Bacteriophage Resistance Mechanisms in the Fish Pathogen Flavobacterium psychrophilum: Linking Genomic Mutations to Changes in Bacterial Virulence Factors

Daniel Castillo, Rói Hammershaimb Christiansen, Inger Dalsgaard, Lone Madsen, Mathias Middelboe

    Research output: Contribution to journalJournal articleResearchpeer-review

    Abstract

    Flavobacterium psychrophilum is an important fish pathogen in salmonid aquaculture worldwide. Due to increased antibiotic resistance, pathogen control using bacteriophages has been explored as a possible alternative treatment. However, the effective use of bacteriophages in pathogen control requires overcoming the selection for phage resistance in the bacterial populations. Here, we analyzed resistance mechanisms in F. psychrophilum after phage exposure using whole-genome sequencing of the ancestral phage-sensitive strain 950106-1/1 and six phage-resistant isolates. The phage-resistant strains had all obtained unique insertions and/or deletions and point mutations distributed among intergenic and genic regions. Mutations in genes related to cell surface properties, gliding motility, and biosynthesis of lipopolysaccharides and cell wall were found. The observed links between phage resistance and the genetic modifications were supported by direct measurements of bacteriophage adsorption rates, biofilm formation, and secretion of extracellular enzymes, which were all impaired in the resistant strains, probably due to superficial structural changes. The clustered regularly interspaced short palindromic repeat (CRISPR) region was unaffected in the resistant isolates and thus did not play a role as a resistance mechanism for F. psychrophilum under the current conditions. All together, the results suggest that resistance in F. psychrophilum was driven by spontaneous mutations, which were associated with a number of derived effects on the physiological properties of the pathogen, including reduced virulence under in vitro conditions. Consequently, phage-driven physiological changes associated with resistance may have implications for the impact of the pathogen in aquaculture, and these effects of phage resistance on host properties are therefore important for the ongoing exploration of phage-based control of F. psychrophilum.
    Original languageEnglish
    JournalAPPLIED AND ENVIRONMENTAL MICROBIOLOGY
    Volume81
    Issue number3
    Pages (from-to)1157-1167
    Number of pages11
    ISSN0099-2240
    DOIs
    Publication statusPublished - 2015

    Keywords

    • Microbial Ecology

    Cite this

    @article{79616553a3364226841ac237e7f1d5a5,
    title = "Bacteriophage Resistance Mechanisms in the Fish Pathogen Flavobacterium psychrophilum: Linking Genomic Mutations to Changes in Bacterial Virulence Factors",
    abstract = "Flavobacterium psychrophilum is an important fish pathogen in salmonid aquaculture worldwide. Due to increased antibiotic resistance, pathogen control using bacteriophages has been explored as a possible alternative treatment. However, the effective use of bacteriophages in pathogen control requires overcoming the selection for phage resistance in the bacterial populations. Here, we analyzed resistance mechanisms in F. psychrophilum after phage exposure using whole-genome sequencing of the ancestral phage-sensitive strain 950106-1/1 and six phage-resistant isolates. The phage-resistant strains had all obtained unique insertions and/or deletions and point mutations distributed among intergenic and genic regions. Mutations in genes related to cell surface properties, gliding motility, and biosynthesis of lipopolysaccharides and cell wall were found. The observed links between phage resistance and the genetic modifications were supported by direct measurements of bacteriophage adsorption rates, biofilm formation, and secretion of extracellular enzymes, which were all impaired in the resistant strains, probably due to superficial structural changes. The clustered regularly interspaced short palindromic repeat (CRISPR) region was unaffected in the resistant isolates and thus did not play a role as a resistance mechanism for F. psychrophilum under the current conditions. All together, the results suggest that resistance in F. psychrophilum was driven by spontaneous mutations, which were associated with a number of derived effects on the physiological properties of the pathogen, including reduced virulence under in vitro conditions. Consequently, phage-driven physiological changes associated with resistance may have implications for the impact of the pathogen in aquaculture, and these effects of phage resistance on host properties are therefore important for the ongoing exploration of phage-based control of F. psychrophilum.",
    keywords = "Microbial Ecology",
    author = "Daniel Castillo and Christiansen, {R{\'o}i Hammershaimb} and Inger Dalsgaard and Lone Madsen and Mathias Middelboe",
    year = "2015",
    doi = "10.1128/AEM.03699-14",
    language = "English",
    volume = "81",
    pages = "1157--1167",
    journal = "Applied and Environmental Microbiology",
    issn = "0099-2240",
    publisher = "American Society for Microbiology",
    number = "3",

    }

    Bacteriophage Resistance Mechanisms in the Fish Pathogen Flavobacterium psychrophilum: Linking Genomic Mutations to Changes in Bacterial Virulence Factors. / Castillo, Daniel; Christiansen, Rói Hammershaimb; Dalsgaard, Inger; Madsen, Lone; Middelboe, Mathias.

