Cholangitis: Bacterial virulence factors that facilitate cholangiovenous reflux and tumor necrosis factor-alpha production

In previous studies we noted that biliary bacteria produce slime and possess P1-fimbriae. The presence of gram-negative bacteria killed by complement correlated with serious biliary infections and induced more tumor necrosis factor-alpha (TNF-α) production in sera, suggesting a role for cytokine production and complement activation in biliary sepsis. This study examined bacterial virulence factors that facilitate cholangiovenous reflux (CVR) and TNF-α production in a rat model. Twenty-one biliary bacteria and two stool isolates were tested for slime production, sensitivity to complement killing, and hemolysin production. 107 Bacterial colony-forming units/ml (or saline control) were injected retrograde into the common bile ducts of Sprague-Dawley rats at a pressure of 30 cm H2O. Blood was obtained at 5 and 60 minutes after infusion for bacterial culture and TNF-α assay, respectively. The magnitude of slime production correlated inversely with the magnitude of bacterial CVR. Average bacterial colony-forming units were 1.4 x 105, 6.8 × 104, or 2.1 × 103 for bacteria with slime production 0 to 10, 11 to 99, or more than 100, respectively (P < 0.0001, analysis of variance). CVR was greater for serum-resistant bacteria (1.2 × 105 vs. 5.5 × 104 [P = 0.007, resistant vs. sensitive]), but TNF-α production was greater in serum-sensitive bacteria. TNF-α production as a function of bacterial reflux followed a logarithmic curve (R2 = 0.75) for serum-sensitive bacteria but was linear (R2 = 0.60) for serum-resistant bacteria. These data show how specific virulence factors explain why some bacterial species colonize without causing illness, whereas others colonize and cause sepsis. Although slime production was necessary for colonization, too much slime inhibited CVR. Although complement killing cleared bacteria from the circulation, it was also associated with increased TNF-α production, which can lead to septic manifestations. The most virulent bacterial species (from patients with sepsis) were killed by complement, but they still had significant CVR and were associated with increased TNF-α production.