title: Efflux pump-deficient mutants as a platform to search for microbes that produce antibiotics.
abstract: Pseudomonas putida DOT-T1E-18 is a strain deficient in the major antibiotic efflux pump (TtgABC) that exhibits an overall increased susceptibility to a wide range of drugs when compared with the wild-type strain. We used this strain as a platform to search for microbes able to produce antibiotics that inhibit growth. A collection of 2400 isolates from soil, sediments and water was generated and a drop assay developed to identify, via growth inhibition halos, strains that prevent the growth of DOT-T1E-18 on solid Luria-Bertani plates. In this study, 35 different isolates that produced known and unknown antibiotics were identified. The most potent inhibitor of DOT-T1E-18 growth was an isolate named 250J that, through multi-locus sequence analysis, was identified as a Pseudomonas sp. strain. Culture supernatants of 250J contain four different xantholysins that prevent growth of Gram-positive bacteria, Gram-negative and fungi. Two of the xantholysins were produced in higher concentrations and purified. Xantholysin A was effective against Bacillus, Lysinibacillus and Rhodococcus strains, and the effect against these microbes was enhanced when used in combination with other antibiotics such as ampicillin, gentamicin and kanamycin. Xantholysin C was also efficient against Gram-positive bacteria and showed an interesting antimicrobial effect against Pseudomonas strains, and a synergistic inhibitory effect with ampicillin, chloramphenicol and gentamicin.
authors.affiliation: Department of Environmental Protection, Consejo Superior de Investigaciones Científicas, E-18008, Granada, Spain.
authors.forename: Carlos
authors.initials: C
authors.lastname: Molina-Santiago
authors.affiliation: Department of Environmental Protection, Consejo Superior de Investigaciones Científicas, E-18008, Granada, Spain.
authors.forename: Zulema
authors.initials: Z
authors.lastname: Udaondo
authors.affiliation: Department of Environmental Protection, Consejo Superior de Investigaciones Científicas, E-18008, Granada, Spain.
authors.forename: Abdelali
authors.initials: A
authors.lastname: Daddaoua
authors.affiliation: Bio-Iliberis R&D, Poligono Juncaril, calle Capileira nº 7, E-18121, Peligros, Granada, Spain.
authors.forename: Amalia
authors.initials: A
authors.lastname: Roca
authors.affiliation: Fundación MEDINA, Centro de Excelencia en Investigación de Medicamentos Innovadores en Andalucía. Avda. del Conocimiento 3, Parque Tecnológico de Ciencias de la Salud, E-18100, Armilla, Granada, Spain.
authors.forename: Jesús
authors.initials: J
authors.lastname: Martín
authors.affiliation: Fundación MEDINA, Centro de Excelencia en Investigación de Medicamentos Innovadores en Andalucía. Avda. del Conocimiento 3, Parque Tecnológico de Ciencias de la Salud, E-18100, Armilla, Granada, Spain.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Pérez-Victoria
authors.affiliation: Fundación MEDINA, Centro de Excelencia en Investigación de Medicamentos Innovadores en Andalucía. Avda. del Conocimiento 3, Parque Tecnológico de Ciencias de la Salud, E-18100, Armilla, Granada, Spain.
authors.forename: Fernando
authors.initials: F
authors.lastname: Reyes
authors.affiliation: Department of Environmental Protection, Consejo Superior de Investigaciones Científicas, E-18008, Granada, Spain.
authors.forename: Juan-Luis
authors.initials: JL
authors.lastname: Ramos
mesh_terms: D000900:Anti-Bacterial Agents / Q000302:isolation & purification / Q000378:metabolism*; D000898:Antibiosis; D001407:Bacillus; D001419:Bacteria / Q000302:isolation & purification / Q000378:metabolism*; D001693:Biological Transport, Active; D004357:Drug Synergism; D004783:Environmental Microbiology; D005658:Fungi; D006801:Humans; D026901:Membrane Transport Proteins / Q000172:deficiency*; D008826:Microbial Sensitivity Tests; D011549:Pseudomonas / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development
chemical_list: D000900:Anti-Bacterial Agents; D026901:Membrane Transport Proteins