Browsing by Issue Date

Sort by: Order: Results:

Now showing items 1-20 of 723
  • Juuri, Juuso (Helsingin yliopisto, 2015)
    Kainate-type of ionotropic glutamate (KA) receptors are associated with the modulation of neuronal excitability, synaptic transmission, and activity of neuronal networks. They are believed to have an important role in the development of neuronal connections. In this thesis, the role of KA receptors in the early brain development was assessed by conducting in vitro electrophysiological recordings from individual neurons at CA3 region in acute slices of neonatal rodent hippocampi. It was found that activation of separate KA receptor populations promoted action potential firing in both glutamatergic pyramidal neurons and GABAergic interneurons. The receptors in pyramidal neurons displayed a high affinity for agonist kainate, appeared to lack subunit GluK1, and promoted spontaneous firing of pyramidal neurons without depolarizing them. The receptors in interneurons contained subunit GluK1 and their activation suppressed afterhyperpolarizing current of medium duration (ImAHP). Receptors in both neuron types appeared to be activated tonically by ambient glutamate, suggesting that their physiological role may be to act as a modulatory mechanism sensitive to changes in extracellular glutamate concentration. Changes in activity of neurons at CA3 by activation of KA receptors were reflected on the network level. Promotion of pyramidal cell firing by pharmacological activation of high-affinity KA receptors lead to enhanced glutamatergic drive and generation of network bursts in the CA3 region. The ImAHP in interneurons was also suppressed by apamin, a blocker of SK potassium channels that mediate majority of this current, and apamin enhanced generation of network bursts. This suggests that also KA receptor mediated regulation of ImAHP may modulate network activity. It was also found that there was an interaction between KA receptors and ethanol in the modulation of hippocampal network: ethanol decreased the occurrence of the network bursts at postnatal days 1 (P1) and P10, whereas it increased bursting at P5. The network effects of ethanol were partially or completely counteracted by specific pharmacological block of GluK1 subunit-containing KA receptors. The findings disclose that via regulation of activity of individual neurons, KA receptors are capable of robust modulation of network activity in immature hippocampus. Additionally, exogenous agents affecting KA receptors may perturb activity dependent developmental processes that are central for the synaptic development. The results shed light on the mechanisms underlying development of hippocampal connectivity, and may help to understand early pathologies of the brain that have developmental origins.
  • Kankainen, Matti (Helsingin yliopisto, 2015)
    Lactobacilli are generally harmless gram-positive lactic acid bacteria and well known for their broad spectrum of beneficial effects on human health and usage in food production. However, relatively little is known at the molecular level about the relationships between lactobacilli and humans and about their food processing abilities. The aim of this thesis was to establish bioinformatics approaches for classifying proteins involved in the health effects and food production abilities of lactobacilli and to elucidate the functional potential of two biomedically important Lactobacillus species using whole-genome sequencing. To facilitate the genome-based analysis of lactobacilli, two new bioinformatics approaches were developed for the systematic analysis of protein function. The first approach, called LOCP, fulfilled the need for accurate genome-wide annotation of putative pilus operons in gram-positive bacteria, whereas the second approach, BLANNOTATOR, represented an improved homology-based solution for general function annotation of bacterial proteins. Importantly, both approaches showed superior accuracy in evaluation tests and proved to be useful in finding information ignored by other homology-search methods, illustrating their added value to the current repertoire of function classification systems. Their application also led to the discovery of several putative pilus operons and new potential effector molecules in lactobacilli, including many of the key findings of this thesis work. Lactobacillus rhamnosus GG is one of the clinically best-studied Lactobacillus strains and has a long history of safe use in the food industry. The whole-genome sequencing of the strain GG and a closely related dairy strain L. rhamnosus LC705 revealed two almost identical genomes, despite the physiological differences between the strains. Nevertheless of the extensive genomic similarity, present only in GG was a genomic region containing genes for three pilin subunits and a pilin-dedicated sortase. The presence of these pili on the cell surface of L. rhamnosus GG was also confirmed, and one of the GG-specific pilins was demonstrated to be central for the mucus interaction of strain GG. These discoveries established the presence of gram-positive pilus structures also in non-pathogenic bacteria and provided a long-awaited explanation for the highly efficient adhesion of the strain GG to the intestinal mucosa. The other Lactobacillus species investigated in this thesis was Lactobacillus crispatus. To gain insights into its physiology and to identify components by which this important constituent of the healthy human vagina may promote urogenital health, the genome of a representative L. crispatus strain was sequenced and compared to those of nine others. These analyses provided an accurate account of features associated with vaginal health and revealed a set of 1,224 gene families that were universally conserved across all the ten strains, and, most likely, also across the entire L. crispatus species. Importantly, this set of genes was shown to contain adhesion genes involved in the displacement of the bacterial vaginosis-associated Gardnerella vaginalis from vaginal cells and provided a molecular explanation for the inverse association between L. crispatus and G. vaginalis colonisation in the vagina. Taken together, the present study demonstrates the power of whole-genome sequencing and computer-assisted genome annotation in identifying genes that are involved in host-interactions and have industrial value. The discovery of gram-positive pili in L. rhamnosus GG and the mechanism by which L. crispatus excludes G. vaginalis from vaginal cells are both major steps forward in understanding the interaction between lactobacilli and host. We envisage that these findings together with the developed bioinformatics methods will aid the improvement of probiotic products and human health in the future.
