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  • Elshibli, Sakina (Helsingin yliopisto, 2009)
    Acquiring sufficient information on the genetic variation, genetic differentiation, and the ecological and genetic relationships among individuals and populations are essential for establishing guidelines on conservation and utilization of the genetic resources of a species, and more particularly when biotic and abiotic stresses are considered. The aim of this study was to assess the extent and pattern of genetic variation in date palm (Phoenix dacttylifera L) cultivars; the genetic diversity and structure in its populations occurring over geographical ranges; the variation in economically and botanically important traits of it and the variation in its drought adaptive traits, in conservation and utilization context. In this study, the genetic diversity and relationships among selected cultivars from Sudan and Morocco were assessed using microsatellite markers. Microsatellite markers were also used to investigate the genetic diversity within and among populations collected from different geographic locations in Sudan. In a separate investigation, fruits of cultivars selected from Sudan, involved morphological and chemical characterization, and morphological and DNA polymorphism of the mother trees were also investigated. Morphological and photosynthetic adjustments to water stress were studied in the five most important date palm cultivars in Sudan, namely, Gondaila, Barakawi, Bitamoda, Khateeb and Laggai; and the mechanism enhancing photosynthetic gas exchange in date palm under water stress was also investigated. Results showed a significant (p < 0.001, t-test) differentiation between Sudan and Morocco groups of cultivars. However, the major feature of all tested cultivars was the complete lack of clustering and the absence of cultivars representing specific clones. The results indicated high genetic as well as compositional and morphological diversity among cultivars; while, compositional and morphological traits were found to be characteristic features that strongly differentiate cultivars as well as phenotypes. High genetic diversity was observed also in different populations. Slight but significant (p < 0.01, AMOVA) divergence was observed for soft and dry types; however, the genetic divergence among populations was relatively weak. The results showed a complex genetic relationships between some of the tested populations especially when isolation by distance was considered. The results of the study also revealed that date palm cultivars and phenotypes possess specific direct or interaction effects due to water availability on a range of morphological and physiological traits. Soft and dry phenotypes responded differently to different levels of water stress, while the dry phenotype was more sensitive and conservative. The results indicated that date palm has high fixation capacity to photosynthetic CO2 supply with interaction effect to water availability, which can be considered as advantageous when coping with stresses that may arise with climate change. In conclusion, although a large amount of diversity exists among date palm germplasm, the findings in this study show that the role of biological nature of the tree, isolation by distance and environmental effects on structuring date palm genome was highly influenced by human impacts. Identity of date palm cultivars as developed and manipulated by date palm growers, in the absence of scientific breeding programmes, may continue to mainly depend on tree morphology and fruit characters. The pattern of genetic differentiation may cover specific morphological and physiological traits that contribute to adaptive mechanisms in each phenotype. These traits can be considered for further studies related to drought adaptation in date palm.
  • Pohjanmies, Tähti (2014)
    Genetic variation within a population is shaped by the life history traits of the species and the properties of the surrounding ecosystem. It is an important factor in the preservation of populations. According to the emerging field of community genetics, genetic variation within a population of one species may also influence the dynamics and diversity of associated species, extending the conservational relevance of intraspecific genetic diversity. Finnish populations of pedunculate oak (Quercus robur) offer an interesting study system for population genetics. Q. robur grows in south-western Finland at the northern limit of its natural range. Here, its distribution has been shaped by long-term climatic and geological changes as well as by human disturbance, and the current populations are small and strongly fragmented. As Q. robur supports a high diversity of associated species, it is considered to have great ecological and conservational importance. In this thesis, I studied the amount and distribution of genetic diversity within and among three Q. robur populations in south-western Finland using population genetic parameters. I also described the spatial and temporal sub-population structure of one population, on the island of Wattkast. The genetic data was based on 15 nuclear microsatellite loci. Additionally, I examined the effect of the genetic diversity and genotypic identity of the oaks within Wattkast on associated herbivore communities. In the analysis, I used observational data from two years. As predicted for widespread, long-lived tree species, the microsatellite loci showed high levels of diversity within the populations, but also significant differentiation among them. This may be due to fragmentation and to the marginality of the populations. Within the population on Wattkast, I observed patterns of spatial and temporal sub-population differentiation. The characteristics of the site, including the ongoing shift to less extensive land use, suggest that the population is in genetic disequilibrium. As both the genetic distance and the community dissimilarity between pairs of trees increased with increasing geographic distance, I could not conclude the genotypic identity of the host trees to have an effect on the herbivore community structure. However, higher heterozygosity was associated with higher richness and abundance of species. This result supports the notion that intraspecific genetic variation may increase associated species richness. Based on the results of my study, both the life history traits of the species and the historic habitat changes may be observed in the genetic structure of Q. robur populations in Finland. The results also suggest that preservation of genetic variation within the remaining stands may be a factor not only in the preservation of these populations, but also in the conservation of associated species diversity.
