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PhDENV / Department of Ecology and Resource Management / See the attached abstract below

The main purpose of this thesis is to use Image Spectroscopy as a tool to monitor the environmental conditions in a region affected by anthropogenic activities via estimating both geochemical and biochemical parameters on a regional scale. The research has been carried on the Sokolov lignite mine, NW Bohemia, a region affected by long-term extensive mining. The thesis is divided into two thematic parts. First part is devoted to applications of Image Spectroscopy into Acid Mine Drainage mapping and its related issues (chapters 2 and 3). In chapter 2 the equivalent mineral end-members were successfully derived from the ASTER image data (Advanced Space-borne Thermal Emission and Reflection Radiometer satellite data). In the chapter 3 the pH was estimated on the basis of mineral and image spectroscopy. The Multi Range Spectral Feature Fitting (MRSFF) technique was utilized for mineral mapping and the multiple regression model using the fit images, the results of MRSFF, as inputs was constructed to estimate the surface pH and statistical significant accuracy was attained. In the second thematic part (chapters 4-6) Image Spectroscopy is applied into monitoring of vegetation stress. A new statistical method was developed to assess the physiological status of macroscopically undamaged foliage of Norway...

Acidophilic iron oxidizing bacteria of the betaproteobacterial genus “Ferrovum” are ubiquitously distributed in acid mine drainage (AMD) habitats worldwide. Since their isolation and maintenance in the laboratory has proved to be extremely difficult, members of this genus are not accessible to a “classical” microbiological characterization with exception of the designated type strain “Ferrovum myxofaciens” P3G. The present study reports the characterization of “Ferrovum” strains at genome and transcriptome level. “Ferrovum” sp. JA12, “Ferrovum” sp. PN-J185 and “F. myxofaciens” Z-31 represent the iron oxidizers of the mixed cultures JA12, PN-J185 and Z-31. The mixed cultures were derived from the mine water treatment plant Tzschelln close to the lignite mining site in Nochten (Lusatia, Germany). The mixed cultures also contain a heterotrophic strain of the genus Acidiphilium. The genome analysis of Acidiphilium sp. JA12-A1, the heterotrophic contamination of the mixed culture JA12, indicates an interspecies carbon and phosphate transfer between Acidiphilium and “Ferrovum” in the mixed culture, and possibly also in their natural habitat. The comparison of the inferred metabolic potentials of four “Ferrovum” strains and the analysis of their phylogenetic relationships suggest the existence of two subgroups within the genus “Ferrovum” (i.e. the operational taxonomic units OTU-1 and OUT-2) harboring characteristic metabolic profiles. OTU-1 includes the “F. myxofaciens” strains P3G and Z-31, which are predicted to be motile and diazotrophic, and to have a higher acid tolerance than OTU-2. The latter includes two closely related proposed species represented by the strains JA12 and PN-J185, which appear to lack the abilities of motility, chemotaxis and molecular nitrogen fixation. Instead, both OTU-2 strains harbor the potential to use urea as alternative nitrogen source to ammonium, and even nitrate in case of the JA12-like species. The analysis of the genome architectures of the four “Ferrovum” strains suggests that horizontal gene transfer and loss of metabolic genes, accompanied by genome reduction, have contributed to the evolution of the OTUs. A trial transcriptome study of “Ferrovum” sp. JA12 supports the ferrous iron oxidation model inferred from its genome sequence, and reveals the potential relevance of several hypothetical proteins in ferrous iron oxidation. Although the inferred models in “Ferrovum” spp. share common features with the acidophilic iron oxidizers of the Acidithiobacillia, it appears to be more similar to the neutrophilic iron oxidizers Mariprofundus ferrooxydans (“Zetaproteobacteria”) and Sideroxydans lithotrophicus (Betaproteobacteria). These findings suggest a common origin of ferrous iron oxidation in the Beta- and “Zetaproteobacteria”, while the acidophilic lifestyle of “Ferrovum” spp. may have been acquired later, allowing them to also colonize acid mine drainage habitats.

