PhD theses

         Consumers' awareness about the nutritional value of different foods and their therapeutic effect on the human body is constantly increasing. The consumer is trying to find low-processed products in which the individual nutrients retain their original properties and thus have a beneficial effect.
      Many scientific sources pay particular attention to the wide use of soluble fiber fractions, for example beta-glucans in the production of a functional food. Some of them have good water solubility and can be used as thickening agent in various food products. In addition to rheological properties, beta-glucans show many pro-health properties. Consumed regularly, they reduce the incidence of diet-related diseases associated with: hyperinsulinemia, hyperlipidaemia, cancer and weakened immune response.
      In the gastrointestinal tract, as in food products, glucans combined with other ingredients create a thick gel, which are not able to be digested by human endogenous enzyme and prevents digestive enzymes from accessing proteins and fats. They can also affect the activity of digestive enzymes. The use of beta-glucans, especially in higher concentrations, may affect protein and fat metabolism during digestion. Therefore, the main objective of the proposed PhD study is to investigate the impact of modification of production technology and chemical composition on the enzymes activity, along with bioavailability of proteins and fats from dairy products.
      The proposed subject of this PhD thesis is interdisciplinary, because it required collaboration of specialists representing different branches of science.

        As a result of rapid development associating with technological progress a demand for the biomaterials with unique properties is increasing. Biopolymers are divided into three categories: chemically synthesized polymers, starch-based biodegradable plastics and polyhydroxyalkanoates (PHAs). Among them, PHAs are the most known, being recognized as completely biosynthetic and biodegradable with zero toxic waste, and completely recyclable into organic waste. For the reason that PHAs have useful properties such as: biodegradability, thermoplasticity, biocompatibility, non-toxicity, they are considered a replacement for petrochemical polymers. PHAs introduction to the world-wide market is currently limited due to their increased production cost compared to their synthetic alternatives. Therefore, there is a growing need for the development of novel microbial processes using inexpensive carbon sources. The promising feedstocks for PHAs production are volatile fatty acids (VFAs) produced from biomass and biowaste.
         Earlier work indicated that VFAs could be used as carbon sources for PHAs production but there is a lack of information on how VFAs of different composition influence the structure and properties of PHAs during microbial fermentation. Therefore, the main goal of the PhD thesis is to investigate the effect of volatile fatty acids obtained during biological routes on the PHAs synthesis process using pure bacteria cultures. The fermentation studies using different feeding strategies will be conducted in the bioreactors. The PHAs will be extracted, then the composition and physical properties of them will be determined. Furthermore, the impact of different VFAs on the molecular level will be carried out.
          The proposed subject of the PhD thesis is interdisciplinary because it links environmental engineering with biotechnology to obtain innovative bioplastics and to make their cost-effective production more feasible.

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          The proposed subject of dissertation is focused on morphology and genetics of fish from family Bramidae together with the identification of specific parasites. Data about species diversity of Bramidae in Southeast Pacific Basinis scarce. There is only a little information about the bathymetric occurrence of Brama  australis and research data on age, growth and mortality B. australis  in the southeastern Pacific Ocean. The fish individuals from Family Bramidae caught in the southeastern Pacific Ocean near Chile were classified as Brama australis, but within caught fish the three different morphological forms were noticed. Probably there are three different species or some hybrids.
          Currently the most fish populations are endangered (some are extincted) and misidentification or false species identification is a serious failure that have a negative impact and decrease the fish biodiversity. In this situation, the detailed fish species identification based on morphological traits, molecular markers and identification of specific parasites should be conducted. The application of proposed analyses allows the precise species or hybrids identification and gain the detailed information about biodiversity of Bramidae in studied geographical region.
          The proposed subject of the dissertation has an interdisciplinary character which cross the boundaries between different fields of research, including:ichthyology, fisheries, fish species identification, morphology, parasitology, application a new genetic tools in fisheries, molecular genetics and biological study of Bramidae that are important in fish biodiversity.

         Transcriptome analysis on high-throughput RNA-sequencing (RNA-Seq) data has been performed to identify sperm transcripts associated with the freezability of boar semen. It is envisaged that such approach will allow the use of a wide range of bioinformatics tools to explore candidate genes that contribute to sperm cryo-survival. In addition, a validation scheme (qRT-PCR, Western blotting) will be employed to reaffirm the biological roles of several different differentially expressed (DE) genes in sperm function following semen cryopreservation. It is likely that the utilization of transcriptome profiling of spermatozoa, in conjunction with advanced bioinformatics screening, will aid in the improvement in boar semen cryopreservation. The subject matter of the doctoral thesis intersects in the study of various disciplines and will fill in critical gaps in the knowledge of sperm-associated gene transcripts in semen freezability.

