The influence of competition on the formation of phage resistance in Flavobacterium columnare
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2021Copyright
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Luonnossa bakteerit elävät monimutkaisissa eliöyhteisöissä, joissa ne vuorovaikuttavat paitsi keskenään, myös bakteereja infektoivien virusten, (bakterio)faagien kanssa. Bakteerien ja faagien välisen koevoluution aikana bakteereille on kehittynyt useita resistenssimekanismeja faagien torjumiseen. Tällaisia mekanismeja ovat muun muassa CRISPR-Cas-välitteinen ja solun pinnan muutokseen perustuva immuniteetti. Kyseisten mekanismien on aiemmissa tutkimuksissa todettu olevan myös kaloja infektoivan Flavobacterium columnare -bakteerin keino torjua faagi-infektioita. Vaikka resistenssimekanismeja onkin tutkittu laajalti, kilpailun vaikutuksia resistenssin kehittymiseen ei vielä juuri tunneta. Tämän tutkimuksen tavoitteena oli saada lisää tietoa resistenssin tyypistä, kun F. columnaren annettiin vuorovaikuttaa Aeromonas sp.- ja Eshcerichia coli -bakteerien kanssa. Uusien CRISPR-toistojaksojen havaitsemiseen käytettiin PCR-menetelmää ja geelielektroforeesia, ja pesäkkeiden morfologiaa tarkasteltiin viljelymaljoilta neljänä ajankohtana 56 vuorokauden aikana. Tulokset näyttivät, että molempien tunnettujen lokusten CRISPR-toistojaksojen määrä kasvoi selvästi kokeen aikana. Faagille alttiiden pesäkkeiden määrä kasvoi, kun taas immuunien pesäkkeiden määrässä havaittiin vastaava lasku. Aeromonas sp:n kanssa vuorovaikuttaneella F. columnarella havaittiin enemmän CRISPR-toistojaksoja kuin kontrolliviljelmissä, mikä viittaa siihen, että vuorovaikutus muiden bakteerien kanssa parantaa CRISPR-immuniteetin muodostumista F. columnarella. Nämä tulokset korostavat luonnollisissa eliöyhteisöissä tehtävien tutkimusten tärkeyttä.
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Bacteria live in complex communities, where they interact with other bacteria and also bacterial viruses, bacteriophages (phages). During the long coevolution between bacteria and phages, bacteria have developed various immunity mechanisms against phage infections. These mechanisms include CRISPR-Cas and surface modification, which have both been found to be the mechanisms utilized by a fish pathogen, Flavobacterium columnare. Although the development of resistance mechanisms has been widely studied, the effect of other bacterial species to their appearance are not yet known. This study aimed to find information about the type of resistance taking place in F. columnare in the presence of Aeromonas sp. and Escherichia coli using a culture condition known to favor CRISPR activity. The study used PCR and gel electrophoresis to detect the acquisition of new spacers in F. columnare CRISPR arrays, besides checking colony morphology changes at four timepoints (7, 14, 32 and 56 days). The results showed a clear increase in the number of new spacers in both CRISPR loci. The abundance of wild-type -like rhizoid colonies increased slightly during the experiment, whereas the typically phage resistant rough colonies had a similar decrease. The number of acquired CRISPR spacers in the presence of Aeromonas sp. was higher than in the control culture, indicating a connection between the presence of a naturally coexisting bacterium and more competent spacer acquisition in F. columnare. These data shows that the occurrence of other bacterial species can influence F. columnare strategies against phages, highlighting the importance of studies made in natural communities.
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