MESMEG / Department of Mining and Environmental Geology / This study entails geochemical and mineralogical characterization of gold tailings of Nestor Mine and Glynn’s Lydenberg Mine of the Sabie-Pilgrim’s Rest goldfields. A total of 35 samples were collected and were analysed for chemical composition (XRF and ICP-MS), mineralogical composition (XRD). In addition, acid-base accounting (ABA) techniques had been conducted to predict the potential for acid mine drainage. Seepage from Nestor tailings dump and water samples from the adjacent Sabie River were also collected and analysed by means of inductively coupled plasma mass spectrometry (ICP-MS) and immediate constituent (IC) -analytical techniques. The study revealed that Sabie-pilgrim’s rest goldfield is characterized by both acid generating and non-acid producing tailings, and this is attributed to variations in the mineralogy of source rocks. Gold occurred within the Black Reef Quartzite Formation in the Nestor Mine and within the Malmani Dolomite in the case of Glynn’s Lydenburg Mine. Mineralogy and bulk geochemical analyses performed in this study showed a clear variation in the chemistry of Nestor Mine and Glynn’s Lydenburg Mine tailings. Predominant oxides in Nestor mine tailings samples are SiO2 (ranging from 66.7-91.25 wt. %; followed by Fe2O3 and Al2O3 (in range of 0.82-15.63 wt. %; 3.21-12.50 wt. % respectively); TiO2 (0.18-10.18 wt. %) and CaO (0.005-3.2 wt. %). Also occurring in small amounts is CaO (0.005-3.2 wt. %), K2O (0.51-2.27 wt. %), MgO (0.005-1.46 wt. %), P2O5 (0.029-0.248), Cr2O3 (0.013-0.042 wt. %) and Na2O (0.005-0.05 wt. %). The samples also contain significant concentrations of As (137-1599 ppm), Cu (34-571 ppm), Cr (43-273 ppm), Pb (12-276 ppm), Ni (16-157 ppm), V (29-255 ppm), and Zn 7-485 ppm). In the Glynn’s Lydenburg Mine tailings SiO2 is also the most dominant oxide ranging between 47.95 and 65.89 w%; followed by Al2O3 (4.31 to 16.19 wt. %), Fe2O3 (8.48 to 11.70 wt %), CaO (2.18 to 7.10 wt. %), MgO (2.74 to 4.7 wt. %). Occurring in small amounts is K2O (1.12-1.70 wt. %), MnO (0.089-0.175 wt. %), P2O5 (0.058-0.144 wt. %) and Cr2O3 (0.015-0.027 wt. %). Arsenic (As), is also occurring in significant amounts (807-2502 ppm), followed by Cr (117-238 ppm), Cu (10-104 ppm), V (56-235 ppm), Ni (45-132 ppm), Pb (13-63 ppm) and Zn (90-240 ppm). Nestor Mine tailings associated with Black Reef Formation mineralization have net neutralizing potential (NPR) <2, hence more likely to generate acid; and their acid potential (AP) ranges 1.56 to 140.31 CaCO3/ton and neutralizing potential (NP) range from -57.75 to -0.3 CaCO3/ton. Glynn’s Lydenburg Mine tailings dump which is vi associated with dolomite mineralization, however, was not leaching acid. Based on acid-base accounting results, these tailings have more neutralizing potential (ranging between 57.6 and 207.88 CaCO3/ton) than acid potential (ranging between 7.5 and 72.1 CaCO3/ton); and their NPR>2, hence unlikely to produce acid. This is confirmed by paste pH which was in the ranges between 7.35 and 8.17. Tailings eroded from Nestor Mine tailings dump were also found to be characterized by high content of metals and oxides, namely, As, Cu, Ni, Pb, V, and Zn with SiO2, Fe2O3 and TiO2. The tailings were observed eroded into the Sabie River where AMD related precipitate (yellow boy) was also observed, indicating further oxidation downstream. Field observations, onsite analyses of water samples and laboratory results revealed that Nestor Mine tailings storage facility discharges acid mine drainage with considerable amounts of Al, As, Cu, Fe, Mn, Zn and SO4 and very low pH exceeding the limit as per South African water quality standards. High concentrations of these metals have toxicity potential on plants, animals and humans. Upon exposure to oxygen and water, tailings from Nestor Mine are more likely to generate acid mine drainage that can cause detrimental effect to the environment and the surrounding communities. Potential pollutants are Fe, Mn, Al, As, Cr, Cu, Ni and Pb. Tailings from Glynn’s Lydenberg showed no potential for acid mine drainage formation. / NRF

