Molecular characterization of sediment bacterial communities affected by fish farming

Fish farming introduces nutrients, microbes and a wide variety of chemicals such as heavy metals, antifoulants and antibiotics to the surrounding environment. Introduction of antibiotics has been linked with the increased incidence of antibiotic resistant pathogenic bacteria in the farm vicinities. In this thesis molecular methods such as quantitative PCR and DNA sequencing were applied to analyze bacterial communities in sediments from fish farms and pristine locations. Altogether four farms and four pristine sites were sampled in the Baltic Sea. Two farm and two pristine locations were sampled over a surveillance period of four years. Furthermore, a new methodology was developed as a part of the study that permits amplifying single microbial genomes and capturing them according to any genetic traits, including antibiotic resistance genes. The study revealed that several resistance genes for tetracycline were found at the sediment underneath the aquaculture farms. The copy number of these genes remained elevated even at a farm that had not used any antibiotics since year 2000, six years before this study started. Similarly, an increase in the amount of mercury resistance gene merA was observed at the aquaculture sediment. The persistence of the resistance genes in absence of any selection pressure from antibiotics or heavy metals suggests that the genes may be introduced to the sediment by the farming process. This is also supported by the diversity pattern of the merA gene between farm and pristine sediments. The bacterial community-level changes in response to fish farming were very complex and no single phylogenetic groups were found that would be typical to fish farm sediments. However, the community structures had some correlation with the exposure to fish farming. Our studies suggest that the established approaches to deal with antibiotic resistance at the aquaculture, such as antibiotic cycling, are fundamentally flawed because they cannot prevent the introduction of the resistance genes and resistant bacteria to the farm area by the farming process. Further studies are required to study the entire fish farming process to identify the sources of the resistance genes and the resistant bacteria. The results also suggest that in order to prevent major microbiological changes in the surrounding aquatic environment, the farms should not be founded in shallow water where currents do not transport sedimenting matter from the farms. Finally, the technique to amplify and select microbial genomes will potentially have a considerable impact in microbial ecology and genomics.

Virtual versus classroom language teaching : A comparative study of the learning outcomes and student responses between two English reading comprehension groups

A 26-hour English reading comprehension course was taught to two groups of second year Finnish Pharmacy students: a virtual group (33 students) and a teacher-taught group (25 students). The aims of the teaching experiment were to find out: 1.What has to be taken into account when teaching English reading comprehension to students of pharmacy via the Internet and using TopClass? 2. How will the learning outcomes of the virtual group and the control group differ? 3. How will the students and the Department of Pharmacy respond to the different and new method, i.e. the virtual teaching method? 4. Will it be possible to test English reading comprehension learning material using the groupware tool TopClass? The virtual exercises were written within the Internet authoring environment, TopClass. The virtual group was given the reading material and grammar booklet on paper, but they did the reading comprehension tasks (written by the teacher), autonomously via the Internet. The control group was taught by the same teacher in 12 2-hour sessions, while the virtual group could work independently within the given six weeks. Both groups studied the same material: ten pharmaceutical articles with reading comprehension tasks as well as grammar and vocabulary exercises. Both groups took the same final test. Students in both groups were asked to evaluate the course using a 1 to 5 rating scale and they were also asked to assess their respective courses verbally. A detailed analysis of the different aspects of the student evaluation is given. Conclusions: 1.The virtual students learned pharmaceutical English relatively well but not significantly better than the classroom students 2. The overall student satisfaction in the virtual pharmacy English reading comprehension group was found to be higher than that in the teacher-taught control group. 3. Virtual learning is easier for linguistically more able students; less able students need more time with the teacher. 4. The sample in this study is rather small, but it is a pioneering study. 5. The Department of Pharmacy in the University of Helsinki wishes to incorporate virtual English reading comprehension teaching in its curriculum. 6. The sophisticated and versatile TopClass system is relatively easy for a traditional teacher and quite easy for the students to learn. It can be used e.g. for automatic checking of routine answers and document transfer, which both lighten the workloads of both parties. It is especially convenient for teaching reading comprehension. Key words: English reading comprehension, teacher-taught class, virtual class, attitudes of students, learning outcomes