  • Halinen, Katrianna (Helsingfors universitet, 2008)
    Cyanobacteria (blue-green algae) form blooms in the Baltic Sea during the warmest summer months. According to paleolimnological data, cyanobacteria have long history in the Baltic Sea, going back at least 7000 years. However, the intensity as well as the expanse of cyanobacterial blooms has increased during recent decades. Blooms attract regular attention in the media because of their visibility and the potential health risk they pose to humans and animals. The Gulf of Finland is the most eutrophied area of the Baltic Sea, and cyanobacterial blooms are widely believed to be the result of intense anthropogenic nutrient loading. Cyanobacterial blooms are formed mainly by species of three genera in the Baltic Sea, Nodularia, Anabaena and Aphanizomenon. The focus of present-day research on Baltic Sea cyanobacteria has been on Nodularia and Aphanizomenon, while the genus Anabaena has been neglected. Anabaena is often considered to play a minor role in cyanobacterial blooms. However, Anabaena can form a significant part of the blooms, especially in the northern part of the Baltic Sea. Cyanobacterial blooms in the Baltic Sea are invariably toxic due to the production of hepatotoxic nodularin by Nodularia spumigena. According to systematic studies, Aphanizomenon flos-aquae was not found to produce hepatotoxins in the Baltic Sea. However, it has been speculated that Baltic Sea Anabaena spp. could produce microcystins. The genetic structure of the Anabaena populations in the Baltic Sea has not been systematically explored. The aim of this present study was to increase our understanding of the Anabaena - a component of the Baltic Sea phytoplankton. Altogether, 49 planktonic Anabaena strains were isolated from the Gulf of Finland, five of which were microcystin-producing. This study provided unequivocal evidence that Baltic Sea Anabaena is able to produce microcystins. Each microcystin-producing Anabaena strain produced two to four dominant microcystin variants, including the highly toxic microcystin-LR. In this study, a culture-independent method was designed to detect putative microcystin and nodularin producers. By means of this DGGE method, microcystin-producing Anabaena populations were detected in cyanobacterial bloom samples from the summers of 2003 and 2004. Microcystin-producing Anabaena populations were detected throughout the Gulf of Finland. This excluded the possibility that the presence of microcystin-producing Anabaena was a chance phenomenon. Results suggest that salinity may limit the distribution of the microcystin-producing Anabaena although further studies are needed to confirm the interdependence of salinity and microcystin production. Microcystin-producing Anabaena populations were found to be highly diverse on analyses of the 16S rRNA, rbcL, rpoC1, and mcyE gene sequences. In previous studies, freshwater microcystin-producing Anabaena strains were grouped together in phylogenetic analyses. All microcystin-producing Baltic Sea Anabaena strains belonged to this hepatotoxic cluster, with the exception of a single strain. Both planktonic and benthic Anabaena populations were genetically heterogeneous and closely related to freshwater Anabaena strains. However, genetic diversity in benthic Anabaena strains was higher than in planktonic strains. In phylogenetic analyses, novel Anabaena lineages, possibly specific to the Baltic Sea, were identified. This suggests ecotypic diversification within Anabaena populations. We found two planktonic Anabaena strains which carried the entire mcy gene cluster, but were nonetheless incapable of producing microcystins. Natural genetic inactivation of the mcy gene cluster was identified in Anabaena strain BIR259. This strain carried insertions which most likely caused the inactivation of the mcy genes. The insertions documented here were surprisingly common in the Baltic Sea bloom samples and they were present in samples from both studied summers, 2003 and 2004. However, these insertions were not identified in freshwater strains or in field samples from freshwater lakes. The aim of this study was to establish a strain collection of Baltic Sea Anabaena and to shed light on the phylogeny, microcystin-production, and genetic diversity of the Baltic Sea Anabaena populations. In addition to strain isolation, these research goals were approached by in situ molecular methods. Systematic toxin screening showed that Anabaena is able to produce microcystins, and this should be taken into account in future toxin monitoring programmes.
  • Palo, Jukka (Helsingin yliopisto, 2003)
  • Säisä, Marjatta (Helsingin yliopisto, 2009)
    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.
