Development and characterization of a fecal-induced peritonitis model of murine sepsis: results from a multi-laboratory study and iterative modification of experimental conditions

Abstract Background: Preclinical sepsis models have been criticized for their inability to recapitulate human sepsis and suffer from methodological shortcomings that limit external validity and reproducibility. The National Preclinical Sepsis Platform (NPSP) is a consortium of basic science researchers, veterinarians, and stakeholders in Canada undertaking standardized multi-laboratory sepsis research to increase the efficacy and efficiency of bench-to-bedside translation. In this study, we aimed to develop and characterize a 72-hour fecal-induced peritonitis (FIP) model of murine sepsis conducted in two independent laboratories. The experimental protocol was optimized by sequentially modifying dose of fecal slurry and timing of antibiotics in an iterative fashion, and then repeating the experimental series at site 1 and site 2. Results: Escalating doses of fecal slurry (0.5 to 2.5 mg/g) resulted in increased disease severity, as assessed by the modified Murine Sepsis Score (MSS), with either minimal ( 60%) mortality outcomes. However, the MSS was a poor predictor of progression to death during the experiments, and mice were found dead without elevated MSS scores. Administration of antibiotics within 4 hours of inoculation rescued the animals from sepsis, as evidenced by 100% survival and reduced bacterial load in peritoneum and blood, compared with late administration of antibiotics after 12 hours. Site 1 and site 2 had statistically significant differences in mortality (60% vs 88%; p < 0.05) for the same dose of fecal slurry (0.75 mg/g) and marked difference in temperature between groups. Microbiome analysis between two batches of fecal slurry revealed differences in beta-diversity and differential abundance analysis, which correlated with differences in mortality in two separate experimental series. Conclusions: We demonstrate a systematic approach to optimizing a 72-hour FIP model of murine sepsis for use in multi-laboratory studies. Alterations to experimental conditions, such as dose of fecal slurry and timing of antibiotics, have clear impact on outcomes. Differences in mortality between sites despite rigorous standardization warrants further investigations to better understand inter-laboratory variation and methodological design in preclinical studies. The use of surrogate mortality endpoints in preclinical sepsis models may introduce bias in evaluation and analysis of outcomes.