Fatigued driving : prevalence, risk factors and groups, and the law

Fatigue and sleepiness are major causes of road traffic accidents. However, precise data is often lacking because a validated and reliable device for detecting the level of sleepiness (cf. the breathalyzer for alcohol levels) does not exist, nor does criteria for the unambiguous detection of fatigue/sleepiness as a contributing factor in accident causation. Therefore, identification of risk factors and groups might not always be easy. Furthermore, it is extremely difficult to incorporate fatigue in operationalized terms into either traffic or criminal law. The main aims of this thesis were to estimate the prevalence of fatigue problems while driving among the Finnish driving population, to explore how VALT multidisciplinary investigation teams, Finnish police, and courts recognize (and prosecute) fatigue in traffic, to identify risk factors and groups, and finally to explore the application of the Finnish Road Traffic Act (RTA), which explicitly forbids driving while tired in Article 63. Several different sources of data were used: a computerized database and the original folders of multidisciplinary teams investigating fatal accidents (VALT), the driver records database (AKE), prosecutor and court decisions, a survey of young male military conscripts, and a survey of a representative sample of the Finnish active driving population. The results show that 8-15% of fatal accidents during 1991-2001 were fatigue related, that every fifth Finnish driver has fallen asleep while driving at some point during his/her driving career, and that the Finnish police and courts punish on average one driver per day on the basis of fatigued driving (based on the data from the years 2004-2005). The main finding regarding risk factors and risk groups is that during the summer months, especially in the afternoon, the risk of falling asleep while driving is increased. Furthermore, the results indicate that those with a higher risk of falling asleep while driving are men in general, but especially young male drivers including military conscripts and the elderly during the afternoon hours and the summer in particular; professional drivers breaking the rules about duty and rest hours; and drivers with a tendency to fall asleep easily. A time-of-day pattern of sleep-related incidents was repeatedly found. It was found that VALT teams can be considered relatively reliable when assessing the role of fatigue and sleepiness in accident causation; thus, similar experts might be valuable in the court process as expert witnesses when fatigue or sleepiness are suspected to have a role in an accident’s origins. However, the application of Article 63 of the RTA that forbids, among other things, fatigued driving will continue to be an issue that deserves further attention. This should be done in the context of a needed attitude change towards driving while in a state of extreme tiredness (e.g., after being awake for more than 24 hours), which produces performance deterioration comparable to illegal intoxication (BAC around 0.1%). Regarding the well-known interactive effect of increased sleepiness and even small alcohol levels, the relatively high proportion (up to 14.5%) of Finnish drivers owning and using a breathalyzer raises some concern. This concern exists because these drivers are obviously more focused on not breaking the “magic” line of 0.05% BAC than being concerned about driving impairment, which might be much worse than they realize because of the interactive effects of increased sleepiness and even low alcohol consumption. In conclusion, there is no doubt that fatigue and sleepiness problems while driving are common among the Finnish driving population. While we wait for the invention of reliable devices for fatigue/sleepiness detection, we should invest more effort in raising public awareness about the dangerousness of fatigued driving and educate drivers about how to recognize and deal with fatigue and sleepiness when they ultimately occur.

Anthropometric measures of obesity-their association with type 2 diabetes and hypertension across ethnic groups

