Quantifying Chlamydia Muridarum Infection

For systemic infection, mice were injected intravenously in the lateral tail vein with 1×10⁵C. muridarum. To enumerate the bacteria burden in tissues, the spleen, liver and kidney, were crushed in 5 mL SPG buffer and tissue homogenate was placed in a tube with glass beads to disrupt cells. After shaking for 5 min, and centrifugation at 500 g for 10 minutes, supernatants were collected and serial dilutions were plated on HeLa 229 cells. For intravaginal infections, estrus was synchronized by subcutaneous injection of 2.5 mg medroxyprogesterone acetate (Greenstone, NJ) 7 days before infection. 1×10⁵ C. muridarum in 5 µL SPG buffer were then deposited into vaginal vaults. To enumerate bacteria, vaginal swabs were collected, shaken with glass beads, and serial dilutions were plated on HeLa 229 cells.