Results
Lyophilization yields were 6.29 and 6.38%, and culturable carbohydrate-utilizing bacterial counts were 3.79 × 10⁷ and 3.93 × 10⁷ CFU/g, respectively, with no significant differences between AHP and CSP. AHP contained higher propionate and valerate concentrations (p < 0.05), whereas CSP contained higher acetate and isovalerate concentrations and had a higher acetate-to-propionate ratio (A/P; p < 0.05). Bacterial, fungal, and methanogen community structures differed significantly between groups (p < 0.01), and CSP showed higher bacterial and fungal Shannon indices (p < 0.01). AHP was significantly enriched in the bacterial genera Xylanibacter, Segatella, Prevotellaceae_UCG-001, and Succinivibrio; the fungal genera Neocallimastix, Piromyces, Feramyces, and Aspergillus; and the methanogen genera Methanoculleus and Methanosphaera. CSP was significantly enriched in the bacterial genera norank_f__F082, Rikenellaceae_RC9_gut_group, norank_f__Bacteroidales_BS11_gut_group, norank_f__[Eubacterium]_coprostanoligenes_group, Ruminococcus, Monoglobus, norank_f__UCG-010, and Candidatus Saccharimonas; the fungal genera Orpinomyces, unclassified_f__Neocallimastigaceae, Cyllamyces, Caecomyces, Pecoramyces, Anaeromyces, Joblinomyces, and Alternaria; and the methanogen genera Methanobrevibacter and Methanolacinia. Cross-domain correlation analysis showed predominantly positive associations among genera enriched within the same roughage-source group and negative associations between AHP- and CSP-enriched genera. Compared with their substrate-free controls, AH and CS showed increased gas production, bacterial crude protein (BCP), total volatile fatty acids (TVFA), acetate, propionate, and butyrate, and decreased pH (p < 0.05). CS showed higher pH, acetate, isovalerate, TVFA, and A/P, whereas AH showed higher gas production, butyrate, and valerate (p < 0.01).