  • Anastasina, Maria (Helsingin yliopisto, 2015)
    Viruses infect all domains of life. They establish complex interactions with their host cells to subvert and hijack multiple cellular processes and warrant their own replication. Understanding virus-host interactions is critical to control spread of pathogenic viruses, develop vaccines and search for antivirals. Besides that, understanding virus-host interactions allows deciphering complex cellular processes and provides useful tools for biotechnology. My research is dedicated to influenza A virus, an important pathogen that infects humans worldwide, represents a constant health care threat and elicits continuous efforts to control the human spread of the disease. Influenza A expresses a non-structural protein NS1 that is a key regulator of viral interactions with the host cell and an important virulence factor. Versatile functions of NS1 modulate multiple cellular functions to secure viral replication. This work addresses several aspects of NS1-mediated modulation of core cellular processes. We discovered that NS1 binds to dsDNA and inhibits transcription of cellular genes, thus limiting antiviral responses. We found that NS1 secures general protein synthesis and mapped several residues within NS1 that are essential for this function. Further, we showed that the length of C-terminal ``tail'' of NS1 is essential for control of cellular antiviral responses and virus pathogenicity. The presented results increase the understanding of influenza A virus-host interactions and can be further utilized in the search for antivirals and vaccine development. In addition, this work provides a biotechnological application of influenza A NS1 protein for improvement of cell-free translation system.
  • Valtanen, Marjo (Helsingin yliopisto, 2015)
    Urbanization is causing a substantial increase in impervious surfaces and thereby alterations in the hydrological cycle, e.g. by increasing runoff volumes and intensities that can lead to urban flooding. Furthermore, urban runoff is considered as one of the most important surface water pollutant sources. The characteristics of urban runoff are known to vary greatly between land use types and climates. However, in cold climates, only a few studies have tackled urban runoff phenomena and these studies rarely include catchment-based and long-term monitoring on both runoff quantity and quality. Yet, to develop successful urban runoff management practices, knowledge on runoff characteristics from various urban sites that are monitored for several pollutants throughout the year is required. The aim of this thesis was to fill gaps in our knowledge on the effects of urbanization on runoff characteristics and to study the mechanisms affecting runoff generation and pollutant transport during vastly divergent seasons in cold climates. This research was conducted in the city of Lahti, southern Finland, and comprised three urban catchments of varying imperviousness: High catchment (city centre, total impervious area [TIA], 89%), Intermediate catchment (city centre, 62%) and Low catchment (residential, 19%). At each catchment, continuous measurements on runoff flow rate and precipitation were carried out for a period of two years. In addition, runoff samples were taken to determine concentrations for total nitrogen (tot-N), total phosphorus (tot-P), total suspended solids (TSS), total and dissolved heavy metals (Zn, Cu, Cr, Mn, Al, Co, Ni, Pb) and total organic carbon (TOC). I showed that in cold climates, annual runoff volumes, most pollutant loads and some of the pollutant concentrations increased with increasing imperviousness. Furthermore, urbanization altered runoff generation (volume, intensity, duration, number of events) more strongly during the warm than the cold period. However, when the summer was dry and the winter wet, the effects of urbanization on annual runoff volumes diminished most likely due to snow removal. The spring snowmelt period began earlier and occurred as several events at the city centres in comparison to the low-development catchment. In contrast with patterns observed during warm periods, cumulative runoff volumes during cold periods decreased with increasing urbanization when TIA reached about 60%. Nevertheless, urbanization increased both warm and cold period loads for most pollutants. However, the mechanisms controlling runoff event quality were distinct between seasons: during cold periods event loads and concentrations were mainly increased by long event durations, yet, during warm periods the peak intensity of runoff was the main increasing factor whereas concentrations tended to decrease as event durations increased. Because of snow removal in the High catchment with TIA of 89%, snowmelt volumes during spring were not affected by the amount of precipitation. In addition, at the High catchment, warm periods contributed more to annual volumes than cold periods, contrary to the pattern observed at the Low catchment. Irrespective of land use, the largest seasonal pollutant loads and concentrations for several pollutants occurred during spring. However, the importance of the warm season in annual pollutant load generation increased with increasing urbanization. This study provides updated data on pollutant unit loads, especially for city centres and for heavy metals. This thesis - applying a catchment-based approach - also showed, for the first time how the seasonality of runoff volumes and loads change at highly impervious city centres.
  • Selonen, Salla (Helsingin yliopisto, 2015)
    Despite the known toxicity of lead (Pb) and the ban on waterfowl hunting, Pb pellets are still used at shotgun shooting ranges around the world. After firing a shotgun, pellets spread across wide areas, ending up in nearby ecosystems, which typically are forests in Finland. Still, little is known about the effects of Pb in these ecosystems and hardly anything about changes in ecosystem structure and function after range abandonment. Thus, ecosystem-level research was conducted in a shotgun shooting range area to evaluate the fate and effects of Pb in a boreal forest ecosystem and the possible recovery of the system after range abandonment. Bioaccumulation, the leaching and vertical distribution of Pb in the soil, soil nutrients and their leaching as well as structure and activity of decomposer community were studied at two contaminated sites (active [NC] and abandoned [OC] shooting ranges) and a control site, each locating in the same pine forest stand. Furthermore, tree growth, nutritional status and litter production were measured. Total Pb pellet burdens at the contaminated sites were similar, reaching up to 4 kg m-2, and shooting activity had lasted for 20 years at both sites, but occurred 20 years earlier at OC. Total Pb concentrations at the shooting ranges were extremely high, and Pb accumulated in the biota. The vertical distribution of Pb in the organic soil horizon differed between the shooting range sites, with total Pb concentrations at NC being higher in the upper F layer than in the lower H layer, but vice versa at OC. Soil fungi and all studied faunal groups (enchytraeid worms, microarthropods and nematodes) except protozoans were affected negatively by Pb. Lead decreased phosphate (PO43-) and increased nitrate (NO3-) concentrations. pH was also increased by Pb, which can further affect the biota directly or indirectly by changing Pb availability and toxicity. In the entire organic soil horizon, the negative effects of Pb were less pronounced at OC than at NC. In addition, pine needle litter decomposed faster at OC than at NC, and tree (Pinus sylvestris) radial growth was suppressed at NC after shooting activity started and increased at OC after shooting activity ceased. However, in the H layer the effects were stronger at OC, enchytraeid worms being completely absent. Furthermore, leaching of Pb through the organic soil horizon was twice as high at OC as at NC. A decrease in total Pb concentrations and toxicity in the topmost soil layer and enhanced litter decomposition rate at the abandoned shooting range indicate an on-going recovery process. In boreal forest soils that are characterized by low decomposition rates and little soil mixing due to the scarcity of earthworms, a less contaminated soil layer is gradually formed when shooting activities cease. This topmost soil layer can provide habitat for the decomposer biota and promote the recovery of soil functions. However, at the same time the dissolution of Pb from pellets deeper in the soil increases toxicity of the humus and the leaching of Pb, increasing risks to the ecosystem and groundwater quality. These findings suggest that communities and functions in Pb-contaminated boreal forest ecosystems depend on contamination history. However, despite Pb-induced changes in the decomposer communities, only slight changes in ecosystem processes were detected. This indicates high resistance of boreal forest ecosystems to this type of stress.
