Characterising the Gut Microbiomes in Wild and Captive Short-Beaked Echidnas Reveals Diet-Associated Changes

Abstract

The gut microbiome plays a vital role in health and wellbeing of animals, and an increasing number of studies are investigating microbiome changes in wild and managed populations to improve conservation and welfare. The short-beaked echidna (Tachyglossus aculeatus) is an iconic Australian species, the most widespread native mammal, and commonly held in zoos. Echidnas are cryptic animals, and much is still unknown about many aspects of their biology. Furthermore, some wild echidna populations are under threat, while echidnas held in captivity can have severe gastric health problems. Here, we used citizen science and zoos to collect echidna scats from across Australia to perform the largest gut microbiome study on any native Australian animal. Using 16S rRNA gene metabarcoding of scat samples, we characterised and compared the gut microbiomes of echidnas in wild (n = 159) and managed (n = 44) populations, which were fed four different diets. Wild echidna samples were highly variable, yet commonly dominated by soil and plant-fermenting bacteria, while echidnas in captivity were dominated by gut commensals and plant-fermenting bacteria, suggesting plant matter may play a significant role in echidna diet. This work demonstrates significant differences between zoo held and wild echidnas, as well as managed animals on different diets, revealing that diet is important in shaping the gut microbiomes in echidnas. This first analysis of echidna gut microbiome highlights extensive microbial diversity in wild echidnas and changes in microbiome composition in managed populations. This is a first step towards using microbiome analysis to better understand diet, gastrointestinal biology, and improve management in these iconic animals.

Keywords: Australia; EchidnaCSI; captive; digestive physiology; herbivore; insects; nutrition.

Figure 1

Location and diet information for faecal samples collected from wild and captive echidnas in this study. Red dots on map indicate locations of faecal samples collected from wild echidnas, diet labelled as ‘insect’ for simplicity; blue circle with star is the location of Perth Zoo, where faecal samples were collected from echidnas that were fed exclusively the Meat diet; green circle with star is the location of Taronga Zoo, where faecal samples were collected from echidnas fed three different diets: Updated Meat Diet (UMD), Vetafarm diet and Wombaroo diet. Map is coloured according to Australian climate zones.

Figure 2

Alpha diversity analyses of gut microbiomes from wild samples collected in different climate regions across Australia. Whisker-box plots depict the following metrics: (A) Faith’s phylogenetic diversity (Faith’s PD); (B) Observed Amplicon Sequence Variants; (C) Shannon’s Diversity index. Horizontal lines indicate median values, upper and lower bounds represent the 25th and 75th percentiles, and top and bottom whiskers indicate maximum and minimum values. Outliers are shown as grey circles. ^*= 0.01 < p < 0.05.

Figure 3

Taxonomy bar plots of relative frequency of bacteria present in all wild and captive echidna scats at the phylum level. Samples are firstly organised into ‘captive’ and ‘wild’ samples, and the wild samples are further organised by climate class (Supplementary Table S1). Top 20 most abundant phyla are coloured in figure; legend is labelled with most abundant taxa on the left to least abundant taxa on the right. Original and interactive QZV files to view bar plots are available in Supplementary Files.

Figure 4

Differences in microbial composition are observed between samples collected in wild compared to samples collected in captivity from echidnas fed four different diets. (A) PCoA plot of unweighted UniFrac distances showing clustering patterns of wild samples (insect diet) and zoo samples (Meat, UMD, Vetafarm and Wombaroo diets). Green dotted circle shows the Taronga Zoo samples mostly clustering together; Blue dotted circle shows the Perth Zoo samples mostly clustering together between the two major clusters of the Wild samples within the red dotted circles. (B) Taxonomy bar plots showing relative frequencies of bacteria present in echidna scats shown at the genus or family level; all samples have been aggregated according to their diet, and an average relative frequency is shown. Samples are labelled by their diet where insect refers to wild collected samples. UMD, Updated Meat Diet. Original and interactive QZV files to view bar plots are available in Supplementary Files.

Figure 5

Differences in microbial composition are observed between Meat diet and all other diets fed in captivity. (A) PCoA plot of unweighted UniFrac distances showing separation between Meat diet clustering further to the left of Axis 1 (blue dotted circle) and other diets clustering to the right of Axis 1 (green dotted circle). (B) Taxonomy bar plots showing relative frequencies of bacteria present in echidna scats shown at the phylum level. Samples are labelled by their sample ID and diet (Supplementary Figure S2). Most prevalent phyla are included in the legend; as bar colours repeat, the legend is labelled with most abundant taxa on top to least abundant taxa on bottom of legend. UMD, Updated Meat Diet. Original and interactive QZV files to view bar plots are available in Supplementary Files.