Hy yest

The strains used in this study are described in Table 1. For general maintenance, K. pneumoniae and P. aeruginosa were grown at 37°C overnight in Hy-Soy (HS) bacteriological media (5 g/L sodium chloride, 10 g/L soytone [Teknova, CA], 5 g/L Hy-yest [Sigma Aldrich, MO]) at 37°C. 1.5% bacteriological agar (AmericanBio, MA) was added to HS for solid media preparations. Fully chemically defined medium (CDM) used for growth of KP in fermentation cultures and to assess guanine auxotrophy has been described previously [19 (link)]. 0.004%-0.025% guanine (Sigma Aldrich, MO) was added for the growth of CVD 3001 to supplement the guaBA mutation. O types were determined by PCR with extracted genomic DNA as described [14 (link)]. K types were determined by sequencing of the wzi or wzc genes, or multiplex PCR [20 (link)–22 (link)].

The strains used in this study are described (S1 Table). All strains were maintained in Hi-Soy (HS) bacteriological media (5 g/L sodium chloride, 10 g/L soytone [Teknova, CA], 5 g/L Hy-yest [Sigma Aldrich, MO]) at 37°C. Growth and preparation of bacteria for in-vitro analyses and in-vivo infection was conducted as described [18 (link)]. Growth media for all guaBA mutants were supplemented with guanine; kanamycin was additionally supplemented for CVD 1925 (pSEC10-wzzB) (S1 Table). STm CVD 1925 (with or without pSEC10-wzzB) (S1 Table) and CVD 1943 (S1 Table) were grown in fully chemically defined media (CDM) in a fermenter. STm D65 (S1 Table) was grown in shake flasks. Fermentation conditions were as follows: 50 mL of CDM supplemented with 0.004% guanine was inoculated with 3–5 colonies from an HS agar plate and grown for 12–18 h at 37°C in a shake flask with agitation at 80 rpm. This culture was then used to inoculate 500 mL of CDM supplemented with 0.004% guanine that was grown under equivalent conditions for 8–10 h. Four liters of CDM containing 0.025% guanine was then inoculated to an OD₆₀₀ nm of 0.15 with the 500 mL shake flask and maintained in a Biostat A-plus fermenter (Sartorius, Germany) culture, for 18–24 h at 400 rpm, 5 LPM ambient air, with an adjustment to pH 7 using 28% ammonium hydroxide.

The strains used in this study are described in Table 1. All strains were grown in Hi-Soy (HS) bacteriological media (5 g/L sodium chloride, 10 g/L soytone [Teknova, Hollister, CA], 5 g/L Hy-yest [Sigma Aldrich, St. Louis, MO]) at 37°C.