4 | DISCUSSION
Feeding habit reflects directly the digestive ability of fish to different nutrient components. The ability of fish to digest and utilize different nutrients in feed is affected by the structure of digestive tract, the secretion of digestive enzymes and the composition and diversity of intestinal microbes. This study explored the relationship among feeding habits of omnivorous, carnivorous, herbivorous and filter-feeder fish species and digestive physiology, intestinal morphology, and intestinal microbial diversity. We also predicted the functions of microbiome from the four fishes. We found clear differences in the enzymes activities of the four fish species depending on their feeding habits. Evidently, the carnivorous fish (S. chuatsi ) had higher trypsin and pancrelipase activities in the hepatopancreas and enteropeptidase in the intestine than herbivorous (C. idella ), omnivorous (C. auratus ) and the filter-feeder fish (S. grahami ). The variations in digestive enzymes are caused by the different feeding habits (Xu et al., 2011). Accordingly, the activities of trypsin, intestinal protease and pancreatic lipase were roughly carnivorous > omnivorous > herbivorous, reflecting the feeding on animal materials with high protein and lipid requiring secretions of related enzymes to digest them. Similarly, Liu et al. (2014) reported higher protease activity in carnivorous fish than omnivorous and herbivorous fish.
However, the intestinal lipase activity in this study was higher in herbivorous and omnivorous fish species than filter-feeder and carnivorous fish species. Contrary, Parrizas et al. (1994) reported higher lipase activity in carnivorous fish than herbivorous and omnivorous fish. The higher lipase activity in stomachless fish is probably due to the relationship between intestinal tissue structure and digestive enzymes (Pan et al., 1996). In our study, the mandarin fish (S. chuatsi ) represents a carnivorous fish with a stomach, grass carp (C. idellus ) and Dianchi high-back crucian carp (C. auratus ) are typical herbivorous and omnivorous species, respectively, without stomach and, Kunming Schizothoracin (S. grahami ) is a filter-feeder fish with an enlarged sac between eosophagus and intestinal tract, which can secrete digestive fluid and perform some stomach functions. Therefore, the higher intestinal lipase activity in the stomachless fish was due to secretions of the enzyme from the intestine, which performs some functions of the stomach. Interestingly, this study found higher pancreatic amylase and intestinal amylase activities in herbivorous than carnivorous species. Similarly, Li et al. (2012) found higher amylase in herbivorous than omnivorous and Liu and Zhang (2001) reported higher amylase activity in omnivorous fish than carnivorous fish. The higher amylase activity in herbivorous fish is related to the utilization of carbohydrates in the feed. Similarly, omnivorous fish can use higher carbohydrate levels than carnivorous fish (Li et al., 2015a).
This study found significant differences in intestinal villi height, villi width and muscularis thickness among the four fish. These variations are due to the morphological and structural characteristics of the fish gut reflected by the different feeding habits (Liu & Zhang, 2001; Zeng & Ye, 1998). The intestinal microvilli of carnivorous fish had many branches and complex structures. The complexity of the microvilli (villi length and villi width) and muscularis thickness, was in the order carnivorous > herbivorous > omnivorous. The intestinal structure accommodates, transports and digests feed and absorbs nutrients. The height, width and complexity of intestinal microvilli increase the surface area for digestion and absorption of digested nutrients (Sun et al., 2019). The muscularis thickness is composed of smooth muscle, which promotes the movement of food in the intestine through rhythmic relaxation and contraction. The thickness of the muscle layer directly reflects the contraction and peristalsis ability of the intestine. Accordingly, strengthening intestinal contraction and peristalsis is an effective means to increase feed digestion and reduces chyme circulation (Bian et al., 2021). Generally, carnivorous fish have shorter intestines (Day et al., 2014). Accordingly, carnivorous increase the complexity of the intestinal structure to reduce circulation rate of chyme and enhance absorption of nutrients. Therefore, the complexity of the intestinal structure is adapted to the feeding habits of fish so as to achieve fully absorption of nutrients.
The intestinal microflora of vertebrates plays an important role in host nutrition, (Liu et al., 2016; Valdes et al., 2018). Many studies have shown that dietary feeding habits (Miyake et al., 2015; Zhou et al., 2021) and host species (Li et al., 2019; Youngblut et al., 2019) are the main factors affecting the gut microbiota of fish. This study also found that the diversity of fish gut microbiota was significantly affected by feeding habits and host species. The diversity of gut microbiota species and abundance determine the stability of the host gut microbiota associated with host nutrition (Kuang et al., 2020). The community diversity of the four fish species followed the order omnivorous > herbivorous > filter-feeder > carnivorous. Contrary, Li et al. (2015b) reported higher bacterial diversity in the gut of the filter-feeding than herbivorous. It has been reported that the higher the Shannon index, the better the stability of the bacterial community and the better the digestion of nutrients (Zhang et al., 2019).
The results of this study showed Proteobacteria and Firmicutes phyla as typical dominant flora in the gut of the four fish species. Proteobacteria and Firmicutes are two phyla typical dominant flora in the intestine of many fish, such as Oncorhynchus mykiss(Ingerslev et al., 2014), Nibea coibor and Nibea diacanthus (Li et al., 2019), Megalobrama terminalis (Liu et al., 2021), Micropterus salmoides (Zhou et al., 2021) andSymphysodon haraldi (Zhang et al., 2021). The four fish species had variations in abundant of microbiota species. The different symbiotic bacteria carried by fish species may be caused by the selective enrichment of different microorganisms due to variations in feeding and host species. A previous study indicated that during evolution, hosts tend to acquire suitable environmental bacteria by recognizing adhesion mechanisms on the cell surface (McFall-Ngai, 2015). In addition, the differential enrichment of specific flora under different feeding conditions may also adapt to the function of the flora, for example, Bacillus species and Cetacea species are potential candidates for probiotics (Larsen et al., 2014),Pseudomonas species produces vitamin B12, andFusobacterium species produces butyrate (Zhou et al., 2019). Accordingly, the carnivorous gut was dominated by A. johnsonii ,A. lückii and P. stutzeri bacteria species, which may contribute to the digestion of proteins, while Bacteroidesspecies were dominant in herbivorous fish gut may help the host to digest cellulose. The presence of these microbiome in the fish species are useful in host nutrition. Indeed, the intestinal microbes of the carnivorous (S. chuatsi ) showed higher protein digestion potential and lower carbohydrate digestion potential, while gut microbes of herbivorous fish (C. idellus ) showed lower protein and high carbohydrate digestion, consistent with their feeding habits.