Novel isolates of Bacillus subtilis improved growth performance and gut health in broilers challenged with Clostridium perfringens
Date
2022
Authors
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Publisher
University of Delaware
Abstract
Probiotics are reported to improve health and performance of broiler chickens by improving intestinal microbiota. They are being used as potential alternatives to in-feed antibiotics and are reported to be protective against some bacterial diseases in poultry including Necrotic enteritis (NE). Better understanding of role of probiotics is critical for development of probiotics-based management of bacterial diseases in poultry, including NE. This study performed a meta-analysis to estimate the effects of Bacillus subtilis probiotic supplementation on Feed conversion ratio (FCR), NE lesion score, and mortality. Further, a systematic review is done for gut microbiota changes on broiler challenged with Clostridium perfringens with or without the probiotic supplementation. Effect sizes were estimated in terms of standardized mean difference (SMD). Random effect models were fit to estimate the pooled effect size and 95% confidence interval of the pooled effect size between the control [probiotic-free C. perfringens challenged group] and the treatment group [Bacillus subtilis supplemented- C. perfringens challenged group]. Overall variance is computed by heterogeneity (Q). ☐ Feeding trial was done to determine effects of our novel Bacillus subtitlis isolates 103a and 62a on growth performance, intestinal lesions and gut microbiota in broiler challenged with Clostridium perfringens. A 2-factor [Infection (10*IBDV vaccine + 10^8 cfu/bird CP vs no IBDV+ no CP) × probiotics (control (no probiotics),103a, 62a, and mixed(103a+62a))] completely randomized feeding trial design was conducted for 28 days starting with day 1 male broiler chicks. Probiotic was given via water at 10^7 cfu/ml dose for 28 days. There were 15 birds in control and 5 in other treatment groups. Body weight and FCR were recorded at days 1,10,15,20, 22,24,26, and 28, and mortality was observed each day. Organ weight, plasma sample and intestine NE lesion score were recorded at day 28. FCR, organ weight and plasma oxygen radical absorbance capacity (ORAC) were analyzed using two-way ANOVA. Cecum microbiota samples were prepared using Shoreline Biome microbiome assay Kit and demultiplexed files were generated by Sbanalyzer. These files were further analyzed in R using DADA2, taxonomy was assigned using Athena database, and alpha and beta diversity indices were explored. ☐ Meta analysis done to observed effect of different strains of Bacillus subtilis supplementation in birds with Clostridium perfringens inoculation showed that Bacillus subtilis probiotic supplementation significantly improved FCR, reduced NE lesion but had no effect on mortality rates. The estimated overall effects of probiotic supplementation on FCR, NE lesion score and mortality percentage in term of SMD were -0.91 (CI=-1.34, -0.49; p<0.001) , -0.67 (CI = -1.11, -0.22; p=0.006) , and -0.32 (CI =-0.70, 0.06; p = 0.08) ,respectively. There was a significant heterogeneity across the studies in the effect of Bacillus subtilis supplementation on FCR (Q=69.66; p<0.001) and NE lesion score (Q=42.35; p<0.001). The heterogeneity was not significant for mortality (Q=2.72; p=0.74). Probiotic supplementation with Bacillus subtilis was reported to have enriched streptococcus thermophilus, Butyricicoccus and Faecalibacterium genera, ruminococcus in gut. These microbiotas were reported to be related to upregulate several gene expression such as TJ proteins occluding, ZO-1 expression, junctional adhesion 2 (JAM2) expression, interferon gamma gene, IL12- β and transform growth factor-β4, and downregulate mucin-2 expression which were involved in restoring intestinal physical barrier, reduce intestinal inflammation, and recover the physiological functions of damaged intestines. ☐ Feeding trial showed increase in body weight in broiler challenged with Clostridium perfringens. There was fluctuation in FCR in all treatment group between day 10 to 28. Infected 62a have significantly (p<0.05) lower FCR compared to non-infected 62a treatment group. 62a probiotic have shown less mortality in both infected and non-infected probiotic supplemented groups. Both of our probiotic isolates were able to inhibit clostridium perfringens in antimicrobial assay. In ileum and cecum sample the relative abundance of Lactobacillus, Butyricicoccus and Erysipelatoclostridium was higher compared to other taxonomic group. Significant difference in genus Lactobacillus was seen in cecum and ileum content suggesting they might be correlated to increased body weight and less FCR which need to be further studied. In-ovo test resulted in higher mortality in probiotic 62a and mixture and less in 103a. Given high mortality 62a and mixed probiotic supplementation is not recommended however, 103a probiotic needs to be further investigated as it has low mortality and higher zone of inhibition. ☐ Overall, dietary supplementation of Bacillus subtilis improved FCR, body weight, NE lesion score, and changed gut microbiota in broilers challenged with C. perfringens.
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Keywords
Bacillus subtilis, Growth performance, Gut health, Broiler chickens, Clostridium perfringens, Probiotics, Necrotic enteritis, Low mortality