Few genomic and metagenomic studies have focused on freshwater systems in the last years. Most of the studies carried out on these particular environments so far rely on microscopy, physiology, phenotypic observations, individual genes and 16S rRNA sequencing. Here, we shed light on microbial communities from oligotrophic and mesotrophic freshwater systems using high-throughput deep sequencing metagenomics and genome-resolved metagenomics. We have focused on the study of ubiquitous and cosmopolitan microbial groups from two temperate Spanish reservoirs (Tous, Amadorio). Among these, we studied freshwater picocyanobacteria from Synechococcus and Cyanobium genera, which so far have not been well characterized at the genomic level, compared to the marine representatives. In particular, we were able to isolate two of the most abundant picocyanobacteria from Tous reservoir, which were previously studied via metagenomics. These picocyanobacteria are not only abundant in this reservoir but are widely distributed in different freshwater and brackish systems. In this work we also shed light on some of the first freshwater representatives of the phylum Verrucomicrobia, that are ecologically uncharacterized in freshwater systems about which relatively little is known. We discovered a wide range of metabolisms in these microbes, ranging from nitrogen fixation and photoheterotrophy via rhodopsin pumps to important contributions in the degradation of recalcitrant matter and polysaccharides. We also include the first metagenomic study of the microbial communities under the ice waters of the largest (by volume) ultraoligotrophic lake in the world, Lake Baikal. This study has provided a first glimpse and a particular microbial composition on the sub-ice, having found an unusual fraction of Verrucomicrobia and new microbial lineages from many typical freshwater phyla, including the first freshwater representative of the groups I/II of SAR11 lineage and novel genomes of Proteobacteria, Thaumarchaeaota, Gemmatimonadetes, Cyanobacteria, Planctomycetes, Bacteroidetes, Acidobacteria, Nitrospirae, Verrucomicrobia or Actinobacteria.

Genetically modified (GM) crops were introduced in South Africa in 1989 and commercially available by 1998. Legislation to control the use of GM crops was only implemented in 1999, with the genetically modified organisms (GMO) act (15 of 1999). In 2012 2.9 million ha of GM crops were planted in South Africa alone. GM Crops, such as Bt maize, are promoted as safer for the environment since no chemical pesticides are needed. However, recently GM crops have been making headlines as more and more studies find adverse effects of these crops on non-target organisms. The effects on aquatic environments have not yet been fully determined, even though traces of Bt residue have been found in water systems surrounding agricultural lands. The aim of this study was to establish the effects of the Bt toxin on fecundity, development and growth of Bulinus tropicus, a freshwater snail. The experiment made use of a static renewal tests to expose B. tropicus to 50 cm2 Bt maize and cotton leaves in 900 ml of synthetic freshwater. The snails were exposed for the duration of one full life cycle (embryo to adult). Endpoints measured included the development, growth, fecundity, and deformities of the reproductive organs. The results obtained showed retarded development and low embryo survival when the snails were exposed to cotton leaves, irrespective of the presence or absence of Bt, indicating to the possibility of trace residues of chemical pesticides may have been present on the leaves. Initial stimulated growth of hatchlings was observed for both Bt cotton and maize exposures, but after sexual maturity has been reached, ‘surplus’ energy was probably shared between growth and fecundity, resulting in a reduction of growth rate. Energy is gained from their diet, thus a sub-optimal diet would result in less energy available to functions such as growth and fecundity. Signs of developmental instability were found in the formation of the shell opening of the snails exposed to Bt. Fecundity decreased significantly after snails had been exposed to Bt maize / cotton leaves. No differences were found in the penis sheath-preputium length ratio, indicating that Bt had no deleterious effects on the reproductive organs. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2014