  • Ronkainen, Tiina (Helsingin yliopisto, 2015)
    Northern peatlands form a large storage of terrestrial carbon and at the same time they provide an important palaeoecological archive to study past climate changes and associated carbon dynamics. One of the most widely used methods to study peatland histories is the plant macrofossil method. However, peat material of the early succession stages, the fens, is often highly decomposed hampering the identification of the fossil plant remains. Thus, current methods may give only a partial view on the past vegetation, and as a result the accuracy of carbon balance estimations and climate implications may remain low. A new promising method to study past plant assemblages from peat is the geochemical plant biomarker method, which has performed well in less decomposed bog peat environments. In my study I assess the applicability of the geochemical plant biomarker method to study past plant assemblages from highly decomposed fen peat. For the first time I apply a living fen plant biomarker training set to study past fen phases. To do this, I collected and analysed two sets of living key fen plants. The training sets included boreal fen, arctic fen and permafrost peat plateau plants. The biomarker analyses on fossil peat were applied in parallel with macrofossil analyses to two boreal and one arctic permafrost peat section, all known to contain highly decomposed peat. The analyses of living plants showed that the biomarker compositions did not differ between the same species collected from different bioclimatic zones, suggesting that, at least to some extent, plant biomarkers can be used universally beyond the geographical areas where the training set was collected. The plant biomarker analyses indicate that the n-alkanes, and their ratios, are the most useful compounds to separate fen plant groups: Sphagnum mosses and vascular plants. Results showed also that biomarker composition of fen plants did not differ substantially from their bog counterparts. However, results indicated that when a wider combination of plants, plant parts and peatland habitats are incorporated into the training set the data interpretation becomes more challenging. For example, the biomarker composition of Sphagnum mosses and sedge roots resembled each other despite their differences in biology. Thus, a larger set of proxies is advisable when plant groups need to be separated more accurately. In the peat sections studied here, the biomarker method performed well in less humified bog peat layers but less well in the highly decomposed fen peat layers. The macrofossil method proved to be most competitive proxy to reconstruct past vegetation assemblages and local environmental conditions through-out the peat sections. However, when macrofossil and biomarker data were interpreted in parallel, it became clear that biomarkers were also able to reflect the major changes in dominating plant groups and in moisture conditions. Accordingly, the analysis separated the most important bog microhabitats and the major regime shifts from fen to bog. I conclude, however, that in fen environments the interpretation of biomarker data can be rather challenging. As a result, it appears that the biomarker method, as applied here, performs the best as a complimentary proxy when used in conjunction with macrofossils, and that the data should be interpreted cautiously.
  • Örmälä-Odegrip, Anni-Maria (Helsingin yliopisto, 2015)
    Bacterial viruses (i.e. phages) are ubiquitous intracellular parasites of bacteria, that along with protist grazers account for majority of bacterial mortality in nature. Phages impose strong selection for bacterial phage-resistance, which is often coupled with fitness costs on bacterial traits such as growth ability, virulence or motility. Traditionally phage-host interactions have been studied with two species systems in the laboratory, neglecting the complex web of interactions present in natural communities. The ability of phages to selectively kill bacteria has ignited an interest on phages as alternative antibacterials. However, in order to develop phage therapy, understanding of phage-host interactions in the eco-evolutionary context is essential. In this thesis I studied the implications of lytic phages on opportunistic pathogenic bacteria, as opportunists often have the ability reproduce and reside in outside-host environments, where they are predisposed to a variety of selection pressures. The role of phages in top-down control of bacterial biomass and the evolution of bacterial phage-resistance were studied in the presence of protist predators with differing feeding modes, in low-resource systems mimicking natural pond environment. Hypothesis of coincidental evolution suggests that virulence is a by-product of selection for traits that maximize bacterial fitness in environmental reservoirs. Yet, disease outbreaks by opportunists are relatively rare, suggesting that something constrains the selection for virulence. To assess the role of lytic phages on the evolution of virulence, bacteria were cultured in low-resource environment, accompanied with changes in temperature regime or changes in composition of the community of interacting bacterial enemy species, and the virulence of bacteria was measured in vivo. To study whether the potential phage-resistant bacteria surviving phage therapy would be coupled with lowered virulence, due to costs associated with phage-resistance, a clinical bacterial isolate was exposed to phage cocktails and the virulence of the phage-resistant bacteria was measured in vivo. given the strong selection for phage- resistance, the prospects of phage therapy depend a great deal on whether new phages infecting pathogenic bacteria can be readily isolated from environment. To address this, an attempt was made to isolate phages against clinical bacterial isolates harboring resistance genes to multiple antibiotics. A single lytic phage was shown to be a non- efficient top-down regulator of bacterial biomass. Rapidly emerging phage-resistant bacteria took over the bacterial populations after initial lysis by phages and protist grazers accounted for most of the long-term negative trophic effects on bacterial biomass. The presence of protist predators selected for bacteria that were less susceptible to infection by lytic phages, which suggests an overlap in the bacterial defense against a parasite and predatory protists. In general, the presence of lytic phages selected for lowered virulence in bacteria. High temperature selected for more virulent and more motile bacteria, but this was constrained by the presence of a lytic phage. In the multispecies communities the presence of all bacterial enemies led to decreased virulence in vivo. Altogether, these results contrast the hypothesis of coincidental evolution, and suggest that the presence of phages in natural reservoirs constrains the evolution of virulence, most likely through fitness costs associated with phage-resistance. Exposure to phage cocktails was also shown to be associated with decreased bacterial virulence in the phage-resistant bacteria. However, exposure to some individual phages resulted in more virulent bacteria, suggesting that the outcome of therapy could depend on the identity of the phage cocktail. Finally, a phage cocktail lysing a wide range of clinical strains was isolated from sewage. This, along with geographical patterns of phage infections suggest that new phages are available in environmental reservoirs for therapy, and the emergence of phage-resistance should not hinder the prospects of phage therapy in the global perspective.
