| Abstract | U ovom radu provedena je karakterizacija autohtone populacije bakterija mliječne kiseline (BMK), prirodno prisutne u tradicionalnim hrvatskim prozvodima, svježem siru i kiselom kupusu, s ciljem definiranja probiotičkih starter kultura. Mikrobiološka analiza pokazala je da BMK predstavljaju dominantnu autohtonu populaciji u ovim proizvodima, a što je potvrđeno s PCR-DGGE metodom. Nakon što su pomoću klasičnih mikrobioloških metoda odabrani autohtoni bakterijski izolati okarakterizirani kao gram-pozitivni, katalaza-negativni, nesporogeni, koki ili štapići, koji sintetiziraju visoke koncentracije mliječne kiseline, provedena je njihova daljna fenotipska karakterizacija pomoću SDS-PAGE elektroforeze ukupnih staničnih proteina i API 50 CHL testa. Kako je preživljavanje probiotičkih sojeva u gastrointestinalnom traktu (GIT) jedan od najvažnijih kriterija pri izboru probiotika, ispitano je preživljavanje svih odabranih autohtonih izolata u simuliranim uvjetima GIT. Iz autohtonog svježeg sira odabrana su 2 različita bakterijska soja, a iz spontano fermentiranog kiselog kupusa 5, za koje je RAPD analizom DNA profila, dobivenih sa specifičnom oligonukleotidnom početnicom, potvrđeno da se radi o različitim sojevima BMK. Autohtoni izolati A7 i A8, te SF1, SF2, SF4, SF9 i SF13 su pokazali visoki stupanj preživljavanja u GIT in vitro. Odabrani izolati su osjetljivi na ispitane antibiotike, osim na vankomicin, za koji predstavnici Lactobacillus vrsta imaju urođenu rezistenciju. Isti sojevi su ispunili važne selekcijske kriterije za probiotike: antimikrobno djelovanje, sposobnost autoagregacije, koagregacije s potencijalnim kontaminantima hrane, te adheziju na Caco-2 stanične linije. Liofilizacijom su pripremljeni suhi aktivni preparati odabranih bakterijskih sojeva. Nakon liofilizacije broj živih stanica primjenjenih kultura se održao visok (cca 1010 CFU/g), što je važno svojstvo s tehnološkog aspekta priprava, čuvanja i primjene sojeva. |
| Abstract (english) | In this research characterization of the autochthonous lactic acid bacteria (LAB) population, naturally present in traditional Croatian products, fresh cheese and sauerkraut, was carried out, aimed to define potential probiotic starter cultures. Microbiological analysis revealed that LAB represent dominant autochthonous population in these high quality products and this was confirmed by the PCR-DGGE. After the autochthonous bacterial isolates were characterized as gram-positive, catalase-negative, asporogenous either cocci or rods that synthesize high concentration of lactic acid, by classical microbiological methods, further phenotypic characterization by SDS-PAGE electrophoresis of the whole cell proteins and by API 50 CHL test was performed. As a high level of probiotic survival in the gastrointestinal tract (GIT) is one of the most important criteria in probiotic selection, all eligible autochthonous strains were examined for their ability to survive harsh simulated GIT conditions. 2 different bacterial strains were selected from the autochthonous fresh cheeses and 5 from spontaneously fermented sauerkraut, which were discriminated as a different LAB strains by the RAPD analysis of DNA profiles obtained with specific oligonucleotides. Autochthonous isolates A7 and A8 but also SF1, SF2, SF4, SF9, SF13 showed high survival rates in the GIT in vitro. Selected isolates were susceptible to the tested antibiotics excluding vancomycin, for which representatives of the Lactobacillus species have an innate resistance. All selected strains fulfilled important selection criteria for probiotics: antimicrobial activity, autoaggregation ability, coaggregation with potential food pathogens, and adhesion to Caco-2 cell line. Finally, dry active preparation of the selected bacterial strains was obtained by freeze-drying. Viable cell count of the freeze-dried cultures was maintained high as it was cca 1010 CFU/g, what is of importance from the technological aspect due to the preparation, storage and application of the probiotic strains. |
