CONTRIBUTION TO DEVELOPMENT OF MESHLESS METHODS FOR FREE AND MOVING BOUNDARY PROBLEMSNAZIA TALAT
, 2018, doctoral dissertation
Keywords: Two-phase flow, free and moving boundaries, computational fluid dynamics, phasefield formulation, 2D problems, axisymmetric problems, diffuse approximate
meshless method, Rayleigh-Taylor instability, Boussinesq approximation, variable
density and viscosity, flow focusing, dripping, jetting
Published: 11.09.2018; Views: 1369; Downloads: 76
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POLYMORPHISMS IN GENES FOR ENDOTHELIN 1, ENDOTHELIN RECEPTORS AND NITRIC OXIDE SYNTHASE 3 IN PATIENTS WITH DIABETIC RETINOPATHY AND DIABETES MELLITUS TYPE 2Dejan Bregar
, 2018, doctoral dissertation
Abstract: The major causes of Diabetes Mellitus Type 2 (T2DM) are multi-factorial consequences of complex interactions between environmental, social and genetic factors. We investigated the genetic risk factors in Slovene patients with T2DM on a model of microvascular complication – Diabetic Retinopathy (DR).
Retrospective case-control study includes a T2DM Slovene population with clinical risk factors for T2DM and DR. Only some of the candidate genes with selected single nucleotid polymorphisms (SNPs) were included: (EDN1 (rs5370, rs3087459, rs1476046), EDNRA (rs5335, rs1801708), EDNRB (rs10507875, rs4885493), NOS3 (rs869109213).
By genotyping with either real-time polymerase chain reaction or standard polymerase chain reaction (PCR) we successfully identified the contribution of variable number of tandem repeats rs869109213 in DR progression (Proliferative Diabetic Retinopathy (PDR)) in Slovene patients with T2DM. The joint effect of individual genotypes of rs10507875 in EDNRB and rs869109213 in NOS3 on DR onset (DR) and DR progression (PDR) was demonstrated as well. The joint effect of the two polymorphisms on DR onset (DR) and DR progression (PDR) was greater than the individual effect of each polymorphism separately in the analyzed genetic models.
Despite genetic research contributions in DR, linkage studies, and Genome-wide association studies the identification of susceptible loci through candidate gene approaches still remains in its early stages. The frequent approach with an ultimate focus on SNP associations with phenotype is likely to underestimate the roles of genetics in human diseases by disregarding not only the joint effect of multiple loci but the complex interaction network between them. By identifying polymorphisms in genetic disorders in a more systematic way, we will be able to deepen our understanding of the regulatory mechanisms and disease etiology which should lead to a more effective development of mechanism-based therapies as well.
Keywords: endothelin 1, EDN1, endothelin receptor A, EDNRA, endothelin receptor B, EDNRB, nitric oxide synthase 3, NOS3, diabetic retinopathy, DR, diabetes mellitus type 2, T2DM, polymorphism, SNP, genetic model
Published: 07.09.2018; Views: 1055; Downloads: 83
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Amorphous nanocomposite of polycarbosilanes and aluminum oxideAndraž Mavrič
, 2018, doctoral dissertation
Abstract: This work presents a paradigm for high temperature stabilization of bulk amorphous aluminium oxide. The thermodynamic stabilization is achieved by preparing a nanocomposite, where polymethylsilane dendritic molecules are dispersed in an aluminium hydroxide gel. Upon heat-treatment the gel transforms to the amorphous aluminium oxide that is stable up to 900°C. The dispersion of the macromolecules and their covalent bonding to the alumina matrix induce homogeneously distributed strain fields that keep the alumina amorphous.
The first part of the thesis focuses on the synthesis, characterization and solubility properties of the dendritic polymethylsilane. The polymethylsilane is synthetized by electrochemical polymerization from trichloromethylsilane monomer. The polymerization mechanism, involving a single polymerization pathway, is identified. The polymer growth proceeds through reduction of the monomers to the silyl anions and their addition to the growing polymer.
The solubility of three chemically related but topologically different polysilanes (linear, dendritic and network) were studied by dynamic light scattering. At room temperature the agglomerates in a range from 500 to 1300 nm are present. They undergo de-agglomeration at slightly elevated temperatures of around 40°C. The de-agglomeration results in formation of stable solutions, where a hydrodynamic diameter of the individual polymer molecules was measured to be in a range from 20 to 40 nm.