  • Rehnström, Karola (Helsingin yliopisto, 2009)
    Positional cloning has enabled hypothesis-free, genome-wide scans for genetic factors contributing to disorders or traits. Traditionally linkage analysis has been used to identify regions of interest, followed by meticulous fine mapping and candidate gene screening using association methods and finally sequencing of regions of interest. More recently, genome-wide association analysis has enabled a more direct approach to identify specific genetic variants explaining a part of the variance of the phenotype of interest. Autism spectrum disorders (ASDs) are a group of childhood onset neuropsychiatric disorders with shared core symptoms but varying severity. Although a strong genetic component has been established in ASDs, genetic susceptibility factors have largely eluded characterization. Here, we have utilized modern molecular genetic methods combined with the advantages provided by the special population structure in Finland to identify genetic risk factors for ASDs. The results of this study show that numerous genetic risk factors exist for ASDs even within a population isolate. Stratification based on clinical phenotype resulted in encouraging results, as previously identified linkage to 3p14-p24 was replicated in an independent family set of families with Asperger syndrome, but no other ASDs. Fine-mapping of the previously identified linkage peak for ASDs at 3q25-q27 revealed association between autism and a subunit of the 5-hydroxytryptamine receptor 3C (HTR3C). We also used dense, genome-wide single nucleotide polymorphism (SNP) data to characterize the population structure of Finns. We observed significant population substructure which correlates with the known history of multiple consecutive bottle-necks experienced by the Finnish population. We used this information to ascertain a genetically homogenous subset of autism families to identify possible rare, enriched risk variants using genome-wide SNP data. No rare enriched genetic risk factors were identified in this dataset, although a subset of families could be genealogically linked to form two extended pedigrees. The lack of founder mutations in this isolated population suggests that the majority of genetic risk factors are rare, de novo mutations unique to individual nuclear families. The results of this study are consistent with others in the field. The underlying genetic architecture for this group of disorders appears highly heterogeneous, with common variants accounting for only a subset of genetic risk. The majority of identified risk factors have turned out to be exceedingly rare, and only explain a subset of the genetic risk in the general population in spite of their high penetrance within individual families. The results of this study, together with other results obtained in this field, indicate that family specific linkage, homozygosity mapping and resequencing efforts are needed to identify these rare genetic risk factors.
  • Jiménez Caldera, Oswalt Rafael (Helsingin yliopisto, 2014)
    The common bean (Phaseolus vulgaris L.) is an important component of food security programs aiming to provide better human nutrition in developing countries. However, low yields, climate change affecting production and variable local demands of specific types of cultivars suggest that we should consider the utilization of local bean genetic resources with high market acceptance in local breeding programs. Those efforts would complement regional breeding agendas. In this study, the genetic diversity of a collection of landraces was found to be considerably higher (mean 8.9 alleles per microsatellite locus) than previously reported, with a population structure of three main clusters grouped according to seed weights. Of these landraces, two promising divergent sources of genetic variation, accessions PV0006 and PV0023, were chosen for single crosses. The hybridity of F1 generation was tested using polymorphic microsatellite markers. Computer simulations demonstrated that the selection of F1 individuals possessing the highest degree of allele recombination following the pedigree method, instead of using the whole set of F1 individuals as is usually done, could improve the selection gains for yield. Between 128 and 1024 pure lines could be obtained after a reasonable number of generations. Subsequently, 420 F2 plants originating from 15 marker-selected F1 plants were established in three augmented blocks together with both parents and check cultivar INTA ROJO . Variables PP, SP, SW and YP were measured for each plant. PP and YP were considered the most appropriate traits for selection based on ANOVAs, high heritability values, and genetic gains obtained for both traits. After conducting plant selection using mixed model analyses, 81 and 74 F2 plants (with 61 plants in common for both groups) were selected based on their superior yield potential compared with both parents and check cultivar. Resistance to BCMV, BGYMV, ANT and rust was confirmed in the segregating population, and their higher potential over 40 bean landraces was validated by computer simulations constructed using genotype frequencies. The level of resistance was unexpectedly found to be similar to that of current cultivars in use. However, further experiments to confirm resistance should be conducted. Moreover, resistance genes for BCMV (bc-3 and bc-12) absent in our segregating plants were detected in bean landraces PV0015, PV0016, PV0017, PV0026, PV0031, PA0001, PA0002, and PA0003. It would be beneficial to pyramid broader resistance to this seed-borne virus in our populations by means of new crosses. All these findings emphasize the feasibility of utilizing local landraces for genetic improvement using efficient statistical methods aided by molecular markers. Additionally, molecular markers were efficiently used to test the genetic purity of cultivar INTA ROJO . The detected significant changes in genotype frequencies in four seed categories were probably caused by inadequate roguing procedures during seed production. Similarly, molecular markers were able to discriminate off-type seeds and plants from INTA ROJO , opening the possibility to complement current phenotypic methods employed in the genetic quality testing of seed lots.
