Large-scale impact of the 2016 Marine Heatwave on the plankton-associated microbial communities of the Great Barrier Reef (Australia).

The Great Barrier Reef (GBR) is the world's largest coral ecosystem and is threatened by climate change. This study investigated the impact of the 2016 Marine Heatwave (MHW) on plankton associated microbial communities along a ∼800 km transect in the GBR. 16S rRNA gene metabarcoding of archived plankton samples collected from November 2014 to August 2016 in this region showed a significant increase in Planctomycetes and bacteria belonging to the genus Vibrio and Synechococcus during and after the heatwave. Notably, Droplet Digital PCR and targeted metagenomic analysis applied on samples collected four months after the MHW event revealed the presence of several potential pathogenic Vibrio species previously associated with diseases in aquatic animals. Overall, the 2016 MHW significantly impacted the surface picoplankton community and fostered the spread of potentially pathogenic bacteria across the GBR providing an additional threat for marine biodiversity in this area.

Metataxonomic analysis of tissue-associated microbiota in grooved carpet-shell (Ruditapes decussatus) and Manila (Ruditapes philippinarum) clams.

Culture-dependent techniques only permit the study of a low percentage of the microbiota diversity in the environment. The introduction of next generation sequencing (NGS) technologies shed light into this hidden microbial world, providing a better knowledge on the general microbiota and, specifically, on the microbial populations of clams. Tissue-associated microbiota of Ruditapes decussatus and Ruditapes philippinarum (mantle, gills, gonad and hepatopancreas) was analysed in two different locations of Galicia (northwest of Spain) during Spring (April) and Autumn (October), employing a metataxonomic approach. High bacterial diversity and richness were found in all samples where a total of 22,044 OTUs were obtained. In most samples, phylum Proteobacteria was most frequently retrieved, although other phyla as Actinobacteria, Bacteroidetes, Tenericutes, Firmicutes or Chlamydiae also appeared at high relative abundances in the samples. At genus level, great variation was found across tissues and sampling periods. A Nonmetric Multidimensional Scaling (NMDS) and a hierarchical clustering analysis allowed to further analyse the factors responsible for the differences among groups of samples in the different sites. Results showed sample ordination based on tissue origin and sampling periods, pointing out that the microbiota was influenced by these factors. Indeed, predominance of certain genera was observed, such as Endozoicomonas or Methylobacterium in gills and gonads, respectively, suggesting that selection of specific bacterial taxa is likely to occur. So far, this study provided a general picture of the tissue associated microbial population structure in R. decussatus and R. philippinarum clams, which, ultimately, allowed the identification of specific tissue-related taxa.

A deep-sea bacterium related to coastal marine pathogens.

Evolution of virulence traits from adaptation to environmental niches other than the host is probably a common feature of marine microbial pathogens, whose knowledge might be crucial to understand their emergence and pathogenetic potential. Here, we report genome sequence analysis of a novel marine bacterial species, Vibrio bathopelagicus sp. nov., isolated from warm bathypelagic waters (3309 m depth) of the Mediterranean Sea. Interestingly, V. bathopelagicus sp. nov. is closely related to coastal Vibrio strains pathogenic to marine bivalves. V. bathopelagicus sp. nov. genome encodes genes involved in environmental adaptation to the deep-sea but also in virulence, such as the R5.7 element, MARTX toxin cluster, Type VI secretion system and zinc-metalloprotease, previously associated with Vibrio infections in farmed oysters. The results of functional in vitro assays on immunocytes (haemocytes) of the Mediterranean mussel Mytilus galloprovincialis and the Pacific oyster Crassostrea gigas, and of the early larval development assay in Mytilus support strong toxicity of V. bathopelagicus sp. nov. towards bivalves. V. bathopelagicus sp. nov., isolated from a remote Mediterranean bathypelagic site, is an example of a planktonic marine bacterium with genotypic and phenotypic traits associated with animal pathogenicity, which might have played an evolutionary role in the origin of coastal marine pathogens.

