Growth of kefir grain in whey and resistance of kefir grain isolates to antibiotics

V magistrskem delu smo želeli ugotoviti, ali je osiromašena kisla sirotka potencialen medij za gojenje kefirnih zrn. Spremljali smo rast biomase 10 vzorcev kefirnih zrn slovenskega, angleškega, nemškega in ruskega porekla. Spremljanje rasti je potekalo 38 dni, kefirna zrna pa so fermentirala pri 25 °C. Pričakovali smo, da bo osiromašena kisla sirotka potencialen medij za uspešno rast in pridobivanje biomase kefirnih zrn. To hipotezo smo tudi potrdili, saj so vsa preiskovana kefirna zrna uspešno pridobivala na biomasi, nekatera bolje, druga pa slabše, čeprav smo za optimizacijo pogojev rasti vrednost pH osiromašene kisle sirotke vsakokrat uravnali na 7. Pri pridobivanju biomase se je najbolje odrezal vzorec A podjetja Kombuchaorganic iz Velike Britanije, najslabše pa vzorec H iz podjetja Arhel d.o.o. Vrednotili smo tudi občutljivost izolatov iz kefirnih zrn za antibiotike, saj je bila raba antibiotikov v preteklosti slabo nadzorovana in so nekatere bakterije lahko razvile odpornost proti antibiotikom. Preverili smo skupno 50 sevov mlečnokislinskih bakterij, ki so bili predhodno osamljeni iz kefirnih zrn: Leuconostoc mesenteroides (8), Lactobacillus helveticus (4), Lactobacillus kefiranofaciens (8), Lactococcus lactis (9), Lentilactobacillus parakefiri (2), Lentilactobacillus kefiri (18) ter en sev vrste Streptococcus thermophilus. Pri tej analizi smo uporabili tehniko mikrodilucije, kjer so na ploščah EUST in EUVENC predhodno nanešene določene koncentracije antibiotikov. Še posebno pozornost smo namenili klinično pomembnim antibiotikom, za katere Evropska agencija za varnost hrane EFSA določa vrednosti MIK: ampicilin, gentamicin, kanamicin, streptomicin, eritromicin, klindamicin, tetraciklin, kloramfenikol in vankomicin. Ugotovili smo, da je med preiskovanimi sevi odpornost najpogosteje prisotna proti naslednjim antibiotikom: kanamicin, kloramfenikol in tetraciklin. Na podlagi pregledane literature lahko trdimo, da ta odpornost proti tem antibiotikom trenutno ne predstavlja tveganja za zdravje ljudi.In the master\u27s thesis, we investigated whether depleted acid whey is a potential medium fort the cultivation of kefir grains. We observed the growth of biomass of 10 samples of kefir grains of Slovenian, English, German and Russian origin. Growth was monitored for 38 days, and kefir grains were fermented at 25 °C. We expected that depleted whey would be a potential medium for successful growth and production of kefir grain biomass. All of the kefir grains tested successfully produced biomass, some better and some worse, although the pH of the depleted whey was adjusted to 7 to optimize growth conditions. In biomass production, sample A Kombuchaorganic from the United Kingdom was the best and sample H from Arhel d.o.o. was the worst. We also evaluated the sensitivity of kefir grain isolates to antibiotics, as antibiotic use has been poorly controlled in the past and bacteria can develop antibiotic resistance. A total of 50 strains of lactic acid bacteria, previously isolated from kefir grains were evaluated: Leuconostoc mesenteroides (8), Lactobacillus helveticus (4), Lactobacillus kefiranofaci (8), Lactococcus lactis (9), Lentilactobacillus parakefiri (2), Lentilactobacillus kefiri (18), and one strain of Streptococcus thermophilus. In this analysis, a microdilution method was used in which specific concentrations of antibiotics were pre-applied to EUST and EUVENC plates. Particular attention was paid to clinically relevant antibiotics for which EFSA determines MIC values: Ampicillin, Gentamicin, Kanamycin, Streptomycin, Erythromycin, Clichamicin, Tetracycline, Chloramphenicol, and Vancomycin. Resistance to the following antibiotics was found most frequently: Kanamycin, Chloramphenicol, and Tetracycline. Based on the literature evaluated, it can be claimed that resistance to these antibiotics does not currently pose a threat to human health