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  • Terhonen, Eeva-Liisa (Helsingin yliopisto, 2015)
    Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) cover large areas in boreal regions with significant economic importance to Finnish forest industry. Approximately 15% of the spruce trees felled in Finland are rotten and thus commercially less valuable. The majority of this conifer wood decay is due to the root and butt rot pathogen Heterobasidion annosum sensu lato. Extensive logging of conifer forests has changed the environment into favouring this pathogen in stands where it originally has been rare. The saprotrophic fungus Phlebiopsis gigantea has for several years been used as a biocontrol agent against H. annosum s.l. in spruce and pine stumps. A major problem is that, although the effectiveness of P. gigantea as a biocontrol agent has empirically been shown, the long-term biological effect of this fungus on other decomposing wood microbiota has not been proven. The first objective of this thesis is to evaluate the impact of the only biocontrol agent used against root and butt rot fungus (H. annosum s.l.) on other resident microflora of Norway spruce stumps. An additional objective is to screen and identify other potentially novel bioagents that can be deployed for the biocontrol of the conifer pathogen. To find out whether the P. gigantea treatment impacts the overall diversity of other non-target stump microbes we used the 454- pyrosequencing approach. Samples were collected from forest sites previously pre-treated with P. gigantea either one, six or 13 years ago. Similarly samples were collected from untreated stumps within the same forest site over the same period of time. The results revealed that initial application of the biocontrol agent influenced the fungal species composition, but the overall fungal diversity was not affected and no statistical differences were observed between treated and non-treated stumps in the mycobiota. The biocontrol treatment significantly decreased the initial bacterial richness in the stumps, but the bacterial community gradually recovered and the negative effect of P. gigantea was attenuated. In parallel to the above studies, I further explored the potential of finding other novel biocontrol agents for use in managing the disease caused by the root rot pathogen Heterobasidion parviporum. This necessitated isolation studies of fungal root endophytes from forestry sites such as pristine mires and drained peatlands where the spread of H. annosum s.l. species have not been commonly reported. A total of 113 isolates of fungal root endophytes were obtained from non-mycorrhizal P. abies roots, which were assigned to 15 different operational taxonomic units (OTUs). Nineteen of the isolates (17%) inhibited the growth of the conifer root rot pathogen in vitro. From these, two isolates were further used to test the potential inhibitory effects during interaction, in vitro, with H. parviporum. Additionally, the metabolites secreted by the selected root endophytes were extracted and the inhibitory effects on these pathogenic fungi were assayed. The secreted metabolites were further chemically analysed using ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS). These results give new information from the new possibilities to protect the valuable seedling roots from root rot. The results of the thesis provide new information to facilitate better management and protection of these forest sites from H. annosum s.l. as well as support the continued used of P. gigantea for stump pre-treatment in Finnish forests.
  • Arovuori, Kyösti (Helsingin yliopisto, 2015)
    This dissertation analyses the effectiveness of the Common Agricultural Policy of the European Union. The motivation of this study arises from the fact that there is a lack of empirical research on the effects of policy instruments on the stated policy objectives. In addition, most of the analysis in the literature has focused on the policy objective to secure farmers incomes and, thus, on the efficiency of income redistribution. The policy effectiveness is defined as the ability of agricultural policy to respond to the stated policy objectives, given the general economic and structural conditions under which the policies operate. In this study, an empirical analysis of the effects of implemented policies and policy reforms on the stated policy objectives in the Common Agricultural Policy of the European Union is conducted. The analysis is carried out at the EU15 level and the time period analysed ranges from 1975 to 2007. In the empirical part, an econometric model utilising panel data for the EU15 countries is built. In the model, the development of the defined policy target variables is explained with policy variables and a set of economic and structural control variables. The results show that policy target variables have, in general, developed in the desired direction. The productivity of agriculture has increased, markets have been stable, self-sufficiency ratios have been achieved and the real term food prices have declined. However, farmers incomes have mostly declined. The implemented agricultural policy reforms have improved the policy effectiveness in general. The main contribution of the implemented reforms has been to the use of resources in agriculture. A policy shift from coupled price support to direct payments has released resources from agriculture to be utilised in other sectors. On the other hand, policy reforms have led to increasing price variation. This is a self-explanatory impact in the sense that administrative price setting was reduced and later on abolished in the policy reforms. According to this study, the impact of agricultural policy on the policy objectives is multifunctional. The implemented policy instruments may also have worked in the opposite direction compared to the targets set. While agricultural policies have clearly contributed to increasing agricultural productivity, they have also absorbed resources into the sector which could have been utilised more efficiently in other sectors. The country-level heterogeneity of economic and agricultural structures has increased especially due to the recent enlargements of the EU from EU15 to EU28. The analysis in this study shows that country-level heterogeneity has a significant impact on the development of policy target variables. Although the implemented policy reforms have made a contribution towards the desired direction and improved the effectiveness of the policies, it is a major challenge for future agricultural policies to effectively tackle the different structures.
