Paralysiella testudinis gen. nov., sp. nov., isolated from the cloaca of a toad-headed turtle (Mesoclemmys nasuta).

A bacterial strain designated 26BT, which had been isolated from the cloaca of a toad-headed turtle, was subjected to a comprehensive taxonomic study. Comparison of 16S rRNA gene sequences demonstrated that strain 26BT is a member of the family Neisseriaceae. Based on highest similarity values, Neisseria animaloris DSM 21642T (95.15 %), Alysiella filiformis ATCC 15532T (95.06 %), Uruburuella testudinis 07_OD624T (94.71 %), Uruburuella suis CCUG 47806T (94.66 %) and Alysiella crassa DSM 2578T (94.64 %) were identified as the closest relatives. Average nucleotide identity values based on the blast algorithm (ANIb) indicated that U. suis (76.10/76.17 %), Neisseria shayeganii 871T (74.34/74.51 %), Stenoxybacter acetivorans (73.30/73.41 %), N. animaloris (72.98/72.80) %, A. filiformis (71.14/71.21 %) and A. crassa (70.53/71.15 %) are the next closest relatives. Like ANIb, genome-based phylogeny did not suggest the affiliation of strain 26BT with any established genus. The polyamine pattern consisted of the major compounds putrescine, 1,3-diaminopropane and spermidine and the major quinone was ubiquinone Q-8. In the polar lipid profile, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and an ornithine lipid were predominant. The fatty acid profile contained predominantly C16 : 1 ω7c, C12 : 0, C14 : 0, C16 : 0 and C12 : 0 3OH. The size of the genome was 2.91 Mbp and the genomic G+C content was 54.0 mol%. Since these data do not demonstrate an unambiguous association with any established genus, we here propose the novel genus Paralysiella with the type species Paralysiella testudinis gen. nov., sp. nov. The type strain is 26BT (=CCM 9137T=LMG 32212T).

The status of the genus Prolinoborus (Pot et al. 1992) and the species Prolinoborus fasciculus (Pot et al. 1992).

On the basis of two other publications (Yarza et al. 2013; Nemec et al. 2019) and on the basis of resequencing of the 16S rRNA gene of Prolinoborus fasciculus CIP 103579T it is concluded that Prolinoborus fasciculus CIP 103579T, which is the only available strain of the species from culture collections, does not conform to the original description given by Pot et al. (1992). The strain investigated is a member of the genus Acinetobacter within the Moraxellaceae, a family of the Gammaproteobacteria and not a member of the Betaproteobacteria as originally proposed. Prolinoborus fasciculus CIP 103579T shared 99.8 % 16S rRNA gene sequence similarity with Acinetobacter lwoffii DSM 2403T. The two strains clustered together by rpoB- and core genome-based phylogenetic analyses and shared an average nucleotide identity of 96.47% (reciprocal, 96.56 %) and a digital genome distance calculation (GGDC) value of 66.9 %. Furthermore, the two strains shared matrix-assisted laser desorption/ionization time of flight MS profiles to a high extent and showed highly similar cellular fatty acid profiles and physiological substrate utilization patterns. It is proposed that the Judicial commission consider (1) that the strain currently deposited as CIP 103579 be recognized as a member of Acinetobacter lwoffii; (2) placing Prolinoborus fasciculus (Pot et al. 1992) on the list of rejected names if a suitable replacement strain, or a neotype strain cannot be found within 2 years of publication of this request; and (3) place the genus name Prolinoborus (Pot et al. 1992) on the list of rejected names [Recommendation 20D (3) of the Code].

Identification of integrative and conjugative elements in pathogenic and commensal Neisseriaceae species via genomic distributions of DNA uptake sequence dialects.

Mobile genetic elements (MGEs) are key factors responsible for dissemination of virulence determinants and antimicrobial-resistance genes amongst pathogenic bacteria. Conjugative MGEs are notable for their high gene loads donated per transfer event, broad host ranges and phylogenetic ubiquity amongst prokaryotes, with the subclass of chromosomally inserted integrative and conjugative elements (ICEs) being particularly abundant. The focus on a small number of model systems has biased the study of ICEs towards those conferring readily selectable phenotypes to host cells, whereas the identification and characterization of integrated cryptic elements remains challenging. Even though antimicrobial resistance and horizontally acquired virulence genes are major factors aggravating neisserial infection, conjugative MGEs of Neisseria gonorrhoeae and Neisseria meningitidis remain poorly characterized. Using a phenotype-independent approach based on atypical distributions of DNA uptake sequences (DUSs) in MGEs relative to the chromosomal background, we have identified two groups of chromosomally integrated conjugative elements in Neisseria: one found almost exclusively in pathogenic species possibly deriving from the genus Kingella, the other belonging to a group of Neisseria mucosa-like commensals. The former element appears to enable transfer of traditionally gonococcal-specific loci such as the virulence-associated toxin-antitoxin system fitAB to N. meningitidis chromosomes, whilst the circular form of the latter possesses a unique attachment site (attP) sequence seemingly adapted to exploit DUS motifs as chromosomal integration sites. In addition to validating the use of DUS distributions in Neisseriaceae MGE identification, the >170 identified ICE sequences provide a valuable resource for future studies of ICE evolution and host adaptation.