     The main concern for the release of antibiotics into the environment is related to the development of antibiotic resistance genes (ARGs) and bacteria (ARB), which reduce the therapeutic potential against human and animal pathogens. Antibiotic resistance is determined by genes located on the bacterial chromosome or mobile elements, such as plasmids, transposons and integrons, which are efficient vectors for the spread of these genes between bacteria. Transfer of genetic information between individuals is achieved by two mechanisms: vertical, from parent to siblings, and horizontal (HGT) between individuals of the same or different species. Whereas the former ensures the maintenance of the identity of species, the latter is a driving force that strongly participates in evolutionary and adaptive processes. HGT plays a key role in the spread of multiple genetic traits, more importantly the resistance to antibiotics, and participates actively in the successful adaptation of bacteria to new niches. In the bacterial kingdom, HGT is mainly mediated by different types of mobile genetic elements (MGE) that constitute the mobilome. Among the MGE, bacterial plasmids represent up to 20% of the commonly shared bacterial genes (the so-called plasmidome) that move through HGT processes.Among the ecological compartments which may be considered as important for the transfer of antibiotic resistance two environments with varied sources of antibiotic pollution were chosen to study:

• environment polluted by human medicine: untreated and treated mixed municipal and hospital wastewater, water bodies which are receivers of WWTPs,
• environment polluted by veterinary medicine: manure from the poultry, dairy and swine farms, soil from fields fertilized with manure from the farms, groundwater and crops from the same fields.

The aim of this study will be to isolate and characterize resistance plasmids in aforementioned samples. Moreover, the study will determine if there is a correlation between concentration of antibiotics, their transformation products (TPs) and the presence of resistance plasmids in analyzed environmants.

In 2015 the World Health Organization has adopted a global antimicrobial action plan, which underlines the need for an effective implementation of the "ONE HEALTH" approach. The "One Health" concept recognizes that human health and animal health are interdependent and bound to the health of the ecosystem in which they (co)exist. In fact, this interconnection favors the transmission of bacteria and other infectious agents, as well as the flow of genetic elements containing antibiotic resistance genes. Therefore, understanding the characteristics and behavior of microorganisms from the most diverse environmental niches is important to take actions in order to attenuate the emergence and dissemination of resistance. The “One Health” approach involves coordination among numerous international sectors and entities, including human and veterinary medicine, agriculture, environment, food and consumer. In order to meet these needs and integrating these efforts, this study will include establishing the link between the various elements of the environment which are under pressure of human activities and the food produced as well as human health in this environment. The main concern for the release of antibiotics into the environment is related to the development of antibiotic resistance genes and bacteria, which reduce the therapeutic potential against human and animal pathogens. A number of reservoir and habitats may be sites for emergence and maintenance of resistant microorganisms. These include hospitals, wastewater treatment plants (WWTPs), farms, aquaculture and habitats to which feces and urine from humans and animals are excreted. Among the ecological compartments which may be considered as important for the transfer of antibiotic resistance we chose to study two different groups of environmental samples:

• the environment where antibiotics originated from human treatment: untreated and treated mixed hospital and municipal wastewater, water bodies which are receivers of WWTPs effluents, WWTPs’workers,
• the environment where antibiotics originated from animals treatment: manure from the poultry, dairy and swine farms, soil from fields fertilized with manure from the farms, groundwater and crops from the same fields, farms’ workers.

The aim of this study will be estimation of the relationship between concentration of the most used beta-lactams antibiotics, their TPs and antibiotic resistance genes and antibiotic resistant bacteria in aforementioned samples. Moreover, the sequencing analysis will enable us to determine changes in types of microorganisms presented in studied microbiome associated with the presence of drugs and their TPs. Additionally, antibiotic resistance of the bacterial population inhabiting the upper respiratory tract of WWTPs' and farms' workers will be estimated. The final results will allow us to determine the degree of environmental pollution by drugs, ARGs, ARB and the risk to humans related to the presence of these micropollutants in the environment.