BC Hydro’s Water Use Planning (WUP) process is one of the world’s most comprehensive hydroelectric dam operational reviews and has served as a model to revise hydropower operating plans with the participation of an inclusive range of stakeholders, rights holders, and the use of up-to-date scientific information, that meets social and environmental goals alongside economic targets. In 2000, BC Hydro initiated a WUP process in the Jordan River watershed. This watershed hosts a wide diversity of water users, including active resource industry stakeholders (mining, forestry, and hydropower), Indigenous rights holders, and rural community citizens; which is representative of watersheds in British Columbia with established WUPs. BC Hydro finalized the Jordan River WUP in 2003, which focuses on establishing critical freshwater flows for fish habitat and achieving specific recreational values of the local community. However, numerous other issues still remain that were beyond the scope of the WUP process, including water quality concerns that were continually brought up by citizens during the consultative process of the WUP. In addition to these concerns, biological monitoring following the implementation of the WUP suggests that contamination from an inactive copper mine has affected and altered sensitive water quality parameters for a healthy Pacific salmon habitat in Jordan River. Yet, there has not been an extensive water quality study conducted that examines the spatial or seasonal water quality extents of the mining contamination in Jordan River, specifically copper. Consequently, fourteen years after the creation of the WUP, local advocates are still struggling to have their concerns heard by the entity responsible for freshwater flow, BC Hydro, alongside federal and provincial government agencies. Advocates are calling for the creation of a watershed-based group as a mechanism for having greater influence in water planning and governance processes. This study explores the research question: if and how has the WUP process contributed to creating watershed governance capacity? This social science thesis project employs a mixed-methods approach using both quantitative and qualitative data. The study includes a document review of relevant water governance literature and focuses on examining the freshwater quality of the Jordan River. Water quality samples were collected over a five-week period from five sites on the Jordan River beginning in September and concluding in October of 2015 during the most sensitive periods of salmon spawning activity in the lower reaches of the Jordan River. Spatial and seasonal water quality trends were identified, and analysis concluded that copper is the primary contaminate affecting the productivity of a healthy salmon habitat in the Jordan River. Acid mine drainage (AMD) processes were identified throughout the water quality data and are strongly influenced by the proximity of existing mine waste piles sourced from an abandoned copper mine, and unnatural anthropogenic flows from the three BC Hydro dams present in the Jordan River system. The final stage of the research project focuses on assessing the adaptive capacity in the watershed to address the issues of concern outlined in the WUP. There is a current movement to create watershed organizations that are formally supported through new legislation in British Columbia, but questions remain about the capacities of these watershed communities to sustain such a formal institution and if these watershed communities are ready to successfully implement a local watershed governance model. The Gupta et al. (2010) six