Whole genome sequencing and inheritance-based variant filtering as a tool for unraveling missing heritability in pediatric cancer.

Survival rates for pediatric cancer have significantly increased the past decades, now exceeding 70-80% for most cancer types. The cause of cancer in children and adolescents remains largely unknown and a genetic susceptibility is considered in up to 10% of the cases, but most likely this is an underestimation. Families with multiple pediatric cancer patients are rare and strongly suggestive for an underlying predisposition to cancer. The absence of identifiable mutations in known cancer predisposing genes in such families could indicate undiscovered heritability. To discover candidate susceptibility variants, whole genome sequencing was performed on germline DNA of a family with two children affected by Burkitt lymphoma. Using an inheritance-based filtering approach, 18 correctly segregating coding variants were prioritized without a biased focus on specific genes or variants. Two variants in FAT4 and DCHS2 were highlighted, both involved in the Hippo signaling pathway, which controls tissue growth and stem cell activity. Similarly, a set of nine non-coding variants was prioritized, which might contribute, in differing degrees, to the increased cancer risk within this family. In conclusion, inheritance-based whole genome sequencing in selected families or cases is a valuable approach to prioritize variants and, thus, to further unravel genetic predisposition in childhood cancer.

In vivo stabilization of a less toxic asparaginase variant leads to a durable antitumor response in acute leukemia.

Asparagine is a non-essential amino acid since it can either be taken up via the diet or synthesized by asparagine synthetase. Acute lymphoblastic leukemia (ALL) cells do not express asparagine synthetase or express it only minimally, which makes them completely dependent on extracellular asparagine for their growth and survival. This dependency makes ALL cells vulnerable to treatment with L-asparaginase, an enzyme that hydrolyzes asparagine. To date, all clinically approved L-asparaginases have significant L-glutaminase co-activity, associated with non-immune related toxic side effects observed during therapy. Therefore, reduction of L-glutaminase co-activity with concomitant maintenance of its anticancer L-asparaginase effect may effectively improve the tolerability of this unique drug. Previously, we designed a new alternative variant of Erwinia chrysanthemi (ErA; Erwinaze) with decreased L-glutaminase co-activity, while maintaining its L-asparaginase activity, by the introduction of three key mutations around the active site (ErA-TM). However, Erwinaze and our ErA-TM variant have very short half-lives in vivo. Here, we show that the fusion of ErA-TM with an albumin binding domain (ABD)-tag significantly increases its in vivo persistence. In addition, we evaluated the in vivo therapeutic efficacy of ABD-ErA-TM in a B-ALL xenograft model of SUP-B15. Our results show a comparable long-lasting durable antileukemic effect between the standard-of-care pegylated-asparaginase and ABD-ErA-TM L-asparaginase, but with fewer co-glutaminase-related acute side effects. Since the toxic side effects of current L-asparaginases often result in treatment discontinuation in ALL patients, this novel ErA-TM variant with ultra-low L-glutaminase co-activity and long in vivo persistence may have great clinical potential.

Prospective, real-time monitoring of pegylated Escherichia coli and Erwinia asparaginase therapy in childhood acute lymphoblastic leukaemia and non-Hodgkin lymphoma in Belgium.

Asparaginase (ASNase) is an important anti-leukaemic drug in the treatment of childhood acute lymphoblastic leukaemia (ALL) and non-Hodgkin lymphoma (NHL). A substantial proportion of patients develop hypersensitivity reactions with anti-ASNase neutralising antibodies, resulting in allergic reactions or silent inactivation (SI), and characterised by inactivation and rapid clearance of ASNase. We report results of a prospective, real-time therapeutic drug monitoring of pegylated Escherichia coli (PEG-)ASNase and Erwinia ASNase in children treated for ALL and NHL in Belgium. Erwinia ASNase was given as second-line after hypersensitivity to PEG-ASNase. In total, 286 children were enrolled in the PEG-ASNase cohort. Allergy was seen in 11·2% and SI in 5·2% of patients. Of the 42 patients treated with Erwinia ASNase, 7·1% experienced allergy and 2·4% SI. The median trough PEG-ASNase activity was high in all patients without hypersensitivity. After Erwinia administration significantly more day 3 samples had activities <100 IU/l (62·5% vs. 10% at day 2 (D2)). The median D2 activity was significantly higher for intramuscular (IM; 347 IU/l) than for intravenous Erwinia administrations (159 IU/l). This prospective, multicentre study shows that monitoring of ASNase activity during treatment of children with ALL and NHL is feasible and informative. Treatment with Erwinia ASNase warrants close monitoring and optimally adherence to a 2-day interval of IM administrations.