    In: APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Vol. 81, No. 3, 2015, p. 1157-1167.

    Research output: Contribution to journalJournal articleResearchpeer-review

    TY - JOUR

    T1 - Bacteriophage Resistance Mechanisms in the Fish Pathogen Flavobacterium psychrophilum: Linking Genomic Mutations to Changes in Bacterial Virulence Factors

    AU - Castillo, Daniel

    AU - Christiansen, Rói Hammershaimb

    AU - Dalsgaard, Inger

    AU - Madsen, Lone

    AU - Middelboe, Mathias

    PY - 2015

    Y1 - 2015

    N2 - Flavobacterium psychrophilum is an important fish pathogen in salmonid aquaculture worldwide. Due to increased antibiotic resistance, pathogen control using bacteriophages has been explored as a possible alternative treatment. However, the effective use of bacteriophages in pathogen control requires overcoming the selection for phage resistance in the bacterial populations. Here, we analyzed resistance mechanisms in F. psychrophilum after phage exposure using whole-genome sequencing of the ancestral phage-sensitive strain 950106-1/1 and six phage-resistant isolates. The phage-resistant strains had all obtained unique insertions and/or deletions and point mutations distributed among intergenic and genic regions. Mutations in genes related to cell surface properties, gliding motility, and biosynthesis of lipopolysaccharides and cell wall were found. The observed links between phage resistance and the genetic modifications were supported by direct measurements of bacteriophage adsorption rates, biofilm formation, and secretion of extracellular enzymes, which were all impaired in the resistant strains, probably due to superficial structural changes. The clustered regularly interspaced short palindromic repeat (CRISPR) region was unaffected in the resistant isolates and thus did not play a role as a resistance mechanism for F. psychrophilum under the current conditions. All together, the results suggest that resistance in F. psychrophilum was driven by spontaneous mutations, which were associated with a number of derived effects on the physiological properties of the pathogen, including reduced virulence under in vitro conditions. Consequently, phage-driven physiological changes associated with resistance may have implications for the impact of the pathogen in aquaculture, and these effects of phage resistance on host properties are therefore important for the ongoing exploration of phage-based control of F. psychrophilum.

    AB - Flavobacterium psychrophilum is an important fish pathogen in salmonid aquaculture worldwide. Due to increased antibiotic resistance, pathogen control using bacteriophages has been explored as a possible alternative treatment. However, the effective use of bacteriophages in pathogen control requires overcoming the selection for phage resistance in the bacterial populations. Here, we analyzed resistance mechanisms in F. psychrophilum after phage exposure using whole-genome sequencing of the ancestral phage-sensitive strain 950106-1/1 and six phage-resistant isolates. The phage-resistant strains had all obtained unique insertions and/or deletions and point mutations distributed among intergenic and genic regions. Mutations in genes related to cell surface properties, gliding motility, and biosynthesis of lipopolysaccharides and cell wall were found. The observed links between phage resistance and the genetic modifications were supported by direct measurements of bacteriophage adsorption rates, biofilm formation, and secretion of extracellular enzymes, which were all impaired in the resistant strains, probably due to superficial structural changes. The clustered regularly interspaced short palindromic repeat (CRISPR) region was unaffected in the resistant isolates and thus did not play a role as a resistance mechanism for F. psychrophilum under the current conditions. All together, the results suggest that resistance in F. psychrophilum was driven by spontaneous mutations, which were associated with a number of derived effects on the physiological properties of the pathogen, including reduced virulence under in vitro conditions. Consequently, phage-driven physiological changes associated with resistance may have implications for the impact of the pathogen in aquaculture, and these effects of phage resistance on host properties are therefore important for the ongoing exploration of phage-based control of F. psychrophilum.

    KW - Microbial Ecology

    U2 - 10.1128/AEM.03699-14

    DO - 10.1128/AEM.03699-14

    M3 - Journal article

    VL - 81

    SP - 1157

    EP - 1167

    JO - Applied and Environmental Microbiology

    JF - Applied and Environmental Microbiology

    SN - 0099-2240

    IS - 3

    ER -