Clinical trials have shown that weight reduction with lifestyles can delay or prevent diabetes and reduce blood pressure. An appropriate definition of obesity using anthropometric measures is useful in predicting diabetes and hypertension at the population level. However, there is debate on which of the measures of obesity is best or most strongly associated with diabetes and hypertension and on what are the optimal cut-off values for body mass index (BMI) and waist circumference (WC) in this regard. The aims of the study were 1) to compare the strength of the association for undiagnosed or newly diagnosed diabetes (or hypertension) with anthropometric measures of obesity in people of Asian origin, 2) to detect ethnic differences in the association of undiagnosed diabetes with obesity, 3) to identify ethnic- and sex-specific change point values of BMI and WC for changes in the prevalence of diabetes and 4) to evaluate the ethnic-specific WC cutoff values proposed by the International Diabetes Federation (IDF) in 2005 for central obesity. The study population comprised 28 435 men and 35 198 women, ≥ 25 years of age, from 39 cohorts participating in the DECODA and DECODE studies, including 5 Asian Indian (n = 13 537), 3 Mauritian Indian (n = 4505) and Mauritian Creole (n = 1075), 8 Chinese (n =10 801), 1 Filipino (n = 3841), 7 Japanese (n = 7934), 1 Mongolian (n = 1991), and 14 European (n = 20 979) studies. The prevalence of diabetes, hypertension and central obesity was estimated, using descriptive statistics, and the differences were determined with the χ2 test. The odds ratios (ORs) or  coefficients (from the logistic model) and hazard ratios (HRs, from the Cox model to interval censored data) for BMI, WC, waist-to-hip ratio (WHR), and waist-to-stature ratio (WSR) were estimated for diabetes and hypertension. The differences between BMI and WC, WHR or WSR were compared, applying paired homogeneity tests (Wald statistics with 1 df). Hierarchical three-level Bayesian change point analysis, adjusting for age, was applied to identify the most likely cut-off/change point values for BMI and WC in association with previously undiagnosed diabetes. The ORs for diabetes in men (women) with BMI, WC, WHR and WSR were 1.52 (1.59), 1.54 (1.70), 1.53 (1.50) and 1.62 (1.70), respectively and the corresponding ORs for hypertension were 1.68 (1.55), 1.66 (1.51), 1.45 (1.28) and 1.63 (1.50). For diabetes the OR for BMI did not differ from that for WC or WHR, but was lower than that for WSR (p = 0.001) in men while in women the ORs were higher for WC and WSR than for BMI (both p < 0.05). Hypertension was more strongly associated with BMI than with WHR in men (p < 0.001) and most strongly with BMI than with WHR (p < 0.001), WSR (p < 0.01) and WC (p < 0.05) in women. The HRs for incidence of diabetes and hypertension did not differ between BMI and the other three central obesity measures in Mauritian Indians and Mauritian Creoles during follow-ups of 5, 6 and 11 years. The prevalence of diabetes was highest in Asian Indians, lowest in Europeans and intermediate in others, given the same BMI or WC category. The  coefficients for diabetes in BMI (kg/m2) were (men/women): 0.34/0.28, 0.41/0.43, 0.42/0.61, 0.36/0.59 and 0.33/0.49 for Asian Indian, Chinese, Japanese, Mauritian Indian and European (overall homogeneity test: p > 0.05 in men and p < 0.001 in women). Similar results were obtained in WC (cm). Asian Indian women had lower  coefficients than women of other ethnicities. The change points for BMI were 29.5, 25.6, 24.0, 24.0 and 21.5 in men and 29.4, 25.2, 24.9, 25.3 and 22.5 (kg/m2) in women of European, Chinese, Mauritian Indian, Japanese, and Asian Indian descent. The change points for WC were 100, 85, 79 and 82 cm in men and 91, 82, 82 and 76 cm in women of European, Chinese, Mauritian Indian, and Asian Indian. The prevalence of central obesity using the 2005 IDF definition was higher in Japanese men but lower in Japanese women than in their Asian counterparts. The prevalence of central obesity was 52 times higher in Japanese men but 0.8 times lower in Japanese women compared to the National Cholesterol Education Programme definition. The findings suggest that both BMI and WC predicted diabetes and hypertension equally well in all ethnic groups. At the same BMI or WC level, the prevalence of diabetes was highest in Asian Indians, lowest in Europeans and intermediate in others. Ethnic- and sex-specific change points of BMI and WC should be considered in setting diagnostic criteria for obesity to detect undiagnosed or newly diagnosed diabetes.

Serum uric acid and metabolic risk factors in three ethnic groups: Asian Indians and Creoles in Mauritius and Chinese in Qingdao, China

In humans with a loss of uricase the final oxidation product of purine catabolism is uric acid (UA). The prevalence of hyperuricemia has been increasing around the world accompanied by a rapid increase in obesity and diabetes. Since hyperuricemia was first described as being associated with hyperglycemia and hypertension by Kylin in 1923, there has been a growing interest in the association between elevated UA and other metabolic abnormalities of hyperglycemia, abdominal obesity, dyslipidemia, and hypertension. The direction of causality between hyperuricemia and metabolic disorders, however, is unceartain. The association of UA with metabolic abnormalities still needs to be delineated in population samples. Our overall aims were to study the prevalence of hyperuricemia and the metabolic factors clustering with hyperuricemia, to explore the dynamical changes in blood UA levels with the deterioration in glucose metabolism and to estimate the predictive capability of UA in the development of diabetes. Four population-based surveys for diabetes and other non-communicable diseases were conducted in 1987, 1992, and 1998 in Mauritius, and in 2001-2002 in Qingdao, China. The Qingdao study comprised 1 288 Chinese men and 2 344 women between 20-74, and the Mauritius study consisted of 3 784 Mauritian Indian and Mauritian Creole men and 4 442 women between 25-74. In Mauritius, re-exams were made in 1992 and/or 1998 for 1 941 men (1 409 Indians and 532 Creoles) and 2 318 non pregnant women (1 645 Indians and 673 Creoles), free of diabetes, cardiovascular diseases, and gout at baseline examinations in 1987 or 1992, using the same study protocol. The questionnaire was designed to collect demographic details, physical examinations and standard 75g oral glucose tolerance tests were performed in all cohorts. Fasting blood UA and lipid profiles were also determined. The age-standardized prevalence in Chinese living in Qingdao was 25.3% for hyperuricemia (defined as fasting serum UA > 420 μmol/l in men and > 360 μmol/l in women) and 0.36% for gout in adults between 20-74. Hyperuricemia was more prevalent in men than in women. One standard deviation increase in UA concentration was associated with the clustering of metabolic risk factors for both men and women in three ethnic groups. Waist circumference, body mass index, and serum triglycerides appeared to be independently associated with hyperuricemia in both sexes and in all ethnic groups except in Chinese women, in whom triglycerides, high-density lipoprotein cholesterol, and total cholesterol were associated with hyperuricemia. Serum UA increased with increasing fasting plasma glucose levels up to a value of 7.0 mmol/l, but significantly decreased thereafter in mainland Chinese. An inverse relationship occurred between 2-h plasma glucose and serum UA when 2-h plasma glucose higher than 8.0 mmol/l. In the prospective study in Mauritius, 337 (17.4%) men and 379 (16.4%) women developed diabetes during the follow-up. Elevated UA levels at baseline increased 1.14-fold in risk of incident diabetes in Indian men and 1.37-fold in Creole men, but no significant risk was observed in women. In conclusion, the prevalence of hyperuricemia was high in Chinese in Qingdao, blood UA was associated with the clustering of metabolic risk factors in Mauritian Indian, Mauritian Creole, and Chinese living in Qingdao, and a high baseline UA level independently predicted the development of diabetes in Mauritian men. The clinical use of UA as a marker of hyperglycemia and other metabolic disorders needs to be further studied. Keywords: Uric acid, Hyperuricemia, Risk factors, Type 2 Diabetes, Incidence, Mauritius, Chinese

