Decoupling Cattle Genetics and Gut Function: Metagenomics Evidence that Breed-Specific Heat Stress Tolerance Shape Functional Faecal Microbiome

The bovine gut microbiome’s role in heat stress tolerance is underexplored, particularly in tropical cattle production systems where heat tolerance is a breeding goal. This study characterized faecal resistome and CAZyome of three breeds with contrasting heat tolerance backgrounds including heat-tolerant Brangus, Kedah-Kelantan and heat-sensitive Holstein-Friesian under uniform tropical field conditions. Faecal samples were collected aseptically by rectal grab. Total microbial DNA was extracted from approximately 250 mg faeces per replicate using a commercial kit. Shotgun metagenomic libraries were prepared (Nextera XT) and sequenced on an Illumina NovaSeq 6000 (150-bp paired-end). After quality trimming and host read removal, assembled contigs were analyzed using the Resistance Gene Identifier (RGI) against CARD and the CAZy database. Non-parametric Kruskal–Wallis tests (p < 0.05) compared breed-level variables. Resistance mechanisms differed significantly: Holstein-Friesian showed greater reliance on antibiotic target alteration (p = 0.039) and lower reliance on efflux (p = 0.027) compared to Brangus and Kedah-Kelantan, CAZyme profiles revealed higher hemicellulose-degrading GH families in Brangus (p = 0.039), more peptidoglycan-remodelling Glycoside Hydrolase (GH) families was observed in Brangus and Kedah-Kelantan versus Holstein-Friesian (p = 0.039), and more pectin-degrading GH families was observed in the Holstein-Friesian (p = 0.027) compared to the other breeds. Integrative analysis indicated broad cross-breed conservation in total abundance, diversity, and dominant family composition, yet specific mechanistic and functional differences aligned with heat-tolerance genetic backgrounds. Thus, while the faecal resistome and CAZyome share a conserved functional core, but the breeds genetic heat-tolerance backgrounds drive distinct functional features under identical tropical management.