Effects of challenge with Clostridium perfringens, Eimeria and both on ileal microbiota of yellow feather broilers

Abstract

In the poultry industry worldwide, Clostridium perfringens has been causing major economic loss as it can cause necrotic enteritis (NE). The coccidial infection has been considered as the most important predisposing factor of NE caused by C. perfringens. In this study, we aimed to advance our knowledge on ileal microbiota of yellow feather broilers under C. perfringens and/or Eimeria challenge. Total of 80 healthy day old yellow feather broilers were randomly assigned to four groups including: Control, C. perfringens challenge group (C. Per), Eimeria challenge group (Cocc), and C. perfringens plus Eimeria challenge group (Comb). On day 14, the Cocc and Comb group broilers were orally gavaged 1 ml PBS solution containing 25,000 oocysts of Eimeria brunetti and 25,000 oocysts of Eimeria maxima. Starting on day 17, the C. Per and Comb group broilers were orally gavaged 10 mL of C. perfringens per bird (4 × 10⁷ CFU/mL, ATCC^® 13124™ Strain) every day for 6 days. 16S rRNA gene sequencing was performed on extracted DNA of ileal digesta samples. The results showed that C. perfringens alone did not affect the alpha diversity of ileal microbiome in yellow feather broilers but co-infection with Eimeria significantly decreased the diversity of ileal microbiota. C. perfringens and Eimeria challenge also decreased the relative abundance of beneficial bacteria including Bacteroidetes at the phylum level and Faecalibacterium at the genus level. At the species level, the relative abundance of Candidatus Arthromitus was significantly decreased in the Eimeria challenged groups. This microbial shift information of ileal microbiota under C. Perfringens and Eimeria challenge provide important reference data for the development of therapeutic approaches to necrotic enteritis in yellow-feather broiler chickens.

Keywords: C. perfringens; Eimeria; challenge; microbiota; yellow feather broiler.

FIGURE 1

Number of Amplicon Sequence Variants (ASVs) in each group. (A) Venn diagram of shared and specific ASVs in the four groups. (B) Rarefaction curve of observed otus in all samples. C. Per, C. perfringens challenge; Cocc, Eimeria challenge; Comb, C. perfringens and Eimeria challenge.

FIGURE 2

Principle component analysis of the ileal microbiota in different groups. C. Per, C. perfringens challenge; Cocc, Eimeria challenge; Comb, C. perfringens and Eimeria challenge.

FIGURE 3

Phylum-level, genus-level, and species level taxonomic composition of the ileal bacterial communities in four groups. C. Per, C. perfringens challenge; Cocc, Eimeria challenge; Comb, C. perfringens and Eimeria challenge.

FIGURE 4

linear discriminant analysis (LDA) effect size (LEfSe) Analysis of differential species among all groups shown in the LDA value distribution histogram. The presented species are biomarkers with statistical differences between groups (LDA > 3.5). The length of the histogram (LDA score) represents the impact of the different species.

FIGURE 5

Function predictions using phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt2). (A) Venn diagram of shared and specific functions predicted in the four groups. (B) Barplot of the top functions predicted against KEGG orthology (KO) database. (C) Principle component analysis (PCA) of the functions predicted in different groups; C. Per, C. perfringens challenge; Cocc, Eimeria challenge; Comb, C. perfringens and Eimeria challenge.