Mutations as molecular tools : The metabolic-rate dependent molecular clock and DNA barcoding of allied species

Mutation and recombination are the fundamental processes leading to genetic variation in natural populations. This variation forms the raw material for evolution through natural selection and drift. Therefore, studying mutation rates may reveal information about evolutionary histories as well as phylogenetic interrelationships of organisms. In this thesis two molecular tools, DNA barcoding and the molecular clock were examined. In the first part, the efficiency of mutations to delineate closely related species was tested and the implications for conservation practices were assessed. The second part investigated the proposition that a constant mutation rate exists within invertebrates, in form of a metabolic-rate dependent molecular clock, which can be applied to accurately date speciation events. DNA barcoding aspires to be an efficient technique to not only distinguish between species but also reveal population-level variation solely relying on mutations found on a short stretch of a single gene. In this thesis barcoding was applied to discriminate between Hylochares populations from Russian Karelia and new Hylochares findings from the greater Helsinki region in Finland. Although barcoding failed to delineate the two reproductively isolated groups, their distinct morphological features and differing life-history traits led to their classification as two closely related, although separate species. The lack of genetic differentiation appears to be due to a recent divergence event not yet reflected in the beetles molecular make-up. Thus, the Russian Hylochares was described as a new species. The Finnish species, previously considered as locally extinct, was recognized as endangered. Even if, due to their identical genetic make-up, the populations had been regarded as conspecific, conservation strategies based on prior knowledge from Russia would not have guaranteed the survival of the Finnish beetle. Therefore, new conservation actions based on detailed studies of the biology and life-history of the Finnish Hylochares were conducted to protect this endemic rarity in Finland. The idea behind the strict molecular clock is that mutation rates are constant over evolutionary time and may thus be used to infer species divergence dates. However, one of the most recent theories argues that a strict clock does not tick per unit of time but that it has a constant substitution rate per unit of mass-specific metabolic energy. Therefore, according to this hypothesis, molecular clocks have to be recalibrated taking body size and temperature into account. This thesis tested the temperature effect on mutation rates in equally sized invertebrates. For the first dataset (family Eucnemidae, Coleoptera) the phylogenetic interrelationships and evolutionary history of the genus Arrhipis had to be inferred before the influence of temperature on substitution rates could be studied. Further, a second, larger invertebrate dataset (family Syrphidae, Diptera) was employed. Several methodological approaches, a number of genes and multiple molecular clock models revealed that there was no consistent relationship between temperature and mutation rate for the taxa under study. Thus, the body size effect, observed in vertebrates but controversial for invertebrates, rather than temperature may be the underlying driving force behind the metabolic-rate dependent molecular clock. Therefore, the metabolic-rate dependent molecular clock does not hold for the here studied invertebrate groups. This thesis emphasizes that molecular techniques relying on mutation rates have to be applied with caution. Whereas they may work satisfactorily under certain conditions for specific taxa, they may fail for others. The molecular clock as well as DNA barcoding should incorporate all the information and data available to obtain comprehensive estimations of the existing biodiversity and its evolutionary history.

Molecular genetics and evolution of Puumala hantavirus

Puumala virus (PUUV) is the causative agent of nephropathia epidemica (NE), a mild form of hemorrhagic fever with renal syndrome. Finland has the highest documented incidence of NE with around 1000 cases diagnosed annually. PUUV is also found in other Scandinavian countries, Central Europe and the European part of Russia. PUUV belongs to the genus Hantavirus in the family Bunyaviridae. Hantaviruses are rodent-borne viruses each carried by a specific host that is persistently and asymptomatically infected by the virus. PUUV is carried by the bank voles (Myodes glareolus, previously known as Clethrionomys glareolus). Hantaviruses have co-evolved with their carrier rodents for millions of years and these host animals are the evolutionary scene of hantaviruses. In this study, PUUV sequences were recovered from bank voles captured in Denmark and Russian Karelia to study the evolution of PUUV in Scandinavia. Phylogenetic analysis of these strains showed a geographical clustering of genetic variants following the presumable migration pattern of bank voles during the recolonization of Scandinavia after the last ice age approximately 10 000 years ago. The currently known PUUV genome sequences were subjected to in-depth phylogenetic analyses and the results showed that genetic drift seems to be the major mechanism of PUUV evolution. In general, PUUV seems to evolve quite slowly following a molecular clock. We also found evidence for recombination in the evolution of some genetic lineages of PUUV. Viral microevolution was studied in controlled virus transmission in colonized bank voles and changes in quasispecies dynamics were recorded as the virus was transmitted from one animal to another. We witnessed PUUV evolution in vivo, as one synonymous mutation became repeatedly fixed in the viral genome during the experiment. The detailed knowledge on the PUUV diversity was used to establish new sensitive and specific detection methods for this virus. Direct viral invasion of the hypophysis was demonstrated for the first time in a lethal case of NE. PUUV detection was done by immunohistochemistry, in situ hybridization and RT-nested-PCR of the autopsy tissue samples.

