters is highlighted. The contribution of bacterial community rearrangement, imposed by water treatment processes, on the increase of antibiotic resistance is discussed.
The streptomycin/spectinomycin resistance determinant of the 29-kb plasmid pCG4 from Corynebacterium glutamicum was found to be a part of a typical class 1 integron. The sequence analysis revealed that the integron (designated InCg) identified in this Gram-positive bacterium is almost identical to the integron InC present on the plasmid pSA1700 from the Gram-negative bacterium Pseudomonas aeruginosa. Differences in only two base pairs were found in the 3.8-kb sequence. One base substitution (G→C) is present in the streptomycin/spectinomycin resistance determinant which is thus identical to the aadA2a gene from the integron In6 of the broad-host-range plasmid pSa. The other one (C→G) is present in the extended −10 region of the integron promoter involved in expression of the antibiotic resistance gene. It was shown that this novel version of the integron promoter displays five times higher activity in both C. glutamicum and Escherichia coli than the original one.
Application of ecological diversity statistics in microbial ecologyJennifer B Hughes1Brendan J. M. Bohannan2(1)Dept. of Ecology and Evolutionary Biology, Brown University, Providence, RI, USA(2)Dept. of Biological Sciences, Stanford University, Stanford, CA, USA IntroductionBiological diversity, or biodiversity, is the variation of organisms at all levels of biological organization, from genes to communities. A fundamental goal of biology is to understand what generates and regulates this diversity. Ecologists, in particular, are often interested in what factors regulate community diversity across temporal and spatial scales and the consequences of this diversity for ecosystem processes. Thus, a key component of many ecological studies is the documentation and comparison of biodiversity patterns in the environment. Ecologists studying the diversity of macroorganisms have developed a range of diversity indices and statistics to document and anal ...
Role of Sewage Treatment Plants in the Transfer of Resistance GenesFate of Antibiotic-Resistant Bacteria and their Resistance DeterminantsBioeffectivities of Low Concentrated AntibioticsReduction of the Dissemination of Antibiotic-Resistant BacteriaStrategy to Evaluate the Resistance Situation and its Biological RiskAcknowledgmentReferences
Integrons are bacterial genetic elements able to capture and express genes embedded within cassettes. Integrons are widely distributed notably among Gram negative bacteria. Resistant integrons are very involved in medical bacteriology due to their ability in recruiting and expressing genes encoding resistance to antibiotics. The aim of this review is to inventory recent works bringing new elements in the knowledge of resistant integrons: both on the mechanistic side with better understanding of their precise function and on the epidemiological aspect with a better appreciation of their impact in the living world.
This chapter examines various concepts in community ecology that are difficult to observe in natural systems and examines some laboratory studies that document the production of ecological structure in multi-species systems. Two aspects of ecology have been explored where laboratory studies are crucial for the understanding of ecological phenomena. The first aspect is parameter (and measurement) oriented—what must be measured and how it should be done to make observations in ecological studies. Secondly, the interactive nature of ecological phenomena and the inability to reduce these processes to their constituent components have been explored. These two discussions implicitly address the relationship between reductionism and holism. It has been argued that laboratory studies are not only valid investigative tools, but may also better capture the essential dynamics of ecological systems compared with field studies. Through exploring the use of microcosms, the dynamic reality of ecological systems has been illustrated, and the systems in which explicit tests of these realities are possible have been examined. The chapter contrasts the laboratory approach with the field and model approaches to identify the prescriptive role for laboratory microcosms in ecological research.
This general review paper with 70 references explains the various aspects of membrane technology for water disinfection. Membrane capabilities for removing bacteria, viruses and other microorganisms are discussed and advantages are shown. Fouling, which is the main problem in membrane based water treatment and its control are explained. Finally the advantages and disadvantages of using membrane for water treatment are summarized.