  • Onkamo, Päivi (Helsingin yliopisto, 2002)
  • Manninen, Outi (Helsingin yliopisto, 2000)
  • Dahlsten, Elias (Helsingin yliopisto, 2013)
    Clostridium botulinum presents a significant hazard to the food processing industry. However, the cellular mechanisms and factors that contribute to their regulation utilized by this feared foodborne pathogen in response and adaptation to food processing and storage-induced stress are poorly characterized. Another major aspect of C. botulinum presenting serious implications on food safety is its capability to produce heat-resistant endospores. Nevertheless, the sporulation cascade of C. botulinum has not been characterized. This study sought to investigate the effects of temperature downshift on the global gene expression pattern of Group I C. botulinum type strain ATCC 3502, and further characterize the roles of regulatory mechanisms identified as cold tolerance-related. Furthermore, the role of a major regulatory element, the alternative sigma factor SigK, in the sporulation cascade of C. botulinum was determined. Additionally, its putative function in stress tolerance was investigated. Transcriptomic analysis of the foodborne pathogen C. botulinum ATCC 3502 upon temperature downshift revealed the induction of several mechanisms previously identified as cold-related in other bacteria, thus suggesting that also C. botulinum utilizes these mechanisms in cold tolerance. Mechanisms with hitherto uncharacterized functions in cold tolerance were also found. The results suggested that secondary oxidative stress was present as a component of cold stress. Additionally, two previously uncharacterized putative DNA-binding regulatory proteins CBO0477 and CBO0558A were shown to play a role in cold tolerance of C. botulinum ATCC 3502. The two-component system (TCS) CBO0366/CBO0365 was shown to be important in the cold tolerance of C. botulinum ATCC 3502. Expression of this TCS was induced upon temperature downshift, but not under optimal temperature and growth conditions. Disruption of either of the TCS components resulted in deteriorated cold tolerance, whereas over-expression of cbo0366 in a wild-type strain resulted in an increase of growth rate at low temperature. Inactivation of the TCS response regulator-encoding cbo0365 markedly altered the transcriptome of C. botulinum ATCC 3502. Totals of 150 and 141 chromosomal coding sequences (CDS) were significantly differently expressed in the cbo0365 mutant at either 37 °C or 15 °C, respectively. There was an overlap of 141 common CDSs between the two temperatures. The genes differentially expressed included ones related to acetone-butanol-ethanol (ABE) fermantion, arsenic resistance, phosphate uptake and flagellar rotation. The involvement of CBO0365-regulated metabolic pathways in cold tolerance was demonstrated by the deteriorated cold tolerance of mutants of the respective pathways. Cold-sensitive phenotypes were observed for mutants of the acetone-butanol-ethanol fermentation pathway components bcd, crt, bdh and ctfA, the arsenic detoxifying machinery components arsC and arsR, and the phosphate uptake mechanism component phoT. Electrophoretic mobility shift assays confirmed transcriptional activation or repression as a means for CBO0365 in regulating itself, and the crt, ars, and pho operons. A dual role for the alternative sigma factor SigK in sporulation and in stress tolerance of C. botulinum ATCC 3502 was demonstrated. Disruption of SigK halted sporulation at an early stage but the phenotype was restored by in trans complementation of the mutation. The expression of sigK was induced upon exposure to low temperature and high salinity, but not upon a downshift in pH. A deteriorated tolerance to low temperature and to high salinity was observed for the sigK mutant strains.
