Conference MendelNet 2015      11 and 12 November, 2015
Contributions

List of Contributions Conference MendelNet 2015

Animal Biology

  • RAPID IDENTIFICATION OF BACTERIA BY BIOBARCODE ASSAY
    CIHALOVA KRISTYNA, DOSTALOVA SIMONA, HEGEROVA DAGMAR, SKALICKOVA SYLVIE, VACULOVICOVA MARKETA, KIZEK RENE, KOPEL PAVEL
    view abstract fulltext kriki.cihalova@seznam.cz

    Abstract: Presence of bacteria with antibiotic resistance is becoming a very large problem throughout the world. Methicillin-resistant Staphylococcus aureus (MRSA) is a dangerous pathogen resistant to β-lactam antibiotics with biofilm-formation ability. Because of an increasing resistance of bacterial species to ATBs, it is necessary to develop new methods for rapid identification of bacteria. Biobarcode assay provides a rapid detection of the antigen presence in the sample. Detection is based on the antibody-antigen interaction. The antibodies were bound to magnetic and non-magnetic particles. Next, immunoglobulin G (IgG) was bounded to magnetic particles. The non-magnetic particles were bound with anti-plasminogen antibody that is a specific antigen for MRSA as well as 20 bp oligonucleotides for detection. The first step involved determination of binding capacity of antibodies for different bacteria by ELISA. The IgG were able to bind 4.6 .104 ± 8% CFU .ml-1 of MRSA, Escherichia coli and Proteus mirabilis and the anti-plasminogen antibody (anti-Pls) was specific for MRSA only with the binding capacity of 5 .103 CFU .ml-1. After binding of antibodies to particles, the bacterial strain MRSA was captured by these antibody-modified particles and the detection oligonucleotide was released and determined by electrochemical method. The results suggest that the IgG is non-specific for MRSA while specificity of the anti-plasmin antibody for MRSA was confirmed. In this study, we developed a method for rapid detection of MRSA in the pooled sample.


     
  • MALDI-TOF MASS SPECTROMETRY IMAGING OF METALLOTHIONEIN IN CHICKEN EMBRYO
    GURAN ROMAN, BLAZKOVA IVA, KOMINKOVA MARKETA, ZITKA ONDREJ, KIZEK RENE, ADAM VOJTECH
    view abstract fulltext r.guran@email.cz

    In last decades the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF MSI) has become an outstanding tool for detecting spatial distribution of different biomarkers in a variety of tissue samples. It utilizes the benefits of MALDI-TOF technique, which are rapid measurements of all mass spectra in a wide mass range and detection of analytes molecular weights. Moreover, the in situ identification of targeted biomarkers can be performed too. In our study, we focused on detection of metallothionein (MT) in chicken embryo. Metallothioneins are low-molecular weight proteins connected with cancer development and protection of organism against environmental pollution. Their main functions are detoxification of heavy metals, maintaining ion homeostasis and protection against the oxidative stress. According to our knowledge, nobody has done MALDI-TOF MSI of MT so far. Therefore, we have selected MT as our studied analyte not only because of this fact but also because a part of team IGA project is aimed on MT.


     
  • POLYMORPHISMS IN PLASMA MEMBRANE CALCIUM-TRANSPORTING ATPASE 1 (ATP2B1) GENE IN HENS
    HORECKA ELISKA, HORECKY CENEK, KOVARIKOVA LENKA, MUSILOVA ANNA, KNOLL ALES, PAVLIK ALES
    view abstract fulltext eliska.horecka@mendelu.cz

    Abstract: Bone fragility in caged laying hens is a severe welfare problem. This fragility has been attributed to osteoporosis, which etiology is multifactorial in birds, as well as in humans, with genetic, environmental, and nutritional components. Plasma membrane calcium-transporting ATPase 1 gene (ATP2B1) is in hens located on chromosome 1, region 43 273 706 – 43 305 815 bp. This gene has 21 exons, three of them were genotyped. In this study we genotyped 110 hens of ISA BROWN hybrids. Genotypes of ATP2B1 gene were determined using PCR-RFLP in exons 10 and 12. Genotypes of two SNPs in exon 8 were determined using sequencing. In our group of animals, only allele without deletion in exon 10 and only allele A in exon 12 was found. In exon 8 subsequent genotypes were detected: in C61Tlocus CC and TT; in C80T locus CC, CT and TT.


