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  • Hautaniemi, Maria (Helsingin yliopisto, 2012)
    Parapoxviruses (PPVs) of the family Poxviridae are zoonotic viruses which cause contagious pustular skin infections of sheep, goats and cattle worldwide. Cases of contagious pustular stomatitis in Finnish reindeer have been reported for many years. The aims of this study were to establish specific and rapid detection methods for the causative agent of the disease and characterise the viruses circulating in Finland. The causative agent of reindeer pustular stomatitis was originally considered to be PPV Orf virus (ORFV). PCR methods amplifying different regions of the PPV genomes were developed to analyse clinical samples obtained from outbreaks of the disease in reindeer and later from viruses isolated from the disease of sheep and cattle in Finland. Subsequent phylogenetic analysis of the partial gene sequences indicated that reindeer virus from the early outbreaks is most closely related to ORFV whereas the PPV strains from the 1999-2000 outbreak is most closely related to the cattle PPV Pseudocowpox virus (PCPV). Furthermore, the phylogenetic analyses of Finnish reindeer, bovine and sheep isolates indicated that the viruses causing the disease in reindeer are very closely related to the PCPVs and ORFVs infecting Finnish cattle and sheep, respectively. Since the initial classification of the viruses causing disease in Finnish reindeer relied solely on the partial sequence analysis of two conserved PPV genes, the genome of PCPV-like reindeer isolate (F00.120R) was sequenced and analysed together with that of a reference strain of PCPV (VR634). The genomes of F00.120R and VR634 viruses were found to consist of a central core region of conserved genes, flanked by more variable terminal regions as seen in other poxvirus genomes. F00.120R was found to have four, possibly fragmented, genes at the left terminus and another near the central region of the genome that are not present in ORFV or Bovine papular stomatitis virus (BPSV; another PPV) genomes. In addition, the F00.120R genome was found to lack six genes seen near the right genome terminus of other PPVs. The protein coding contents and G+C profile of the genomes, as well as gene order and predicted protein homologies indicated that F00.120R is an isolate of PCPV and that PCPV is correctly classified as a member of the genus Parapoxvirus. These results expand the host range of PCPV to reindeer. The observed six gene deletion at the right terminus of the F00.120R genome was further investigated. The results showed that a 5431 bp sequence containing genes 116-121 was likely to have been deleted from the F00.120R genome prior to the 7th cell culture passage. These findings conclude that the genome of reindeer PCPV is 140 kbp in length and has 137 genes.
  • Kukkonen, Sami (Helsingin yliopisto, 2004)
  • Byholm, Patrik (Helsingin yliopisto, 2003)
  • Latva-Karjanmaa, Tarja (Helsingin yliopisto, 2006)
    The European aspen (Populus tremula) is a keystone species for biodiversity in boreal forests. However, the future of aspen may be threatened, because large aspens have mostly been removed from managed forests, whereas regeneration and the long-term persistence of mature trees are subjects of concern in protected areas. Aspen is a pioneer tree, and it can reproduce both sexually by seed and asexually by root suckers. Through asexual reproduction aspen forms clones, groups of genetically identical trees (ramets). In my thesis, I have studied the structure of aspen populations in terms of number, size, clonal and demographic properties. Additionally, I have investigated the emergence and survival of seedlings as well as the seed quantity and quality in crosses between the European and hybrid aspen. To study the regeneration and population structure, mature aspens were recorded in old-growth and managed forests in eastern Finland based on a large-scale inventory (11 400 ha). In addition, small aspen trees were surveyed on sample plots. Clonal structure was investigated both by morphological characters and by DNA-based markers (microsatellites). Seedling emergence and survival was studied with two sowing experiments. With crosses between European and hybrid aspens we wanted to study whether elevated temperatures due to climate change would benefit the different crosses of European and hybrid aspen unequally and thus affect the gene flow between the two species. The average volumes of mature aspen were 5.3 m3/ha in continuous old-growth, and 0.8 m3/ha in managed forests. Results indicate also that large aspen trees in managed forests are a legacy of the past less intensively managed forest landscapes. Long-term persistence of aspen in protected areas can only be secured by restoration measures creating sufficiently large gaps for regeneration. More emphasis should be given to sparing aspens in thinnings and to retaining of mature aspens in regeneration cutting in managed forests. Aspen was found to be spatially aggregated in the landscape. This could be explained by site type, disturbance history and / or limitations in seed dispersal. Clonal structure does not explain the spatial aggregation, since average size of the clones was only 2.3 ramets, and most clones (70 %) consisted of just one ramet. The small size of the clones suggests that most of them are relatively young. Therefore, sexual reproduction may be more common than has previously been thought. Seedling emergence was most successful in mineral soil especially, when the site had been burned. Only few seedlings occurred on humus. Survival of the seedlings was low, and strongly dependent on moisture, but also on seedbed conditions. The seeds were found to maintain their germinability longer than has earlier been thought to be possible. Interspecific crosses produced more seeds with higher quality than intraspecific crosses. When temperature was elevated, germination of hybrid aspen seeds increased more than seeds from P. tremula x P. tremula crosses. These results suggest that hybrid aspen may have a significant genetic impact on the European aspen, and this effect may become strengthened by climate warming.
