Odpornost mlečnokislinskih bakterij in bifidobakterij iz črevesne sluznice proti antibiotikom

Kastelec, Lea (author), Bogovič Matijašić, Bojana (mentor), Mohar Lorbeg, Petra (co-mentor)

Raziskovali smo antibiotično odpornost mlečnokislinskih bakterij in bifidobakterij izoliranih iz črevesne sluznice. Zanimali sta nas morebiten pojav fenotipske in genotipske odpornosti. Po homogenizaciji črevesnih bioptov smo izolirali mlečnokislinske bakterije, bifidobakterije smo že imeli izolirane in smo jih vzeli iz mikrobiološke zbirke. Izolate smo z metodo RAPD pregledali za vrstno pestrost in za nadaljnjo analizo izbrali tiste z unikatnimi vzorci pomnožkov. Po izolaciji DNA in namnožitvi 16S rDNA, smo identifikacijo izvedli glede na rezultate sekvenciranja pomnožkov. Sledilo je preverjanje fenotipske odpornosti z metodo razredčevanja v mikrotitrski ploščici. Glede na mejne vrednosti, ki jih je postavila EFSA, je previsoke MIC izkazalo 14 sevov od skupno 104, devet proti eritromicinu, štirje proti tetraciklinu in eden proti kanamicinu. Sev B. longum 34-7 v svoji dednini nosi zapis za gen erm(B), sevu B. adolescentis IM677 smo potrdili prisotnost gena tet(O) in sevu B. longum 34-8 pa prisotnost gena tet(W). Pri nobenem izmed testiranih sevov nismo potrdili prisotnosti genov za odpornost proti kanamicinu. Ker odpornostnih genov nismo našli pri vseh sevih, ki so izkazali fenotipsko odpornost je ta najverjetneje posledica mutacij, ali pa genov, ki jih ne poznamo oz. jih nismo vključili v raziskavo.

• Language: Slovenian
• Keywords: mlečnokislinske bakterije, bifidobakterije, črevesna mikrobiota, črevesna sluznica, antibiotiki, odpornost proti antibiotikom, eritromicin, tetraciklin, kanamicin, odpornostni geni
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Publisher: [L. Kastelec]
• Year: 2018
• PID: 20.500.12556/RUL-100161
• UDC: 579.24:615.33:612.33
• COBISS.SI-ID: 4887160
• Publication date in RUL: 10.03.2018

Secondary language

• Language: English
• Title: Antibiotic resistance of lactic acid bacteria and bifidobacteria from intestinal mucosa

Abstract

The aim of the study was to search for antibiotic (erythromycin, tetracycline and/or kanamycin) resistance among lactic acid bacteria (LAB) and bifidobacteria isolated from children's intestinal mucosa. We were searching for signs of phenotypic or genotypic antibiotic resistance. First we homogenized intestinal biopsy samples and isolated LAB, bifidoacteria strains had already been isolated. LAB isolates were screened with RAPD to see how related they are, for further analysis we chose the ones with uniqe M13 bands. After DNA isolation and 16S rDNA amplification isolates were sent for sequencing to determine their genus or species. Further on, we tested the LAB and bifidobacterial isolates for phenotypic resistance with microdilution method. At microdilution test 14 out of 104 strains showed resistance against antibiotics, nine strains against erythromycin, four strains against tetracycline and one strain against kanamycin. Those 14 strains with high MIC were later on screened with PCR to see if they contain any of resistance genes. Out of 14 strains that showed phenotypic resistance we discovered that one strain of B. longum (34-7) carries gene for erythromycin resistance erm(B) in his DNA, one strain of B. adolescetnis (IM677) carries gene for tetracycline resistance tet(O), strain B. longum (34-8) carries gene tet(W). None of the strains tested carried genes for resistance against kanamycine.

• Keywords: lactic acid bacteria, bifidobacteria, intestinal microbiota, intestinal mucosa, antibiotics, antibiotic resistance, erythromycin, tetracycline, kanamycin, resistance genes