     
  • EXTENSION OF THE MICROSATELLITE PANEL FOR DIVERSITY STUDIES IN THE EQUINE Ly49 GENES REGION
    HORECKY CENEK, HORECKA ELISKA, FUTAS JAN, JANOVA EVA, HORIN PETR, KNOLL ALES
    view abstract fulltext cool.czenda@gmail.com

    Abstract: The genetic variability and different expression of genes for receptors underlies functional variability of individual natural killer cells (NK). Like in the mouse model, the Ly49 receptors on the horse NK cells are believed to bind MHC class I molecules of target cells. Six Ly49 genes constitute a gene family located on the horse chromosome 6, between 38 200 Kbp – 38 520 Kbp. Immune-response genes represent a functionally important region of the vertebrate genome subject to selection pressure. NK cells are involved in the antigen recognition process through their highly variable receptors. Our work may contribute to better estimating the genetic diversity of this functionally important region. In this work, we identified and genotyped three new polymorphic microsatellite markers that expand the original panel of microsatellites and are located in the Ly49 region. This methodology will be used for assessment genetic diversity and association analyses with selected diseases of horses.


     
  • EFFECT OF EXTRUDED AND NO EXTRUDED SOYBEANS SUPPLEMENTS IN FODDER ON ANTIOXIDANT LIVER ACTIVITY IN BROILERS
    KABOURKOVA ELISKA, LICHOVNIKOVA MARTINA, ADAM VOJTECH
    view abstract fulltext eliska.kabourkova@mendelu.cz

    Abstract: The aim of the trial was to evaluate the effect of extruded and no extruded soybeans substitute of soybean meal and soybean oil in fodder on the broiler liver antioxidant activity. Metallothionein was used to measure oxidative stress in liver. There were used the substitute of 15% soybean meal and soybean oil by extruded soybeans, the substitute of 10% soybean meal and soybean oil by extruded soybeans and  the substitute of 10% soybean meal and soybean oil by no extruded soybeans of a feed ration from 10 to 35 day of age. The trial was carried out on the 156 female chickens Ross 308. The chickens were kept in the double-floor cage technology. All of them were fed by the same complete feed mixture BR1 (Broiler No. 1) for the first 10 days. After 10 days of age, the birds were randomly allocated to 12 cages in both tiers corresponding to 4 dietary treatments with three replicates of each treatment. Each group comprised 3 cages containing 13 broilers each. The dietary treatments included a control diet and the diets containing 15% extruded soybean substitute, 10% extruded soybean substitute and 10% no extruded soybean substitute of soybean meal and soybean oil with the other components remaining the same as in the control diet. The level of the metallothionein content was determined in samples obtained from 35 days old animals by adsorptive transfer stripping differential pulse voltammetry. The fodder substitute of the extruded and no extruded soybeans had significant (P<0.05) effect on the level of the metallothionein level in the liver of the broiler chickens.

    The highest level of the metallothionein was measured in the 10% substitute of extruded soybeans. The lowest level of metallothionein level was measured in the control group. The difference found was significant (P<0.05).


     
  • EFFECT OF PC-3 PROSTATE CANCER CELL LINE SUPERNATANT ON APOPTOSIS IN MACROPHAGES
    MAZALOVA LENKA, SLADEK ZBYSEK
    view abstract fulltext lenka.mazalova@mendelu.cz

    Abstract: Particular types of cell death, for example apoptosis, play important role in metastatic processes. Apoptosis is programmed cell death, which is characterised by specific morphological changes. In this study we aimed on topic of affecting of supernatant from prostate cancer cell line PC-3 on a healthy cells of immune system, macrophages concretely. Peripheral blood monocytes were cultivated for 7 days to macrophages. Macrophages were stimulated for 24 hours by lipopolysaccharide (LPS) and cultivated with supernatant for another 24 hours. Preparations were prepared and analysed by light microscopy. Macrophages with normal morphology show the largest rate in our experiment. In samples macrophages + supernatant there was an increase mainly in late stage of apoptosis. A lot of observed cells showed feature of rupture and spillage of cell contents into the microenvironment. In samples macrophages + supernatant + LPS there is a noticeable decline in the incidence of normal morphology compared to the control. This means that effect of cancer cells and their supernatants on macrophages can induce apoptosis on macrophages and thus prevent proper course of immune responses.


