Lonicerae flos and turmeric extracts alleviate necrotic enteritis in broilers by modulating gut-liver health and microbiota

Abstract

Background: Necrotic enteritis (NE) can cause intestinal barrier dysfunction in broilers, leading to secondary liver injury (SLI). In this process, the gut-liver axis plays a crucial role. Lonicerae flos and turmeric extracts (LTE), containing chlorogenic acid and curcumin, have been reported to possess anti-inflammatory, and antioxidant properties. Based on these potential biological benefits, this study aims to investigate the reparative effects of LTE on the intestinal barrier dysfunction in NE-infected broilers and assess its therapeutic efficacy in alleviating SLI. By elucidating the regulatory mechanisms of LTE on gut-liver axis health, this research provides new insights into the prevention and treatment of NE in broilers.

Results: LTE improved body weight and average daily gain while reducing intestinal lesion scores, coccidia oocysts, and Clostridium perfringens counts in NE broilers (P < 0.05). LTE enhanced intestinal morphology and up-regulated the expression of tight junction protein genes (CLDN1, TJP1) and MUC2, suppressed pro-inflammatory cytokine and myeloperoxidase (MPO) levels, and minimized endotoxin (ET) accumulation in NE broilers (P < 0.05). Furthermore, LTE alleviated oxidative stress in ileal cells and protected mitochondrial structure and function in NE broilers. NE infection induced intestinal permeability in broilers, leading to increased serum pro-inflammatory cytokines and intestinal-derived endotoxin levels, which caused liver damage. LTE significantly reduced liver pathologic damage, pro-inflammatory cytokine levels, aspartate transaminase, alanine aminotransferase, and ROS levels in NE broilers (P < 0.05). Additionally, 16S rRNA sequencing revealed that NE significantly increased the relative abundance of Barnesiella and decreased the relative abundance of Bacteroidota, Desulfobacterota and Bacteroides in the cecum of broilers. LTE enhanced intestinal microbiota diversity and reduced the segregation of intestinal microbiota induced by NE infection.

Conclusions: In summary, LTE can alleviate NE and SLI by modulating the microbiota, inhibiting inflammation and oxidative stress, and ameliorating mitochondrial dysfunction, thereby enhancing gut-liver axis health and growth performance.

Keywords: Chlorogenic acid; Curcumin, Microbiota; Mitochondrial dysfunction; Necrotic enteritis; Secondary liver injury.

Fig. 1

Representative image of coccidiosis oocysts in feces on d 18. Black arrows indicate coccidial oocysts. Scale bar: 100 μm. NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6)

Fig. 2

Effects of LTE on Clostridium perfringens in cecum of NE broilers on d 28 (Black dots indicate Clostridium perfringens). NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6)

Fig. 3

Effects of LTE on intestinal barrier function of NE broilers on d 28. A Representative images of ileum H&E and AB-PAS staining. B Number of ileal goblet cells. C MUC2 mRNA expression level. D mRNA expression level of ileal tight junction-related genes. E Endotoxin content of cecal. F MPO content of ileum. G Inflammatory factor level of ileum. NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6). ^a,bThe different lowercase letters on the bar charts indicate significant differences (P < 0.05)

Fig. 4

Effects of LTE on oxidative stress and mitochondrial function in the ileum of NE broilers on d 28. A CAT, T-SOD, and GSH-Px content of ileal tissue. B ROS content of ileal tissue. C Nrf2, HO-1, and NQO-1 mRNA expression levels. D Mitochondrial CAT, T-SOD, and GSH-Px content. E ROS content of mitochondrial. F ATP content of mitochondrial. G Transmission electron micrographs of mitochondria (Scale bar: 5 μm, 2 μm, 500 nm; Magnification: × 5.0 k, × 15.0 k, × 40.0 k). NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6). ^a–cThe different lowercase letters on the bar charts indicate significant differences (P < 0.05)

Fig. 5

Effects of LTE on secondary liver injury in NE broilers on d 28. A Serum endotoxin levels. B Serum inflammatory cytokine levels. C Serum AST and ALT levels. D Liver endotoxin levels. E Liver inflammatory cytokine levels. F Liver weight. G Liver ROS levels. H Nrf2, HO-1, NQO-1 mRNA expression levels. I Liver index. J Liver H&E staining. NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6). ^a,bThe different lowercase letters on the bar charts indicate significant differences (P < 0.05)

Fig. 6

Effects of LTE on microbial diversity in the cecum of NE broilers on d 28. A Rarefaction curve. B Rank abundance curve. C Venn diagram. D The α-diversity parameters including ACE, Chao, Shannon, and Simpson indices. NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6). The different lowercase letters on the bar charts indicate significant differences (P < 0.05)

Fig. 7

Effects of LTE on microbial community structure and composition in the cecum of NE broilers on d 28. A Principal co-ordinate analysis (PCoA) plot. B Non-metric multidimensional scaling (NMDS) plot. C Microbiota in the top 10 species of relative abundance at the phylum level. D Microbiota in the top 10 species of relative abundance at the genus leel. NC, unchallenged and without additive; PC, challenged and without additives; LTE300, challenged and dietary supplementation with 300 mg/kg LTE (n = 6)

Fig. 8

Heatmap of the correlation analysis between intestinal microbiota (at the genus level) parameters related to growth performance and intestinal barrier function on d 28. ^*P < 0.05, ^**P < 0.01