  • Siitonen, Paula (Helsingin yliopisto, 2003)
  • Nieminen, Tiina Maileena (Helsingin yliopisto, 2005)
  • Roitto, Marja (Helsingin yliopisto, 2006)
    The goal of this thesis was to examine the ecophysiological responses of Scots pine (Pinus sylvestris L.), with an emphasis on the oxidative enzyme peroxidase and plant phenolics to environmental stresses like elevated levels of nickel (Ni) and copper (Cu), and herbivory. The effects of Ni and Cu were studied in a gradient survey at a sulphur dioxide contaminated site in the Kola Peninsula, and with experiments in which seedlings were exposed to Ni mist or to Ni and Cu amended into the soil. In addition, experimental Ni exposure was combined with disturbance of the natural lichen cover of the forest ground layer. Pine sawfly attack was simulated in the early season defoliation experiment, in which mature Scots pine were defoliated (100 %) during two successive years in a dry, nutrient-poor Scots pine stand. In addition, the effect of previous defoliation on the growth of sawfly (Diprion pini L.) larvae was studied. Apoplastic peroxidase activity was elevated in the needles of pine in a Ni- , Cu- and SO2- polluted environment, which indicated an increased oxidative stress. Increased foliar peroxidase activity due to Ni contamination was shown in the experiment, in which Ni was added as mist. No such response was found in peroxidase acitivity of the roots exposed to elevated Ni and/or Cu in the soil. Elevated Ni in the soil increased the concentration of foliar condensed tannins, which are able to bind heavy metals in the cells. Addition of low levels of Ni in the soil appeared to benefit pine seedlings, which was seen as promoted shoot growth and better condition of the roots. Wet Ni deposition of 2000 mg m-2 reduced growth and survival of pine seedlings, whereas deposition levels 200 mg m-2 or 20 mg m-2 caused no effects in a 2-y lasting experiment. The lichen mat on the forest floor did not act as an effective buffer against the adverse impacts of heavy metals on pine seedlings. However, some evidence was found indicating that soil microbes profited from the lichen mat. Artificial defoliation increased peroxidase activity in the Scots pine needles. In addition, defoliation decreased nitrogen, diamine putrescine and glucose concentrations in the needles and increased the concentrations of several phenolic compounds, starch and sucrose. Previous artificial defoliation led to poor growth of sawfly larvae reared on the pines, suggesting delayed induced resistance in Scots pine. However, there was no consistent relationship between inducibility (proportional increase in a compound following defoliation) and adverse effects on the growth of pine sawfly larvae. The observed inducible responses in needle phenolics due to previous defoliation thus appear to represent non-specific responses against sawflies.
