Insights into antimicrobial resistance patterns in Lower Austrian rivers along gradients of faecal pollution

Abstract

Antibiotic resistance (AR) is increasing worldwide causing several hundred thousands of deaths per year. The clinical environment has been demonstrated to be a hot-spot of development, transmission and spread of AR in developed countries such as Austria. However, in the past years, there have been increasing discussions about the role of the natural environment in the transmission, evolution and spread of AR, and therefore environmental AR surveillance and monitoring is recommended. Antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) predominantly enter surface waters such as lakes and rivers through faecal pollution via the wastewater pathway but also diffuse pollution sources such as agricultural run-offs. The aim of the thesis is to evaluate the AR and faecal pollution situation in Lower Austrian rivers in water and submerged biofilms with strong focus on the human wastewater pathway and also whether clinical resistance situation is reflected in the environment. The first chapter presents a large-scale screening of AR in three tributaries and a small stretch of the Danube River in Lower Austria over a one-year cycle. Beside river water samples, submerged biofilms were investigated to find out whether in these habitats is an accumulation of AR and if clinically relevant ARB and ARGs are likely to establish in the environment. All samples were investigated with a harmonised toolbox and study design to elucidate potential driving factors of the resistome in these rivers. For a quantitative assessment, the abundance of nine ARGs and the resistance ratios of 5197 Escherichia coli (E. coli) isolates were linked to quantitative faecal pollution data and environmental, chemical and hydrological parameters. The key parameter influencing AR abundance in the investigated catchment was anthropogenic faecal pollution and strongly connected to wastewater discharges. In all rivers the faecal pollution levels were clearly higher in the water compared to biofilm indicating that faecal bacteria do not establish in the riverine biofilm communities. However, for ARB and ARGs neither water nor biofilm could be identified as generally more important AR habitat with partly contrasting results for the individual rivers and at the sampling sites. This clearly indicates that general conclusions about the prevalence and potential driving factors of ARGs or phenotypic resistances in rivers and the role of biofilms should be drawn with caution, specifically when investigations were done at small spatial scales and with different methods. In the second chapter, the main results from first chapter concerning the occurrence and abundances of ARB and ARG in the four investigated rivers were summarized for the national water management sector and water authorities. This chapter also contains a detailed comparison of the measured ARB and ARG concentrations in river water with results from other countries with focus on Europe and shows that the investigated rivers in Austria display a comparably moderate pollution status concerning ARB and ARG.The third chapter represents a comparison of environmental E. coli isolates sampled in the Danube River upstream and downstream of a wastewater treatment plant receiving wastewater from a hospital with isolates obtained in the same hospital from urine of patients suffering from urinary tract infections or colonised urethral catheters. Patient isolates had significantly higher resistance ratios than the environmental ones and higher resistance ratios were found in biofilm compared to water. Seven extended-spectrum beta-lactamase (ESBL) producing isolates were isolated from the patients, two ESBL-E.coli were obtained from biofilm. In addition, one multi-drug resistant isolate carrying a tigecycline resistance, slightly higher imipenem and significantly higher amoxicillin/clavulanic acid resistance levels were detected in the biofilm isolates compared to the patient isolates.The fourth chapter evaluated the impact of a short-term cultural event on the faecal pollution status of the Traisen River. Enterococci and E. coli were quantified as standard faecal indicators before, during and after the music festival. During the event a massive increase in enterococci was observed, while the E. coli concentrations did not show such clear elevation. The difference of the two indicators may be explained by different persistence to many of the substances of personal care products which were found in high levels water. The different sensitivity may have been additionally supported by the high water temperatures and solar radiation during the festival period. In concern of human health, for upcoming festivals a more effective management, i.e., improved sanitation infrastructure and better waste management, is recommended to prevent such massive faecal pollution in the Traisen River.Although this thesis has a strong focus on rivers in Lower Austria, the developed investigation strategy including study design and quantitative methodology allows comparing AR prevalences internationally. In addition to the investigation strategy which can be applied on different environmental waters, the thesis provides suggestions for future management strategies, e.g., application of more efficient wastewater treatment and creation of awareness for AR, to prevent dissemination and mitigation of AR and faecal pollution in the riverine environment.