Search

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 (N-2) 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 abundanc

Vibrionaceae is one of the most diverse bacterial families and is currently classified into over 50 clades, some members of which play an important role in the symbiotic relationships with humans and animals. Halioticoli clade, which currently consists of 10 species: 8 species associated with the gut of abalone (symbiotic), 1 species (V. breoganii) from bivalves, and 1 species (V. ishigakensis) from subtropical seawater (planktonic). To accelerate studies in the evolution, ecogenomics, and biotechnology of Halioticoli clade species, the genomic backbones and pangenome analyses based on complete genome sequences are needed. Genome sizes of Halioticoli clade species ranged from 3.5 Mb to 4.8 Mb, with V. ishigakensis the biggest. The evolutionary relationships using multilocus sequence analysis based on eight housekeeping genes and 125 single-copy core genes revealed a division of five sub-clades in this clade; 1) V. breoganii, V. comitans, V. inusitatus and V. superstes, 2) V. ezurae, V. neonatus, and V. halioticoli, 3) V. rarus, 4) V. gallicus, and 5) V. ishigakensis. The pan-genomic analysis combined with function and metabolism estimations showed that the planktonic group (sub-clade 5) contained the greatest number of specific genes, and more genes responsible for carbohydrate metabolisms, especially the genes encoding D-galactonate degradation. These results demonstrated that the genome expanded by acquiring more abilities for utilizing various carbohydrates during the evolution from symbiotic to a planktonic lifestyle. Moreover, according to Carbohydrate-Active enZYmes (CAZy) profiling, genes encoding alginate degrading enzymes (aly), classified into PL6, PL7, PL15, and PL17 were common in the ten genomes, but sub-clade 1 had the most. Meanwhile, sub-clade 1and 5 also possessed abundant genes related to macroalgae substrates degradation (GHs), which are also responsible for the genome expansion of sub-clade 1 and 5.

Light is one of the most critical stimuli in the majority of living organisms. In the last two decades, blue light (BL) has become a major subject of attention because of developments in light-emitting diodes (LED). The effects of BL on eukaryotic organisms and phototrophic prokaryotes have been well studied, but the knowledge of its effects on non-phototrophic prokaryotes remains unclear. Since BL can penetrate seawater, it is expected that most prokaryotes living in the ocean possess molecular mechanisms which protect against BL. The aim of this study is to assess the molecular mechanisms of Vibrio parahaemolyticus cells against BL as a marine bacterial model compared to other wavelength light exposures. Physiological and transcriptomic analyses of BL-exposed cells compared to other light treated cells revealed the highest ROS fold change, the highest number of differentially expressed genes (DEGs), and up-regulation in the gene responsible to not only compatible solute such as glycine betaine and ectoine but also iron-sulfur biosynthesis related to ROS formation. Furthermore, red light (RL) up-regulated the expression of cryptochrome DASH, a protein known to be excited by BL, and orange light (OL) decreased the expression of thermostable direct hemolysin (TDH), suggesting that OL attenuates the virulence of V. parahaemolyticus. In addition, the expression of VtrA (V. parahaemolyticus type III secretion system 2 (T3SS2) regulator A) but not VtrB (V. parahaemolyticus T3SS2 regulator B) increased under both light treatments, indicating that light exposure is unlikely to be involved in T3SS2-mediated pathogenicity. These results expand our knowledge on unique light responses in non-phototrophic marine prokaryotes.

Currently, over 190 species in family Vibrionaceae, including not-yet-cultured taxa, have been described and classified into over nine genera, in which the number of species has doubled compared to the previous vibrio evolutionary update (Vibrio Clade 2.0) (Sawabe et al. 2014). In this study, "Vibrio Clade 3.0," the second update of the molecular phylogenetic analysis was performed based on nucleotide sequences of eight housekeeping genes (8-HKGs) retrieved from genome sequences, including 22 newly determined genomes. A total of 51 distinct clades were observed, of which 21 clades are newly described. We further evaluated the delineation powers of the clade classification based on nucleotide sequences of 34 single-copy genes and 11 ribosomal protein genes (11-RPGs) retrieved from core-genome sequences; however, the delineation power of 8-HKGs is still high and that gene set can be reliably used for the classification and identification of Vibrionaceae. Furthermore, the 11-RPGs set proved to be useful in identifying uncultured species among metagenome-assembled genome (MAG) and/or single-cell genome-assembled genome (SAG) pools. This study expands the awareness of the diversity and evolutionary history of the family Vibrionaceae and accelerates the taxonomic applications in classifying as not-yet-cultured taxa among MAGs and SAGs.