  • Tuittila, Eeva-Stiina (Helsingin yliopisto, 2000)
  • Loponen, Heidi (Helsingin yliopisto, 2012)
    Sensory hair cells and supporting cells of the mammalian inner ear are non-dividing cells, in contrast to supporting cells of non-mammalian species that have a natural capacity for cell cycle re-entry and production of new hair cells following traumas. The mechanisms regulating the maintenance of the postmitotic state of hair cells and supporting cells are not fully understood. In this thesis, the focus has been on understanding the involvement of the core cell cycle machinery in the regulation of the postmitotic state in the inner ear sensory epithelia. Cyclin dependent kinase inhibitors (CKIs) are negative cell cycle regulators that inhibit the progression of cell cycle. In several tissues they have an important role in regulating the postmitotic state of quiescent cells. We showed that two CKIs, p19Ink4d and p21Cip1, act in co-operation to maintain the postmitotic state of hair cells. When p19Ink4d and p21Cip1 were simultaneously inactivated in mouse, auditory hair cells re-entered the cell cycle during a restricted period in early postnatal life. Furthermore, part of the hair cells that had re-entered the cell cycle also progressed to mitosis. However, cell cycle re-activation led rapidly to the activation of DNA damage response pathway and finally to p53-mediated apoptosis. These results demonstrated that postnatal hair cells do not tolerate forced cell cycle re-entry. We also analyzed the expression of other cell cycle regulators in the inner ear and found that cyclin D1 (cD1) is expressed transiently in postnatal auditory hair cells. This transient expression pattern corresponded temporarily and spatially to the abnormal proliferation pattern seen in the p19Ink4d/p21Cip1 mutant mice. This suggested that both CKIs and cD1 are involved in the control of the postmitotic state of hair cells. We tested this hypothesis by ectopically expressing cD1 in vestibular hair cells where it is not normally expressed. Only vestibular hair cells from p19Ink4d/p21Cip1 mutant mice, but not from wild type mice re-entered the cell cycle in response to ectopic cD1 expression, confirming our hypothesis. These results emphasized that the control of the postmitotic state is a complex interplay between negative and positive cell cycle regulators. Neonatal vestibular supporting cells are able to respond to mitogenic stimuli and re-enter the cell cycle. However, this response is rapidly lost during postnatal life. We found that cD1 is strongly expressed in neonatal vestibular supporting cells during the time period when these cells are able to re-enter the cell cycle in response to mitogens. This correlation suggested that the inability of mature supporting cells to re-enter the cell cycle could be caused by the absence of cD1 in these cells. We expressed cD1 ectopically in mature vestibular supporting cell in tissue culture and showed that ectopic cD1 expression triggered robust cell cycle re-entry. However, only a small fraction of supporting cells progressed to mitosis. The majority of cell cycle reactivated supporting cells showed DNA damage and arrested at the G2/M boundary. This thesis work shows that differentiated hair cells and supporting cells can be forced to re-enter the cell cycle. However, there are severe restrictions in their proliferative potential.
  • Jääskeläinen, Marko (Helsingin yliopisto, 2012)
    Retrotransposons are major components of most eukaryotic genomes. They resemble retroviruses except that their lifecycle is limited to within the boundaries of the cell. In this thesis work, the goal was to understand the replication of the BARE1 retrotransposon which belongs to the Class I LTR (Long Terminal Repeat) transposable elements (TEs) of the Copia superfamily. The BARE family constitutes about 2.9% of the barley (Hordeum vulgare L.) genome. We systemically searched Expressed Sequence Tag (EST) databases for transcriptionally active TEs, and found that BARE and LTR -retrotransposons in general are shared and widely active in grasses. We raised antibodies to the BARE1 capsid (GAG), integrase (INT), and reverse-transcriptase-RNaseH (RT-RH) proteins and showed that the BARE1 is expressed as one polyprotein of 150 kDa, processed into a shorter 90 kDa form after cleavage of the RT-RH, and further into mature-sized GAG and INT. These proteins were present in barley almost constitutively; only drought-stress had a positive effect on the levels of the GAG protein. Our results show, for the first time, that pools of retrotransposon polyproteins in plant cells are conserved and abundant enough to be detected immunologically in wide selection of species in the Gramineae. We studied the potential sites for BARE replication in barley by immune and in situ localizations. Both root and shoot apical meristems showed the presence of the BARE proteins, supporting the strategy that newly inserted copies must be carried clonally to the cells giving rise to gametes in order to survive in succeeding generations. Moreover, the cells in phloem companion cells showed localization that suggests the BARE may be able to move within the vasculature of the barley. Density gradients were used to study whether the BARE forms VLPs in barley cells. Fractions positive for BARE1 GAG and INT were visualized in transmission electron microscopy using negative staining. This was the first time VLPs were demonstrated in any plant. The VLP assembly requires excess amounts of GAG, whereas the BARE1 encodes its proteins in a single open reading frame. We observed three different pools of RNA transcripts, one for the polyprotein translation, one for encapsidation and reverse transcription, and a third one that is spliced so that it can translate only GAG, thus contributing to a comparatively larger pool of GAG. The transcript dedicated for encapsidation lacks both the cap and poly(A) for translatability, but contains the R regions critical for replication into the complementary DNA that can be inserted back into the host genome. Evidence for these new BARE insertions was found by comparing polymorphism revealed by PCR methods between several grass species. Taken together, we found that the BARE elements are capable to accomplish their lifecycle not only in barley, but in grass species in general, and thus contribute to the growth of their genome sizes.