  • Selim, Shaimaa (Helsingin yliopisto, 2015)
    The objective of the research described in this thesis was to increase the understanding of the transcriptional adaptations of genes encoding proteins, which have key roles in lipid and glucose metabolism, insulin signalling and inflammation, in situations of overfeeding energy during the dry period in dairy cows and summer grazing season in mares. Emphasis was placed on the potential to decrease metabolic disorders and to improve animal nutritional management and health. In the experiment I, dairy cows were fed a grass silage based diet either on a controlled energy level (on average 99 MJ/d metabolizable energy (ME), n = 8) during the last six weeks of the dry period or high energy level (on average 141 MJ/d ME, n = 8) for the first three weeks and then gradually decreasing energy allowance during three weeks to 99 MJ/d ME by parturition. In experiment II, dairy cows were fed ad libitum either grass silage (144 MJ/d ME, n = 8) or a mixture of grass silage, wheat straw and rapeseed meal (TMR, 55%: 40%: 5%, 109 MJ/d ME, n = 8) during the 8-wk dry period. Liver and adipose tissue biopsies and blood samples were collected during the transition period in experiments I and II. In the experiment III, the impact of grazing either on cultivated high-yielding pasture (CG) or semi-natural grassland pasture (NG) on fat deposition, insulin resistance status and adipose tissue gene expression of Finnhorse mares was studied. Body measurements, intravenous glucose tolerance tests (IVGTT), and sampling for the determination of neck and tailhead adipose tissue gene expressions were conducted in May and September. In experiments I and II, overfed cows had greater total dry matter and ME intakes and ME balance before parturition than control cows, but no differences were observed after calving. Increases in body weight and body condition score were greater in the overfed cows of the experiment II during the dry period. In the experiment I, there was greater plasma insulin and lower glucagon/insulin ratio in overfed cows than in control cows, while in the experiment II, there were no differences in blood parameters between overfed and TMR group during the transition period. Down-regulation of key genes linked to hepatic gluconeogenesis and fatty acid β-oxidation in the overfed group of cows in the experiment I was found, suggesting impaired liver function compared to a controlled energy diet. In the experiment II, ad libitum feeding of grass silage throughout the dry period may have attenuated the increase of hepatic gluconeogenic capacity from propionate compared to a controlled TMR diet. However, there was no difference in the expression of genes related to hepatic glucose release during the transition period (II). In adipose tissue, there was some evidence that the level of energy overfeeding may have exacerbated the inflammatory status postpartum and temporarily decreased lipogenesis very near parturition relative to control energy diet (I). In the experiment II, prepartal ad libitum feeding of grass silage decreased lipogenesis and insulin sensitivity early postpartum compared to the TMR control group. In the experiment III, CG mares had higher median body condition score and body weight, and larger waist circumference than NG mares at the end of grazing. In September, greater basal and peak insulin concentrations, and faster glucose clearance rate during IVGTT were observed in CG mares than in NG mares. In addition, a greater decrease in plasma non-esterified fatty acids during IVGTT was noticed in CG mares. There were no differences in the expression of genes related to insulin resistance, inflammation and lipogenesis between the two groups. Significant temporal differences in the expression profiles of genes related to insulin resistance and lipogenesis were observed during the grazing season. Grazing on CG had moderate effects on responses during IVGTT, but did not exacerbate insulin resistance. In conclusion, overfeeding energy with concomitant weight gain moderately altered the expression of genes related to insulin resistance, inflammation and lipogenesis in adipose tissue of dairy cows. Prepartal overfeeding energy affected the expression of genes related to hepatic gluconeogenesis and fatty acid oxidation in dairy cows, but the extent of these effects differed depending on the dietary composition during the close-up period (e.g. feeding of cereal grain). In mares, the diets with variable energy content did not affect the expression of insulin resistance- or inflammation-related genes, although mares were different in their body condition scores.
  • Österman, Janina (Helsingin yliopisto, 2015)
    Nitrogen is an indispensable element for plants and animals to be able to synthesise essential biological compounds such as amino acids and nucleotides. Although there is plenty of nitrogen in the form of nitrogen gas (N2) in the Earth s atmosphere, it is not readily available to plants but needs to be converted (fixed) into ammonia before it can be utilised. Nitrogen-fixing bacteria living freely in the soil or in symbiotic association with legume plants, fix N2 into ammonia used by the plants. This is known as biological nitrogen fixation (BNF). In contrast to industrial nitrogen fixation, an energy-demanding process using high temperature and pressure to produce chemical fertilizers, BNF makes use of solar energy alone to complete the same reaction. However, the requirements on compatibility of plants and nitrogen-fixing micro-organism, the rate of conversion and the ability of the micro-organisms to survive in stressful environments are limiting factors of this system. The current demand for more sustainable food production makes BNF an attractive alternative. However, optimization of existing BNF systems as well as development of new highly productive ones is necessary, to be able to replace the use of chemical fertilisers. In order to develop new alternatives, we need to gain more knowledge on the requirements set by both plants and micro-organisms for successful and efficient nitrogen fixation to occur. In this thesis, the nitrogen-fixing legume host Galega (goat s rue) and its symbiotic microbial partner Neorhizobium galegae were used as a model system to investigate the features defining good symbiotic nitrogen fixation. Studies of genetic diversity within the host plant showed that there are genetic traits making a distinction between the two species G. orientalis and G. officinalis, both at a whole-genome level and at the level of specific symbiosis-related genes. Genome sequencing of ten strains of N. galegae provided a useful dataset for studying i) the genomic features separating N. galegae from related nitrogen-fixing bacteria (rhizobia) and ii) the genetically encoded characteristics that divide strains of N. galegae into two separate symbiovars (symbiotic variants that show different phenotypes on the two different Galega host plant species). These studies provided new information on genes possibly involved in determining host specificity and efficiency of nitrogen fixation. In addition, previously unrecognised genetic contents provided insight into the ecology of N. galegae. Most importantly, genome sequencing enabled identification of the noeT gene, responsible for acetylation of the N. galegae Nod factor (signal molecule required for symbiosis). Although the noeT gene did not turn out to be the crucial determinant enabling nodulation of Galega spp. as previously anticipated, these results are important for future studies on mechanisms behind the selectiveness (host specificity) observed in nitogen-fixing symbioses between Galega and N. galegae.
  • Tuulos, Antti (Helsingin yliopisto, 2015)
    Winter turnip rape is a biennial crucifer oilseed. A problem associated with winter turnip rape is early sowing time in July, when farms are short of available land. The aim of this study was to determine if winter turnip rape can be established by undersowing with spring cereals without decreasing cereal and winter turnip rape yields. It was also studied, if cutting the vegetative leaves of winter turnip rape in autumn could be performed without affecting the yield and whether the composition of leaf material was nutritionally acceptable to be used as forage. The ability of winter turnip rape to act as a mineral nitrogen scavenging catch crop was studied as well. Field experiments were conducted at University of Helsinki experimental farm in Finland during 2009-2011. Winter turnip rape was either undersown with spring cereal or as pure stands. Cereals were six-row barley, two-row barley, oat and wheat. One third of the winter turnip rape plots were cut after cereal harvest to simulate forage harvest. Winter turnip rape yield and its quality was not compromised due to undersowing with a cereal, when the overwintering conditions following cereal harvest were favorable. Cutting the winter turnip rape stands in autumn decreased seed yield in the following year. Winter turnip rape leaf forage has very high crude protein content and low crude fibre content. The glucosinolate content of winter turnip rape leaf forage is comparable to other forage crucifers. Cereal yield was not decreased by the undersown winter turnip rape and quality of wheat was only slightly affected. Seed yield of six-row barley and oat was increased by the undersown winter turnip rape, indicating a facilitative interaction between the species. Undersown winter turnip rape decreased subsoil nitrate content effectively in late autumn under moist conditions that favored mineralization. Winter turnip rape can be established by undersowing to a cereal without decreasing the yields of both crops. Some cultivars of barley and oat may even benefit from the undersown winter turnip rape possibly due to root interactions. Even though winter turnip rape is nutritionally suitable as forage, autumn forage cuts should be avoided. Winter turnip rape seems to be an effective catch crop, which may decrease the leaching of soil nitrate.
