Publication: Peltigera cinsine ait liken ve endolikenik fungus örneklerinin pseudomonas aeruginosa üzerine anti-quorum sensing ve anti-biyofilm etkinliklerinin in vitro ve in siliko yöntemlerle incelenmesi
Abstract
Pseudomonas aeruginosa akut ve kronik enfeksiyonlardan sorumlu biyofilm formları nedeniyle çoklu antibiyotiklere dirençli olabilen fırsatçı patojen bakteridir. Quorum Sensing (QS) mekanizması, antibiyotik direnci, biyofilm formunun oluşumu ile virülans faktörlerin salınması ile ilişkilidir. Bu sebeple, bakterilerin gelişimini baskılamadan, ölüm stresi yaşatmadan hücre içi ve hücreler arası iletişimi ve QS ilişkili biyofilm formunu inhibe edebilecek yeni QS inhibitörlerine (QSI) ihtiyaç duyulmaktadır. Bu çalışmada, özgün tıbbi ilaç özellikleri bildirilen sekonder metabolitleri üreten likenlerin n-Hekzan ve endolikenik fungusların (ELF) etil asetat özütlerinin QSI potansiyelleri araştırılmıştır. Liken sekonder metabolit kütüphanesi oluşturularak, terapötik ve toksisite testleri gerçekleştirilmiş ve toksisite göstermeyen 179 metabolitin P. aeruginosa QS sistemine dahil olan LasI/ LasR, RhlR ve PqsR reseptör proteinleriyle moleküler kenetlenme simülasyonları gerçekleştirilmiştir. En başarılı bağlanma enerjisine sahip Peltigera cinsi likenlere ait 10 metabolit saptanmıştır. In siliko bulgular sonucunda, in vitro etkinliklerinin yüksek olabileceği düşünülen Peltigera cinsi liken türleri Artvin ve Bursa bölgelerinden toplanarak morfolojik tür tayinleri gerçekleştirilmiş daha sonra bu likenlerin talluslarından ELF izolasyonu ve moleküler tanımlamaları gerçekleştirilmiştir. Peltigera cinsi likenlerin ve izole edilen ELF’lerin özütlerinin P. aeruginosa PAO1 CECT4122 suşu üzerinde anti-bakteriyel, anti-biyofilm etkileri ve lasB-gfp, rhlA-gfp ve pqsA-gfp biyosensör suşları ile anti-QS etkileri incelenmiştir. Özellikle P. horizontalis likeninin ve bu türden izole edilerek Penicillium sp. olduğu saptanan ELF28’in özütlerinin P. aeruginosa üzerinde yüksek QSI ve anti-biyofilm potansiyelleri saptanmıştır. Bulgular değerlendirildiğinde, in siliko ve in vitro çalışmaların sonuçlarının birbirleriyle tutarlılıkları görülmüştür. Peltigera likeni ve içerdiği ELF özütlerinin alternatif olarak antibiyotik direnci problemine karşı etkili yeni QSI molekülleri içerme potansiyeline sahip olabileceği ve bu tezin gelecekteki çalışmalar için literatüre katkıda bulunacağı düşünülmektedir.
Pseudomonas aeruginosa is an opportunistic pathogen bacteria that can be resistant to multiple antibiotics due to its biofilm forms responsible for acute and chronic infections. The Quorum Sensing (QS) mechanisms is associated with antibiotic resistance, biofilm formation, and the release of virulence factors. Therefore, there is a need for new QS inhibitors (QSI) that can inhibit intracellular and intercellular communication and the QS-related biofilm form without suppressing the growth of bacteria and causing death stress. In this study, the QSI potentials of n-Hexane extracts of lichens and ethyl acetate extracts of endolichenic fungus (ELF), which produce secondary metabolites with unique medicinal drug properties, were investigated. By creating a lichen secondary metabolite library, therapeutic and toxicity tests were performed, and then molecular docking simulations of 179 metabolites that do not show any toxicity were performed with the LasI/ LasR, RhlR and PqsR receptor proteins involved in the P. aeruginosa QS system. Ten lichen metabolites belonging to the genus of Peltigera were determined to have the most successful binding energy. As a result of in silico findings, the species belonging to Peltigera genus, which are thought to have high in vitro activities, were collected from Artvin and Bursa regions, and morphological identifications were performed, and then ELF isolation and molecular identifications of ELF’s were made from the thalli of these lichen species The extracts of lichens and ELFs, obtained from Peltigera species, were examined against P. aeruginosa PAO1 CECT4122 for anti-bacterial, anti-biofilm effects and lasB-gfp, rhlA-gfp and pqsA-gfp biomonitor strains for anti-QS effects. In particular, high QSI and anti-biofilm potentials against P. aeruginosa were detected for the extracts of P. horizontalis and ELF28 isolated from P. horizontalis and identified as Penicillium sp. When the results were evaluated, the consistency in in silico and in vitro studies was observed. It is thought that the extracts of Peltigera and its ELFs may alternatively have the potential to contain new QSI molecules against antibiotic resistance problem and thus, in aid for future studies, this thesis is expected to contribute to academic literatüre.
Pseudomonas aeruginosa is an opportunistic pathogen bacteria that can be resistant to multiple antibiotics due to its biofilm forms responsible for acute and chronic infections. The Quorum Sensing (QS) mechanisms is associated with antibiotic resistance, biofilm formation, and the release of virulence factors. Therefore, there is a need for new QS inhibitors (QSI) that can inhibit intracellular and intercellular communication and the QS-related biofilm form without suppressing the growth of bacteria and causing death stress. In this study, the QSI potentials of n-Hexane extracts of lichens and ethyl acetate extracts of endolichenic fungus (ELF), which produce secondary metabolites with unique medicinal drug properties, were investigated. By creating a lichen secondary metabolite library, therapeutic and toxicity tests were performed, and then molecular docking simulations of 179 metabolites that do not show any toxicity were performed with the LasI/ LasR, RhlR and PqsR receptor proteins involved in the P. aeruginosa QS system. Ten lichen metabolites belonging to the genus of Peltigera were determined to have the most successful binding energy. As a result of in silico findings, the species belonging to Peltigera genus, which are thought to have high in vitro activities, were collected from Artvin and Bursa regions, and morphological identifications were performed, and then ELF isolation and molecular identifications of ELF’s were made from the thalli of these lichen species The extracts of lichens and ELFs, obtained from Peltigera species, were examined against P. aeruginosa PAO1 CECT4122 for anti-bacterial, anti-biofilm effects and lasB-gfp, rhlA-gfp and pqsA-gfp biomonitor strains for anti-QS effects. In particular, high QSI and anti-biofilm potentials against P. aeruginosa were detected for the extracts of P. horizontalis and ELF28 isolated from P. horizontalis and identified as Penicillium sp. When the results were evaluated, the consistency in in silico and in vitro studies was observed. It is thought that the extracts of Peltigera and its ELFs may alternatively have the potential to contain new QSI molecules against antibiotic resistance problem and thus, in aid for future studies, this thesis is expected to contribute to academic literatüre.