  • Holopainen-Mantila, Ulla (Helsingin yliopisto, 2015)
    Barley (Hordeum vulgare L.) is a globally important grain crop. The composition and structure of barley grain is under genotypic and environmental control during grain development, when storage compounds (mainly starch and protein), are accumulated. Grain structure plays a significant role in malting and feed- and food-processing quality of barley. Hordeins, the major storage proteins in barley grains, are centrally located in the endosperm forming a matrix surrounding starch granules, but their role in the structural properties of barley grain is not completely understood. Thus, the main aim of the current study was to demonstrate the role of hordeins in barley grain structure. The dependence of the grain structure on the growth environment, in particular with respect to day-length and sulphur application relevant to northern growing conditions, was studied. The effects of the grain structure on end use properties in milling as well as in hydration and modification during malting were characterized. The longer photoperiod typical to latitudes in Southern Finland resulted in a C hordein fraction, entrapped by aggregated B and D hordeins, being more deeply located in the endosperm of barley cultivar Barke. Thus the impact of the growing environment on hordein deposition during grain filling was observed both at the tissue and subcellular level. However, the mechanism behind the differential accumulation of C hordein remains unclear. The deeper localization of entrapped C hordein was linked to improved hydration of grains during malting in three barley cultivars. Thus, the role of the subaleurone region in barley grain was found to be significant with respect to end use quality. Moreover, the results suggest that the growing environment affects the end-use properties of barley and that especially the northern growing conditions have a positive impact on barley processing quality. The influence of sulphur application on hordein composition in the Northern European growing conditions was demonstrated for the first time. Asparagine and C hordein served as nitrogen storage pools when the S application rate was lower than 20 mg S / kg soil, whereas total hordein and B hordein contents increased with higher S application rates. The current study also showed that even when sulphur is sufficiently available in field conditions, the hordein composition may react to sulphur application. The observed sulphur responses were in accordance with those reported earlier for hordein composition. This indicates that the more intensive growth rhythm induced in northern growing conditions does not alter greatly the effect of sulphur on grain composition. The current study confirmed that the main grain components: starch, protein and β-glucan, influence grain processing properties including milling, hydration and endosperm modification. However, their influence on endosperm texture (hardness or steeliness), which also affects the performance of barley grains in these processes, cannot be directly derived or estimated on the basis of the grain composition. The results obtained suggest that hordeins should also be taken into account in the evaluation of the processing behaviour of barley grains.
  • Broberg, Martin (Helsingin yliopisto, 2015)
    The interactions between phytopathogenic bacteria and their host plants can be characterized as an intricate web of signals and appropriate responses. Phytopathogenic soft rot bacteria occur globally, causing disease in Solanum tuberosum (potato) and other tubular staple foods in both the field and storage. One widely studied soft rot bacterium is Pectobacterium wasabiae, which has been identified in Eutrema wasabi (wasabi) plants in Japan and in potatoes in Finland. Generally, the interactions between this type of bacterium and host plants are characterized by maceration of plant tissue, due to the actions of secreted plant cell wall degrading enzymes (PCWDE), and the induction of phytohormone dependent defenses in the plants. The maceration of plant tissue involves the release of pectic oligogalacturonides (OGs) from plant cell walls. OGs have been identified as important signaling compounds, inducing the expression of a variety of defense-related genes. As the bacterial infection advances, the bacteria coordinate the production of virulence factors by utilizing regulatory proteins that modulate the transcriptome. Transcriptomic analyses have been used extensively in past studies to identify regulatory networks and signaling pathways, and these studies have provided insights into the processes underlying plant-pathogen interactions. The novel scientific results of this dissertation are derived from a combination of transcriptomic, genomic, genetic, and phenotypic analyses. This study analyzed various aspects of plant-pathogen interactions. The central bacterial model used was P. wasabiae, and the model plant of interest was Arabidopsis thaliana. This study characterized the genome of P. wasabiae via sequencing and bioinformatics analysis. Various virulence associated genes and operons, such as two distinct type 6 secretion systems, were identified and annotated. The bacterium was found to in fact be more related to P. wasabiae than Pectobacterium carotovorum, which the strain originally had been named after. Furthermore, a combination of functional genetics and transcriptomic methods, such as reverse transcription quantitative PCR (RT-qPCR) and microarrays, were used to determine the regulons controlled by the proteins ExpA and RsmA in P. wasabiae. These two proteins have been identified as important for the virulence of several γ-proteobacterial pathogens. This study analyzed the regulons via the use of three mutants: expA, rsmA, and an expA rsmA double mutant (DM). Overlapping and independently regulated targets were identified between ExpA and RsmA. Phenotypic assays for motility, growth, PCWDE activity, and virulence confirmed the transcriptomic data for the mutant strains. Novel findings included reduction of swimming motility in agar medium for P. wasabiae expA and rsmA mutants. In addition, the DM exhibited enhanced virulence and fitness in planta compared to either single mutant. Via analysis of transcriptomic data, a subset of genes was identified as affected in expression by an expA mutation independently of the presence of rsmA. The relatively unexplored role of short OGs (with a degree of polymerization (DP) less than 10) in damage-associated molecular pattern (DAMP) signaling in A. thaliana was characterized in this study. Comparative gene expression profiling based on RNA sequencing and RT-qPCR was performed on RNA harvested from plants treated with short OGs or with a mock suspension. Phenotypic assays confirmed the gene expression data. In a meta-data analysis, the resulting RNA sequencing and RT-qPCR data were compared with gene expression data from previous studies, in which long OGs (DP more than 10) were used to treat plants. This work demonstrated that short and long OGs induce genes and genesets associated with pathogen defense and phytohormone signaling, whereas reducing plant growth and development. The transcriptomic data of this study suggests that plant treatment with a mixture of short or long OGs yields a more pronounced and varied modulation of global gene expression, compared to treatment with only trimeric OGs. The regulation of the virulence of P. wasabiae, and the DAMP signaling triggered by plant cell wall damage in A. thaliana, are elements of the interactions between the plant and pathogen. The studies presented in this dissertation provide novel information about these two biological processes and highlights their connection.