Human and veterinary drugs are continually being released in the environment mainly as a result of manufacturing processes, disposal of unused or expired products, and excreta. Because of their physical and chemical properties, many of these substances or their bioactive metabolites end up in soils and sediments, where they can accumulate and induce adverse effects in terrestrial or aquatic organisms. Among these effects, bacterial resistance is increasingly observed and is caused by the extensive use of antibiotics in animal and fish farming and the growing practice of adding manure and sewage sludge to agricultural fields, which is of particular concern. Literature on the environmental analysis and occurrence of drugs has addressed a very small percentage of these compounds, so very little information is available about the fate and the potential effects of drugs in the environment. This article presents an overview of recent developments in the determination of veterinary and human drugs in solid environmental matrices, including soil, sediment and sludge. The analysis of pharmaceuticals in the such samples has always been carried out by high-performance liquid chromatography coupled to ultraviolet detection, and, to a lesser extent, to mass spectrometry and fluorescence detection. In most cases, sample pretreatment includes extraction of the solid sample and further purification of the extract by solid phase extraction with C18 sorbents. In addition to analytical articles, this overview includes papers concerning usage of drugs, as well as sources, fate, persistence, and effects of pharmaceuticals in solid environmental matrices.
This study was based on the hypothesis that the occurrence of antimicrobial residues and antibiotic resistant bacteria in the sewage could be correlated with the structure and composition of the bacterial community and the antibiotic resistance loads of the final effluent. Raw and treated wastewater composite samples were collected from an urban treatment plant over 14 sampling dates. Samples were characterized for the i) occurrence of tetracyclines, penicillins, sulfonamides, quinolones, triclosan, arsenic, cadmium, lead, chromium and mercury; ii) antibiotic resistance percentages for tetracycline, sulfamethoxazole, ciprofloxacin and amoxicillin and iii) 16S rRNA gene-DGGE patterns. The data of corresponding samples, taking into account the hydraulic residence time, was analyzed using multivariate analysis. Variations on the bacterial community structure of the final effluent were significantly correlated with the occurrence of tetracyclines, penicillins, sulfonamides, quinolones and triclosan in the raw inflow. Members of the class Epsilonproteobacteria presented positive correlations with those antimicrobials, whereas negative correlations were observed with Beta and Gammaproteobacteria and Firmicutes. Antibiotic resistance percentages presented different trends of variation in heterotrophs/enterobacteria and in enterococci, varied over time and after wastewater treatment. Antibiotic resistance was positively correlated with the occurrence of tetracyclines residues and high temperature. A relationship between antibiotic residues, bacterial community structure and composition and antibiotic resistance is demonstrated. Further studies, involving more wastewater treatment plants may help to elucidate this complex relationship.
This bench-scale study investigated whether strains of Escherichia coli that are resistant to two common types of antibiotics, ampicillin and trimethoprim, possess increased resistance to two common disinfectants in water and wastewater treatment, free chlorine and ultraviolet disinfection, relative to an antibiotic-sensitive strain of E. coli isolated from sewage sludge. Trimethoprim-resistant E. coli was slightly more resistant to chlorine than the antibiotic-sensitive isolate and the ampicillin-resistant E. coli under the study conditions (95% confidence), however this difference would not be important under normal chlorination conditions applied in practice. There were no statistically significant differences between the ultraviolet dose-response profiles of the antibiotic-resistant and antibiotic-sensitive E. coli strains over the ultraviolet dose range tested.La présente étude à l'échelle du laboratoire examinait si des souches d'Escherichia coli qui sont résistantes à deux types fréquents d'antibiotiques, l'ampicilline et la triméthoprime, possèdent une résistance accrue à deux désinfectants fréquents dans le traitement des eaux et des eaux usées, le chlore libre et la désinfection aux ultraviolets, par rapport à une souche E. coli sensible aux antibiotiques qui provient de boues d'épuration. E. coli résistante à la triméthoprime a été légèrement plus résistante au chlore que l'isolat sensible aux antibiotiques et que E. coli résistante à l'ampicilline sous les conditions de l'étude (confiance de 95 %); toutefois, cette différence ne serait pas significative sous les conditions de chloration normales utilisées dans la pratique. Il n'existait aucune différence statistique importante entre les profils de réponse aux doses d'ultraviolets des souches E. coli résistantes aux antibiotiques et celles sensibles aux antibiotiques pour la plage de doses d'ultraviolets utilisée dans les essais.