The obtained diameters of two dendritic polymethylsilane macromolecules, synthesized under different electrolysis conditions, are much larger than the theoretical size estimated for an ideal dendrimer. We determined by 29Si NMR that the reason for this is in a large number of branching irregularities (defects) contained in the molecular structure. Combining the experimental values obtained by DLS and density measurements with a structural model that considers the branching irregularities, it is shown that the inclusion of the defects allows the dendritic polymer to exceed the sterical limitations and form the hyperbranched dendritic structure. The final size depends on a relative amount of the branching defects.
In the second part, the synthetized polymethylsilane molecules were successfully used for the nanocomposite formation. The aluminium hydroxide gel with the dispersed polymethylsilane molecules was prepared as a precursor. Upon heat-treatment it gives the amorphous aluminium oxide stable up to 900°C. The dispersed macromolecules induce homogeneously distributed strain fields that keep the aluminium oxide amorphous during the thermal treatment the dispersed macromolecules covalently bind to the matrix, inducing the interface strain. The amorphous state was confirmed by the presence of penta-coordinated aluminium detected by 27Al NMR and a low bandgap measured by UV-vis absorption spectroscopy.
Keywords: amorphous aluminium oxide, polymethylsilane, nanocomposite, electropolymerization, solubility, agglomeration, de-agglomeration, dendrimer, hyperbranched dendritic structure, dynamic light scattering, thermal analysis, transmission electron microscopy, scanning electron microscopy, X-ray diffraction, infrared spectroscopy, UV-Vis spectroscopy
Published: 19.07.2018; Views: 2125; Downloads: 110
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DISTRIBUTION OF ENTERIC VIRUSES IN THE GULF OF TRIESTE AND THEIR INTERACTIONS WITH ENVIRONMENTAL AND BIOLOGICAL PARAMETERSJosé Manuel Carita Gonçalves
, 2018, doctoral dissertation
Abstract: The available classical diagnostic methods, due to many disadvantages, do not allow effective detection of pathogenic enteric viruses in environmental samples. Due to low concentrations of pathogenic viruses in the sea, it is important to develop an effective concentration procedure for their successful detection.
In the first part of the doctoral thesis, we focused primarily on the development of a protocol for an effective concentration of pathogenic enteric viruses in coastal water samples. Monolithic chromatographic columns (BIAseparations) were used for the concentration of rotaviruses and noroviruses, prior to the detection with reverse transcription quantitative PCR in real time (RT-qPCR). We tested the efficiency of concentration using columns of various chemical properties and selected pathogenic enteric viruses (rotavirus and norovirus). Among them, hydrophobic interaction monolithic column (CIM® C4) was the most effective. CIM C4 was used to optimize the concentration step and tested in waters with different salinities.
The presence of concentrated viruses was confirmed by RT-qPCR and transmission electron microscope. We have developed a protocol that enables rapid concentration of viruses in coastal waters of various salinities and can be used on-site.
The presence of RoV and NoV was surveyed, using the developed concentration protocol, prior to one-step RT-qPCR molecular detection, in the inner part of the Bay of Koper, in mussel farming areas and a swimming area. Rotaviruses, noroviruses and fecal indicator bacteria were frequently detected in the inner part of the Bay of Koper.
Rotaviruses and noroviruses were detected in the studied area, with higher rates close to the outfall of the wastewater treatment plant in the estuary of river Rižana and were also detected in the middle of the Bay of Koper and in areas used for recreation and mussel farming.
The results show that water bodies, which are otherwise defined as suitable for bathing or mussel farming, based on the results of fecal indicator bacteria, still contain low concentrations of pathogenic enteric viruses.
In addition to human pathogenic enteric viruses and faecal coliforms, changes in abundance of bacteria and virus particles were studied in relation to temperature, salinity, inorganic and organic nutrient concentrations in the organically polluted Rižana estuary. Preliminary results showed spatially and seasonally changes in bacterial and viral particles abundance, and bacterial composition spatially and seasonally. However, seasonality plays a greater role in bacterial dynamics.