Impact of nanoplastics on hemolymph immune parameters and microbiota composition in Mytilus galloprovincialis.

Ocean contamination by micro- and nanoplastics represents a potential threat to marine biota, from bacterial communities to higher organisms. In this work, the effect of in vivo exposure of Mytilus galloprovincialis to amino modified nanopolystyrene (PS-NH2) (10 µg/L, 96 h) on hemolymph immune parameters and microbiota composition were investigated. Nanoplastics significantly affected immune parameters (decreased phagocytosis, increased ROS and lysozyme activity, inhibition of NO production). These changes were associated with a shift in hemolymph microbiota composition, with increase in some genera (Arcobacter-like, Psychrobium, Vibrio), and decreases in others (Shewanella, Mycoplasma). The results indicate that exposure to nanoplastics can impact on the microbiome of marine bivalves, and suggest that downregulation of immune defences induced by PS-NH2 may favour potentially pathogenic bacteria. These data underline how exposure to nanoplastics may represent a potential threat to the complex interplay between innate immunity and host microbiota, thus affecting the homeostatic processes involved in maintenance of organism health.

Dynamics of the Pacific oyster pathobiota during mortality episodes in Europe assessed by 16S rRNA gene profiling and a new target enrichment next-generation sequencing strategy.

Infectious agents such as the bacteria Vibrio aestuarianus or Ostreid herpesvirus 1 have been repeatedly associated with dramatic disease outbreaks of Crassostrea gigas beds in Europe. Beside roles played by these pathogens, microbial infections in C. gigas may derive from the contribution of a larger number of microorganisms than previously thought, according to an emerging view supporting the polymicrobial nature of bivalve diseases. In this study, the microbial communities associated with a large number of C. gigas samples collected during recurrent mortality episodes at different European sites were investigated by real-time PCR and 16SrRNA gene-based microbial profiling. A new target enrichment next-generation sequencing protocol for selective capturing of 884 phylogenetic and virulence markers of the potential microbial pathogenic community in oyster tissue was developed allowing high taxonomic resolution analysis of the bivalve pathobiota. Comparative analysis of contrasting C. gigas samples conducted using these methods revealed that oyster experiencing mortality outbreaks displayed signs of microbiota disruption associated with the presence of previously undetected potential pathogenic microbial species mostly belonging to genus Vibrio and Arcobacter. The role of these species and their consortia should be targeted by future studies aiming to shed light on mechanisms underlying polymicrobial infections in C. gigas.

Exposure to TiO2 nanoparticles induces shifts in the microbiota composition of Mytilus galloprovincialis hemolymph.

It is now recognized that host microbiome, the community of microorganisms that colonize the animal body (e.g. microbiota) and their genomes, play an important role in the health status of all organisms, from nutrient processing to protection from disease. In particular, the complex, bilateral interactions between the host innate immune system and the microbiota are crucial in maintaining whole body homeostasis. The development of nanotechnology is raising concern on the potential impact of nanoparticles-NPs on human and environmental health. Titanium dioxide-nTiO2, one of the most widely NP in use, has been shown to affect the gut microbiota of mammals and fish, as well as to potentially alter microbial communities. In the marine bivalve Mytilus galloprovincialis, nTiO2 has been previously shown to interact with hemolymph components, thus resulting in immunomodulation. However, no information is available on the possible impact of NPs on the microbiome of marine organisms. Bivalves host high microbial abundance and diversity, and alteration of their microbiota, in both tissues and hemolymph, in response to stressful conditions has been linked to a compromised health status and susceptibility to diseases. In this work, the effects of nTiO2 exposure (100 µg/L, 4 days) on Mytilus hemolymph microbiota were investigated by 16S rRNA gene-based profiling. Immune parameters were also evaluated. Although hemolymph microbiota of control and nTiO2-treated mussels revealed a similar core composition, nTiO2 exposure affected the abundance of different genera, with decreases in some (e.g. Shewanella, Kistimonas, Vibrio) and increases in others (e.g. Stenotrophomonas). The immunomodulatory effects of nTiO2 were confirmed by the increase in the bactericidal activity of whole hemolymph. These represent the first data on the effects of NPs on the microbiome of marine invertebrates, and suggest that the shift in hemolymph microbiome composition induced by nTiO2 may result from the interplay between the microbiota and the immune system.

