Lb agar

The ETEC strain EC43-Ph (EC43), previously serotyped as O9:H9, comprises the enterotoxin (STa) and two adhesins (F5 and F41). This diarrheagenic E. coli was recovered from a neonatal pig in a Spanish swine farm [3 (link)]. EC43 was cultivated in Lysogeny Broth (LB, NZYTech) or on LB agar (12 g L⁻¹, VWR), at 37 °C. The phage vB_EcoM_FJ1 (FJ1) and its host HFJ1 were provided by the company ALS. Although FJ1 has been isolated from chickens’ litter, from an enrichment with the avian pathogenic E. coli strain HFJ1 (non-published data), this phage infects EC43, a swine-originating E. coli strain. FJ1 was then propagated on a mid-log grown host strain, HFJ1, with a multiplicity of infection (MOI) of 0.01. After 6 h of incubation (37 °C), the culture was centrifuged, treated with chloroform (10% v/v) and filtered (0.2 µm). Phage concentration was assessed by plaque counting (PFU mL⁻¹) and phage stock was stored at 4 °C.

S. Typhimurium SL1344 was grown overnight in LB broth (Invitrogen) in a shaker at 37°C. CFUs were estimated by optical density at an absorbance (A) of 600 and confirmed by growth on LB agar (VWR) with 50ug/ml streptomycin (Sigma).
Mice 8–10 weeks of age were housed 2–4 mice per cage in disposable cages and fasted in the morning for 4 hours prior to infection in the afternoon. No pre-treatment with streptomycin or sodium bicarbonate was used. Mice were gavaged with 10⁹ CFU in 150ul 100mM HEPES buffer (pH = 8.0) (Fischer Scientific) and food was replaced. Mice were followed for 2 weeks for survival.
To measure colonization, mice were sacrificed on day 7 post-infection by isoflurane and cardiac puncture. The spleen, liver, cecum, and mesenteric lymph nodes were collected and weighed for determination of bacterial load. Briefly, tissues were collected into 1ml thioglycolate (USP Alternative) using sterile technique. A 1/8” steel bead was added to each sample, and samples dissociated using a TissueLyser (Qiagen) for 3 minutes at 30 Hz for 4 cycles. Samples were diluted and plated on LB agar with 50ug/ml streptomycin. Plates were incubated for 48 hours at 37°C and counted.
For IV Salmonella infection, the tail vein was injected with 200ul of PBS containing 10²⁻³ CFU and mice were followed daily for survival.

C. difficile and E. coli strains are described in Supplementary Table 4. E. coli strains were routinely grown at 37 °C in LB broth and on LB agar (VWR or Fisher Scientific). C. difficile strains were routinely grown under anaerobic conditions at 37 °C on brain heart infusion (BHI, Oxoid) or supplemented BHI (BHI-S)³⁵ agar and in TY broth. Growth media were supplemented with chloramphenicol (15 μg ml⁻¹), thiamphenicol (15 μg ml⁻¹) or kanamycin (50 μg ml⁻¹) as required.