A Gram-staining-negative, aerobic bacterium, designated strain PT3T was isolated from laboratory-reared larvae of the Japanese sea cucumber Apostichopus japonicus. Phylogenetic analysis based on the 16S rRNA gene nucleotide sequences revealed that PT3T was closely related to Amphritea ceti RA1T (= KCTC 42154T = NBRC 110551T) and Amphritea spongicola MEBiC05461T (= KCCM 42943T = JCM 16668T) both with 98.3% sequence similarity, however, average nucleotide identity (ANI) and in silico DNA-DNA hybridization (in silico DDH) values among these three strains were below 95% and 70%, respectively, confirming the novelty of PT3T. Furthermore, the average amino acid identity (AAI) values of PT3T against other Amphritea species were on the reported genus delineation boundary (64–67%). Multilocus sequence analysis using four protein-coding genes (recA, mreB, rpoA, and topA) further demonstrated that PT3T, Amphritea ceti and Amphritea spongicola formed a monophyletic clade clearly separate from other members of the genus Amphritea. Three strains (PT3T, A. ceti　KCTC 42154T and A. spongicola JCM 16668T) also showed higher similarities in their core genomes compared to those of the other Amphritea spp. Based on the genome-based taxonomic approach, Aliamphritea gen. nov. was proposed together with the reclassification of the genus Amphritea and Aliamphritea ceti comb. nov. (type strain RA1T = KCTC 42154T = NBRC 110551T), Aliamphritea spongicola comb. nov. (type strain MEBiC05461T = KCCM 42943T = JCM 16668T), and Aliamphritea hakodatensis sp. nov. (type strain PT3T = JCM 34607T = KCTC 82591T) were suggested.

The genus Thalassotalea is ubiquitous in marine environments, and up to 20 species have been described so far. A Gram-staining-negative, aerobic bacterium, designated strain PTE2T was isolated from laboratory-reared larvae of the Japanese sea cucumber Apostichopus japonicus. Phylogenetic analysis based on the 16S rRNA gene nucleotide sequences revealed that PTE2T was closely related to Thalassotalea sediminis N211T (= KCTC 42588T = MCCC 1H00116T) with 97.9% sequence similarity. ANI and in silico DDH values against Thalassotalea species were 68.5–77.0% and 19.7–24.6%, respectively, indicating the novelty of PTE2T. Based on genome-based taxonomic approaches, strain PTE2T (= JCM 34608T = KCTC 82592T) is proposed as a new species, Thalassotalea hakodatensis sp. nov.

Sea cucumbers possess the remarkable capacity to regenerate their body parts or organs. Regeneration of host organs and/or body parts involves reconstruction of the host associated microbiota, however, the dynamics and contribution of microbiota to the regeneration process are largely unknown due to a lack of experimental models. To track the dynamics of individual gut microbiomes during gut regeneration, both caged mariculture and laboratory isolator systems of sea cucumbers (Apostichopus japonicus) were developed and longitudinal meta16S analyses were performed. Under natural environmental conditions in the caged mariculture system, both bacterial and eukaryotic communities in sea cucumbers’ guts appeared to be reconstructed within 4 months after evisceration. Using the laboratory isolator, which can trace daily dynamics, we found that fecal microbiota collected before evisceration were clearly different from those collected after evisceration. We also identified eight key bacteria, belonging to Alteromonadaceae, Rhodobacteraceae, Oceanospirillaceae and family-unassigned Gammaproteobacteria, suggesting that these bacteria might interact with the host during the gut regeneration process. Six of the eight key bacteria were isolated for further bioassay using the isolator developed in this study to test whether these isolates affect gut regeneration.

Sea urchin is an indicator of coastal environmental changes in the global warming era, and is also a model organism in developmental biology and evolution. Due to the depletion of wild resources, new aquaculture techniques for improving stocks have been well studied. The gut microbiome shapes various aspects of a host’s physiology. However, these microbiome structures and functions on sea urchins, particularly Mesocentrotus nudus and Strongylocentrotus intermedius which are important marine bioresources commonly found in Japan, have not been fully investigated yet. Using metagenomic approaches including meta16S and shotgun metagenome sequencings, the structures, functions, and dynamics of the gut microbiome of M. nudus and S. intermedius, related to both habitat environment and host growth, were studied. Firstly, a broad meta16S analysis revealed that at the family level, Psychromonadaceae and Flavobacteriaceae reads (38–71%) dominated in these sea urchins, which is a unique feature observed in species in Japan. Flavobacteriaceae reads were more abundant in individuals after rearing in an aquarium with circulating compared to one with running water. Campylobacteraceae and Vibrionaceae abundances increased in both kinds of laboratory-reared sea urchins in both types of experiments. 2-weeks feeding experiments of M. nudus and S. intermedius transplanted from the farm to laboratory revealed that these gut microbial structures were affected by diet rather than rearing environments and host species. Secondly, further meta16S analysis of microbial reads related to M. nudus growth revealed that at least four Amplicon Sequence Variant (ASV) affiliated to Saccharicrinis fermentans, which is known to be a nitrogen (N2) fixing bacterium, showed a significant positive correlation to the body weight and test diameter. Interestingly, gut microbiome comparisons using shotgun metagenome sequencing of individuals showing higher and lower growth rates revealed a significant abundance of “Nitrate and nitrite ammonification” genes in the higher-grown individuals under the circulating water rearing. These findings provide new insights on the structure-function relationship of sea urchin gut microbiomes beyond previously reported nitrogen fixation function in sea urchin in 1950s; we discovered a nitrate reduction function into ammonium for the growth promotion of sea urchin.

There is a lot of evidence indicating pioneer microbes in early life having various effects on later host biology. Because of the influential phylogenetic position of sea cucumber, which is a deep branching clade in Deuterostomia, the attention on the microbiome in sea cucumber has been increasing. Although microbes in sea cucumber have been reported in several studies, there is a lack of knowledge regarding the pioneer microbiota in the early life stages of sea cucumber. In this study, microbiota changes during the larval development of sea cucumber were assessed using a laboratory rearing system. Microbial community structure was likely to be related to the developmental stage and significant alterations were detected in the late auricularia stage. The relative abundances of Oceanospirillales, Alteromonadales, and Rhodobacterales significantly varied after gut formation. A total of 257 strains were isolated from larval developmental stages of sea cucumber and affiliated to 124 ASVs in the metagenomic analysis. This data demonstrates for the first-time dynamic changes of sea cucumber microbiota in the developmental stages in early life.

A Gram-staining-negative, oxidase-positive, strictly aerobic rod-shaped bacterium, designated strain PT1T, was isolated from the laboratory-reared larvae of the sea cucumber Apostichopus japonicus. A phylogenetic analysis based on the 16S rRNA gene nucleotide sequences revealed that PT1T was closely related to Neptuniibacter marinus ATR 1.1T (= CECT 8938T = DSM 100783T) and Neptuniibacter caesariensis MED92T (= CECT 7075T = CCUG 52065T) showing 98.2% and 98.1% sequence similarity, respectively. However, the average nucleotide identity (ANI) and in silico DNA-DNA hybridization (DDH) values among these three strains were 72.0%-74.8% and 18.3%-19.5% among related Neptuniibacter species, which were below 95% and 70%, respectively, confirming the novel status of PT1T. The average amino acid identity (AAI) values of PT1T showing 74–77% among those strains indicated PT1T is a new species in the genus Neptuniibacter. Based on the genome-based taxonomic approach, Neptuniibacter victor sp. nov. is proposed for PT1T. The type strain is PT1T (JCM 35563T = LMG 32868T).