Kiloniella majae sp. nov., isolated from spider crab (Maja brachydactyla) and pullet carpet shell clam (Venerupis pullastra).

Ten Gram-negative, rod-shaped and motile bacterial strains were isolated from spider crab (M27.10, M27.11a, F36.1, F36.4, M56.1, F76.17b, M146.1, M166.3 and M166.6) and pullet carpet shell clam (SBRF 1.10) collected in the coast of Galicia. Analyses of the 16S rRNA genes showed that the strains belong to the genus Kiloniella and have high similarity with the species Kiloniella spongiae (99.44-99.86%) and Kiloniella litopenaei (99.0-99.5%). Strains M56.1T (=CECT 9195, =LMG 29925), M146.1 (=CECT 9193, =LMG 29926) and SBRF 1.10 (=CECT 9194, =LMG 29927) were selected on the basis of genotyping by enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). Phylogenetic analysis based on concatenated sequences of the genes gyrB, ftsZ, rpoD and mreB showed that the isolates form a differentiated branch within the genus Kiloniella. Moreover, the average nucleotide identity (ANIm, ANIb and OrthoANI) and in silico estimated DNA-DNA reassociation values between selected Galician isolates and Kiloniella species were below the established cut-off for species deliniation. The results obtained in the genetic and phenotypical analyses indicate that the isolates represent a new species of the genus Kiloniella, for which the name Kiloniella majae sp. nov. is proposed with strain M56.1T (=CECT 9195T, =LMG 29925T) as the type strain.

Draft Genome Sequence of the Fish Strain Edwardsiella tarda NCIMB 2034.

Edwardsiella tarda is an important pathogen for fish. The strain NCIMB 2034, obtained from the National Collection of Industrial Food and Marine Bacteria, was isolated from unknown diseased fish in the United States. The draft genome sequence has 3.79 Mb with a G+C content of 57.1% and >3,340 protein-coding genes.

Genome sequence of three Psychrobacter sp. strains with potential applications in bioremediation.

To date, the genus Psychrobacter consists of 37 recognized species isolated from different sources, however they are more frequently found in cold and other non-polar environments of low water activity. Some strains belonging to the genus have shown different enzymatic activities with potential applications in bioremediation or food industry. In the present study, the whole genome sequences of three Psychrobacter-like strains (C 20.9, Cmf 22.2 and Rd 27.2) isolated from reared clams in Galicia (Spain) are described. The sequenced genomes resulted in an assembly size of 3,143,782 bp for C 20.9 isolate, 3,168,467 bp for Cmf 22.2 isolate and 3,028,386 bp for Rd 27.2 isolate. Among the identified coding sequences of the genomes, mercury detoxification and biogeochemistry genes were found, as well as genes related to heavy metals and antibiotic resistance. Also virulence-related features were identified such as the siderophore vibrioferrin or an aerobactin-like siderophore. The phylogenetic analysis of the 16S rRNA gene suggested that these strains may represent novel species of the Psychrobacter genus. The genome sequences of the Psychrobacter sp. strains have been deposited at DDBJ/EMBL/GenBank under the accession numbers MRYA00000000 (Cmf 22.2), MRYB00000000 (Rd 27.2) and MRYC00000000 (C 20.9), and the sequences could be found at the site https://www.ncbi.nlm.nih.gov/bioproject/PRJNA353858.

