A comprehensive comparison of fecal microbiota in three ecological bird groups of raptors, waders, and waterfowl

Abstract

Gut microbiota plays a vital role in maintaining the health and immunity of wild birds. However, less is known about the comparison of fecal microbiota between different ecological groups of wild birds, particularly in the Yellow River National Wetland in Baotou, China, an important transit point for birds migrating all over the East Asia-Australian and Central Asian flyways. In this study, we characterized the fecal microbiota and potential microbial function in nine bird species of raptors, waders, and waterfowl using 16S rRNA gene amplicon sequencing to reveal the microbiota differences and interaction patterns. The results indicated that there was no significant difference in α-diversity, but a significant difference in β-diversity between the three groups of birds. The fecal bacterial microbiota was dominated by Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes in all groups of birds. Furthermore, we identified five bacterial genera that were significantly higher in raptors, five genera that were significantly higher in waders, and two genera that were more abundant in waterfowl. The bacterial co-occurrence network results revealed 15 and 26 key genera in raptors and waterfowls, respectively. The microbial network in waterfowl exhibited a stronger correlation pattern than that in raptors. PICRUSt2 predictions indicated that fecal bacterial function was significantly enriched in the antibiotic biosynthesis pathway in all three groups. Metabolic pathways related to cell motility (bacterial chemotaxis and flagellar assembly) were significantly more abundant in raptors than in waders, whereas waders were enriched in lipid metabolism (synthesis and degradation of ketone bodies and fatty acid biosynthesis). The fecal microbiota in waterfowl harbored more abundant vitamin B6 metabolism, RNA polymerase, and tyrosine and tryptophan biosynthesis. This comparative study revealed the microbial community structure, microbial co-occurrence patterns, and potential functions, providing a better understanding of the ecology and conservation of wild birds. Future studies may focus on unraveling metagenomic functions and dynamics along with the migration routine or different seasons by metagenomics or metatranscriptomics.

Keywords: co-occurrence network; fecal microbiota; metabolic pathways; raptors; waders; waterfowl.

FIGURE 1

Alpha and beta diversity of fecal microbiota among three ecological groups of birds (MQ, SQ, and YQ). Alpha diversity was characterized by Chao1 index (A), Shannon index (B), and Simpson indices (C). Solid horizontal line within a box represents the median, the dots indicate the observed value, the box margins are the interquartile range (50% of the observations), and whisker lines extend for 1.5 times the interquartile range. There was no significant difference in Chao1 index, Shannon index, and Simpson indices (p > 0.05). Venn diagram of ASVs overlapping across MQ, SQ, and YQ based on ASV presence and absence (D). Principal coordinate analysis of fecal bacterial communities from the three groups of birds (E). MQ, raptors; SQ, waders; YQ, waterfowl.

FIGURE 2

Bar chart of relative abundance. The relative abundance (%) of the top 10 abundant bacteria phyla (A) and genera (B) among the three ecological bird groups. The dominant phyla in three groups of birds consisted of Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes. Others, bacteria taxa with ≤1% abundance. MQ, raptors; SQ, waders; YQ, waterfowl.

FIGURE 3

Bacterial co-occurrence network in MQ (A) and YQ (B) based on positive correlation analysis at the genus level. Nodes correspond to genus and edges to the correlation. Node size is proportional to the degree number. Node color represents the associated phylum for each genus. Edge width displays the strength of correlation. The blue edge indicates a positive correlation. Each large circle (e.g., MQ-M1 or YQ-M1) represents a module detected by Louvain method. MQ, raptors; SQ, waders; YQ, waterfowl.

FIGURE 4

Linear discriminant analysis effect size (LEfSe) analysis. The linear discriminant analysis identified significantly different taxon between MQ, SQ, and YQ at the phylum (A) and genus (B) levels with a threshold of LDA score ≥ 4.0 and p < 0.05. Left, logarithm score of LDA analysis for each taxon. Right, relative abundance of different taxon; MQ, raptors; SQ, waders; YQ, waterfowl; LDA, linear discriminant analysis; LEfSe, LDA Effect Size.

FIGURE 5

Microbial functional difference analysis between groups. (A) KOs principal coordinate analysis (PCoA) based on Bray–Curtis distance of predicted KO abundance. (B) Difference in the KEGG pathway identified by LEfSe (LDA score ≥ 3.0 and p < 0.05) between MQ, SQ, and YQ. Heat maps of differential pathways enrichment analysis based on KEGG. MQ, raptors; SQ, waders; YQ, waterfowl.