Humanizing Zophobas morio larvae microbiota for rapid screening of emerging decolonization strategies against multidrug-resistant bacteria

Abstract Purpose Designing efficient strategies against the gut colonization due to multidrug-resistant (MDR) bacteria is an important task. However, scalable and reliable in vivo models that possess a human-like gut microbiota are not yet available. Here, we tested whether Zophobas morio larvae (ZmL) could be used as a humanized microbiota model. Methods A pooled fecal transplant material from 7 human donors was administered via contaminated food to a group of ZmL every 48-h for 28 days (T28), followed by a 28-day washout phase. A control group received the standard diet. Gut microbiota composition was assessed at 9 timepoints (from T0 to T56) by 16S rRNA gene amplicon sequencing across 3 independent runs. Results Fecal microbiota transplants (FMTs) increased the richness of the larval gut bacterial population, with 8 of 10 top human-associated genera increasing during this phase. The experimental group exhibited higher observed richness than the control across T7-T56 (median 116 vs. 62 amplicon sequence variants; p < 0.001). A humanization score, calculated as the sum of the 10 top human genera detected in the transplanted ZmL, peaked at T14, and declined to near zero by the end of the washout phase (T56). Community composition differed significantly between groups, with the variable diet accounting for more variation than the run (PERMANOVA, R2 = 13.3%; p = 0.001). Conclusions Under repeated FMTs, ZmL underwent a humanization of their microbiota making this model a promising tool to study novel decolonization strategies against MDR bacteria. Future efforts should focus on the stabilization of the human-like microbiota without repeated FMTs.