Comparative Genomic Analysis of Two Vibrio toranzoniae Strains with Different Virulence Capacity Reveals Clues on Its Pathogenicity for Fish.

Vibrio toranzoniae is a Gram-negative bacterium of the Splendidus clade within the Vibrio genus. V. toranzoniae was first isolated from healthy clams in Galicia (Spain) but recently was also identified associated to disease outbreaks of red conger eel in Chile. Experimental challenges showed that the Chilean isolates were able to produce fish mortalities but not the strains isolated from clams. The aim of the present study was to determine the differences at the genomic level between the type strain of the species (CECT 7225T) and the strain R17, isolated from red conger eel in Chile, which could explain their different virulent capacity. The genome-based comparison showed high homology between both strains but differences were observed in certain gene clusters that include some virulence factors. Among these, we found that iron acquisition systems and capsule synthesis genes were the main differential features between both genomes that could explain the differences in the pathogenicity of the strains. Besides, the studied genomes presented genomic islands and toxins, and the R17 strain presented CRISPR sequences that are absent on the type strain. Taken together, this analysis provided important insights into virulence factors of V. toranzoniae that will lead to a better understanding of the pathogenic process.

Isolation of Vibrio tapetis from two native fish species (Genypterus chilensis and Paralichthys adspersus) reared in Chile and description of Vibrio tapetis subsp. quintayensis subsp. nov.

A group of seven Chilean isolates presumptively belonging to Vibrio tapetis was isolated from diseased fine flounders (Paralichthys adspersus) and red conger eel (Genypterus chilensis) experimentally reared in Quintay (Chile). All isolates were confirmed as members of V. tapetis on the basis of matrix-assisted laser desorption ionization time-of-flight MS, 16S rRNA gene sequencing, DNA-DNA hybridization values and G+C content. The ERIC-PCR and REP-PCR patterns were homogeneous among those isolates recovered from the same host (red conger or fine flounders), but distinct from the type strains V. tapetis subsp. tapetis CECT 4600T and V. tapetis subsp. britannicus CECT 8161T. On the basis of atpA, rpoA, rpoD, recA and pyrH gene sequence similarities (99.7-100 %) and clustering in the phylogenetic trees, the red conger isolates (Q20, Q047, Q48 and Q50) were confirmed as representing V. tapetis subsp. tapetis. However, they differed from V. tapetis subsp. tapetis CECT 4600T in their lipase, alpha quimiotripsin and non-acid phosphatase production. On the other hand, the fine flounder isolates (QL-9T, QL-35 and QL-41) showed rpoD, recA and pyrH gene sequence similarities ranging from 91.6 to 97.7 % with the type strains of the two V. tapetis subspecies (CECT 4600T and CECT 8161T) and consistently clustered together as an independent phylogenetic line within V. tapetis. Moreover, they could be differentiated phenotypically from strains CECT 4600T and CECT 8161T by nine and three different biochemical tests, respectively. In conclusion, the presence of V. tapetis in diseased red conger eel and fine flounder was demonstrated, extending the known host range and geographical location for this pathogen. Furthermore, this study demonstrates that the three isolates from fine flounder represent a novel subdivision within V. tapetis, for which the name V. tapetis subsp. quintayensis subsp. nov. is proposed and with QL-9T (=CECT 8851T=LMG 28759T) as the type strain. Although QL-9T was isolated from kidney of diseased fine flounder specimens, the challenge assays showed that it was non-pathogenic for this species.

Characterization of the microbiota associated to Pecten maximus gonads using 454-pyrosequencing.