Genetic diversity of Atlantic salmon (Salmo salar) and European whitefish (Coregonus lavaretus) in the Baltic Sea

We described the patterns and extent of microsatellite DNA variation in historical and present-day Atlantic salmon (Salmo salar L.) stocks in the Baltic Sea and neighbouring areas, and in European whitefish (Coregonus lavaretus) ecotypes, populations and run-timing types in Finland. Moreover, the amount and pattern of genetic diversity in historical salmon populations before human impact were described, and the proportion of diversity maintained in the present hatchery stocks evaluated. Salmon populations in the Baltic Sea were, on average, significantly less variable than eastern Atlantic populations, and the diversity of landlocked populations (Lakes Vänern, Saimaa, Onega and Ladoga) was in turn significantly lower than that of anadromous salmon populations in the Baltic Sea populations. Within the Baltic Sea, the anadromous populations of Atlantic salmon formed three clear groups, corresponding to the northern (Gulf of Bothnia), eastern (Gulf of Finland and eastern Baltic Main Basin) and southern (western Baltic Main Basin) regions. Based on microsatellite data, three salmon population groups in the Baltic Sea were considered potentially different colonization lineages. In short- and long-term breeding programmes of Atlantic salmon, the average observed rate of loss of alleles was 4.9% and 2.0% per generation and the average rate of loss of heterozygosity was 1.4% and 1% per generation, respectively. When comparing the genetic parameters of stocks before and after hatchery breeding of several successive generations (Rivers Iijoki and Oulujoki), statistically significant changes in allele frequencies were common, while large wild stock in the Teno River has remained temporally very stable over 56 years. Despite the observed losses of genetic diversity in broodstock breeding, a large proportion of the genetic resources of the extirpated stocks are still conserved in the broodstocks. Genetic differentiation among European whitefish ecotypes was generally low, thus giving support to the hypothesis of one native European whitefish species in Fennoscandia. Among the ecotypes, the northern, large sparsely rakered, bottom-dwelling whitefish was the most unique. The known genetic differences in quantitative traits have thus either developed independently of potential phylogenetic lineages, or the lineages have mixed and the quantitative traits of the ecotypes, like gill-raker number, have later changed according to environment and selection pressures. Overall, genetic distances between the anadromous whitefish populations along the Finnish coast, especially in the Bothnian Bay area, were small. Wild whitefish populations studied had slightly higher allelic diversity than hatchery-reared populations in corresponding rivers.

Rhizoctonia solani as a potato pathogen : Variation of isolates in Finland and host response

