Isolation and Cultivation of Marine Microbial Strains

The 827 strains tested in this study were previously isolated from the microbiota and faeces of marine invertebrates including anemones (A. sulcata and A. equina) and holothurians (H. tubulosa and H. forskali). The samples were collected and processed in a previous study at iMARE Natural S.L (http://www.imarenatural.com) aquaculture facilities located in Motril, Granada (36°44′33.4″N 3°31′12.1″W), in southern Spain [42 (link)]. All marine strains used were routinely grown at 28 °C in marine broth (MB, BD Difco^®, Franklin Lakes, NJ, USA). The human-related pathogen P. aeruginosa PAO1 was cultured in LB medium at 37 °C. C. violaceum ATCC 12472 [93 (link)], C. violaceum CV026 [94 (link)], and C. violaceum VIR07 [93 (link)] were grown at 28 °C in LB medium. In the case of strains CV026 and VIR07, the medium was supplemented with kanamycin (50 µg mL⁻¹). The synthetic AHLs used were C6-HSL (N-hexanoyl-dl-homoserine lactone) and C10-HSL (N-decanoyl-dl-homoserine lactone) (Sigma^®, Madrid, Spain).