Vrednotenje sposobnosti tvorbe biofilma različnih sevov Pseudomonas aeruginosa in določanje zaviralnega učinka na tvorbo biofilma knjižnice spojin

Podlesnik, Maja (author), Hrast Rambaher, Martina (mentor), Golob, Majda (comentor)

Magistrska naloga obravnava problematiko bakterije Pseudomonas aeruginosa, ki predstavlja enega najpomembnejših oportunističnih patogenov v sodobni medicini. Njena sposobnosti tvorbe biofilma ter izrazita odpornosti proti številnim antibiotikom pomembno prispevata k razvoju kroničnih in težko ozdravljivih okužb. Biofilm predstavlja kompleksno tridimenzionalno strukturo mikroorganizmov, obdano z zunajceličnim matriksom, ki bakterijam omogoča zaščito pred neugodnimi okoljskimi pogoji, imunskim sistemom gostitelja in protimikrobnimi učinkovinami. Posledično so okužbe, povezane z biofilmi, pogosto dolgotrajne, zahtevajo intenzivno zdravljenje in predstavljajo veliko breme za zdravstveni sistem. Glavni namen raziskave je bil ovrednotiti sposobnost tvorbe biofilma pri različnih sevih bakterije P. aeruginosa ter določiti zaviralni učinek izbranih spojin iz knjižnice spojin na nastanek biofilma. V raziskavo je bilo vključenih 1136 spojin, ki so bile testirane z uporabo standardizirane metode v mikrotitrskih ploščah. Tvorbo biofilma smo kvantificirali z metodo barvanja s kristal vijoličnim in merjenjem absorbance pri 584 nm. Rezultati so pokazali, da različni sevi P. aeruginosa izkazujejo različno stopnjo tvorbe biofilma, kar nakazuje pomembno vlogo genetskih in okoljskih dejavnikov. Nekateri klinični in okoljski sevi iz mleka so tvorili izrazito več biofilma kot referenčni sev PAO1, kar dodatno poudarja kompleksnost obravnavanega problema. Pri presejalnem testiranju spojin smo identificirali več kandidatov, ki so zavirali tvorbo biofilma za 50 % in več. Ponovno testiranje izbranih spojin je potrdilo njihovo potencialno protibiofilmsko delovanje, kar kaže na možnost nadaljne uporabe v razvoju novih terapevtskih pristopov. Razprava rezultatov poudarja, da je inhibicija tvorbe biofilma obetavna strategija za izboljšanje učinkovitosti zdravljenja okužb, saj ne deluje neposredno baktericidno, temveč le zmanjšuje virulentnost bakterij in njihovo sposobnost preživetja. Takšen pristop lahko prispeva k zmanjšanju razvoja odpornost na antibiotike. Kljub spodbudnim rezultatom raziskava izpostavlja tudi omejitve, kot so pogoji testiranja in vitro, ki ne odražajo v celoti kompleksnosti okolja in vivo. Zaključki naloge poudarjajo, da je razumevanje mehanizmov nastajanja biofilma ter identifikacija učinkovitih inhibitorjev ključnega pomena za razvoj novih strategij zdravljenja okužb, povezanih s P. aeruginosa. Nadaljnje raziskave bodo morale vključevati optimizacijo identificiranih spojin ter njihovo testiranje v bolj kompleksnih, klinično relevantnih modelih.

• Language: Slovenian
• Keywords: Pseudomonas aeruginosa, protibiofilmske spojine, biofilm, okužbe
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2026
• PID: 20.500.12556/RUL-182560
• Publication date in RUL: 16.05.2026

Secondary language

• Language: English
• Title: Evaluation of biofilm forming potential of different Pseudomonas aeruginosa strains and determination of antibiofilm activity of a compound library

Abstract

The master's thesis addresses the challenges associated with Pseudomonas aeruginosa, one of the most important opportunistic pathogens in modern medicine. Its ability to form biofilms and its high level of resistance to antibiotics significantly contribute to persistent and difficult treatment of infections. A biofilm is a complex three-demensional structure of microoganisms embedded in an extracellular polimeric matrix, which protects bacteria from enviromental stress, host immune responses, and antimicrobal agents. Consequently, biofilm-associated infections are often chronic, require prolonged treatment, and represent a substantial burden on healthcare systems. The main objective of this research was to evaluate the biofilm-forming capacity of different strains of P. aeruginosa and to determine the inhibitory effect of selected compounds from compound library on biofilm formation. A total of 1136 compounds were screened using a standardized microtiter plate assay. Biofilm formation was quantified using crystal violet staining and absorbance measurments at 584 nm. The results demonstrated significant variability in biofilm formation among different P. aeruginosa strains, highlighting the importance of genetic background and isolate origin. Some clinical and enviromental strains from milk exhibited a higher biofilm-forming capacity than the reference strain PAO1, emphasizing the complexity of the problem. Screening identified several compounds capable of inhibiting biofilm formation by 50 % or more. Subsequent validation confirmed the antibiofilm potential of several candidates, indicating their promise for further development. The discussion highlights that targeting biofilm formation represents a promising strategy for improving infection treatment, as it reduces bacterial virulence rather than directly killing bacteria, potentially limiting the development of antibiotic resistance. However, the study also acknowledges limitations, particularly the use of in vitro models, that may not fully reflect in vivo conditions. In conclusion, understanding the mechanisms of biofilm formation and identifying effective inhibitors are crucial steps toward developing new therapeutic approaches for managing P. aeruginosa infections. Further research should focus on optimizing the identified compounds and evaluating their efficacy in more complex and clinically relevant models.

• Keywords: Pseudomonas aeruginosa, antibiofilm compounds, biofilm, infections