Keywords: Concentration of viruses, Enteric viruses, Rotavirus, Norovirus, Feacal coliforms, Feacal contamination, qPCR, RT-qPCR, Monolithic columns, Sewage, Seasonal dynamics, Concentration, Coastal environment, Gulf of Trieste
Published: 02.07.2018; Views: 1846; Downloads: 130
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STRUCTURE AND FUNCTIONING OF THE HYPORHEIC ZONES IN THE GRAVEL-BEDS OF FIVE RIVERS IN RELATION TO CATCHMENT LAND USEBarbara Debeljak
, 2018, doctoral dissertation
Abstract: The hyporheic zone has been recognised as a functionally important component of streams and rivers. Due to increasing human impact on ecosystems, studies that assess ecological responses of the hyporheic zone are of great importance. The objectives of the thesis are to study the responses of abiotic parameters, sediment biofilm characteristics (the in situ respiration (R), potential respiratory activity (ETSA), protein content), and invertebrate assemblages (analysed as total assemblages, EPT assemblages and Copepoda assemblages) in the hyporheic zones related to different land use patterns (forest, agricultural and urban areas). The focus of study also includes the impact of clogging. This research is comprised of three sampling campaigns conducted in summer (2013), winter (2013) and spring (2014) in five pre-Alpine Slovenian rivers. For each river, three or six sampling locations were chosen in the downwelling hyporheic sections of three dominant land uses (forest, agriculture and urban) within a 250 m wide impact zone. Three sampling points per location were sampled from two depths (5 –15 cm and 20 – 40 cm) using PVC tube and Bou-Rouch method. The results indicate a significant spatial and temporal heterogeneity of measured hyporheic zone components. Physical and chemical parameters of water revealed moderate response to land use. The effect of land use on the amounts of suspendable fine sediments in the hyporheic zone was observed only in the spring season. The effect of land use was significant for in situ R during summer and spring and for ETSA and protein content in all seasons, indicating that land use, such as agriculture, near a stream can affect biogeochemical processes. A relatively high diversity of invertebrates was recognised in the hyporheic zone. Groups such as Nematoda, Oligochaeta, Copepoda and Chironomidae were found in all samples. Within EPT taxa as representatives of occasional hyporheic invertebrates, 35 taxa were identified. Within Copepoda, as representatives of permanent hyporheic invertebrates, 14 Cyclopoida and 19 Harpacticida taxa were identified. Both assemblages were characterised by high abundance of widespread taxa, such as Baetis sp., Leuctra sp., Diacylops cladestinus and Acanthocyclops hispanicus. Statistically significant differences were observed among land uses in Copopoda assemblages but not for EPT assemblages. Calculated metrics on EPT assemblages showed statistically significant differences in land use in the number of EPT taxa, Simpson index and Shannon-Wiener index. Within Copepoda assemblages, the numbers of Copepoda taxa were significantly higher at forest sites. The hyporheic assemblages were relatively well explained by environmental parameters. This thesis presents a comprehensive study of the hyporheic zone, where both structural and functional measures reflected the ecological integrity of the hyporheic zone. The in situ R and ETSA were generally higher in agriculture stream reaches, indicating that hyporheic functioning responded to nutrients and carbon runoff-derived inputs from agricultural activities within the 250 m impact zone. Thus functional parameters in situ R and ETSA were more efficient indicators of land use impacts in the hyporheic zone than physical and chemical parameters. The potential of using hyporheic invertebrates as bioindicators of environmental conditions was confirmed but should be tested with wider gradients of environmental parameters. The results obtained by this study underlined that the hyporheic zone should be integrated as an additional ecological element by assessing the ecological conditions of surface water bodies.
Keywords: hyporheic zone, ecosystem functioning, land use, biofilm characteristics, invertebrates
Published: 02.07.2018; Views: 1452; Downloads: 55
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MODIFIED U1 RNAs AS SPLICING CORRECTORS IN HUMAN GENETIC DISORDERSKatarzyna Rajkowska
, 2018, doctoral dissertation
Abstract: The experimental work of this thesis was performed at the International Centre
for Genetic Engineering and Biotechnology (ICGEB) in the Human Molecular
Genetics Group, under the scientific direction of Prof. Franco Pagani. The project
was developed during the academic years 2014-2017.