Rhizoctonia solani is a soil inhabiting basidiomycetous fungus able to induce a wide range of symptoms in many plant species. This genetically complex species is divided to 13 anastomosis groups (AG), of which AG-3 is specialized to infect potato. However, also a few other AGs are able to infect or live in close contact with potato. On potato, R. solani infection causes two main types of diseases including stem canker observed as a dark brown lesions on developing stems and stolons, and black scurf that develops on new tubers close to the time of harvest. These disease symptoms are collectively called a ‘Rhizoctonia disease complex’. Between the growing seasons R. solani survives in soil and plant debri as sclerotia or as the sclerotia called black scurf on potato tubers which when used as seed offer the main route for dispersal of the fungus to new areas. The reasons for the dominance of AG-3 on potato seem to be attributable to its highly specialization to potato and its ability to infect and form sclerotia efficiently at low temperatures. In this study, a large nationwide survey of R. solani isolates was made in potato crops in Finland. Almost all characterized isolates belonged to AG-3. Additionally, three other AGs (AG-2-1, AG-4 and AG-5) were found associated with symptoms on potato plants but they were weaker pathogens on potato than AG-3 as less prone to form black scurf. According to phylogenetic analysis of the internal transcribed sequences (ITS) of the ribosomal RNA genes the Finnish AG-3 isolates are closely related to each other even though a wide variation of physiological features was observed between them. Detailed analysis of the ITS regions revealed single nucleotide polymorphism in 14 nucleotide positions of ITS-1 and ITS-2. Additionally, compensatory base changes on ITS-2 were detected which suggests that potato-infecting R. solani AG-3 could be considered as a separate species instead of an AG of R. solani. For the first time, molecular defence responses were studied and detected during the early phases of interaction between R. solani AG-3 and potato. Extensive systemic signalling for defence exploiting several known defence pathways was activated as soon as R. solani came into close contact with the base of a sprout. The defence response was strong enough to protect vulnerable sprout tips from new attacks by the pathogen. These results at least partly explain why potato emergence is eventually successful even under heavy infection pressure by R. solani.

Biodiversity and Phylogeny of Planktic Cyanobacteria in Temperate Freshwater Lakes

Currently, the classification used for cyanobacteria is based mainly on morphology. In many cases the classification is known to be incongruent with the phylogeny of cyanobacteria. The evaluation of this classification is complicated by the fact that numerous strains are only described morphologically and have not been isolated. Moreover, the phenotype of many cyanobacterial strains alters during prolonged laboratory cultivation. In this thesis, cyanobacterial strains were isolated from lakes (mainly Lake Tuusulanjärvi) and both morphology and phylogeny of the isolates were investigated. The cyanobacterial community composition in Lake Tuusulanjärvi was followed for two years in order to relate the success of cyanobacterial phenotypes and genotypes to environmental conditions. In addition, molecular biological methods were compared with traditional microscopic enumeration and their ability and usefulness in describing the cyanobacterial diversity was evaluated. The Anabaena, Aphanizomenon, and Trichormus strains were genetically heterogeneous and polyphyletic. The phylogenetic relationships of the heterocytous cyanobacteria were not congruent with their classification. In contrast to heterocytous cyanobacteria, the phylogenetic relationships of the Snowella and Woronichinia strains, which had not been studied before this thesis, reflected the morphology of strains and followed their current classification. The Snowella strains formed a monophyletic cluster, which was most closely related to the Woronichinia strain. In addition, a new cluster of thin, filamentous cyanobacterial strains identified as Limnothrix redekei was revealed. This cluster was not closely related to any other known cyanobacteria. The cyanobacterial community composition in Lake Tuusulanjärvi was studied with molecular methods [denaturant gradient gel electrophoresis (DGGE) and cloning of the 16S rRNA gene], through enumerations of cyanobacteria under microscope, and by strain isolations. Microcystis, Anabaena/Aphanizomenon, and Synechococcus were the major groups in the cyanobacterial community in Lake Tuusulanjärvi during the two-year monitoring period. These groups showed seasonal succession, and their success was related to different environmental conditions. The major groups of the cyanobacterial community were detected by all used methods. However, cloning gave higher estimates than microscopy for the proportions of heterocytous cyanobacteria and Synechococcus. The differences were probably caused by the high 16S rRNA gene copy numbers in heterotrophic cyanobacteria and by problems in the identification and detection of unicellular cyanobacteria.

Traditionally Protected Forests and Sacred Forests of Zigua and Gweno Ethnic Groups in Tanzania

In Tanzania, indigenous forests can still be found whose existence is based on the management systems of precolonial society. This study covers material from over 900 forests. There are similar types of forests elsewhere in Africa, and similar forests can also be found in indigenous cultures on every continent. In this study they are called traditionally protected forests (TPFs). They have a high level of endemism and a rich biodiversity. The field study was carried out during the years 1997-2003 using participatory methods. An active debate is going on concerning the capacity of local communities to manage their environment. The role of indigenous people and their institutions in the development of the physical environment is a central issue in the debate. This study discusses the opportunities that the local people have had to decide on how to conserve, maintain, utilise, and manage their environment during different political periods. The study explains what kinds of changes have taken place in these forests and institutions in northeastern Tanzania among the matrilinear Zigua and patrilinear Gweno ethnic groups. About 2% of the land area of the communities was still protected by the precolonial structures. The communities have established their protection systems for different reasons, not only because of their beliefs but also because of different secular and clearly environmentally motivated reasons. There are different TPF types. Less than half of them are directly related to spirituality, and more than half are not. In earlier research elsewhere, it has been commonly understood that spiritual reasons played the main role in the protection of these environments. This study is also part of the postcolonial geographical discussion on the precolonial landscape and environmental management which was started by Carl Sauer. In the Zigua case study villages, only two out of five first comer clans have performed rain rituals in the past 30 years. Many of the most respected sacred sites do not have a ritual maker or even a person who knows how to perform rituals any longer. The same is happening with male initiation rites. In all case study villages there have been illegal cuts in the TPFs, but variations can be seen between the communities. The number of those who neither respect indigenous regulations nor accept indigenous penalties is growing. Positive developments have also taken place. Nowadays, the Forest Act of 2002 is in effect, which works as a cornerstone of community-based land ownership and also allows elders to protect TPFs, and by-laws are created with the support of different projects. Moreover, during the field study it was found that many young people are ignorant about their village's TPF sites, but interested in learning about their history and values.