  • Turja, Raisa (Helsingin yliopisto, 2015)
    Biomarkers measured in organisms are sensitive molecular, cellular or individual level biological effects, which can be applied as early-warning signals of environmental contamination before damage occurs at population, community or ecosystem levels. In this thesis, a suite of biomarkers and tissue concentrations of chemicals were measured in mussels (Mytilus trossulus) as indicators of environmental pollution. The mussels were transplanted in specially made cages in coastal areas of the northern Baltic Sea influenced by different types of contamination and environmental factors. The aims of the research were to apply the biomarker approach to (1) assess the impact of contaminants on the health status of mussels, (2) investigate the effects of seasonal variability in biotic and abiotic factors, and low salinity, and (3) validate the usefulness of the mussel caging method for biomonitoring of chemical contamination in the northern Baltic Sea. The results showed marked biomarker responses coinciding with higher concentrations of contaminants, such as polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organotins and trace metals, in mussels caged at the most contaminated study sites. At the contaminated sites, for example the enzymatic biomark¬ers of the antioxidant defense system indicated higher stress level, which was closely linked to elevated level of DNA-damage, increased biotransformation activity and decreased general health status of the cells. Biomarker responses and tissue contaminant concentrations were also related to the seasonal variability in growth and especially in soft tissue weight of the mussels. In the northern Baltic Sea the mussels almost fully deplete their energy stores during the winter; in the spring mussels efficiently feed on the fresh phytoplankton, which forms the main source of energy for their reproduction, energy metabolism and growth. Natural environmental factors caused less variation in the biomarker responses in the late summer and autumn suggesting that this time period is the most suitable for studying contaminant induced effects in mussels. More pronounced effects were observed in mussels exposed jointly to low salinity and chemical contamination, indicating that increased environmental stress reduces the tolerance of mussels towards anthropogenic pressures. This work showed that the mussel caging approach is an efficient biomonitoring method to assess biological effects and tissue accumulation of complex mixtures of contaminants as long as the effects of seasonal variability and low salinity are taken into account.
  • Gabryelczyk, Bartosz (Helsingin yliopisto, 2015)
    The possibility of controlling interactions at interfaces and surfaces of solid materials is highly interesting for a wide range of materials-related nanotechnological applications, for example, colloidal systems, adhesives, biosensors, biomimetic composite and biomedical materials. In Nature, many proteins and peptides possess the ability to recognize, specifically bind, and modify the surfaces of solid materials through sophisticated mechanism of molecular recognition. These properties have been developed during evolution via successive cycles of random mutations and selection. The natural evolution processes can be mimicked in the laboratory scale with the use of a directed evolution approach, for instance, based on the selection of short material-specific peptides from the combinatorial libraries displayed on the surface of bacteriophages or bacterial cells. Selected from billions of different variants, material-specific peptides can be studied by experimental and computational methods to define their sequence, structure, and binding properties. Subsequently, they can be engineered in order to improve their binding affinity and tailor their function for practical applications. The studies presented in this thesis show how phage display was used to identify peptides binding to diamond-like carbon (DLC). DLC is an amorphous form of carbon, with chemical and physical properties resembling natural diamond. It is used as a coating material in many industrial and biomedical applications. Peptides binding to DLC were selected form a combinatorial phage display library. Their binding and molecular basis of the function were investigated in different molecular contexts (when displayed on the phage surface, forming fusion proteins, or present in free soluble form), using multiple independent methods. It was also demonstrated that the peptides can be used in nanotechnological applications, i.e., as a self-assembling coating on the DLC surface, and for controlling properties of a colloidal form of DLC. Besides finding and characterizing peptides binding to DLC, the thesis also highlights different challenges of the directed evolution techniques, for example, selection of target unrelated peptides during biopanning, and the necessity of multiple independent ways of analyzing the functionality of selected peptides.
  • Pulkkinen, Henni (Helsingin yliopisto, 2015)
    Overfishing and environmental changes impose high risks on the wellbeing of the world's fish stocks. It is commonly acknowledged that fisheries management should be risk averse, following the principles of the precautionary approach, but unfortunately the statistical stock assessment methods often lack the ability to estimate the uncertainties related to their results. Further challenges arise from the fact that stocks which are in the most desperate need of a stock assessment are often data poor and resources to gather new data from them are scarce. Bayesian statistical inference can be utilized to conduct stock assessments cost-efficiently since these methods provide a formal way to combine information from various sources including databases, literature and expert knowledge. Bayesian inference is essentially a learning process where existing information is combined into a prior distribution, which is further updated with the most recent data. The result, a posterior distribution, expresses the best available knowledge about the phenomenon, including the related uncertainty. Furthermore, Bayesian hierarchical models enable learning between similar units, for example, stocks of the same or related species. This thesis consists of four studies that use Bayesian hierarchical models to improve knowledge in the fisheries stock assessment. Correlations between biological parameters, arising from different life history strategies among species, are utilized in paper [I] so that a data rich set of length-weight parameters can reduce the uncertainty of length-fecundity parameters for a data poor species. In paper [II], similarities between stocks of Atlantic salmon are used to estimate the stock specific key parameters of eggs-to-smolts relationship. A predictive distribution of this key parameter is also estimated, and could be used as an informative prior in a subsequent study of another Atlantic salmon stock. In addition, a hierarchical model is built to study structural uncertainty, estimating posterior probabilities of competing functional forms that describe the eggs-to-smolts survival at different juvenile densities. The Bayesian approach makes it possible to conduct analyses sequentially, as in the case of Baltic salmon stock assessment reviewed in paper [III]. Sequential analysis is useful if a stock assessment is complex and computational power does not enable analysis of all observation models at the same time. Thus, the Bayesian methods permit the creation of complex model frameworks, where incentives for structural choices arise rather from the biological process, than from the requirements or limitations of the datasets. The analysis of acoustic survey information to estimate herring resources at Bothnian Sea (paper [IV]) is another example of a computationally intensive model that has been built on the basis of available background information. This includes on one hand, the technical knowledge about the survey, and on the other hand, the biological knowledge about the spatial distribution of the fish.
