I. INTRODUCTION
The ban on the use of AGPs together with good antibiotic stewardship practices by intensive livestock industries resulting in a decline of antimicrobial resistance globally (Salim et al., 2018). Probiotics have been widely studied partially as a replacement of AGPs. Some probiotics are able to improve production performance whilst improving gut health and decreasing the likelihood of mortality due to infection. This study investigated the effects of various novel Bacillus probiotic formulations selected through various in vitro metrics including enzyme production, on broiler chicken growth performance, microbial profile, gut morphology, and nutrient digestibility to determine the most viable AGP alternative.
Table 1 - Bacillus species formulation combinations (6x108 CFU/g).
II. METHOD
The animal experiments developed in this project were approved by The University of Queensland (UQ) Production and Companion Animal Ethics Committee with the certificate number 2020/AE000235. A total of 576-day-old ROSS308 broiler chicks (mixed sex) were transferred to the Queensland Animal Science Precinct (QASP) facility at UQ, Gatton for a 21- day preliminary trial. All chicks were weighed, and randomly distributed to 72 floor pens, 8 chicks per pen. Pens were randomly allocated to six corn-soybean-based treatments; T1: Negative control (NC), T2: Positive control (NC + Tylosin), T3: T1 + formulation 1, 0.15%, T4: T1 + formulation 2, 0.15%, T5: T1 + formulation 3, 0.15%, T6: T1 + formulation 4, 0.15% (12 pens per treatment, n=96) (Table 1). Feed intake and body weight were measured weekly, and mortality was recorded daily. Ileum, duodenum, jejunum, and ileal contents were collected for gut morphology, microbial profiling, and nutrient digestibility, respectively. In the assay for protease, clearance zones were observed on the skim milk agar; whereas the assay for αamylase was conducted on starch agar.
III. RESULTS
The performance results have been summarised in Table 2. Formulation 1 (T3) compared to formulation 3 (T5) showed significantly (P < 0.05) higher body weight at day 21 and higher average daily gain d1-21. The European feed efficiency index demonstrated that T3 had a significantly higher performance than T5 (P < 0.05). In addition, F1 strain (T3) tended (P< 0.10) to show the lowest mortality among other treatments. The gut morphology, however, did not reveal significant differences (P > 0.05). Microbial profiling revealed that at the genus level, Lactobacillus, Alistipes, Akkermansia, Turicibacter and Bacteroides have low abundance in T1 compared to the remaining treatment groups (P < 0.05), suggesting the administration of Tylosin had a significant impact on the growth of these genera. However, the addition of probiotic strains Bacillus seemed to restore the genera abundance (T3, T4, T5, T6.) T3 revealed the highest N digestibility, which was significantly higher than the negative control, T1. T2, T5, and T6 also had significantly higher N than T1 (P < 0.05). The same result was also reflected in the crude protein digestibility (P < 0.05).
Table 2 - Effect of different Bacillus strains on performance parameters of broiler chickens from day 1 to 21 (n=96).
IV. DISCUSSION
This trial revealed that the inclusion of Bacillus based probiotics into broiler diets can improve growth performance, gut health, and nutrient digestibility compared to a positive control and AGP diet. Formulation 1, a combination of 3 Bacillus amyloliquefaciens strains, was the most effective in broiler diets, which was also demonstrated by Ahmat et al. (2021) who found that broiler production performance and immune response was improved in broilers fed this species of Bacillus. As Bacillus amyloliquefaciens produce protease and amylase as by-products, the digestibility of nutrients such as crude protein are improved, as well as increasing resistance to gut infection (Stefanello et al., 2017). The administration of probiotic strains Bacillus can be used to recover the abundance of helpful genera, which play an important role in fibre fermentation and promote health such as Lactobacillus (Alloui et al., 2013). The lack of differences between treatments in gut morphology contradicts the findings by Rodjan et al (2018). The discrepancy might be due to short-term termination at 21 days of our study. This study aimed at screening several probiotic formulations hence the shortened reading period. Thus, this study has identified formulation 1 to be a candidate for further testing to improve broiler production performance, gut health, and nutrient digestibility in the absence of in-feed antimicrobials.
Presented at the 34th Annual Australian Poultry Science Symposium 2023. For information on the next edition, click here. 