Cortinarius subgenus Telamonia p.p. in North Europe

Cortinarius is the largest genus of Agaricales with a worldwide distribution. So far, over 4000 Cortinarius names and combinations have been published. Cortinarius spp. form ectomycorrhizae with different trees and shrubs. A majority of the Cortinarius species have narrow ecological preferences and many form ectomycorrhiza with only one or few host species. The subgenus Telamonia sensu lato (s. lat.), comprising the greatest number of species, is the most poorly known of the subgenera of Cortinarius. The centre of diversity is in the northern hemisphere, although some species of the group are also recognized in the southern hemisphere. The aim of this thesis was to study the taxonomy of Cortinarius subgenus Telamonia p.p. species based on morphological and molecular data, as well as to study the ecology and distribution of the species in North Europe. The taxonomical problems encountered and the difficulty in finding and studying all the relevant names and types slowed down the study. The diversity of the subgenus Telamonia s. lat. in North Europe (excluding sect. Hydrocybe, Icrustati and Anomali) was found to be far greater than previously thought. Even many of the common species have not yet been described. So far, ca. 200 species have been recognised from the Nordic countries, but the sampling in most groups does not cover the whole diversity and especially the southern deciduous forest species are underrepresented in our study. In most cases phylogenetic (only based on ITS data) and morphological species recognition were in concordance, but in a few cases morphologically delimited species had almost identical ITS sequences, raising the question as to whether ITS is always variable enough for species recognition. The opposite situation, in which a morphologically uniform species included two phylogenetically distinct lineages, however, was also encountered, suggesting the possibility of cryptic species in Cortinarius. In our studies no taxa below species level were recognised and the aforementioned results indicate that presumably they can only be recognised genetically. Based on our preliminary results a revision of the infrageneric classification in Cortinarius subgenus Telamonia s. lat. is needed, and more sections should be established for a meaningful and functional classification. Many groups have turned out to be artificial, and it seems evident that many characteristics have been over- or underemphasised. Many morphological characteristics, however, are useful in the identification of telamonioid species and e.g. some spore characteristics have often been overlooked. Our studies have concentrated on North Europe, but we have found some similarities with North European and North American taxa.

