Circadian regulation of the gut microbiome in ruminants: opportunities for targeted microbiota manipulation and optimized timing for interventions

Precise manipulation of the gut microbiota is complex. This study explored its circadian regulation in ruminants via time-restricted feeding. We collected 287 fecal samples (every 6 h for 48 h) from 12 lactating cows under ad libitum, daytime-restricted, and nighttime-restricted feeding. Microbiota analysis revealed rhythmic fluctuations in hindgut bacterial populations. Under feeding restrictions, 39.25% of bacterial genera (e.g., Bifidobacterium , Prevotella ) shifted rhythms in response to feeding time, while 3.90% (e.g., Succinivibrio ) did not. Fecal short-chain fatty acids (SCFA) and ammonia nitrogen (NH 3 -N) concentrations also exhibited circadian rhythms, peaking 6 h apart. Metatranscriptomics showed daily dynamics in active bacteria, archaea, eukaryotes, and bacteriophages: active bacteria and bacteriophages peaked at 08:00 and 02:00, whereas active archaea peaked at 14:00. A more active microbial community coincided with high SCFA concentration (08:00), reflected by elevated expression of genes for carbohydrate-active enzymes, antibiotic resistance, virulence, and quorum sensing. Conversely, gene expression for nitrogen metabolism, sulfur metabolism, and bacterial secretion peaked with NH 3 -N (14:00). Our findings reveal circadian rhythm-responsive features and daily shifts in the active microbiome, offering a mechanistic basis for hindgut microbiome dynamics and fermentation in ruminants, with implications for microbiota manipulation in other mammals.