Dulbeccos modified eagle s media dmem

BS-C-1 cells (ATCC CCL 26) were grown in Dulbeccos’ modified eagle’s media DMEM (Lonza, Basel, Switzerland) containing 10% fetal calf serum (FCS) (Hyclone III, Logan, UT, USA), 2 mM l-glutamine (GIBCO, Grand Island, NY, USA), 100 U/mL penicillin (GIBCO), and 100 μg/mL streptomycin (GIBCO). Virus was purified through a sucrose cushion as described elsewhere [33 ]. Animals were sacrificed and bled by heart-sticks to determine viral titers. Tissues were removed and ground in phosphate buffered saline (PBS) (10% FBS w/v). Footpads were removed with a surgical blade (Henry Schein, Melville, NY, USA) and processed in the same manner as tissue. Following grinding, samples were freeze–thawed thrice interrupted by 20 s sonications. Virus infectivity was calculated by titration on BS-C-1 monolayers for seven days (TATV) or four days (ECTV) at 37 °C [34 (link)]. Plaques were visualized by addition of 0.5 mL 0.3% crystal violet/10% formalin to each well. Arithmetic means above the limit of detection (1 × 10² PFU/mL) were calculated as plaque forming units (PFU)/g or PFU/mL [35 (link)].
TATV was a gift from Geoffrey Smith (Imperial College, London, UK). The virus was isolated from a wild gerbil (Tatera Kempi) caught in Dahomey (now Republic of Benin), Africa in 1968 [28 (link)]. Plaque purified isolates of TATV, ECTV-MOS and VACV-COP were propagated in BS-C-1 cells.

The bacterial strains E. coli ATCC 25922, Shiga-toxin producing E. coli (STEC) O91 EC20010076, Mh 276, Mh 13, Mh 136, Mh 535, Mh 587, and Mh 330 were used in the study (Table 2 [23 (link),24 (link),38 (link)]). Glycerol stocks of isolates were streaked onto tryptic soy agar (TSA; EMD Millipore Corporation, Billerica, MA, USA) supplemented with 5% defibrinated sheep blood (QUAD FIVE, Ryegate, MT, USA) and incubated at 37 °C in the presence of 5% CO₂ for 20–24 h. A single colony was inoculated into 10 mL Mueller Hinton broth (MHB; Oxoid, Basingstoke, Hants, UK) and incubated at 37 °C with shaking at 180 rpm for 20–22 h. Bovine turbinate cells (BTs) were grown on Dulbecco’s Modified Eagle’s Media (DMEM) (BioWhittaker, Walkersville, MD, USA) with phenol red, 10% fetal bovine serum (Gibco by Life Technologies, Grand Island, NY, USA), and 1% Antibiotic-Antimycotic (Gibco, Life Technologies, Grand Island, NY, USA) at 37 °C with 5% CO₂.