Pinpointing a potential role for CLEC12B in cancer predisposition through familial exome sequencing.

Predisposition to cancer is only partly understood, and thus, the contribution of still undiscovered cancer predisposing variants necessitates further research. In search of such variants, we performed exome sequencing on the germline DNA of a family with two children affected by ganglioneuroma and neuroblastoma. Applying stringent selection criteria, we identified a potential deleterious, missense mutation in CLEC12B, coding for a lectin C-type receptor that is predicted to regulate immune function. Although further screening in a larger population and functional characterization is needed, we propose CLEC12B as a candidate cancer predisposition gene.

Effect of resveratrol on cultured skin fibroblasts from patients with oxidative phosphorylation defects.

Few therapeutic options are available to patients with oxidative phosphorylation disorders. Administering pharmacological agents that are able to stimulate mitochondrial biogenesis have been put forward as a possible treatment, yet the approach remains in need of thorough testing. We investigated the effect of resveratrol in an in vitro setting. Mitochondrial enzymatic activities were tested in cultured skin fibroblasts from patients harboring a nuclear defect in either complex II or complex IV (n = 11), and in fibroblasts from healthy controls (n = 11). In the latter, preincubation with resveratrol resulted in a significant increase of citrate synthase, complex II and complex IV enzyme activity. In patients with complex II or complex IV deficiency, however, activity of the deficient complex could not be substantially augmented, and response was dependent upon the residual activity. We conclude that resveratrol is not capable of normalizing oxidative phosphorylation activities in deficient cell lines.

Erwinia oleae sp. nov., isolated from olive knots caused by Pseudomonas savastanoi pv. savastanoi.

Three endophytic bacterial isolates were obtained in Italy from olive knots caused by Pseudomonas savastanoi pv. savastanoi. Phenotypic tests in combination with 16S rRNA gene sequence analysis indicated a phylogenetic position for these isolates in the genera Erwinia or Pantoea, and revealed two other strains with highly similar 16S rRNA gene sequences (>99 %), CECT 5262 and CECT 5264, obtained in Spain from olive knots. Rep-PCR DNA fingerprinting of the five strains from olive knots with BOX, ERIC and REP primers revealed three groups of profiles that were highly similar to each other. Multilocus sequence analysis (MLSA) based on concatenated partial atpD, gyrB, infB and rpoB gene sequences indicated that the strains constituted a single novel species in the genus Erwinia. The strains showed general phenotypic characteristics typical of the genus Erwinia and whole genome DNA-DNA hybridization data confirmed that they represented a single novel species of the genus Erwinia. The strains showed DNA G+C contents ranging from 54.7 to 54.9 mol%. They could be discriminated from phylogenetically related species of the genus Erwinia by their ability to utilize potassium gluconate, l-rhamnose and d-arabitol, but not glycerol, inositol or d-sorbitol. The name Erwinia oleae sp. nov. (type strain DAPP-PG 531(T)= LMG 25322(T) = DSM 23398(T)) is proposed for this novel taxon.

Transfer of Pantoea citrea, Pantoea punctata and Pantoea terrea to the genus Tatumella emend. as Tatumella citrea comb. nov., Tatumella punctata comb. nov. and Tatumella terrea comb. nov. and description of Tatumella morbirosei sp. nov.