Archaea in the Mycorrhizosphere of Boreal Forest Trees

Archaea were long thought to be a group of ancient bacteria, which mainly lived in extreme environments. Due to the development of DNA sequencing methods and molecular phylogenetic analyses, it was shown that the living organisms are in fact divided into three domains; the Archaea, Bacteria and the Eucarya. Since the beginning of the previous decade, it was shown that archaea generally inhabit moderate environments and that these non-extremophilic archaea are more ubiquitous than the extremophiles. Group 1 of non-extreme archaea affiliate with the phylum Crenarchaeota. The most commonly found soil archaea belong to the subgroup 1.1b. However, the Crenarchaeota found in the Fennoscandian boreal forest soil belong to the subgroup 1.1c. The organic top layer of the boreal forest soil, the humus, is dominated by ectomycorrhizal fungal hyphae. These colonise virtually all tree fine root tips in the humus layer and have been shown to harbour distinct bacterial populations different from those in the humus. The archaea have also been shown to colonise both boreal forest humus and the rhizospheres of plants. In this work, studies on the archaeal communities in the ectomycorrhizospheres of boreal forest trees were conducted in microcosms. Archaea belonging to the group 1.1c Crenarchaeota and Euryarchaeota of the genera Halobacterium and Methanolobus were detected. The archaea generally colonised fungal habitats, such as ectomycorrhizas and external mycelia, rather than the non-mycorrhizal fine roots of trees. The species of ectomycorrhizal fungus had a great impact on the archaeal community composition. A stable euryarchaeotal community was detected especially in the mycorrhizas, of most of the tested Scots pine colonising ectomycorrhizal fungi. The Crenarchaeota appeared more sporadically in these habitats, but had a greater diversity than the Euryarchaeota. P. involutus mycorrhizas had a higher diversity of 1.1c Crenarchaeota than the other ectomycorrhizal fungi. The detection level of archaea in the roots of boreal trees was generally low although archaea have been shown to associate with roots of different plants. However, alder showed a high diversity of 1.1c Crenarchaeota, exceeding that of any of the tested mycorrhizas. The archaeal 16S rRNA genes detected from the non-mycorrhizal roots were different from those of the P. involutus mycorrhizas. In the phylogenetic analyses, the archaeal 16S rRNA gene sequences obtained from non-mycorrhizal fine roots fell in a separate cluster within the group 1.1c Crenarchaeota than those from the mycorrhizas. When the roots of the differrent tree species were colonised by P. involutus, the diversity and frequency of the archaeal populations of the different tree species were more similar to each other. Both Cren- and Euryarchaeota were enriched in cultures to which C-1 substrates were added. The 1.1c Crenarchaeota grew anaerobically in mineral medium with CH4 and CO2 as the only available C sources, and in yeast extract media with CO2 and CH4 or H2. The crenarchaeotal diversity was higher in aerobic cultures on mineral medium with CH4 or CH3OH than in the anaerobic cultures. Ecological functions of the mycorrhizal 1.1c Crenarchaeota in both anaerobic and aerobic cycling of C-1 compounds were indicated. The phylogenetic analyses did not divide the detected Crenarchaeota into anaerobic and aerobic groups. This may suggest that the mycorrhizospheric crenarchaeotal communities consist of closely related groups of anaerobic and aerobic 1.1c Crenarchaeota, or the 1.1c Crenarchaeota may be facultatively anaerobic. Halobacteria were enriched in non-saline anaerobic yeast extract medium cultures in which CH4 was either added or produced, but were not detected in the aerobic cultures. They may potentially be involved in anaerobic CH4 cycling in ectomycorrhizas. The CH4 production of the mycorrhizal samples was over 10 times higher than for humus devoid of mycorrhizal hyphae, indicating a high CH4 production potential of the mycorrhizal metanogenic community. Autofluorescent methanogenic archaea were detected by microscopy and 16S rRNA gene sequences of the genus Methanolobus were obtained. The archaeal community depended on both tree species and the type of ectomycorrhizal fungus colonising the roots and the Cren- and Euryarchaeota may have different ecological functions in the different parts of the boreal forest tree rhizosphere and mycorrhizosphere. By employing the results of this study, it may be possible to isolate both 1.1c Crenarchaeota as well as non-halophilic halobacteria and aerotolerant methanogens from mycorrhizospheres. These archaea may be used as indicators for change in the boreal forest soil ecosystem due to different factors, such as exploitations of forests and the rise in global temperature. More information about the microbial populations with apparently low cell numbers but significant ecological impacts, such as the boreal forest soil methanogens, may be of crucial importance to counteract human impacts on such globally important ecosystems as the boreal forests.

Phylogenetic studies of cyanobacterial lichens

Phylogenetic studies of cyanobacterial lichens Lichens are symbiotic assemblages between fungi (mycobiont) and green algae (phycobiont) or/and cyanobacteria (cyanobiont). Fossil records show that lichen-like symbioses occurred already 600 million years ago. Lichen symbiosis has since then become an important life strategy for the Fungi, particularly for species in the phylum Ascomycota as approximately 98% of the lichenized fungal species are ascomycetes. The taxonomy of lichen associations is based on the mycobiont. We reconstructed, using DNA sequence data, hypotheses of phylogenetic relationships of lichen-forming fungi that include species associated with cyanobacteria. These hypotheses of phylogeny should form the basis for the taxonomy. They also allowed studies of the origin and the evolution of specific symbioses. Genetic diversity and phylogenetic relationships of symbiotic cyanobionts were also studied in order to examine selectivity of cyanobionts and mycobionts as well as possible co-evolution between partners involved in lichen associations. The suggested circumscription of the family Stereocaulaceae to include Stereocaulon and Lepraria is supported. The recently described crustose Stereocaulon species seem to be correctly placed in the genus, although Stereocaulon traditionally included only fruticose species. The monospecific crustose genus Muhria is also shown to be best placed in Stereocaulon. Family Lobariaceae as currently delimited is monophyletic. Within Lobariaceae genus Sticta including Dendriscocaulon dendroides form a monophyletic group while the genera Lobaria and Pseudocyphellaria are non-monophyletic. A new classification of Lobariaceae is obviously needed. Further studies are however required before a final proposal for a new classification can be made. Our results show that the cyanobacterial symbiotic state has been gained repeatedly in the Ascomycota while losses of symbiotic cyanobacteria appear to be rare. The symbiosis with green algae is confirmed to have been gained repeatedly in Ascomycota but also repeatedly lost. Cyanobacterial symbioses therefore seem to be more stable than green algal associations. Cyanobacteria are perhaps more beneficial for the lichen fungi and therefore maintained. The results indicate a dynamic association of the lichen symbiosis. This evolutionary instability will perhaps be important for the lichen fungi as the utilization of options will perhaps enable lichens to colonize new substrates and survive environmental changes. Some cyanobacterial lichen genera seem to be highly selective towards the cyanobiont while others form symbioses with a broad spectrum of cyanobacteria. No evidence of co-evolution between fungi and cyanobacteria in cyanolichens could be demonstrated.