A next-generation sequencing (NGS) approach was used to study the microbiota associated to Pecten maximus broodstock, applying pyrosequencing of PCR-amplified V1-V4 16S rRNA gene regions. We analysed the resident bacterial communities in female and male scallop gonads before and after spawning. DNA samples were amplified and quality-filtered reads were assigned to family and genus taxonomic levels using the Ribosomal Database Project classifier. A total of 18,520 sequences were detected, belonging to 13 phyla, including Proteobacteria (55%), Bacteroidetes (11,7%), Firmicutes (3%), Actinobacteria (2%) and Spirochaetes (1,2%), and 110 genera. The major fraction of the sequences detected corresponded to Proteobacteria, Beta- and Gammaprotebacteria being the most abundant classes. The microbiota of P. maximus gonad harbour a wide diversity, however differences on male and female samples were observed, where female gonad samples show a larger number of genera and families. The dominant bacterial genera appeared to be Delftia, Acinetobacter, Hydrotalea, Aquabacterium, Bacillus, Sediminibacterium, Sphingomonas, and Pseudomonas that were present among the four analysed samples. This next generation sequencing technique, applied for the first time in P. maximus (great scallop) gonads was useful for the study of the bacterial communities in this mollusc, unravelling the great bacterial diversity in its microbiota. [Int Microbiol 19(2): 93-99(2016)].

Marinomonas gallaica sp. nov. and Marinomonas atlantica sp. nov., isolated from reared clams (Ruditapes decussatus).

Three Gram-negative bacterial strains (Cmf 17.2T, Rd 20.33 and Cmf 18.22T) isolated from reared clams in Galicia were subjected to a taxonomic study, based on genetic and phenotypic characterization. Analysis of the 16S rRNA gene allowed the identification of the strains as members of the genus Marinomonas, sharing the highest similarity with Marinomonas aquimarina CECT 5080T (97.8 %-98.5 % 16S rRNA gene sequence similarity). Phylogenetic analysis of the sequences showed that the three isolates formed two different groups distantly related to their closest relative, M. aquimarina. DNA-DNA hybridizations were performed to confirm the taxonomic position and the results were below the recommended threshold for species delimitation, specifically 44.5 % (Cmf 17.2T with M. aquimarina CECT 5080T) and 55 % (Cmf 18.22Twith M. aquimarina CECT 5080T). Furthermore, the average nucleotide identity (ANIb, ANIm and OrthoANI) and in silico estimated DNA-DNA reassociation values among Cmf 17.2T, Cmf 18.22T and M. aquimarina CECT 5080T were in all cases below the respective threshold for species differentiation. The estimated G+C content of the genomic DNA was found to be 45.3 % (Cmf 17.2T) and 44.6 % (Cmf 18.22T). The principal fatty acids of the strains were found to be summed feature 3 (C16 : 1 ω7c/C16 : 1ω6c), summed feature 8 (C18 : 1ω7c/C18 : 1ω6c), C16 : 0, C12 : 0 and C10 : 0 3-OH. The results obtained on the characterization of the clam isolates indicate that they represent two novel species of the genus Marinomonas, for which the names Marinomonas gallaica sp. nov. (type strain Cmf 17.2T=CECT 9049T=LMG 29243T) and Marinomonas atlantica sp. nov. (type strain Cmf 18.22T=CECT 9050T=LMG 29244T) are proposed.

Draft Genome Sequence of Vibrio toranzoniae Strain CECT 7225T.

Vibrio toranzoniae(CECT 7225(T)) was isolated from healthy reared carpet shell clams in Galicia (Northwest Spain). In addition, this species has been recently identified as a potential pathogen of red conger eel in Chile. The draft genome sequence has 4.5 Mbp, a G+C content of 43.9%, and >3,800 protein-coding genes.

Isolation and identification of Vibrio toranzoniae associated with diseased red conger eel (Genypterus chilensis) farmed in Chile.

