Xenopus laevis tadpoles can regenerate functional tails, containing the spinal cord, notochord, muscle, fin, blood vessels and nerves, except for a brief refractory period at around 1 week of age. At this stage, amputation of the tadpole's tail may either result in scarless wound healing or the activation of a regeneration programme, which replaces the lost tissues. We recently demonstrated a link between bacterial lipopolysaccharides and successful tail regeneration in refractory stage tadpoles and proposed that this could result from lipopolysaccharides binding to Toll-like receptor 4 (TLR4). Here, we have used 16S rRNA sequencing to show that the tadpole skin microbiome is highly variable between sibships and that the community can be altered by raising embryos in the antibiotic gentamicin. Six Gram-negative genera, including Delftia and Chryseobacterium, were over-represented in tadpoles that underwent tail regeneration. Lipopolysaccharides purified from a commensal Chryseobacterium spp. XDS4, an exogenous Delftia spp. or Escherichia coli, could significantly increase the number of antibiotic-raised tadpoles that attempted regeneration. Conversely, the quality of regeneration was impaired in native-raised tadpoles exposed to the antagonistic lipopolysaccharide of Rhodobacter sphaeroides. Editing TLR4 using CRISPR/Cas9 also reduced regeneration quality, but not quantity, at the level of the cohort. However, we found that the editing level of individual tadpoles was a poor predictor of regenerative outcome. In conclusion, our results suggest that variable regeneration in refractory stage tadpoles depends at least in part on the skin

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Background Limb buds develop as bilateral outgrowths of the lateral plate mesoderm and are patterned along three axes. Current models of proximal to distal patterning of early amniote limb buds suggest that two signals, a distal organizing signal from the apical epithelial ridge (AER, Fgfs) and an opposing proximal (retinoic acid [RA]) act early on pattern this axis. Results Transcriptional analysis of stage 51 Xenopus laevis hindlimb buds sectioned along the proximal‐distal axis showed that the distal region is distinct from the rest of the limb. Expression of capn8.3, a novel calpain, was located in cells immediately flanking the AER. The Wnt antagonist Dkk1 was AER‐specific in Xenopus limbs. Two transcription factors, sall1 and zic5, were expressed in distal mesenchyme. Zic5 has no described association with limb development. We also describe expression of two proximal genes, gata5 and tnn, not previously associated with limb development. Differentially expressed genes were associated with Fgf, Wnt, and RA signaling as well as differential cell adhesion and proliferation. Conclusions We identify new candidate genes for early proximodistal limb patterning. Our analysis of RA‐regulated genes supports a role for transient RA gradients in early limb bud in proximal‐to‐distal patterning in this anamniote model organism.

Here, we report the genome sequences of three bacterial isolates, Kinneretia sp. strain XES5, Shinella sp. strain XGS7, and Vogesella sp. strain XCS3, which were cultured from skin of adult female laboratory-bred Xenopus laevis .

Limb buds develop as bilateral outgrowths of the lateral plate mesoderm and are patterned along three axes. Current models of proximal to distal patterning of early amniote limb buds suggest the presence of two signals, a distal organising signal from the apical epithelial ridge (AER) and an opposing proximal signal, that act early on to form the stylopod and zeugopod. Here, we have used transcriptional analysis of early Xenopus laevis hindlimb buds sectioned along the proximal-distal axis, to show support for this model in anamniotes. The distal limb, which contains AER and distal mesenchyme, is transcriptionally distinct from the rest of the limb. Expression of capn8.3.L, a novel calpain, was located in cells immediately flanking the AER suggesting a novel role in determining the boundary of this critical signalling centre. The Wnt antagonist Dkk1 was AER-specific in Xenopus limbs, which differs from the mesenchymal expression in amniotes. Two transcription factors with no known role in limb development, sall1 and zic5, were expressed in distal mesenchyme. We also describe expression of two proximal genes, gata5.L and tnn.L, not previously associated with limb development. Differentially expressed genes were associated with Fgf, Wnt and retinoic acid (RA) signalling as well as differential cell adhesion and proliferation. We find evidence to support transcriptional gradients of RA-regulated genes across the early hindlimb, suggesting that proximal RA signals regulate proximal to distal patterning.