Is<Emphasis Type="Italic">Pseudobutyrivibrio xylanivorans</Emphasis> strain Mz5<Superscript>T</Superscript> suitable as a probiotic? An<Emphasis Type="Italic">in Vitro</Emphasis> study

Rumen bacterium Pseudobutyrivibrio xylanivorans strain Mz5T possessed a potent xylanolytic enzyme system consisting of at least 7 different xylan hydrolases with molar mass 27-145 kDa. Three of them were successfully isolated in active native form. This strain produced butyrate and lactate on different saccharides. cis-9, trans-11-Conjugated linoleic acid was also detected in the culture medium. Bacteriocin-like inhibitory substances of Mz5T were active against some strains of rumen bacteria and against selected Salmonella and E. coli isolates from poultry meat. The strain Mz5T retained viability and xylanolytic activity also under not fully anaerobic conditions; its cells attached to the Caco-2 cells so that its successful association with gut epithelial cells may be expected. These in vitro results confirmed several probiotic traits of the isolate Mz5T and justified further in vivo experiments to test its ability to improve animal health and performance.     

Recombinant single-chain Fv antibodies that recognize the p25 protein of the Maedi-Visna virus

Single chain Fv (scFv) antibodies (generated by phage display technology, molecules representing new and efficient tools in the research and diagnostics of infectious diseases) against the capsid protein (p25) of Maedi-Visna virus were selected. Several clones of p25 specific scFv antibodies were identified; one of them was expressed as a soluble scFv molecule, purified by immobilized metal-affinity chromatography and further characterized by sequencing and determination of the kinetic equilibrium association constant. Sequence analysis showed that the rearranged VL and VH domains of the analyzed scFv clone used sequences from the VL3 family (germline DPL16/VL3.1) and VH1 family (germline VH20), respectively. The kinetic equilibrium association constant was determined as KA = 1.12 +/- 0.52 L/mumol.     

Thank You from the Tuček Family

Note

Thank You from the Tuek Family     

Random Fields Approach to the Study of DNA Chains

We apply the random field theory tothe study of DNA chains which we assume tobe trajectories of a stochastic process. Weconstruct statistical potential betweennucleotides corresponding to theprobabilities of those trajectories thatcan be obtained from the DNA data basecontaining millions of sequences. It turnsout that this potential has aninterpretation in terms of quantitiesnaturally arrived at during the study ofevolution of species i.e. probabilities ofmutations of codons. Making use of recentlyperformed statistical investigations of DNAwe show that this potential has differentqualitative properties in coding andnoncoding parts of genes. We apply ourmodel to data for various organisms andobtain a good agreement with the resultsjust presented in the literature. We alsoargue that the coding/noncoding boundariescan corresponds to jumps of the potential.     

Antigenotoxic potential of glucomannan on four model test systems

Antimutagenic, anticlastogenic, and bioprotective effect of polysaccharide glucomannan (GM) isolated fromCandida utilis was evaluated in four model test systems. The antimutagenic effect of GM against 9-aminoacridine (9-AA)- and sodium azide (NaN₃)-induced mutagenicity was revealed in theSalmonella typhimurium strains TA97 and TA100, respectively. GM showed anticlastogenic effect against N-nitroso-N-methylurea (NMU) induced chromosome aberrations in theVicia sativa assay. The bioprotective effect of GM co-treated with methyl-methane-sulphonate (MMS) was also established inChlamydomonas reinhardtii repair deficient strainsuvs10 anduvs14. The statistically significant antimutagenic potential of GM was not proved against 4-nitro-quinoline-1-oxide (4-NQO)-induced mutagenicity inSaccharomyces cerevisiae D7 assay. It may be due to bioprotectivity of -mannan and -glucan, which are integral part ofS. cerevisiae cell walls. Due to the good water solubility, low molecular weight (30 kDa), antimutagenic/anticlastogenic, and bioprotective activity against chemical compounds differing in mode of action, GM appears to be a promising natural protective (antimutagenic) agent.     

