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Contaminants in dust and risks to human health
Šafaříková, Linda ; Erban, Tomáš (advisor) ; Hubert, Jan (referee)
This bachelor's thesis addresses the impact of contaminant occurrence in dust and how these contaminants can affect human health. The focus is primarily on allergens and potential allergens. The theoretical part introduces major allergens and methods for their separation and identification. Identifying the spectrum of allergens in the environment is crucial for discerning the causes of allergic reactions. Some methods based on antibody detection, such as Western blot or ELISA, enable the identification of specific individual allergens in samples. In cases where antibodies are not available, alternative approaches are necessary. An appropriate approach is proteomic analysis, which holds significant potential for identifying the entire spectrum of allergens and other proteins in dust. Especially, high-capacity proteomic approaches enable the identification and characterization of multiple proteins, including allergens, simultaneously, providing a comprehensive view of dust sample composition. In the practical part, high-capacity proteomic methods were employed. A combination of gel-based and gel-free analysis of overall heterogeneous dust samples and band analysis after 1D-E separation was used for household dust analysis. Samples were collected from beneath the bed and behind the wardrobe, where the...
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Methodology for assessing the effect of sublethal doses of pesticides on solitary bees employing the OMICs approach
Erban, Tomáš ; Shcherbachenko, Elena ; Šlachta, Martin ; Cudlín, Pavel ; Chalupníková, Julie ; Halešová, Taťána ; Tomešová, Daniela ; Václavíková, Marta ; Votavová, Alena
Pollinators are potentially exposed to a number of pesticides in the environment. The most important group of hazardous substances comes from the category of plant protection products (PPPs), which are applied not only by farmers but also by small gardeners. Each PPP including the active substance and formulation undergoes a very detailed and thorough evaluation, which is in the European Union (EU) based on Regulation (EC) No 1107/2009 of the European Parliament and of the Council. Although the negative effects of PPPs on the environment and non-target organisms are minimized with proper use, we cannot rule out possible hidden effects, especially those of a sublethal nature. Modern analytical approaches, such as the application of high-throughput methods, help to identify hidden side effects. This methodology is focused on the evaluation of the effect of sublethal doses of pesticides on solitary bees employing the OMICs approach. In the risk assessment of pesticides, the honey bee has long been preferred as a model species of pollinator, while solitary bees stay in the background. The methodology contains innovative aspects for experiments, such as a modified isolator, preparation of plants for exposure, the timing of the experiment, monitoring of active substance distribution or proteomic analysis of solitary bees to identify hidden effects of the test substance or its formulation. In this methodology, the model of the solitary bee, red mason bee (Osmia bicornis), is considered. However, the methodology is also applicable to other, especially related species of solitary bees. The methodology can be used in areas of state administration, private laboratories and research activities in the assessment of environmental risks of pesticides on solitary bees. The methodological procedure can confirm or even eliminate environmental risks when registering new products or reevaluating existing products. This methodology therefore has the potential for use in testing new substances intended for plant protection before their registration. In an exemplary embodiment, the effect of acetamiprid in a Careo sticks formulation was tested. The distribution of acetamiprid in the soil substrate and the plant was evaluated. The hazardous metabolite acetamiprid IM-2-1 was identified in plant tissues. However, proteomic analysis indicated a very low risk of acetamiprid for O. bicornis. This result is in agreement with the fact that acetamiprid has been assessed by EFSA to be a low risk for bees and is registered in the EU until 28 February 2033.