adaptive capacity dimensions provide a logical framework to explore if these capacities are present such that it could be expected that local watershed organizations would be effective as society adapts to more watershed-based governance approaches. Thirteen semi-structured interviews were conducted from October 2016 to February 2017. Interviews and observational data focused on the WUP process and prospective and current members of the Jordan Watershed Round Table (JWRT). The research evaluated whether these six adaptive capacity dimensions are present in watershed communities that have been subjected to water management processes, specifically the WUP program. Overall, the research concluded that the WUP has contributed to some adaptive capacity for watershed governance in the Jordan River, specifically on building the adaptive capacity dimensions: variety, learning capacity, room for autonomous change, leadership, and resources within the JWRT. / Graduate

Acidophilic iron oxidizing bacteria of the betaproteobacterial genus “Ferrovum” are ubiquitously distributed in acid mine drainage (AMD) habitats worldwide. Since their isolation and maintenance in the laboratory has proved to be extremely difficult, members of this genus are not accessible to a “classical” microbiological characterization with exception of the designated type strain “Ferrovum myxofaciens” P3G. The present study reports the characterization of “Ferrovum” strains at genome and transcriptome level. “Ferrovum” sp. JA12, “Ferrovum” sp. PN-J185 and “F. myxofaciens” Z-31 represent the iron oxidizers of the mixed cultures JA12, PN-J185 and Z-31. The mixed cultures were derived from the mine water treatment plant Tzschelln close to the lignite mining site in Nochten (Lusatia, Germany). The mixed cultures also contain a heterotrophic strain of the genus Acidiphilium. The genome analysis of Acidiphilium sp. JA12-A1, the heterotrophic contamination of the mixed culture JA12, indicates an interspecies carbon and phosphate transfer between Acidiphilium and “Ferrovum” in the mixed culture, and possibly also in their natural habitat. The comparison of the inferred metabolic potentials of four “Ferrovum” strains and the analysis of their phylogenetic relationships suggest the existence of two subgroups within the genus “Ferrovum” (i.e. the operational taxonomic units OTU-1 and OUT-2) harboring characteristic metabolic profiles. OTU-1 includes the “F. myxofaciens” strains P3G and Z-31, which are predicted to be motile and diazotrophic, and to have a higher acid tolerance than OTU-2. The latter includes two closely related proposed species represented by the strains JA12 and PN-J185, which appear to lack the abilities of motility, chemotaxis and molecular nitrogen fixation. Instead, both OTU-2 strains harbor the potential to use urea as alternative nitrogen source to ammonium, and even nitrate in case of the JA12-like species. The analysis of the genome architectures of the four “Ferrovum” strains suggests that horizontal gene transfer and loss of metabolic genes, accompanied by genome reduction, have contributed to the evolution of the OTUs. A trial transcriptome study of “Ferrovum” sp. JA12 supports the ferrous iron oxidation model inferred from its genome sequence, and reveals the potential relevance of several hypothetical proteins in ferrous iron oxidation. Although the inferred models in “Ferrovum” spp. share common features with the acidophilic iron oxidizers of the Acidithiobacillia, it appears to be more similar to the neutrophilic iron oxidizers Mariprofundus ferrooxydans (“Zetaproteobacteria”) and Sideroxydans lithotrophicus (Betaproteobacteria). These findings suggest a common origin of ferrous iron oxidation in the Beta- and “Zetaproteobacteria”, while the acidophilic lifestyle of “Ferrovum” spp. may have been acquired later, allowing them to also colonize acid mine drainage habitats.:EIDESSTATTLICHE ERKLÄRUNG ... 