Kinetics of endotoxin release by gram-negative bacteria in the intestinal tract of mice during oral administration of bacitracin and during in vitro growth.

The release of endotoxin by gram-negative bacteria was studied during bacitracin-induced intestinal colonization in C3H/Law mice and during in vitro growth. The "free" endotoxin concentration was determined by the Limulus amebocyte lysate test in faecal and in culture supernatants, respectively. After oral administration of bacitracin for 2 days a significant (p less than 0.001) increase of the faecal concentration of gram-negative bacteria of 3-4 logs accompanied a significant (p less than 0.001) increase of the faecal endotoxin concentration from 10(1.8 +/- 0.2) to 10(3.3 +/- 0.2) micrograms endotoxin/g faeces. In vitro, however, an increase of the concentration of gram-negative bacteria of 3-4 logs resulted in a 3-4 log increase of the concentration of endotoxin during the exponential and early stationary growth phase. The faecal endotoxin level after 8 days of bacitracin treatment dropped to a value not significantly different from the initial value, regardless of a high faecal level of gram-negative bacteria. Endotoxin determination by the Limulus amebocyte lysate test appeared to be unaffected by the amount of bacitracin present in faecal preparations after oral administration of this antibiotic. In addition, the in vitro release of endotoxin was not influenced by bacitracin. From these results we conclude, that "free" endotoxin is a product of extensive proliferation of gram-negative bacteria. Therefore, the intestinal endotoxin level does not necessarily correlate with the level of gram-negative bacteria, but corresponds with the proliferative activity of these bacteria.