Gut health and microbiology expert, CERN, Adisseo France S.A.S.
Global Scientific Support - Health by Nutrition, Adisseo France S.A.S.
Gut health is usually considered as the symbiotic relationship between the host and its microbiota. Recent research has proposed a four-pillar scheme to describe healthy, pre-disease and disease states from dynamic interactions among the microbiota, mucosal barrier, immune system and redox balance (van de Gutchte et al., 2018) (Figure 1).
In case of an imbalance in these interactions, inflammation will occur, leading to 'leaky gut' – increased permeability of the intestinal barrier, and other gut-related disorders. Nowadays, probiotics are gaining importance as nutritional strategies to improve gut health through their influence on microbiota, gut morphology, inflammatory responses and overall animal performance.
Probiotics are live microorganisms, when administered in adequate amount, can confer beneficial physiological effects to the host (WHO, 2001). Probiotics can improve intestinal microbiota balance, leading to an optimum environment for digestion and nutrients absorption. These more balanced microbiota will also help to reduce the growth and/or the pathogenicity of harmful bacteria and positively influence the host immune system, contributing to overall better animal health and performance. Compared with antibiotics which have a specific effect on pathogens and maybe some anti-inflammatory properties, probiotics instead can deliver global benefits to the host through multiple and complementary modes of action.
In this article, recent trials with a poultry specific probiotic solution, unique spore forming Bacillus subtilis (strain DSM 29784, Alterion®) are presented. This unique specific strain was discovered and selected after extensive in vitro research at Novozymes's Laboratories, followed by further in vivo research in independent scientific institutions, universities and Adisseo' research farm (CERN, Centre for Expertise and Research on Nutrition). With all efforts above, Alterion is proved to deliver consistent benefits for poultry production from its research-supported modes of action, including positive changes in the microbiota, intestinal integrity and inflammatory responses. As such, Alterion enhances dialogues between the microbiota and the host, creating sustainable effects.
The balanced and varied microbiota is key to develop and maintain robust gut functions. The effect of Bacillus subtilis on bacterial species in the gut of broilers was investigated at Southern Poultry Research, Inc (Athens, GA, USA) in collaboration with Dr Greg F. Mathis. In a 42-day trial, 1600 Cobb 500 male broiler chicks were fed a control or treatment diet with Alterion supplemented at 1x108 CFU/kg of feed. At day 42, intestinal contents were sampled to determine ileal and cecal microbiota composition by rRNA gene sequencing.
At the cecal level (Figure 2), five genera (out of 116) were observed at a significantly higher relative abundance in Alterion treatment group, mainly from the Firmicutes phylum including Ruminococcus. These bacteria are known to breakdown polysaccharides to oligosaccharides, assisting the digestion of cereals and vegetable proteins in poultry diets.
In addition, increased relative abundance of Lachnoclostridium was also observed. These bacteria degrade oligosaccharides and produce short chain fatty acids (SCFAs) – butyrate in particular. It is well known that as a very important microbial metabolite, butyrate represents the main source of energy for enterocytes, cells from gut wall, and a very potent anti-inflammatory compound at the same time.
The gut mucosa, or alternatively named gut barrier or frontier, is where many dynamic interactions occur between the host and the "outside environment". The integrity and robustness of gut mucosa is essential for efficient nutrients absorption, innate immune system stimulation and prevention of bacteria and toxins from entering the bloodstream, therefore accounting for the optimal bird health and performance.
In the same study reported above, histological samples were also collected from broilers and gut morphology were assessed. Those birds in Alterion group had significantly longer intestinal microvilli (+18% increased length in the ileum and +17% in the cecum), suggesting a positive impact of Alterion on gut morphology (Figure 3).
In addition to longer microvilli, Alterion also has direct positive influence on the intestinal barrier integrity, demonstrated by an in vitro model (Caco-2 monolayer cells in Transwell System). A pro-inflammatory cytokine (TNF-α) was introduced to impair intestinal frontier integrity. Transepithelial electrical resistance (TEER, voltage between the apical and basolateral side) was measured, corresponding to the passage of the molecules. The higher the TEER, the better the integrity. In addition, paracellular pathway with D-mannitol was also monitored to further evaluate the integrity of the intestinal barrier. More D-mannitol passing from apical to basolateral means that the integrity of the barrier is damaged.
As expected, the stimulation with TNF-α decreased TEER and increased D-Mannitol flux. Alterion was able to restore both parameters to levels similar to non-stimulated cells (Figure 4).
Probiotics have been recognized and widely used as one of most effective AGP alternative solutions. In order to check their direct anti-inflammatory properties, the same in vitro model as above is used. Alterion was tested and compared with two other Bacillus subtilis strains found in commercial probiotic products (named Bs A and Bs B), benchmarked by epigallocatechin gallate (EGCG) as strong anti-inflammatory compound.
The Caco-2 cells were then stimulated by exposing to interleukin 1 beta (IL-1 beta), an inducer to mimic inflammatory response. The level of inflammatory response was measured against the production of interleukin 8 (IL-8), a marker of acute inflammation. As shown in Figure 5, upon stimulation of the Caco-2 cells with IL-1, a huge increase in IL8 production was observed (negative control – non-stressed cells vs positive control – stressed cells), and EGCG was very effective in preventing the inflammatory response. Even though all Bacillus strains tended to reduce inflammatory response, Alterion was much more effective, decreasing the production of IL-8 to the level obtained with EGCG.
In the broiler study mentioned previously, Alterion supplementation led to a 5.4% improvement in FCR, 5.7% increase in final body weight and an unchanged feed intake in growing broilers (Table 1). These economically important parameters are therefore greatly influenced by gut health, through in particular changes in the gut microbiota and mucosa.
As summary, Alterion, when administrated to broilers, improves the microbiota balance, strengthen mucosal barrier function and prevent inflammation, which all together contribute to a healthier gut and optimal bird performance.