Effect of water deficit on oxidative stress and degradation of cell membranes in needles of Norway spruce (<Emphasis Type="Italic">Picea abies</Emphasis>)

We studied the impact of mild and severe drought stresses for 42 days and rehydration for 21 days on 4-year-old seedlings of Norway spruce. Water relations in spruce tissues were determined on the basis relative water content of needles and shoot water potential (Ψ_shoot). During the stress, we measured the level of: reactive oxygen species (ROS), antioxidants, and degradation of cell membranes. In the seedlings subjected to severe stress, Ψ_shoot decreased to −2.4 MPa, while in those subjected to mild stress, to −0.8 MPa. After rehydration, shoot water potential increased, but did not reach the control level. Water deficit caused oxidative stress, reflected in an increased production of ROS: superoxide anion radical ( ) and hydrogen peroxide (H₂O₂). Their concentrations in needles were the highest in seedlings subjected to severe stress, where they exceeded the control level by 116% and 30%, respectively. During rehydration, the differences in ROS levels between treated and control seedlings diminished. Oxidative stress causing degradation of cell membranes included: de-esterification of phospholipids, oxidation of fatty acids, and increase in concentration of malondialdehyde, as their permeability to ions increased by 125%. In the defence against the oxidative stress in needles, an important role was played by low-molecule antioxidants such as glutathione, ascorbic acid, flavonoids, α-tocopherol and antioxidant enzymes. An increase in intensity of water deficit caused a significant reductio in the level of low-molecular antioxidants, which attests to their utilization during the process of scavenging for free radicals. Water deficit at Ψ_shoot=−1.7 MPa caused a decline in ascorbic acid level by 37% in needle cells. An effective defensive mechanism removing the excess of ROS was also reflected in the activity of the main enzymes of oxidative stress: superoxide dismutase (SOD) and guaiacol peroxidase (PO). As a result of water deficit, SOD activity increased by 80 %, while PO activity decreased by 82 %.     

Dynamics of Cadmium Distribution in the Intercellular Space and Inside Cells in Soybean Roots, Stems and Leaves

Soybean (Glycine max L.) plants grown in nutrient solution were exposed to 1 mM Cd(NO₃)₂ for 24 h. Dynamics of distribution of cadmium among its different forms (water soluble, Ca-exchangeable and complexed) in the intercellular space and the ratio of the intercellular and intracellular cadmium in roots, stems and leaves were studied. In roots, in the beginning of treatment the largest portion of Cd was found in the intercellular space and 1 h later Cd content started to decrease, so that between 13- and 24-h treatment an equilibrium was reached in which about 70 % of Cd was found inside cells. In stems, already after 1-h treatment, the Cd concentrations in the cells and intercellular space were similar, the equilibrium being disturbed after 13 h, so that after 24-h treatment 80 % of Cd was found inside cells. In leaves, up to the 13 h Cd distribution showed fluctuation, after that equilibrium was reached, with 70 % of intracellular Cd. The highest contents of all Cd forms in the intercellular space was observed in roots.     

Research into Plant Invasions in a Crossroads Region: History and Focus

The Czech Republic is a central European country whose geographical location, natural conditions, history of human settlement, and present land-use management make it relatively prone to plant invasions, hence it represents a convenient model for their study. Research in plant invasions, which date to the late 19th century, is reviewed in the present paper. A long-term floristic tradition allowed for the accumulation of a large body of floristic data on alien plants. During the 1960s–1970s, the main research focus was on their distribution. In this period, attempts were also made to predict potential invasiveness of weeds of arable land. The success rate of this prediction was about 39%. Considerable effort was put into a detailed classification of human-accompanying plants and the terminology associated with the issue. There is a high level of taxonomic research conducted in the country, and the new Flora of the Czech Republic treats the immigration status of taxa with reasonable care and detail. A complete, serious catalogue of the alien plants of the country has been published recently: there are currently 1378 alien plants (33.4% of the total flora). The core of present research in plant invasions is in ecological, biological, and biogeographical studies, focussed on (i) the history of invasion of particular species since their introduction, (ii) the role and importance of alien species in vegetation, including their participation in succession, and (iii) the major invasive species of the Czech flora and comparison of congeners. Reynoutria spp., Heracleum mantegazzianum, Oenothera spp., Pinus strobus, and Bidens frondosa are among the taxa most intensively studied in recent time. A complete list of 69 invasive plants in the Czech flora introduced after the year 1500 [following the definition of Richardson et al. (2000a) Diversity and Distributions 6: 93–107] is given. The available legal instruments relevant to the issue are reviewed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Local and regional patterns of species richness in Central European vegetation types along the pH/calcium gradient

