Developmental, Dietary, and Geographical Impacts on Gut Microbiota of Red Swamp Crayfish (Procambarus clarkii)

Abstract

Red swamp crayfish (Procambarus clarkii) breeding is an important economic mainstay in Hubei province, China. However, information on the gut microbiota of the red swamp crayfish is limited. To address this issue, the effect of developmental stage, diet (fermented or non-fermented feed), and geographical location on the gut microbiota composition in the crayfish was studied via high-throughput 16S rRNA gene sequencing. The results revealed that the dominant phyla in the gut of the crayfish were Proteobacteria, Bacteroidetes,Firmicutes, Tenericutes, and RsaHF231. The alpha diversity showed a declining trend during development, and a highly comparable gut microbiota clustering was identified in a development-dependent manner. The results also revealed that development, followed by diet, is a better key driver for crayfish gut microbiota patterns than geographical location. Notably, the relative abundance of Bacteroidetes was significantly higher in the gut of the crayfish fed with fermented feed than those fed with non-fermented feed, suggesting the fermented feed can be important for the functions (e.g., polysaccharide degradation) of the gut microbiota. In summary, our results revealed the factors shaping gut microbiota of the crayfish and the importance of the fermented feed in crayfish breeding.

Keywords: Procambarus clarkii; fermented feed; gut microbiota; high-throughput sequencing; red swamp crayfish; shaping factors.

Figure 1

Sample locations: Jingzhou of Hubei province at the middle reaches of Yangtze river, and Yangzhou and Xuyi of Jiangsu province at lower reaches of Yangtze river. The distance between the sampling locations was indicated.

Figure 2

Relative abundances of the dominant bacteria at phyla level. Proteobacteria, Bacteroidetes, Firmicutes, Tenericutes and RsaHF231 were most dominant across samples. JZ: Jingzhou, XY: Xuyi, and YZ: Yangzhou.

Figure 3

Trajectory of the gut microbiota in crayfishes across developmental stages. (A) The α-diverdity indexes (including observed species, Chao1, Shannon and Simpson) of crayfish during developmental stages. (B). Venn diagram showing OUT numbers change according to sample types. (C) Non-metric Multi-Dimensional Scaling ordination (NMDS) showing that bacterial communities cluster by sample types using the unweighted UniFrac distance. Adonis test, R² = 0.382, p = 0.001. (D) The phylogenetic tree of top 50 OTUs relative to the relative abundance.

Figure 4

Heat map of the top 30 OTUs relative abundance at different developmental stages at the phyla level (A) or at the family level (B).

Figure 5

Dominant bacteria at different developmental stages. (A) Relative abundances of the dominant bacteria at the phyla level. (B) Relative abundances of the dominant bacteria at the family level.

Figure 6

Impact of geography and diet on gut microbiota of crayfish. (A) Non-metric Multi-Dimensional Scaling ordination analysis of gut microbiota in crayfishes related to geographic locations. Adonis test, R² = 0.122, p = 0.001. Red dots: gut samples of crayfishes from JZ fed with fermented feed; green dots: gut samples of crayfishes from XY fed with fermented feed; blue dots: gut samples of crayfish from YZ fed with non-fermented feed. (B) Relative abundances of the dominant bacteria in the guts of crayfish samples from different geographic locations (JZ: Jingzhou; XY: Xuyi; YZ: Yangzhou) at the family level.

Figure 7

Changes of Bacteroidetes and Proteobacteria during developments. The relative abundance of Bacteroidetes (A) and Proteobacteria (B) phyla. Green bars: samples from Jingzhou; orange bars: samples from Xuyi; blue bars: samples from Yangzhou. (C) The composition of gut microbiota from three locations at the genus level. Significance: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ***** p < 0.00001, one-way ANOVA.