In situ transfer of a self-transmissible, antibiotic-multiresistant plasmid RP4 from a laboratory Escherichia coli strain C600 to indigenous activated sludge bacteria was investigated using filter mating. The transfer frequency of RP4 from the donor E. coli to the bacteria that was sampled from two wastewater treatment plants was 5.1x10(-2) to 7.5x10(-1) and 4.6x10(-3) to 1.3x10(-2)/potential recipient. The isolated transconjugants showed resistance to Ap, Km, and Tc and the presence of a plasmid with a similar size to RP4. The traG gene on RP4 was also detected from all transconjugants. Reverse-transfer experiments from the transconjugants to E. coli HB101 indicated that RP4 maintained self-transmissibility in the transconjugants. The transconjugant strains were dominant bacteria in activated sludge including Pseudomonas fluorescens, P. putida, and Ochrobactrum anthropi and minor populations of enteric bacterial strains including Citrobacter freundii, E. coli, Enterobacter cloacae, E. asburiae, and Klebsiella pneumoniae ssp. pneumoniae. The transconjugant strains K. pneumoniae ssp. pneumonia, E. cloacae, and E. asburiae had several naturally occurring plasmids. These results suggest that in situ transfer of plasmids and the exchange of antibiotic-resistant genes can occur between released and indigenous bacteria in activated sludge.
This work investigated the application of a solar driven advanced oxidation process (solar photo-Fenton), for the degradation of antibiotics at low concentration level (μg L(-1)) in secondary treated domestic effluents at a pilot-scale. The examined antibiotics were ofloxacin (OFX) and trimethoprim (TMP). A compound parabolic collector (CPC) pilot plant was used for the photocatalytic experiments. The process was mainly evaluated by a fast and reliable analytical method based on a UPLC-MS/MS system. Solar photo-Fenton process using low iron and hydrogen peroxide doses ([Fe(2+)](0) = 5 mg L(-1); [H(2)O(2)](0) = 75 mg L(-1)) was proved to be an efficient method for the elimination of these compounds with relatively high degradation rates. The photocatalytic degradation of OFX and TMP with the solar photo-Fenton process followed apparent first-order kinetics. A modification of the first-order kinetic expression was proposed and has been successfully used to explain the degradation kinetics of the compounds during the solar photo-Fenton treatment. The results demonstrated the capacity of the applied advanced process to reduce the initial wastewater toxicity against the examined plant species (Sorghum saccharatum, Lepidium sativum, Sinapis alba) and the water flea Daphnia magna. The phytotoxicity of the treated samples, expressed as root growth inhibition, was higher compared to that observed on the inhibition of seed germination. Enterococci, including those resistant to OFX and TMP, were completely eliminated at the end of the treatment. The total cost of the full scale unit for the treatment of 150 m(3) day(-1) of secondary wastewater effluent was found to be 0.85 € m(-3).
A GFPmut3b-tagged derivative of broad host-range plasmid RP4 was used to monitor the conjugative transfer of the plasmid from a Pseudomonas putida donor strain to indigenous bacteria in activated sludge. Transfer frequencies were determined to be in the range of 4×10−6 to 1×10−5 transconjugants per recipient. In situ hybridisation with fluorescently labeled, rRNA-targeted oligonucleotides was used to phylogenetically affiliate the bacteria that had received the plasmid.
The distribution of tetracycline resistance determinants Tet A–E was studied by PCR in 40 tetracycline-resistant Salmonella enterica serotype Hadar (S. hadar) isolates collected from human patients in 1996 and 1997, as well as in the microbial community originating from activated sludges of hospital and urban wastewater treatment facilities. A fast DNA extraction and purification method from activated sludges was used to provide amplifiable DNA. The method is based on the direct lysis of bacteria improved by bead-beating followed by DNA purification on polyvinylpolypyrrolidone spin columns to remove PCR inhibitors. The purified DNAs from salmonellae and activated sludges were characterized for the presence of tetracycline determinants with specific primer pairs designed on the basis of published sequences. The Tet A determinant was present in all clinical isolates and DNAs extracted from the bacterial community of the selected activated sludges. The Tet C determinant was identified in only one of the 40 clinical isolates and in six of the seven environmental samples. No signal was detected for Tet B, D and E determinants. This study revealed a high and stable prevalence of the Tet A determinant in both salmonellae clinical isolates and the microbial community of activated sludges from hospital and urban wastewater treatment facilities over a 2-year period.
