During WELLMA-DNA, 13 diploma and bachelor theses along with several internships have been completed. A sampling system for biofilm samples as well as a sampling device for water samples have been designed and tested. More than 400 DNA samples of different well sites have been collected and analyzed. Microbiological and molecular methods have been combined to gain a better understanding of the community composition of the ochre forming biofilms inside the wells. Molecular methods included PCR, DGGE, cloning and sequencing. During the project, the bacterial populations of an unprecedented number of wells have been analyzed and several indicator bacteria for iron-related well clogging have been identified. Alongside iron-oxidizing bacteria, iron-reducing bacteria have been found in the wells and their potential for ochre-solubilization was confirmed. Alongside the molecular experiments, microbiological trials included the isolation of pure cultures, microscopic analysis and physiological tests. The morphology of the encountered iron bacteria could be classified into four different groups, which may have an impact on the rigidity of the biofilms on a macroscopic level. We were able to cultivate several of these indicator organisms, which could play an important role in the formation of ochreous deposits in the Berlin wells. During experiments utilizing microscopic flow cells, differences in growth rate and patterns of these ochre-forming bacteria have been observed. For several of the identified indicator bacteria, primers have been calculated. These primers will allow for the first time to quantify the amount of indicator bacteria in a water sample and to derive operational pointers. In addition, several experiments regarding the effect of hydrogen peroxide on ochre forming biofilms have been conducted and the effect of an additional electron donor (ethanol) on the communities has been tested. For future data acquisition and documentation, a guideline for classifying the degree of pump clogging has been developed.

Trinkwasserbrunnen unterliegen natürlichen Alterungsprozessen, unter anderem der Verockerung (Wiacek 2006). Es wurden mehrere Biofilmproben aus der Rohwasserleitung des Wasserwerks Stolpe-Berlin entnommen (Kapitel 1.4/ 2.2.1). Die Proben die von der TU-Berlin zur Verfügung gestellt wurden, wurden kultiviert und anschließend mit mikrobiologischen Methoden charakterisiert (Kapitel 3.4). Vier Stämme, die verstärkt Eisenablagerungen und dunkle Kolonien aufwiesen, wurden für Folgeversuche (Kapitel 3.2/ 3.3) eingesetzt. Alle Versuchsergebnisse deuten daraufhin, dass Eisenhydroxid einen großen Einfluss auf die Wirkung des H2O2 auf Biofilme hatte (Kapitel 3). Es hat sich ergeben, dass eisenoxidierende Bakterien und mit ihnen assoziierte Bakterien (Kapitel 3.6) effektiv zu bekämpfen sind, wenn die sie umgebenden Eiseninkrustierungen vorher gelöst werden (Kapitel 4.1.5). Sowohl in den Plattenversuchen, als auch in den Versuchen mit den Biofilmen, haben Oxalsäure und die EDTA-Lösung die besten Ergebnisse erzielt, in bezug auf die sich lösenden Eiseninkrustierung und der nachfolgenden Reduzierung der Zellzahlen mit H2O2. Auch wenn diese Stoffe nicht in verockerten Trinkwasserbrunnen eingesetzt werden können, dienten sie doch dazu, die negative Wirkung des Eisenhydroxids bei der Biofilmbehandlung mit H2O2 zu verdeutlichen.

Well biofouling is a complex and not yet sufficiently understood process. Water wells represent a unique habitat, since they create a link between the anaerobic ground water containing Fe(II) and the aerobic surface. This groundwater is rich in soluble Fe(II) and the presence of trace amounts of free oxygen in the well screens presents ideal conditions for the growth of iron bacteria. The ochreous deposits produced block not only the filter area, but also the adjacent gravel pack or even parts of the aquifer, and result in a steady decrease of well performance. In this project, the bacterial communities of several Berlin wells have been compared using standard microscopic techniques and molecular techniques like DGGE. The aim of this study is to identify the responsible bacteria and associated processes that lead to well clogging. The sampling system allows easy and effective collection of undisturbed biofilm samples with minimal impact on normal well operation. Fingerprinting analysis indicates the presence of bacterial populations that are ubiquitous in the wells and certain indicator bacteria which can be found in only few wells. Free water and biofilm populations show distinct similarities. Some well populations can be grouped in clusters, depending on the location of the well. Further comparison with chemical data of the wells is planned to asses the impact of chemical conditions of the respective groundwater on the bacteria responsible for clogging. A sampling device was designed and built for exposure of carrier materials into different Berlin drinking water abstraction wells. In addition, samples from well components (pumps and pipes) and water samples were collected. The DNA was extracted using the FastDNA SPIN Kit for Soil (MP). 16S rDNA polymerase chain reaction (PCR) of the V3 region and denaturing gradient gel electrophoresis (DGGE) analyses were performed on the DNA samples (Muyzer et al., 1993).

Well biofouling is a complex and yet not sufficiently understood process. Water wells represent a unique habitat, since they create a link between the anaerobic ground water, containing Fe(II) and the aerobic surface. These special conditions set ideal conditions for the growth of iron bacteria (Stuetz and McLaughlan, 2004). Some of these bacteria are known to be responsible for well clogging by precipitation of iron hydroxides (Cullimore, 1999). The consistency of the ochres can range from soft and bulky to solid and compact. The type of deposit strongly depends on the dominant species of bacteria at the well screen and inside the gravel pack. Within this project (WellMa) a sampling system was created, which allowed the collection of unaffected biofilm samples from inside the wells. The samples were microscopically examined, DNA was extracted and community profiles were created.

Bacterial induced well clogging is a common problem in water wells. The well represents a unique habitat by creating a link between the anaerobic ground water, containing Fe(II) and the aerobic surface. The presence of trace amounts of free oxygen in the well screens, sets ideal conditions for the growth of iron bacteria (Stuetz and McLaughlan, 2004). These bacteria precipitate iron hydroxides (Cullimore, 1999), that not only block the filter area, but also the adjacent gravel pack or even parts of the aquifer and result in a steady decrease of well performance. Each well has it’s own distinct chemical conditions, which impact the type of bacterial community that forms in the gravel pack. Within this project a novel sampling system was developed, which allowed the collection of intact biofilm samples from a selected range of Berlin water wells. The resulting biofilms were microscopically examined to gain a first rough overview of the different sampling sites. Subsequently, the bacterial DNA was extracted and used for a population comparison utilizing denaturing gradient gel electrophoresis, cloning and sequencing.