Circadian Rhythm Disruptions and Health

The doctoral thesis defined connections between circadian rhythm disruptions and health problems. Sleep debt, jet-lag, shift work, as well as transitions into and out of the daylight saving time may lead to circadian rhythm disruptions. Disturbed circadian rhythm causes sleep deprivation and decrease of mood and these effects may lead to higher accident rates and trigger mental illnesses. Circadian clock genes are involved in the regulation of the cell cycle and metabolism and thus unstable circadian rhythmicity may also lead to cancer development. In publications I-III it was explored how transitions into and out of the daylight saving time impact the sleep efficiency and the rest-activity cycles of healthy individuals. Also it was explored whether the effect of transition is different in fall as compared to spring, and whether there are subgroup specific differences in the adjustment to transitions into and out of daylight saving time. The healthy participants of studies I-III used actigraphs before and after the transitions and filled in the morningness-eveningness and seasonal pattern assessment questionnaires. In publication IV the incidence of hospital-treated accidents and manic episodes was explored two weeks before and two weeks after the transitions into and out of the daylight saving time in years 1987-2003. In publication V the relationship between circadian rhythm disruption and the prevalence of Non-Hodgkin lymphoma was studied. The study V consisted of all working aged Finns who participated in the national population census in 1970. For our study, all the cancers diagnosed during the years 1971-1995 were extracted from the Finnish Cancer Register and linked with the 1970 census files. In studies I-III it was noticed that transitions into and out of the daylight saving time disturbs the sleep-wake cycle and the sleep efficiency of the healthy participants. We also noticed that short sleepers were more sensitive than long sleepers for sudden changes in the circadian rhythm. Our results also indicated that adaptation to changes in the circadian rhythm is potentially sex, age and chronotype-specific. In study IV no significant increase in the occurrence of hospital treated accidents or manic episodes was noticed. However, interesting observations about the seasonal fluctuation of the occurrence rates of accidents and manic episodes were made. Study V revealed that there might be close relationship between circadian rhythm disruption and cancer. The prevalence of Non-Hodgkin lymphoma was the highest among night workers. The five publications included in this thesis together point out that disturbed circadian rhythms may have adverse effect on health. Disturbed circadian rhythms decrease the quality of sleep and weaken the sleep-wake cycle. A continuous circadian rhythm disruption may also predispose individuals to cancer development. Since circadian rhythm disruptions are common in modern society they might have a remarkable impact on the public health. Thus it is important to continue circadian rhythm research so that better prevention and treatment methods can be developed. Keywords: Circadian rhythm, daylight saving time, manic episodes, accidents, Non-Hodgkin lymphoma 11

