Transcriptomic profiles of the ruminal wall in Italian Mediterranean dairy buffaloes fed green forage

Background Green feed diet in ruminants exerts a beneficial effect on rumen metabolism and enhances the contentof milk nutraceutical quality. At present, a comprehensive analysis focused on the identification of genes, and therefore,biological processes modulated by the green feed in buffalo rumen has never been reported. We performedRNA-sequencing in the rumen of buffaloes fed a total mixed ration (TMR) + the inclusion of 30% of ryegrass greenfeed (treated) or TMR (control), and identified differentially expressed genes (DEGs) using EdgeR and NOISeq tools.Results We found 155 DEGs using EdgeR (p-values < 0.05) and 61 DEGs using NOISeq (prob >=0.8), 30 of which areshared. The rt-qPCR validation suggested a higher reliability of EdgeR results as compared with NOISeq data, in ourbiological context.Gene Ontology analysis of DEGs identified using EdgeR revealed that green feed modulates biological processesrelevant for the rumen physiology and, then, health and well-being of buffaloes, such as lipid metabolism, response tothe oxidative stress, immune response, and muscle structure and function. Accordingly, we found: (i) up-regulation ofHSD17B13, LOC102410803 (or PSAT1) and HYKK, and down-regulation of CDO1, SELENBP1 and PEMT, encoding factorsinvolved in energy, lipid and amino acid metabolism; (ii) enhanced expression of SIM2 and TRIM14, whose productsare implicated in the immune response and defense against infections, and reduced expression of LOC112585166(or SAAL1), ROR2, SMOC2, and S100A11, encoding pro-inflammatory factors; (iii) up-regulation of NUDT18, DNAJA4and HSF4, whose products counteract stressful conditions, and down-regulation of LOC102396388 (or UGT1A9) andLOC102413340 (or MRP4/ABCC4), encoding detoxifying factors; (iv) increased expression of KCNK10, CACNG4, andATP2B4, encoding proteins modulating Ca2+homeostasis, and reduced expression of the cytoskeleton-related MYH11and DES.Conclusion Although statistically unpowered, this study suggests that green feed modulates the expression ofgenes involved in biological processes relevant for rumen functionality and physiology, and thus, for welfare and qualityproduction in Italian Mediterranean dairy buffaloes.These findings, that need to be further confirmed through the validation of additional DEGs, allow to speculate a roleof green feed in the production of nutraceutical molecules, whose levels might be enhanced also in milk.