The studies in this thesis were conducted with Baltic Sea protists. The Baltic Sea is a subarctic brackish-water basin that partially freezes over every winter. If the salinity of parent water is higher than 0.6, the forming ice has a semi-solid structure with solid ice crystals and saline water (brine) channels. The brine channels offer habitats for small-sized organisms. Due to the low salinity of the Baltic Sea, the brine channels are small, and therefore, the Baltic Sea ice eukaryotic community is dominated by protists. Studies on Baltic sea-ice biology have been accumulating since the 1980’s, but there are still gaps on knowledge; for example, what protistan species and how many there are associated with sea ice. In this thesis, morphological, molecular and ecological information was combined to delineate species of a Baltic Sea cryptomonad, haptophyte and dinoflagellate. Protistan community composition in Baltic Sea ice was assessed with environmental sequencing, and diversity estimates were compared in different types of ice. The taxonomic and environmental-sequencing studies were linked using the gathered taxonomic information to evaluate the accuracy of the molecular diversity-measurement method.

A new cryptomonad species, Rhinomonas nottbecki, was described based on morphological characters distinguished by light- and electron-microscopy together with molecular evidence from 18S rRNA gene and ITS region. The same approach was applied to the identification of the alternate stage Prymnesium polylepis (Haptophyta), which bloomed in the whole Baltic Proper during autumn–spring 2007–2008. Also, a novel cold-water and sea-ice associated dinoflagellate subspecies, Heterocapsa arctica subsp. frigida, was described. Environmental 18S-rRNA-gene sequencing revealed that the richest eukaryotic lineages inhabiting the Baltic Sea ice were ciliates, cercozoa, dinoflagellates and diatoms. The different developmental stages and types of ice had different community composition. Protistan richness was higher in ice than water even though water included more divergent lineages. The Baltic fast ice had higher richness than pack and drift ice.

The results of this thesis showed that there remains novelty to be described in the Baltic Sea, and what we know about the protistan community in Baltic Sea ice now is very incomplete. Although the environmental sequencing produced data that met the requirements of calculation of comparable diversity indices (all taxa defined at the same level), revealed cryptic taxa, and gave higher protistan richness than basic light microscopy of fixed samples, the lack of taxonomic detail was not restricted to the light-microscopic surveys but was also a result of the environmental-sequencing approach. This was shown when the environmental-sequencing approach was applied on the 18S-rRNA-gene data of the cryptomonad family Pyrenomonadaceae and the haptophyte genus Prymnesium. Only one Pyrenomonadaceae and two Prymnesium OTUs were found although both data sets included 15 distinct taxa. Errors in environmental sequencing and alignment make the use of high similarity levels in the OTU definition questionable, and the variability in the 18S rDNA is not equal within different eukaryotic lineages. Consequently, use of lower similarity level (97 %) is justifiable in the environmental-sequencing, but the approach used gave conservative estimates of the protistan richness in the Baltic Sea ice.

The overall conclusion is that we need to apply all available techniques when assessing the diversity of protists, as each technique provides a biased perspective on nature. A labor intensive taxonomic approach that includes the study of live cells by light microscopy, detailed morphological description based on electron microscopy and phylogenetic analysis of suitable genetic markers gives us the best chance of finding out how many different species of protists live within the Baltic Sea ice or any other environment, and what they do.

This thesis was undertaken to investigate the potential of tailings from apatite ore beneficiation at the Siilinjärvi phosphate mine, Finland, in the dephosphorization of sewage and in the remediation of metal-contaminated areas. The material is a mixture of minerals, mainly phlogopite [KMg3(Si3Al)O10(OH)2] and calcite (CaCO3), accompanied by apatite [Ca5(PO4)3F] residues. Based on the versatile chemical properties, this geomaterial was hypothesized to act as a sorbent for P and Pb, rendering the tailings a potential agent for environmental remediation. A part of the original tailings material was artificially weathered by treating with a strong acid to create reaction-active aluminium (Al) and iron (Fe) (hydr)oxide sites. Some of the acidified material was further subjected to partial neutralization by treating with a strong base to precipitate any metals dissolved from the mineral structure during the acidification. Furthermore, all of the tailings materials were sieved into two particle-size fractions somewhat differing in their mineralogical composition and investigated as separate amendments.

