Effects of Propionibacterium on the growth and mycotoxin production by some species of Fusarium and Alternaria.

The aim of this research was to study the antifungal properties of propionibacteria. Three fractions from cultures of Propionibacterium freudenreichii ssp. shermanii 41 and ssp. freudenreichii 111 (i.e. culture containing viable bacteria, cell-free supernatant and bacteriocin preparation) were tested for their ability to inhibit the growth and mycotoxin production of Alternaria alternata, Fusarium culmorum, Fusarium graminearum and Fusarium verticillioides. The growth of the fungi was monitored during cultivation using a plating method. The concentration of toxins produced was measured by HPLC on the 14th day of culture. Altenuene and tenuazonic acid were determined in cultures of A. alternata whilst concentration of nivalenol, deoxynivalenol, fumonisin B1 and zearalenone was measured in Fusarium cultures. The strongest inhibition of growth and toxin production was observed in the presence of cultures containing viable cells and supernatants obtained from propionibacteria cultures. The bacteriocin extracts generally had a weak fungistatic effect on the growth of A. alternata, F. culmorum and F. graminearum. Despite the fact that growth was slower in the presence of bacteriocin extracts than in control trials, none of the preparations prepared from the propionibacteria significantly reduced the level of mycotoxin production. The ability of P. freudenreichii ssp. freudenreichii 111 to remove zearalenone from liquid medium was also evaluated. It was shown that both viable and non-viable cells caused a decrease in zearalenone concentration in the medium.

Detection of bar gene encoding phosphinothricin herbicide resistance in plants by electrochemical biosensor.

An electrochemical biosensor for the detection of bar gene coding phosphinothricin herbicide resistance is presented. The detection was based on hybridization reaction between the specific to bar gene 19-mer probe immobilized on the electrode surface and complementary DNA in a sample. Single-stranded DNA probe specific to bar gene was covalently attached by 5'-phosphate end to the surface of carbon paste electrode. Outer layer of a conventional CPE was provided with carboxyl groups of stearic acid. ssDNA was coupled to the electrode through ethylenediamine with the use of water-soluble 1-ethyl-3(3'-dimethylaminopropyl)-carbodiimide and N-hydroxy-sulfosuccinimide as activating reagents. Hybridization reaction at the electrode surface was detected via Co(bpy)(3)(3+), which possess a much higher affinity to the resulting DNA duplex compared to ssDNA probe. Detection limit of the sensor was 0.1 microM of target DNA fragments and its response was linear from 5 to 20 microM. Hybridization event was also detected by measuring guanine peak but this approach presented distinctly higher detection limit (1 muM) and lower reproducibility. Complete time of one measurement with the use of the biosensor including covalent attachment of ethylenediamine (linker) and ssDNA probe to the electrode, hybridization with target and interaction with electroactive indicator was about 70 min.

Application of DNA Hybridization Biosensor as a Screening Method for the Detection of Genetically Modified Food Components.

An electrochemical biosensor for the detection of genetically modified food components is presented. The biosensor was based on 21-mer single-stranded oligonucleotide (ssDNA probe) specific to either 35S promoter or nos terminator, which are frequently present in transgenic DNA cassettes. ssDNA probe was covalently attached by 5'-phosphate end to amino group of cysteamine self-assembled monolayer (SAM) on gold electrode surface with the use of activating reagents - water soluble 1-ethyl-3(3'- dimethylaminopropyl)-carbodiimide (EDC) and N-hydroxy-sulfosuccinimide (NHS). The hybridization reaction on the electrode surface was detected via methylene blue (MB) presenting higher affinity to ssDNA probe than to DNA duplex. The electrode modification procedure was optimized using 19-mer oligoG and oligoC nucleotides. The biosensor enabled distinction between DNA samples isolated from soybean RoundupReady® (RR soybean) and non-genetically modified soybean. The frequent introduction of investigated DNA sequences in other genetically modified organisms (GMOs) give a broad perspectives for analytical application of the biosensor.