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Emerging research suggests that bacteriophages (phages) may exhibit alternative infection strategies that deviate from the preconceived lytic or lysogenic life cycles. Carrier cell infection is an alternative phage life cycle where complete virus particles are formed and remain within host cells, without cell lysis or integration into the host genome. Phage Φ6 (Φ6), the type member of the double-stranded RNA (dsRNA) virus family Cystoviridae, is a lytic phage that can also establish a carrier cell within its plant pathogenic host, Pseudomonas syringae pathovar (pv.) phaseolicola strain HB10Y (HB10Y). This thesis contributes to current limited knowledge and provides an insight on the underlying mechanisms of the Φ6 carrier cell infection. This study has agricultural and ecological relevance and may contribute to future plant therapeutic options. Synthetic carrier cell lines harboring Φ6 tri-segmented genome or Φ6 genomic constructs in which the coding regions in the S- and/or M- segments were replaced by heterologous sequences from tobacco mosaic virus (TMV) were created using a reverse genetics method. Spontaneous Φ6 carrier cell lines were also isolated from HB10Y after exposure of the host to excess phage. Spontaneous carrier cells were not stable, but rather occasionally released phage into liquid culture. Synthetic carrier cell lines were subjected to secondary phage infection and were found to be less susceptible than wild type (WT) to Φ6 but not Φ8, a more distant member of Cystoviridae. Studies suggest that carrier cell resistance to secondary infection (superinfection exclusion) is exhibited through the Φ6 S-segment gene 8. To test how temperature affects the stability of Φ6 carrier cells, spontaneous carrier cell line culture was incubated at RT and 30°C, and phage productivity was compared. Elevated temperature induced carrier cell stability. Comparison of the growth curves between Φ6 synthetic and spontaneous carrier cell lines and their respective WT strains showed that Φ6 carrier cell infection does not greatly affect host growth.

Introduction In addition to families and home environments, preschools can shape children’s eating habits. Children’s fruit and vegetable (FV) consumption remains below recommendations. Nutrition interventions should be aimed at young children when eating habits develop and be implemented in influential settings, like preschools. The preferred method to study and measure consumption is by weighing foods. However, weighing is hardly used in preschool-settings especially in group-level research. Aims The aim was to examine the effect of the DAGIS intervention on the consumption of FV in preschool meals. The secondary aim was to determine whether the results differed from the impact of DAGIS intervention on the FV consumption at group level when the amount of plate waste was considered. Materials and methods The DAGIS intervention was a randomized controlled trial (2017–2018) aiming to promote lifestyle and self-regulation skills affecting the energy-balance of Finnish 3–6-year-old. The 23-week intervention included 32 preschools. Vegetable consumption was measured at group level for three days in preschool before and after the intervention by weighing vegetables served, serving waste and plate waste. Data analyses were performed using Chi-square test, t-test, Wilcoxon signed-rank test and linear regression. Results In the follow-up, the median consumption of fresh FV, salad mixes, warm vegetables as a side dish and canned FV (25th; 75th quartile) per child was 81.3 g (55.2; 120.6) in the control group and 72.1 g (65.7; 119.3) in the intervention group. The total consumption of FV per child in the intervention group was not higher than in the control group after intervention. The inclusion of plate waste did not change the results adjusted by municipality. Conclusions There was no increase in FV consumption in the intervention group after intervention compared to the control group. Apart from highlighting FV consumption, future group-level interventions should be targeted at serving of vegetables. They should also critically consider the necessity of collecting plate-waste, especially if it increases research cost and burden.

Oat protein is a valuable cereal protein with high protein content and a good balance of amino acids profile. However, the inflexible molecular structure and poor watersolubility of oat protein limit its application in food industry. To exploit the food use of oat, the functionalities of oat proteins should be improved. ?-chymotrypsin, a proteolytic enzyme, has been proven as being able to deamidate some food proteins without severe hydrolysis under alkaline pH conditions, hence to improve the protein functionalities. The aim of this research was to test whether oat protein could be deamidated by ?- chymotrypsin under alkaline pH conditions. The extent of ?-chymotrypsin-induced proteolysis on oat protein was also studied. Oat protein was treated by ?-chymotrypsin under alkaline conditions from pH 8 to 11 at room temperature for 2 hours. The deamidation degrees of the proteins were measured by ammonia quantification. The hydrolysis degrees of proteins were analyzed by trichloroacetic acid precipitation and protein quantification method. The changes of molecular weights were analyzed by SDS-PAGE and SE-HPLC. The action of ?-chymotrypsin under alkaline pH conditions induced slight deamidation of oat protein to low deamidation degrees (9-12%). On the other hand, ?-chymotrypsin mainly caused hydrolysis of oat protein to hydrolysis degrees between 49% and 62%. SDS-PAGE and SE-HPLC analysis also revealed that significant hydrolysis of oat protein occurred during the reaction. The hydrolysis mainly caused the shift of oat protein fractions from molecular weight above 20 kDa to molecular weight below 15 kDa. Change in pH did not cause significant differences on deamidation degrees and hydrolysis degrees of oat protein In conclusion, oat protein could not be effectively deamidated by ?-chymotrypsin under alkaline pH conditions, while the proteolysis was severe.

