Bacterial viability and direct contact between bacteria and the host cells affect upregulation of EGR1. Induction of EGR1 was monitored by qPCR after infection of the host epithelial cells with live or dead bacteria. Dead bacteria were obtained by heat treatment at 95°C for 10 min. Viable counts were used to ensure complete killing of the bacteria. The role of bacterial contact with the epithelial cells in the EGR1 induction was studied using a 0.4 μm Millicell filter, which helps to physically separate the bacteria and host cells, but still allows diffusion of secreted factors in the cell growth media (Sup.). (A) Pharyngeal FaDu cells infected with N. meningitidis (Nm-C). (B) Pharyngeal FaDu cells infected with S. pyogenes (Sp-M6). (C) Gastric AGS cells infected with H. pylori (Hp-J99). (D) Cervical ME180 cells infected with N. gonorrhoeae (Ng). (E) Intestinal Caco-2 cells infected with S. Enteritidis (SE3439). Bacteria were added to a MOI of 100 in all experiments. The data was analyzed at 2 h post infection for Nm-C, Sp-M6, SE3439, and Hp-J99. The data was analyzed at 4 h post infection of Ng.