Kvantifikacija prisotnosti, adhezivnosti in filmotvornosti bakterij vrste Listeria innocua z genom za luciferazo na polistirenski površini

Vidic, Natalija (author), Klančnik, Anja (mentor), Berlec, Aleš (comentor)

Bakterije vrste Listeria innocua so nepatogene bakterije, prisotne v okolju ter hrani, ki smo jih zaradi podobnosti s patogenimi bakterijami vrste Listeria monocytogenes v raziskovanju uporabili kot modelni organizem. Bakterije vrste L. innocua in L. monocytogenes tvorijo biofilm, ki povečuje preživetje mikroorganizmov znotraj živilsko predelovalne industrije. Biofilm je lahko vir kontaminacije delovnih površin, posledično pa tudi kot vir prenosa patogenov na živila. Zaradi resne problematike tvorbe biofilmov znotraj živilske predelovalne industrije, je nujno potreben razvoj novih alternativnih in cenejših metod s hitro, natančno in specifično analizo. V magistrskem delu smo vpeljali novo metodo kvantifikacije prisotnosti, adhezivnosti (čas inkubacije 4, 8 in 24 ur) in filmotvornosti (čas inkubacije 48 in 72 ur) z uporabo seva L. innocua NLUC, z vstavljenim genom za luciferazo ter ovrednotenjem kvantifikacije preko merjenja bioluminiscence. Rezultate smo primerjali z rezultati že uveljavljenih metod – metodo določanja števila adheriranih celic in biofilmskih celic ter metodo določanja biomase po barvanju z barvilom kristal vijolično. Kvantifikacijo prisotnosti, adhezivnost ter filmotvornosti smo preverili tudi po tretiranju celic z vodnimi izvlečki iz višjih gliv. Ugotovili smo, da je vpeljana kvantitativna metoda z merjenjem bioluminiscence ponovljiva, občutljiva, hitra in specifična, ter da omogoča realnejšo oceno prisotnosti, adhezivnosti ter filmotvornosti.

• Language: Slovenian
• Keywords: Listeria innocua, Listeria monocytogenes, adhezija, biofilm, luciferaza, bioluminiscenca
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: BF - Biotechnical Faculty
• Place of publishing: Ljubljana
• Publisher: [N. Vidic]
• Year: 2021
• PID: 20.500.12556/RUL-131478
• UDC: 579.24/.26:577.336
• COBISS.SI-ID: 78609667
• Publication date in RUL: 28.09.2021

VIDIC, Natalija, 2021, Kvantifikacija prisotnosti, adhezivnosti in filmotvornosti bakterij vrste Listeria innocua z genom za luciferazo na polistirenski površini [online]. Master’s thesis. Ljubljana : N. Vidic. [Accessed 31 March 2025]. Retrieved from: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=eng&id=131478

Secondary language

• Language: English
• Title: Quantification of viability, adhesion and biofilm formation on polystyrene surface of Listeria innocua with luciferase gene

Abstract

Bacteria of the species Listeria innocua are non-pathogenic bacteria found in the environment and in food that have been used as model organisms in research because of their similarity to pathogenic bacteria of the species Listeria monocytogenes. Bacteria of the species L. innocua and L. monocytogenes form a biofilm that increases the survival of microorganisms in the food processing industry. Biofilm can be a source of contamination of work surfaces and hence transmission of pathogens to food. Due to the serious problems of biofilm formation within the food processing industry, the development of new alternative and cost-effective methods with rapid, accurate and specific analysis is urgently needed. In this master thesis, a new method was presented to quantify the presence, adhesion (incubation time 4, 8 and 24 hours) and film formation (incubation time 48 and 72 hours) of L. innocua bacteria with inserted luciferase gene and evaluation of quantification by measuring bioluminescence. The results were compared with those of previously established methods - the method for determining the number of adherent cells and biofilm cells, and the method for determining biomass after staining with the dye crystal violet. Quantification of presence, adhesion and film formation were also checked after treatment of cells with aqueous extracts from higher fungi. It was shown that the presented quantitative method by measuring bioluminescence is reproducible, sensitive, fast and specific and allows a more realistic assessment of presence, adhesion and film formation.

• Keywords: Listeria innocua, Listeria monocytogenes, adhesion, biofilm, luciferase, bioluminescence