Phylogenetics of Myrmica ants and their social parasites

Understanding the overwhelming diversity of life calls for complex organisational schemes. The field of systematics may thus be seen as the cornerstone of evolutionary biology. In the last few decades, systematics has been rejuvenated through the introduction of molecular methods such as DNA barcoding and multi-gene phylogenetic approaches. These methods may shed new light on established taxonomic ideas and problems. For example, the classification of ants has aroused much debate due to reinterpretation of morphological characters or contradictions between molecular data and morphology. Only in the last few years a consensus was reached regarding the phylogeny of ant subfamilies. However, the situation remains deplorable for lower taxonomic ranks such as subfamilies, tribes and genera. This thesis describes the systematics and evolution of the Holarctic ant genus Myrmica and the tribe to which it belongs, Myrmicini. Using barcoding, molecular-phylogenetic data and divergence time estimations, it addresses questions regarding the taxonomy, morphology and biogeography of this group. Furthermore, the interrelationships between socially parasitic Myrmica species and their hosts (other species in the genus) were inferred. The phylogeny suggests that social parasitism evolved several times in Myrmica. Finally, this thesis investigated whether coevolution shaped the phylogeny of socially parasitic Maculinea butterflies that live inside Myrmica colonies. No evidence was found for coevolution.

Actin Dynamics in Muscle Cells

In every cell, actin is a key component involved in migration, cytokinesis, endocytosis and generation of contraction. In non-muscle cells, actin filaments are very dynamic and regulated by an array of proteins that interact with actin filaments and/or monomeric actin. Interestingly, in non-muscle cells the barbed ends of the filaments are the predominant assembly place, whereas in muscle cells actin dynamics was reported to predominate at the pointed ends of thin filaments. The actin-based thin filament pointed (slow growing) ends extend towards the middle of the sarcomere's M-line where they interact with the thick filaments to generate contraction. The actin filaments in muscle cells are organized into a nearly crystalline array and are believed to be significantly less dynamic than the ones in other cell types. However, the exact mechanisms of the sarcomere assembly and turnover are largely unknown. Interestingly, although sarcomeric actin structures are believed to be relatively non-dynamic, many proteins promoting actin dynamics are expressed also in muscle cells (e.g ADF/cofilin, cyclase-associated protein and twinfilin). Thus, it is possible that the muscle-specific isoforms of these proteins promote actin dynamics differently from their non-muscle counterparts, or that actin filaments in muscle cells are more dynamic than previously thought. To study protein dynamics in live muscle cells, I used primary cell cultures of rat cardiomyocytes. My studies revealed that a subset of actin filaments in cardiomyocyte sarcomeres displays rapid turnover. Importantly, I discovered that the turnover of actin filaments depends on contractility of the cardiomyocytes and that the contractility-induced actin dynamics plays an important role in sarcomere maturation. Together with previous studies those findings suggest that sarcomeres undergo two types of actin dynamics: (1) contractility-dependent turnover of whole filaments and (2) regulatory pointed end monomer exchange to maintain correct thin filament length. Studies involving an actin polymerization inhibitor suggest that the dynamic actin filament pool identified here is composed of filaments that do not contribute to contractility. Additionally, I provided evidence that ADF/cofilins, together with myosin-induced contractility, are required to disassemble non-productive filaments in developing cardiomyocytes. In addition, during these studies we learned that isoforms of actin monomer binding protein twinfilin, Twf-1 and Twf-2a localise to myofibrils in cardiomyocytes and may thus contribute to actin dynamics in myofibrils. Finally, in collaboration with Roberto Dominguez s laboratory we characterized a new actin nucleator in muscle cells - leiomodin (Lmod). Lmod localises towards actin filament pointed ends and its depletion by siRNA leads to severe sarcomere abnormalities in cardiomyocytes. The actin filament nucleation activity of Lmod is enhanced by interactions with tropomyosin. We also revealed that Lmod expression correlates with the maturation of myofibrils, and that it associates with sarcomeres only at relatively late stages of myofibrillogenesis. Thus, Lmod is unlikely to play an important role in myofibril formation, but rather might be involved in the second step of the filament arrangement and/or maintenance through its ability to promote tropomyosin-induced actin filament nucleation occurring at the filament pointed ends. The results of these studies provide valuable new information about the molecular mechanisms underlying muscle sarcomere assembly and turnover. These data offer important clues to understanding certain physiological and pathological behaviours of muscle cells. Better understanding of the processes occurring in muscles might help to find strategies for determining, diagnosis, prognosis and therapy in heart and skeletal muscles diseases.