  • Isomaa, Marleena (Helsingin yliopisto, 2015)
    The purpose of this thesis was to increase knowledge of heavily harvested cod stock dynamics and attain information for more suitable and sustainable management strategies. Achieving sustainable management strategies requires sufficient knowledge about the population dynamics. One tool for research this is to introduce stochastic variation models in investigated species stock dynamics. As a model species we use eastern Baltic Sea cod (Gadus morhua callarias), where the population dynamics are strongly influenced by environmental factors and fishing. We consider the stock dynamics and recovery potential under three different harvest strategies (proportional, threshold) and environmental noise scenarios. We developed age-structured discrete-time population models that include interactions with the environmental factors, e.g. salinity, fishing and natural mortality. Our results show that under current environmental conditions and threshold harvest methods cod fishing is at a sustainable level. Precautionary management ensures the best economical incomes and sustainable stocks in the long run, despite environmental fluctuations or moratoria. Results also reveal how the unpredictability of stock size, catch, and recovery increase with increasing environmental autocorrelation. Initial age structure has a strong impact on population recovery capacity in the short run, especially in over-harvested populations and under strongly fluctuating environmental conditions. Protection of younger age groups will substantially increase the recovery potential and will increase the maximum sustainable yield considerably. As a whole, precautionary harvesting strategies will increase the stock resilience against adverse environmental conditions and harvesting, guaranteeing sustainable yields in the future.
  • Voutilainen, Maria (Helsingin yliopisto, 2015)
    Mammary gland development begins during embryogenesis with the formation of species-typical number of mammary placodes that emerge along the flanks of the embryo at conserved positions. By birth, the mammary primordium has undergone branching morphogenesis and displays a small ductal tree with several branches. The organ development and growth continues throughout postnatal life and the mammary gland matures to functional form only during pregnancy and following lactation. Ectodysplasin (Eda), a member of the tumour necrosis factor family, is one of the key regulators of epithelial appendage development in all vertebrates. In humans, mutations in the Eda gene, or in other components of the signalling pathway, cause hypohidrotic ectodermal dysplasia (HED), a disorder characterized by sparse hair, missing teeth, and defects in several exocrine glands including the breast. Previous studies have shown that transgenic overexpression of Eda (K14-Eda mice) in the developing ectoderm leads to formation of ectopic mammary placodes, which give rise to supernumerary glands in the adult mice. Otherwise, effects of Eda signalling in the mammary gland have been fairly unknown. Here I have analysed the role of Eda in prepubertal mammary gland development. Characterization of the mammary glands of Eda gain- (K14-Eda) and loss-of-function (Eda−/−) mice revealed that the branching morphogenesis of the organs correlated with Eda levels. Overexpression of Eda induced precocious and accelerated branching whereas lack of Eda reduced number of ductal tips. Furthermore, Eda induced supernumerary mammary placode formation not only on the flank but also in the neck region. Analysis of the mouse line with suppressed NF-kappaB signaling (IκBαΔN mice) revealed that the transcription factor is a major mediator of Eda in the mammary gland. NF-kappaB activity was shown to be necessary for the ability of Eda to induce supernumerary mammary primordia and to accelerate branching morphogenesis. With a candidate gene approach and genome wide-profiling several potent Eda target genes were identified in the mammary gland. Among them were members of the Wnt/beta-cat pathway. The obtained results suggest that Eda promotes mammary cell fate by enhancing canonical Wnt pathway activity and other effects of Eda are cooperatively mediated by certain Wnt family members in addition to other factors. To study mammary placode formation and branching morphogenesis and to assess roles of individual downstream factors or pathways, ex vivo culture systems were developed and utilized in this thesis work.
  • Budria, Alexandre (Helsingin yliopisto, 2015)
    Recent research has strongly linked disease emergence and resurgence with human-induced environmental change. Habitat alteration, biodiversity loss, pollution, climate change and introduced species are nowadays considered potential threats to human and wildlife health. However, the impacts of these changes are not always so clear. Recent findings indicate that anthropogenic disturbances may favour the transmission of some parasites, but can also be disadvantageous to others. The aim of this thesis is to determine how human-induced environmental change affects host-parasite interactions by using anthropogenic eutrophication of the habitat of the threespine stickleback, Gasterosteus aculeatus, and its macroparasites as a model system. The different parts of the thesis focus on the influence of anthropogenic disturbances on epidemics, and on the ecological mechanisms underlying the observed patterns. After reviewing current knowledge about the influence of human-induced environmental change on host-parasites interactions, I focus on epidemiological patterns in fish populations inhabiting eutrophied waters, which vary in the extent of eutrophication. By analysing the data in respect to the degree of environmental disturbance and the knowledge about the biology of the parasites and their hosts, I develop hypotheses regarding the different mechanisms explaining the observed patterns. To test these hypotheses, I perform statistical modelling and experimental work focusing on the two most prevalent endoparasites in the studied stickleback populations, i.e. the eye fluke Diplostomum spp. and the tapeworm Schistocephalus solidus. The different approaches used in my thesis reflect the complexity of host-parasite systems and the need to mix theory, monitoring and experimental work to estimate the impact of human-induced environmental change on host-parasite interactions. Finally I point out areas that are in need of further investigation in order to elucidate the consequences of human activities for ecosystems and to build a predictive framework for how global change can affect parasitic diseases.