ARGs are often associated with mobile genetic elements (MGEs), and thus horizontal gene transfer (HGT) is a major concern in understanding the spread of antibiotic resistance and developing strategies for mitigation. The identification of hotspots of ARGs dissemination should rely on the detection of the common genetic structures hosting the ARGs (typically, mobile genetic elements) rather than the ARGs themselves. There are different types of mobile genetic elements, namely plasmids, transposons, bacteriophages, integrons. Integrons are of particular interest because: (i) are the simply elements involved in the mobility of gene cassettes, (ii) all have a common structure, (iii) can be associated to other mobile genetic elements and (iv) are particularly efficient in trapping ARGs.Among the ecological compartments which may be considered as important for the transfer of antibiotic resistance two environments with varied sources of antibiotic pollution will be chosen to study:

• environment polluted by human medicine: untreated and treated mixed hospital and municipal wastewater, water bodies which are receivers of WWTPs,
• environment polluted by veterinary medicine: manure from the poultry, dairy and swine farms, soil from fields fertilized with manure from the farms, groundwater and crops from the same fields.

 The aim of this study will be estimation of the relationship between concentration antibiotics, their transformation products (TPs) and integrase genes (intI1, intI2, intI3) in aforementioned samples. Moreover, the sequencing analysis will enable us to detect integron structures. By determination of structures of gene cassettes, information on antibiotic resistance and ARGs spread will be provided.

         Although there is a large variety of laboratory methods that can be used during the isolation of specific cell and tissue components, we still have problems in protein isolation with the right quality and quantity in order to obtain adequate results in the next stage of analysis. Electrophoretic studies are an important step in the analysis of cell and tissue proteome, and the degree of sample purification is of key importance here. The search for biological markers of reproductive processes is based on the analysis of both the qualitative and quantitative set of proteins isolated from spermatozoa, fluids or tissues of the male reproductive system. Choosing the optimal method of protein isolation is always difficult. The aim of the research will be to discover the relationship between the used isolation method and the quality and quantity of individual protein fractions in analyzed material.
         As a result of interdisciplinary activities combining research methods such as proteomic analysis of proteins from sperm, fluids and tissues of the reproductive system of selected animal species typical of the scientific discipline which is biology of reproduction and technological solutions using the phenomena of sound wave propagation in the liquid typical for the scientific discipline which is agricultural engineering is assumed to acquire new knowledge about the usefulness of ultrasounds to support the processes of protein isolation from the male reproductive system as well as purity and recovery rate of individual protein fractions from the initial preparation. Supporting ultrasound extraction may have a significant effect on solubility, diffusion, solvent penetration, analyte transport, extraction time and efficiency.

         It is generally agreed upon that hydrogen is an ultimate fuel of the future. This is because H2 is the only known ecologically clean fuel (its combustion leads to the formation of water molecule) with practically unlimited resources, as well as it has superior to other commonly known fuels heat of combustion (ca. 120-130 MJ/kg). The process of alkaline water electrolysis is one of the most promising technologies for the production of ultra-high purity hydrogen gas (powered e.g. by photovoltaic or wind energy sources), which is considered an ideal energy carrier for the renewable energy storage. The latter aspect is especially important with respect to rapid development of PEM (Proton Exchange Membrane), hydrogen/oxygen fuel cell technologies. This thesis aims at the development of advanced catalyst materials for the production of highly effective cathode (hydrogen generation) and anode (oxygen generation) electrodes, with optimized (minimized) overpotential parameters for a laboratory-scale alkaline water electrolyzer unit. Such-produced hydrogen and oxygen gases will then be properly purified and compressed in order to serve as fuel and oxidizing agent for a small size PEM-type (Ballard) fuel cell device. In this sense, the subject of water electrolysis becomes interdisciplinary science and technology, as it is closely related to a number of important areas of technical knowledge and industry, including: Environmental Management and Protection, Energy Generation and Storage (e.g. Energy Clusters), and Automotive Manufacturing.

The use of lignocellulosic biomass is important for the national bioeconomy. On agricultural land, mainly marginal, the number of plantations used for industrial and energy purposes increases. It is interesting how it will affect the biodiversity of rural areas and the entire landscape. Are there any effects on biodiversity within the plantations of perennial industrial crops ,including short rotation coppice (eg. willow, poplar) and other plants using to a production of lignocellulosic biomass (miscanthus, sida) and in their close surroundings? Preliminary research of biodiversity in and around willow plantations in Łężany and Samławki (a lakeland landscape) indicated that no negative effects on biodiversity within the plantations and in theirs surroundings. The preliminary researches was carried out in 2011-2013 (short period) and covered  mammals, birds, amphibians and some group of aquatic insects. For some invertebrates willow plantations cane serve as a temporary shelter. The planned research will use the island ecology model and the metapopulation model. Poisons (e.g. butterflies) and aquatic invertebrates will be investigated as indicator’s species. The field research will be carried out on the experiments owned by the Department of Plant Breeding and Seed Production, Faculty of Environmental Management and Agriculture. The laboratory research (identification of species) and statistical analysis  will be carried out on the Department of Ecology and Environmental Protection, Faculty of Biology and Biotechnology.