Modified U1 RNAs, also named Exon Specific U1s (ExSpeU1s) represent a novel
class of small RNA-based molecules that correct exons splicing defects. To evaluate
their therapeutic potential focused on Familial Dysautonomia (FD), a rare autosomal
recessive disorder characterized by progressive degeneration of the sensory and
autonomic nervous system. More than 99% of patients are homozygous for the T to
C transition in position 6 of the IKBKAP intron 20 (c.2204+6T>C). This substitution
modifies the exon 20 5’ splice site (5’ss) inducing exon skipping in a tissue-specific
manner and reducing the total amount of IKAP protein. The molecular mechanisms
underlying the IKBKAP mis-splicing are not completely clear and there
are no effective treatments.
In this thesis, I investigated the therapeutic potential of ExSpeU1s and the role
of cis- and trans-acting factors that regulates IKBKAP splicing. Using a splicing
functional assay, I identified ExSpeU1s that bind to intron 20 sequences and rescue
the exon 20 skipping defect. Interestingly, their rescue activity was modulated
by several splicing factors and requires a critical exonic splicing enhancer element.
Transfection experiment showed the involvement of both enhancing (TIA1, PTBP1
and PTB4) and inhibitory (SRSF3, hnRNPA1, FOX and FUS) splicing factors
in IKBKAP splicing. To better evaluate the ExSpeU1s therapeutic efficacy,
I transduced FD patient’s fibroblasts with a lentiviral vector expressing the most
active ExSpeU1. This resulted in a complete rescue of the exon skipping defect and
improvement in IKAP protein expression. Most importantly, intraperitoneal delivery
of ExSpeU1s by AAV9 into a transgenic mouse model, that recapitulates
the tissue-specific mis-splicing seen in FD patients, corrected the aberrant splicing
patterns in several tissues increasing the amount of the corresponding IKAP protein.
All together, these results identify novel regulatory splicing factors involved in the
IKBKAP exon 20 regulation and provide the proof of principle that ExSpeU1s
delivered in vivo by AAV vectors represent a novel therapeutic strategy for FD.
Keywords: Familial Dysautonomia, IKBKAP, IKAP, splicing, splicing defects, ExSpeU1, U1 snRNA, mouse model, AAV
Published: 26.03.2018; Views: 1815; Downloads: 52
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Speleogenetic factors and processes in the karst conduits of Zagorska Mrežnica Spring Cave (Croatia)Petra Kovač Konrad
, 2018, doctoral dissertation
Abstract: Zagorska Mrežnica spring cave is in the Desmerice village, 7.59 km SW from the town of Ogulin. The coordinates of the entrance are X: 399563 m, Y: 5006974 m, and Z: 314 m (HTRS96). The cave system is positioned in the contact zone of Jurassic limestone and the thrust front composed of Triassic Dolomite. The karst drainage system has elements of point recharge through a set of ponors in the hinterland karst poljes and diffuse infiltration through numerous dolines on Velika Kapela Mountain. The karst of Ogulinsko Zagorje area and its epiphreatic and phreatic cave systems have been intensively explored for the last eight years, resulting in the mapping of 1134 m of submerged passages of the Zagorska Mrežnica spring cave. Until 2014 it was the longest mapped submerged system in Croatia explored by cave diving techniques only. Altogether, over 3.5 km of submerged passages in seven caves of Ogulinsko Zagorje (Zagorska Mrežnica spring cave, Spring of Rupečica, Ponor of Rupečica, Cave system Pećine-Veliko vrelo, Spring of Bistrac, Cave Zagorska Peć and Pit Klisura) were explored during 500 hours of diving over a ten-year period. The goal of the research was to determine speleogenetic factors and processes in phreatic conditions. A new methodology for mapping of cave cross-sections, microrelief forms, and structural elements was developed, sediment and petrographic analysis were done, hydrological analysis before and after the building of the accumulation lake Sabljaci, 48 cross-sections of the cave passaged and a 3D model of the cave system was created, water chemistry as well as geomorphological analysis was carried out. In the end, a simplified (modified) vulnerability assessment was also done.
The results of my research showed that the distribution of the cave passages is a result of complex tectonic activities that are reflected in the orientation of cave passages. The general orientation of the measured fissures, measured during cave diving, show a dominance of a NWW-SEE direction but also the pattern of cave passages shows a significant dominance of the NE-SW orientation indicating the existence of multiple secondary traverse faults originating from main NW-SE faults. The morphology of the cave passages shows a transition between epiphreatic (possibly vadose) and phreatic phases as well as the presence of paragenetic developments in the ceiling. This indicates the existence of several epiphreatic (possibly vadose) and phreatic speleogenetic phases. The initial shape of the cross-sections was identified by sub-horizontal beds and bedding planes. The mapping of microrelief forms showed that the mechanical erosion was a very intensive process shaping the morphology of the cave passages.