  • Woldesenbet, Sebsebe Lemeta (Helsingin yliopisto, 2009)
    The von Hippel-lindau (VHL) disease is a dominantly inherited neoplastic disorder which predisposes patients to multiple tumours including capillary haemangioblastomas (CHBs), pheochromocytomas (PCCs), renal cell carcinomas (RCCs). CHBs are the most common manifestations of VHL disease, occurring sporadically or as a manifestation of VHL disease. Inactivation of the VHL gene at 3p25-26 is believed to cause both familial and sporadic VHL-associated tumours and germ-line mutation of the VHL gene have been detected in 100% of the CHBs studied. However, a limited number of sporadic CHBs, PCCs display VHL inactivation. Other molecular alterations involved in tumourigenesis of sporadic CHBs, PCCs remain largely unknown. The purpose of the present work was to search for genetic alterations, or other mechanisms of inactivation, in addition to the VHL gene, that may be important in the development of VHL-associated tumours. Though less satisfactory than cure, prevention and early detection are the most promising and feasible means reducing cancer morbidity and mortality. This work is based on the view that increasing knowledge about the molecular events underlying tumour development will eventually aid in early detection and lead to improved treatment. We evaluated a large set of VHL-associated patients, searched for a clinical and radiologic signs of the disease. We succesfully performed a germ-line mutation analysis and characterised three patient groups, VHL, suspect VHL and sporadic, a germ-line mutation analysis revealed a 50% mutation rate only in the VHL groups, no sporadic or suspect cases displayed any mutation. We also utilized comparative genomic hybridization (CGH) to screen for DNA copy number changes in both sporadic and VHL-associated CHB. Our analysis revealed (27%) DNA copy number losses. The most common finding was loss of chromosomal arm 6q, seen in (23%) cases, No differences were noted between VHL-associated and sporadic tumours. Furthermore a loss of heterozygosity (LOH) study on chromosome 3p and 6q was done with the purpose to determine allele losses not observable by CGH, and to uncover the location of putative tumour suppressor genes important in CHB and PCC tumourigenesis. We identified loss of chromosome 6q and a minimal deleted area at 6q23-24 in CHBs. We also showed LOH at 6q23-24 in PCCs and identified the ZAC1 (6q24-25) as a candidate gene, ZAC1 is a maternally imprinted tumour suppressor gene with anti proliferative properties. To study further the role of ZAC inactivation in CHBs, we investigated LOH, promoter hypermethylation and expression status of the ZAC1 gene in mainly sporadic CHBs. Our LOH analysis revealed that the majority of the tumours with allele loss. The gene promoter methylation analysis similarly detected predominance of the methylated ZAC sequence in almost all tumours. Immunohistochemistry exhibited a strongly reduced expression of ZAC in stromal cells of all CHBs studied. Our current results indicate that the absence of the unmethylated, ZAC1 promoter sequence was highly concurrent with LOH for the ZAC1 region or 6q loss. This observation together with lack of ZAC expression, points to preferential loss of the non imprinted, expressed ZAC allele in CHB, in summary, our series of studies reveal a new chromosomal region 6q, emphasizes the importance of ZAC1 gene in the development of CHB and PCC, particularly in non-VHL associated cases.
  • Kilpinen, Helena (Helsingin yliopisto, 2010)
    Autism is a childhood-onset developmental disorder characterized by deficits in reciprocal social interaction, verbal and non-verbal communication, and dependence on routines and rituals. It belongs to a spectrum of disorders (autism spectrum disorders, ASDs) which share core symptoms but show considerable variation in severity. The whole spectrum affects 0.6-0.7% of children worldwide, inducing a substantial public health burden and causing suffering to the affected families. Despite having a very high heritability, ASDs have shown exceptional genetic heterogeneity, which has complicated the identification of risk variants and left the etiology largely unknown. However, recent studies suggest that rare, family-specific factors contribute significantly to the genetic basis of ASDs. In this study, we investigated the role of DISC1 (Disrupted-in-schizophrenia-1) in ASDs, and identified association with markers and haplotypes previously associated with psychiatric phenotypes. We identified four polymorphic micro-RNA target sites in the 3 UTR of DISC1, and showed that hsa-miR-559 regulates DISC1 expression in vitro in an allele-specific manner. We also analyzed an extended autism pedigree with genealogical roots in Central Finland reaching back to the 17th century. To take advantage of the beneficial characteristics of population isolates to gene mapping and reduced genetic heterogeneity observed in distantly related individuals, we performed a microsatellite-based genome-wide screen for linkage and linkage disequilibrium in this pedigree. We identified a putative autism susceptibility locus on chromosome 19p13.3 and obtained further support for previously reported loci at 1q23 and 15q11-q13. To follow-up these findings, we extended our study sample from the same sub-isolate and initiated a genome-wide analysis of homozygosity and allelic sharing using high-density SNP markers. We identified a small number of haplotypes shared by different subsets of the genealogically connected cases, along with convergent biological pathways from SNP and gene expression data, which highlighted axon guidance molecules in the pathogenesis of ASDs. In conclusion, the results obtained in this thesis show that multiple distinct genetic variants are responsible for the ASD phenotype even within single pedigrees from an isolated population. We suggest that targeted resequencing of the shared haplotypes, linkage regions, and other susceptibility loci is essential to identify the causal variants. We also report a possible micro-RNA mediated regulatory mechanism, which might partially explain the wide-range neurobiological effects of the DISC1 gene.