     
  • EFFECT OF MELITTIN ON INFLUENZA-INFECTED CHICKEN EMBRYOS
    MICHALEK PETR, ZITKA ONDREJ, KREJCOVA LUDMILA, PRIDAL ANTONIN, KOMINKOVA MARKETA, GURAN ROMAN, MILOSAVLJEVIC VEDRAN, KOPEL PAVEL, HEGER ZBYNEK, ADAM VOJTECH, KIZEK RENE
    view abstract fulltext petrmichalek85@gmail.com

    Abstract: Antimicrobial peptides are peptides isolated from a wide range of organisms that exert microbicidal activity against a wide spectrum of targets, including bacteria, viruses, fungi, and parasites. These peptides are aimed directly on the phospholipid bilayer and do not target on the cellular or metabolic activities of the cells as antibiotics and other drugs do. But thanks to many vital proteins associated with the membrane, these peptides can also influence these structures and therefore facilitate and accelerate their death. Melittin is a well-characterized pore-forming lytic amphiphilic peptide (consisting of 26 amino acids) found in bee venom. The amphiphilic property of this peptide makes it water-soluble and yet it spontaneously associates with natural and artificial membranes. This integration leads to the distortion and permeabilization of the membrane. In this study melittin has been utilized to study antiviral properties against influenza A virus. Melittin was used in the mixture with the influenza A virus and together were inoculated into chicken embryo. Living conditions were monitored during infection. After death of chicken, biochemistry of allantoic fluid was performed.


     
  • THE EFFECT OF LIGHT INTENSITY UPON HEMATOLOGICAL PARAMETERS OF BROWN RATS’ BLOOD
    PECINOVA HANA, BROUSKOVA ELISKA, KVASNOVSKY MICHAL, HODULIKOVA LUCIA, HAVLICEK ZDENEK
    view abstract fulltext hana.pecinova@mendelu.cz

    Abstract: The main idea of this topic is to assess the effect of light intensity upon selected elements of animals’ blood (brown rat) as the influence of day and night cycles, or any other variations in light intensity affecting an organism have been proven by several studies. Three groups of animals have been observed in terms of an impact of varying light intensity (increased intensity, natural intensity and darkness). Obtained blood samples served for determining the amount of erythrocytes and hemoglobin, share of hematocrit and the number of leukocytes. Regarding erythrocytes, no significant increase that could be caused by heightened light intensity has been noticed. On the other hand, zero intensity has reduced the amount of erythrocytes down to 7.09 T·l-1 (compared to 8.29 T·l-1 when the intensity level got higher). The highest hemoglobin levels (172.68 g·l-1) as well as the amount of leukocytes (9.7 g·l-1) have been observed upon the control group. Heightened light intensity has not taken any effect on increased levels of blood parameters, although all observed parameters went down as a result of total lack of light (i.e. darkness).


     
  • DETECTION OF TET GENES IN TETRACYCLINE RESISTANT MASTITIS PATHOGENS
    PYATOV VLADIMIR
    view abstract pyatov.vladimir@gmail.com

    The abundant use of antibiotics for the treatment of human and animal diseases as well as for
    prophylaxis has contributed to the spread of antimicrobial resistance genes among bacteria.
    This leads to microorganisms being non-sensitive to treatment with the corresponding
    antibiotics. Tetracycline belongs to the broad-spectrum antibiotics and is widely used in
    animal husbandry.
    The aim of this study was to evaluate the frequency of different tetracycline resistance genes
    (tet genes) in pathogens causing mastitis. Five of the most common pathogens, S.uberis,
    S.aureus, E.coli, S.agalactiae and S.dysgalactiae were chosen and a total of 30 samples was
    analysed in order to investigate the abundance and correlation of various tet genes.


     
  • STEAROYL-COA DESATURASE GENE AND HIS ASSOCIATION WITH FATTY ACIDS IN BEEF
    SCHMIDTOVA ANNA, KNOLL ALES
    view abstract fulltext anna.schmidtova@mendelu.cz

    Abstract: The quality of meat in cattle is influenced by many genes; one of them is SCD1 gene (stearoyl-CoA desaturase). This gene is associated with composition of fatty acids in meat and milk. In this study, the total of 260 bulls of Czech Fleckvieh breed were genotyped using the PCR-RFLP method. The frequencies of alleles and genotypes were determined in this population and the association analysis between fatty acids in fat extracted from musculus longissimus dorsi and genotypes was performed. Statistically significant (p˂0.0001) association between genotypes and myristoleic acid (C14:1) was found. CC genotype had higher median value. No other associations were found.