The Emerging Phenotype: Ecological Genetics of Color, Form and Sex in the Common Frog

One of the main aims of evolutionary biology is to explain why organisms vary phenotypically as they do. Proximately, this variation arises from genetic differences and from environmental influences, the latter of which is referred to as phenotypic plasticity. Phenotypic plasticity is thus a central concept in evolutionary biology, and understanding its relative importance in causing the phenotypic variation and differentiation is important, for instance in anticipating the consequences of human induced environmental changes. The aim of this thesis was to study geographic variation and local adaptation, as well as sex ratios and environmental sex reversal, in the common frog (Rana temporaria). These themes cover three different aspects of phenotypic plasticity, which emerges as the central concept for the thesis. The first two chapters address geographic variation and local adaptation in two potentially thermally adaptive traits, namely the degree of melanism and the relative leg length. The results show that although there is an increasing latitudinal trend in the degree of melanism in wild populations across Scandinavian Peninsula, this cline has no direct genetic basis and is thus environmentally induced. The second chapter demonstrates that although there is no linear, latitudinally ordered phenotypic trend in relative leg length that would be expected under Allen s rule an ecogeographical rule linking extremity length to climatic conditions there seems to be such a trend at the genetic level, hidden under environmental effects. The first two chapters thus view phenotypic plasticity through its ecological role and evolution, and demonstrate that it can both give rise to phenotypic variation and hide evolutionary patterns in studies that focus solely on phenotypes. The last three chapters relate to phenotypic plasticity through its ecological and evolutionary role in sex determination, and consequent effects on population sex ratio, genetic recombination and the evolution of sex chromosomes. The results show that while sex ratios are strongly female biased and there is evidence of environmental sex reversals, these reversals are unlikely to have caused the sex ratio skew, at least directly. The results demonstrate that environmental sex reversal can have an effect on the evolution of sex chromosomes, as the recombination patterns between them seem to be controlled by phenotypic, rather than genetic, sex. This potentially allows Y chromosomes to recombine, lending support for the recent hypothesis suggesting that sex-reversal may play an important role on the rejuvenation of Y chromosomes.

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.

Habitat use and population dynamics of brown bears (Ursus arctos) in Scandinavia

Large carnivore populations are currently recovering from past extirpation efforts and expanding back into their original habitats. At the same time human activities have resulted in very few wilderness areas left with suitable habitats and size large enough to maintain populations of large carnivores without human contact. Consequently the long-term future of large carnivores depends on their successful integration into landscapes where humans live. Thus, understanding their behaviour and interaction with surrounding habitats is of utmost importance in the development of management strategies for large carnivores. This applies also to brown bears (Ursus arctos) that were almost exterminated from Scandinavia and Finland at the turn of the century, but are now expanding their range with the current population estimates being approximately 2600 bears in Scandinavia and 840 in Finland. This thesis focuses on the large-scale habitat use and population dynamics of brown bears in Scandinavia with the objective to develop modelling approaches that support the management of bear populations. Habitat analysis shows that bear home ranges occur mainly in forested areas with a low level of human influence relative to surrounding areas. Habitat modelling based on these findings allows identification and quantification of the potentially suitable areas for bears in Scandinavia. Additionally, this thesis presents novel improvements to home range estimation that enable realistic estimates of the effective area required for the bears to establish a home range. This is achieved through fitting to the radio-tracking data to establish the amount of temporal autocorrelation and the proportion of time spent in different habitat types. Together these form a basis for the landscape-level management of the expanding population. Successful management of bears requires also assessment of the consequences of harvest on the population viability. An individual-based simulation model, accounting for the sexually selected infanticide, was used to investigate the possibility of increasing the harvest using different hunting strategies, such as trophy harvest of males. The results indicated that the population can sustain twice the current harvest rate. However, harvest should be changed gradually while carefully monitoring the population growth as some effects of increased harvest may manifest themselves only after a time-delay. The results and methodological improvements in this thesis can be applied to the Finnish bear population and to other large carnivores. They provide grounds for the further development of spatially-realistic management-oriented models of brow bear dynamics that can make projections of the future distribution of bears while accounting for the development of human activities.

