Supplemental thymol and carvacrol increases ileum Lactobacillus population and reduces effect of necrotic enteritis caused by Clostridium perfringes in chickens

Abstract

Necrotic enteritis (NE) caused by Clostridium perfringens is one of the most detrimental infectious diseases in poultry. This study examined the effect of blends of essential oils (BEOs) (25% thymol and 25% carvacrol) on NE and bacterial dynamics and functions in chicks challenged with C. perfringens. Chicks were assigned to a Control diet and BEOs diet (Control diet + 120 mg/kg BEOs), were challenged with C. perfringens from days 14 to 20 and were killed on day 21 for assessment. Supplementation with BEOs decreased the mortality, alleviated gut lesions, and decreased the virulence factors of pathogenic bacteria (VF 0073-ClpE, VF0124-LPS, and VF0350-BSH). Lack of supplementation also changed the nutrient and immunological dynamics of host microbiota in responding to C. perfringens infection. Adding BEOs changed the host ileum microbial population by increasing the numbers of Lactobacillus crispatus and Lactobacillus agilis, and decreasing Lactobacillus salivarius and Lactobacillus johnsonii. The functional roles of these changing host bacterial populations coupled with the putative reduced pathogenicity of C. perfringens by BEOs contributed to the reduction in gut lesions and mortality in infected chickens. It suggests that dietary supplementation with BEOs could significantly reduce the impact of NE caused by C. perfringens on broilers.

Figure 1

Effects of challenge and supplemented BEOs on the mortality of broiler chickens during d 14–21 (Ctrl: C. perfringens challenge; BEOs: C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 2

Effects of challenge and supplemented BEOs on the intestinal lesion scores of broiler chickens (Ctrl: C. perfringens challenge; BEOs: C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 3

Compositions of the ileal microbiota of the broilers. Relative taxa abundance (%) of ileal bacteria of broilers at phylum level (A) and order taxonomic level (B) (Ctrl: C. perfringens challenge; BEOs: C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 4

Relative abundance of different annotated species in ileal microbiota of broilers (Top 10)(Ctrl: C. perfringens challenge; BEOs: C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 5

Comparison of the compositions of the ileal microbiota of the broilers. A Principal coordinate analysis (PCoA) based on the weighted unifrac diatance of 16S rRNA of ileal bacteria of broilers (A). Principal component analysis (PCA) of microbiota community by Bray-Curtis distance (B). The circles were drawn around microbiota from the same treatment.

Figure 6

Phylum and genus differentially represented between BEOs and Ctrl samples identified by linear discriminant analysis coupled with effect size (LEfSe) (LDA >2, P < 0.05). (Ctrl: (green): C. perfringens challenge; BEOs (red): C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 7

The different functions of the ileal microbiota of the broilers. Statistics of the number of annotated genes at KEGG meatabolic pathway level one (A) and eggNOG level one (B). (Ctrl: (blue): C. perfringens challenge; BEOs (red): C. perfringens challenge and supplemental BEOs 120 mg/kg).

Figure 8

Comparison of the gene pathways of the ileum microbiota of broilers annotated genes at KEGG pathways at level two (Ctrl: (blue): C. perfringens challenge; BEOs (red): C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 9

Comparison of immune system pathways in ileum microbiota annotated by KEGG pathways at level three (Ctrl: (blue): C. perfringens challenge; BEOs: (red): C. perfringens challenge and supplemental blends of essential oils 120 mg/kg).

Figure 10

Comparison of relative abundance of annotated genes at VFDB in ileum microbiota of broilers (Ctrl: (blue): C. perfringens challenge; BEOs (red): C. perfringens challenge and supplemental blends of essential oils 120 mg/kg) * Means significantly diffference (P < 0.05).