Search

Bakterije vrste C. jejuni so najpogostejše povzročiteljice kampilobakterioze ter gastroenteritisov pri človeku. Za svoje razmnoževanje in rast potrebujejo optimalne pogoje, zato so znani kot zelo zahtevni in občutljivi mikroorganizmi. Sposobnost tvorbe biofilmov jim omogoča večjo odpornost na protimikrobne učinkovine ter s tem lažje preživetje. Nadzor bakterij v odporni obliki biofilmov zahteva uporabo še večjih koncentracij antibiotikov. Problem naraščajoče odpornosti bakterij zato zahteva vpeljavo novih alternativnih strategij za nadzor nad odpornimi sevi in odpornimi oblikami biofilma. Ena od alternativnih strategij je uporaba protimikrobnih učinkovin naravnega izvora in sicer v njihovih sub-inhibitornih koncentracijah. V teh koncentracijah učinkovine ne vplivajo na rast, ampak na druge lastnosti bakterij in tako ne povzročajo razvoja odpornosti. Namen naloge je bil: (i) predstaviti različne učinkovine naravnega izvora in njihovo dokazano delovanje na inhibicijo tvorbe in redukcijo nastalega biofilma bakterij C. jejuni, (ii) opisati mehanizme delovanja na različnih celičnih nivojih C. jejuni ter (iii) primerjati tarčno delovanje antibiotikov in učinkovin naravnega izvora pri bakterijah C. jejuni. V zaključku naloge smo opisali modulacijo genov oz- proteinov, ključnih za nastanek biofilma ter vključene v mehanizme gibljivosti, adhezije in medcelične komunikacije, v celicah tretiranimi z učinkovinami naravnega izvora. Na ta način smo dokazali tarčne mehanizme delovanja protimikrobnih učinkovin naravnega izvora na C. jejuni. Bacteria of the species C. jejuni are the most common causative agents of campylobacteriosis and gastroenteritis in humans. They require optimal conditions for their replication and growth, which is why they are known to be very fastidious and sensitive microorganisms. The ability to form biofilms makes them more resistant to antimicrobial agents and thus more survivable. To control bacteria in the resistant form of biofilms require the use of even higher concentrations of antibiotics. Therefore increasing bacterial resistance requires the introduction of new alternative strategies to combat resistant strains and resistant forms of biofilms. One of these alternative strategies is the use of antimicrobial agents of natural origin at their subinhibitory concentrations. At these concentrations, the agents do not affect the growth but other properties of the bacteria and thus do not lead to development of resistance. The purpose of this thesis was: (i) to present different compounds of natural origin and to demonstrate their ability to inhibit biofilm formation or reduce the formed biofilm of C.jejuni, (ii) to describe the mechanisms of action at different cellular levels of C. jejuni, and (iii) to compare the target effect of antibiotics and compounds of natural origin on C. jejuni. At the end, we described the modulation of genes or proteins crucial for biofilm formation and their involvement in the mechanisms of motility, adhesion and intercellular communication, in cultures treated with natural antimicrobial agents. In this way, we have explained the target mechanisms of action of antimicrobial agents of natural origin in C. jejuni cells.

Zaradi neustrezne in nekritične uporabe protimikrobnih zdravil (PZ) pri živalih, ki vstopajo v prehransko verigo človeka, število odpornih bakterijskih sevov, osamljenih iz živali, kakor tudi živil, narašča. Kampilobakterioza, ki jo povzroča Campylobacter jejuni je ena izmed najpogosteje prijavljenih gastrointestinalnih bolezni pri ljudeh, večja prevalenca odpornih bakterijskih sevov pa povečuje tveganje za neučinkovito zdravljenje bolezni. Zato je pomembno na ta problem opozarjati vse deležnike živilske proizvodno-oskrbovalne verige in tudi zdravnike, da se redno izvajajo testi bakterijske odpornosti na najpomembnejša protimikrobna zdravila (PZ). V eksperimentalnem delu smo določali odpornost 8 sevov C. jejuni iz proizvodnega (klavniškega) in 8 sevov iz naravnega (površinskih voda) okolja proti šestim PZ (eritromicinu, ciprofloksacinu, tetraciklinu, gentamicinu, nalidiksni kislini in streptomicinu) na komercialno pripravljenih mikrotitrskih ploščicah SensititreTM EUCAMP2. Tako klavniški (KS) kot vodni (VS) sevi so pokazali odpornost proti istim vrstam PZ – ciprofloksacinu (CIP), tetraciklinu (TET) in nalidiksni kislini (NAL). Delež odpornih sevov proti posameznim PZ je bil višji pri klavniških (CIP 87,5 %, TET 50 %, NAL 50 %) kot vodnih (CIP 25 %, TET 12,5 %, NAL 25 %) sevih. Zasledili smo tudi večkratno odpornost, ki je bila pri klavniških sevih 87,5 % pri vodnih pa 25 %, in sicer med CIP in TET (KS: 37,5 %), ter CIP in NAL (KS: 50 %, VS: 25 %). Prišli smo do sklepa, da je vzrok večje odpornosti KS večja izpostavljenost PZ, vzrok odpornosti VS pa je verjetno prenos odpornih sevov iz klavnice v njeno okolico. Due to inadequate and uncritical use of antimicrobial drugs in animals, entering the human food chain, the number of resistant bacterial strains, isolated from humans as well as from the food, is increasing. Campylobacteriosis caused by Campylobacter jejuni is one of the most frequently reported gastrointestinal diseases in humans. The appearance of resistant bacterial strains increases the risk of ineffective treatment of the disease. Therefore, it is important to alert all stakeholders in the food production and supply chain, as well as doctors, to regularly carry out bacterial resistance tests on the most important antibiotics. In our experimental work, we tested the resistance of 8 strains of C. jejuni from the production (slaughterhouse) and 8 strains from the natural (surface water) environment against six antimicrobial drugs (erythromycin, ciprofloxacin, tetracycline, gentamicin, streptomycin nalidixic acids) on commercially prepared microtiter plates SensititreTM EUCAMP2. Both slaughtering (SS) and water (WS) strains have shown resistance to the same groups of antibiotics - ciprofloxacin (CIP), tetracycline (TET) and nalidixic acid (NAL). The proportion of resistant strains against individual AM was higher in slaughterhouse (CIP 87,5 %, TET 50%, NAL 50%) than water (CIP 25 %, TET 12,5 %, NAL 25 %) strains. Multidrug resistance was also present, which is in the case of slaughterhouse strains 87,5 % and 25 % in water strains, namely between CIP and TET (SS: 37,5 %) and CIP and NAL (SS: 50 %, WS: 25 %). We have come to the conclusion that the cause for the higher resistance levels of SS might be a higher exposure to antibiotics, and the cause of resistant WS is the transfer of resistant strains from the slaughterhouse to its surroundings.