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  • Determination of Bacillus cereus and its toxins from milk and milk products
    Godič Torkar, Karmen ; Smole Možina, Sonja
    The aim of our study was to review and compare certain classical, biochemical,immunological and molecular-genetic methods used in determination and characterizing of the bacteria Bacillus cereus from ... milk and milk products. About 446 bacterial strains were isolated from raw bulk milk, milk from mastitis cows and from milk products. The samples were inoculated in a tryptic soy broth using the MPN technique. After incubation, the cultures were inoculated on PEMBA and MYP agar, where detected. Hemolytic activity was determined on blood agar. The presumptive strains belonging to the Bacillus cereus species which could not ferment mannitol and gave positive lecithinase and hemolytic reactions were confirmed with the API 50 CHB and API 20 E test system using the identification program V.2.0. (BioMerieux). For further analysis, we chose 82 Bacillus cereus strains. The PCR amplification of specific DNA regions for cereolysin cerAB (for phospholipase C and sphingomyelinase activity) and hemolysin hblA genes (for the B component of the hemolysin BL) was carried out using the primer sets Pf-Cr and HblAl-HblA2,respectively. All strains were analyzed using the BCET-RPLA (Oxoid) immunoassay method to determine the production of the L2 component of hemolytic diarrheal enterotoxin, while the TECRA-BDEVIA (Tecra) kit was used to determine the production of nonhemolytic diarrhealenterotoxin (NHE). After confirmation with API biotyping, only 79%o of all presumptive strains which produced a positive reaction on selective media truly belonged to the Bacillus cereus species. The remaining 21% belonged to the other Bacillus spp. With the primer set Pf and Vr, cerAB gene fragment in 78 out of the 82 strainstested was amplified. All PCR products were of the expected size of 1460 bp. About 873 bp long hblA amplification product was obtained from 66 strains, which corresponds to the results from the BCET-RPLA test kit. About 80% of 82 strains produced hemolytic while 96% of them produced nonhemolytic diarrheal enterotoxin, and some 78% of the strains produced both enterotoxin types. Almost all Bacillus cereus isolates from milk and milk products produced one of the two tested diarrheal enterotoxins. The isolated strains showed a large degree of heterogeneity after biotyping with the API kits. The comparative analysis of phenotypic expression and PCR amplification of genes coding for lecithinase and hemolytic diarrheal entertoxin synthesis revealed a high level of correlation and confirmed the usefullness of rapid molecular detection and/or identification methods for toxigenic. Bacillus cereus strainsfrom milk and milk products.
    Vir: Periodicum biologorum : an interdisciplinary international journal of the Societas Scientiarum Naturalium Croatica established 1885. - ISSN 0031-5362 (Vol. 103, no. 2, 2001, str. 169-173)
    Vrsta gradiva - članek, sestavni del
    Leto - 2001
    Jezik - angleški
    COBISS.SI-ID - 2539896

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