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Browsing by Subject "faagi"

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  • Pankka, Salla (2023)
    The objective of this thesis was to isolate and characterize new bacteriophages (phages) against clinical Klebsiella pneumoniae strains for phage therapy. K. pneumoniae is causing an emerging threat to global health due to its broad antibiotic resistance profile and hypervirulent strains. New treatment options are urgently needed to defeat the crisis. Phage therapy could provide one option to treat multiresistant K. pneumoniae infections. In this thesis, five new phages were isolated and characterized from Finnish wastewater and Georgian river water against two clinical K. pneumoniae strains. The three phages from Georgian river water, fMtkKpn01, fMtkKpn03, and fMtkKpn04, resembled Drulisviruses based on phylogenetic analysis. The two phages from Finnish wastewater, fJoKpn03 and fJoKpn05 were phylogenetically distinct. fJoKpn03 couldn’t be classified. fJoKpn05 resembled Weberviruses. Based on sequence analysis, none of the phage genomes included any harmful genes that would prevent their use in phage therapy. All phages demonstrated a 6-hour total inhibition to host bacterial growth. Their host range was determined to be narrow, only infecting their respective host strains from the 80 bacterial strains tested. All the phages tolerated high pH well. fJoKpn03 was the only one tolerating very low pH. All phages showed a synergistic effect on the inhibition of bacterial growth when applied together with piperacillin. In conclusion, all five phages proved potential for phage therapy. They demonstrated inhibitory action against K. pneumoniae strains with capsule types against which there previously were no phages in our collection. Due to their narrow host range, they could be suited for personalized phage therapy or used in combination therapy with antibiotics to increase efficacy and duration of action. fJoKpn03 could provide an opportunity for oral administration due to its broad pH stability profile.
  • Mustonen, Markus (2024)
    The increase of antibiotic resistance is one of the major healthcare threats globally. One potential way to battle against antibiotic resistant bacterial infections is to treat them with the natural opponents of bacteria, bacteriophages, known as phage therapy. The aim of this thesis was to identify new bacteriophages against clinically notable bacterial species such as Escherichia coli, Burkholderia cepacia, Enterococcus faecalis and Enterococcus faecium. Bacteriophages were screened from various origins such as hospital sewage samples, soil samples and manure samples, collected in between 2019 and 2022. The isolated bacteriophages were then initially characterized to evaluate their potential use in phage therapy. In this thesis, two phages (fHo-Eco16, fHo-Eco17) against clinical E. coli isolate and one phage (fHo-Efa06) against clinical E. faecalis isolate were found from the recently collected Finnish hospital sewage sample pool. Both E. coli phages were classified as Felixounaviruses belonging to family of Ounavirinae and class of Caudoviricetes. Enterococcus phage fHo-Efa06 was characterized as Saphexavirus belonging to class of Caudoviricetes. Preliminary genome annotation did not reveal any characteristics of lysogenic lifecycle, or antibiotic resistance or bacterial toxin genes, which would prevent the use of phages in phage therapy. Both E. coli phages (fHo-Eco16, fHo-Eco17) showed narrow host range infecting only the primary host bacterial isolate but none of 29 other tested clinical E. coli isolates. Phage fHo-Efa06 showed relatively broad host range properties infecting nine tested E. faecalis isolates out of 20 tested E. faecalis isolates but no infection capabilities against six tested clinical E. faecium isolates. In conclusion, freshly collected hospital sewage seemed to be optimal environment to find bacteriophages against clinical bacterial isolates. Furthermore, phages fHo-Eco16, fHo-Eco17 and fHo-Efa06 did not display any strictly unsuitable properties which could prevent their use in phage therapy. In turn, to obtain the definitive certainty on the usability of the phages in therapeutic use, in-depth host range screening together with detailed functional and structural annotation for the phage genomes of fHo-Efa06, fHo-Eco16 and fHo-Eco17 should be completed.
  • Trivedi, Milla-Maaria (2020)
    Tämä maisterintutkielma on osa tutkimusohjelmaa, jossa on tarkoitus tunnistaa uusia antibioottien vaikutuskohteita bakteereissa. Tavoitteena oli pystyttää Haartman-instituutissa molekyylibiologinen menetelmä, jolla voitaisiin selvittää faagigeenien tuottamien proteiinien toksisuutta. Menetelmän kehityksessä korostui tarve pystyä analysoimaan tehokkaasti mahdollisimman monta tuntematonta faagigeeniä ja niiden toimintaa. Työssä kehitetty menetelmä perustui bioluminesenssin käyttöön toksisuuden havaitsemiseksi. Ensimmäisessä vaiheessa valmistettiin plasmidi, joka sisälsi luxAB-geenit. Työn toisessa vaiheessa muodostettiin itsemurhavektori, joka sisälsi luxCDE-geenit. Itsemurhavektorin sisältämät luxCDE-geenit integroitiin bakteerin genomiseen deoksiribonukleiinihappoon (gDNA), jonka jälkeen luxAB-geenit sisältävä plasmidi voitiin elektroporoida kyseiseen bakteeriin. Yhdessä nämä muodostivat geeniyhdistelmän luxCDABE, joka pystyi tuottamaan valoa bioluminesenssin avulla. Jos luxAB-plasmidiin kloonattavan geenin tuote on toksinen, eivät plasmidin saaneet transformantit elektroporaation jälkeen säily hengissä, minkä seurauksena transformantit eivät tuota valoa, kun taas, jos geenin tuote ei ole toksinen, tapahtuu valon tuottoa. Toisin sanoen, luminesenssin puuttuessa menetelmä toimisi halutulla tavalla eli indikoisi geenien tuottamien proteiinien toksisuutta. Menetelmän toimivuutta testattiin käyttämällä tunnettuja bakteerikantoja, jotka sisälsivät luxCDE-geenikasetin. Tarkoituksena oli varmistaa luxAB-geenien toimivuus komplementoida minkä tahansa bakteerin luxCDE-geenit. Tarkoitus oli saada aikaan menetelmä, jolla voidaan nopeasti ja tehokkaasti selvittää tuntemattomien faagigeenien toimintaa. Bioluminesenssin tuottama valo erottuu selkeästi ja menetelmä on helposti toistettavissa. Työssä kehitettyä luxAB-geenit sisältävää plasmidia voidaan jatkotutkimuksissa testata ensin tunnetuilla kontrolligeeneillä ja lopuksi käyttää toksisia proteiineja tuottavien faagigeenien tunnistamisessa.