Co-Signaling by Neurotrophic Factors and the Extracellular Matrix for Axonal Growth and Neuronal Survival

Neurotrophic factors (NTFs) and the extracellular matrix (ECM) are important regulators of axonal growth and neuronal survival in mammalian nervous system. Understanding of the mechanisms of this regulation is crucial for the development of posttraumatic therapies and drug intervention in the injured nervous system. NTFs act as soluble, target-derived extracellular regulatory molecules for a wide range of physiological functions including axonal guidance and the regulation of programmed cell death in the nervous system. The ECM determines cell adhesion and regulates multiple physiological functions via short range cell-matrix interactions. The present work focuses on the mechanisms of the action of NTFs and the ECM on axonal growth and survival of cultured sensory neurons from dorsal root ganglia (DRG). We first examined signaling mechanisms of the action of the glial cell line-derived neurotrophic factor (GDNF) family ligands (GFLs) on axonal growth. GDNF, neurturin (NRTN) and artemin (ART) but not persephin (PSPN) promoted axonal initiation in cultured DRG neurons from young adult mice. This effect required Src family kinase (SFK) activity. In neurons from GFRalpha2-deficient mice, NRTN did not significantly promote axonal initiation. GDNF and NRTN induced extensive lamellipodia formation on neuronal somata and growth cones. This study suggested that GDNF, NRTN and ARTN may serve as stimulators of nerve regeneration under posttraumatic conditions. Consequently we studied the convergence of signaling pathways induced by NTFs and the ECM molecule laminin in the intracellular signaling network that regulates axonal growth. We demonstrated that co-stimulation of DRG neurons with NTFs (GDNF, NRTN or nerve growth factor (NGF)) and laminin leads to axonal growth that requires activation of SFKs. A different, SFK-independent signaling pathway evoked axonal growth on laminin in the absence of the NTFs. In contrast, axonal branching was regulated by SFKs both in the presence and in the absence of NGF. We proposed and experimentally verified a Boolean model of the signaling network triggered by NTFs and laminin. Our results put forward an approach for predictable, Boolean logics-driven pharmacological manipulation of a complex signaling network. Finally we found that N-syndecan, the receptor for the ECM component HB-GAM was required for the survival of neonatal sensory neurons in vitro. We demonstrated massive cell death of cultured DRG neurons from mice deficient in the N-syndecan gene as compared to wild type controls. Importantly, this cell death could not be prevented by NGF the neurotrophin which activates multiple anti-apoptotic cascades in DRG neurons. The survival deficit was observed during first postnatal week. By contrast, DRG neurons from young adult N-syndecan knock-out mice exhibited normal survival. This study identifies a completely new syndecan-dependent type of signaling that regulates cell death in neurons.

Stress and developmental responses in Arabidopsis thaliana: regulation through the transcription factor-interacting protein RCD1

Plants constantly face adverse environmental conditions, such as drought or extreme temperatures that threaten their survival. They demonstrate astonishing metabolic flexibility in overcoming these challenges and one of the key responses to stresses is changes in gene expression leading to alterations in cellular functions. This is brought about by an intricate network of transcription factors and associated regulatory proteins. Protein-protein interactions and post-translational modifications are important steps in this control system along with carefully regulated degradation of signaling proteins. This work concentrates on the RADICAL-INDUCED CELL DEATH1 (RCD1) protein which is an important regulator of abiotic stress-related and developmental responses in Arabidopsis thaliana. Plants lacking this protein function display pleiotropic phenotypes including sensitivity to apoplastic reactive oxygen species (ROS) and salt, ultraviolet B (UV-B) and paraquat tolerance, early flowering and senescence. Additionally, the mutant plants overproduce nitric oxide, have alterations in their responses to several plant hormones and perturbations in gene expression profiles. The RCD1 gene is transcriptionally unresponsive to environmental signals and the regulation of the protein function is likely to happen post-translationally. RCD1 belongs to a small protein family and, together with its closest homolog SRO1, contains three distinguishable domains: In the N-terminus, there is a WWE domain followed by a poly(ADP-ribose) polymerase-like domain which, despite sequence conservation, does not seem to be functional. The C-terminus of RCD1 contains a novel domain called RST. It is present in RCD1-like proteins throughout the plant kingdom and is able to mediate physical interactions with multiple transcription factors. In conclusion, RCD1 is a key point of signal integration that links ROS-mediated cues to transcriptional regulation by yet unidentified means, which are likely to include post-translational mechanisms. The identification of RCD1-interacting transcription factors, most of whose functions are still unknown, opens new avenues for studies on plant stress as well as developmental responses.