Plasmid transfer was investigated in microbial populations associated with different types of surfaces. The general strategy behind these investigations was to label the transferable plasmid with a gene encoding a fluorescent protein in order to make it a transfer reporter. This was achieved by fusing the reporter gene with a lac promoter expression cassette and combining this with a donor cell-associated lacI repressor cassette. After construction of a range of strains and plasmids with combinations of genes expressing fluorescent proteins from constitutive (cell tagging) or regulated promoters (transfer reporters) it was thus possible to detect transfer events in situ and correlate these with either the location of donor and recipient cells or with the growth activity of the cells. In some cases, expression of unstable Gfp from a growth-controlled promoter, rrnB from Escherichia coli, was used to monitor bacterial growth activity in situ. Differential tagging of mobilizing and mobilizable plasmids with different genes encoding fluorescent proteins with varying emission wavelengths allowed in situ detection of plasmid mobilization and detection of retro-transfer on agar surfaces. The obtained data show that the several different types of fluorescent reporters, which are now available, allow more informative in situ investigations of horizontal gene transfer to be carried out, and by combining these genes with various expression systems it is possible to simultaneously monitor donor/recipient positioning, cellular activity and appearance of transconjugants.
An integron is a genetic unit that includes the determinants of the components of a site-specific recombination system capable of capturing and mobilizing genes that are contained in mobile elements called gene cassettes. An integron also provides a promoter for expression of the cassette genes, and integrons thus act both as natural cloning systems and as expression vectors. The essential components of an integron are an int gene encoding a site-specific recombinase belonging to the integrase family, an adjacent site, attI, that is recognized by the integrase and is the receptor site for the cassettes, and a promoter suitably oriented for expression of the cassette-encoded genes. The cassettes are mobile elements that include a gene (most commonly an antibiotic-resistance gene) and an integrase-specific recombination site that is a member of a family of sites known as 59-base elements. Cassettes can exist either free in a circularized form or integrated at the attI site, and only when integrated is a cassette formally part of an integron. A single site-specific recombination event involving the integron-associated attI site and a cassette-associated 59-base element leads to insertion of a free circular cassette into a recipient integron. Multiple cassette insertions can occur, and integrons containing several cassettes have been found in the wild. The integrase also catalyses excisive recombination events that can lead to loss of cassettes from an itegron and generate free circular cassettes. Due to their ability to acquire new genes, integrons have a clear role in the evolution of the genomes of the plasmids and transposons that contain them.
Naturally-occurring inhibitory compounds are a major concern during qPCR and RT-qPCR analysis of environmental samples, particularly large volume water samples. Here, a standardized method for measuring and mitigating sample inhibition in environmental water concentrates is described. Specifically, the method 1) employs a commercially available standard RNA control; 2) defines inhibition by the change in the quantification cycle (C(q)) of the standard RNA control when added to the sample concentrate; and 3) calculates a dilution factor using a mathematical formula applied to the change in C(q) to indicate the specific volume of nuclease-free water necessary to dilute the effect of inhibitors. The standardized inhibition method was applied to 3,193 large-volume water (surface, groundwater, drinking water, agricultural runoff, sewage) concentrates of which 1,074 (34%) were inhibited. Inhibition level was not related to sample volume. Samples collected from the same locations over a one to two year period had widely variable inhibition levels. The proportion of samples that could have been reported as false negatives if inhibition had not been mitigated was between 0.3% and 71%, depending on water source. These findings emphasize the importance of measuring and mitigating inhibition when reporting qPCR results for viral pathogens in environmental waters to minimize the likelihood of reporting false negatives and under-quantifying virus concentration.
Antibiotic resistance genes (ARGs), in association with antibiotic resistant bacteria (ARB), have been identified as widespread contaminants of treated drinking waters and wastewaters. As a consequence, concerns have been raised that ARB or ARG transport between aquatic compartments may enhance the spread of antibiotic resistance amongst non-resistant bacterial communities by means of horizontal gene transfer processes. Most often, discussion of horizontal gene transfer focuses on the probable role of conjugative plasmid or transposon exchange, which requires live ARB donor cells. Conventional water and wastewater disinfection processes generally provide highly effective means for mitigating the transport of live ARB; thereby minimizing risks of conjugative gene transfer. However, even if ARB present in a treated water are fully inactivated during a disinfection process, the possibility remains that intact remnants of DNA contained within the resulting cell debris could still confer resistance genotypes to downstream bacterial populations by means of natural transformation and/or transduction, which do not require live donor cells. Thus, a systematic evaluation of the capability of common disinfection technologies to ensure the destruction of bacterial DNA, in addition to pathogen inactivation, seems warranted. With that objective in mind, this review seeks to provide a concise introduction to the significance of ARB and ARG occurrence in environmental systems, coupled with a review of the role that commonly used water and wastewater disinfection processes may play in minimizing ARG transport and dissemination.