  • Österman, Janina (Helsingin yliopisto, 2015)
    Nitrogen is an indispensable element for plants and animals to be able to synthesise essential biological compounds such as amino acids and nucleotides. Although there is plenty of nitrogen in the form of nitrogen gas (N2) in the Earth s atmosphere, it is not readily available to plants but needs to be converted (fixed) into ammonia before it can be utilised. Nitrogen-fixing bacteria living freely in the soil or in symbiotic association with legume plants, fix N2 into ammonia used by the plants. This is known as biological nitrogen fixation (BNF). In contrast to industrial nitrogen fixation, an energy-demanding process using high temperature and pressure to produce chemical fertilizers, BNF makes use of solar energy alone to complete the same reaction. However, the requirements on compatibility of plants and nitrogen-fixing micro-organism, the rate of conversion and the ability of the micro-organisms to survive in stressful environments are limiting factors of this system. The current demand for more sustainable food production makes BNF an attractive alternative. However, optimization of existing BNF systems as well as development of new highly productive ones is necessary, to be able to replace the use of chemical fertilisers. In order to develop new alternatives, we need to gain more knowledge on the requirements set by both plants and micro-organisms for successful and efficient nitrogen fixation to occur. In this thesis, the nitrogen-fixing legume host Galega (goat s rue) and its symbiotic microbial partner Neorhizobium galegae were used as a model system to investigate the features defining good symbiotic nitrogen fixation. Studies of genetic diversity within the host plant showed that there are genetic traits making a distinction between the two species G. orientalis and G. officinalis, both at a whole-genome level and at the level of specific symbiosis-related genes. Genome sequencing of ten strains of N. galegae provided a useful dataset for studying i) the genomic features separating N. galegae from related nitrogen-fixing bacteria (rhizobia) and ii) the genetically encoded characteristics that divide strains of N. galegae into two separate symbiovars (symbiotic variants that show different phenotypes on the two different Galega host plant species). These studies provided new information on genes possibly involved in determining host specificity and efficiency of nitrogen fixation. In addition, previously unrecognised genetic contents provided insight into the ecology of N. galegae. Most importantly, genome sequencing enabled identification of the noeT gene, responsible for acetylation of the N. galegae Nod factor (signal molecule required for symbiosis). Although the noeT gene did not turn out to be the crucial determinant enabling nodulation of Galega spp. as previously anticipated, these results are important for future studies on mechanisms behind the selectiveness (host specificity) observed in nitogen-fixing symbioses between Galega and N. galegae.
  • Vihervuori , Liisa (Helsingin yliopisto, 2015)
    Many risks and environmental concerns have been linked with the cultivation of genetically modified (GM) trees. Among the most frequently mentioned risks are the unintentional/pleiotropic effects of transgenes on organisms or plant properties that are not the targets of genetic modification. Risks in forest ecosystems are difficult to predict, due to the long life cycles of trees and their complex ecological interactions. This thesis is focused on the interactions between insect and mammal herbivores and transgenic trees. The studied silver birch (Betula pendula Roth) carried a sugar beet chitinase IV gene and the aspen and hybrid aspen (Populus sp.) carried a pine pinosylvin synthase gene, both aiming to improve the trees resistance to fungal diseases, but also potentially affecting insect and mammal herbivores. Tree growth and quality, and insect density and composition were studied during the field trial. The palatability of transgenic trees was studied in the feeding experiments using the lepidopteran herbivores rusty tussock moth and buff-tip (Orgyia antiqua L., Phalera bucephala L.) and the mammalian herbivores mountain hare and roe deer (Lepus timidus L., Capreolus capreolus L.). The studies revealed that sugar beet chitinase IV caused unintentional effects on silver birch: the transgenic trees were smaller and their red leaf colour indicated stress. There were differences in herbivore pressure between transgenic and wild-type birches: GM trees had a higher aphid density but a lower diversity of insect species and visible leaf damages. The relative growth rate of O. antiqua on transgenic birches was lower in comparison to wild-type birches. No difference in the palatability to mammals was found between transgenic and wild-type trees. The results suggest that the impact on herbivores is species- dependent. The pleiotropic effects of the transgenes affecting plant-herbivore interaction can explain these impacts. These results should be taken into account when considering the biosafety of GM trees.
  • Yaghi, Nader (Helsingin yliopisto, 2015)
    In terms of chemistry, phosphate (PV) and arsenate (AsV) are similar, but their biochemical roles are completely different. Phosphorus (P) is an important nutrient for all forms of life but when entering to watercourses it increases the eutrophication risk. Arsenic (As) is a metalloid, the inorganic species of which are toxic, with AsIII classified as a first group carcinogen. This work was undertaken to investigate the potential of Light Expanded Clay Aggregates (LECAs) to act as sorbents to remove P and As species from the aqueous phase. Detailed studies were undertaken to determine if the efficiency of LECAs can be improved by coating them with aluminum (Al) or iron (Fe) oxide. The ability of various LECA materials to remove P from water was compared at different pH values, and the reversibility and kinetics of the reactions were investigated. The results revealed that the oxide-coated LECAs were superior to LECAs as P sorbents. Furthermore, Al-LECAs was superior to Fe-LECAs, which is redox-sensitive and therefore may lose its sorption properties in temporarily prevailing anoxic conditions, e.g., in constructed wetlands (CWs) used for wastewater purification. Studies of the reaction kinetics, impact of the degree of P saturation (DPS) and P desorption revealed that the oxide-coated materials retained P more strongly than LECAs. The experiments with As were designed to simulate the conditions in groundwater where As exists as AsV or AsIII, depending on the redox potential. The optimal pH for the sorption of both As species, the effect of oxidizing agents on the sorption of AsIII and the effect of competing phosphate anions on As sorption were investigated in systematic studies, which revealed that the oxide coatings decisively increased the sorption of both As species. For AsV at a pH of 4, Al-LECAs appeared to be a better sorbent than Fe-LECAs. However, Fe-LECAs were superior to Al-LECAs at a pH of 6. The sorption of AsIII increased with elevated pH. At a pH of 9, sorption was greater onto Al-LECAs than Fe-LECAs. However, the opposite trend was obtained at a pH of 10. The oxidation of AsIII to AsV enhanced sorption by LECAs. Moreover, the oxidation-reduction reaction created new sorption sites for AsV on the manganese dioxide (MnO2) surface. The competition with P for sorption sites relatively diminished the retention of As onto LECAs compared to the oxide-coated LECAs. In the Al-LECAs, the P-induced decrease in the AsV sorption was larger than that in the Fe-LECAs. The competition between P and AsIII lowered the sorption by oxide-coated LECAs most markedly in acidic media. Thus, LECAs seem to be the best option for removing AsIII and AsV from groundwater with high P. The sorption properties of LECA materials were assessed by means of sorption isotherms describing the sorption as a function of the sorbate concentration in the solution after equilibration The actual P sorption capacity was assessed as the P retained at a P concentration of 20 μg L−1 in the equilibrium solution (X20), which is close to the upper limit (25 µg L-1) allowed in surface water to avoid eutrophication. The corresponding actual sorption capacity for the As species was determined at an equilibrium concentration of 10 μg As L−1 (X10), which is the highest allowable level in drinking water.
