Experimentally induced metamorphosis in highly regenerative axolotl (ambystoma mexicanum) under constant diet restructures microbiota - PubMed (original) (raw)

Experimentally induced metamorphosis in highly regenerative axolotl (ambystoma mexicanum) under constant diet restructures microbiota

Turan Demircan et al. Sci Rep. 2018.

Abstract

Axolotl (Ambystoma mexicanum) is a critically endangered salamander species and a model organism for regenerative and developmental biology. Despite life-long neoteny in nature and in captive-bred colonies, metamorphosis of these animals can be experimentally induced by administering Thyroid hormones (THs). However, microbiological consequences of this experimental procedure, such as host microbiota response, remain largely unknown. Here, we systematically compared host bacterial microbiota associated with skin, stomach, gut tissues and fecal samples, between neotenic and metamorphic axolotls based on 16S rRNA gene sequences. Our results show that distinct bacterial communities inhabit individual organs of axolotl and undergo substantial restructuring through metamorphosis. Skin microbiota among others, shifted sharply, as highlighted by a major transition from Firmicutes-enriched to Proteobacteria-enriched relative abundance and precipitously decreased diversity. Fecal microbiota of neotenic and metamorphic axolotl shared relatively higher similarity, suggesting that diet continues to shape microbiota despite fundamental transformations in the host digestive organs. We also reproduced the previous finding on reduction in regenerative capacity in limbs of axolotl following metamorphosis, highlighting the need to investigate whether shifts in microbiota is causally linked to regenerative capacity of axolotl. The initial results on axolotl microbiota provide novel insights into microbiological aspects of axolotl metamorphosis and will establish a baseline for future in-depth studies.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1

Figure 1

Experimental design for the comparison of neotenic and metamorphic axolotl. Of 48 siblings, a subset of 24 axolotls (9 axolotls for metamorphosis and 15 axolotls for regeneration experiments) were randomly selected and induced metamorphosis by T4 hormone administration while the rest kept untreated in neoteny. 30 animals (15 neotenic and 15 metamorphosed) were used in regeneration experiments and 18 animals (9 neotenic and 9 metamorphic) were housed individually for microbiome analysis. Each sample groups for skin, gut, stomach and fecal samples consisted of 3 replicates following randomization and pooling.

Figure 2

Figure 2

Effect of metamorphosis on bacterial diversity between neotenic and metamorphic axolotl. Box plots illustrate the comparison of diversity indices; Observed (a), Chao1 (b), Shannon (c) and Faith’s Phylogenetic Diversity (PD) measures.

Figure 3

Figure 3

Beta diversity analysis based on Bray-Curtis distance matrix showing separation of neotenic and metamorphic bacterial communities. Samples were compared using PCO (a) and Canonical Analysis of Principal Coordinates (CAP) (b) methods.

Figure 4

Figure 4

Mean relative abundances of 16S rRNA sequences. Phylum level relative abundance as bar chart (a), genus level relative abundances shown as heatmap (Individual taxa displayed if the its abundance in any sample ≥5%). Samples and bacterial taxa were clustered using average linkage hierarchical clustering of a distance matrix based on Bray–Curtis distance and taxa abundances, respectively. Samples from each group were color coded on the column side bar as follows: aqua samples (brown); samples from neotenic axolotl (light slate blue), metamorphic axolotl (magenta) (b).

Figure 5

Figure 5

Differentially enriched genus level taxa and indicator species in samples from neotenic and metamorphic axolotl. The color scale bar indicates log2 fold changes for absolute OTU abundances (DESeq2 analysis (q < 0.01) (**a**), Bubble plot showing indicator species. Only highly significant indicator values (IndVal >0.7, q < 0.01) are displayed. Size of bubble symbol is proportional to the mean relative abundance of indicator OTUs and the color scale bar shows indicator value for each OTU (b).

Figure 6

Figure 6

Venn diagrams showing the number of unique and shared OTUs. Skin and aqua samples (a), and gut and fecal samples (b), collected from neotenic and metamorphic axolotls as indicated in the diagram.

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