We investigated the relationship between soil pH/calcium content and species richness of vascular plants in seven broadly defined Central European vegetation types, using Ellenberg indicator values for soil reaction and a phytosociological data set of 11,041 vegetation sample plots from the Czech Republic. The vegetation types included (A) broad-leaved deciduous forests, (B) meadows, (C) dry grasslands, (D) reed-bed and tall-sedge vegetation, (E) fens and transitional mires, (F) perennial synanthropic vegetation and (G) annual synanthropic vegetation. Relationships between local species richness (alpha diversity) and pH/calcium were positive for vegetation types A and C, negative for D and G, unimodal for E, and insignificant for B and F. Ellenberg soil reaction values explained 37% of variation in local species richness for vegetation type E, 24% for A, 13% for D, but only less than 4% for the others. Species pool size, i.e., the number of species that can potentially occur in a given habitat, was calculated for each plot using Beals index of sociological favourability applied to a large phytosociological database. For most vegetation types, the relationships between species pool size and pH/calcium were similar to the relationships between local species richness and pH/calcium, with the exception of meadows (weak unimodal) and perennial synanthropic vegetation (weak negative).These patterns suggest that for those types of Central European vegetation that developed independently of human influence in the Pleistocene or early Holocene (dry grasslands, deciduous forests), there are larger pools of calcicole than calcifuge species. This pattern is also found at the level of local species richness, where it is, however, less clearly pronounced, possibly due to the predominance of a few widespread and generalist calcifuges in acidic habitats. The unimodal pattern found in mires may result from similar underlying mechanisms, but in high pH environments mineral-rich spring waters probably decrease species richness by having toxic effects on plant growth. By contrast, vegetation types developed under direct human influence (meadows, synathropic vegetation) show weak negative or no relationships of local species richness or species pool to pH/calcium gradient. These results support the hypothesis ofPärtel (Ecology 83: 2361–2366, 2002) andEwald (Folia Geobot. 38: 357–366, 2003), that the modern calcicole/calcifuge disparity in the species pool of Central European flora has resulted from historical and evolutionary processes that took place on high pH soils. In the Pleistocene, calcareous soils dominated both the dry continental landscapes of Central Europe and glacial refugia of temperate flora, which were mostly situated in southern European mountain ranges with abundant limestone and dolomite. The negative pattern of species richness along the pH/calcium gradient found in reed-bed and tall-sedge vegetation, however, is not consistent with this historical explanation.     

Generation of functional dendritic cells for potential use in the treatment of acute lymphoblastic leukemia

Immunotherapy of malignant diseases mediated by dendritic cells (DC) pulsed with tumor antigens ex vivo is a promising new tool in the individual treatment of malignant diseases. The present study focuses on the problem of how to optimize in vitro culture conditions and induce the maturation of DC with the capacity to induce antitumor immunity toward leukemic cells. DC were generated from peripheral mononuclear cells by co-cultivation with granulocyte/macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). Tumor antigens were added for 2 h after 7 days in culture. Irradiated leukemic blasts, blast lysate, apoptotic cells from the Jurkat cell line (T ALL) and their lysate were used in various concentrations for antigen pulsing. Harvested DC were phenotyped by flow cytometry, and viability was assessed using trypan blue exclusion (Annexin test). After the cells had been pulsed with tumor antigens and co-cultured with autologous lymphocytes, the production of interferon-gamma (IFN-gamma) and IL-12 was analyzed, and lymphocyte proliferative response and cytotoxicity against the target tumor cell line were assessed. The cultivation of monocytes under the described conditions led to the expression of surface markers typical of DC (i.e. CD83, CD86, HLA-DR, CD11c and CD40). Pulsation by antigens from leukemic cells further increased the cell populations expressing these markers. Antigen pulsation decreased the viability of generated DC depending on the increase in concentration of tumor antigens. Pulsed DC-lymphocyte interaction increased the proliferative response of lymphocytes and IFN-gamma production depending on the type of tumor antigens used for pulsation. The highest proliferative response was detected with DC pulsed with Jurkat cell-line lysate. Similarly to the proliferation assay, cytotoxic testing showed the highest efficiency of DC pulsed with Jurkat cell-line lysate in killing the target malignant cells. Our results show that an appropriate antigen concentration used for DC pulsing is one of the crucial factors in an effective treatment strategy, as high concentrations of tumor antigens induce apoptosis of DC, thereby rendering them non-functional. Under optimal conditions, pulsation by lysate from leukemic blasts induced the maturation of DC and led to an increase in the proliferation of autologous lymphocytes, to the production of Th1-cytokines and to the induction of cytotoxicity toward the leukemic cell line. These results are encouraging for the possible application of pulsed DC in the therapy of acute lymphoblastic leukemia.