  • Untiveros Lazaro, Milton (Helsingin yliopisto, 2015)
    Sweet potato feathery mottle virus (SPFMV, genus Potyvirus) infects sweet potato wherever it is cultivated. In single infections, SPFMV often causes only mild symptoms a situation that changes dramatically when it co-infects the plants with Sweet potato chlorotic stunt virus (SPCSV, genus Crinivirus). Co-infection generates the sweet potato virus disease (SPVD), the most devastating viral disease of sweet potato that can cause total loss of yield. Previous studies have described RNase3 as the SPCSV protein responsible for the occurrence of SPVD. However, attempts to find resistance to SPVD based on this knowledge have failed, suggesting possible SPFMV determinants involved in the development of SPVD. This study aimed to contribute to the molecular understanding of SPFMV, in particular regarding its phylogenetic relationships, genomic structure, and ability to suppress RNA interference (RNAi). Deployment of adequate control measures requires proper characterization of the pathogen. Phylogenetic relationship of SPFMV isolates and closely related potyviruses was ambiguous due to misidentification of viruses often found in mixed infections, and lack of specific methods of detection. In this study, we sequenced the complete genome of SPFMV strain C and found phylogenetic evidence to support its reclassification as a different species, Sweet potato virus C (SPVC). Furthermore, we demonstrated that Sweet potato feathery mottle virus and Sweet potato virus C together with Sweet potato virus G (SPVG) and Sweet potato virus 2 (SPV2) were found to cluster into one unique monophyletic subgroup among the potyviruses, the so-called SPFMV-group . Members of the SPFMV-group contain some unique genomic features. Based on pairwise comparisons of partial and complete genome sequences we found recombinant isolates in SPFMV and SPVC. We also identified, novel viral determinants characteristic for the SPFMV-group of potyviruses, mainly in the P1 cistron, a region known for its high variability among potyviruses. An additional domain at the N terminus of P1 (P1-N) which is not found in other potyvirus, but is found in sweet potato mild mottle virus (genus Ipomovirus) is invariably found in members of the SPFMV-group . The P1-N domain is followed by a hypervariable region which contains specific hallmarks for each member of the SPFMV-group , and is becoming a promising region for their rapid detection and characterization. However, perhaps the most remarkable finding was the identification of an extra open reading frame (ORF) overlapping the C-terminal part of P1, and which was designated as pispo (Pretty Interesting Sweet potato Potyvirus ORF). pispo is translated as a result of a transcriptional slippage mechanism occurring in the members of the SPFMV-group of potyviruses and results in a novel protein P1N-PISPO. Plants rely on RNAi, an antiviral defense machinery, to destroy viral ribonucleic acid (RNA) molecules recognized inside cells, and spread the alert signal to neighboring cells. To evade RNAi, viruses encode proteins termed suppressors. Our analyses revealed that both P1 and P1N-PISPO contain RNAi suppressor (RSS) activity. Hence, SPFMV utilizes novel suppressors expressed from the same P1 region in different reading frames. The protein P1N-PISPO suppresses cell-to-cell movement of silencing, probably by blocking the spread of signaling to neighboring cells. In contrast, P1 protein suppresses silencing only locally. In both cases, a conserved Glycine/Tryptophan (GW) motif located in the P1N part of P1 plays a crucial role in the RSS activity. On the other hand, HC-Pro, a widely known RSS protein of potyviruses was not able to suppress silencing in SPFMV. This particular arrangement, where the RSS activity resides on P1 region and not in HC-Pro, is not reported in potyviruses, but is known, e.g., in the members of the genus Ipomovirus of the Potyviridae family. Taxonomic, structural and functional findings of this study will contribute greatly to the understanding of the evolution of SPFMV, and characterization of diseases it causes.
  • Kankare, Ville (Helsingin yliopisto, 2015)
    The precise knowledge of forest structural attributes play an essential role in decision-making, forest management procedure planning and in wood supply chain optimization. Laser scanning (LS) is one of the most promising remote sensing techniques, which can be used to estimate forest attributes at all levels, from single trees to global applications. The main objectives of the present thesis were to develop LS-based methodologies for mapping and measuring single trees. More specifically, new high-density LS-based models and methodologies were developed for the prediction of aboveground biomass (AGB), logging recovery, stem curve and external tree quality estimation. Multisource remote sensing methodologies were additionally introduced for the detailed next generation forest-inventory process. Substudies I and II concentrated on developing LS-based biomass models. Total AGB was estimated with the relative root mean squared errors (RMSE%) of 12.9% and 11.9% for Scots pine (Pinus sylvestris L.) and Norway spruce (Picea abies (L.) H.Karst.), respectively using terrestrial LS (TLS) -derived predictors in multiple regression modelling. TLS-based AGB models significantly improved the estimation accuracy of AGB components compared to state-of-the-art allometric biomass models. Airborne LS (ALS) resulted in slightly higher RMSE% values of 26.3% and 36.8% for Scots pine and Norway spruce compared to results obtained with TLS. The goal of substudies III and IV was to predict timber assortment and tree quality information using high-density LS data. Sawlog volumes were estimated with RMSE% of 17.5% and 16.8% with TLS and a combination of TLS and ALS, respectively. Results in IV showed that trees could be successfully classified in different quality classes based on TLS-measured attributes with accuracies between 76.4% and 83.6% depending on the amount of quality classes. Substudies V and VI presented new automatic processing tools for TLS data and a multisource approach for the more detailed prediction of diameter distribution. Automatic processing of TLS data was demonstrated to be effective and accurate and could be utilized to make future TLS measurements more efficient. Accuracies of ~1 cm were achieved using the automatic stem curve procedure. The multisource single-tree inventory approach combined accurate treemaps produced automatically from the TLS data, and ALS individual tree detection technique. Results from diverse forest conditions were promising, resulting in diameter prediction accuracies between 1.4 cm and 4.7 cm depending on tree density and main tree species. Each substudy (I VI) presented new methods and results for single-tree AGB modelling, external tree quality classification, automatic stem reconstruction and multisource approaches.