The effect of solar radiation on the inactivation of multidrug resistant Escherichia coli (MDR) strains selected from an urban wastewater treatment plant (UWWTP) effluent and the change of their resistance to a mixture of three antibiotics (evaluated in terms of minimum inhibit concentration (MIC)) in wastewater polluted stream were investigated. The solar photodegradation of the mixture of the three target antibiotics (amoxicillin (AMX), ciprofloxacin (CPX), and sulfamethoxazole (SMZ)) was also evaluated. Additionally, since UWWTP effluents are possible sources of antibiotics and antibiotic resistant bacteria, the disinfection by conventional chlorination process of the UWWTP effluent inoculated with MDR strains was investigated too. Solar radiation poorly affected the inactivation of the two selected antibiotic resistant E. coli strains (40 and 60% after 180 min irradiation). Moreover, solar radiation did not affect strain resistance to AMX (MIC>256 μg/mL) and SMZ (MIC>1024 μg/mL), but affected resistance of the lower resistance strain to CPX (MIC decreased by 33% but only after 180 min of irradiation). Chlorination of wastewater sample strongly decreased the number of the two selected antibiotic resistant E. coli strains (99.667 and 99.999%), after 60 min of contact time at 2.0 mg/L initial chlorine concentration, but the resistance of survived colonies to antibiotics was unchanged. Finally, the solar photodegradation rate of the antibiotic mixture (1mg/L initial concentration respectively) resulted in the following order (half-life time): CPX (t(1/2)=24 min)<AMX (t(1/2)=99 min)<SMZ (t(1/2)=577 min). Accordingly, the risk of the development of resistance to SMZ in surface water is significantly higher compared to CPX and AMX.
Transferable (R+) and non-transferable (R−) resistance of coliform bacteria in settled sewage and after biofiltration, secondary sedimentation, chlorination and sand filtration was investigated. The percentage of R− coliforms resistant to ampicillin (A), chloramphenicol (C) or streptomycin (S), but not kanamycin (K) or tetracycline (T), was slightly reduced through the purification system. On the average the percentage of R+ coliforms resistant to one or more of the same drugs was reduced by about 50%. This reduction was mainly accomplished by biofiltration and sand filtration. The ratio of transferable to non-transferable resistance for A, C, K or T increased during secondary sedimentation and chlorination. R factor transfer in water may explain these findings. Rapid passage over stony surfaces in biological and sand filters is unfavourable for conjugation and may damage sex pilli while R factor transfer can be expected under the relatively stagnant conditions in sedimentation and chlorination tanks. The incidence of R factors conferring resistance to all five drugs simultaneously increased through the purification system. These R factors are probably large and may carry markers which determine resistance to sewage purification processes. Between 30 and 40% of all R factors mediated resistance to at least four of the five drugs studied. The incidence of Escherichia coli I among R+ coliforms varied but did not exceed 50%. The finding that conventional sewage purification has a limited effect on the incidence of drug resistance in bacteria supports the view that sewage should be treated by more advanced methods prior to discharge into the environment.
This work compares antibiotic resistance of 870 E. coli strains isolated from domestic raw sewage, in the effluent from aerobic lagoons and activated sludge plants. Efficiency of both treatment systems in removing faecal coliforms (FC) has also been evaluated. Reduction of FC was greater in the aerobic lagoon (99.99% in summer) than in the activated sludge system (91.30% in summer). Although the aerobic lagoon is efficient in removing faecal coliforms in comparisons with the activated sludge plant, it discharges these bacteria into receptor environments in larger (30.103 CFU/100 ml) or smaller quantities (9–102 CFU/100 ml) according to season. Among these FC, E. coli which was relatively high (on average 80%) constitutes a health problem when it is resistant to antibiotics. As for percentage of global antibiotic-resistance (evaluated by Gallery ATBG-method), the E. coli strains isolated from the effluent of the aerobic lagoon showed significantly higher antibiotic-resistance (34.66%) than those isolated from domestic sewage (23%). In the activated sludge, the percentage of antibiotic resistant strains (resistance to at least one antibiotic) showed seasonal changes in the inflow and outflow water samples. The increase of the percentage of antibiotic resistant strains of E. coli in the outflow of the lagoon system is probably related to the selection of antibiotic resistant strains by this kind of treatment. Transfer of antibiotic-resistance between E. coli strains can occur in the wastewaters treated by this system. Survival experiments on two strains of E. coli, E. coli resistant to seven antibiotics and E. coli O126:B16 sensitive to the 15 tested antibiotics, isolated from the environment, show the ability of resistant bacteria to survive better than sensitive ones in waste water treated in lagoons.
