Combining genotoxicity/mutagenicity tests and physico-chemical methodologies can be useful for determining the potential genotoxic contaminants in soil samples. The aim of our study was to evaluate ...the genotoxicity of soil by applying an integrated physico-chemical-biological approach. Soil samples were collected at six sampling points in a Slovenian industrial and agricultural region where contamination by heavy metals and sulphur dioxide (SO₂) are primarily caused by a nearby power plant. The in vitro alkaline version of the comet assay on water soil leachates was performed with Caco-2 and HepG2 cells. A parallel genotoxicity evaluation of the samples was performed by Ames test using Salmonella typhimurium and the Tradescantia micronucleus test. Pedological analyses, heavy metal content determination, and different physico-chemical analyses, were also performed utilizing standard methodology. Water leachates of soil samples were prepared according to standard methods. Since only a battery of biotests with prokaryotic and eukaryotic organisms or cells can accurately estimate the effects of (geno)toxicants in soil samples and water soil leachates, a combination of three bioassays, with cells or organisms belonging to different trophic levels, was used. Genotoxicity of all six water soil leachates was proven by the comet assay on both human cell lines, however no positive results were detected by bacterial assay, Ames test. The Tradescantia micronucleus assay showed increase in micronuclei formation for three samples. According to these results we can assume that the comet assay was the most sensitive assay, followed by the micronucleus test. The Ames test does not appear to be sensitive enough for water soil leachates genotoxicity evaluations where heavy metal contamination is anticipated.
Issue Title: ISAM 7, 7th International Symposium of Anaerobic Microbiology, Smolenice, Slovakia, 15- 18 June 2011 One of the main mechanisms of nanoparticle toxicity is known to be the generation of ...reactive oxygen species (ROS) which primarily damage cell membranes. However, very limited data on membrane effects in anaerobic environments (where ROS could not be the cause of membrane damage) are available. In the following study, rumen anaerobe Ruminococcus flavefaciens 007C was used as a bacterial model to assess the potential effects of Al^sub 2^O3 and TiO2 nanoparticles on membranes in an anaerobic environment. Fatty acid profiles of cultures after exposure to Al^sub 2^O3 or TiO2 nanoparticles were analyzed and compared with the profiles of non-exposed cultures or cultures exposed to bulk materials. Analysis revealed dose-effect changes in membrane composition exclusively when cells were exposed to Al^sub 2^O3 nanoparticles in a concentration range of 3-5 g/L, but were not present in cultures exposed to bulk material. On the other hand, the tested concentrations of nano-TiO2 did not significantly affect the membrane profile of the exposed bacterium. The results suggest the possibility that Al^sub 2^O3 induces changes in bacterial membranes by direct physical interaction, which was supported by TEM image analysis.PUBLICATION ABSTRACT
One of the main mechanisms of nanoparticle toxicity is known to be the generation of reactive oxygen species (ROS) which primarily damage cell membranes. However, very limited data on membrane ...effects in anaerobic environments (where ROS could not be the cause of membrane damage) are available. In the following study, rumen anaerobe Ruminococcus flavefaciens 007C was used as a bacterial model to assess the potential effects of Al sub(2)O sub(3) and TiO sub(2) nanoparticles on membranes in an anaerobic environment. Fatty acid profiles of cultures after exposure to Al sub(2)O sub(3) or TiO sub(2) nanoparticles were analyzed and compared with the profiles of non-exposed cultures or cultures exposed to bulk materials. Analysis revealed dose-effect changes in membrane composition exclusively when cells were exposed to Al sub(2)O sub(3) nanoparticles in a concentration range of 3-5 g/L, but were not present in cultures exposed to bulk material. On the other hand, the tested concentrations of nano-TiO sub(2) did not significantly affect the membrane profile of the exposed bacterium. The results suggest the possibility that Al sub(2)O sub(3) induces changes in bacterial membranes by direct physical interaction, which was supported by TEM image analysis.
Uporaba ksilanaz kot krmnih dodatkov je pri monogastričnih živalih zelo obetavna, saj so njihov ugoden učinek na zdravstveno stanje in prirast živali potrdile številne raziskave. Pred kratkim je bila ...opisana nova vrsta vampne bakterije P. xylanivorans s tipskim sevom Mz5T, ki je močno ksilanolitična. Nekaj njenih ksilanolitičnih encimov je že opisanih, preiskati pa želimo še ostale, da bi ugotovili vzrok visoke ksilanolitične aktivnosti in možnosti uporabe bakterije kot probiotika pri monogastričnih živalih (prašiči, perutnina) ali encimov kot krmnih dodatkov. Iz celičnega izvlečka smo uspeli delno očistiti ksilanazi, veliki 44 kDa in 81 kDa in delno proučiti njuno encimsko delovanje. Primerjali smo tudi encimsko delovanje na celično površino vezanih ksilanaz s ksilanazami celičnega izvlečka. 44 kDa ksilanaza je eksoksilanaza brez β-ksilozidazne aktivnosti, pri ksilanazi velikosti 81 kDa predvidevamo, da gre za endoksilanazo. Ksilanolitični encimi celične površine učinkovito razgrajujejo ksilan, vendar niso sposobni cepiti ksilobioze. Le-to razgradijo ksilanolitični encimi v notranjosti celice.