  • Eronen-Rasimus, Eeva (Helsingin yliopisto, 2015)
    Sea ice, at its maximum extent, is one of the largest biomes on Earth. In addition to the polar oceans, it covers extensive sea areas at lower latitudes such as the Baltic Sea and the Sea of Okhotsk. During ice formation, organic and inorganic components in the parent seawater are concentrated into saline brines within the ice, which serve as a habitat for diverse auto- and heterotrophic organisms, including bacteria. Sea-ice bacteria are responsible for many biogeochemical processes, such as decomposition of particulate organic matter, recycling of dissolved organic matter and remineralization of nutrients, analogously to bacterially driven biogeochemical processes in the water column. Since bacterial groups vary by their metabolic traits and participation in biogeochemical processes, knowledge of the bacterial community structure and its seasonal variation is essential for an understanding of ice biogeochemistry. This thesis characterises sea-ice bacterial communities during ice formation and during the winter/spring transition phase when the community composition is poorly known. Bacterial communities in Arctic and Baltic sea ice during the winter/spring transition were studied and compared. In addition, the effect of the dissolved organic matter regime on bacterial community formation was investigated in an experimental sea-ice system with North Sea water. The main methods applied were terminal-restriction fragment length polymorphism and/or Illumina Miseq sequencing together with bacterial production and abundance measurements. During the early stages of sea-ice formation, the bacterial communities were similar to the parent water communities, suggesting that the parent water determines the initial sea-ice bacterial community composition. After congealment of the sea ice, the bacterial communities changed towards communities typical of sea ice in spring. During the winter/spring transition, members of the classes Flavobacteriia (formerly Flavobacteria), Gammaproteobacteria and Alphaproteobacteria were predominant both in Baltic and Arctic sea ice. The Baltic and Arctic sea-ice bacterial communities were significantly different; however, a few members of common sea-ice bacterial genera, such as Polaribacter and Shewanella, were closely related, pointing to similar selection in ice, regardless of differences in the prevailing environmental conditions. In the experimental system, the bacterial communities were able to respond to altered substrate availability immediately after ice formation. This indicates successful adaptation of sea-ice bacteria to major shifts in temperature and salinity during ice formation. The results of this thesis suggest that sea-ice bacterial community formation and dynamics is defined by a combination of changes in environmental conditions during sea-ice maturation and its associated substrate availability, as well as resource competition. The sea-ice habitat provides an example of the enormous capacity of bacteria to adapt to changing environments and how minor members of the bacterial community can become predominant when environmental conditions change.
  • Shishido Joutsen, Tânia Keiko (Helsingin yliopisto, 2015)
    Cyanobacteria have a long evolutionary history, dating back to 3500 million years ago. They are an ancient lineage of photosynthetic bacteria that contribute to global nitrogen and carbon cycles. Cyanobacteria can be found in diverse environments, from aquatic to terrestrial systems, with specimens detected and isolated from geothermal, hypersaline, polar and desert regions. Cyanobacteria are infamous for the production of toxins such as microcystin, cylindrospermopsin, saxitoxin and anatoxin-a. However, many different types of cyanobacterial compounds with e.g. antibacterial, antifungal, anticancer, antiviral and antiprotozoal activity have also been found. This study aimed at investigating the evolution, biosynthesis, chemical variety and antifungal activity of cyanobacterial compounds. The results indicate that distantly related cyanobacteria converged on the ability to produce a rare variant of microcystin. Microcystins are commonly produced by cyanobacteria during blooms, but have recently also been found in benthic and lichen-associated cyanobacteria. A benthic, a lichen-associated cyanobacterium and two planktonic strains were shown to produce [D-Leu1] microcystin-LR. Bioinformatic analyses indicated that different evolutionary events, i.e. point mutations and gene conversion, were involved in this convergent evolution. Over 400 cyanobacterial strains were screened for the production of antifungal compounds. Genome mining allowed the discovery of the biosynthetic genes involved in the synthesis of the antifungal compounds hassallidin and anabaenolysin. Anabaena sp. SYKE748A produced more than 40 glycolipopeptide hassallidins in addition to the two main variants. Hassallidins were also identified from Aphanizomenon, Cylindrospermopsis, Nostoc and Tolypothrix species. The lipopeptides anabaenolysins were detected only in Anabaena strains. New variants of anabaenolysins C and D were chemically characterized. The antifungal activity of hassallidin D and anabaenolysin B were investigated through disc diffusion and microdilution bioassays. Synergistic antifungal activity was surprisingly observed through the production of anabaenolysin and cyclodextrins by Anabaena strains. The macrolide scytophycin was identified from Anabaena strains in this study, the first report of scytophycin from this genus. In addition, Nostoc and Scytonema strains from benthic habitats in the Finnish coastal area in the Baltic Sea were found to produce scytophycins. Unidentified antifungal compounds from the strains Fischerella sp. CENA 298, Scytonema hofmanni PCC 7110 and Nostoc sp. N107.3 were detected in the present study. Further chemical characterizations of these compounds are needed. Cyanobacteria are a prolific source for bioactive compounds, which could be toxic or potentially new drug leads. In this study, we show evidence of cyanobacterial biosynthetic genes and their evolution. We also detected new variants of the cyanobacterial compounds and their bioactivity. Furthermore, this study showed the potential of utilizing cyanobacteria for drug discovery.