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A method for identification of foreign amylases in honey
Erban, Tomáš ; Shcherbachenko, Elena ; Talacko, Pavel ; Harant, Karel
Honey is a unique natural product. Honey has been used as a sweet and delicious foodstuff since ancient times. However, it is also valued for its multifaceted currative properties. Unfortunately, honey is one of the most adulterated foods. Nothing may be added to or modified from the honey. Honey also needs to be handled with care. Honey quality can negatively be affected by the way of processing such as heating and storage. Despite great progress in analytical methods, it is not possible to prove all adulterated honeys. Some methods of adulteration are quite sophisticated. Therefore, it is necessary find new approaches and methods for identification of honey adulteration. To be sold, honey must comply with internationally valid standards, which are also valid at national levels with possible minor modifications. One of the important parameters for honey is the level of diastase or amylase enzyme activity, which is a recognized indicator of the freshness and quality of honey. Lower diastase activity below the established level may indicate old honey, but it may also be the result of careless handling of honey. Last but not least, diastase activity may be reduced due to adulteration of honey such as its dilution with sugar substitutes. It is possible that amylase activity can be artificially adjusted by the addition of enzymes. Honey adulterated by the artificially added amylase meets the legislative requirements for placing honey on the market, but this violates the rules laid down by law. The methods used so far have not made it possible to prove this way of honey adulteration. Therefore, this methodology focuses on the identification of foreign amylases that may occur in honey. The methodology enables to identify practically any foreign amylase in honey by bottom-up shotgun proteomic approach. Based on the obtained results with specific peptides can be further used for the development of a targeted method for the identification of foreign amylases.
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Measures to support populations of solitary bees in agroecosystems
Šlachta, Martin ; Erban, T. ; Votavová, A. ; Cudlín, Ondřej ; Cudlín, Pavel ; Halešová, T.
Solitary bees participate in the pollination of crops. According to our field\nsurvey, this involves 88 species of solitary bees in spring, mainly mining bees\n(Andrenidae) and sweat bees (Halictidae) nesting in the soil. The species\nnesting above soil were present less due to the lack of nesting opportunities\nand generally summer nesting activity. Among them, mason bees (Osmia sp.),\nnesting in spring, are used abroad for the pollination of fruit trees. The aim of\nthis methodology is to provide farmers with instructions for the use of mason\nbees for pollination of orchards by means of artificial nesting sites. We propose\na convenient construction of nesting sites and the methodology for rearing the\nEuropean orchard bee, Osmia cornuta (Latreille, 1805), which is, due to its earlyspring nesting activity, a suitable substitute for the currently used buff-tailed\nbumblebee. Breeding can be supplemented by the red mason bee, O. bicornis\n(Linnaeus, 1758), with later nesting activity. The book contains also suggested\nmeasures to support solitary bees in the landscape and new research data on\nthe diversity of solitary bees and pollen preferences of mason bees.
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Methodology for assessing the effect of sublethal doses of pesticides on solitary bees\nemploying the OMICs approach
Erban, T. ; Shcherbachenko, E. ; Šlachta, Martin ; Cudlín, Pavel ; Chalupníková, J. ; Halešová, T. ; Tomešová, D. ; Václavíková, M. ; Votavová, A.
Pollinators are potentially exposed to a number of pesticides in the environment. The\nmost important group of hazardous substances comes from the category of plant protection\nproducts (PPPs), which are applied not only by farmers but also by small gardeners. Each PPP\nincluding the active substance and formulation undergoes a very detailed and thorough evaluation,\nwhich is in the European Union (EU) based on Regulation (EC) No 1107/2009 of the European\nParliament and of the Council. Although the negative effects of PPPs on the environment and nontarget organisms are minimized with proper use, we cannot rule out possible hidden effects,\nespecially those of a sublethal nature. Modern analytical approaches, such as the application of\nhigh-throughput methods, help to identify hidden side effects. This methodology is focused on the\nevaluation of the effect of sublethal doses of pesticides on solitary bees employing the OMICs\napproach. In the risk assessment of pesticides, the honey bee has long been preferred as a model\nspecies of pollinator, while solitary bees stay in the background. The methodology contains\ninnovative aspects for experiments, such as a modified isolator, preparation of plants for exposure,\nthe timing of the experiment, monitoring of active substance distribution or proteomic analysis of\nsolitary bees to identify hidden effects of the test substance or its formulation. In this methodology,\nthe model of the solitary bee, red mason bee (Osmia bicornis), is considered. However, the\nmethodology is also applicable to other, especially related species of solitary bees. The\nmethodology can be used in areas of state administration, private laboratories and research\nactivities in the assessment of environmental risks of pesticides on solitary bees. The\nmethodological procedure can confirm or even eliminate environmental risks when registering new\nproducts or reevaluating existing products. This methodology therefore has the potential for use in\ntesting new substances intended for plant protection before their registration. In an exemplary\nembodiment, the effect of acetamiprid in a Careo sticks formulation was tested. The distribution of\nacetamiprid in the soil substrate and the plant was evaluated. The hazardous metabolite\nacetamiprid IM-2-1 was identified in plant tissues. However, proteomic analysis indicated a very\nlow risk of acetamiprid for O. bicornis. This result is in agreement with the fact that acetamiprid\nhas been assessed by EFSA to be a low risk for bees and is registered in the EU until 28 February\n2033.