         Biomass has become the main renewable energy source (RES) in Poland and it accounted for 76% of all RES in 2015. Moreover, biomass is a raw material for the production of biogas and transport biofuels, which accounted for 5.5% and 6.7% of RES, respectively. The majority of biomass as energy and industrial feedstock is derived from forests. However, domestic policy has indicated an increased importance of agricultural biomass, including woody shrubs grown in short harvest rotations, such as willow, poplar and perennial herbaceous crops (PHC), e.g. cup plant, Virginia fanpetals. These plants could be grown on soils of poorer quality, unusable for growing crops for food or fodder. The area of land in Poland which meets the usability criteria for perennial energy and industrial crops, without providing competition or having a negative impact on the production of food and fodder, is estimated to be 1.6 million ha. In consequence, there are a number of opportunities in Poland for: (i) cultivation of perennial crops, (ii) utilization of soils of poor quality, and (iii) production of biomass as energy feedstock. Depending on the species and the harvest date, biomass of perennial plants can be used directly as a solid fuel or transformed into briquettes or pellets. It can also be used in biorefineries as feedstock for various bioproducts as well as liquid and gaseous biofuels. Currently, maize is the most frequently used substrate for the production of biogas in Poland, Germany and other EU countries. Development of biogas plants boosts RES production growth, it also stimulates the competition for maize as feedstock for fodder and food production. Therefore, cultivation of PHC creates opportunities: (i) to use soils of poor quality, (ii) it can be a source of biomass as a substrate or co-substrate for a biogas plant, i.e. it is a potential supplement and replacement for maize and (iii) it can also be a feedstock for other industrial purposes. Not much is known on the utilization of PHC for biogas production. Therefore, the aim of the study will to determine the biomass and biogas productivity and LCA analysis of biomass production of two PHC species. The planned research will be a combination of research in the field of Agronomy and Environmental Management and Protection. Biomass production and LCA will be implemented as part of research conducted in the Department of Plant Breeding and Seed Production (Faculty of Environmental Management and Agriculture). Research in the field of biogas efficiency will be carried out in cooperation with the Department of Environmental Engineering (Faculty of Environmental Sciences).

         The part of the economy that uses biomass, crops and residues from households, agriculture and food industry for the manufacturing of materials, chemicals, transportation fuels and energy is defined as biobased economy. The biobased economy consists of all options to produce food and non-food products and energy services from biomass. The biobased economy needs sustainable supply of biomass (including residues) for bioproduct generation and multiple uses. In this concept, no biomass should be used for energy generation unless other options have been considered of using it to produce higher value added products (hierarchy of biomass utilization). Therefore, the use of biomass residues from households for feeding insects for production of protein and fat is important for the national and world bioeconomy development. The planned research will be a combination of research in the field of agricultural and biological sciences. Biomass from agricultural residues, as a source of feed for selected insect species, will be provided from research carried out in the Department of Plant Breeding and Seed Production (Faculty of Environmental Management and Agriculture). Assessment of the suitability and growth rate of selected insect species, etc. will be carried out in cooperation of the Department of Plant Breeding and Seed Production and the Department of Ecology and Environmental Protection (Faculty of Biology and Biotechnology).

 The aim of the experiment will be to verify the scientific hypothesis indicating the possibility of increasing the efficiency of enzymatic hydrolysis of selected lignocellulose substrates used in the production of bioethanol of the second generation by repeated use of enzymes absorbed in the substrate. The prospect of restrictions of access to the main fossil energy sources forces to restrict their use and the search for renewable, environmentally friendly sources of energy. New, alternative and renewable energy sources are needed, but due to the demand for foodstuffs it is also necessary to stop using food raw materials for the production of biofuels. Currently the cheapestand the most commonly used for the production of biofuels is lignocellulosic biomass, which comes from grasses, energy crops, agricultural waste, waste from wood processing.

The problem that arises when using lignocellulosic biomass is its complex structure limiting access to saccharides used in the next stage of ethanol fermentation. The key to the commercial success of using lignocellulosic material is to find an efficient and economically justified pre-treatment method that facilitates further processes. It is also necessary to increase the efficiency of the enzymatic hydrolysis process to release as much as possible and use the available saccharides.

