magistrsko delo
Maja Tabor (Author), Hrvoje Petković (Reviewer), David Stopar (Mentor), David Stopar (Thesis defence commission member), Tjaša Danevčič (Thesis defence commission member), Mojca Narat (Thesis defence commission member), Hrvoje Petković (Thesis defence commission member), Tjaša Danevčič (Co-mentor)

Abstract

Prodigiozin je rdeči pigment, ki sodi v skupino prodigininov. Zanimiv je, ker deluje protimikrobno, protiglivno, algicidno in proti praživalim. Odkrili so tudi, da ima prodigiozin velik potencial kot protirakasta in imunosupresivna molekula. Protimikrobni mehanizem delovanja prodigiozina je slabo poznan, zato smo v magistrskem delu želeli določiti tarčne celične komponente bakterije B. subtilis, na katere deluje prodigiozin in pri tem povzroči celično smrt. Z dilucijsko metodo v mikrotitrskih ploščah smo različnim sevom Bacillus sp. določili MIK. MIK prodigiozina smo določili tudi bakterijam pri spremembi temperature inkubacije in slanosti v gojišču. Protibakterijsko aktivnost smo določali z merjenjem OD650 bakterijskih kultur v tekočem gojišču LB, kjer smo ob različnih fazah rasti kulturam dodali ustrezno koncentracijo prodigiozina. Hkrati smo celice spremljali tudi z uporabo mikroskopije z diferencialnim interferenčnim kontrastom in merjenjem aktivnosti β – galaktozidaze. V kontrolnih poskusih smo B. subtilis PS-216 tretirali z različnimi bakterolitičnimi in bakteriostatskimi sredstvi. Ugotovili smo, da so bili vsi testirani sevi Bacillus občutljivi na dodatek prodigiozina. Vrednosti MIK so bile različne in so znašale od 2,82 ± 1,81 mg/L pri Bacillus sp. 5A do 7,83 ± 2,86 mg/L pri B. subtilis NCIB 3610. V stresnih pogojih se je občutljivost bakterij na prodigiozin še povečala. Z opazovanjem celic pod mikroskopom in spremljanjem OD650 smo ugotovili, da prodigiozin povzroči avtolizo celic. Največji vpliv ima na hitro deleče se celice v eksponentni fazi rasti. Pokazali smo, da delovanje prodigiozina povzroči večjo prepustnost celične membrane. Iz dobljenih rezultatov sklepamo, da prodigiozin najverjetneje inducira avtolizine, ki povzročijo hitro lizo bakterijske populacije.

Keywords

prodigiozin;Bacillus;mehanizem delovanja;protimikrobno sredstvo;liza celic;

Data

Language: Slovenian
Year of publishing:
Typology: 2.09 - Master's Thesis
Organization: UL BF - Biotechnical Faculty
Publisher: [M. Tabor]
UDC: 579.852.11:579.24:604.4:615.33(043.2)
COBISS: 8760185 Link will open in a new window
Views: 1413
Downloads: 960
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Other data

Secondary language: English
Secondary title: Antibacterial action of prodigiosin on Bacillus genus
Secondary abstract: Prodigiosin is a natural red pigment from a family of prodiginins. It is interesting because of its antibacterial, antifungal, algicidal and antiprotozoal activity. In addition, the anticancer and immunosuppressive properties of prodigiosin were described. The mechanism of prodigiosin's antibacterial action is still unknown. The aim of the study was to determine cellular target components which prodigiosin affects and causes bacterial cell lysis. Minimal inhibitory concentration (MIC) of Bacillus sp. was evaluated by using broth dilution method in microtiter plates. MIC was also evaluated at different incubation temperatures and different medium salinity. The antibacterial activity was evaluated with measuring optical density of bacterial strains in the LB medium at 650 nm, where prodigiosin was added at different cell growth phases. For cell viability evaluation differential interference contrast microscopy was used and cytoplasmic membrane leakage was determined by measuring β – galactosidase activity. As a control, B. subtilis PS-216 was treated with different bacteriolytic and bacteriostatic agents. We have demonstrated, that all tested Bacillus strains are susceptible to prodigiosin. MIC value was between 2,82 ± 1,81 mg/L against Bacillus sp. 5A and 7,83 ± 2,86 mg/L against B. subtilis NCIB 3610. Exposure of bacterial strains to stress caused by temperature fluctuation and NaCl concentraton increased susceptibility to prodigiosin. Microscopy and optical density data indicate that prodigiosin causes celly lysis. The highest impact of prodigiosin was observed on actively dividing cells in the exponental growth phase. We have observed that prodigiosin increases permeability of cytoplasmic membrane. Based on our results, we have proposed that prodigiosin induces autolysins, leading to fast autolysis of bacterial culture.
Secondary keywords: prodigiosin;mechanism of action;antimicrobial action;cell lysis;
Type (COBISS): Master's thesis/paper
Study programme: 0
Thesis comment: Univ. v Ljubljani, Biotehniška fak., Študij biotehnologije
Pages: XI, 44 f., [17] f. pril.
ID: 10850850