Crenobacter cavernae sp. nov., isolated from a karst cave, and emended description of the genus Crenobacter.

A Gram-stain-negative, rod-shaped, motile and strictly aerobic novel bacterial isolate, designated strain K1W11S-77T, was obtained from a water sample that was collected from a karst cave in Guizhou province, PR China. The results of a phylogenetic analysis based on 16S rRNA gene sequences indicated that K1W11S-77T represented a member of the genus Crenobacter within the family Neisseriaceae of the phylum Proteobacteria. K1W11S-77T was phylogenetically closely related to Crenobacter luteusYIM 78141T (Their 16S rRNA gene sequence similarity is 95.02 %). Growth of K1W11S-77T occurred at 10-30 °C, at pH 7-9, and in the presence of 0-1 % (w/v) NaCl. The major cellular fatty acids were C12 : 0, C16 : 0, C18:1ω7c and summed feature 3. The major isoprenoid quinone was Q-8. The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol and one unidentified phospholipid. The genome of K1W11S-77T was 3.27 Mb long and encoded 3167 annotated genes. The DNA G+C content of the genomic DNA was 65.3 mol%. On the basis of phylogenetic, phenotypic and chemotaxonomic characteristics, K1W11S-77T is considered to represent a novel species of the genus Crenobacter, for which the name Crenobactercavernae sp. nov. is proposed. The type strain is K1W11S-77T (=CGMCC 1.13527T=NBRC 113452T).

Identification of active denitrifiers by DNA-stable isotope probing and amplicon sequencing reveals Betaproteobacteria as responsible for attenuation of nitrate contamination in a low impacted aquifer.

Groundwater reservoirs constitute important freshwater resources. However, these ecosystems are highly vulnerable to contamination and have to rely on the resident microbiota to attenuate the impact of this contamination. Nitrate is one of the main contaminants found in groundwater, and denitrification is the main process that removes the compound. In this study, the response to nutrient load on indigenous microbial communities in groundwater from a low impacted aquifer in Uruguay was evaluated. Denitrification rates were measured in groundwater samples from three different sites with nitrate, acetate and pyrite amendments. Results showed that denitrification is feasible under in situ nitrate and electron donor concentrations, although the lack of readily available organic energy source would limit the attenuation of higher nitrate concentrations. DNA-stable isotope probing, combined with amplicon sequencing of 16S rRNA, nirS and nirK genes, was used to identify the active denitrifiers. Members of the phylum Betaproteobacteria were the dominant denitrifiers in two of three sites, with different families being observed; members of the genus Vogesella (Neisseriaceae) were key denitrifiers at one site, while the genera Dechloromonas (Rhodocyclaceae) and Comamonas (Comamonadaceae) were the main denitrifiers detected at the other sites.

Diversification of Type VI Secretion System Toxins Reveals Ancient Antagonism among Bee Gut Microbes.

Microbial communities are shaped by interactions among their constituent members. Some Gram-negative bacteria employ type VI secretion systems (T6SSs) to inject protein toxins into neighboring cells. These interactions have been theorized to affect the composition of host-associated microbiomes, but the role of T6SSs in the evolution of gut communities is not well understood. We report the discovery of two T6SSs and numerous T6SS-associated Rhs toxins within the gut bacteria of honey bees and bumble bees. We sequenced the genomes of 28 strains of Snodgrassella alvi, a characteristic bee gut microbe, and found tremendous variability in their Rhs toxin complements: altogether, these strains appear to encode hundreds of unique toxins. Some toxins are shared with Gilliamella apicola, a coresident gut symbiont, implicating horizontal gene transfer as a source of toxin diversity in the bee gut. We use data from a transposon mutagenesis screen to identify toxins with antibacterial function in the bee gut and validate the function and specificity of a subset of these toxin and immunity genes in Escherichia coli Using transcriptome sequencing, we demonstrate that S. alvi T6SSs and associated toxins are upregulated in the gut environment. We find that S. alvi Rhs loci have a conserved architecture, consistent with the C-terminal displacement model of toxin diversification, with Rhs toxins, toxin fragments, and cognate immunity genes that are expressed and confer strong fitness effects in vivo Our findings of T6SS activity and Rhs toxin diversity suggest that T6SS-mediated competition may be an important driver of coevolution within the bee gut microbiota.IMPORTANCE The structure and composition of host-associated bacterial communities are of broad interest, because these communities affect host health. Bees have a simple, conserved gut microbiota, which provides an opportunity to explore interactions between species that have coevolved within their host over millions of years. This study examined the role of type VI secretion systems (T6SSs)-protein complexes used to deliver toxic proteins into bacterial competitors-within the bee gut microbiota. We identified two T6SSs and diverse T6SS-associated toxins in bacterial strains from bees. Expression of these genes is increased in bacteria in the bee gut, and toxin and immunity genes demonstrate antibacterial and protective functions, respectively, when expressed in Escherichia coli Our results suggest that coevolution among bacterial species in the bee gut has favored toxin diversification and maintenance of T6SS machinery, and demonstrate the importance of antagonistic interactions within host-associated microbial communities.

