## 5 Discussion  
Microbiota shape the health status of pets. Subsequently, dog owners play a role in determining the microbiota of dogs through their food. Pet foods are formulated to contain the carbohydrates, proteins, fats, minerals, and vitamins, etc. Presently, some pet foods include probiotics in their formulations. Several probiotic supplements have been reported to have health-promoting properties for both healthy and diseased dogs. Probiotics isolated from fresh fecal swabs of healthy dogs could also be used as a potent probiotic candidate as a dietary supplement for dogs [73; 74]. Positive regulation of the microbial ecology in the GI tract has systemic effects. Studies have confirmed that the intervention of probiotics could change the microbiota of dogs. The supplementation of probiotic preparations was shown to increase the abundance of beneficial microbes, such as _Lactobacillus_ spp. and _Bifidobacterium_ spp., and suppress the growth of harmful microbes, such as _Enterobacteriaceae_ and _C. perfringens_[70]. The additional functional foods that are nutritive or nonnutritive and required dietary components to promote the host health are referred as functional nutrition. Functional nutrients and functional foods with immune-regulatory properties are termed as immunohistochemistry. Vitamins (D and E), minerals (Zinc), omega-3 polyunsaturated fatty acids, phytochemicals such as epigallocatechin-3-gallate, microbial metabolites such as SCFA, and probiotics have immunomodulating potentials. The vitamins, zinc, and SCFA are involved in intestinal homeostasis and maintain intestinal integrity [75; 76; 77]. Supplementation of vitamin E and selenium enhanced the immune response in vaccinated dogs, and especially increased the IgG concentration compared to the control, and offered the highest protection [78]. The supplementation of eicosapentaenoic acid (EPA) and docosahexaenoicenoic acid showed significant health improvement in diseased dogs [79]. The intestinal dysbiosis affects the availability of beneficial microbial metabolites, such as SCFA. The use of antibiotics might deplete the anaerobic bacterial load, and drain the SCFA content, particularly butyrate concentrations. Besides, over-immunomodulation has to be taken into account so that the naive functions, such as eliminating microbial infections, of the immune system can work accordingly. Probiotic supplementation resumes the healthy intestinal microbiota and promotes the ideal immune regulation. Thus, factors such as selection of the probiotic strain, dose, and duration have to be optimized properly before practical applications [77]. Dogs cannot synthesize vitamins, but vitamins can be obtained from their diet and the gut microbiota. The gut microbiota has been considered as a rich source of vitamins [80]. The deficiency of folate and cobalamin is considerably related to and maintains gastrointestinal homeostasis. The deficiency of folate might lead to the reduction of regulatory T (Treg) cells (subpopulation of T cells involved in the immune homeostasis), which cause impaired gut homeostasis, and are prone to intestinal inflammation [81]. Cobalamin is a marker of gastrointestinal diseases in companion animals due to the availability and detectability of cobalamin in serum, even in the serum of animals fed with a low cobalamin diet. Hypothetically, reduced serum cobalamin is due to the utilization of available cobalamin by the increased number of intestinal bacteria [82]. The dogs with diarrhea were reported for the low serum cobalamin concentration [83]. [PERSON] et al. [84] reported that a decrease in serum cobalamin concentration was observed in dogs with chronic enteropathies [84], and the oral supplementation of cobalamin rectified the deficiency [85]. [PERSON] et al. [21] reported that the probiotic mixture intervention did not significantly affect the cobalamin concentration, while the serum folate concentration was increased [21]. Hypocobalaminemia is associated with negative outcomes in dogs with chronic gastrointestinal disease [86].