The Splendidus clade is the largest clade in Vibrionaceae, and its members are often related to mortality of marine animals with huge economic losses. The molecular bases of their pathogenicity and virulence, however, remain largely unknown. In particular, the complete genome sequences of the Splendidus clade species are rarely registered, which is one of the obstacles to predict core and/or unique genes responsible for their adaptation and pathogenicity, and to perform a fine scale meta-transcriptome during bacterial infection to their hosts. In this study, we obtained the complete genomes of all type strains in the Splendidus clade and revealed that (1) different genome sizes (4.4-5.9 Mb) with V. lentus the biggest and most of them had several big plasmids, likely because of the different features on mobilome elements; (2) the Splendidus clade consists of 19 species except V. cortegadensis, and 3 sub-clades (SC) were defined with the 15 most closely related members as SC1; (3) different carbohydrate degradation preferences may be the result of environmental adaptation; and (4) a broad prediction of virulence factors (VFs) revealed core and species unique VF genes.

Vibriosis accounts for 66.7% of diseases reported in groupers' cultures and affects almost all stages of growth. The disease could lead up to mortality up to 50% mortality, and it was reported that high stocking density and poor fish handling were among the factors that contributed to the disease dissemination. V. harveyi has been reported to be among the causative agent and has caused acute mortality in cage groupers. In this study, we report the genome of V. harveyi VH1 isolated from a diseased tiger grouper Epinephelus fuscoguttatus, reared in a cage farm located in the coastal area of Langkawi. (c) 2022 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)

Light is one of the most critical stimuli in the majority of living organisms. In the last two decades, blue light (BL) has become a major subject of attention because of developments in light-emitting diodes (LED). The effects of BL on eukaryotic organisms and phototrophic prokaryotes have been well studied, but the knowledge of its effects on non-phototrophic prokaryotes remains unclear. Since BL can penetrate seawater, it is expected that most prokaryotes living in the ocean possess molecular mechanisms which protect against BL. The aim of this study is to assess the molecular mechanisms of Vibrio parahaemolyticus cells against BL as a marine bacterial model compared to other wavelength light exposures. Physiological and transcriptomic analyses of BL-exposed cells compared to other light treated cells revealed the highest ROS fold change, the highest number of differentially expressed genes (DEGs), and up-regulation in the gene responsible to not only compatible solute such as glycine betaine and ectoine but also iron-sulfur biosynthesis related to ROS formation. Furthermore, red light (RL) up-regulated the expression of cryptochrome DASH, a protein known to be excited by BL, and orange light (OL) decreased the expression of thermostable direct hemolysin (TDH), suggesting that OL attenuates the virulence of V. parahaemolyticus. In addition, the expression of VtrA (V. parahaemolyticus type III secretion system 2 (T3SS2) regulator A) but not VtrB (V. parahaemolyticus T3SS2 regulator B) increased under both light treatments, indicating that light exposure is unlikely to be involved in T3SS2-mediated pathogenicity. These results expand our knowledge on unique light responses in non-phototrophic marine prokaryotes.

Currently, over 190 species in family Vibrionaceae, including not-yet-cultured taxa, have been described and classified into over nine genera, in which the number of species has doubled compared to the previous vibrio evolutionary update (Vibrio Clade 2.0) (Sawabe et al. 2014). In this study, "Vibrio Clade 3.0," the second update of the molecular phylogenetic analysis was performed based on nucleotide sequences of eight housekeeping genes (8-HKGs) retrieved from genome sequences, including 22 newly determined genomes. A total of 51 distinct clades were observed, of which 21 clades are newly described. We further evaluated the delineation powers of the clade classification based on nucleotide sequences of 34 single-copy genes and 11 ribosomal protein genes (11-RPGs) retrieved from core-genome sequences; however, the delineation power of 8-HKGs is still high and that gene set can be reliably used for the classification and identification of Vibrionaceae. Furthermore, the 11-RPGs set proved to be useful in identifying uncultured species among metagenome-assembled genome (MAG) and/or single-cell genome-assembled genome (SAG) pools. This study expands the awareness of the diversity and evolutionary history of the family Vibrionaceae and accelerates the taxonomic applications in classifying as not-yet-cultured taxa among MAGs and SAGs.

