Quercetin Increases Growth Performance and Decreases Incidence of Diarrhea and Mechanism of Action in Weaned Piglets

Abstract

This study aimed to investigate the mechanism of quercetin increasing growth performance and decreasing incidence of diarrhea in weaned piglets. Forty-eight Duroc × Landrace × Large White weaned piglets with similar body weight (7.48 ± 0.20 kg, 28 days of age) were randomly divided into four treatments (control, 250 mg/kg quercetin, 500 mg/kg quercetin, and 750 mg/kg quercetin treatments) and fed with basal diet or experimental diet supplemented with quercetin. Performance, diarrhea rate and index, and content of serum anti-inflammatory factors were determined and calculated in weaned piglets; colonic flora and signaling pathways related to anti-inflammation were measured using 16S rDNA sequencing and RNA-seq, respectively. The results showed that compared with control, feed-to-gain ratio and content of serum interferon gamma (IFN-γ) were significantly decreased in the 500 and 750 mg/kg quercetin treatments (P < 0.05); quercetin significantly decreased diarrhea rate and diarrhea index (P < 0.05) and significantly increased the content of serum transforming growth factor (TGF-β) in weaned piglets (P < 0.05); the content of serum NF-κB was significantly decreased in the 750 mg/kg quercetin treatment (P < 0.05); moreover, quercetin significantly increased diversity of colonic flora (P < 0.05), and at the phylum level, the relative abundance of Actinobacteria in the 500 and 750 mg/kg treatments was significantly increased (P < 0.05), and the relative abundance of Proteobacteria in the three quercetin treatments were significantly decreased (P < 0.05) in the colon of weaned piglets; at the genus level, the relative abundance of Clostridium-sensu-stricto-1, Turicibacter, unclassified_f_Lachnospiraceae, Phascolarctobacterium, and Family_XIII _AD3011_group was significantly increased (P < 0.05); the relative abundance of Subdollgranulum and Blautia was significantly decreased in the 500 and 750 mg/kg treatments (P < 0.05); the relative abundance of Eschericha-Shigella, Terrisporobacter, and Eubacterium-coprostanoligenes was significantly increased (P < 0.05); the relative abundance of Streptocococcus, Sarcina, Staphylococcus, and Ruminococcaceae_UCG-008 was significantly decreased in the three quercetin treatments (P < 0.05); the relative abundance of Ruminococcaceae_UCG_014 was significantly increased in the 250 mg/kg quercetin treatment in the colon of weaned piglets (P < 0.05). The results of Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that differentially expressed genes (DEGs) from the quercetin treatments were significantly enriched in nuclear transcription factor-κB (NF-κB) signal pathway (P < 0.05); mRNA expression of tumor necrosis factor-α (TNF-α), interleukin-1R1 (IL-1R1), conserved helix-loop-helix ubiquitous kinase (CHUK), toll-like receptor 4 (TLR4), and IL-1β from quercetin treatments were significantly decreased in colonic mucosa of weaned piglets (P < 0.05). In summary, quercetin increased feed conversion ratio and decreased diarrhea through regulating NF-κB signaling pathway, controlling the balance between anti-inflammatory and proinflammatory factors, and modulating intestinal flora, thus promoting the absorption of nutrients in weaned piglets. These results provided the theoretical foundation for applying quercetin in preventing weaning piglets' diarrhea and animal husbandry practices.

Figure 1

Description of flora community in the colon of weaned piglets: (a) rarefaction curves based on the observed taxonomic units (OTUs) in the colon, (b) Venn diagrams of shared genes, (c) principal component analysis (PCA) of flora in the colon, (d, e) the relative abundance of phyla and genera in the colon, and (f) the significant influence of genus level.  ^∗,  ^∗∗, and  ^∗∗∗ represent P < 0.05, n = 6. Con represents control; Q250 represents 250 mg/kg quercetin treatment; Q500 represents 500 mg/kg quercetin treatment; and Q750 represents 750 mg/kg quercetin treatment.

Figure 2

Effects of quercetin on differentially expressed genes in the colon of weaned piglets. The scatter plot showed the correlation of gene abundance. Red points represented the upregulated genes. Green points represented the downregulated genes, and gray dots represented significantly unchanged transcripts: (a) 250 mg/kg treatment, (b) 500 mg/kg treatment, and (c) 750 mg/kg treatment.

Figure 3

GO analyses of differentially expressed genes in control and 250 mg/kg quercetin treatment, 500 mg/kg quercetin treatment, and 750 mg/kg quercetin treatment. Figure 0.025% shown the GO analysis of control and 250 mg/kg quercetin treatment differentially expressed genes. Figure 0.05% shown the GO analysis of control and 500 mg/kg quercetin treatment differentially expressed genes. Figure 0.075% shown GO analysis of control and 750 mg/kg quercetin treatment differentially expressed genes: (a) 250 mg/kg quercetin treatment, (b) 500 mg/kg quercetin treatment, and (c) 750 mg/kg quercetin treatment.

Figure 4

Correlation of mRNA expression of 10 random DEG sequences of anti-inflammatory factors in NF-κB pathway using RNA-seq and RT-qPCR. The three diagrams shown the results of comparing Q250, Q500, and Q750 with Con. Black represented the RNA-seq result, and gray represented the RT-qPCR result. Con represented control; Q250 represented 250 mg/kg quercetin treatment; Q500 represented 500 mg/kg quercetin treatment; and Q750 represented 750 mg/kg quercetin treatment.

Figure 5

The expression of 10 DEGs was detected using RT-qPCR. Mean values without a common letter are significantly different, P < 0.05. Values were mean ± SEM, n = 6. Con represents control; Q250 represents 250 mg/kg quercetin treatment; Q500 represents 500 mg/kg quercetin treatment; and Q750 represents 750 mg/kg quercetin treatment.