The role of MADS and TCP transcription factors in Gerbera hybrida flower development

Angiosperms represent a huge diversity in floral structures. Thus, they provide an attractive target for comparative developmental genetics studies. Research on flower development has focused on few main model plants, and studies on these species have revealed the importance of transcription factors, such as MADS-box and TCP genes, for regulating the floral form. The MADS-box genes determine floral organ identities, whereas the TCP genes are known to regulate flower shape and the number of floral organs. In this study, I have concentrated on these two gene families and their role in regulating flower development in Gerbera hybrida, a species belonging to the large sunflower family (Asteraceae). The Gerbera inflorescence is comprised of hundreds of tightly clustered flowers that differ in their size, shape and function according to their position in the inflorescence. The presence of distinct flower types tells Gerbera apart from the common model species that bear only single kinds of flowers in their inflorescences. The marginally located ray flowers have large bilaterally symmetrical petals and non-functional stamens. The centrally located disc flowers are smaller, have less pronounced bilateral symmetry and carry functional stamens. Early stages of flower development were studied in Gerbera to understand the differentiation of flower types better. After morphological analysis, we compared gene expression between ray and disc flowers to reveal transcriptional differences in flower types. Interestingly, MADS-box genes showed differential expression, suggesting that they might take part in defining flower types by forming flower-type-specific regulatory complexes. Functional analysis of a CYCLOIDEA-like TCP gene GhCYC2 provided evidence that TCP transcription factors are involved in flower type differentiation in Gerbera. The expression of GhCYC2 is ray-flower-specific at early stages of development and activated only later in disc flowers. Overexpression of GhCYC2 in transgenic Gerbera-lines causes disc flowers to obtain ray-flower-like characters, such as elongated petals and disrupted stamen development. The expression pattern and transgenic phenotypes further suggest that GhCYC2 may shape ray flowers by promoting organ fusion. Cooperation of GhCYC2 with other Gerbera CYC-like TCP genes is most likely needed for proper flower type specification, and by this means for shaping the elaborate inflorescence structure. Gerbera flower development was also approached by characterizing B class MADS-box genes, which in the main model plants are known regulators of petal and stamen identity. The four Gerbera B class genes were phylogenetically grouped into three clades; GGLO1 into the PI/GLO clade, GDEF2 and GDEF3 into the euAP3 clade and GDEF1 into the TM6 clade. Putative orthologs for GDEF2 and GDEF3 were identified in other Asteraceae species, which suggests that they appeared through an Asteraceae-specific duplication. Functional analyses indicated that GGLO1 and GDEF2 perform conventional B-function as they determine petal and stamen identities. Our studies on GDEF1 represent the first functional analysis of a TM6-like gene outside the Solanaceae lineage and provide further evidence for the role of TM6 clade members in specifying stamen development. Overall, the Gerbera B class genes showed both commonalities and diversifications with the conventional B-function described in the main model plants.

