E. huxleyi strains were purchased from the National Center for Marine Algae (NCMA) and the Roscoff Culture Collection (RCC) and maintained in filtered seawater (FSW). NCMA379, RCC1216, and NCMA373 were cultured in f/2 medium (-Si) (63), and NCMA2090 was cultured in k/2 medium (-tris, -Si) (64). Cultures were incubated at 18°C with a 16-hour light/8-hour dark illumination cycle. A light intensity of 100 μmol photons m−2 s−1 was provided by cool white light-emitting diode lights. Cultures were made axenic by the following treatment: Cells were gently washed with autoclaved FSW on sterile 1.2-μm nitrocellulose membrane filters (Millipore). Cells were transferred to algal growth media containing the following antibiotic mix: chloramphenicol (20 μg ml−1), polymyxin B (120 U ml−1), penicillin (40 U ml−1), and streptomycin (40 μg ml−1). After 7 days, the cultures were diluted into fresh algal growth media, and the antibiotics mix was replenished. After another 7 days, the cultures were diluted again into fresh algal growth media without antibiotics. For strains 1216, 373, and 2090, cultures were treated again with the following antibiotics mix: ampicillin (50 μg ml−1), streptomycin (25 μg ml−1), and chloramphenicol (5 μg ml−1). Cultures were transferred one to two times a week. After 2 weeks, the cultures recovered and no bacteria could be detected by flow cytometry (see full description in the following section) or by plating on marine agar 2216 plates. Cultures were maintained with antibiotics and were transferred every 7 to 10 days. Before infection, E. huxleyi cultures were transferred three to four times to antibiotic-free algal growth media. For all experiments, E. huxleyi cultures were infected at early exponential growth phase (2 × 105 to 4 × 105 cells ml−1). For CAM infection, algal cultures were inoculated with 104 bacteria ml−1. For Sulfitobacter D7 infection, bacteria were inoculated from a glycerol stock (kept at −80°C) into 1/2 YTSS [2 g of yeast extract, 1.25 g of tryptone, and 20 g of sea salts (Sigma-Aldrich) dissolved in 1 liter of double distilled water (DDW)] and grown overnight at 28°C at 150 rpm. Bacteria were washed three times in FSW by centrifugation (10,000g, 1 min). Algal cultures were inoculated at t = 0 days with 103 bacteria ml−1. In the experiment presented in Fig. 5, E. huxleyi 2090 cultures were inoculated with 106 bacteria ml−1. When noted, DMSP, glycerol, or propionate were added at t = 0 days. DMSP was synthesized according to Steinke et al. (26).

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