Genetically modified (GM) crops were introduced in South Africa in 1989 and commercially available by 1998. Legislation to control the use of GM crops was only implemented in 1999, with the genetically modified organisms (GMO) act (15 of 1999). In 2012 2.9 million ha of GM crops were planted in South Africa alone. GM Crops, such as Bt maize, are promoted as safer for the environment since no chemical pesticides are needed. However, recently GM crops have been making headlines as more and more studies find adverse effects of these crops on non-target organisms. The effects on aquatic environments have not yet been fully determined, even though traces of Bt residue have been found in water systems surrounding agricultural lands. The aim of this study was to establish the effects of the Bt toxin on fecundity, development and growth of Bulinus tropicus, a freshwater snail. The experiment made use of a static renewal tests to expose B. tropicus to 50 cm2 Bt maize and cotton leaves in 900 ml of synthetic freshwater. The snails were exposed for the duration of one full life cycle (embryo to adult). Endpoints measured included the development, growth, fecundity, and deformities of the reproductive organs. The results obtained showed retarded development and low embryo survival when the snails were exposed to cotton leaves, irrespective of the presence or absence of Bt, indicating to the possibility of trace residues of chemical pesticides may have been present on the leaves. Initial stimulated growth of hatchlings was observed for both Bt cotton and maize exposures, but after sexual maturity has been reached, ‘surplus’ energy was probably shared between growth and fecundity, resulting in a reduction of growth rate. Energy is gained from their diet, thus a sub-optimal diet would result in less energy available to functions such as growth and fecundity. Signs of developmental instability were found in the formation of the shell opening of the snails exposed to Bt. Fecundity decreased significantly after snails had been exposed to Bt maize / cotton leaves. No differences were found in the penis sheath-preputium length ratio, indicating that Bt had no deleterious effects on the reproductive organs. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2014

In Australia, significant effort goes into reducing the amount of nitrogen and phosphorus entering inland waters from point sources. However, little is known of the extent to which riparian organic matter may act as a source of these nutrients. Also, whilst the relationships between the nitrogen, phosphorus and carbon cycles are broadly known, there is little quantitative data regarding the release of these elements from Australian riparian organic matter and their subsequent microbial mineralisation within aquatic environments. In particular, comparatively little is known of their comparative role in nutrient and organic matter cycling within anoxic zones, and the influence that different riparian organic matter may have on stream water quality. This lack of such data presently hampers the ability of water managers to make educated decisions regarding the management of riparian zones in Australia. In order to improve understanding in this area, a combination of laboratory and in situ experiments were carried out in order to compare the abiotic release and aerobic/ anaerobic mineralisation of leaf derived dissolved organic carbon (DOC), dissolved nitrate/nitrite (NOx) and soluble reactive phosphorus (SRP) under different environmental conditions. Four plants common to Australian riparian zones were investigated: two native species, Eucalyptus camaldulensis (gum) and Phragmites australis (common reed), and two exotic species, Salix babylonica (willow) and Lolium multiflorum (rye grass). After 30 days, formaldehyde inhibited 1g willow and rye grass extracts contained the most SRP (0.7 mg/L), whilst gum extracts contained 0.3 mg/L and common reed 0.1 mg/L of SRP.Willow and rye grass abiotically released twice as much NOx than gum and common reed, although concentrations were only between 0.05-0.1 mg/L. Gum and common reed released the most DOC per gram of leaf matter (14 and 12 mmol/g of leaf matter respectively), but based on the initial carbon content of each leaf type, the largest percentage contributor of DOC under abiotic conditions was common reed and rye grass (both 38% mass/mass), with gum (33% mass/mass) and willow (30% mass/mass) being smaller contributors. The most bioavailable DOC was released by rye grass and common reed, with between 83 and 94% of this DOC microbially mineralised after 30 days in oxic conditions. When conditions were not inhibited, microbial growth was evident almost immediately in willow, rye grass and common reed leaf extracts. However, microbial growth was suppressed for the first 48 hours in gum leaf extracts. After this suppression period, the rate of DOC mineralisation was equal in willow and gum leaf extracts (0.1 day-1). Under anoxic conditions, the rate and extent of DOC mineralisation of willow and gum leaves depended on the type of electron acceptor provided. Added nitrate and iron III enhanced the mineralisation of both willow and gum leaves relative to no terminal electron acceptors (from zero to 0.01-0.04 and 0.002- 0.004 moles/day respectively), but added sulphate only enhanced the mineralisation of gum leaves (0.04 moles/day). When no additional electron acceptors were provided, particulate leaf mineralisation was more extensive under oxic than anoxic conditions. However, the mineralisation of leaf derived DOC were the same regardless of oxygen availability, and after 35 days in either condition the percentage of leaf DOC mineralised for each leaf type was of the order common reed > rye grass > willow > gum. All the leaf types tested were able to sustain the caddis fly larvae Triplectides australis under controlled laboratory conditions, and survival rates were high using all four leaf types as a food source. Triplectides australis did not significantly increase the amount of DOC released from each type of leaf matter, but they did consistently increase the proportion of simple carbohydrates present within the DOC fraction. The results of these experiments suggest that changes to riparian vegetation, particularly from the native to exotic species used in this study, will inherently alter in-stream concentrations of dissolved carbon and nutrients (particularly SRP). This potentially will affect in-stream, hyporheic and subsurface processes, particularly in areas where surface water flow is low and riparian leaf inputs are high.