  • Uusitalo, Matti (Helsingin yliopisto, 2010)
    The thesis provides a proposal to divide Alycidae G. Canestrini & Fanzago into two subfamilies and four tribes. This new hierarchy is based on a reassessment and reranking of new and previously known synapomorphies of the clusters concerned by cladistic analysis, using 60 morphological characters for 48 ingroup species. The basic characters of the taxa are illustrated either by SEM micrographs (Scanning Electron Microscopy) or by outline drawings. The presented classification includes the definitions of Alycini G. Canestrini & Fanzago new rank; Bimichaeliini Womersley new rank; Petralycini new rank; and the (re)descriptions of Alycus C.L. Koch, Pachygnathus Dugès, Amphialycus Zachvatkin, Bimichaelia Thor and Laminamichaelia gen. nov. The species described or redescribed are: Pachygnathus wasastjernae sp. nov. from Kvarken (Merenkurkku), Finland; Pachygnathus villosus Dugès (in Oken); Alycus roseus C.L. Koch; Alycus denasutus (Grandjean) comb. and stat. nov.; Alycus trichotus (Grandjean) comb. nov.; Alycus marinus (Schuster) comb. nov.; Amphialycus (Amphialycus) pentophthalmus Zachvatkin; Amphialycus (Amphialycus) leucogaster (Grandjean); and Amphialycus (Orthacarus) oblongus (Halbert) comb. nov.; Bimichaelia augustana (Berlese); Bimichaelia sarekensis Trägårdh; Laminamichaelia setigera (Berlese) comb. nov.; Laminamichelia arbusculosa (Grandjean) comb. nov.; Laminamichelia subnuda (Berlese) comb. nov. and Petralycus unicornis Grandjean. Fourteen nominal species were found to be junior synonymies. The importance of sensory organs in taxonomy is well recognized, but inclusion of the elaborate skin pattern seemed to improve essentially the usefulness of the prodorsal sensory area. The detailed pictures of the prodorsa of the European alycids could be used like passport photographs for the species. A database like this of prodorsa of other mite taxa as well might be an answer to future needs of species identification in soil zoology, ecology and conservation.
  • Grönberg, Henrietta (Helsingin yliopisto, 2008)
    Rhizoctonia spp. are ubiquitous soil inhabiting fungi that enter into pathogenic or symbiotic associations with plants. In general Rhizoctonia spp. are regarded as plant pathogenic fungi and many cause root rot and other plant diseases which results in considerable economic losses both in agriculture and forestry. Many Rhizoctonia strains enter into symbiotic mycorrhizal associations with orchids and some hypovirulent strains are promising biocontrol candidates in preventing host plant infection by pathogenic Rhizoctonia strains. This work focuses on uni- and binucleate Rhizoctonia (respectively UNR and BNR) strains belonging to the teleomorphic genus Ceratobasidium, but multinucleate Rhizoctonia (MNR) belonging to teleomorphic genus Thanatephorus and ectomycorrhizal fungal species, such as Suillus bovinus, were also included in DNA probe development work. Strain specific probes were developed to target rDNA ITS (internal transcribed spacer) sequences (ITS1, 5.8S and ITS2) and applied in Southern dot blot and liquid hybridization assays. Liquid hybridization was more sensitive and the size of the hybridized PCR products could be detected simultaneously, but the advantage in Southern hybridization was that sample DNA could be used without additional PCR amplification. The impacts of four Finnish BNR Ceratorhiza sp. strains 251, 266, 268 and 269 were investigated on Scot pine (Pinus sylvestris) seedling growth, and the infection biology and infection levels were microscopically examined following tryphan blue staining of infected roots. All BNR strains enhanced early seedling growth and affected the root architecture, while the infection levels remained low. The fungal infection was restricted to the outer cortical regions of long roots and typical monilioid cells detected with strain 268. The interactions of pathogenic UNR Ceratobasidium bicorne strain 1983-111/1N, and endophytic BNR Ceratorhiza sp. strain 268 were studied in single or dual inoculated Scots pine roots. The fungal infection levels and host defence-gene activity of nine transcripts [phenylalanine ammonia lyase (pal1), silbene synthase (STS), chalcone synthase (CHS), short-root specific peroxidase (Psyp1), antimicrobial peptide gene (Sp-AMP), rapidly elicited defence-related gene (PsACRE), germin-like protein (PsGER1), CuZn- superoxide dismutase (SOD), and dehydrin-like protein (dhy-like)] were measured from differentially treated and un-treated control roots by quantitative real time PCR (qRT-PCR). The infection level of pathogenic UNR was restricted in BNR- pre-inoculated Scots pine roots, while UNR was more competitive in simultaneous dual infection. The STS transcript was highly up-regulated in all treated roots, while CHS, pal1, and Psyp1 transcripts were more moderately activated. No significant activity of Sp-AMP, PsACRE, PsGER1, SOD, or dhy-like transcripts were detected compared to control roots. The integrated experiments presented, provide tools to assist in the future detection of these fungi in the environment and to understand the host infection biology and defence, and relationships between these interacting fungi in roots and soils. This study further confirms the complexity of the Rhizoctonia group both phylogenetically and in their infection biology and plant host specificity. The knowledge obtained could be applied in integrated forestry nursery management programmes.