Pantoea citrea, Pantoea punctata and Pantoea terrea were described for strains isolated from fruit and soil originating in Japan. These three 'Japanese' species have been shown to be phylogenetically distant from other species of the genus Pantoea. It has been observed previously that, using multilocus sequence analysis (MLSA), the 'Japanese' species consistently formed a distinct clade with an extended branch length, casting doubt on the inclusion of these species within the genus Pantoea. Furthermore, the 'Japanese' species are closely related to Tatumella ptyseos, strains of which originate from human clinical specimens. DNA-DNA hybridization and phenotypic tests confirmed the observed phylogenetic distance of P. citrea, P. punctata and P. terrea from the genus Pantoea and the affiliation of these species with Tatumella. In addition, strains causing pink disease of pineapple, identified previously as P. citrea , were shown to represent a separate species by using 16S rRNA gene sequence analysis, and MLSA and DNA-DNA hybridization data. The name Tatumella morbirosei sp. nov. with the type strain LMG 23360(T) (=BD 878(T)=NCPPB 4036(T)=CMC6(T)) is proposed to accommodate these strains. The new combinations Tatumella citrea (Kageyama et al. 1992) comb. nov. (type strain, SHS 2003(T)=ATCC 31623(T)=BD 875( T)=CCUG 30156(T)=CIP 105599(T)=DSM 13699(T)=JCM 8882(T)=LMG 22049(T)), Tatumella punctata (Kageyama et al. 1992) comb. nov. (type strain, SHS 2006(T)=ATCC 31626(T)=BD 876( T)=CCUG 30159(T)=CIP 105598(T)=DSM 13700(T)=JCM 8885(T)=LMG 22050(T)) and Tatumella terrea (Kageyama et al. 1992) comb. nov. (type strain, SHS 2008(T)=ATCC 31628(T)=BD 877(T)=CCUG 30161(T)=CIP 105600(T)=DSM 13701(T)=JCM 8887(T)=LMG 22051(T)) are proposed for P. citrea, P. punctata and P. terrea , respectively.

Validation of the (GTG)(5)-rep-PCR fingerprinting technique for rapid classification and identification of acetic acid bacteria, with a focus on isolates from Ghanaian fermented cocoa beans.

Amplification of repetitive bacterial DNA elements through the polymerase chain reaction (rep-PCR fingerprinting) using the (GTG)(5) primer, referred to as (GTG)(5)-PCR fingerprinting, was found a promising genotypic tool for rapid and reliable speciation of acetic acid bacteria (AAB). The method was evaluated with 64 AAB reference strains, including 31 type strains, and 132 isolates from Ghanaian, fermented cocoa beans, and was validated with DNA:DNA hybridization data. Most reference strains, except for example all Acetobacter indonesiensis strains and Gluconacetobacter liquefaciens LMG 1509, grouped according to their species designation, indicating the usefulness of this technique for identification to the species level. Moreover, exclusive patterns were obtained for most strains, suggesting that the technique can also be used for characterization below species level or typing of AAB strains. The (GTG)(5)-PCR fingerprinting allowed us to differentiate four major clusters among the fermented cocoa bean isolates, namely A. pasteurianus (cluster I, 100 isolates), A. syzygii- or A. lovaniensis-like (cluster II, 23 isolates), and A. tropicalis-like (clusters III and IV containing 4 and 5 isolates, respectively). A. syzygii-like and A. tropicalis-like strains from cocoa bean fermentations were reported for the first time. Validation of the method and indications for reclassifications of AAB species and existence of new Acetobacter species were obtained through 16S rRNA sequencing analyses and DNA:DNA hybridizations. Reclassifications refer to A. aceti LMG 1531, Ga. xylinus LMG 1518, and Ga. xylinus subsp. sucrofermentans LMG 18788(T).

Identification of lactobacilli by pheS and rpoA gene sequence analyses.

The aim of this study was to evaluate the use of the phenylalanyl-tRNA synthase alpha subunit (pheS) and the RNA polymerase alpha subunit (rpoA) partial gene sequences for species identification of members of the genus Lactobacillus. Two hundred and one strains representing the 98 species and 17 subspecies were examined. The pheS gene sequence analysis provided an interspecies gap, which in most cases exceeded 10 % divergence, and an intraspecies variation of up to 3 %. The rpoA gene sequences revealed a somewhat lower resolution, with an interspecies gap normally exceeding 5 % and an intraspecies variation of up to 2 %. The combined use of pheS and rpoA gene sequences offers a reliable identification system for nearly all species of the genus Lactobacillus. The pheS and rpoA gene sequences provide a powerful tool for the detection of potential novel Lactobacillus species and synonymous taxa. In conclusion, the pheS and rpoA gene sequences can be used as alternative genomic markers to 16S rRNA gene sequences and have a higher discriminatory power for reliable identification of species of the genus Lactobacillus.

Acetobacter ghanensis sp. nov., a novel acetic acid bacterium isolated from traditional heap fermentations of Ghanaian cocoa beans.