The ability of the tailings to retain P and Pb from aqueous solutions as well as the tailings-induced changes in the Pb retention capacity of a mineral soil were studied by means of an isotherm technique. A sequential fractionation procedure was undertaken to investigate (a) the distribution of inherent and added P between various chemical pools in the tailings and (b) the tailings-induced changes in the distribution of Pb between various chemical pools in a mineral soil artificially contaminated with Pb as well as in an organic shooting range soil contaminated with pellet-derived Pb. Because the toxicity of dissolved Pb depends on its chemical speciation, the tailings-induces changes in the chemical speciation of water-extractable Pb in contaminated shooting range soil was tested separately by means of a cation exchange resin. The tailings retained both P and Pb efficiently. The removal of soluble P was primarily due to specific sorption by Al and Fe (hydr)oxides and possibly to retention to calcite. Lead sorption by the untreated tailings was a combination of various sorption mechanisms taking place simultaneously, primarily through precipitation and surface complexation. All tailings materials increased the Pb sorption capacity of a mineral soil and transferred Pb from the NH4NO3-extractable pool to the more strongly bound forms. In a contaminated shooting range soil, the pellets were found to undergo continuous weathering processes that released Pb into the soil. Amending the soil with the untreated tailings (a) reduced the solubility of the pellet-derived Pb through the formation of sparingly soluble fluorpyromorphite and cerussite, (b) reduced the bioavailability of Pb by transferring it from the water-soluble and NH4NO3-extractable pools into the NaOH-extractable one and (c) transferred the most toxic cationic Pb species to the less toxic non-cationic form.

The results suggest that the tailings may serve as an agent for dephosphorization of sewage and for Pb immobilization in polluted soil. The sorption properties of the material may be further optimized by chemical and physical pre-treatments. At present, the tailings material represents an uneconomic fraction of the ore deposit, but its components may render it a natural, environmentally sound and cost-effective remediation agent.

The aim of this study was to develop novel instrumental techniques for the determina-tion of organic compounds without any analyte preselection and to identify unknown anthropogenic contaminants in different water matrices. The methods developed were based on analytical separation, using gas and liquid chromatography combined with accurate mass measurement using time-of-flight mass spectrometry. The data produced were then processed with a deconvolution program to locate the chromatographic peaks and to extract their mass spectra. The measured accurate masses were then used to confirm the elemental compositions of the detected ions. The identification processes were validated, using spiked water samples, and finally the methods were applied to the identification of organic xenobiotics from wastewater effluent, stormwater, surface water and landfill leachate samples.

The results showed that analysis using time-of-flight mass spectrometry enables screening of large analyte groups without previous information on sample composition. The most comprehensive knowledge is yielded by analysing the sample with both gas and liquid chromatography. In many cases, tentative compound identification can be obtained if the deconvoluted spectra and accurate mass data are complemented with information, e.g. from spectral libraries and peak isotope patterns. This tentative identifications must, however, always be confirmed with a pure standard compound. The main limitations of the methods were related to insufficient features of the deconvolution program used. Most of the data-processing stages had to be performed manually or visually, which slows down the data processing and hinders their applicability, especially with large sample sets. Dozens of compounds were tentatively identified from water samples and several of them were also confirmed with a standard compound. The highest numbers of compounds were identified from wastewater effluent, stormwater and landfill leachate samples. The results confirmed the fact that anthropogenic waste streams are an important route for organic xenobiotics into the environment. Since the future volumes of chemicals will increase, the control and efficient treatment of these fluxes becomes evermore essential.

Cellular uptake has proven to be even more challenging. Oligonucleotides are internalized into cells by endocytosis, after which they often remain trapped in endosomes. Therefore, it would be advantageous to develop delivery vectors capable of bypassing endocytic routes of uptake or enhancing endosomal escape. Cell-penetrating peptides, for example, exploit several mechanisms of uptake, some of which lead to rapid entry without endosomal localization. In addition, encouraging results have been achieved using liposomes, gold nanoparticles, and other nanocarriers, which also shield the oligonucleotide from degrading enzymes.

The aim of this work was to improve the in vitro delivery of oligonucleotides by employing chemical modifications and nanoparticle carriers. The synthesis of the compounds, their characterization by various analytical methods, and the evaluation of biological effects are described. Antisense oligonucleotides covalently linked to cell-penetrating peptides via convergent conjugation displayed improved cellular uptake but failed to inhibit reporter genes due to endosomal entrapment in cells. Circular oligonucleotides exhibited enhanced selectivity of mismatch detection and increased stability in biological fluids compared to linear oligonucleotides. Altogether 44 compounds were analyzed by electrospray ionization mass spectrometry and liquid chromatography–mass spectrometry, which were found to be excellent methods for the characterization of modified oligonucleotides. Finally, we synthesized cationic gold nanoparticles modified with a Tat-related peptide, which did not adversely affect cell viability and effectively delivered short interfering RNA into cells as non-covalent complex.

The aim of this theses was to study the diversity, ecology and evolutionary relationships of mycocalicioid fungi, especially the resinicolous species. The study is based on specimens collected from different parts of the world, and analyzed using both molecular and morphological methods.

In this study seven new resinicolous Chaenothecopsis species are described from boreal North America and Europe, tropical Africa, and temperate China, four from angiosperm exudates and three from conifer resins. Two newly found fossils of Chaenothecopsis from Eocene and Oligocene ambers are described and their relations to extant species are elucidated. The phylogenetic relationships of several species and lineages, and the evolution of the resinicolous ecology are discussed in considerable detail. In addition, new morphological and chemical characters that can be used in future taxonomic studies are described.