  • Suonsyrjä, Timo (Helsingin yliopisto, 2011)
    Hypertension is one of the major risk factors for cardiovascular morbidity. The advantages of antihypertensive therapy have been clearly demonstrated, but only about 30% of hypertensive patients have their blood pressure (BP) controlled by such treatment. One of the reasons for this poor BP control may lie in the difficulty in predicting BP response to antihypertensive treatment. The average BP reduction achieved is similar for each drug in the main classes of antihypertensive agents, but there is a marked individual variation in BP responses to any given drug. The purpose of the present study was to examine BP response to four different antihypertensive monotherapies with regard to demographic characteristics, laboratory test results and common genetic polymorphisms. The subjects of the present study are participants in the pharmacogenetic GENRES Study. A total of 208 subjects completed the whole study protocol including four drug treatment periods of four weeks, separated by four-week placebo periods. The study drugs were amlodipine, bisoprolol, hydrochlorothiazide and losartan. Both office (OBP) and 24-hour ambulatory blood pressure (ABP) measurements were carried out. BP response to study drugs were related to basic clinical characteristics, pretreatment laboratory test results and common polymorphisms in genes coding for components of the renin-angiotensin system, alpha-adducin (ADD1), beta1-adrenergic receptor (ADRB1) and beta2-adrenergic receptor (ADRB2). Age was positively correlated with BP responses to amlodipine and with OBP and systolic ABP responses to hydrochlorothiazide, while body mass index was negatively correlated with ABP responses to amlodipine. Of the laboratory test results, plasma renin activity (PRA) correlated positively with BP responses to losartan, with ABP responses to bisoprolol, and negatively with ABP responses to hydrochlorothiazide. Uniquely to this study, it was found that serum total calcium level was negatively correlated with BP responses to amlodipine, whilst serum total cholesterol level was negatively correlated with ABP responses to amlodipine. There were no significant associations of angiotensin II type I receptor 1166A/C, angiotensin converting enzyme I/D, angiotensinogen Met235Thr, ADD1 Gly460Trp, ADRB1 Ser49Gly and Gly389Arg and ADRB2 Arg16Gly and Gln27Glu polymorphisms with BP responses to the study drugs. In conclusion, this study confirmed the relationship between pretreatment PRA levels and response to three classes of antihypertensive drugs. This study is the first to note a significant inverse relation between serum calcium level and responsiveness to a calcium channel blocker. However, this study could not replicate the observations that common polymorphisms in angiotensin II type I receptor, angiotensin converting enzyme, angiotensinogen, ADD1, ADRB1, or ADRB2 genes can predict BP response to antihypertensive drugs.
  • Broberg, Martin (Helsingin yliopisto, 2015)
    The interactions between phytopathogenic bacteria and their host plants can be characterized as an intricate web of signals and appropriate responses. Phytopathogenic soft rot bacteria occur globally, causing disease in Solanum tuberosum (potato) and other tubular staple foods in both the field and storage. One widely studied soft rot bacterium is Pectobacterium wasabiae, which has been identified in Eutrema wasabi (wasabi) plants in Japan and in potatoes in Finland. Generally, the interactions between this type of bacterium and host plants are characterized by maceration of plant tissue, due to the actions of secreted plant cell wall degrading enzymes (PCWDE), and the induction of phytohormone dependent defenses in the plants. The maceration of plant tissue involves the release of pectic oligogalacturonides (OGs) from plant cell walls. OGs have been identified as important signaling compounds, inducing the expression of a variety of defense-related genes. As the bacterial infection advances, the bacteria coordinate the production of virulence factors by utilizing regulatory proteins that modulate the transcriptome. Transcriptomic analyses have been used extensively in past studies to identify regulatory networks and signaling pathways, and these studies have provided insights into the processes underlying plant-pathogen interactions. The novel scientific results of this dissertation are derived from a combination of transcriptomic, genomic, genetic, and phenotypic analyses. This study analyzed various aspects of plant-pathogen interactions. The central bacterial model used was P. wasabiae, and the model plant of interest was Arabidopsis thaliana. This study characterized the genome of P. wasabiae via sequencing and bioinformatics analysis. Various virulence associated genes and operons, such