Twenty-three acetic acid bacteria, isolated from traditional heap fermentations of Ghanaian cocoa beans, were subjected to a polyphasic taxonomic study. The isolates were catalase-positive, oxidase-negative, Gram-negative rods. They oxidized ethanol to acetic acid and were unable to produce 2-ketogluconic acid, 5-ketogluconic acid and 2,5-diketogluconic acid from glucose; therefore, they were tentatively identified as Acetobacter species. 16S rRNA gene sequencing and phylogenetic analysis confirmed their position in the genus Acetobacter, with Acetobacter syzygii and Acetobacter lovaniensis as their closest phylogenetic neighbours. (GTG)(5)-PCR fingerprinting grouped the strains in a cluster that did not contain any type strains of members of the genus Acetobacter. DNA-DNA hybridization with the type strains of all recognized Acetobacter species revealed DNA-DNA relatedness values below the species level. The DNA G+C contents of three selected strains were 56.9-57.3 mol%. The novel strains had phenotypic characteristics that enabled them to be differentiated from phylogenetically related Acetobacter species, i.e. they were motile, did not produce 2-ketogluconic acid or 5-ketogluconic acid from glucose, were catalase-positive and oxidase-negative, grew on yeast extract with 30 % glucose, grew on glycerol (although weakly) but not on maltose or methanol as carbon sources, and did not grow with ammonium as sole nitrogen source and ethanol as carbon source. Based on the genotypic and phenotypic data, the isolates represent a novel species of the genus Acetobacter for which the name Acetobacter ghanensis sp. nov. is proposed. The type strain is R-29337(T) (=430A(T)=LMG 23848(T)=DSM 18895(T)).

Pediococcus stilesii sp. nov., isolated from maize grains.

A Gram-positive, coccus-shaped, lactic acid bacterium, strain LMG 23082T, was isolated from steeped maize grains. The organism is homofermentative and produces D- and L-lactic acid from glucose. 16S rRNA gene sequence analysis revealed that the organism belongs to the genus Pediococcus, with Pediococcus pentosaceus and Pediococcus acidilactici as nearest neighbours. Genotypic fingerprinting, whole-cell protein electrophoresis, DNA-DNA hybridizations and physiological and biochemical tests allowed differentiation of strain LMG 23082T from other established Pediococcus species. A remarkable feature was that, unlike other pediococci, this bacterium was capable of growth at pH 9.0. The strain studied represents a novel species for which the name Pediococcus stilesii sp. nov. is proposed with the type strain LMG 23082T (=BFE 1652T=FAIR-E 180T=CCUG 51290T), the only currently known isolate of the species.

Reclassification of Enterococcus flavescens Pompei et al. 1992 as a later synonym of Enterococcus casseliflavus (ex Vaughan et al. 1979) Collins et al. 1984 and Enterococcus saccharominimus Vancanneyt et al. 2004 as a later synonym of Enterococcus italicus Fortina et al. 2004.

The taxonomic relatedness between the species Enterococcus casseliflavus and Enterococcus flavescens and between Enterococcus italicus and Enterococcus saccharominimus was investigated. Literature data had already indicated the synonymy between E. casseliflavus and E. flavescens, but this observation had not been formally published. Additional evidence that the two taxa represent a single species was provided by comparison of the partial sequences for three housekeeping genes, phenylalanyl-tRNA synthase alpha subunit (pheS), RNA polymerase alpha subunit (rpoA) and the alpha subunit of ATP synthase (atpA). Additional genomic data derived from DNA-DNA hybridization demonstrated that the two species are synonymous. For E. italicus and E. saccharominimus, two recently described taxa, a high 16S rRNA gene sequence similarity of >99% and analogous phenotypic features indicated a close taxonomic relatedness. The same multilocus sequence analysis scheme for the three housekeeping genes was also applied for E. italicus and E. saccharominimus and indicated possible conspecificity, an observation that was also confirmed by a high DNA-DNA hybridization value (>or=78%). Data from the present study led to the proposal that E. flavescens should be reclassified as a later synonym of E. casseliflavus and that E. saccharominimus should be reclassified as a later synonym of E. italicus.

Larkinella insperata gen. nov., sp. nov., a bacterium of the phylum 'Bacteroidetes' isolated from water of a steam generator.