In recent years, there has been a tremendous amount of research and development in the area of photocatalysis (heterogeneous and homogeneous), a process included in a special class of oxidation techniques defined as Advanced Oxidation Processes (AOPs), all characterized by the same chemical feature, production of OH radicals. This paper reviews the use of sunlight to produce the OH radicals by TiO2 photocatalysis and photo-Fenton process. The reacting systems necessary for performing solar photocatalysis are described. The paper also summarizes most of the research carried out related to solar photocatalytic degradation of water contaminants, and how it could significantly contribute to the treatment of persistent toxic compounds. It outlines how to enhance the process efficiency by integration with biotreatment. Various solar reactors for photocatalytic water treatment mainly based on non-concentrating collectors built during the last few years are also described in detail in this review, as well as the use of the solar photocatalytic processes to inactivate microorganisms present in water, placing special emphasis on experimental systems made to optimize this disinfection technique.
The fate of antibiotics in the environment, and especially antibiotics used in animal husbandry, is subject to recent studies and the issue of this review. The assumed quantity of antibiotics excreted by animal husbandry adds up to thousands of tonnes per year. Administered medicines, their metabolites or degradation products reach the terrestrial and aquatic environment by the application of manure or slurry to areas used agriculturally, or by pasture-reared animals excreting directly on the land, followed by surface run-off, driftage or leaching in deeper layers of the earth. The scientific interest in antimicrobially active compounds in manure and soil, but also in surface and ground water, has increased during the last decade. On the one side, scientific interest has focused on the behaviour of antibiotics and their fate in the environment, on the other hand, their impact on environmental and other bacteria has become an issue of research. Analytical methods have now been developed appropriately and studies using these new techniques provide accurate data on concentrations of antimicrobial compounds and their residues in different organic matters. Some antibiotics seem to persist a long time in the environment, especially in soil, while others degrade very fast. Not only the fate of these pharmaceuticals but their origin as well is an object of scientific interest. Besides human input via wastewater and other effluents, livestock production has been recognised as a source of contamination. One main concern with regard to the excessive use of antibiotics in livestock production is the potential promotion of resistance and the resulting disadvantages in the therapeutic use of antimicrobials. Since the beginning of antibiotic therapy, more and more resistant bacterial strains have been isolated from environmental sources showing one or multiple resistance. There have been several attempts to use antibiotic resistance patterns in different bacteria as indicators for various sources of faecal pollution. This review gives an overview of the available data on the present use of veterinary antibiotics in agriculture, on the occurrence of antibiotic compounds and resistant bacteria in soil and water and demonstrates the need for further studies.
In this study, the impact of tertiary-treated municipal wastewater on the quantity of several antibiotic resistance determinants in Duluth-Superior Harbor was investigated by collecting surface water and sediment samples from 13 locations in Duluth-Superior Harbor, the St. Louis River, and Lake Superior. Quantitative PCR (qPCR) was used to target three different genes encoding resistance to tetracycline (tet(A), tet(X), and tet(W)), the gene encoding the integrase of class 1 integrons (intI1), and total bacterial abundance (16S rRNA genes) as well as total and human fecal contamination levels (16S rRNA genes specific to the genus Bacteroides ). The quantities of tet(A), tet(X), tet(W), intI1, total Bacteroides , and human-specific Bacteroides were typically 20-fold higher in the tertiary-treated wastewater than in nearby surface water samples. In contrast, the quantities of these genes in the St. Louis River and Lake Superior were typically below detection. Analysis of sequences of tet(W) gene fragments from four different samples collected throughout the study site supported the conclusion that tertiary-treated municipal wastewater is a point source of resistance genes into Duluth-Superior Harbor. This study demonstrates that the discharge of exceptionally treated municipal wastewater can have a statistically significant effect on the quantities of antibiotic resistance genes in otherwise pristine surface waters.