A novel thermophilic bacterium, strain SSM-sur55(T), was isolated from a chimney structure at the Urashima site on the Southern Mariana Trough in the Pacific Ocean. Growth was observed at temperatures between 25 and 60 degrees C (optimum, 55 degrees C; 180 min doubling time), at pH values between 5.3 and 7.2 (optimum, pH 5.9) and in the presence of between 1.6 and 5.6% (w/v) NaCl (optimum, 3.2%). The isolate used molecular hydrogen as its sole energy source, carbon dioxide as its sole carbon source, ammonium as its sole nitrogen source, and elemental sulfur as its sole sulfur source. Thiosulfate, molecular oxygen (0.1%, v/v) or elemental sulfur was utilized as its sole electron acceptor. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain SSM-sur55(T) belonged to the genus Hydrogenimonas of the class "Campylobacteria", and its closest relative was Hydrogenimonas thermophila EP1-55-1%(T) (94.9%). On the basis of the phylogenetic, physiological and molecular characteristics, strain SSM-sur55(T) represents a novel species within the genus Hydrogenimonas, for which the name Hydrogenimonas urashimensis sp. nov. is proposed, with the type strain SSM-sur55(T) (JCM 19825 = KCTC 15926).

Nitrous oxide (N2O) is a potent greenhouse gas and has significantly increased in the atmosphere. Deep-sea hydrothermal fields are representative environments dominated by mesophilic to thermophilic members of the class Campylobacteria that possess clade II nosZ encoding nitrous oxide reductase. Here, we report a strain HRV44(T) representing the first thermophilic campylobacterium capable of growth by H-2 oxidation coupled to N2O reduction. On the basis of physiological and genomic properties, it is proposed that strain HRV44(T) (=JCM 34002 = DSM 111345) represents a novel species of the genus Nitratiruptor, Nitratiruptor labii sp. nov. The comparison of the N2O consumption ability of strain HRV44(T) with those of additional Nitratiruptor and other campylobacterial strains revealed the highest level in strain HRV44(T) and suggests the N2O-respiring metabolism might be the common physiological trait for the genus Nitratiruptor. Our findings provide insights into contributions of thermophilic Campylobacteria to the N2O sink in deep-sea hydrothermal environments.

Deposit‐feeding sea cucumbers repeat ingestion of sediments and excretion of faeces daily and consequently increase bacterial abundance in sediments and promote organic matter mineralization. Such ecological roles are expected to be collaborative activities of sea cucumbers and the gut microbiota. Here, we performed a spatiotemporally broad 16S rRNA gene analysis using 109 samples from sea cucumber faeces and habitat sediments to explore potential contribution of their gut microbiota to the ecological roles. Most operational taxonomic units (OTUs) observed in the faecal samples were shared with the sediment samples, nevertheless faecal and sediment microbiota differed from each other in UniFrac analysis. Lower bacterial diversity and increased relative abundance of specific OTUs in the faecal microbiota strongly suggest selective enrichment of ingested sediment microbiota in their guts. Interestingly, representative faecal OTUs were more abundant in sea cucumber‐populated sediments than in un‐inhabited sediments, indicating bacteria selectively enriched in the guts were spread on ambient sediments via faeces. Moreover, the predicted microbial community metabolic potential showed a higher abundance of genes related to carbohydrate and xenobiotics metabolisms in faeces than in sediments. Our study suggests the repeated selective enrichment transforms ambient sediment microbial communities and maintains the host's ecological roles by promoting organic matter mineralization.

Biohydrogen is one of the most suitable clean energy sources for sustaining a fossil fuel independent society. The use of both land and ocean bioresources as feedstocks show great potential in maximizing biohydrogen production, but sodium ion is one of the main obstacles in efficient bacterial biohydrogen production. Vibrio tritonius strain AM2 can perform efficient hydrogen production with a molar yield of 1.7 mol H-2/mol mannitol, which corresponds to 85% theoretical molar yield of H-2 production, under saline conditions. With a view to maximizing the hydrogen production using marine biomass, it is important to accumulate knowledge on the effects of salts on the hydrogen production kinetics. Here, we show the kinetics in batch hydrogen production of V. tritonius strain AM2 to investigate the response to various NaCl concentrations. The modified Han-Levenspiel model reveals that salt inhibition in hydrogen production using V. tritonius starts precisely at the point where 10.2 g/L of NaCl is added, and is critically inhibited at 46 g/L. NaCl concentration greatly affects the substrate consumption which in turn affects both growth and hydrogen production. The NaCl-dependent behavior of fermentative hydrogen production of V. tritonius compared to that of Escherichia coli JCM 1649 reveals the marine-adapted fermentative hydrogen production system in V. tritonius. V. tritonius AM2 is capable of producing hydrogen from seaweed carbohydrate under a wide range of NaCl concentrations (5 to 46 g/L). The optimal salt concentration producing the highest levels of hydrogen, optimal substrate consumption and highest molar hydrogen yield is at 10 g/L NaCl (1.0% (w/v)).

