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Coccidiosis is a well-known poultry disease that causes the severe destruction of the intestinal tract, resulting in reduced growth performance and immunity, disrupted gut homeostasis and perturbed gut microbiota. Supplementation of probiotics were explored to play a key role in improving growth performance, enhancing innate and adaptive immunity, maintaining gut homeostasis and modulating gut microbiota during enteric infection. This study was therefore designed to investigate the chicken gut whole microbiota responses to Bacillus subtilis (B. subtilis) probiotic feeding in the presence as well as absence of Eimeria infection. For that purpose, 84 newly hatched chicks were assigned into four groups, including (1) non-treated non-challenged control group (CG − ET), (2) non-treated challenged control group (CG + ET), (3) B. subtilis-fed non-challenged group (BS − ET) and (4) B. subtilis-fed challenged group (BS + ET). CG + ET and BS + ET groups were challenged with Eimeria tenella (E. tenella) on 21 day of housing. Our results for Alpha diversity revealed that chickens in both infected groups (CG + ET and BS + ET) had lowest indexes of Ace, Chao 1 and Shannon, while highest indexes of Simpson were found in comparison to non-challenged groups (CG − ET and BS − ET). Firmicutes was the most affected phylum in all experimental groups following Proteobacteria and Bacteroidota, which showed increased abundance in both non-challenged groups, whereas Proteobacteria and Bacteroidota affected both challenged groups. The linear discriminant analysis effect size method (lEfSe) analysis revealed that compared to the CG + ET group, supplementation of probiotic in the presence of Eimeria infection increased the abundance of some commensal genera, included Clostridium sensu stricto 1, Corynebacterium, Enterococcus, Romboutsia, Subdoligranulum, Bacillus, Turicibacter and Weissella, with roles in butyrate production, anti-inflammation, metabolic reactions and the modulation of protective pathways against pathogens. Collectively, these findings evidenced that supplementation of B. subtilis probiotic was positively influenced with commensal genera, thereby alleviating the Eimeria-induced intestinal disruption.

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In this study, we performed transcriptome analysis in the cecum tissues of negative control untreated non-challenged (NC), positive control untreated challenged (PC), and Bacillus subtilis (B. subtilis) fed challenged chickens (BS + ET) in order to examine the underlying potential therapeutic mechanisms of Bacillus based probiotic feeding under an experimental Eimeria tenella (E. tenella) infection. Our results for clinical parameters showed that birds in probiotic diet decreased the bloody diarrhea scores, oocyst shedding, and lesion scores compared to positive control birds. RNA-sequencing (RNA-seq) analysis revealed that in total, 2509 up-regulated and 2465 down-regulated differentially expressed genes (DEGs) were detected in the PC group versus NC group comparison. In the comparison of BS + ET group versus PC group, a total of 784 up-regulated and 493 down-regulated DEGs were found. Among them, several DEGs encoding proteins involved in immunity, gut barrier integrity, homeostasis, and metabolism were up-regulated by the treatment of probiotic. Functional analysis of DEGs also revealed that some gene ontology (GO) terms related with immunity, metabolism and cellular development were significantly affected by the exposure of probiotic. Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis showed that the DEGs in the cecum of B. subtilis-fed challenged group were mainly participated in the pathways related with immunity and gut barrier integrity, included mitogen-activated protein kinase (MAPK) signaling pathway, toll-like receptor (TLR) signaling pathway, extracellular matrix (ECM)–receptor interaction, tight junction, and so on. Taken together, these results suggest that Bacillus based probiotic modulate the immunity, maintain gut homeostasis as well as barrier system and improve chicken metabolism during E. tenella infection.