White shrimp (Litopenaeus vannamei), one of the most widely farmed crustaceans in the world, can be cultured in fresh seawater due to its eurysalinity (1). As one of the high density breeding species in China, white shrimp farming has been plagued by intestinal dysfunction caused by an inflammatory response, which can be induced by the infection of pathogenic bacteria from Aeromonas, gram-negative microbes (2). The intestinal dysfunction could ulteriorly lead to metabolic dysfunction and trigger various diseases, followed by substantial economic losses to farmers. As we know, the intestine inhabits a complex micro-ecological system, and intestinal inflammation is confirmed to be closely related to microbial dysbiosis (3). The intestine is a crucial digestion organ, while it is also an essential immunity organ for animals. And thus, finding an effective way to solve the problem of intestinal dysfunction would promote the rapid development of shrimp culture.

β-glucan is a well-known immunostimulant, which can be extracted from cell walls of fungi, algae, and natural bioactive polysaccharides and contains three different specific glycoside linkage sources, including β- (1, 3)/β- (1, 4)/β- (1, 5) (4). The β-glucan has been widely applied to the aquaculture industry due to its positive impact on the health of aquatic animals (5, 6). Numerous studies in β-1,3-glucan have confirmed its property of anti-inflammatory, immune-enhancing, and anti-oxidation in aquatic animals such as white shrimp (7–9), marine swimming crab (Portunus trituberculatus), hybrid striped bass (Morone chrysops × M. saxatilis) (10), and large yellow croaker (Pseudosciaena crocea) (11). Evidence from human hepatocytes suggested that β-1,3-glucan could attenuate inflammation and accumulation of reactive oxygen species (ROS) induced by lipopolysaccharide (12). Similarly, the results in rainbow trout (Oncorhynchus mykiss) (13), silver catfish (Rhamdia quelen) (14), and common carp (Cyprinus carpio L) (15) suggested that β-glucan (including β-1,3-glucan) could reduce the intestinal inflammatory response and oxidative stress induced by the challenge of Aeromonas hydrophila or tissue damage. Additionally, the study in Indian major carp (Labeo rohita) suggested that β-1,3-glucan supplementation could dramatically elevate antibody response after being challenged with the Edwardsiella tarda vaccine (16).

Multitudinous evidence has confirmed that a vital role of intestinal microbiota in the intestinal health of the host, and β-glucan, a complex polysaccharide, is authorized to possess various health-promoting properties through intestinal microbiota regulation (17–19). Several studies explored the effects of β-glucan on the intestinal microbiota of aquatic animals including common carp (20), tilapia (21), rainbow trout (1), and white shrimp (22), mainly focusing on microbial composition. Undoubtedly, the number and richness of species are essential elements for a microbial ecosystem, while complex interspecific interactions are crucial to maintaining intestinal microbiota homeostasis (23). And thus, β-glucan supplementation altered the composition of intestinal microbial community and would further affect interactions between different species within the microbial community. It is well accepted that the microbes perform different roles due to their ecological niches, and the critical microbes were considered essential in maintaining intestinal microbiota homeostasis (24). Since 100-fold more genes than hosts possessed by microbes could synthesize many enzymes and other products, these symbiotic microbes inhabiting the intestine could participate in host nutrition metabolism and affect intestinal health through their metabolites (25–27). Hence, the intestinal microbiota is considered as an auxiliary metabolic organ involved in the metabolic process of the host (28).

Although β-1,3-glucan displayed positive effects on intestinal health, mechanisms explaining observed immunomodulatory in white shrimp have mainly remained obscure; hence, the purpose of the current study was to explore the underlying mechanism of β-1,3-glucan improving intestinal health of white shrimp through the investigation of its effects on the intestinal microbiota, inflammatory response, immunity, and antioxidant capacity. The results from this study would provide a better understanding of the mechanism of β-1,3-glucan ameliorating the intestinal health of white shrimp and further promote the application of β-1,3-glucan in the shrimp culture.

Multitudinous pieces of evidences from the animal model and human studies have verified the critical role played by intestinal microbiota in intestinal health, and the diet consumed by the host could shape the microbial composition (25, 28, 35, 36). β-glucan serves as a prebiotic and is confirmed to positively impact the intestinal microbiota and the host’s health (19). Here, β-1,3-glucan supplementation did affect the intestinal microbiota and significantly increased the microbial diversity in white shrimp. Similar results were also observed in tilapia (21), carp (Cyprinus carpio) (37) and white shrimp from a recent study (22). Although the composition of the intestinal microbial community can be dramatically affected by the diets, the core microbiota is essentially stable in the host (38). Here, results indicated that white shrimp fed basal and β-1,3-glucan diet also shared the same core microbiota from Proteobacteria and Firmicutes, even though intestinal microbiota structure was notably altered by dietary supplementation of β-1,3-glucan. The current results suggested that shrimp fed with commercial feed displayed a significant increase in the percentage of Aeromonas species from Gammaproteobacteria, which were confirmed to be opportunistic pathogens in freshwater-farmed white shrimp (2); β-1,3-glucan diet was found to inhibit these Aeromonas species significantly, and dramatically increase the proportion of microbes from Alphaproteobacteria, Betaproteobacteria, Deltaproteobacteria, Clostridiaceae_1, and scarce class groups like Actinobacteria, Chloroflexi, Verrucomicrobia, and Acidobacteria. The increased ratio of these low-class groups led to the rise of the microbial diversity in shrimp.