  • Maanavilja, Liisa (Finnish Society of Forest Science, Natural Resources Institute Finland, Faculty of Agriculture and Forestry of the University of Helsinki, School of Forest Sciences of the University of Eastern Finland, 2015)
    Drainage to increase timber production has drastically decreased the area of undrained spruce swamp forests in northern Europe. In restoration by rewetting, drainage ditches are blocked to restore the original hydrology and, ultimately, the structure, function and ecosystem services of undrained boreal spruce swamp forests. This study quantifies the restoration success of rewetting regarding plant community composition, moss community carbon assimilation potential, Sphagnum biomass production and surface peat biogeochemistry, and aims to determine the main controls of success. The study sites comprised 18 rewetted, nine undrained and nine drained spruce swamp forests in southern Finland, complemented by sites in the umava Mountains, Czech Republic. Drainage had taken place decades prior; the rewetted sites varied in their rewetting age from 1 to 15 years. The results show that rewetting has to raise the water table above a threshold to initiate any changes in the drained ecosystem. If the threshold is crossed, the changes that occur will be rapid. Two strands of development emerged throughout the different components of the ecosystem: development towards the undrained reference state and development towards a new direction, different from both the undrained and the drained state. Rewetting created favourable conditions for Sphagnum photosynthesis. Sphagnum mosses recovered in cover and biomass production rapidly. The new growth started the accumulation of the porous surface organic matter layer characteristic of mires, which increased microbial decomposition activity in the surface organic layer towards undrained levels. Meanwhile, rewetting applied on the compacted, physicochemically altered peat created wet, unstable hydrological conditions, which increased the cover of opportunistic plant species in the understory and caused high NH4 mobilization and CH4 production in the surface organic layer. Demanding spruce swamp forest species were lacking at the rewetted sites, but rewetting was successful in restoring the common species and directing the ecosystem towards mire-like functioning.
  • Santala, Johanna (Helsingin yliopisto, 2015)
    The spraing symptoms on potato tubers caused by Potato mop-top virus (PMTV) are considered as one of the most important quality problems in potato production in the Nordic countries. PMTV has a tripartite single stranded positive sense RNA genome. RNA-RdRp encodes the viral replicase. PMTV-CP codes for a coat protein (CP) and a read through part of CP associated with the vector transmission. RNA-TGB encodes three movement related triple gene block proteins (TGBp) and an 8 kDa cysteine rich protein (8K) implicated as suppressor of RNA silencing. The aim of this study was to find new host and viral factors affecting PMTV infection and detection. As only few PMTV sequences were available in databanks prior to this study, 28 PMTV isolates obtained from symptomatic and asymptomatic potato tubers were partially sequenced. Two distinguishable types of RNA-CP and RNA-TGB were found, each showing only little genetic variability. Occurrence of tuber symptoms did not correlate with the types of PMTV RNA but seemed to depend on potato cultivar and environmental conditions. Indeed, in some cultivars almost all PMTV infections were asymptomatic. Potato sprouts grown in dark were found to contain high titers of PMTV and can be used to test potatoes for PMTV. However, viral titers decreased rapidly in sprouts exposed to light. Accumulation of PMTV specific small interfering RNAs (siRNA) in both dark grown and light exposed sprouts indicated that RNA silencing was acting against the virus in the sprouts. Moreover, the amount of viral siRNA, as opposed to viral RNA, did not decrease in the light-exposed sprouts indicating that RNA silencing was enhanced in the light exposed sprouts. PMTV TGBp1 was found, and PMTV 8K confirmed, to act as weak suppressor of silencing. PMTV TGBp3 interacts with TGBp2 to target viral ribonucleoprotein complexes to plasmodesmata and further to adjacent cells, but TGBp2 and TGBp3 are recycled through endocytosis. PMTV TGBp3 was found to be phosphorylated on two tyrosine residues, both of which are located within tyrosine based sorting motifs implicated in endocytosis. Mutation of one of the sites led to strengthening of the TGBp2-TGBp3 interaction and restricted the virus to the originally infected cell. Therefore, tyrosine phosphorylation of TGBp3 seemed to affect PMTV infection by regulating the interaction between TGBp2 and TGBp3. According to this study, the current reliance on symptoms in seed potato inspections for PMTV is unreliable and risks spreading the virus to new fields. Light was found to accelerate antiviral silencing in sprouts, which may explain why PMTV and its symptoms are rarely detected in aboveground parts of potato plants. Tyrosine phosphorylation of TGBp3 marked the first evidence that viral proteins can be tyrosine-phosphorylated in plants. This find could offer means for breeding PMTV resistant potato cultivars.  
  • Damerau, Annelie (Helsingin yliopisto, 2015)
    The consumption of whole grain foods high in fibre is of interest because of the health-promoting effects associated with dietary fibre. Therefore, there is interest in developing new fibre-rich cereal foods. However, these kinds of foods also contain polyunsaturated lipids, which are prone to oxidation. Further, lipids are dispersed in a heterogeneous matrix of starch, proteins and fibre, which increases their tendency to oxidize because of a large surface area and possible contact with prooxidants. The oxidation of lipids decreases nutritional quality and causes the formation of undesirable flavours. Knowledge of the oxidation behaviour of dispersed lipids in solid cereal foods, and of how factors like process parameters, structural features of the products and storage conditions affect lipid oxidation, is limited. In this thesis, the oxidative behaviour of foods with dispersed lipids was studied using two model systems. The first model system was a spray-dried emulsion containing sunflower oil encapsulated in a Na-caseinate-maltodextrin matrix, with either non-cross-linked or cross-linked proteins. The stability of the total and surface lipid fractions was determined during storage under different relative humidities (RHs). Further, the effect of RH on the amount of volatiles released from oxidized spray-dried emulsions was studied. The second model system consisted of extruded cereals produced from either whole grain oats or rye bran (coarse or fine) using different extrusion parameters. Their oxidative stability was studied during storage at 40 ºC, after milling and standardization to RH 33%. The primary oxidation was measured by peroxide values in the spray-dried emulsions and by losses of tocopherols and tocotrienols in the spray-dried emulsions and rye bran extrudates. Secondary oxidation was determined based on volatile secondary lipid oxidation products analysed by static head space (SHS-GC-FID) in the spray-dried emulsions and by head space solid-phase micro extraction (HS-SPME-GC-MS) in the extruded cereals. In addition to the oxidation parameters, enzymatic hydrolysis of lipids in the oat extrudates and the fatty acid composition of all models were studied by measuring the neutral lipid and fatty acid profiles, respectively. Increasing the RH improved the oxidative stability of both the total and surface lipid fractions of the stored spray-dried emulsions. This behaviour was mainly linked to the loss of individual powder particles upon caking and collapsing of the matrix at RH 75%. In addition, excess protein may have delayed oxidation via its radical scavenging activity. At RH 54%, cross-linking of the protein slightly improved the oxidative stability. The profiles of the volatile oxidation products from the spray-dried emulsions analysed by HS-SPME were also influenced by the RH. The effect was related to water-induced changes in hydrophilicity, structure and binding ability of the matrix, and to partitioning and solubility of the volatiles. The highest overall amount of volatiles released was obtained at water contents of 3.1% and 5.2% (RH 11% and 33%). The enzymatic hydrolysis of lipids in oats was effectively prevented by extrusion, even at the lowest temperature of 70 °C. The extrusion temperature could be increased to 110 °C without subjecting the lipids to non-enzymatic oxidation. However, by increasing the temperature to 130 °C, lipid oxidation was promoted, which also yielded losses of neutral lipids over time. In the case of the rye bran, the low water content (13% or 16%) in the extrusion of coarse or fine bran led to the most stable lipids during storage. The improved oxidative stability at low water contents in extrusion was connected with the higher formation of Maillard reaction products, which could have acted as antioxidants. The grinding of rye bran prior to extrusion caused a loss of tocols and increased the amounts of Maillard reaction products formed. The oxidative stability of the dispersed lipids was shown to be highly related to water induced physical changes in the matrix structure, which makes controlling the RH in the surrounding atmosphere an important factor in storage. Further, the RH affected the amount of volatile lipid oxidation products released, and this needs to be considered in determining lipid oxidation by HS-SPME. Extrusion was shown to inactivate lipases in oats. For the lipid stability in cereal extrudates, low temperature and low water content during extrusion were shown to be beneficial.