The taxonomic diversity and antibiotic resistance phenotypes of aeromonads were examined in samples from drinking and waste water treatment plants (surface, ground and disinfected water in a drinking water treatment plant, and raw and treated waste water) and tap water. Bacteria identification and intra-species variation were determined based on the analysis of the 16S rRNA, gyrB and cpn60 gene sequences. Resistance phenotypes were determined using the disc diffusion method. Aeromonas veronii prevailed in raw surface water, Aeromonas hydrophyla in ozonated water, and Aeromonas media and Aeromonas puntacta in waste water. No aeromonads were detected in ground water, after the chlorination tank or in tap water. Resistance to ceftazidime or meropenem was detected in isolates from the drinking water treatment plant and waste water isolates were intrinsically resistant to nalidixic acid. Most of the times, quinolone resistance was associated with the gyrA mutation in serine 83. The gene qnrS, but not the genes qnrA, B, C, D or qepA, was detected in both surface and waste water isolates. The gene aac(6')-ib-cr was detected in different waste water strains isolated in the presence of ciprofloxacin. Both quinolone resistance genes were detected only in the species A. media. This is the first study tracking antimicrobial resistance in aeromonads in drinking, tap and waste water and the importance of these bacteria as vectors of resistance in aquatic environments is discussed.
Viable but non-culturable (VBNC) bacteria are common in nutrient poor and/or stressed environments as planktonic cells and biofilms. This article discusses approaches to researching VBNC bacteria to obtain knowledge that is lacking on their gene expression while in the VBNC state, and when they enter into and then recover from this state, when provided with the necessary nutrients and environmental conditions to support growth and cell division. Two-dimensional gel electrophoresis of proteins, global gene expression, reverse-transcription polymerase chain reaction (PCR) analysis and sequencing by synthesis coupled with data on cell numbers, viability and species present are central to understanding the VBNC state.
Antibiotic resistance in Gram-negative bacteria is often due to the acquisition of resistance genes from a shared pool. In multiresistant isolates these genes, together with associated mobile elements, may be found in complex conglomerations on plasmids or on the chromosome. Analysis of available sequences reveals that these multiresistance regions (MRR) are modular, mosaic structures composed of different combinations of components from a limited set arranged in a limited number of ways. Components common to different MRR provide targets for homologous recombination, allowing these regions to evolve by combinatorial evolution, but our understanding of this process is far from complete. Advances in technology are leading to increasing amounts of sequence data, but currently available automated annotation methods usually focus on identifying ORFs and predicting protein function by homology. In MRR, where the genes are often well characterized, the challenge is to identify precisely which genes are present and to define the boundaries of complete and fragmented mobile elements. This review aims to summarize the types of mobile elements involved in multiresistance in Gram-negative bacteria and their associations with particular resistance genes, to describe common components of MRR and to illustrate methods for detailed analysis of these regions.
The usage of antibiotics in animal husbandry has promoted the development and abundance of antibiotic resistance in farm environments. Manure has become a reservoir of resistant bacteria and antibiotic compounds, and its application to agricultural soils is assumed to significantly increase antibiotic resistance genes and selection of resistant bacterial populations in soil. The genome location of resistance genes is likely to shift towards mobile genetic elements such as broad-host-range plasmids, integrons, and transposable elements. Horizontal transfer of these elements to bacteria adapted to soil or other habitats supports their environmental transmission independent of the original host. The human exposure to soil-borne resistance has yet to be determined, but is likely to be severely underestimated.
To characterize the molecular diversity of class 1 integrons and antibiotic resistance (AR) genes of Enterobacteriaceae strains recovered from aquatic habitats in Jinan, Shandong Province, China.
Six hundred and thirty-eight antimicrobial-resistant Enterobacteriaceae isolated from wastewater were examined for class 1 integron. Of these, 293 were positive for the class 1 integrase gene intI1; among these, 34 gene cassettes and 29 AR genes were detected. Twenty-nine distinct gene cassette arrays were identified by restriction fragment length polymorphism (RFLP). Seven strains harboring novel gene cassette arrays were subjected to further study, in which antimicrobial susceptibility profiles were determined, and the presence of other AR genes outside of the integrons was assayed. Several of the resistance determinants were found to be transferable by conjugation or transformation.
This study established the assessment of class 1 integron and antimicrobial resistance gene patterns among environmental Enterobacteriaceae. Also, a restriction enzyme EcoRII was employed to develop a rapid and simple method for characterizing gene cassette arrays by RFLP analysis, which facilitated further study of novel gene cassette arrays.
These data not only illustrated the diversity of class 1 integron gene cassettes but also provided direct evidence that integrons mobilized gene cassettes, generating new linkages of resistance genes, and they could be integrated in gene transfer units such as conjugative plasmids to contribute to the dissemination of AR genes by horizontal gene transfer (HGT) in aquatic environments.