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Secondary metabolite production in actinomycetes izolated from potato rhizosphere.
Komžák, Ondřej ; Bosáková, Zuzana (advisor) ; Erban, Tomáš (referee)
This bachelor thesis is focused on searching for secondary metabolite producers suppressing Streptomyces scabies (synonym scabiei), which is a causative agent of common potato scab affecting some species of agricultural crops. By the 16S rRNA gene sequencing strains of bacteria from potato rhizosphere were identified and strains effectively suppressing Streptomyces scabies were determined by a simple cultivation test. The work on this project will continue by focusing on identification of those secondary metabolites by HPLC.
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Study of physiology and searching of proteomic tools useful in suppression and detection of Varroa destructor
Holenková, Martina ; Erban, Tomáš (advisor) ; Titěra, Dalibor (referee)
This work is focused on the study of physiology and proteome of the the mite Varroa destructor and on comparison with the honeybee Apis mellifera. Varroa is currently a major problem for beekeeping, because infects most of the colonies. The control of the mite can not be done without residues both in the hive and for example in the honey or other resources used by the man. Another problem can also be the simultaneously discussed issue of the connection with the Colony Collapse Disorder. The internal anatomy of V. destructor was studied by using paraffin histology. On histological sections stained using hematoxylin and PAS was possible to observe the mite digestive system, but also egg, ovaries or brain. The primary aim of this study was to identify the proteins of mite V. destructor and bee A. mellifera as a host of this parasite. For the electrophoretic separation was used two- dimensional gel electrophoresis, where the second dimension was carried out using 12% and 15% SDS-PAGE. The most abundant spots were selected for analysis using MALDI TOF/TOF mass spectrometry. The most abundant protein identified in samples of V. destructor was hexamerin, arginine kinase or hemelipoglycoprotein precursor. Hexamerins were also identified as the major proteins in the pupae samples. On the contrary the main...
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Nutritional biology of synanthropic mites (Acari: Acaridida)
Erban, Tomáš
Ph.D. THESIS TITLE Nutritional Biology of Synanthropic Mites (Acari: Acaridida) ABSTRACT Several attempts to describe the nutritional biology of acaridid mites were undertaken, however full understanding of these processes remains incomplete. The objective of this Ph.D. thesis was to expand our knowledge concerning digestive physiology of stored product and house dust mites and to apply this knowledge to their nutritional biology. The research approach adopted in this Ph.D. thesis includes in vitro characterization of enzymatic activity in whole mite extracts (WME) and spent growth medium extracts (SGME), evaluation of the enzyme activities with respect to the gut physiological pH, enzyme inhibition experiments, in vivo localization of enzyme activities in the mite gut, determination of effects of nutrient or antifeedant additives in experimental diets on mite population growth and determination of the feeding preferences of synanthropic mites as assessed by in vitro and in vivo analyses. The gut contents of twelve species of synanthropic acaridid mites were determined to be within a pH range of 4 to 7 and showed a pH gradient from the anterior to the posterior midgut. The pH in digestive tract of synanthropic acaridid mites corresponds to the activity of proteases, α-glucosidases, α-amylases and...
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