A Gram-negative bacterium, designated strain LMG 22510T, was isolated from water of a pharmaceutical company steam generator. The cells had a ring-like and horseshoe-shaped morphology and possessed gliding motility. Phylogenetic analysis of the 16S rRNA gene sequence showed that the strain was a member of the Flexibacter group within the phylum 'Bacteroidetes'; its nearest neighbour was Spirosoma linguale (88.8 % sequence similarity). DNA base content, fatty acid composition and biochemical characteristics were determined. Genotypic and phenotypic data indicated that strain LMG 22510T could not be assigned to any recognized genus; therefore, a novel genus and species is proposed, Larkinella insperata gen. nov., sp. nov., with LMG 22510T (= NCIMB 14103T) as the type strain.

Surface microflora of four smear-ripened cheeses.

The microbial composition of smear-ripened cheeses is not very clear. A total of 194 bacterial isolates and 187 yeast isolates from the surfaces of four Irish farmhouse smear-ripened cheeses were identified at the midpoint of ripening using pulsed-field gel electrophoresis (PFGE), repetitive sequence-based PCR, and 16S rRNA gene sequencing for identifying and typing the bacteria and Fourier transform infrared spectroscopy and mitochondrial DNA restriction fragment length polymorphism (mtDNA RFLP) analysis for identifying and typing the yeast. The yeast microflora was very uniform, and Debaryomyces hansenii was the dominant species in the four cheeses. Yarrowia lipolytica was also isolated in low numbers from one cheese. The bacteria were highly diverse, and 14 different species, Corynebacterium casei, Corynebacterium variabile, Arthrobacter arilaitensis, Arthrobacter sp., Microbacterium gubbeenense, Agrococcus sp. nov., Brevibacterium linens, Staphylococcus epidermidis, Staphylococcus equorum, Staphylococcus saprophyticus, Micrococcus luteus, Halomonas venusta, Vibrio sp., and Bacillus sp., were identified on the four cheeses. Each cheese had a more or less unique microflora with four to nine species on its surface. However, two bacteria, C. casei and A. arilaitensis, were found on each cheese. Diversity at the strain level was also observed, based on the different PFGE patterns and mtDNA RFLP profiles of the dominant bacterial and yeast species. None of the ripening cultures deliberately inoculated onto the surface were reisolated from the cheeses. This study confirms the importance of the adventitious, resident microflora in the ripening of smear cheeses.

Reclassification of Lactobacillus ferintoshensis as a later heterotypic synonym of Lactobacillus parabuchneri.

Lactobacillus ferintoshensis has recently been described as a novel species, distinct from its close phylogenetic neighbours Lactobacillus buchneri, Lactobacillus kefiri and Lactobacillus hilgardii. Two highly related species with validly published names, Lactobacillus parakefiri and Lactobacillus parabuchneri, were not considered in the study due to the lack of 16S rRNA gene sequence data at that time. Since the publication of the study, the sequences have become available and have revealed that L. ferintoshensis and L. parabuchneri share 99.7% 16S rRNA gene sequence similarity. Further genomic and phenotypic data, derived from fluorescent amplified fragment length polymorphism, DNA-DNA hybridization and API 50 CHL analyses, have demonstrated that the species are synonymous.

Grouping of streptomycetes using 16S-ITS RFLP fingerprinting.

A total of 463 Streptomyces and Kitasatospora type strains were screened using 16S-ITS RFLP fingerprinting (combined restriction digest using enzymes BstUI and HaeIII). In total, 59 clusters could be delineated, each comprising multiple strains with nearly identical patterns. Good correlation was found in general with phylogeny, as revealed by 16S rDNA sequencing. Most strains assigned to a particular 16S-ITS RFLP cluster were classified into the corresponding 16S sequencing cluster whether a 16S similarity cut-off value of 97 or 98% was used. We conclude that the taxonomic resolution of 16S-ITS RFLP fingerprinting is higher than that of 16S rDNA sequencing; this may provide a tool for reducing the number of laborious DNA-DNA hybridizations necessary for discovering potentially new species within Streptomyces.

Reclassification of [Cytophaga] marinoflava Reichenbach 1989 as Leeuwenhoekiella marinoflava gen. nov., comb. nov. and description of Leeuwenhoekiella aequorea sp. nov.