  • Miettinen, Juho (Helsingin yliopisto, 2014)
    Innate immunity provides the first line of defence against invading pathogens, and can also be activated by endogenous danger signals released upon tissue damage or injury. Macrophages play an important role in innate immunity. They perform the immune surveillance of their immediate surroundings seeking out tissue damage and/or invading pathogens via pattern recognition receptors. Macrophages are activated upon detection of an invading pathogen or tissue damage, leading to the expression, and secretion of the proteins required for an efficient innate immune response. Herpes simplex virus-1 (HSV-1) is a common human pathogen. It is a master of evading the host immune response leading to its ability to cause a life-long infection in its host. In most cases, HSV-1 does not cause any symptoms in the host, or causes nothing more than an occasional painful blister in the orofacial region. However, in certain cases when the host immune system is compromised, it can cause severe, even lethal, infections. Uric acid can act as an endogenous danger signal that cells can produce and release into the extracellular space after encountering a stress, including that caused by certain viral infections. A high amount of uric acid in cells can also be attributed to certain dietary habits. In the extracellular space if present at high enough concentrations and in the presence of sodium, uric acid can become crystallized into monosodium urate (MSU). MSU is the causative of gouty inflammation in the joints and is known to be phagocytosed by macrophages. In this work, human monocyte-derived macrophages infected with HSV-1, or stimulated with MSU were studied using several proteomic methods combined with functional assays. The main goals were: to characterize the protein secretion pathways and the proteins being secreted from macrophages upon (1) HSV-1 infection, and (2) MSU stimulation; and (3) to characterize the functional mechanisms of HSV-1 infected cell protein 27 (ICP27) in the inhibition of the interferon (IFN) response and also inhibition of apoptosis during HSV-1 infection. The results revealed that robust extracellular vesicle-mediated unconventional protein secretion is induced by IFN-beta priming in HSV-1-infected, and by MSU stimulation in Toll-like receptor 4 ligand, bacterial cell wall component lipopolysaccharide-primed human macrophages. The secreted proteins included endogenous danger signal proteins, and interferon-stimulated gene-encoded proteins. In addition, HSV-1 ICP27 protein requires both the nuclear localization sequence (NLS) and nuclear export sequence (NES) in order to inhibit the IFN response, whereas the NES, but not NLS, is required for the inhibition of apoptosis. The results also demonstrate that pattern recognition receptors RIG-I and/or MDA5 are involved in the HSV-1-induced activation of apoptosis. To conclude, this thesis provides new information on the innate immune response induced by HSV-1 infection, and MSU stimulation, and also on the evasion mechanisms deployed by HSV-1 to avoid the innate immune response in human macrophages. These new results presented in this thesis may be exploited in several ways; in the development of new therapies against viral infections, and in the treatment of autoinflammatory diseases.
  • Heikkilä, Maria (Helsingin yliopisto, 2014)
    Lepidoptera (butterflies and moths) are one of the most diverse and species-rich groups of organisms. Currently, the number of described species is close to 160.000, but it is estimated that the true number of species could be as high as half a million. Nearly 99% of all Lepidoptera belong to the more advanced subclade Ditrysia. The focus of this thesis is the evolutionary history of the enormous ditrysian clade. The relatedness between the 30 ditrysian superfamilies has been a mystery that has only recently begun to unfold. The aim of the thesis is to find new information on the evolutionary relationships between these superfamilies, and within two of the largest of them, Gelechioidea and Papilionoidea, both with over 18.000 described species. Evidence on the evolutionary relationships is sought from both morphological and genetic data. The morphological datasets are based on characters coded from larval, pupal and adult stages, and are thus far the largest in terms of the number of coded characters and of the number of exemplar species. In the analyses of chapters II-IV, morphological data are combined to molecular data from eight gene regions sequenced from corresponding species or genera. Morphological characters supporting phylogenetic affinities are discussed in detail. Characters described in the articles of this thesis can be used in the identification of extant and fossil taxa, and in future studies focusing on the evolution of specific morphological traits. Chapter I elucidates the evolutionary affinities of the mysterious family Lypusidae, the phylogenetic position of which has not been known. It is demonstrated that this family belongs within the megadiverse superfamily Gelechioidea. Chapter II focuses on the phylogenetic relationships between the families of the superfamily Papilionoidea (Papilionidae, Pieridae, Nymphalidae, Lycaenidae, Riodinidae, Hesperiidae and Hedylidae). The results show that the traditional concept of butterflies, i.e. including only the first five above-mentioned families is paraphyletic. Hesperiidae and Hedylidae are sister-groups and are internested within the rest of the butterflies. A divergence time analysis indicates that this superfamily originated in the Cretaceous and diversified in the Tertiary. Chapter III explores the evolutionary relationships within one of the largest but least studied groups of Lepidoptera, Gelechioidea. Based on the results, a new classification into 16 families is proposed. The aim of chapter IV is to find morphological evidence on the relationships between the ditrysian superfamilies. Several morphological characters claimed to define subgroups of Ditrysia are tested. New characters supporting evolutionary affinities of clades are described and discussed. By combining morphological and sequence data, some taxa that have been unstable in analyses based on either type of data only, find a stable position, e.g. Epipyropiidae and Cyclotornidae, Urodidoidea + Schreckensteinioidea.
