Izolacija i karakterizacija bakteriofaga specifičnih za vrste Acinetobacter baumannii i Escherichia coli iz efluenta zagrebačkog uređaja za pročišćavanje otpadnih voda

master's thesis

• Title (english): Isolation and characterization of bacteriophages specific for Acinetobacter baumannii and Escherichia coli species from effluent of the Zagreb wastewater treatment plant
• Author: Marina Mihaljević
• Mentor: Silvija Černi (mentor)
• Committee member: Silvija Černi (predsjednik povjerenstva)
• Committee member: Mladen Krajačić (član povjerenstva)
• Committee member: Sofia Ana Blažević (član povjerenstva)
• Committee member: Tomislav Ivanković (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2022-02-18, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Bakteriofagi (fagi) su virusi koji inficiraju bakterije, te ih posljedično često liziraju. Širenjem
antibiotske rezistencije je porastao interes za terapijsko korištenje faga, osobito kod infekcija
rezistentnim sojevima patogenih bakterija, poput Acinetobacter baumannii. Jedna od osnovnih
metoda proučavanja i izoliranja faga je uzgoj plakova na bakterijskim livadama, a različiti fagi se
često razlikuju po morfologiji plakova koje stvaraju. Cilj ovog istraţivanja bio je izolirati i
karakterizirati fage specifične za bakteriju A. baumannii, te visoko zastupljenu bakteriju
Escherichia coli iz efluenta centralnog ureĎaja za pročišćavanje otpadnih voda grada Zagreba.
Namjera je bila proučiti plakove koje fagi stvaraju na bakterijskim livadama te fage izolirane iz
plakova različite morfologije proučiti transmisijskim elektronskim mikroskopom. Uzorci eluenta
obogaćeni su inkubacijom s navedenim bakterijama pri različitim temperaturama, a prisutnost faga
potvrĎena je konfluentnom lizom bakterijske livade te metodom čistina i plakova tehnikom
nakapavanja. Fagi izolirani iz plakova, uzgojenih metodom čistina u dvostrukom agaru, umnoţeni
su metodom konfluentne lize bakterijske livade te im je odreĎen titar. Na livadama E. coli utvrĎene
su četiri različite morfologije plakova iz kojih su izolirani fagi, dok fagi specifični za A. baumannii
nisu izolirani iz testiranih uzoraka. Korištenom metodom pročišćavanja i koncentriranja uzoraka
nisu dobiveni uzorci zadovoljavajuće kvalitete za karakterizaciju faga elektronskom
mikroskopijom.

Abstract (english)

Bacteriophages (phages) are viruses that infect bacteria, and consequently often lyse host cells.
Interest in the therapeutic usage of phages has increased due to the spread of antibiotic resistance,
especially in infections caused by resistant strains of pathogenic bacteria, such as Acinetobacter
baumannii. One of the fundamental methods for studying and isolating phages is growing plaques
on bacterial lawns, where different phages can often be distinguished by the morphology of plaques
they produce. The aim of this research was to isolate and characterize phages specific for A.
baumannii and highly abundant Escherichia coli from effluent of the Zagreb wastewater treatment
plant; to study phage produced plaques on bacterial lawns; and to observe phages isolated from
plaques of different morphology by transmission electron microscopy. Effluent samples were
enriched by incubation with the mentioned bacteria at different temperatures, and the presence of
phages was confirmed by both confluent lysis of bacterial lawn and combined spot and plaque
assay. Phages, isolated from plaques grown by double agar overlay, were multiplied by confluent
lysis of bacterial lawns and their titer was determined. Four plaques of different morphology were
observed on E. coli bacterial lawns, from which specific phages were isolated, while phages
specific for A. baumannii were not isolated from tested samples. The methodology used for phage
purification and concentration was not of satisfactory quality for their characterization by electron
microscopy.

• Keywords: plak; metoda plakova u dvostrukom agaru; terapija fagima
• Keywords (english): plaque; double agar overlay; phage therapy
• Language: croatian
• URN:NBN: urn:nbn:hr:217:046004
• Study programme: Title: Experimental Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra eksperimentalne biologije (magistar/magistra eksperimentalne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access; Embargo expiration date: 2023-02-18
• Created on: 2022-03-24 12:59:48