DRama Queen (DRQ) is an unknown conserved gene that might be involved with sensing cytokinin signalling in the roots of Arabidopsis thaliana. It was originally found in a mutant screen when a mutated version of it – mDRQ – was found to improve the cytokinin sensitivity of plants in a desensitized genetic background. This thesis was done to define the protoxylem phenotype of mDRQ single mutant in Columbia (Col-0) background and to test the functionality of the gene by analyzing the complementation lines. The protoxylem phenotypes were scored from fuchsin stained samples by DIC microscopy. Protoxylem differentiation in the root of A. thaliana is an outcome of a mutually inhibitory signalling mechanism of auxins and cytokinins. Analysis of protoxylem status is a good tool for studying the cytokinin signalling because the differentiation is affected both by increased and reduced signalling levels. High signalling causes loss of protoxylem phenotypes whereas whereas low cytokinin signalling leads to ectopic protoxylem formation. The data of this thesis was analysed by SPSS version 22 software using cumulative logit modelling for the analysis. It was considered to be the most suitable alternative for analysis since the protoxylem phenotype data is ordinal by its nature. A protoxylem phenotype distribution was defined for mDRQ line and it was statistically different from Col-0. Most of the complementation lines were functional at a statistically significant level though the phenotype distributions of the complementation lines were not identical with original background. The difference could possibly be explained by the fact that the mDRQ gene is partially functional and may influence the phenotype distributions.

The present literature was reviewed to understand the recent indications of lipid degradation affecting β glucan molecular weight in aqueous solutions, the main topics being oat lipid reactions, oat β glucan, and oat bran/β glucan fortified yogurts, as well as the possible interplay of these three. A non-heat treated, a heat treated, and a heat treated and defatted oat bran concentrate (OBC) were produced, all with ~20% β glucan. OBCs were subjected to a storage trial in dark for 0–12 weeks at 4, 22 and 40 °C. Lipid degradation was followed during storage by assessing volatile compounds (HS-SPME-GC-MS), neutral lipid classes (NP-HPLC-ELSD) and tocopherol and tocotrienol levels (NP-HPLC-FLD) as well as β glucan molecular weights (HP-SEC). Acid milk gels were prepared as a model system for yogurts with added OBCs, and analysed for spontaneous and forced syneresis and textural changes. Lipid oxidation was observed in all OBCs, and hydrolysis in the non-heat treated OBC, but no changes in β glucan molecular weight were observed in the dry stored OBCs. OBC addition caused structural changes in an acid milk gels, the water retaining properties were improved, and the gels were softer and smoother than without added OBCs. Time and temperature affected OBC lipid oxidation and hydrolysis behaviours. Adding OBCs to gels changed gel textures compared to both without vs. with OBCs and adding fresh vs. stored OBCs.

The literature review dealed with the production of, and factors affecting the optimal processing of, traditional sauce base. Aspects of manufacturing relating to recycling and energy, such as recycling of animal by-products, were also discussed. The aim of the experimental work was to find a solution for the problem related to the opacity of the meat stock caused by a pressure cooking method. The key aim was to find the causes of opacity when the bones were pressure cooked (up to 1.5 bar) and to identify a method to prevent the occurrence of turbidity. Different cooking times, pressures and proteolytic enzyme combinations were evaluated during experimental studies. The goal was to obtain a beef-flavoured demi-glace sauce base broth with a bright appearance and as high a dry matter content as possible. Dry matter, total protein and connective tissue protein concentrations, pH and turbidity values were evaluated when comparing the results of the manufacturing methods and conditions. In addition, potential methods to improve the heat recovery systems were investigated. The dry matter of the broth consisted mainly of proteins. There was slightly faster increase in dry matter concentration at higher pressure than lower pressure. Addition of the proteolytic enzyme had the same effect. Based on this study the broth manufacturer must choose between a high gelatin content and turbidity when pursuing high dry matter content. The dry matter hydrolysed from bones by pressure cooking was almost exclusively collagen. Collagen dissolved in water while the rest of the bone proteins precipitated when heated. The construction of the heat recovery system in a small food processing company may be unprofitable. Improving energy efficiency in a small food company is challenging but possible if professionals accurately evaluate the conditions.

