Chemical profiling of manure: toward source inference in environmental fraud cases?

With 6% of Europe's freshwater resources, Switzerland is often considered the ‘water tower of Europe’. Over 8 millions residents rely on more than 57’600 farms to provide meat, milk, or eggs, with 1.58 million cattle, 1.58 million swine or 9.39 million poultry. Livestock farming produce the manure, which is a mixture of urine, faeces and infrastructure washing water. Manure is stockpiled in over 100’000 pits in Switzerland (total capacity of 18 million m3), and then used as fertilizer. When manure enters aquatic environment, such pollution could be responsible of major health risks, contaminating drinkable water sources, or ecological disasters with the death of biota, including fishes. This kind of accident occurs frequently, and authors are in many cases not found by environmental law enforcement. This study aimed at exploring the possibilities of chemical markers to differentiate manures, with the purpose of estimating whether it would be possible to perform source inference in manure pollution cases in a chemical profiling approach. An exploratory market research was first conducted to understand practices of farming chemicals use, including antibiotics and detergents, in order to select potential chemical markers. Manure samples were analysed using ASE-SPE-UPLC-MS/MS (Accelerated Solvent Extraction – Solid Phase Extration – Ultra Performance Liquid Chromatography - tandem Mass Spectrometry) to detect 17 selected chemical markers with Multiple Reaction Monitoring mode: 11 antibiotics (benzylpenicillin, cloxacillin, colistin, danofloxacin, enrofloxacin, neomycin, sufadoxin, sulfaguanidin, sulfamethoxazol, tetracycline and trimetroprim), 1 vermifuge (flubadenzol), 2 local anesthetics (lidocaine and procain), 1 detergent (dodecylbenzenesulfonic acid) and 2 manure/faecal chemical markers (skatole and indole). Analysis in scan mode, extraction of several m/z and statistical testing (PCA and hierarchical cluster analysis) permitted to differentiate manure from cattle and pig farms, and some markers may also be useful to identify a farm among others. Finally, analysis of few specimens collected during pollution events showed that some chemical markers can be found in the water. The results of this study suggest that the applied methodology could allow to identify some types of pollution (including faecal pollution such as manure or wastewater treatment plant malfunctioning), and may help to achieve source inference in environmental fraud cases.

On-field chemical profiling of volatile and odorous compounds in water pollution cases: toward a forensic in situ approach

Luiz Felippe De Alencastro, William Lacour

With 6% of Europe's freshwater resources, Switzerland is often considered the ‘water tower of Europe’. There are 57’600 farms and 750 wastewater treatment plants (WWTP) in the country. When manure or raw wastewater enters aquatic environment, accidently or intentionally, such pollution can provoke major health risks, contaminating drinkable water sources, or ecological disasters with the death of biota. Among the detected water pollutions, the sources of 25% remain unknown. On one side, it is difficult for environmental law enforcement to initially classify a water pollution (e.g. manure vs WWTP dysfunction), because there is no quick method routinely used on the field. This often hinders following investigation to identify toxic chemicals in the water or potential geographical source in order to target adequate sampling. This works aims at comparing actual practice with potential portable detectors in order to define a forensic approach in water pollution cases investigated by the police.

2015

Assainissement de la décharge de Colliare à Penthaz – Etude de variantes

Gabrielle Hack

The landfill of Colliare is situated on the left bank of the river « la Venoge », at the place of a former gravel pit in the municipality of Penthaz. It was exploited until 1995 and many different wastes were deposited in it, such as incineration slag of household waste, grinding of cars, residues of biostabilizer, waste of construction site and materials of excavation. A water pollution due to the presence of this landfill was noticed at the beginning of the nineties, mainly by ammonium and nitrite. In one measure chromium was also found in rather important concentration. The presence of these compounds in water reflects the existence of biodegradation of the organic matter present in the mass of waste. The concentration of ammonium being higher than the limit imposed by Swiss law, a remediation is necessary. Various projects of remediation were made during the last twenty years, the main being to clean up this former landfill in order to have the possibility of creating a new one for bioactive wastes. This project was rejected by the « Grand Conseil » and the remediation of the landfill was set aside during several years. Today, the question is to study various variants of remediation to define which one would be the best for Colliare. The choice of the variants was limited either to a containment of the landfill’s surface or to bioventing. This work studies the possible implantation of these two variants, by analysing their advantages and disadvantages so as to facilitate the choice of the final solution. For that purpose, laboratory analyses were made to define the list of contaminants present in the mass of waste that risk to contaminate the water near the landfill. It was demonstrated that beside the already turn out organic pollution, the landfill contains a huge quantity of PCB and PAH as well as several heavy metals such as chromium, lead or copper. Furthermore analyses to obtain information on the quantity of biodegradable organic matter were made. It was shown that organic matter constitues only a small percentage of the mass of waste. Landfill containment is not considered as a real remediation because the contaminants stay in the site. The solution is studied here because of the various types of contaminants present in the waste. Furthermore the time it takes to implement is relatively short and it is possible to put vegetation on the site just after the installation of the mineral cover. There are thus few impacts on the fauna and the flora and the contaminants are normally not a risk anymore for the water during all the life of the containment. Bioventing has as main advantage to be a real method of remediation. The problem of its implementation in the case of Colliare is the presence of PCB and PAH in the mass of waste. Indeed biodegradation would have no effect on these compounds and the risk of water contamination would not decrease thanks to bioventing. Furthermore the small percentage of organic matter does not justify the implementation of such an installation. Finally the costs of both variants were compared. Containment costs less than bioventing. Thus the cost of bioventing is too high to implement it in addition to containment except if the decrease of greenhouse gases emissions is enough important for being exchanged on the market of carbon. The solution chosen here is landfill containment, although it is not ideal for a real remediation of the site.