Whole genome sequence comparisons have become essential for establishing a robust scheme in bacterial taxonomy. To generalize this genome-based taxonomy, fast, reliable, and cost-effective gnome sequencing methodologies are required. MinION, the palm -sized sequencer from Oxford Nanopore Technologies, enables rapid sequencing of bacterial genomes using minimal laboratory resources. Here we tested the ability of Nanopore sequences for the genome-based taxonomy of Vibrionaceae and compared Nanopore- only assemblies to complete gnomes of fiw Rumoiensis clade species: Vibrio aphrogenes, V. algivorus, V. casei, V. litoralis, and V. rumoiensis. Comparison of overall gnome relatedness indices (OGRI) and multilocus sequence analysis (MLSA) based on Nanopore-only assembly and Illumina or hybrid assemblies revealed that errors in Nanopore-only assembly do not Influence average nucleotide identity (ANI), in silico DNA -DNA hybridization (DDH), G+C content, or MLSA tree topology in Vibrionaceae. Our results show that the genome sequences frorn Nanopore-based approach can be used for rapid species identification based on the OGRI and MLSA.

A novel strain Vibrio aphrogenes sp. nov. strain CA-1004(T) isolated from the surface of seaweed collected on the coast of Mie Prefecture in 1994 [1] was characterized using poly phasic taxonomy including multilocus sequence analysis (MLSA) and a genome based comparison. Both phylogenetic analyses on the basis of 16S rRNA gene sequences and MLSA based on eight protein-coding genes (gapA, gyrB, ftsZ, mreB, pyrH, recA, rpoA, and topA) showed the strain could be placed in the Rumoiensis Glade in the genus Vibrio. Sequence similarities of the 16S rRNA gene and the multilocus genes against the Rumoiensis Glade members, V. rumoiensis, V. algivorus, V. casei, and V. litoralis, were low enough to propose V. aphrogenes sp. nov. strain CA-1004(T) as a separate species. The experimental DNA-DNA hybridization data also revealed that the strain CA-1004(T) was separate from four known Rumoiensis Glade species. The G+C content of the V. aphrogenes strain was determined as 42.1% based on the genome sequence. Major traits of the strain were non motile, halophilic, fermentative, alginolytic, and gas production. A total of 27 traits (motility, growth temperature range, amylase, alginase and lipase productions, and assimilation of 19 carbon compounds) distinguished the strain from the other species in the Rumoiensis Glade. The name V. aphrogenes sp. nov. is proposed for this species in the Rumoiensis Glade, with CA-1004(T) as the type strain (JCM 31643(T) = DSM 103759(T)).

Gut microbiome shapes various aspects of a host's physiology, but these functions in aquatic animal hosts have yet to be fully investigated. The sea cucumber Apostichopus japonicus Selenka is one such example. The large growth gap in their body size has delayed the development of intensive aquaculture, nevertheless the species is in urgent need of conservation. To understand possible contributions of the gut microbiome to its host's growth, individual fecal microbiome comparisons were performed. High-throughput 16S rRNA sequencing revealed significantly different microbiota in larger and smaller individuals; Rhodobacterales in particular was the most significantly abundant bacterial group in the larger specimens. Further shotgun metagenome of representative samples revealed a significant abundance of microbiome retaining polyhydroxybutyrate (PHB) metabolism genes in the largest individual. The PHB metabolism reads were potentially derived from Rhodobacterales. These results imply a possible link between microbial PHB producers and potential growth promotion in Deuterostomia marine invertebrates.