Kin selection, social polymorphism, and reproductive allocation in ants

Social groups are common across animal species. The reasons for grouping are straightforward when all individuals gain directly from cooperating. However, the situation becomes more complex when helping entails costs to the personal reproduction of individuals. Kin selection theory has offered a fruitful framework to explain such cooperation by stating that individuals may spread their genes not only through their own reproduction, but also by helping related individuals reproduce. However, kin selection theory also implicitly predicts conflicts when groups consist of non-clonal individuals, i.e. relatedness is less than one. Then, individual interests are not perfectly aligned, and each individual is predicted to favour the propagation of their own genome over others. Social insects provide a solid study system to study the interplay between cooperation and conflict. Breeding systems in social insects range from solitary breeding to eusocial colonies displaying complete division of reproduction between the fertile queen and the sterile worker caste. Within colonies, additional variation is provided by the presence of several reproductive individuals. In many species, the queen mates multiply, which causes the colony to consist of half-sib instead of full-sib offspring. Furthermore, in many species colonies contain multiple breeding queens, which further dilutes relatedness between colony members. Evolutionary biology is thus faced with the challenge to answer why such variation in social structure exists, and what the consequences are on the individual and population level. The main part of this thesis takes on this challenge by investing the dynamics of socially polymorphic ant colonies. The first four chapters investigate the causes and consequences of different social structures, using a combination of field studies, genetic analyses and laboratory experiments. The thesis ends with a theoretical chapter focusing on different social interactions (altruism and spite), and the evolution of harming traits. The main results of the thesis show that social polymorphism has the potential to affect the behaviour and traits of both individuals and colonies. For example, we found that genetic polymorphism may increase the phenotypic variation between individuals in colonies, and that socially polymorphic colonies may show different life history patterns. We also show that colony cohesion may be enhanced even in multiple-queen colonies through patterns of unequal reproduction between queens. However, the thesis also demonstrates that spatial and temporal variation between both populations and environments may affect individual and colony traits, to the degree that results obtained in one place or at one time may not be applicable in other situations. This opens up potential further areas of research to explain these differences.

Differentiation of neural stem cells in fragile X syndrome

Multipotent stem cells can self-renew and give rise to multiple cell types. One type of mammalian multipotent stem cells are neural stem cells (NSC)s, which can generate neurons, astrocytes and oligodendrocytes. NSCs are likely involved in learning and memory, but their exact role in cognitive function in the developing and adult brain is unclear. We have studied properties of NSCs in fragile X syndrome (FXS), which is the most common form of inherited mental retardation. FXS is caused by the lack of functional fragile X mental retardation protein (FMRP). FMRP is involved in the regulation of postsynaptic protein synthesis in a group I metabotropic glutamate receptor 5 (mGluR5)-dependent manner. In the absence of functional FMRP, the formation of functional synapses is impaired in the forebrain which results in alterations in synaptic plasticity. In our studies, we found that FMRP-deficient NSCs generated more neurons and less glia than control NSCs. The newborn neurons derived from FMRP-deficient NSCs showed an abnormally immature morphology. Furthermore, FMRP-deficient NSCs exhibited aberrant oscillatory Ca2+ responses to glutamate, which were specifically abolished by an antagonist of the mGluR5 receptor. The data suggested alterations in glutamatergic differentiation of FMRP-deficient NSCs and were further supported by an accumulation of cells committed to glutamatergic lineage in the subventricular zone of the embryonic Fmr1-knockout (Fmr1-KO) neocortex. Postnatally, the aberrant cells likely contributed to abnormal formation of the neocortex. The findings suggested a defect in the differentiation of distinct glutamatergic mGluR5 responsive cells in the absence of functional FMRP. Furthermore, we found that in the early postnatal Fmr1-KO mouse brain, the expression of mRNA for regulator of G-protein signalling-4 (RGS4) was decreased which was in line with disturbed G-protein signalling in NSCs lacking FMRP. Brain derived neurotrophic factor (BDNF) promotes neuronal differentiation of NSCs as the absence of FMRP was shown to do. This led us to study the effect of impaired BDNF/TrkB receptor signaling on NSCs by overexpression of TrkB.T1 receptor isoform. We showed that changes in the relative expression levels of the full-length and truncated TrkB isoforms influenced the replication capacity of NSCs. After the differentiation, the overexpression of TrkB.T1 increased neuronal turnover. To summarize, FMRP and TrkB signaling are involved in normal differentiation of NSCs in the developing brain. Since NSCs might have potential for therapeutic interventions in a variety of neurological disorders, our findings may be useful in the design of pharmacological interventions in neurological disorders of learning and memory.