Crop monitoring in commercial indoor farming is a commonly used method in assessing the general productivity of the cultivated plants. This assessment practice is typically conducted manually by greenhouse workers and is sometimes supplemented by certain hand-held or stationary devices. An interesting example of novel device-assisted crop monitoring technologies utilizes digital imaging devices and computer-driven image analysis algorithms that have been prominently employed within the field of plant phenotyping. In the context of botanical studies, they have been used in e.g. characterizing various complex interactions between the genotypes of important food crops and their agronomic traits in specific prevailing environmental conditions. Additionally, image-based data acquisition technologies also present very interesting prospects for precision agriculture management practices. They could be harnessed to scan entire greenhouse compartments continuously and acquire massive amounts of data on multiple morphological and physiological aspects of crop growth and development in a non-destructive fashion. The acquired data could be implemented into mathematical greenhouse control models and utilized in a plethora of useful applications, including e.g. estimating and predicting biomass production and yield, detecting and localizing potential abiotic/biotic stress symptoms at an early stage, and ultimately enhancing overall crop production efficiency. In this thesis, these imaging technologies were explored in practice by designing and constructing a growth chamber embedded with automatic climate control and a low-cost multispectral imaging subsystem. The final assembly was tested by conducting a simple experiment involving drought-stressed sweet basil plants (Ocimum basilicum L. cv. ‘Genovese’) to determine how early drought-stress related symptoms could be detected purely from multispectral images. While the system carried out the tasks of automated climate control and continuous image capture adequately, the implemented approach in drought-stress detection was deemed unsuccessful. Significant differences between drought-stressed plants and their respective controls were not observed until visible symptoms were present. This was assumed to be due to incompatibility of the camera module’s spectral sensitivity in detecting changes in water content in plant tissue.

Yeasts are a major spoilage threat in carbonated and fermented beverages, causing considerable economic losses for the manufacturers. Dekkera bruxellensis and Zygosaccharomyces bailii are the two most common spoilage yeast in beverages due to their high tolerance towards beverage-related stress factors. For industry, early and reliable detection of contamination is necessary to minimize spoilage potential and maintain product quality. Cultivation on selective/differential media remains the main method for detection of these organisms, with incubation times from 3 to 15 days. Beverage-related stresses may generate sub-population of injured yeast cells and further delay or even prevent the detection in regular media. PCR, flow cytometry and other alternative detection methods also rely on enrichment cultivation to achieve the required sensitivity for the industry. Therefore, reduced incubation time of sample enrichment and improved detection of injured cells is crucial for a more rapid and reliable detection method. Modification of specific compounds in the culture medium composition has been reported to improve recovery of bacteria after stress. As analogue studies have not been performed on spoilage yeast, modification of the culture medium composition offers a possibility to improve the growth of injured and healthy yeast cells. The aim of this study is to reduce cultivation time required for detection of healthy and injured Dekkera bruxellensis and Zygosaccharomyces bailii cells. Initially, conditions for inducing organic acid and heat injury in D. bruxellensis, D. anomala and Z. bailii cells were studied in an artificial beverage containing basic components of soft drinks. Selective and non-selective plate cultivation and fluorescent viability stains were used to assess the level of injury. The organic acid treatments resulted in inconsistent injury of spoilage yeasts, and thus, recovery from organic acid injury could not be screened. The heat treatments resulted in consistent 1-3 log reduction of viable cell counts. Altogether, 46 potential injury-relieving or growth-enhancing supplements were screened for their effects on the growth rate and lag time of heat-treated and untreated cells in non-selective YM broth using high-throughput automated turbidometry. During individual screening, the growth of Z. bailii strains was significantly improved (p<0.05) only by supplementation with three ion sources: calcium chloride, potassium chloride, and magnesium sulphate. Synergistic effects of the three ion sources was optimized for D. bruxellensis and Z. bailii individually using surface response analysis. Optimized D. bruxellensis YM medium showed no consistent impact on healthy or heat-treated D. bruxellensis strains. On the other hand, two out of the three Z. bailii strains showed significant lag time reduction of 63-66% in untreated cells and 34% in heat-treated cells when incubated in optimized Z. bailii YM medium. The lack of differentiation between improvement of growth of untreated and heat-treated cells point to a generalized ionic deficiency in YM medium. In conclusion, the optimized Z. bailii YM medium is a promising candidate for reducing the detection time of the common spoilage yeast, but it would still require validation with additional Z. bailii strains and quality control samples. It would be also interesting to study the benefits of the medium for cultivation of other spoilage yeasts and in the presence of Z. bailii selective compounds. The information about the importance of various salts for growth of Z. bailii may also prove useful in biotechnological applications of this yeast.

