title: Contributions of tropodithietic acid and biofilm formation to the probiotic activity of Phaeobacter inhibens.
abstract: BACKGROUND
  The probiotic bacterium Phaeobacter inhibens strain S4Sm, isolated from the inner shell surface of a healthy oyster, secretes the antibiotic tropodithietic acid (TDA), is an excellent biofilm former, and increases oyster larvae survival when challenged with bacterial pathogens. In this study, we investigated the specific roles of TDA secretion and biofilm formation in the probiotic activity of S4Sm.
  
  
  RESULTS
  Mutations in clpX (ATP-dependent ATPase) and exoP (an exopolysaccharide biosynthesis gene) were created by insertional mutagenesis using homologous recombination. Mutation of clpX resulted in the loss of TDA production, no decline in biofilm formation, and loss of the ability to inhibit the growth of Vibrio tubiashii and Vibrio anguillarum in co-colonization experiments. Mutation of exoP resulted in a ~60% decline in biofilm formation, no decline in TDA production, and delayed inhibitory activity towards Vibrio pathogens in co-colonization experiments. Both clpX and exoP mutants exhibited reduced ability to protect oyster larvae from death when challenged by Vibrio tubiashii. Complementation of the clpX and exoP mutations restored the wild type phenotype. We also found that pre-colonization of surfaces by S4Sm was critical for this bacterium to inhibit pathogen colonization and growth.
  
  
  CONCLUSIONS
  Our observations demonstrate that probiotic activity by P. inhibens S4Sm involves contributions from both biofilm formation and the production of the antibiotic TDA. Further, probiotic activity also requires colonization of surfaces by S4Sm prior to the introduction of the pathogen.
authors.affiliation: Department of Cell and Molecular Biology, University of Rhode Island, 120 Flagg Rd., Kingston, RI, 02881, USA. wenjing_zhao@hms.harvard.edu.|Present Address: Department of Microbiology and Immunology, Harvard Medical School, Boston, MA 02115, USA. wenjing_zhao@hms.harvard.edu.
authors.forename: Wenjing
authors.initials: W
authors.lastname: Zhao
authors.affiliation: Biomedical and Pharmaceutical Sciences, University of Rhode Island, Kingston, RI, 02881, USA. Christine.Dao@umassd.edu.|Present Address: Department of Chemistry and Biochemistry, University of Massachusetts Dartmouth, Darmouth, MA 02747, USA. Christine.Dao@umassd.edu.
authors.forename: Christine
authors.initials: C
authors.lastname: Dao
authors.affiliation: Fisheries, Animal and Veterinary Sciences, University of Rhode Island, Kingston, RI, 02881, USA. murnimarlina@upm.edu.my.|Present Address: Department of Aquaculture, Faculty of Agriculture, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia. murnimarlina@upm.edu.my.
authors.forename: Murni
authors.initials: M
authors.lastname: Karim
authors.affiliation: Fisheries, Animal and Veterinary Sciences, University of Rhode Island, Kingston, RI, 02881, USA. gomezchi@uri.edu.
authors.forename: Marta
authors.initials: M
authors.lastname: Gomez-Chiarri
authors.affiliation: Biomedical and Pharmaceutical Sciences, University of Rhode Island, Kingston, RI, 02881, USA. drowley@uri.edu.
authors.forename: David
authors.initials: D
authors.lastname: Rowley
authors.affiliation: Department of Cell and Molecular Biology, University of Rhode Island, 120 Flagg Rd., Kingston, RI, 02881, USA. dnelson@uri.edu.
authors.forename: David R
authors.initials: DR
authors.lastname: Nelson
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D018441:Biofilms; D010125:Ostreidae / Q000254:growth & development / Q000382:microbiology*; D019936:Probiotics / Q000008:administration & dosage* / Q000378:metabolism; D041902:Rhodobacteraceae / Q000235:genetics / Q000502:physiology*; D014334:Tropolone / Q000031:analogs & derivatives* / Q000378:metabolism; D014733:Vibrio / Q000502:physiology; D014735:Vibrio Infections / Q000188:drug therapy / Q000382:microbiology / Q000662:veterinary*
chemical_list: D001426:Bacterial Proteins; C527336:tropodithietic acid; D014334:Tropolone