     
  • EFFECTS OF PROBIOTIC ON MORPHOLOGICAL CHANGES IN PORCINE MACROPHAGES DURING IN VITRO CULTIVATION
    SUSTROVA TEREZA, LEVA LENKA, ONDRACKOVA PETRA, KOLAROVA MIROSLAVA, SLADEK ZBYSEK
    view abstract fulltext tereza.sustrova@mendelu.cz

    Abstract: Nowadays, Bifidobacterium bifidum, Lactobacillus rhamnosus and Enterococcus faecium are frequently used probiotics in porcine nutrition. The probiotics-immunobiotics positively influence function of gastrointestinal tract and can also modulate function of immune system. The probiotics interact with immune cells as macrophages, neutrophils, dendritic cells or other immune cells and stimulate them to produce cytokines or to enhance phagocytosis. The aim of this study was to determine whether interactions of probiotics with porcine monocyte derived macrophages (MDMF) lead to structural changes of these cells during in vitro cultivation. We used the light microscopy and our findings suggest that probiotics affected the structural changes of porcine MDMF. Part of MDMF underwent apoptosis or necrosis and it was described the different stadia leading to the cell death. In some MDMF numerous vacuoles are accumulated in cytoplasm. The most pronounced structural changes of MDMF were caused by Enterococcus faecium. Finally, interactions of probiotics with MDMF were associated with phagocytosis all used probiotics.


     
  • THE EFFECT OF PROBIOTICS ON THE VIABILITY OF THE PORCINE AND HUMAN MONOCYTES
    VEJRYCHOVA SARKA, SUSTROVA TEREZA, SLADEK ZBYSEK
    view abstract fulltext sarka.vejrychova@seznam.cz

    Abstract: The aim of this study was evaluate the effect of probiotics on the viability of the cells of immune system. For experiment we have chosen the monocytes, which were isolated from porcine and human blood. The population of monocytes was cultivated in vitro conditions with probiotics strains such as Bifidobacterium bifidum, Lactobacillus rhamnosus and Enterococcus faecium. The monocytes were incubated without (control sample) or with probiotics for 2 and 4 hours. The percentage of apoptosis and necrosis of monocytes was analysed by flow cytometry. The results have shown statistically significant differences in proportion of porcine apoptotic monocytes cultivated with Bifidobacterium bifidum. Enterococcus faecium showed statistically significant effect on necrosis of porcine monocytes. The statistically significant differences in proportion of human apoptotic monocytes were observed in cultivation with Lactobacillus rhamnosus and Enterococcus faecium andthe highest percentage of necrotic cells was seen in human monocytes cultivated with Bifidobacterium bifidum. It is obvious that these selected strains of probiotics had immunomodulatory effect on immune cells and induced apoptosis and necrosis of porcine and human monocytes in vitro condition.


     
  • DISTRIBUTION OF MERCURY IN TISSUES OF THE COMMON CARP (CYPRINUS CARPIO L.)
    VICAROVA PETRA, PELCOVA PAVLINA, KLECKEROVA ANDREA, MARES JAN, KOPP RADOVAN, POSTULKOVA EVA, DOCEKALOVA HANA
    view abstract fulltext petra.vicarova@mendelu.cz

    Abstract: The aim of the experiment was to determine the distribution of mercury in ten selected tissues (muscle, skin, fish scales, biliary vesicle, brain, eyes, kidneys, spleen, liver and gills) of common carp (Cyprinus carpio L.). Carp fingerlings weighed 47.67 ± 4.61 g. Carps were exposed to increasing concentrations of mercury (0 µg.l-1 (control), 0.5 µg.l-1, 1.5 µg.l-1 and 3.0 µg.l-1) in fish tanks for 14 days. The concentrations of mercury in fish tanks were continuously monitored and in case of a change they were adjusted to an acceptable value. The fish were not fed during the experiment and mercury got accumulated in fish tissues from fish tank water only. Five fish were collected on the 0th, 4th, 9th and 14th day of experiment from each concentration for the analysis of total mercury content in selected tissues. Total mercury content in water and in selected tissues was determined by the atomic absorption spectrometer AMA 254. The increase of mercury in all tested tissues was not observed in the control group during the 14-day experiment. The time linear increase of mercury content was observed in the muscles, skin, fish scales, biliary vesicle, eyes, kidneys, spleen and gills in all three mercury concentrations under testing. The lowest mercury concentrations were determined in the control group in the range of 0.004 ̶ 0.052 mg.kg-1. Compared to this group, the highest concentration of mercury was found in kidneys (for fish tank with 0.5 μg.l-1 the mercury concentration was 1.405 ± 0.300 mg.kg-1, for fish tank with 1.5 μg.l-1 the mercury concentration was  5.537 ± 0.027 mg.kg-1 and for fish tank with 3.0 μg.l-1 the mercury concentration was 25.209 ± 2.152 mg.kg-1 on day 14 of the experiment).