**1. Introduction**

Human emotional susceptibility in an inter-group is associated with individual differences in the functions of the hormonal and/or neurochemical systems in response to internal and external stimulations. Aggression, as one of emotional disorders, has been commonly defined as feeling of anger or antipathy of an organism during social interaction, leading to hostile of violent behavior or attitude, provoking attack or confront toward another individual [1]. Aggression affects millions of people's health and welfare around the world annually, resulting in significant social destruction and economic costs.

Intestinal bacteria, as a virtual endocrine organ, release various neuroactive compounds directly or indirectly regulating brain function, including the serotonergic system, to regulate host health and behavior through the gut-brain axis [2–10]. Intestinal bacteria, as well as *Bacillus subtilis*, used as probiotics are involved in tryptophan metabolism [11–14]. Tryptophan metabolites have roles in protecting intestinal mucosa from inflammation and regulating gut immune homeostasis [15–18]. Tryptophan, a precursor of 5-HT, directly affects brain 5-HT synthesis as that tryptophan can pass the brain-blood barrier [19, 20]. The brain serotonergic system plays a critical role in regulating behaviors, especially aggression [21, 22]. Concentrations of 5-HT and its metabolites, as well as the density of 5-HT receptors, have been used as major indicators of aggressive behaviors in humans and experimental animals [23–26].

Probiotics (also called psychobiotics or bio-friendly agents), defined "as a source of live (viable) naturally occurring microorganisms (direct-fed microbials, DFMs)", have been used as dietary supplements to target gut microbiota (microbiome) for a novel promising therapeutic approach of various diseases including social stressinduced mental disorders in humans and various experimental animals [27–36]. Different probiotic strains, for example, have been investigated as functional food or therapeutic treatment of various diseases, including *Bifidobacterium bifidum* [37–39]; *Bifidobacterium bifidum* (BGN4) and *Bifidobacterium longum* (BORI) [40, 41]; *Bifidobacterium pseudocatenulatum* [42]; *Lactobacillus helveticus* [31, 32]; *Lactobacillus plantarum* [43]; *Lactobacillus paracasei* (KW3110, [44]); *Lactobacillus rhamnosus* [45]; and *Clostridium butyricum* [46]. The results collected from these studies indicate that the effects of probiotics on physiological homeostasis, immunity, stress resistance, and related health status are affected by multiple factors, including the probiotic species, its concentration, and duration as well as the host's age and health status [47, 48].

*Bacillus subtilis*, as one of the three most common species of probiotic products in the United States [49], has been widely used as functional feed supplements, such as in a number of dairy and non-dairy fermented foods, for improving human health and well-being [50–52]. Similarly, *Bacillus subtilis*, as growth promoters, has been demonstrated to improve chickens' growth performance [53–55]; regulate intestinal microstructure [56] and digestive enzymes [57, 58]; synthesize and release antimicrobial and antibiotic compounds [15, 59–63]; increase immunity [57, 64] and neurochemical activities including serotonin [8, 65, 66] as well as affect animal behavior [67, 68] following various stressors. In addition, *Bacillus subtilis* can overproduce l-tryptophan [11, 69], consequently increase 5-HT in the hypothalamus [70], function as an antidepressant and anti-anxiety agent [71, 72], and eliminate nervous tension in mice [73]. In the current study, chickens were used as an animal model to assess if dietary supplementation of probiotic, *Bacillus subtilis,* reduces aggressive behaviors following social challenge.