  • Kupiainen, Kaarle (Helsingin yliopisto, 2007)
    Vehicles affect the concentrations of ambient airborne particles through exhaust emissions, but particles are also formed in the mechanical processes in the tire-road interface, brakes, and engine. Particles deposited on or in the vicinity of the road may be re-entrained, or resuspended, into air through vehicle-induced turbulence and shearing stress of the tires. A commonly used term for these particles is road dust . The processes affecting road dust emissions are complex and currently not well known. Road dust has been acknowledged as a dominant source of PM10 especially during spring in the sub-arctic urban areas, e.g. in Scandinavia, Finland, North America and Japan. The high proportion of road dust in sub-arctic regions of the world has been linked to the snowy winter conditions that make it necessary to use traction control methods. Traction control methods include dispersion of traction sand, melting of ice with brine solutions, and equipping the tires with either metal studs (studded winter tires), snow chains, or special tire design (friction tires). Several of these methods enhance the formation of mineral particles from pavement wear and/or from traction sand that accumulate in the road environment during winter. When snow and ice melt and surfaces dry out, traffic-induced turbulence makes some of the particles airborne. A general aim of this study was to study processes and factors underlying and affecting the formation and emissions of road dust from paved road surfaces. Special emphasis was placed on studying particle formation and sources during tire road interaction, especially when different applications of traction control, namely traction sanding and/or winter tires were in use. Respirable particles with aerodynamic diameter below 10 micrometers (PM10) have been the main concern, but other size ranges and particle size distributions were also studied. The following specific research questions were addressed: i) How do traction sanding and physical properties of the traction sand aggregate affect formation of road dust? ii) How do studded tires affect the formation of road dust when compared with friction tires? iii) What are the composition and sources of airborne road dust in a road simulator and during a springtime road dust episode in Finland? iv) What is the size distribution of abrasion particles from tire-road interaction? The studies were conducted both in a road simulator and in field conditions. The test results from the road simulator showed that traction sanding increased road dust emissions, and that the effect became more dominant with increasing sand load. A high percentage of fine-grained anti-skid aggregate of overall grading increased the PM10 concentrations. Anti-skid aggregate with poor resistance to fragmentation resulted in higher PM levels compared with the other aggregates, and the effect became more significant with higher aggregate loads. Glaciofluvial aggregates tended to cause higher particle concentrations than crushed rocks with good fragmentation resistance. Comparison of tire types showed that studded tires result in higher formation of PM emissions compared with friction tires. The same trend between the tires was present in the tests with and without anti-skid aggregate. This finding applies to test conditions of the road simulator with negligible resuspension. Source and composition analysis showed that the particles in the road simulator were mainly minerals and originated from both traction sand and pavement aggregates. A clear contribution of particles from anti-skid aggregate to ambient PM and dust deposition was also observed in urban conditions. The road simulator results showed that the interaction between tires, anti-skid aggregate and road surface is important in dust production and the relative contributions of these sources depend on their properties. Traction sand grains are fragmented into smaller particles under the tires, but they also wear the pavement aggregate. Therefore particles from both aggregates are observed. The mass size distribution of traction sand and pavement wear particles was mainly coarse, but fine and submicron particles were also present.
  • Molotkov, Dmitry (Helsingin yliopisto, 2014)
    Among other glial cell types such as microglia, oligodendrocytes and radial glia, astrocytes are known to be involved in brain function; metabolically supporting neurons, regulating blood flow dynamics, participating in the development of pathological states, sensing and modulating synaptic activity. At the same time the complex astrocytic morphology, with a number of highly ramified peripheral processes located near the synaptic terminals, suggests them as a possible source for morpho-functional plasticity in the brain. This thesis summarizes the work on the in vitro development and further in vivo implementation, using a gene delivery system, of a tool for suppressing activity-dependent astrocytic motility. Calciuminduced astrocyte process outgrowth and its dependence on Profilin-1, novel in vivo gene delivery approaches, a demonstration of astrocytic motility in vivo and the independence of visual processing from astrocytic motility rates are the main findings of the project. The results described in this work increase our understanding of the interactions occurring between astrocytes and neurons as well as the consequences for brain function.