Five heterotrophic, aerobic, halotolerant and pigmented bacterial strains with gliding motility were isolated from Antarctic sea water; one other isolate was collected from the sea urchin Strongylocentrotus intermedius in the Gulf of Peter the Great in the Sea of Japan. 16S rRNA gene sequence analysis indicated that the strains are members of the family Flavobacteriaceae, the nearest neighbour (with 97.1 % sequence similarity) being the misclassified species [Cytophaga] marinoflava. DNA-DNA hybridization experiments and chemotaxonomic and phenotypic analyses demonstrated that the six novel isolates represent a single species distinct from [C.] marinoflava. On the basis of its separate phylogenetic lineage (the nearest neighbours show 92 % sequence similarity), [C.] marinoflava is reclassified as Leeuwenhoekiella marinoflava gen. nov., comb. nov. A second species of this new genus, Leeuwenhoekiella aequorea sp. nov., is proposed for the six novel isolates, with strain LMG 22550(T) (=CCUG 50091(T)) as the type strain.

Corynebacterium mooreparkense, a later heterotypic synonym of Corynebacterium variabile.

Strains of a Gram-positive bacterium were isolated from the Irish smear-ripened cheese Gubbeen, and assigned to a new species, Corynebacterium mooreparkense, in 2001. During a further study on the same cheese, no additional isolates from this species could be found. Instead, multiple isolates of its nearest phylogenetic neighbour, Corynebacterium variabile, were found. A first screening with rep-PCR and SDS-PAGE pointed to a similarity between C. mooreparkense and C. variabile. Following this peculiar result, attempts were made to collect all type strains deposited at different culture collections and all strains described by Brennan et al. [Int J Syst Evol Microbiol (2001) 51, 843-852]. Subsequently, 16S rRNA gene sequencing and DNA-DNA hybridizations were performed. All C. mooreparkense strains had a 16S rRNA gene sequence similarity of at least 99.5 % with C. variabile and the DNA-DNA relatedness was 95 %. On the basis of these results, it is concluded that C. mooreparkense is a later heterotypic synonym of C. variabile.

Description of Algoriphagus aquimarinus sp. nov., Algoriphagus chordae sp. nov. and Algoriphagus winogradskyi sp. nov., from sea water and algae, transfer of Hongiella halophila Yi and Chun 2004 to the genus Algoriphagus as Algoriphagus halophilus comb. nov. and emended descriptions of the genera Algoriphagus Bowman et al. 2003 and Hongiella Yi and Chun 2004.

Four marine heterotrophic, aerobic, pink-pigmented and non-motile bacterial strains were isolated from sea water and algae collected in the Sea of Japan. In a polyphasic taxonomic study, 16S rRNA gene sequence analysis indicated that the strains were phylogenetically highly related to Algoriphagus ratkowskyi LMG 21435T, a member of the phylum Bacteroidetes. Further phenotypic, chemotaxonomic and genomic analyses revealed that the strains represent three novel species of the genus Algoriphagus, for which the following names are proposed: Algoriphagus aquimarinus sp. nov., Algoriphagus chordae sp. nov. and Algoriphagus winogradskyi sp. nov., with type strains KMM 3958T (= LMG 21971T = CCUG 47101T), KMM 3957T (= LMG 21970T = CCUG 47095T) and KMM 3956T (= LMG 21969T = CCUG 47094T), respectively. The species Hongiella halophila Yi and Chun 2004 is transferred to the genus Algoriphagus as Algoriphagus halophilus comb. nov. because of its close phylogenetic relatedness to Algoriphagus species and analogous phenotypic and chemotaxonomic properties. The above-mentioned novel species descriptions and species reclassification justify emended descriptions of the genera Algoriphagus and Hongiella.

Reclassification of Vibrio hollisae as Grimontia hollisae gen. nov., comb. nov.

The taxonomic positions of three representative strains of Vibrio hollisae (LMG 17719(T), LMG 21416 and LMG 21538) were investigated by means of 16S rDNA sequences and phenotypic data. V. hollisae strains (GenBank/EMBL accession nos AJ514909-AJ514911) shared 99.5 % 16S rDNA sequence similarity, but had only 94.6 % similarity to their closest phylogenetic neighbour, Enterovibrio norvegicus. 16S rDNA sequence similarity of V. hollisae and Vibrio cholerae was only 91 %. These results suggest that V. hollisae should be placed into a novel genus, for which the name Grimontia gen. nov. is proposed.