2 CONTENT ... 4 SUMMARY ... 9 CHAPTER I ... 11 ORIGIN AND MICROBIOLOGY OF ACID MINE DRAINAGE ... 11 ACIDOPHILIC IRON OXIDIZING BACTERIA OF THE GENUS “FERROVUM” ... 12 APPLICATION OF OMICS-BASED APPROACHES TO CHARACTERIZE ACIDOPHILES ... 14 AIMS OF THE PRESENT WORK ... 15 CHAPTER II ... 17 ABSTRACT ... 18 INTRODUCTION ... 18 METHODS ... 19 GENOME PROJECT HISTORY ... 19 GROWTH CONDITIONS AND GENOMIC DNA PREPARATION ... 20 GENOME SEQUENCING AND ASSEMBLY ... 20 GENOME ANNOTATION ... 21 RESULTS ... 21 CLASSIFICATION AND FEATURES ... 21 GENOME PROPERTIES ... 24 INSIGHTS FROM THE GENOME SEQUENCE ... 24 COMPARATIVE GENOMICS ... 28 CONCLUSIONS ... 30 ACKNOWLEDGMENTS ... 32 AUTHOR CONTRIBUTIONS ... 32 CHAPTER III ... 33 ABSTRACT ... 34 INTRODUCTION ... 34 METHODS ... 36 ORIGIN AND CULTIVATION OF “FERROVUM” STRAIN JA12 ... 36 GENOME SEQUENCING, ASSEMBLY AND ANNOTATION ... 37 VISUALIZATION OF THE NEARLY COMPLETE GENOME ... 38 PHYLOGENETIC ANALYSIS ... 39 PREDICTION OF MOBILE GENETIC ELEMENTS ... 39 NUCLEOTIDE SEQUENCE ACCESSION NUMBER ... 39 RESULTS AND DISCUSSION ... 39 PHYLOGENETIC CLASSIFICATION OF “FERROVUM” STRAIN JA12 ... 39 GENOME PROPERTIES ... 40 NUTRIENT ASSIMILATION AND BIOMASS PRODUCTION ... 44 Carbon dioxide fixation ... 44 Central carbon metabolism ... 45 Nitrogen ... 47 Phosphate ... 49 Sulfate ... 50 ENERGY METABOLISM ... 50 Ferrous iron oxidation ... 50 Other redox reactions connected to the quinol pool ... 54 Predicted formate dehydrogenase ... 55 STRATEGIES TO ADAPT TO ACIDIC ENVIRONMENTS, HIGH METAL LOADS AND OXIDATIVE STRESS ... 55 Acidic environment ... 55 Strategies to cope with high metal and metalloid loads ... 58 Oxidative stress ... 59 HORIZONTAL GENE TRANSFER ... 60 CONCLUSIONS ... 61 ACKNOWLEDGMENTS ... 62 AUTHORS\' CONTRIBUTIONS ... 62 CHAPTER IV ... 63 ABSTRACT ... 64 INTRODUCTION ... 64 METHODS ... 66 ORIGIN AND CULTIVATION OF “FERROVUM” STRAINS PN-J185 AND Z-31 ... 66 GENOME SEQUENCING, ASSEMBLY AND ANNOTATION ... 66 PREDICTION OF MOBILE GENETIC ELEMENTS ... 67 COMPARATIVE GENOMICS ... 68 Phylogenomic analysis ... 68 Assignment of protein-coding genes to the COG classification ... 68 Identification of orthologous proteins ... 68 Comparison and analysis of genome architectures ... 69 RESULTS ... 69 GENERAL GENOME FEATURES AND PHYLOGENETIC RELATIONSHIP OF THE FOUR “FERROVUM” STRAINS ... 69 COMPARISON OF INFERRED METABOLIC TRAITS ... 71 Identification of core genes and flexible genes ... 71 Comparison of the central metabolism ... 74 Central carbon metabolism ... 74 Nitrogen metabolism ... 77 Energy metabolism ... 78 Cell mobility and chemotaxis ... 78 Diversity of predicted stress tolerance mechanisms ... 78 Maintaining the intracellular pH homeostasis ... 78 Coping with high metal loads ... 79 Oxidative stress management ... 79 IDENTIFICATION OF POTENTIAL DRIVING FORCES OF GENOME EVOLUTION ... 80 Prediction of mobile genetic elements ... 81 Linking the differences in the predicted metabolic profiles to the genome architectures ... 82 Gene cluster associated with flagella formation and chemotaxis in “F. myxofaciens” ... 84 Gene clusters associated with the utilization of alternative nitrogen sources ... 86 Gene cluster associated with carboxysome formation in “F. myxofaciens” and OTU-2 strain JA12 ... 87 Putative genomic islands in the OTU-strain JA12 ... 89 CRISPR/Cas in “F. myxofaciens” Z-31: a defense mechanism against foreign DNA ... 91 DISCUSSION ... 92 THE COMPARISON OF THEIR METABOLIC PROFILES INDICATES THE EXISTENCE OF OTU- AND STRAIN-SPECIFIC FEATURES ... 92 GENOME EVOLUTION OF THE “FERROVUM” STRAINS APPEARS TO BE DRIVEN BY HORIZONTAL GENE TRANSFER AND GENOME REDUCTION ... 94 Horizontal gene transfer ... 94 Mechanisms of genome reduction ... 95 CONCLUDING REMARKS ... 98 ACKNOWLEDGMENTS ... 98 AUTHOR CONTRIBUTIONS ... 98 CHAPTER V ... 99 ABSTRACT ... 100 INTRODUCTION ... 