  • Cui, Fuqiang (Hansaprint, 2014)
    To face the constant challenges from numerous pathogens in the environment, sophisticated defense systems have evolved in plants. Reactive oxygen species (ROS) and phytohormones are important cellular compounds that regulate plant defense systems to overcome biotic stresses from different pathogens. Against biotrophic pathogens, which require living host cells, hypersensitive cell death response (HR), a type of programed cell death mediated by ROS and salicylic acid (SA), is effective for immunity. However, to necrotrophic pathogens, which take host cell death as a hallmark of a successful colonization, the roles of ROS and phytohormones in the manipulation of cell death during plant defense are more complex. In this work, we utilized the model necrotrophic pathogen Botrytis cinerea (Botrytis; grey mold) and the model plant Arabidopsis thaliana (Arabidopsis), using mutants in reverse genetic screens, especially radical-induced cell death1 (rcd1) and botrytis susceptible1 (bos1), were used to study the functions of ROS and phytohormones in plant-Botrytis interactions. It was found that Botrytis-triggered signaling in Arabidopsis mostly overlapped with the signaling triggered by apoplastic ROS but not intracellular ROS. However, rcd1 and bos1 exhibited opposite symptoms in response to Botrytis and apoplastic ROS. This suggested that the resistance signaling regulated by RCD1 or BOS1 were distinct from a more common signaling programs induced by Botrytis and apoplastic ROS. Further study revealed that RCD1 negatively regulated Botrytis resistance independent of stress-hormones. RCD1 positively regulated Botrytis-toxin sensitivity and brassinosteroid (BR) signaling, which was demonstrated to negatively regulate plant resistance to Botrytis. In the BOS1 study, suppression of abscisic acid (ABA)-elicited cell death and control of cell death spread were identified as pivotal functions of BOS1 in its regulation of host resistance to Botrytis. This work emphasized the negative roles of both BR and ABA in response to Botrytis infection. Considering the established facts that: 1) ABA promoted plant cell death, 2) BR deficiency leads to delayed senescence, and 3) the ROS burst causes damage to both host and Botrytis; this work supports the view that cell death control plays a pivotal role in plant-Botrytis interactions, where defense combined with less cell death confers plants with an advantage in the battle against Botrytis.
  • Perkola, Noora (Helsingin yliopisto, 2014)
    The newly detected chemicals, the environmental distribution, fate, and effects in the environment of which are not well known, are called emerging compounds. Artificial sweeteners are one group of emerging compounds. The consumption of artificial sweeteners is high, and because they do not significantly metabolise, all that is consumed finds its way to wastewater treatment plants. Two artificial sweeteners, acesulfame and sucralose, do not degrade in wastewater treatment either, leading to elevated concentrations in the receiving water bodies. Another group of emerging compounds is perfluoroalkyl acids. They have been used both in industry and consumer products since the 1950s. The fluorine-carbon chain of perfluoroalkyl acids makes them extremely resistant to biological, chemical and physical degradation. They are ubiquitous in the environment and are suspected to be carsinogenic, immunotoxic and to interfere with reproduction. The aim of this study was to add to knowledge about the environmental distribution, fate and effects of artificial sweeteners and perfluoroalkyl acids. The occurrence of artificial sweeteners and perfluoroalkyl compounds was surveyed in surface waters. Wastewater effluents and sludge, storm water, and landfill leachate were analysed to evaluate the fluxes of perfluoroalkyl acids into the aquatic environment. Artificial sweeteners and perfluorooctanoic acid were irradiated under artificial sun to investigate their potential to transform via direct and indirect photochemical reactions in surface waters and ultraviolet radiation and during germicidal ultraviolet water treatment. Furthermore, Daphnia magna were exposed to artificial sweeteners to evaluate the ecotoxicological effects. It was discovered that artificial sweeteners and perfluoroalkyl acids are ubiquitous in the Finnish aquatic environment. Of the studied emission sources, wastewater effluents were the most important source of perfluoroalkyl acids in environmental waters. Based on the irradiation of perfluorooctanoic acid with a solar simulator, perfluorocarboxylic acids do not transform via direct or indirect photochemical reactions in the environment. Although a decrease in acesulfame was observed under irradiation with artificial sun, the photolytic half-life in surface water is at least one year. The photolytic half-lives of the other sweeteners were estimated to be 3 to 6 years. The ecotoxicological tests suggest that cyclamic acid might hinder the reproduction of Daphnia magna. Perfluorocarboxylic acids, sucralose and acesulfame are persistent and ubiquitous in surface waters.
  • Koskinen, Mikko (Helsingin yliopisto, 2014)
    Dendritic spines are small bulbous protrusions extending from dendritic shafts of neurons. These compartments house most of the postsynaptic terminals of excitatory synapses in the mammalian central nervous system. Dendritic spines are formed during early development and their density and morphology undergoes significant changes during maturation. After maturation dendritic spines are not static structures but display constant changes in their morphology and stability. The shape and size of dendritic spines have been linked to synaptic transmission, coupling the form of spines to neuron function. Several neurological diseases and disabilities are characterized by abnormal spine density and morphology. The main structural component of the dendritic spines is the actin filament, F-actin. Actin filaments are dynamic polymers of the monomeric protein actin. The filaments are constantly turning over and reorganizing. Both processes are regulated by actin binding proteins. All structural changes and the maintenance of dendritic spines are dependent on actin dynamics. Current research indicates that the dynamics of actin filaments do not change following spine maturation. Maturation does lead to a decrease in the movement of spines and an increase in stability, indicating changes in F-actin dynamics. In this study I have shown that the dynamics of F-actin do change during maturation. The stable pool of F-actin increases in size and the turnover of the dynamic pool increases. One of the actin binding proteins with a potential to regulate actin stabilization is myosin IIb, a motor protein with capabilities to bind F-actin and to introduce contractility into the filament network. Myosin IIb has been shown to regulate dendritic spine development, size and shape and play a role in memory consolidation. In this study I have shown that myosin IIb regulates dendritic spine F-actin via two distinct mechanisms. Myosin IIb can bind F-actin and stabilize it without affecting the turnover of the dynamic filaments. Myosin IIb-mediated contractility on the other hand can facilitate the turnover of the dynamic filaments. These findings help us to understand the molecular mechanism behind dendritic spine structure regulation and possibly in the future how it is related to synaptic transmission and different pathological states. Due to their small size, dendritic spines pose unique challenges for the study of actin dynamics. Most of the available methods are based on advanced fluorescence microscopy. In this study I have made a critical evaluation of the methods used to measure F-actin turnover in dendritic spines and the analysis of the data. I have also developed a novel approach to use fluorescence anisotropy to measure the level of actin bundling. The method has been previously applied to measure actin polymerization. My findings have led to the conclusion that in actin-dense compartments, such as the dendritic spines, fluorescence anisotropy reflects actin bundling rather than polymerization and that conclusions based on earlier research using similar techniques should be re-evaluated.