as two distinct type 6 secretion systems, were identified and annotated. The bacterium was found to in fact be more related to P. wasabiae than Pectobacterium carotovorum, which the strain originally had been named after. Furthermore, a combination of functional genetics and transcriptomic methods, such as reverse transcription quantitative PCR (RT-qPCR) and microarrays, were used to determine the regulons controlled by the proteins ExpA and RsmA in P. wasabiae. These two proteins have been identified as important for the virulence of several γ-proteobacterial pathogens. This study analyzed the regulons via the use of three mutants: expA, rsmA, and an expA rsmA double mutant (DM). Overlapping and independently regulated targets were identified between ExpA and RsmA. Phenotypic assays for motility, growth, PCWDE activity, and virulence confirmed the transcriptomic data for the mutant strains. Novel findings included reduction of swimming motility in agar medium for P. wasabiae expA and rsmA mutants. In addition, the DM exhibited enhanced virulence and fitness in planta compared to either single mutant. Via analysis of transcriptomic data, a subset of genes was identified as affected in expression by an expA mutation independently of the presence of rsmA. The relatively unexplored role of short OGs (with a degree of polymerization (DP) less than 10) in damage-associated molecular pattern (DAMP) signaling in A. thaliana was characterized in this study. Comparative gene expression profiling based on RNA sequencing and RT-qPCR was performed on RNA harvested from plants treated with short OGs or with a mock suspension. Phenotypic assays confirmed the gene expression data. In a meta-data analysis, the resulting RNA sequencing and RT-qPCR data were compared with gene expression data from previous studies, in which long OGs (DP more than 10) were used to treat plants. This work demonstrated that short and long OGs induce genes and genesets associated with pathogen defense and phytohormone signaling, whereas reducing plant growth and development. The transcriptomic data of this study suggests that plant treatment with a mixture of short or long OGs yields a more pronounced and varied modulation of global gene expression, compared to treatment with only trimeric OGs. The regulation of the virulence of P. wasabiae, and the DAMP signaling triggered by plant cell wall damage in A. thaliana, are elements of the interactions between the plant and pathogen. The studies presented in this dissertation provide novel information about these two biological processes and highlights their connection.
  • Jiménez Caldera, Oswalt Rafael (2009)
    Common bean (Phaseolus vulgaris L.) is one of the most important crops in Nicaraguan agriculture. Bean production is carried out on small scale farms, where farmers generally lack key inputs. Seeds have been identified as the most critical input in bean production. For this reason, national seed program will be a priority during next ten years. Among the four main seed quality components, genetic component has been the least studied. The occurrence of offtype seeds and plants in the cultivar ‘INTA ROJO’ during seed production has hindered the seed certification process and questioned the genetic quality of the cultivars produced in Nicaragua. The present study aimed to compare the genetic composition of different seed categories in the cultivar ‘INTA ROJO’, and to confirm the genetic identity of offtype seeds and plants found in this cultivar. The genotype frequencies of fourty individuals were analyzed using ten microsatellite markers in the following seed categories: Breeder’s seed, foundation seed, registered seed and certified seed. The genotype frequencies were analyzed using Fisher’s exact test, where breeder’s seed was assigned as a reference population. Additionally, twenty offtype seeds and plants, among them the offtype seeds known as “frijol viterra” and “frijol rojo oscuro”, were contrasted with breeder’s seed through pairwise comparisons of genetic distances. The results suggest that changes in genotype frequencies take place during seed production and they are ascribed to the selection pressures caused by environmental differences among production regions and inadequate varietal depuration procedures during seed production. In addition, the offtypes denominated “frijol rojo oscuro” were identified as an unknown cultivar probably derived from natural segregations, mutations and out-crossing among bean seed lots, and in less degree from accidental seed mixtures. In contrast, “frijol viterra” was confirmed to be the same cultivar ‘INTA ROJO’. Its differences in seed weights were associated rather to environmental effects than to genotypic ones. The seed technology implications of these findings and further perspectives are discussed.