The sediment mineralogy coincides with the lithology of the catchment area, and the sub- angular grains reveal relatively short transport distances. Further analysis of the catchment area’s geomorphology and its relevance for the speleogenesis of Zagorska Mrežnica cave show influence of relief structures that have a Dinaric orientation (NW-SE) with cave structures and passage orientations, great relative relief, steep slopes on Velika Kapela, and flat areas of karst poljes dictate the allogenic and autogenic hydrological regime of the cave. A 3D model of the cave system reveals a relationship between the cave system and the landscape, as well as a possible connection with the nearby Zagorska Peć cave. The modified karst vulnerability assessment shows that the catchment area of Zagorska Mrežnica spring cave has a high vulnerability ranking, mainly due to high doline density and the potential large diffuse capacity of infiltration of pollutants.
Keywords: Ogulin Zagorje, karst, spring, speleogenesis, Zagorska Mrežnica spring cave
Published: 02.03.2018; Views: 1926; Downloads: 92
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Ecological evaluation of aquatic and terrestrial subterranean fauna in a karst cavePeter Kozel
, 2018, doctoral dissertation
Abstract: Studies on environmental parameters−subterranean fauna relationship and spatial and temporal patterns of subterranean fauna have dealt with either aquatic or terrestrial faunas. So far, no simultaneous ecological evaluation of aquatic and terrestrial faunas has been performed. To address this issue, we conducted a pilot study dealing in parallel with these two faunas in the cave Zguba jama near Postojna, Slovenia, applying monthly sampling over one year. Aquatic fauna was sampled from permanent water drips, and terrestrial fauna by visual inspection and advanced baited pitfall trapping, along with recording the main abiotic parameters. In percolating water, we recorded aquatic fauna, Copepoda being the dominant taxon, and species-rich troglobiotic fauna, which most probably coexist with aquatic species in a semi-aquatic epikarst environment. Analyses revealed that temperature, distance from the entrance and ceiling thickness are the most important parameters that influence the presence of stygobionts; higher values of these parameters result in a higher probability of the presence of stygobionts in percolation water. The recorded spatial distribution of stygobionts varied distinctly among the sampling sites. This indicates that fauna in sampled drips originated from aquatic or semiaquatic epikarst microhabitats, being completely or nearly completely separated from each other, and probably varied according to environmental characteristics. The temporal pattern of the stygobiotic fauna dynamics presumably occurred because of variable water flow rates and the specific physical and chemical characteristics of the water. The abundance of troglobionts showed distinctive spatial distribution pattern over the seasons. In spring and summer, the highest abundances and the highest probability of presence were found near the entrance. In autumn and winter, unfavorable conditions in the entrance zone most likely triggered the migration of troglobionts towards the climatically more stable deep cave zone, and from the cave into the adjacent fissure network. The highest abundance of troglobionts was found at 7–9°C and relatively high ground substrate moisture. Additionally, we found a higher probability of the presence of troglobionts in cave sections with a larger passage cross-section size, which is probably due to the greater variability of microhabitat types in spacious cave sections. In addition, the higher probability of presence relates to higher substrate pH. One potential explanation for this could be that the specific microorganism communities present in such conditions support the nutritional needs of troglobionts. Most troglobiotic species preferred the deep cave zone where the highest species richness and diversity were also observed. Species richness, abundance and diversity of troglophiles and trogloxenes were highest in the entrance zone and showed relatively similar patterns within the cave. However, troglophiles were more abundant in the transition zone and the initial part of the inner zone. The temporal pattern of trogloxenes and presumably of most troglophiles occurred because of migration between the surface and the cave. The probability of the presence of trogloxenes was highest in the entrance zone and in winter, owing to massive overwintering in the cave. A modified technique of pitfall trapping by placing traps in two parallel sets, a ground and an upper one along the cave, resulted in recording greater species diversity in comparison with the traditional method, i.e., ground pitfall trapping alone. It turned out that such sampling contributes at the same time to both more effective and less invasive inventory of subterranean fauna. The dynamics of relative abundance and species richness of stygobionts and troglobionts within the cave showed partly comparable annual patterns. Nevertheless, our findings point toward the need for a long-term and more detailed study in future to address this question properly.
