Endotoxic activity in faeces of mice from different microbiological environments

Area of Science:

• Microbiology
• Immunology
• Biochemistry

Background:

• Endotoxins, potent immune activators, are crucial in various biological processes.
• The Limulus amoebocyte lysate (LAL) test is a standard method for detecting endotoxins.
• Microbiota composition can influence host physiology and biochemical markers.

Purpose of the Study:

• To evaluate the performance of chromogenic LAL tests for measuring endotoxic activity in mouse feces.
• To assess the impact of different microbiological environments (germ-free, SPF, conventional) on LAL test results.
• To determine if fecal microbiota influences endotoxin levels and LAL test specificity.

Main Methods:

• Chromogenic Limulus amoebocyte lysate (LAL) assays were performed on mouse fecal samples.
• Fecal samples were collected from BALB/c mice from germ-free (GF), specific-pathogen-free (SPF), and conventional (CV) colonies.
• Heat inactivation and other treatments were used to investigate interfering substances.

Main Results:

• Feces from GF and SPF mice exhibited heat-labile, non-specific amidase activity, causing false-positive LAL reactions.
• CV mouse feces did not show this interfering activity.
• GF feces also demonstrated a heat-stable, non-endotoxic effect on the LAL system.
• Average fecal endotoxin levels were 0.2 µg/g (GF), 0.8 µg/g (SPF), and 17-24 µg/g (CV).

Conclusions:

• The microbiological status of mice critically affects fecal endotoxin quantification using chromogenic LAL tests.
• Non-specific enzymatic activities in the gut microbiota can lead to inaccurate endotoxin measurements.
• The LAL test's specificity is compromised by the host's microbiological environment, necessitating careful interpretation of results.