100 METHODS ... 102 CULTIVATION OF THE “FERROVUM”-CONTAINING MIXED CULTURE JA12 ... 102 Up-scaling of pre-cultures for the transcriptome study ... 103 Experimental setup of the transcriptome study ... 103 Cell harvest from large culture volumes ... 106 EXTRACTION OF TOTAL RNA ... 106 LIBRARY CONSTRUCTION AND SEQUENCING ... 107 DATA ANALYSIS ... 107 Processing of raw data ... 107 Quantification of gene expression levels ... 108 Functional analysis ... 108 RESULTS ... 108 CULTIVATION OF THE MIXED CULTURE JA12 IN THE MULTIPLE BIOREACTOR SYSTEM ... 108 Growth monitoring ... 108 Microbial composition ... 111 RNA SEQUENCING (RNA-SEQ) ... 112 FUNCTIONAL CATEGORIZATION OF EXPRESSED GENES ... 113 Functional assignment of highly expressed genes ... 117 Functional assignment of poorly expressed genes ... 121 COMPARISON OF EXPRESSION LEVELS OF GENES PREDICTED TO BE INVOLVED IN OXIDATIVE STRESS MANAGEMENT ... 122 DISCUSSION ... 124 METABOLIC PATHWAYS RELEVANT UNDER CULTURE CONDITIONS MIMICKING THE NATURAL CONDITIONS IN THE MINE WATER TREATMENT PLANT ... 125 Novel insights into the energy metabolism of “Ferrovum” sp. JA12 ... 125 Insights from poorly expressed genes ... 126 VARIATION OF GENE EXPRESSION PATTERNS UNDER THE DIFFERENT CONDITIONS ... 128 EVALUATION OF THE EXPERIMENTAL SET-UP INVOLVING THE MULTIPLE BIOREACTOR SYSTEM ... 129 CONCLUDING REMARKS: SIGNIFICANCE OF THE PRESENT TRANSCRIPTOME STUDY ... 130 ACKNOWLEDGMENTS ... 131 AUTHOR CONTRIBUTIONS ... 131 CHAPTER VI ... 133 ABSTRACT ... 133 EXTENDED INSIGHTS INTO THE FERROUS IRON OXIDATION IN BETAPROTEOBACTERIA ... 133 MECHANISMS OF PHYLOGENETIC AND METABOLIC DIVERSIFICATION WITHIN THE GENUS “FERROVUM” ... 136 INFERRED ROLES OF “FERROVUM” SPP. IN THE MICROBIAL NETWORK OF THE MINE WATER TREATMENT PLANT ... 138 PERSPECTIVES ... 143 REFERENCES ... 145 SUPPLEMENTARY MATERIAL ... 170 DATA DVD ... 170 SUPPLEMENTARY MATERIAL FOR CHAPTER III ... 171 NUCLEOTIDE ACCESSION NUMBERS ... 171 PHYLOGENETIC ANALYSIS ... 171 GENOME PROPERTIES ... 173 NUTRIENT ASSIMILATION ... 174 Carbon metabolism ... 174 FERROUS IRON OXIDATION ... 176 HORIZONTAL GENE TRANSFER ... 179 SUPPLEMENTARY MATERIAL FOR CHAPTER IV ... 180 PHYLOGENETIC ANALYSIS ... 180 ASSIGNMENT OF PROTEIN-CODING GENES TO THE COG CLASSIFICATION ... 180 COMPARISON OF THE CENTRAL METABOLISM ... 181 Predicted metabolic potential of the four “Ferrovum” strains ... 181 Genes predicted to be involved in the central metabolism, energy metabolism, cell motility and stress management in the four “Ferrovum” strains ... 183 PREDICTED MOBILE GENETIC ELEMENTS IN THE GENOMES OF THE FOUR “FERROVUM” STRAINS ... 184 THE FLAGELLA AND CHEMOTAXIS GENE CLUSTER ... 184 THE UREASE GENE CLUSTER ... 185 THE CARBOXYSOME GENE CLUSTER ... 186 PUTATIVE GENOMIC ISLANDS IN “FERROVUM” SP. JA12 ... 187 Gene content of the genomic islands ... 187 Flanking sites of the putative genomic islands 1 and 2 ... 188 SUPPLEMENTARY MATERIAL FOR CHAPTER V ... 189 ORGANIZATION AND OPERATION OF THE LABFORS 5 MULTIPLE BIOREACTOR SYSTEM ... 189 INVESTIGATION OF THE MICROBIAL COMPOSITION IN THE IRON OXIDIZING MIXED CULTURE JA12 ... 192 SUPPLEMENTARY DATA OF THE TRANSCRIPTOME DATA ANALYSIS ... 193 RNA-Seq statistics ... 193 Expression strength of protein-coding genes ... 194 Expression of genes involved in carboxysome formation ... 197 Expression of a ribosomal proteins-encoding gene cluster ... 199 Expression of a gene cluster presumably involved in ferrous iron oxidation ... 202 Lowest expressed genes ... 205 Expression of genes predicted to be involved in oxidative stress response ... 206 ACKNOWLEDGMENTS ... 208 COLLEAGUES ... 208 ERFOLGSTEAM “JUNGE FRAUEN AN DIE SPITZE” (“YOUNG WOMEN TO THE TOP“) ... 208 FAMILY AND FRIENDS ... 209 FUNDING ... 209 CURRICULUM VITAE ... 210 LIST OF PUBLICATIONS ... 212 RESEARCH ARTICLES ... 212 CONFERENCE PROCEEDINGS ... 212 ORAL PRESENTATIONS AND POSTERS ... 213