Coral reefs perform a major role in regulating marine biodiversity and serve as hotspot for highly dynamic and diverse microbiomes as holobionts. Corals around Ishigaki, however, are at risk due to tremendous stressors including elevation of seawater temperature, eutrophication and so on. However, no information is currently available on how Vibrio diversity fluctuates spatially and temporally due to environmental determinants in Ishigaki coral reef ecosystems. The aim of this study is to elucidate spatiotemporal Vibrio diversity dynamic at both community and population levels and to assess the environmental drivers correlated to Vibrio abundance and diversity. The Vibrio community identified based on pyrH gene phylogeny of 685 isolates from seawater directly connecting to Ishigaki coral holobionts consisted of 22 known and 12 potential novel Vibrionaceae species. The most prominent species were V. hyugaensis, V. owensii and V. harveyi followed by V. maritimus/V. variabillis, V. campbellii, V. coralliilyticus, and Photobacterium rosenbergii. The Vibrio community fluctuations, assessed by PCoA with UniFrac distance and clustering with Euclidiean distance were varied less not only by year but also by site. Interestingly, significant positive correlation was observed between rising seawater temperature and the abundance of V. campbellii (r = 0.62; P < 0.05) whereas the opposite was observed for V. owensii (r = -0.58; P < 0.05) and the C6 group of V. hyugaensis (r = -0.62; P < 0.05). AdaptML-based microhabitat differentiation revealed that V. harveyi, V. campbellii, P. rosenbergii, and V. coralliilyticus populations were less-ecologically distinctive whereas V. astriarenae and V. ishigakensis were ecologically diverse. This knowledge could be important clue for the future actions of coral conservation.

Carotenoids are a class of naturally occurring pigment, carrying out important biological functions in photosynthesis and involved in environmental responses including nutrition in organisms. Saproxanthin and myxol, which have monocyclic carotenoids with a gamma-carotene skeleton, have been reported to show a stronger antioxidant activity than those with beta-carotene and zeaxanthin. In this research, a yellow-orange bacterium of strain 11shimoA1 (JCM19538) was isolated from a seaweed collected at Nabeta Bay (Shizuoka, Japan). The 16S rRNA gene sequence of strain 11shimoA1 revealed more than 99.99 % similarity with those of Jejuia pallidilutea strains in the family Flavobacteriaceae. Strain 11shimoA1 synthesized two types of carotenoids. One of them was (3R, 3'R)-zeaxanthin with dicyclic structure and another was identified as (3R, 2'S)-2'-isopentenylsaproxanthin, a novel monocyclic carotenoid with pentenyl residue at C-2' position of saproxanthin, using FAB-MS, H-1 NMR, and CD analyses. Culturing strain 11shimoA1 in an alkaline medium at pH 9.2 resulted in a markedly increased in production of 2'-isopentenylsaproxanthin per dry cell weight, but a decreased in zeaxanthin production as compared to their respective production levels in medium with pH 7.0. These carotenoids are likely to play some roles in the adaptation of the bacterium to the environmental conditions.

Recombinant protein production in Escherichia coli is well suited to applications in the basic and applied sciences due to its associated simplicity, cost-effectiveness and the large number of genetic strategies available. However, the active form of the marine bacterial enzyme, alginate lyase, was difficult to express in E. coli cells under standard culture conditions. In this study, we found various calcium compounds that enhanced the expression of the active enzyme. The alyPEEC gene encoding extracellular alginate lyase of Pseudoalteromonas elyakovii IAM 14594 was comprised of a 1197 bp open reading frame encoding 398 amino acid residues with the domain G165 to N398 functioning as the mature enzyme. Three clones, pTPB24 with a 2.7 kb insert containing alyPEEC gene and paeX and their respective promoter regions, pTPB31 with a 1.6 kb insert containing only the alyPEEC gene its own promoter, and pCD11 containing the truncated domain encoding G165 to N398 of AlyPEEC inserted into the pTrcHisB expression vector were constructed and their expression was analyzed. Alginate lyase activity for the three clones was detected in cell-free extract cultured in LB broth containing 50% artificial seawater (ASW), but not in media with LB broth alone. Maximum activity was observed in the clones cultured in broth containing 50–100% ASW, respectively. Further expression analysis using one-by-one element deficient ASW showed that calcium sulfate affected active AlyPEEC expression. Furthermore, in contrast to inorganic calcium, calcium lactate, glyceric acid calcium and calcium propionate enhanced active AlyPEEC expression markedly.

We performed the first broad study aiming at the reconstruction of the evolutionary history of vibrios by means of multilocus sequence analysis of nine genes. Overall, 14 distinct clades were recognized using the SplitsTree decomposition method. Some of these clades may correspond to families, e.g., the clades Salinivibrio and Photobacteria, while other clades, e.g., Splendidus and Harveyi, correspond to genera. The common ancestor of all vibrios was estimated to have been present 600 million years ago. We can define species of vibrios as groups of strains that share >95% gene sequence similarity and >99.4% amino acid identity based on the eight protein-coding housekeeping genes. The gene sequence data were used to refine the standard online electronic taxonomic scheme for vibrios (http://www.taxvibrio.lncc.br).

Chapter 15