High-throughput pyrosequencing of the 16S rRNA gene has been used to identify the intestinal microbiota of many animal species. Nonetheless, little comparative data is available for aquatic invertebrate organism. In particular, only one high-throughput study has been performed on the intestinal microbiota of sea cucumbers, a non-vertebrate deuterostome. We have now studied the intestinal microbiota of the sea cucumber Holothuria glaberrima, an echinoderm well-known for its remarkable power of regeneration. We characterized and compared the microbiota associated to the main three components of H.glaberrima's digestive tract: the anterior descending intestine, the medical intestine (these two comprimse the small intestine) and the posterior descending intestine (or large intestine) in two different environments, the natural marine environment and sea water aquaria. A total of 8,172 OTU's were grouped in 10 bacterial phyla, 23 classes, 44 orders, 83 families, 127 genera and 1 group of unknown bacteria, present across the digestive tract of 10 specimens. The results showed that the anterior intestine is dominated by Proteobacteria (61%) and Bacteroidetes (4%), and the posterior intestine was remarkably different, dominated by Firmicutes (48%) and Bacteroidetes (35%). On the other hand, results from intestine segments samples from specimens kept in indoor sea water aquaria showed that all three intestines segments (anterior, medium, and posterior) are dominated by Firmicutes (54%, 62%, and 57% respectively) and Bacteroidetes (39%, 29%, and 16% respectively), similar to phyla that were observed in the posterior intestine segments from specimens in natural marine environment. Our results evidence differences in the microbiota of several intestinal segments (P= 0.005), and changes in the bacterial composition associated with the animal environment (P= 0.001). Finally, based on our findings, we suggest that the bacteria associated to the posterior segment of the gut could reflect the core microbiota of the h.glaberrima's intestine. This is the first step toward understanding the underlying principles that govern bacterial community assembly in the intestine of H.glaberrima and of other sea cucumbers.