This thesis investigates the ecological and evolutionary implications of a recent secondary contact between two closely related bird species: collared (Ficedula albicollis) and pied (F. hypoleuca) flycatchers. Collared flycatchers started to colonize the Swedish island of Öland, where pied flycatchers were already present, in the late 1950s-early1960s. My major aims were to investigate which factors are acting against versus for long-term coexistence between the two species. Specifically, I investigated the relative importance of allopatric divergence, interspecific competition, hybridization and learning in promoting or inhibiting coexistence. The combined effects of interspecific competition and hybridization drives pied flycatchers towards local extinction in their preferred deciduous habitat. However, my results also show that pied flycatchers are better able to tolerate harsh environmental conditions. This trade-off between competitive ability and resilience in the face of harsh conditions facilitates a regional coexistence between the species. Coexistence is furthermore favoured by competition-mediated divergence in breeding habitat choice, timing of breeding and male breeding plumage colouration. Due to interspecific competition, male pied flycatchers are forced to breed in a more mixed forest type with a later peak in food abundance, which is accompanied by a divergence in breeding time between the two species. In areas shared with collared flycatchers, male pied flycatchers with brown plumage coloration, most divergent from that of collared flycatchers, are favoured by selection. In addition to facilitating coexistence, the observed shift in habitat occupancy increases reproductive isolation between the two species. By using cross-fostering experiments I demonstrate that natal habitat imprinting has the potential to additionally speed up habitat segregation. Finally I show that hybrid nestlings express an intermediate response to harsh environments, indicating that another aspect of ecological-based selection may be important in reproductive isolation between the species. In summary, my results show that adaptations during historic allopatry are important both in facilitating coexistence as well as in providing a foundation for further ecological divergence at secondary contact. This is of relevance today as many species are shifting their distributions in response to habitat disturbance and global warming.

The methods presented in this study are aimed at the identification of subpopulations (clones) of genetically similar cells within tissue samples through measurement of loci-specific Fluorescence in-situ hybridization (FISH) spot signals for each nucleus and analyzing cell spatial distributions by way of Voronoi tessellation and Delaunay triangulation to robustly define cell neighbourhoods. The motivation for the system is to examine lung cancer patient for subpopulations of Non-Small Cell Lung Cancer (NSCLC) cells with biologically meaningful gene copy-number profiles: patterns of genetic alterations statistically associated with resistance to cis-platinum/vinorelbine doublet chemotherapy treatment. Current technologies for gene-copy number profiling rely on large amount of cellular material, which is not always available and suffers from limited sensitivity to only the most dominant clone in often heterogeneous samples. Thus, through the use of FISH, the detection of gene copy-numbers is possible in unprocessed tissues, allowing identification of specific tumour clones with biologically relevant patterns of genetic aberrations. The tissue-wide characterization of multiplexed loci-specific FISH signals, described herein, is achieved through a fully automated, multicolour fluorescence imaging microscope and object segmentation algorithms to identify cell nuclei and FISH spots within. Related tumour clones are identified through analysis of robustly defined cell neighbourhoods and cell-to-cell connections for regions of cells with homogenous and highly interconnected FISH spot signal characteristics. This study presents experiments which demonstrate the system’s ability to accurately quantify FISH spot signals in various tumour tissues and in up to 5 colours simultaneously or more through multiple rounds of FISH staining. Furthermore, the system’s FISH-based cell classification performance is evaluated at a sensitivity of 84% and specificity 81% and clonal identification algorithm results are determined to be comparable to clone delineation by a human-observer. Additionally, guidelines and procedures to perform anticipated, routine analysis experiments are established.