Dung beetle radiations in Madagascar

Speciation on islands is affected by island size and the range of habitats and resources available and often also by limited interactions with other taxa. An ancestral population may evolve into a large number of species via an adaptive radiation. In Madagascar, most groups of animals and plants have radiated on the island, having arrived via oceanic dispersal during the long isolation of Madagascar. Characteristic features of Malagasy biota are exceptionally high level of endemism, high species richness as well as lack of many higher taxa that are dominant on the African mainland. Malagasy dung beetles are dominated by two tribes, Canthonini and Helictopleurina, with more than 250 endemic species. In this thesis I have reconstructed molecular phylogenies for the two tribes using several gene regions and different phylogenetic methods. Evolution of closely related species and among populations of the same species was examined with haplotype networks. The Malagasy Canthonini consists of three large lineages, while Helictopleurina forms a monophyletic group. The ancestors of each of the four clades colonised Madagascar at different times during Cenozoic. The subsequent radiations differ in terms of the number of extant species (from 37 to more than 100) and the level of ecological differentiation. In addition, Onthophagini (6 species) and Scarabaeini (3) have colonised Madagascar several times, but they have not radiated and the few species have not entered forests where Canthonini and Helictopleurina mostly occur. Among the three Canthonini radiations, speciation appears to have been mostly allopatric in the oldest and the youngest clades, while in the Epactoides clade sister species have diverged in their ecologies but have similar geographical distributions, indicating that speciation may have occurred in regional sympatry. The most likely isolating mechanisms have been rivers and forest refugia during dry and cool geological periods. Most species are generalists feeding on both carrion and dung, and competition among ecologically similar species may prevent their coexistence in the same communities. Some species have evolved to forage in the canopy and a few species have shifted to use cattle dung, a new resource in the open habitats following the introduction of cattle 1500 years ago. The latter shift has allowed species to expand their geographical ranges.

Population structure and evolution in the ant Plagiolepis pygmaea and its two social parasites Plagiolepis xene and Plagiolepis grassei

Social behaviour affects dispersal of animals and is an important modifier of genetic population structures. The female sex is often philopatric, which maintains coancestry within the breeding groups and promotes cooperative behaviours. This enables also inclusive fitness returns from altruism and explains why some individuals sacrifice personal reproduction for the good of others in social insects such as ants. However, reduced dispersal and population substructuring at the level of colonies may also entail inbreeding, loss of genetic diversity, and vulnerability. In addition, the most vulnerable ants are species that are evolved to parasitize colonies of other ants, and which compromise between abilities to disperse and the efficiency to parasitize the host. On the other hand, certain social organisations of ant colonies may facilitate a species to disperse outside its natural range and become a pest. Altogether, knowledge on genetic structuring of ant populations, as well as the evolution of their life histories can contribute to conservation biology and population management. The aim of this thesis was to investigate population structures and phylogenetic evolution of the ant Plagiolepis pygmaea and its two obligatory, workerless social parasites (inquilines) P. xene and P. grassei with genetic markers and DNA sequence data. The results support the general assumption that populations of inquiline parasites are highly fragmented and genetically vulnerable. Comparison of the two parasites suggests that differences in their relative abundance may follow from their interaction with the host, i.e. how well the species is adapted to reproduce in the host colonies. The results also indicate that the most recent free living ancestor to these two parasite species is their common host. This is considered to provide evidence for the controversial issue of sympatric speciation. Further, given that the level of adaptations to parasitic life history depends on the evolutionary time since the free-living ancestor, the results establish a link between species rarity and its evolutionary age. The populations of the host species P. pygmaea displayed significantly reduced dispersal both among the females (queens) and males, and high levels of inbreeding which may enhance worker altruism. In addition, the queens were found to mate with multiple males. Given the high relatedness between the queens and their mates, this occurs probably for non-genetic reasons, e.g. without benefits associated in genetically more diverse offspring. The results hence caution that the contribution of non-genetic factors to the prevailing mating patterns and genetic population structures should not be underestimated.