2011

Sources, Spreading and Fate of Antibiotic Resistance Genes and Resistant Bacteria in Vidy Bay, Lake Geneva, Switzerland

Nadine Czekalski

Increasing numbers of multiresistant bacterial pathogens cause severe health problems on a global scale. Large quantities of bacteria and their resistance genes are discharged into natural environments via human and animal waste. Not much is known so far about the fate and further spread of resistance determinants in the environment. There is evidence that aquatic environments may function as important compartments for preserving and exchanging antibiotic resistance genes (ARG) from sewage and drinking water. Lake Geneva is the largest fresh water reservoir of Western Europe and supplies the surrounding cities. At the same time these cities discharge sewage into the lake. Lausanne is the largest city in the lake’s catchment and discharges the highest amount of treated, as well as in case of heavy rain events, partly untreated waste waters. Also two rivers, Chambering and Flown, transport untreated sewage (bypass), in case of storm events. Only 3.4 km from the sewage discharge point lake water is pumped for drinking water preparation for Lausanne. Many studies have reported on contamination of the bay with heavy metals, inorganic nutrients, organic carbon and fecal indicator bacteria due to the sewage discharge. The present thesis aimed on investigating the sources, spreading and fate of antibiotic resistant bacteria and antibiotic resistance genes in the Vida bay, Lake Geneva. In the first part of this dissertation, a screening for antibiotic- and specifically multi-resistant bacteria was carried out as one of the first studies in a Swiss waste water stream and the receiving fresh water lake. For this purpose, a combined set of culture based (viable bacterial counts on nutrient media supplemented with antibiotics) and molecular tools (detection and quantitation of antibiotic resistance genes from environmental DNA-extracts using quantitative real-time PCR) was applied. Raw hospital and communal sewage, as well as partly treated/mixed sewage, which is discharged into the lake via an effluent pipe 700 m off shore, were taken into account. Furthermore lake water samples nearby the end of the waste water discharge pipe and the drinking water pump as well as close to the two river mouths were analyzed. Finally, the role of Lausannes WTP was evaluated for its potential to remove resistant and multiresistant bacteria, as well ARG. Results show highest levels of multiresistant bacteria and ARG in both hospital and communal sewage. Incidence was found that the WTP is a selective environment for multiresistant bacteria and resistance genes. Moreover, lake water and sediments nearby the end of the WTP discharge pipe and the two river mouths exhibited significantly higher levels of resistant determinants as compared to samples from the drinking water uptake. If the level of antibiotic resistance close to the drinking water uptake represents the natural background level of the lake or whether it is impacted by the WTP plume remained open during this first survey. The spatial distribution and persistence of ARG discharged via waste water into lake systems has been evaluated for the first time in the second part of this thesis. 22 sediment cores were sampled along two major transects (leading either into the deep lake or along the south-western shore line to the drinking water uptake) and in the vicinity of the WTP discharge pipe in the Vidy bay. A subset of samples was also analyzed for contamination with a “conventional” pollutant of sewage (mercury), for which a similar transport behavior (particle bound) as for bacteria and their resistance genes is assumed. The microbial community composition in all sediment as well as in waste water samples was analyzed using ARISA. Strong pollution with both ARG and mercury was detected in close proximity of the WTP discharge pipe and both contaminants significantly decreased along the transect into the deep lake. In general however, the spatial contamination of mercury and ARG was only moderately correlated. While the mercury contaminated sediment surface area tended to have a pollution tail in south eastern direction, ARG showed only a slow decrease along the south western transect. These findings imply differences in transport and fate of mercury and ARG in the bay, or additional sources of ARG contamination. Analysis of the microbial diversity in the samples support these hypotheses. This study revealed feasibility of qPCR as a tool to semi-quantitatively track spatial ARG contamination in fresh water lake sediments. Not much is known yet on the antibiotic resistance background level of fresh water lakes and how this may be influenced by anthropogenic activities in the surrounding catchment. Water samples from 21 Swiss lakes have been collected and were analyzed for the prevalence of different ARG (2 sulfonamide resistance genes sul1 & sul2, 3 tetracycline resistance genes tetB, tetM, tetW and one plasmid mediated fluoroquinolone resistance gene qnrA). In order to document the abundance of these genes in the bacterial populations, they were normalized to bacterial 16S rRNA gene copy numbers. The lakes differ in size, geographical position, land use pattern, and trophical status. The composition of the microbial community of all lakes was analyzed using ARISA. Sulfonamide resistance genes were detected in all investigated lakes (0.2-4%, sul1 and 0.01-0.3%, sul2), with highest levels in Lake Baldegger (21%). Lake Baldegger also contained all of the three tetracycline resistance genes, which were found in only 4 other lakes. None of the lakes was positive for qnrA. Important anthropogenic catchment activities linked to sul gene prevalence could not be identified based on our data and may require further analysis and an additional variables to complete our data set. However, our study demonstrates contamination of Swiss lakes with sulfonamide resistance genes and the potential of fresh water lakes preserve these genes, even remote from direct sources of pollution.