  • Nemirov, Kirill (Helsingin yliopisto, 2003)
  • Pereira, Carlos Manuel Navarro (Helsingin yliopisto, 2002)
  • Tikkanen, Emmi (Helsingin yliopisto, 2013)
    Coronary heart disease (CHD) is a major burden for public health worldwide. Several factors are known to be associated with the disease risk, including high levels of low-density lipoprotein (LDL) cholesterol and blood pressure. The established risk factors do not, however, fully predict an individual s risk for the disease. In recent years, new candidate risk factors, including genetic markers, have been extensively studied. Genome-wide association studies (GWASs) have mapped over 40 genetic regions for CHD risk and hundreds of loci for CHD risk factors. The impact of these findings on public health remains obscure. In this study, we utilized the findings from large-scale GWASs and constructed genetic risk scores (GRSs) based on panels of single-nucleotide polymorphisms (SNPs). The aim was to estimate the joint effects of common genetic markers on CHD and its risk factors, and to evaluate the incremental value of genetic information in CHD risk assessment. In Projects I and II, we studied longitudinal effects of genetic loci associated with lipids and blood pressure, and evaluated the prediction of dyslipidemia and hypertension in young adults by using the genetic information in addition to clinical measurements. Our results show that the GRSs were significantly associated with longitudinal measurements of lipid traits and blood pressure throughout childhood, adolescence and adulthood. For some traits, the genetic effect was not consistent across age groups. For example, the GRS effect for high-density lipoprotein (HDL) cholesterol was considerably larger in children than in adults, and the proportion of variance explained by the SNPs in children was twice as much as in adults. The GRS for triglycerides improved the prediction of dyslipidemia in young adults when added to childhood lipid measurement. The blood pressure GRS increased the risk of hypertension, but did not improve risk discrimination over other risk factors. In Projects III and IV, we found that the GRSs based on CHD SNPs predicted CHD events. The estimated relative risk for the GRS was similar in magnitude to the relative risk of other risk factors such as systolic blood pressure. The genetic effect was independent of family history of the disease, which has been used as a surrogate for genetic risk in many prediction algorithms. The GRS based on 28 SNPs improved the prediction of CHD events beyond traditional risk factors and family history when evaluated with reclassification or discrimination metrics. Genetic screening could be especially useful for individuals in the intermediate-risk group (10-year risk 10-20%), as current preventive strategies are focused mainly on the high-risk group (>20%). In conclusion, these findings suggest that the genetic information obtained from GWASs could be used in early identification of individuals at increased risk for lipid disorders, hypertension and CHD. Keywords: cardiovascular disease, genetic association, genetic epidemiology, risk factor
  • Soronen, Pia (Juvenes Print - Tampereen yliopistopaino Oy, 2012)
    Mood disorders (major depressive disorder and bipolar disorder) are complex psychiatric disorders that are the leading cause of disability to work in Finland. Their specific genetic background remains widely unknown. Recent genome wide association (GWA) studies have revealed some novel susceptibility genes and risk variants as the field has moved into the era of genome-wide genotyping of huge samples sizes from collaborative studies. The risk variants revealed with this approach, however, cover only a small fraction of the total heritability of these diseases. Thus, it is still highly relevant to study samples from more genetically and environmentally homogenous populations, since the amount of risk variants that are enriched in isolated populations is smaller than in more confounded populations. Detailed characterization of the phenotype has also become an important factor in psychiatric genetics, since the psychiatric disease phenotypes themselves are so broad and heterogeneous. The principal goal of this work was to survey functionally relevant candidate genes for mood disorders and risk variants revealed from the first GWA studies in Finnish samples. The bipolar family sample comprises 723 individuals (227 affected for BD). Part of this sample has been characterized in more detail enabling the search for genetic susceptibility genes for potential endophenotypes of mood disorders. The clinical cohort sample is comprised of 272 MDD and 178 BD patients, who were characterized for clinical comorbidities and followed for long term clinical outcome. In the bipolar family sample, the highly studied variant val66met of BDNF was found to be associated with BD with the allelic replication observed in previous association studies. The most statistically significant result demonstrated the association of variants of the DAOA gene and the general intellectual function of visuospatial ability. The clinical cohort sample revealed statistically significant association of two functional variants from P2RX7 gene with familial form of mood disorder, and both variants affected also the clinical outcome of the patients by lengthening the time of illness. Our effort to replicate the results from the first GWA studies in the bipolar family sample revealed five genomic areas that associated with mood disorder also in Finland. Most of these variants also associated with some potential mood-disorder endophenotype, such as neurocognitive trait or seasonality in fluctuation of sleep, social activity, or mood. In conclusion, the use of detailed characterized samples may provide an advantage to the genetic studies, since it increases the possibility of finding the susceptibility genes behind more homogenous phenotypes. This work has also revealed some potential endophenotypes, or trait features, that might convey the effect of the susceptibility genes to the end diagnosis of mood disorder.