<i>APOE</i> genotype and sex drive microbiome divergence after microbiome standardization in <i>APOE</i> -humanized mice

The <i>APOE4</i> allele is the greatest known genetic factor for sporadic or late-onset Alzheimer's Disease (LOAD). Gut microbiome (GMB) dysbiosis can lead to poorer outcomes in disease. The intersection of sex, <i>APOE</i> genotype, inflammation, and gut microbiota is incompletely understood. Previous studies in humans and humanized <i>APOE</i> mice have demonstrated <i>APOE</i>-genotype-specific differences in the GMB. However, most of these studies were unable to resolve bacteria to the species level. It remains unclear how GMB changes with age and sex in the context of <i>APOE</i> genotype. In this study, humanized male mice with either <i>APOE</i> 2, 3, or 4 genotype were bred with the same two C57BL/6J sisters to standardize microbiomes across lines and monitor divergence based on <i>APOE</i> allele. Stool samples were collected at breeder set up and from the heterozygous (F1) and homozygous (F2) generations at wean and 6 months old. Stool was assessed via shallow shotgun sequencing to enable species and strain-level taxonomic resolution. The heterozygous pups' microbiome resembled each other at wean across all genotypes. However, the heterozygous pups and their homozygous offspring continued to diverge, particularly the <i>APOE2</i> females. In homozygous mice, the GMB demonstrated significant divergence at 6 months of age based on sex and <i>APOE</i> genotype. In comparison to their <i>APOE3</i> and <i>APOE4</i> counterparts, <i>APOE2</i> females and males demonstrated an increased quantity of bacteria associated with anti-inflammatory profiles, including in the <i>Lachnospiraceae</i> family (<i>Lachnospiraceae</i> bacterium UBA3401) and decreased quantities in the <i>Turicibacteraceae</i> family (higher levels are associated with LOAD).IMPORTANCEThe <i>APOE4</i> allele is implicated as a significant risk factor for many diseases, including cardiovascular disease (responsible for more deaths than any other disease) and sporadic or late-onset Alzheimer's Disease (accounts for an estimated 60%-80% of all dementia cases). It is known that the gut microbiome (GMB) is affected by different genotypes and disease states. Mouse model studies have environmental and genetic controls, allowing a specific gene to be studied. This study aims at discovering key GMB species differences allowing for future therapeutic targets. The GMB of the experimental mice was standardized, and genotype and sex-specific divergence was observed with species and even strain level taxonomic resolution. Reported here are the first data demonstrating GMB divergence over time driven by <i>APOE</i> genotype from an inherited source and the first data to identify APOE genotype-specific bacteria species that may serve as therapeutic targets in <i>APOE</i>-driven disease.