  • Holopainen, Sari (Helsingin yliopisto, 2015)
    The boreal biome harbours a large share of the world s wetlands, and is the main breeding area for several duck species. Breeding habitats strongly influence duck reproduction. In this study, the habitat use of ducks and their breeding success was estimated in a boreal landscape in southern Finland. A review of duck habitat use in boreal wetlands was additionally made. Boreal forest lakes are normally considered stable environments, but from a duck s point of view both resource abundance and habitat quality in the lakes may differ from one year to another. In this study the vegetation of boreal freshwater lakes was found to slowly change during the 20-year study period. Beavers (Castor canadensis) caused pronounced alterations in the lakes by flooding them. Flooding rendered lake vegetation structure more luxuriant and increased the lake use by ducks. In general, duck brood lake use was positively associated with both the coverage of wide belts of tall emergent plants and the abundance of emerging insects and aquatic invertebrates. The relative importance of these factors varied between species. The brood production of common goldeneyes (Bucephala clangula) was regulated by density dependence per se, but fluctuated with food availability per capita. No spatial density-dependent effects were found for the common teal (Anas crecca), but the breeding success was instead explained well by habitat quality variation. Both food and flood abundance had a positive effect, but different variables seemed to operate during different phases of the breeding season. Many aspects of boreal breeding ducks are still poorly studied. Especially lacking are studies concerning duckling survival and the effects of anthropogenic actions on duck habitat use. This research underlines the importance of high quality habitats for breeding duck populations in the boreal landscape. The varying habitat requirements of duck species should be considered when implementing wetland management.
  • Majasalmi, Titta (Helsingin yliopisto, 2015)
    The aim of this dissertation is to assess the accuracy of different ground reference methods used to validate satellite based leaf area index (LAI) and the fraction of absorbed photosynthetically active radiation (fPAR) products. LAI and fPAR are strongly linked, although they principally and practically measure different properties: LAI quantifies the areal interphase between soil and atmosphere, whereas fPAR quantifies the energy available for photosynthesis. Until now, the development of remote sensing based methods to estimate LAI and fPAR in a boreal forest has been hindered by the scarcity of ground data, which is required to validate and develop existing algorithms. The aim of the first part of this dissertation was to assess the impacts of different methodological approaches to estimate LAI in boreal forests, and to validate satellite based LAI products. Results showed that the accuracy of ground based LAI estimates is sensitive to both the retrieval methods and sampling scheme used to collect the optical LAI data. The satellite based measurements of LAI demonstrated a large temporal variability in LAI. The second part of the dissertation focused on measuring and modeling fPAR in a boreal forest. A new scheme for measuring and modeling ground reference fPAR based on photon recollision probability was presented in this dissertation. Ground reference fPAR is usually estimated only for the forest canopy layer. This study is among the first ones to validate the new global satellite based fPAR product called GEOV1 using data of both the forest canopy and understory layers from boreal forests. Results showed that satellite based fPAR products may correspond better with the total fPAR, instead of only the forest canopy fPAR as has often been presumed.
  • Hakojärvi, Mikko (Helsingin yliopisto, 2015)
    Precision farming is a set of advanced technologies and cultivation practices that aim to enhance crop growing conditions in a site-specific manner. Current machinery offers many technical tools for such actions, but information about when and where it is necessary to use the developed machinery effectively is lacking. The use of crop models has been proposed to combine the effects of growing season conditions and field spatial properties. However, in a timely and spatially variable environment, which a field often is, much information about site-specific growing conditions should be available in order for an advanced crop model to reproduce the site-specific growth in a detailed manner. Unfortunately the information from fields has often been very limited, and insufficient for such purposes. Furthermore the set of precision farming tools and the number of growth factors that can be managed is limited. For describing maximal biomass accumulation, a simple crop model was introduced and evaluated in this thesis. The model is mechanistic, and it uses a minimal number of parameters that all are based on physics, chemistry or physiology. The model can be used for calculating the radiation or radiation and water limited biomass accumulation of a C3-crop. A field experiment equipped with continuous measurements was used for model establishment and after model establishment the model was evaluated with a field experiment with various radiation, nitrogen fertilization and precipitation conditions. In both the studies the crop model was found to produce the maximal biomass accumulation when parameter values measured in the experiment were used. The model was applied in a study evaluating the effects of selected site- and depth-specific soil properties on yield variation on three different clay soil fields located in southern Finland. In order to evaluate the effects of selected soil properties under various weather conditions a Monte Carlo method was used with the biomass accumulation model and generated precipitations. The yield variation was evaluated according to temporal mean biomass yield and temporal standard deviation of the biomass yield. For studying the use of the model in a spatial environment, the introduced biomass accumulation model was applied in a simulator built for simulating a fully automated crop farm. In addition, the use of continuous soil moisture measurements for measuring the crop water use and further for biomass accumulation was tested. According to the results, the crop model was capable of simulating the highest biomass accumulation of the crops used in the experiments. This was the case for all radiation-limited simulations and for most of the water-limited simulations. In a few cases the values of the observed soil properties were found to cause too low biomass accumulation in simulations but in such cases the problem was also present in the comparisons of observed soil properties and observed soil water content during the growing season. For future research with the model, the next phase will be to test the model use in precision farming-related decision making. The structure of the model enables its use with other C3-crops than small grain cereals. Therefore testing of the model with other C3-crops could be performed in future research.