In the past 20 years, three known disease emergence events of highly pathogenic coronaviruses have highlighted the importance of monitoring wildlife for the presence of these viruses. Their peculiar characteristics, like high mutation and recombination rate, have increased their potential for species adaptation and interspecies transmission. Understanding the diversity of these viruses in wildlife and increased surveillance might be key to predicting and preventing future spillovers and pandemics. Studies on wildlife coronaviruses commonly focus on the order Chiroptera, mainly in temperate and tropical regions of the Asian continent. Even though animals belonging to this order are considered the main reservoir, the importance of other small terrestrial mammals should not be overlooked. Rodents, for instance, are animals of great interest for many zoonoses, as they often host parasites, bacteria and other groups of viruses that cause diseases in humans. A recent description of several lineages of coronaviruses recovered from rodents from China highlighted and suggested the presence of these viruses in small terrestrial rodents. In this project, we aimed to investigate the presence of coronaviruses in small mammals from France. Samples were collected during spring 2021 in twelve different locations, within two regions of eastern France, Auvergne Rhône-Alpes and Franche Comté. A total of 448 rodents, 13 shrews and 416 bat samples were collected. The samples were screened and coronaviruses sequences were recovered in 20 different samples. Nine Betacoronavirus genus sequences were recovered from rodent colon samples, and one Alpha- and ten Betacoronavirus sequences from bat guano. These results confirmed previous evidence of these viruses’ presence in small mammals from France and provide the first evidence of betacoronaviruses circulating in wild French bats. The study covers two eastern regions that have not been surveilled in previously released studies therefore this highlights the need to increase the efforts in monitoring these viruses and their wildlife host

Molecular biology has created a new pathway for plant breeding in cut flower industry. It focuses on studying flower gene functions and provides a more direct and effective way of breeding new flower cultivars using genetic transformation. Besides flower color, disease resistance, quality and vase life, modification of flower architecture is an important target for flower breeding. Previous studies have showed that various transcription factors encoded by the corresponding genes are involved regulating flower development and flower architecture. The most studied are MADS domain and TCP domain transcription factors. For targeted breeding, it is crucial to study the functions of the corresponding genes in detail. For both MADS and TCP domain proteins, previous studies have indicated that protein-protein interactions are important for their function. GhCYC1, GhCYC2, GhCYC3 and GhCYC4, isolated from gerbera (Gerbera hybrida), are CYCLOIDEA –like genes affecting inflorescence development. The protein-protein interactions among these four genes have previously been studied by yeast two-hybrid system. The aim of this thesis was to verify the interactions in living plant cells, using both BiFC and split luciferase assays. Protoplast electroporation and agroinfiltration were used to introduce the genes in planta. The results from the two assays were compared in order to find an effective in planta method for detecting protein-protein interactions. The experiment also provided information about DNA transformation efficiency using protoplast electroporation and agroinfiltration. The results of the split luciferase assay showed that GhCYC1+GhCYC4, GhCYC3+GhCYC4 as well as GhCYC4+GhCYC4 interacted quite strongly in plant cells while GhCYC1+GhCYC1, GhCYC2+GhCYC2 as well as GhCYC4+GhCYC2 had almost no interactions. The interactions between GhCYC3+GhCYC4, and GhCYC4+ GhCYC4 were also shown in yeast two-hybrid, but the other results were different. According to the BiFC assay, no signals of interactions were detected from GhCYC2+GhCYC2, while strong signals were observed from GhCYC2+GhCYC3, and weak signals were seen from GhCYC2+GhCYC4. The interactions between GhCYC2+GhCYC3, GhCYC2+GhCYC4 were also observed in yeast two-hybrid, but the other results were unconfirmed. Large standard deviations were observed in the split luciferase assay and thereby reliable conclusions cannot be drawn from it. However, BiFC turned out to be a better method to detect the protein-protein interactions in planta and clear signals from interactions could be observed. Comparison of the transformation methods indicated that agroinfiltration is a better way of introducing DNA into plant cells than protoplast electroporation. For further study, BiFC assay still needs to be repeated to confirm the efficiency of this assay, and factors affecting the transformation efficiency in protoplast electroporation need to be optimized in the future studies.