The interspecific interactions capacitate the microbial community to form an ecological network through which the micro-ecological ecosystem maintains its dynamic balance in intestine (23). In the present study, shrimp fed basal and β-1,3-glucan diets displayed two completely different patterns of microbial interactions referred to as competition and cooperation dominated in submodules of the ecological network, respectively. Based on the r/K selection theory in ecology, the r-strategy species would be the representative community appearing in nutrient-rich environments with characteristics of low competition, high nutrient utilization capacity, and high growth rate (39). Although artificial feeding provided a nutrient-rich environment for the intestinal microbiota, the proliferation of Aeromonas species, the pathogens, intensified the competition within the microbial community for the nutrients in this study. In spatial conditions, the cooperation-dominated communities are more stable because the cooperative interactions are more robust to population disturbance, while the competitive interactions are susceptible to perturbations (40, 41). And thus, the β-1,3-glucan diet promoted intestinal microbiota homeostasis by inhibiting the competition within the microbial community through the suppression of Aeromonas. The dominant microbiota is the main component of the ecological network, in which the generalists referred to connectors and module hubs serve as structural and functional keystones and execute a key role in maintaining the property of the web (42). Therefore, β-1,3-glucan promoted the stability of ecological network by increasing the number of module hubs. Taken together, dietary supplementation of β-1,3-glucan could improve the homeostasis of intestinal microbiota in white shrimp through the inhibition of composition caused by Aeromonas and the increase in number of module hubs.

The intestinal microbiota plays a vital role in host nutrient metabolism since microbial fermentation and nutrient synthesis provide numerous metabolites (25–27). Undoubtedly, the variation in microbial composition would further cause a change in microbial metabolism. The microbial carbohydrate metabolism was more active in the white shrimp fed a β-1,3-glucan diet in response to polysaccharide supplementation. Meanwhile, due to the significant reduction in the ratio of Aeromonas, the microbial lipopolysaccharide biosynthesis was significantly inhibited in shrimp fed β-1,3-glucan diet in this study. As we know, lipopolysaccharide could induce intestinal inflammation by triggering TLR4-mediated inflammatory pathway (43, 44), and this may explain why β-1,3-glucan diet displayed an anti-inflammatory capacity in white shrimp in terms of the significant decrease in expression of inflammatory factor genes in this study. Additionally, β-1,3-glucan also could suppress intestinal inflammation by elevating the activity of intestinal AKP in the present study since the AKP can remove the phosphoric acid groups from lipopolysaccharide, thereby reducing its inflammatory effects (45, 46). Therefore, dietary supplementation of β-1,3-glucan could benefit intestinal health by recovering intestinal dysfunction induced by an inflammatory response.

The β-glucan, as an ideal immunostimulant, is widely applicated in aquaculture and positively impacts the immunity and antioxidant system of aquatic animals. Pieces of evidence from previous studies in white shrimp have confirmed the immune-enhancing effect of β-1,3-glucan (8, 47). The lipopolysaccharide and β-1,3-glucan binding protein (LGBP), vital pattern recognition proteins, can recognize lipopolysaccharide and β-1,3-glucan and subsequently trigger innate immunity (48). Indeed, the present results suggested that the expression of intestinal LGBP was significantly increased in shrimp fed β-1,3-glucan diet, together with an enhancement in immunity parameters in intestinal tissue and serum. Additionally, β-1,3-glucan supplementation enhanced the antioxidant capacity of shrimp, and similar results were also observed in white shrimp fed β-1,3-glucan (7). HSP70 serves as an important oxidative stress biomarker (49), and higher expression of HSP70 in the intestinal tissue shrimp fed β-1,3-glucan confirmed the antioxidant property of β-glucan again. Along with the improvement of intestinal function, the growth performance of white shrimp was also dramatically promoted by β-1,3-glucan finally.

In conclusion, β-1,3-glucan supplementation significantly altered the microbial composition and promoted the homeostasis of intestinal microbiota, and inhibited intestinal inflammatory through the suppression of pathogenic bacteria Aeromonas, concurrent with an enhancement in the immunity, antioxidant capacity, and growth performance of white shrimp.

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: https://www.ncbi.nlm.nih.gov/, PRJNA908810.

The animal study was reviewed and approved by Nanchang University.

GY and MP designed the experiments and supervised the manuscript. KS and LB carried out the animal experiment and sample analysis with the help of ML, WL, QZ, JD, PF, BH, CW, and wrote the manuscript. VK revised the manuscript. All authors read and approved the final manuscript. All authors contributed to the article and approved the submitted version.