Occurrence of intervertebral disc calcifications was higher in Finnish miniature Dachshunds than previously reported; 76% of longhaired and 87% of wirehaired variety had at least one calcification. Therefore screening protocol excluding every dog with calcifications from breeding would not be possible. The number of calcifications varied between none and 13, and therefore radiographic screening, together with favouring dogs with fewer calcified discs in breeding, is suggested.

Accuracy of the Finnish screening protocol for mild (grade 1) ED differed between the two breeds studied. In Belgian shepherd dogs, the Finnish protocol based mainly on evaluation of osteophytes was inaccurate, as 47% of the joints free of ED were incorrectly graded as dysplastic and 40% of dysplastic joints were graded as normal. On the other hand, assessment of the radiographic signs indicative of medial coronoid process disease proved to be accurate in this breed; sensitivity and specificity of blurring of the cranial border of the medial coronoid process were 80% and 90%, respectively. In Labrador retrievers, the Finnish protocol proved to be accurate, as sensitivity of grading was 79% and specificity 92%. Osteophytes seen in the supplemental craniocaudal oblique radiographic projection was the most reliable sign of ED in Labrador retrievers with sensitivity of 93% and specificity of 92%. Adding this view might be a valuable addition to the screening protocol. Labrador retrievers with grade 1 ED should not be used for breeding since most of them have ED, as was clearly demonstrated in the study.

LTV was common in Finnish German shepherd dogs studied with occurrence of 40%. A laterolateral (LL) radiograph appeared to be a useful supplement to the currently used ventrodorsal (VD) radiograph in diagnosis of LTV. Sensitivity of the diagnosis based on the VD projection alone was 90% increasing to 100% when LL projection was used together with the VD projection. Additionally, diagnosis of the eighth lumbar vertebra (L8) was only possible based on the LL projection. It is therefore suggested for inclusion in the radiographic screening protocol. The L8 resembled an LTV, and is therefore proposed as a part of the LTV-complex and for consideration when screening for LTV.

The fear of soil preparation accelerating GHG emissions, particularly CO2, from peat into the atmosphere appears unwarranted at least on nutrient-poor, boreal forestry-drained peatland sites. The overall climatic impact of soil preparation, in the forms of mounding and scalping, three years after application expressed in terms of CO2 equivalents (100-year GWP), was neutral compared to leaving soil unprepared.

The core findings of this research support mounding as the best alternative on nutrient-poor, drained peatland sites when the goal is to maximize the regeneration success of Scots pine after clearcutting with minimal impact on soil GHG emissions. In the future, development of soil preparation methodology is particularly deserving of further attention. While it may not be the sexiest research topic in the worldwide rat race of the modern day, it is nonetheless of substantial importance in a country highly specialized not only in the utilization but also the rejuvenation of wood resources on drained peatlands.

Keywords: forestry-drained peat soil, clearcutting, mounding, scalping, CO2, CH4, and N2O fluxes, drought and waterlogging stress

In addition to chemical testing, environmental monitoring of endocrine disrupting compounds is important to assess the level of exposure and possible adverse effects of chemicals on humans and wildlife. Chemical analysis methods used in environmental monitoring are sensitive, but they are also laborious, expensive, and require specialized instruments. Consequently, robust biological methods have become valuable tools to measure endocrine disrupting potency of chemicals and environmental samples. For this purpose, several Saccharomyces cerevisiae yeast cell-based bioreporters utilizing different nuclear receptors have been developed. Yeast-cell-based bioreporter assays have several advantages in environmental analytics. In addition to being inexpensive, they are particularly useful in determining the bioavailability of contaminants. Yeast is also very tolerant towards toxicity of different sample matrices. Yeast nuclear receptor bioreporter assays have been used to determine the total hormonal activity of samples containing unknown mixture of chemicals. However, these assays cannot identify the chemicals in the sample, and thus, monitoring of a single chemical has not been possible.

Many cell-based assays have already been adapted to high throughput screening plate formats of 384 and 1536 wells and even higher. However, nearly all yeast nuclear receptor bioreporter assays are still performed in 96-well plates. Consequently, yeast bioreporter assays should be adapted to automated liquid handling and high density well plates to enable screening of large chemical libraries and high number of samples.

In this thesis study, a yeast nuclear receptor bioreporter assay for specific detection of a single chemical, bisphenol A (BPA) was developed. The creation of the BPA-targeted receptor included application of a oligonucleotide-based mutation method and a positive-negative genetic selection method for human estrogen receptor α (publication I). Chemical specificity of the BPA-targeted receptor (BPA-R) bioreporter assay was characterized, and its use demonstrated with chemical mixture and waste water samples (publication II). In addition, the existing battery of yeast bioreporters was adapted to automated liquid handling and high density 384 and 1536 well plates to meet the requirements of high throughput screening (publication III). Finally, a new yeast-based bioreporter utilizing a chimeric human retinoid X receptor was constructed and characterized (publication IV). This bioreporter can be used to measure organotin compounds such as tributyl tin in environmental samples.