The first stage of experiments will include the use of various methods of pre-treatment of the raw material. In the next stage, these raw materials will be subjected to enzymatic hydrolysis using various enzymatic preparations, e.g. cellulolytic enzymes, pectinolytic enzymes. The effects of hydrolysis will be determined using colorimetric methods and using high-performance liquid chromatography.

The possibility of re-use of the insoluble solid residue after enzymatic hydrolysis will also be explored to increase enzyme hydrolysis efficiency using enzyme recycling. Individual stages of experiments will be illustrated by the use of electron microscopy and atomic force microscopy. The efficiency and productivity of processes, quality and method of using solid biomass residues, selection of enzyme doses will be assessed. The use of statistical methods to optimize the process is envisaged.

The supervisor of the biotechnology discipline will be responsible for aspects related to the selection of enzymes, evaluation of their properties, catalysis effect, evaluation of enzyme sorption and the possibility of immobilization, stability of activity, evaluation of synergistic effects between enzymes, and recycling, while supervisor of the discipline of food technology and human nutrition for aspects related to the evaluation of substrates: characterization of the residue after hydrolysis and ethanol fermentation, analysis of reaction products, the possibility of valorisation of by-products.

Continuously stricter EU legal regulations regarding animal feeds, encourage searching for new solutions that help maintain the health and productivity of poultry. Increasing awareness and consumer demands direct the attention of farmers towards herbs and natural feed additives. They are finding increasingly application in animal nutrition.. Various herbal mixtures are added to feed to improve the health and productivity of animals. Herbs can be anti-bacterial, anti-inflammatory, antioxidant, etc., they can also reduce the stress associated with pre-slaughter period. Various biologically active compounds - phytobiotics, such as alkaloids, tannins, flavonoids, glucosides, saponins, terpenes, etc. - are responsible for these herbal activities.  However, high demand for herbs in the food and medical industry, makes them quite expensive. Therefore, it seems reasonable to look for alternative sources of phytobotics. They may be by-products of the food and herbal industry, such as pomace after the production of juices or parts of herbal plants, unfit for human consumption. New sources of phytobiotics can also be found in plants less popular or recently introduced in the EU, such as Gumi (Elaeagnus multiflora) or actinidia (Actinidia arguta). Consumers expect healthy and natural food whose production is part of the bio-economy canon. The increasing pollution of the environment with antibiotics and the growing resistance of bacteria also speaks for the use of phytobotics in animal nutrition. All these factors lead to the search for novel sources of phytobotics for poultry.

        Platform substances are a group of chemicals that serve as starting materials for the production of different valuable chemicals. Currently, petroleum-based platform chemicals dominate in the industry. Therefore, to decrease both the environmental pollution and the use of fossil fuels, the idea of the production of platform chemicals from renewable source has been gaining interest. In this study, distillery residue that contains high amount of organic matter will account for a sustainable feedstock for the production of selected platform chemicals using a biorefinery concept. Based on the literature study and the analysis of distillery residue, the potential chemicals that can be produced from this waste source will be first selected. Next, the methods of environmental engineering, biotechnology and chemical technology will be used for the optimization of the production of these platform substances, including the searching for a method of substrate pretreatment. The goal of the study will be to investigate the effective technological solution for the production of the selected platform chemicals from distillery residue. The combination of the interdisciplinary methodologies will allow achieving the assumed research goals.

         With the increasing production of tvarogs (acid coagulated cheese) and even greek yoghurts, there is a need for optimized acid whey processing. Many dairy plants, especially small and medium, suffer with lack of technology to process acid whey into valuable food products. Additional problem create the necessity to manage the wastes produced during whey processing, i.e. UF permeate.
The research will focus on identification of optimal parameters of acid whey treatment. The aim of this project is to develop several technologies to process acid whey into added value dairy based ingredients. Several schemes for whey treatment will be developed depending on the size of the dairy processing plant. The threshold values of critical ingredients (i.e. dry matter, protein) will be determined in order to calculate the economic feasibility of the process. The technological determinants (i.e. type of equipment, process parameters) of acid whey processing will be selected. The technology to concentrate and/or fractionate acid whey ingredients will be developed. It is necessary to choose the most relevant sequence of fractionation methods and conditions applied during processing to produce tailored dairy products with desired functional properties. The by-products of acid whey processing that cannot be used for the production of food products, will be utilized to produce i.e. biofuels. The proposed project guarantees the complex and interdisciplinary approach to acid whey management in dairy plants.