Functional characterization of MutL homologue mismatch repair proteins and their variants

Mismatch repair (MMR) mechanisms repair DNA damage occurring during replication and recombination. To date, five human MMR genes, MSH2, MHS6, MSH3, MLH1 and PMS2 are known to be involved in the MMR function. Human MMR proteins form 3 different heterodimers: MutSα (MSH2 and MSH6) and MutSβ (MSH2 and MSH3), which are needed for mismatch recognition and binding, and MutLα (MLH1 and PMS2), which is needed for mediating interactions between MutS homologues and other MMR proteins. The other two MutL homologues, MLH3 and PMS1, have been shown to heterodimerize with MLH1. However, the heterodimers MutLγ (MLH1and MLH3) and MutLβ (MLH1 and PMS1) are able to correct mismatches only with low or no efficiency, respectively. A deficient MMR mechanism is associated with the hereditary colorectal cancer syndrome called hereditary nonpolyposis colorectal cancer (HNPCC) or Lynch syndrome. HNPCC is the most common hereditary colorectal cancer syndrome and accounts for 2-5% of all colorectal cancer cases. HNPCC-associated mutations have been found in 5 MMR genes: MLH1, MSH2, MSH6, PMS2 and MLH3. Most of the mutations have been found in MLH1 and MSH2 (~90%) and are associated with typical HNPCC, while mutations in MSH6, PMS2 and MLH3 are mainly linked to putative HNPCC families lacking the characteristics of the syndrome. More data of MLH3 mutations are needed to assess the significance of its mutations in HNPCC. In this study, were functionally characterized 51 nontruncating mutations in the MLH1, MLH3 and MSH2 genes to address their pathogenic significance and mechanism of pathogenicity. Of the 36 MLH1 mutations, 22 were deficient in more than one assay, 2 variants were impaired only in one assay, and 12 variants behaved like the wild type protein, whereas all seven MLH3 mutants functioned like the wild type protein in the assays. To further clarify the role and relevance of MLH3 in MMR, we analyzed the subcellular localization of the native MutL homologue proteins. Our immunofluorescence analyses indicated that when all the three MutL homologues are natively expressed in human cells, endogenous MLH1 and PMS2 localize in the nucleus, whereas MLH3 stays in the cytoplasm. The coexpression of MLH3 with MLH1 results in its partial nuclear localization. Only one MSH2 mutation was pathogenic in the in vitro MMR assay. Our study on MLH1 mutations could clearly distinguish nontruncating alterations with severe functional defects from those not or only slightly impaired in protein function. However, our study on MLH3 mutations suggest that MLH3 mutations per se are not sufficient to trigger MMR deficiency and the continuous nuclear localization of MLH1 and PMS2 suggest that MutLα has a major activity in MMR in vivo. Together with our functional assays, this confirms that MutLγ is a less efficient MMR complex than MutLα.

Carabid beetles (Coleoptera, Carabidae) as indicators of environmental change in Ranomafana National Park, Madagascar

Growing human populations and increasing exploitation of natural resources threaten nature all over the world. Tropical countries are especially vulnerable to human impact because of the high number of species, most of these endemic and still unknown. Madagascar is one of the centers of high biodiversity and renowned for its unique species. However, during the last centuries many endemic species have gone extinct and more are endangered. Because of high natural values, Madagascar is one of the global conservation priorities. The establishment of Ranomafana National Park (RNP) was intended to preserve the unique nature of Madagascar. Containing several endemic and threatened species, Ranomafana has been selected as one of UNESCO’s World Natural Heritage sites. However, due to strong human pressures the region immediately surroundings the protected area has severely degraded. Aims of this thesis were to inventory carabid fauna in RNP and evaluate their use as indicators of the environmental change. Carabid beetles were collected from protected area (secondary and primary forests) and from its degraded surrounding area. Collecting was mostly conducted by hand during years 2000-2005. Species compositions between the protected area and its surroundings were compared, and species habitat preferences and seasonal variations were studied. In total, 4498 individuals representing 127 carabid species (of which 38 are new species) were collected. Species compositions within and outside of the protected area were markedly different. Most of the species preferred forest as their primary habitat and were mainly collected from trees and bushes. Their value as indicators is based on their different habitat requirements and sensitivity to environmental variables. Some of the species were found only in the protected forest, some occupied also the degraded forests and some preferred open areas. Carabid fauna is very species rich in Ranomafana and there are still many species to be found. Most of the species are arboreal and probably cannot survive in the deforested areas outside the park. This is very likely also the case for other species. Establishment and continued protection of RNP is probably the only way to conserve this globally important area. However, new occupations and land use methods are urgently needed by the local people for improving their own lives while maintaining the forest intact.