The present study deals with the first isolation of Vibrio toranzoniae from cultured red conger eel (Genypterus chilensis). During the summer season of 2011, mortalities were observed in young red conger eel at one aquaculture experimental rearing system in Quintay, Valparaiso, Chile. The microbiological analysis of the diseased fish resulted in the isolation of three dominant and representative isolates, designated as R.17, R.18 and R.19, which were obtained from gill, fin and external lesions from three different fish, respectively. All isolates were identified as V. toranzoniae by means of a polyphasic taxonomic approach, including phenotypic characterization, sequencing of 16S rRNA and housekeeping genes, and DNA-DNA hybridization. Inoculation of a representative strain (R18) in turbot as model fish species demonstrated the pathogenic potential for fish of the Chilean isolates. Results obtained indicate that the geographical and host distribution of V. toranzoniae is wider than expected, and that this species may have negative incidence in the culture of marine organisms.

Vibrio mexicanus sp. nov., isolated from a cultured oyster Crassostrea corteziensis.

A bacterial strain was taxonomically characterised by means of a genomic approach comprising 16S rRNA gene sequence analysis, multilocus sequence analysis (MLSA), the DNA G+C content, whole genome analyses (ANI and GGDC) and phenotypic characterisation. The strain CAIM 1540(T) was isolated from a cultured oyster Crassostrea corteziensis in La Cruz, Sinaloa state, México. The isolate was found to be catalase and oxidase positive, cells were observed to be motile, O/129-sensitive and facultatively anaerobic. The almost-complete 16S rRNA gene sequence placed this strain within the genus Vibrio; the closest related species were found to be Vibrio aestivus, Vibrio marisflavi, Vibrio maritimus and Vibrio variabilis with similarity values of 99.02, 97.05, 96.70, and 96.59 % respectively. MLSA of four housekeeping genes (ftsZ, gapA, recA, and topA) was performed with the closely related species. A draft genome sequence of strain CAIM 1540(T) was obtained. The DNA G+C content of this strain was determined to be 43.7 mol%.The ANI values with V. aestivus were 89.6 % (ANIb), 90.6 % (ANIm) and a GGDC value of 39.5 ± 2.5 % was obtained; with V. marisflavi the genomic similarities were 71.5 % (ANIb), 85.5 % (ANIm) and 20.2 ± 2.3 % (GGDC); with V. maritimus 72.6 % (ANIb), 85.7 % (ANIm) and 22.0 ± 2.0 % (GGDC); and with V. variabilis 72.6 % (ANIb), 85.8 % (ANIm) and 21.6 ± 1.6 % (GGDC). These ANI and GGDC values are below the threshold for the delimitation of prokaryotic species, i.e. 95-96 and 70 %, respectively. Phenotypic characters also showed differences with the closest related species analysed. The results presented here support the description of a novel species, for which the name Vibrio mexicanus sp. nov. is proposed, with strain CAIM 1540(T) (= CECT 8828(T), = DSM 100338(T)) as the type strain. In addition, we found that the recently described species Vibrio thalassae and Vibrio madracius might be a single species because the values of ANIb 95.8 %, ANIm 96.6 % and GGDC 70.2 ± 2.9 % are above the accepted species thresholds.

Description of Lacinutrix venerupis sp. nov.: a novel bacterium associated with reared clams.

A group of five Gram-negative strains isolated from clams (Venerupis decussata and Venerupis philippinarum) in Galicia (NW Spain) were analyzed using a polyphasic taxonomic approach. The strains were non-motile, strictly aerobic rods, and formed yellow-pigmented colonies. Based on 16S rRNA gene sequence similarity, the clam isolates belonged to the Lacinutrix genus, within the Flavobacteriaceae family, and showed the highest sequence similarities with Lacinutrix mariniflava (97.0%) and Lacinutrix algicola (96.9%). The DNA G+C content of the designated type strain Cmf 20.8(T) was 34.1%, which was within the range of the other species in the genus (29.0-37.0%). The main cellular fatty acids of the type strain Cmf 20.8(T) were iso-C15:0 3-OH, iso-C15:1 G, iso-C15:0 and iso-C17:0 3-OH. The DNA-DNA analysis value of strain Cmf 20.8(T) with L. mariniflava AKS292(T) was 51% and with L. algicola AKS293(T) it was 56%. Phenotypic data showed that the isolates could be differentiated easily from the other species of the genus. On the basis of these results, the strains represent a novel species of the genus Lacinutrix, for which the name Lacinutrix venerupis sp. nov. is proposed, with Cmf 20.8(T) (=CECT 8573(T)=DSM 28755(T)) as the designated type strain.

