Rasprostranjivanje gena za rezistenciju na cefalosporine treće generacije među bakterijama komunalnih otpadnih voda

master's thesis

• Title (english): Dissemination of genes encoding resistance to third generation cephalosporins among bacteria from municipal wastewater
• Author: Ivan Kodela Pacenti
• Mentor: Nikolina Udiković Kolić (mentor)
• Mentor: Jasna Hrenović (komentor)
• Committee member: Jasna Hrenović (predsjednik povjerenstva)
• Committee member: Tomislav Ivanković (član povjerenstva)
• Committee member: Goran Klobučar (član povjerenstva)
• Committee member: Željka Vidaković-Cifrek (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2023-09-22, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Bakterijska otpornost na antibiotike predstavlja značajnu prijetnju za ljudsko zdravlje zbog
ograničenih mogućnosti u liječenju infektivnih bolesti. Cefalosporini treće generacije su
antibiotici koji se koriste u liječenju mnogih infekcija, ali Gram-negativne bakterije ih mogu
uspješno hidrolizirati pomoću β-laktamaza proširenog spektra (ESBL), čiji geni se uglavnom
nalaze na plazmidima. Važnu ulogu u širenju te otpornosti imaju uređaji za pročišćavanje
otpadnih voda (UPOV) unutar kojih bakterije, pod pritiskom antibiotika i drugih zagađivala,
izmjenjuju gene za otpornost putem plazmida. U ovom istraživanju korištena je metoda
egzogene izolacije plazmida za analizu prijenosa ESBL gena iz bakterija pročišćenih otpadnih
voda iz sedam UPOV-a u bakteriju primatelj Escherichia coli CV601. Dobiveno je 18
pozitivnih transkonjuganata (E. coli CV601 s ESBL genom na plazmidu) iz pet analiziranih
UPOV-a. Većina transkonjuganata je ispoljavala višestruko otporan fenotip potvrđen
metodama disk difuzije i mikrodilucije. Primjenom metoda lančane reakcije polimeraze (PCR)
i agarozne gel elekroforeze, na svim plazmidima pozitivnih transkonjuganata potvrđena je
prisutnost ESBL gena blaCTX-M-1 i blaTEM, dok je na jednom plazmidu dodatno detektiran gen
blaGES. Osam plazmida je dodatno nosilo gen za karbapenemazu blaVIM ili blaNDM. Primjenom
PCR metode za određivanje replikona, u svim pozitivnim transkonjugantima potvrđena je
prisutnost jednog do tri plazmida koji su po tipu replikona većinom pripadali skupini
inkopatibilnosti IncL/M.

Abstract (english)

Bacterial resistance to antibiotics poses a significant threat to human health due to limited
treatment options in the treatment of infectious diseases. Third-generation cephalosporins are
antibiotics used to treat many infections, but Gram-negative bacteria can successfully hydrolyze
them using extended-spectrum β-lactamases (ESBL), whose genes are usually found on
plasmids. Wastewater treatment plants (WWTPs) play an important role in the spread of this
resistance because bacteria there exchange resistance genes via plasmids under the pressure of
antibiotics and other contaminants. In this study, the exogenous plasmid isolation method was
used to analyze the transfer of ESBL genes from bacteria in treated wastewater from seven
WWTPs to the recipient bacterium, Escherichia coli CV601. Eighteen positive transconjugants
(E. coli CV601 with ESBL gene on plasmid) were obtained from five WWTPs studied. Most
of the transconjugants exhibited a multidrug-resistant phenotype, which was confirmed by disk
diffusion and microdilution methods. PCR and agarose gel electrophoresis confirmed the
presence of the ESBL genes blaCTX-M-1 and blaTEM on all plasmids of the positive
transconjugants, with the additional detection of the blaGES gene on one plasmid. Eight plasmids
also carried the carbapenemase genes blaVIM or blaNDM. Using the polymerase chain reactionbased replicon typing (PBRT) method, the presence of one to three plasmids belonging mainly
to the IncL/M incompatibility group was confirmed for all positive transconjugants.

• Keywords: plazmidi; horizontalni prijenos gena; β-laktamaze proširenog spektra; otpadna voda; karbapenemaze; cefalosporini treće generacije
• Keywords (english): plasmids; horizontal gene transfer; extended-spectrum β-lactamases; wastewater; carbapenemases; 3rd generation cephalosporins
• Language: croatian
• URN:NBN: urn:nbn:hr:217:701563
• Project: Number: IP-2019-04-5539; Title: Antibiotička rezistencija u uređajima za obradu otpadnih voda u Hrvatskoj: naglasak na ß-laktamaze proširenog spektra i karbapenemaze; Title: Antibiotic Resistance in Wastewater Treatment Plants in Croatia: Focus on Extended-Spectrum ß-Lactamases and Carbapenemases; Acronym: WasteCare; Leader: Nikolina Udiković Kolić; Jurisdiction: Croatia; Funder: HRZZ; Funding stream: IP
• Study programme: Title: Ecology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra ekologije i zaštite prirode (magistar/magistra ekologije i zaštite prirode)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2023-10-25 08:34:47