Taxonomic, Genomic, and Functional Variation in the Gut Microbiomes of Wild Spotted Hyenas Across 2 Decades of Study

The gut microbiome provides vital functions for mammalian hosts, yet research on the variability and function of the microbiome across adult lifespans and multiple generations is limited in large mammalian carnivores. Here we use 16S rRNA gene and metagenomic sequencing to profile the taxonomic composition, genomic diversity, and metabolic function of the gut microbiome of 12 wild spotted hyenas (Crocuta crocuta) residing in the Masai Mara National Reserve, Kenya over a 23-year period spanning three generations. We determined the extent to which host factors predict variation in the gut microbiome and identify the core microbes present in the guts of hyenas. We also investigate novel genomic diversity in the mammalian gut by reporting the first metagenome-assembled genomes (MAGs) for hyenas. We found that gut microbiome taxonomic composition was highly variable across the two decades of sampling, but despite this, a core set of 14 bacterial genera and 19 amplicon sequence variants were identified. The strongest predictors of microbiome alpha and beta-diversity were host identity and age, suggesting that hyenas possess individualized microbiomes, and that these may change with age during adulthood. Gut microbiome functional profiles were also individual-specific, and were moderately correlated with antelope prey abundance, indicating that the functions of the gut microbiome vary with host diet. We recovered 149 high-quality MAGs from the hyena gut, spanning 25 bacterial orders and 51 genera. Some MAGs were classified as taxa previously reported for other carnivores, but many were novel and lacked species level matches to genomes in existing reference databases. Importance There is a gap in knowledge regarding the genomic diversity and variation of the gut microbiome across a host’s lifespan and across multiple generations of hosts in wild mammals. Using two types of sequencing approaches, we demonstrate that although gut microbiomes are individualized and temporally variable among hyenas, they correlate similarly to large-scale changes in their host’s ecological environment. We also recovered 149 high-quality MAGs from the hyena gut, greatly expanding the microbial genome repertoire known for hyenas, carnivores and wild mammals in general. Some MAGs came from genera abundant in the gastrointestinal tracts of canid species and other carnivores but over 80% of MAGs were novel and from species previously not represented in genome databases. Collectively, our novel body of work illustrates the importance of surveying the gut microbiome of non-model wild hosts, using multiple sequencing methods and computational approaches, and at distinct scales of analysis.