Yrittäjyyden arvottaminen akateemisten nuorten argumentoinnissa

Evaluation of entrepreneurship in the speech of academic students and newly qualified young academics a summary of a qualitative attitude study. In Finland very few university students plan to become entrepreneurs. The aim of this research was to examine entrepreneurial attitudes expressed in speech. The material was gathered from interviews with university students and newly qualified young academic adults. The interviewees commented on twelve different sentences with claims formulated using research literature and views that have appeared in public discussions. The interviewees were divided into three different groups based on their self-expressed entrepreneurial intentions. The method of qualitative attitude research (Vesala & Rantanen 1999, 2007) was used in the interviews. The research material was studied using two interpretative theories: (1) The planned behaviour theory (Ajzen 1985, 1991a, b), which makes it possible to focus on the separate elements (attitude towards an act, subjective norms and perceived feasibility) necessary for intentions to develop; and (2) The theory of the two images of entrepreneurship (Vesala 1996), where individualism and relationism can be seen as resources for evaluating entrepreneurship. The subject of the research was how university students and newly qualified young adults viewed entrepreneurship as a general phenomen and in relation to the academic world. A second focus was on the attitudes expressed toward entrepreneurial university education and the possibility of combining entrepreneurship and academic knowledge. Of interest were also questions such as whether academic studies, knowledge and the university itself are resources or barriers to entrepreneurial intentions and entrepreneurship whether university students received any support for their entrepreneurial ambitions from the university and their fellow academic students. The problems tackled by this research were thus the following: How was entrepreneurship seen, both as a general phenomen and in an academic context, when it was evaluated positively, negatively or neutrally by the interviewees? In what way was entrepreneurship constructed in the interviewees attitudes? How were entrepreneurship and the academic world related in the interviewees attitudes? What kind of role did the university as an academic context play in the interviewees attitudes for example were university education and academic knowledge seen as resources or barriers to their entrepreneurial intentions. Traditional attitude studies claim that attitudes are a stable property of an individual. In contrast, rhetorical social psychological and qualitative attitude studies emphasize the contextual and linguistic aspects of attitude, and they offered an alternative viewpoint for this research. The study was based on two general assumptions: attitudes have objects and are evaluative. Here attitude was defined as an evaluative interpresentation made towards an object; adopting an attitude is a contextual process in the sense that attitudes are always concerned with the action context of the persons presenting them. Entrepreneurship, both as a general phenomen and in an academic context, was specified as the object to which an attitude was taken. From a theoretical point of view, qualitative methods suited the general structure of this research well. In a particular, qualitative approach which emphasized contextual elements proved to be both empirically valid and useful for avoiding the problematic assumptions associated with traditional attitude study. The subject of the analysis was the argumentative speech produced by the interviewees. The results of the study show the subjects responses to three main ways of viewing entrepreneurships. The first was an individualistic, ideal image of entrepreneurship. This was mostly evaluated positively and gained wide approval especially among interviewees who included entrepreneurship among their employment choices. Entrepreneurship was seen as the decision to earn one s living independently. In this individualistic image of entrepreneurship, the social context was hardly ever mentioned. Elements which were seen to threaten this ideal image were evaluated negatively. When entrepreneurship was evaluated negatively using the individualistic image of entrepreneurship, it was mentioned that it forced one into a never ending cycle of work and uninterested duties. The relationistic image of entrepreneurship was used as a speech resource when the social context was constructed as an economic resource or a threat to the ideal image of entrepreneurship. In the second view, entrepreneurship was characteristically seen as being based on economics, which was seen as a threat to the ideal individualistic image of entrepreneurship. The risk of economic failure was seen as a limiting factor to entrepreneurial ambitions as it forced entrepreneurs to work around the clock. The third view concerned the relationship between entrepreneurship and the academic world. Entrepreneurship as an employment choice for university educated persons was evaluated as relevant, and thus positively, when university education was constructed as a resource for entrepreneurship - and irrelevant and thus negatively when it was construed as an obstacle, too wide, or when successful entrepreneurship was seen as being mostly based on an individual s personal characteristics. The interviewees with no entrepreneurial intentions expressed the view that academic education didn t provide the proper skills and knowledge for entrepreneurship. The interviewees also expressed interest in university entrepreneurship education, although none had experience on this. The interviewees emphasized the fact that the University didn t encourage them to consider entrepreneurship as a relevant employment choice. The assumption made by this study was that becoming an entrepreneur is a conscious decision, the environment may influence an individual s decisions on how to make a living as it tends to socialise people to act in accordance with cultural traditions. Keywords: Entrepreneurship, Attitudes towards entrepreneurship, Intentional behaviour, Entrepreneurial intention, University entrepreneurship education, Qualitative attitude research (Vesala & Rantanen 1999, 2007), Rhetorical social psychology (Billig 1986), The theory of entrepreneuship s two images: individualism and relationism (Vesala 1996 ), The planned behaviour theory (Ajzen 1985, 1991a, b)

Domestic labour relations in India : Vulnerability and gendered life courses in Jaipur

This study explores labour relations between domestic workers and employers in India. It is based on interviews with both employers and workers, and ethnographically oriented field work in Jaipur, carried out in 2004-2007. Combining development studies with gender studies, labour studies, and childhood studies, it asks how labour relations between domestic workers and employers are formed in Jaipur, and how female domestic workers trajectories are created. Focusing on female part-time maids and live-in work arrangements, the study analyses children s work in the context of overall work force, not in isolation from it. Drawing on feminist Marxism, domestic labour relations are seen as an arena of struggle. The study takes an empirical approach, showing class through empiria and shows how paid domestic work is structured and stratified through intersecting hierarchies of class, caste, gender, age, ethnicity and religion. The importance of class in domestic labour relations is reiterated, but that of caste, so often downplayed by employers, is also emphasized. Domestic workers are crucial to the functioning of middle and upper middle class households, but their function is not just utilitarian. Through them working women and housewives are able to maintain purity and reproduce class disctinctions, both between poor and middle classes and lower and upper middle classes. Despite commodification of work relations, traditional elements of service relationships have been retained, particularly through maternalist practices such as gift giving, creating a peculiar blend of traditional and market practices. Whilst employers of part-time workers purchase services in a segmented market from a range of workers for specific, traditional live-in workers are also hired to serve employers round the clock. Employers and workers grudgingly acknowledged their dependence on one another, employers seeking various strategies to manage fear of servant crime, such as the hiring of children or not employing live-in workers in dual-earning households. Paid domestic work carries a heavy stigma and provide no entry to other jobs. It is transmitted from mothers to daughters and working girls were often the main income providers in their families. The diversity of working conditions is analysed through a continuum of vulnerability, generic live-in workers, particularly children and unmarried young women with no close family in Jaipur, being the most vulnerable and experienced part-time workers the least vulnerable. Whilst terms of employment are negotiated informally and individually, some informal standards regarding salary and days off existed for maids. However, employers maintain that workings conditions are a matter of individual, moral choice. Their reluctance to view their role as that of employers and the workers as their employees is one of the main stumbling blocks in the way of improved working conditions. Key words: paid domestic work, India, children s work, class, caste, gender, life course