Keywords: biological inventories, epikarst fauna, faunal dynamics, microhabitats, stygobionts, troglobionts, troglophiles, trogloxenes, Zguba jama
Published: 16.02.2018; Views: 1549; Downloads: 198
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SIGNALING STUDIES IN THE EMERGING KIWIFRUIT PATHOGEN Pseudomonas syringae pv. actinidiaeSree Gowrinadh Javvadi
, 2017, doctoral dissertation
Abstract: In the past two decades emerging and re-emerging plant pathogens have caused new threats to the
production of several economically important crops, one among them is P. syringae pv. actinidiae
(PSA) which causes canker or leaf spot on kiwifruit plants. PSA enters plant through wounds and
remains dormant in cortex tissue of the branches, and spreads in the tissue to cause severe
symptoms from winter to early spring. The disease can be visualized by brown discoloration of
buds, dark brown angular spots surrounded by yellow haloes on leaves, cankers with white to
reddish (oxydation) exudate on twigs and trunks, fruit collapse, wilting and eventually plant
mortality. Current control methods have their own significance in disease control, however there
is considerable lack of clear understanding of PSA pathogenicity. Virulence of plant pathogens
often relies on the synchronized/coordinated expression of pathogenicity factors via quorum
sensing (QS). Therefore, investigations on QS in PSA may lead to develop novel disease control
strategies and reliable methods to curb the disease. It is currently unknown whether PSA produces
a QS signal molecule thus the aim of this thesis is to investigate whether PSA possesses a QS
system. As genome mining did not reveal the presence of any currently known QS system, this
study initially by metabolomics was aimed at identifying potentially low molecular weight
secondary metabolite QS molecules produced by PSA. Azelaic acid was discovered to be produced
by PSA, this is the first report of azelaic acid production by bacteria. The characterization and
possible role of azelaic acid in QS is presented. Since azelaic acid is ubiquitous in nature, in
addition to determining its biological role, the catabolism of azelaic acid in bacteria using the
efficient degrader Pseudomonas nitroreducens DSM 9128 was also studied.
Keywords: Quorum sensing, Azelaic acid, Metabolomics, Catabolism
Published: 19.12.2017; Views: 2026; Downloads: 117
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Development of Slovenian linguistic terminology in Slovenian grammars in the 18th and 19th centuriesMitja Trojar
, 2017, doctoral dissertation
Abstract: The dissertation describes the problems of metalanguage and peculiarities of linguistic terminology. Special attention is paid to autonymy, which plays a central role in metalanguage.
The dissertation presents the life and work of Eugen Wüster, the founder of traditional terminology, and the principal tenets of his approach to terminology. Traditional terminology is critically evaluated; we show its inability to describe linguistic terminology: it lacks a coherent theoretical apparatus enabling description, its semantic theory is based on classical definition, which in many cases cannot be formulated, which means that traditional terminology is also unable to account for translation of terms and comparison of concepts across different concept systems.
A sample of 50 basic linguistic terms from 19 grammars from the 18th, 19th and 20th centuries was used to examine the development of Slovenian linguistic terminology and to ascertain who played the most important role in it. The research shows that Anton Janežič made by far the most important contribution to Slovenian linguistic terminology: he introduced 27 terms (in both his 1854 and 1863 grammars), which represents 54% of the 50 terms retained by Breznik.
The dissertation also contains some remarks on the semantics of linguistic terms.
The scientific relevance of the thesis lies in its critical evaluation of traditional terminology; the results show that traditional terminology faces a number of theoretical problems; a new theory of terminology is needed, one that will be able to account for translation of terms as well as for diachronic conceptual change. The scientific relevance of results related to the second research goal (determining the contributions of individual grammarians to the development of Slovenian linguistic terminology) consists in proposing a model of research that could be used in order to measure the influence of a specific author on the introduction of new terms.
Keywords: metalanguage, linguistic terminology, history of terminology, development of linguistic terminology, diachrony
Published: 27.11.2017; Views: 3290; Downloads: 217
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