Soil samples were obtained from Slovenian industrial region Šaleška dolina and aqueous leachates were prepared. The toxicity and genotoxicity potential of these aqueous fractions have been evaluated ...by in vitro toxicity and genotoxicity bioassays. Freshwater toxicity test (PROTOXKIT FTM) with a ciliate protozoan Tetrahymena thermophila, and genotoxicity comet assay using Tetrahymena thermophila, Caco-2 and HepG2 cell lines were performed. Biological data were completed through chemical analyses. It has been shown that physico-chemical analyses alone may not be sufficient to characterize soil hazards. To study soil ecotoxcity it is therefore necessary to take into consideration both, the physico-chemical analyses, toxicity and genotoxicity assays.
Bacteria belonging to or resembling the genus Butyrivibrio represent a significant proportion of bacterial isolates from different mammalian gastro-intestinal tracts from all over the world. The main ...characteristic of this genus is production of butyric acid, which has an important role in maintaining colonic health. The rumen strains – B. fibrisolvens are also known producers of conjugated linoleic acid (CLA), bacteriocins and several hydrolytic enzymes that contribute to effective forage digestion. Genus Butyrivibrio contains isolates that are very diverse in both genomic and phenotypic properties. Different researchers have proposed the grouping of isolates on the basis of some specific characteristics but up to now none was recognised as a new species. Therefore, a complex approach is needed to set basis for new species delineation and characterisations and to define a set of reasonable diagnostic methods for their discrimination.
DNA damage within single mammalian cells can be one of the earliest signs of a whole range of health problems, including disease, diet and exposure to occupational or environmental toxins. The Comet ...assay is a relatively simple and inexpensive method for measuring this damage. It works by determining the number of breaks in the strands of DNA within the cell. Cells are embedded in agarose gel on a microscopic slide and washed to remove the cell membranes, soluble cell contents and histones from the nucleus. An electric field is then activated across and the loops of damaged DNA are pulled towards the anode. Epifluorescence microscopy of stained gels shows the image which gives the techique its name: a clump of undamaged DNA (the head) with the loops pulled away, forming a tail of a comet. We report herein the checking of the Comet assay for the evaluation of oxidative stress on chicken, mouse and pig blood cells, pig sperm cells and chicken hepatocytes. We intend to use the Comet assay as a quantitative method in some nutritional experiments with domestic animals including oxidative food components and antioxidants and for the evaluation of some environmental pollutants effects on animal and human cells.
Butirat je pomemben probiotski dejavnik, ki ugodno vpliva na delovanje in preživetje kolonocitov ter v debelem črevesu deluje tudi antikancerogeno. Pred kratkim je bila na osnovi 17 sevov, izoliranih ...iz vampa, opisana nova vrsta P. xylanivorans s tipskim sevom Mz5T. Sev Mz5T je na ksilanu ovsenih plev produciral butirat in laktat kot glavna produkta fermentacije in pri tem porabljal acetat. Produkcija butirata je močno narasla med logaritemsko fazo in se je še v pozni stacionarni fazi počasi povečevala. Na drugih ogljikovih hidratih (na brezovem in bukovem ksilanu, ksilozi, arabinozi in glukozi) je bila kinetika tvorbe butirata podobna. Največ butirata glede na laktat je nastajalo med zgodnjo logaritemsko fazo neodvisno od substrata. P. xylanivorans Mz5T bi torej lahko uporabili kot probiotik v prehrani živali tako za izboljšanje razgradnje ksilana kot tudi za povečano produkcijo butirata v debelem črevesu. Najbolje bi bilo uporabiti kulturo v zgodnji logaritemski fazi rasti, ki bi jo lahko namnožili na cenenem ogljikohidratnem viru.
U probavnom se sustavu biljojeda nalaze vrlo učinkoviti mikroorganizmi što razgrađuju celulolozu i hemicelulozu iz sažvakanog biljnog materijala, te ih opskrbljuju hranjivim tvarima. Osim protozoa i ...gljivica, razgradnji otporne (hemi)celulozne biomase u buragu znatno pridonose i bakterije. U fokusu je ovog preglednog rada opis enzimskog sustava triju predstavnika bakterija iz buraga što proizvode celulazu, i to: Ruminococcus flavefaciens, Prevotella bryantii i Pseudobutyrivibrio xylanivorans. R. flavefaciens je poznat po proizvodnji jedne od najkompleksnijih enzimskih struktura, pa bi se mogao upotrijebiti za dizajn enzimskog kompleksa za razgradnju stanične stijenke biljaka. S druge strane, bakterije Prevotella bryantii i Pseudobutyrivibrio xylanivorans proizvode jednostavne, slobodne i vrlo aktivne ksilanaze. Bakterija P. xylanivorans ima i probiotička svojstva, pa se može upotrijebiti za proizvodnju bioplina i kao dodatak krmivu. Ispitivanje je genoma i proteoma bakterija što razgrađuju celulozu i hemicelulozu usmjereno na identifikaciju novih enzima, koji se nakon kloniranja i ekspresije u odgovarajućim domaćinima mogu upotrijebiti za izradu vrlo aktivnih rekombinantnih hidrolitičkih mikroorganizama, te primijeniti u različitim biotehnološkim procesima.