Copies of author's previously published articles inserted. / Includes corrigendum. / Bibliography: leaves 97-106. / xiii, 106, [69] leaves, [31] leaves of plates : ill. (some col.), maps ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / This thesis reports an investigation of the etiology of ScSMD (sugarcane striate mosaic disease) by biological and microscopic studies and by nucleic acid studies. Studies of ScSMD affected sugarcane are followed by the detection, isolation, cloning and partial sequencing of a disease specific dsRNA, and the tentative classification of the putative viral agent from the partial sequence. This study also describes the ScSMD associated virion and an improved protocol for the purification of dsRNA. / Thesis (Ph.D.)--University of Adelaide, Dept. of Crop Protection, 1997

Doctor of Philosophy / Benign tumours grow by expanding and displacing the surrounding tissues, while malignant tumours replace and destroy the surrounding tissues by invasion. Although there is extensive literature on mechanisms of tumour invasion and metastasis, with an emphasis on angiogenesis, adhesion, degradation of the extracellular matrix and migration, an important question not clearly addressed by the literature, but nonetheless approached in this thesis, is that of the fate of normal cells during tissue replacement by migrating invasive malignant cells. Earlier work in the laboratory where this PhD candidature was carried out, investigated the effect of osteosarcoma cells on endothelium. In contrast to the expected angiogenic effect of malignant cells for endothelium, it was found that the human osteosarcoma cell line (SAOS-2) induced apoptosis in human umbilical vein endothelial cells (HUVEC) in contact dependent manner (McEwen et al., 2003). It was suggested that apoptosis of endothelium by malignant tumour cells may facilitate tumour invasion and metastasis (McEwen et al., 2003), and one of the aims of the current study was to extend these findings to include human gingival fibroblasts (HGF) and human umbilical artery smooth muscle cells (HUASMC). The major finding of this thesis was that SAOS-2 induced a reduction in the apparent cell culture density of HGF and HUASMC in a contact-dependent manner. The SW480 colorectal carcinoma cell line did not have any clear effect upon the apparent stromal cell culture density of either HGF or HUASMC, suggesting that the effect under investigation was tumour cell line specific. Surprisingly and in contrast to the similar effect reported for endothelium (Chen et al., 2005; McEwen et al., 2003), the effect of SAOS-2 upon HGF and HUASMC was not due to stromal cell apoptosis. Apoptosis was ruled out as a possible mechanism for the reduced apparent culture density under study, by using widely accepted methods which are dependent upon intermucleosomal fragmentation of DNA, the permeability of plasma membranes to dyes in advanced apoptosis and necrosis, phosphatidylserine translocation as well as inhibitor studies blocking both caspase dependent and independent pathways. While apoptosis was not demonstrated, the possibility emerged that reduced apparent stromal cell culture density reflected fusion events rather than the simple removal of cells as had been earlier reported for HUVEC (McEwen et al., 2003). This idea was supported by reduced SAOS-2 circularity in co-culture. Confocal microscopy of cells pre-labelled with fluorescent dyes further supported this idea, with dual-labelling as evidence of cell fusion. Although occasional homotypic fusion of stromal cells was seen, heterotypic fusion of stromal cells with SAOS-2 was much more prevalent. Time lapse microscopy was performed to further characteristic cell fusion in co-cultures, and revealed multiple transient fusions between SAOS-2 and HGF. To work towards determining the biological relevance of the key observation, two stable SAOS-2 GFP clones were generated for future planned studies using human gingival explants and nude mice. Importantly, the clones were similar to native SAOS-2 with regard to alkaline phosphatise expression and reducing apparent stromal cell culture density. Transient fusions between HGF and SAOS-2, may be a mechanism for cooption of stromal cells into the malignant process, facilitating tumour invasion. Additionally, heterocellular fusion of SAOS-2 with stromal cells may facilitate immune evasion, while it seems likely that despite the absence of an identical activity in SW480 cells, other malignant tumour cells may also express similar activity.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2006. / Härtill 5 uppsatser.

Diss. (sammanfattning) Alnarp : Sveriges lantbruksuniv., 2002. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2002. / Härtill 7 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2004. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2004. / Härtill 5 uppsatser.