New Vibrio species associated to molluscan microbiota: a review.

The genus Vibrio consists of more than 100 species grouped in 14 clades that are widely distributed in aquatic environments such as estuarine, coastal waters, and sediments. A large number of species of this genus are associated with marine organisms like fish, molluscs and crustaceans, in commensal or pathogenic relations. In the last decade, more than 50 new species have been described in the genus Vibrio, due to the introduction of new molecular techniques in bacterial taxonomy, such as multilocus sequence analysis or fluorescent amplified fragment length polymorphism. On the other hand, the increasing number of environmental studies has contributed to improve the knowledge about the family Vibrionaceae and its phylogeny. Vibrio crassostreae, V. breoganii, V. celticus are some of the new Vibrio species described as forming part of the molluscan microbiota. Some of them have been associated with mortalities of different molluscan species, seriously affecting their culture and causing high losses in hatcheries as well as in natural beds. For other species, ecological importance has been demonstrated being highly abundant in different marine habitats and geographical regions. The present work provides an updated overview of the recently characterized Vibrio species isolated from molluscs. In addition, their pathogenic potential and/or environmental importance is discussed.

Vibrio cortegadensis sp. nov., isolated from clams.

A group of four strains isolated from clams (Venerupis decussata and Venerupis philippinarum) in Galicia (NW Spain) were subjected to a polyphasic characterization, based on the phenotypic characteristics, the analysis of chemotaxonomic features, the sequencing of the 16S rRNA and five housekeeping (atpA, pyrH, recA, rpoA and rpoD) genes, as well as DNA-DNA hybridization (DDH). The analysis of the phenotypic and chemotaxonomic characteristics and the results of a phylogenetic study, based on the 16S rRNA gene sequence analysis and multilocus sequence analysis, clearly indicated that these strains belong to the genus Vibrio and were allocated between the Splendidus and Anguillarum clades showing a close relationship with the type strains of Vibrio tapetis (98.8 %), Vibrio pomeroyi (98.0 %) and Vibrio crassostreae (97.9 %). DNA-DNA hybridization results confirmed that these isolates constitute a new species. The name Vibrio cortegadensis sp. nov. is proposed with C 16.17(T) (=CECT 7227(T)=LMG 27474(T)) as the type strain.

Vibrio toranzoniae sp. nov., a new member of the Splendidus clade in the genus Vibrio.

Four motile facultative anaerobic marine isolates (Vb 10.8(T) [CECT 7225(T), CAIM 1869(T)], CMJ 9.4 [CECT 8091, CAIM 1870], CMJ 9.11 and Cmf 13.9), were obtained from cultured clams (Venerupis philippinarum and Venerupis decussata) in Galicia (NW Spain). These isolates were studied by a polyphasic approach, including a phylogenetic analysis based on sequences of 16S rRNA and five housekeeping genes atpA, recA, pyrH, rpoA and rpoD, that supported their inclusion in the Splendidus clade of the genus Vibrio, forming a well-defined group separated from the others species of the clade. DNA-DNA hybridizations with the type strains of species showing more than 98.5% 16S rRNA gene sequence similarity rendered values of hybridization below 60%. These isolates could be differentiated from the closest relatives on the basis of several phenotypic and chemotaxonomic features. These results demonstr8ated that the strains constitute a novel specie of the genus for which the name Vibrio toranzoniae sp. nov. is proposed, with Vb 10.8(T) (=CECT 7225(T)CAIM 1869(T)) as the type strain.