There is a necessity for the implementation of in-feed probiotics in the poultry production industry, following strict regulations around the use of antibiotic growth promoters (AGP). Bacillus spp. are becoming an attractive alternative because of their functionality and stability. This study aims to evaluate the effect of a novel multi-strain Bacillus based probiotic on growth performance and gut health in male Ross 308 broiler chickens challenged with Clostridium perfringens Type A. Broilers on a 4 phase feeding program were fed diets containing either a standard metabolizable energy (ME) (100%) or a reduced ME (98%) level. The test probiotic was compared to an un-supplemented negative control and a commercial benchmark product as positive control over a 35 D feeding trial, using a 2 × 3 factorial experimental design. Chicks were inoculated with a once-off dose of C. perfringens on day 14. Growth performance was measured weekly to calculate body weight (BW), feed intake (FI) and feed conversion ratio (FCR). Villi histomorphology, gut lesions, and liver weight were assessed at day 35. Broilers fed the reduced ME diet with the test probiotic achieved higher final BWs (P = 0.037) and FCR (P = 0.014) than the negative control. Broilers fed the standard ME diet with the test probiotic showed improved (P = 0.001) FCR than the negative control from day 21 onwards. Increased duodenal villi height (P = 0.012) and villi height to crypt depth ratio in the duodenum (P < 0.0001) and jejunum (P = 0.0004) were observed in broilers fed the reduced ME diet containing the test probiotic. Additionally, the test probiotic resulted in significantly reduced relative liver weights in both ME groups. Consequently, the results suggest that the novel multi-strain Bacillus based probiotic enhanced broiler performance and improved gut health and is thus attractive as an alternative to AGP's in broiler production.