  • Itkonen, Suvi (Helsingin yliopisto, 2015)
    Phosphorus (P) intake in Western countries exceeds the nutritional recommendations 2- to 3-fold, and the increased use of food additive phosphates (FAPs) in the food industry has augmented total P (TP) intake; an estimated 10-50% of TP intake comes from additives. However, in food composition databases, the amounts of FAPs may have not been taken into account. Difficulties in measuring true P intake occur also because bioavailability of P differs between foodstuffs; inorganic P from additives is absorbed better in the intestine than natural, organic P. The harmfulness of high P intake to kidney patients has been known for years, and dietary P restriction is used in the treatment of the disease to avoid vascular calcification, which occurs due to calcium-phosphate deposition in the vascular vessel wall. However, recently, concerns have been raised about the role of elevated serum phosphate concentrations in cardiovascular health in the general population, but data on dietary P is scarce. This thesis aims to offer new insights into the potential health risks related to high dietary P intake in Finland by providing knowledge on the bioavailability of P in foodstuffs, and on the association of high dietary P intake with cardiovascular disease risk factors in the general population. In Study I, a new method was developed for analysis of in vitro digestible P (DP), indicating bioavailable and absorbable P. DP and TP contents of certain differently processed cereals were analyzed. The calculated uncertainty of the new analysis method had little effect on the DP contents in the samples. Soured cereals contained more DP than unsoured cereals, and the long processing time increased the amounts of DP. In Study II, the DP contents of selected plant-based foodstuffs with or without FAPs were determined by the analysis method developed in Study I. The analyzed plant-based products contained varying amounts of P, but most P was not digestible, except in FAP-containing foodstuffs (cola drinks and baking powder-leavened muffins). In Study III, the associations between TP intake, P density of the diet (TP intake related to energy intake), FAP intake, and a risk factor of cardiovascular disease, carotid intima-media thickness were investigated in a middle-aged Finnish population (n=546). Linear associations among P density of the diet, FAP intake, and carotid intima-media thickness were found when all subjects were included in the analysis, and between FAP intake and carotid intima-media thickness in females (n=370). However, among males (n=176) no significant associations were found. In conclusion, results of this thesis reinforce the previous understanding about the importance of the source of P when assessing P intake in a population. The plant-based foodstuffs generally contain low amounts of DP, but the FAP-containing plant-based products have relatively higher DP contents than FAP-free products. The method developed for the analysis of DP is reliable but requires validation against an in vivo method before these results can be generalized. Albeit high dietary P intake (P density of the diet and FAP intake) was associated with a cardiovascular disease risk factor, carotid intima-media thickness, due to the cross-sectional design of the study, no causal relationships can be inferred. Thus, the potential adverse effects of highly absorbable FAPs should be further investigated both in intervention and in follow-up studies before final conclusions about the harmfulness of high P intake for the general population can be drawn. To find this out, data on use of FAPs in the food industry and updated information on P contents in foodstuffs are also needed.
  • Nousiainen, Aura (Helsingin yliopisto, 2015)
    The use of pesticides has allowed the efficient use of agricultural soil and provided humans with greater yields and agri-food security. Unfortunately, many pesticides have also adverse effects to the environment or human health, and may end up where they were not intended: the precious groundwater reserves. The use of atrazine, a herbicide used for controlling broad-leaf weeds, was banned in the EU for this reason in 2004, but is still globally one of the most widely used herbicides today. Although atrazine can be completely mineralized by microbes, in the subsurface, slow or incomplete degradation of atrazine is often observed. The ability of atrazine degradation by microbes can be utilized in bioremediation, a technique in which contaminants are removed by microbial activity. This study was undertaken to elucidate the potential use of genetic tools, such as quantitative PCR (qPCR), radiorespirometry, microautoradiography (MAR), clone libraries and genetic fingerprinting methods, in atrazine contaminated soils, and to apply them in atrazine bioremediation. Collaboration with our Indian partner permitted comparison between atrazine treated, cropped agricultural soils and boreal subsoil contaminated two decades ago with residual atrazine from weed control in municipal areas. Four different bioremediation methods, natural attenuation, bioaugmentation, biostimulation, and their combination, were used to reduce atrazine concentration in soil. Atrazine degradation copy numbers often reflected the atrazine degradation potential, indicating their robustness as monitoring tools in different soils. The most efficient bioremediation treatment was bioaugmentation by atrazine degrading bacterial strains Pseudomonas citronellolis or Arthrobacter aurescens, or by an atrazine degrading bacterial consortium: in the agricultural soil, up to 90% of atrazine was degraded in less than a week, whereas in the boreal subsoil, 76% of atrazine was mineralized. In the clone library constructed from boreal soil, several clones related to taxa which include known atrazine degraders were found. In this soil, biostimulation with additional carbon was an efficient treatment at reduced temperature. In general, the efficiency of atrazine removal in different treatments was bioaugmentation and biostimulation > bioaugmentation > biostimulation > natural attenuation. Previous exposure to atrazine was the most influential factor in atrazine disappearance from soil, as recent exposure always correlated with faster atrazine degradation, and greatly affected the composition of the microbial community, elucidated by LH-PCR. These results serve as an example on how soil origin, exposure history, organic content and use must be taken into account while choosing the best bioremediation method. Knowledge on the presence of genetic degradation potential can be helpful in choosing the treatment method. While bioaugmentation removed 90% of atrazine from soil, its application in field scale may be challenging. Our results show, that biostimulation alone may serve as the treatment method of choice, even in the challenging subsoil surroundings where atrazine concentrations are low.