Multi-locus sequence typing of Laribacter hongkongensis isolates from freshwater animals, environment and diarrhea patients in southern China.

Laribacter hongkongensis is a novel emerging bacterium associated with gastroenteritis and invasive bacteremic infections. Freshwater fish and edible frogs have been identified as major reservoirs of L. hongkongensis. Currently one of the main challenges in L. hongkongensis research is to identify their sources and possible transmission routes. The aim of this study was to determine the genetic diversity and relatedness of these L. hongkongensis isolates to their hosts in the hope of shedding light on these issues. In this study, multi-locus sequence typing (MLST) was used to determine the genetic characteristics of 114 L. hongkongensis strains, including 113 isolated from humans, fish, frogs, Amazonian snails and water sample in Guangzhou and Jiangmen, Southern China, and one reference isolate HZ242, recovered from a diarrhea patient in Hangzhou. The relationships among the STs and the relatedness among the isolates were assessed by phylogenetic and eBURST analysis. A total of 72 different sequence types (STs) from 114 isolates of L. hongkongensis were identified by MLST analysis, and ST99-ST161were novel. Significant difference of the prevalence of different STs between fish isolates (41.8%, 23/55) and frog isolates (82.4%, 42/51) was revealed (p=0.000). The most frequent ST (ST45) was identified 28 times and only found in fish isolates. In addition, 10 groups were identified by eBURST in this study. Combined the MLST data from Hong Kong and the present study, there were eight eBRUST lineages (group A-H) included the isolates (49.2%, 128/260) from either numerous hosts or multiple geographic origins, which contained 33.1% (53/160) of all the STs. Group A (n=57, STs=20) consisted exclusively of isolates from fish and 92.9% (39/42) of isolates in group B (n=42, STs=16) were only from fish. Group C-F (n=22, STs=14) were found to be associated with human, apart from other hosts. In this study, extensive genetic heterogeneity among the L. hongkongensis isolates from various hosts was observed. Specifically, there is higher genetic diversity of L. hongkongensis isolates of frog-origin than those of fish-origin. This study indicated some isolates exhibited a preference for specific hosts or geographic areas. ST45 was revealed to be the most frequent ST, which was only found in the fish isolates in Southern China, but might be irrelative to human infection. This MLST study further revealed that frog was likely to be another major source for human infection with L. hongkongensis apart from fish.

Microvirgula curvata sp. nov., isolated from hydrocarbon-contaminated soil, and emended description of the genus Microvirgula.

A novel Gram-stain-negative, small curved-rod-shaped, motile strain, designated L6T, was isolated from hydrocarbon-contaminated soils collected from Kuwait. Strain L6T was able to grow at 10-40 °C (optimum, 27-32 °C), pH 6.1-8.8 (optimum, 6.5-7.5) and 0-4.5 % (w/v) NaCl (optimum, 0-0.5). C18 : 1ω6c/C18 : 1ω7c, C16 : 0, C16 : 1ω6c/C16 : 1ω7c, C12 : 0 and C12 : 0 3-OH were predominant fatty acids with minor amounts of C14 : 0 and C17 : 0 cyclo. Phosphatidylglycerol and phosphatidylethanolamine were major polar lipids. The genomic G+C content was 61.2 mol%. 16S rRNA gene sequence comparisons indicated that strain L6T represents a member of the genus Microvirgula within the family Neisseriaceae of the class Betaproteobacteria. Strain L6T has a sequence similarity of 99.2 % with Microvirgula aerodenitrificans SGLY2T and <93.8 % with other members of the family Neisseriaceae. However, strain L6T showed only 56.5±2 % relatedness (based on DNA-DNA hybridization) with M. aerodenitrificans KACC 12055T (=SGLY2T). Distinct morphological, physiological and genotypic differences from the previously described taxa support the classification of strain L6T as a representative of a novel species in the genus Microvirgula, for which the name Microvirgula curvata sp. nov. is proposed. The type strain is L6T (=KEMB 2255-471T=JCM 31223T). An emended description of the genus Microvirgula is also proposed.

Chitinibacter fontanus sp. nov., isolated from a spring.

A bacterial strain, designated STM-7T, was isolated from a spring in Taiwan and characterized using a polyphasic taxonomy approach. Cells of strain STM-7T were Gram-staining-negative, aerobic, poly-ß-hydroxybutyrate-accumulating, motile by a single polar flagellum, rod-shaped, surrounded by a thick capsule and formed milky-white colonies. Growth occurred at 15-37 °C (optimum, 25-30 °C), at pH 6-8 (optimum, pH 6-7) and with 0-2 % NaCl (optimum, 0-1 %). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain STM-7T belonged to the genus Chitinibacter and was most closely related to Chitinibacter tainanensis S1T with a sequence similarity of 97.3 %. Strain STM-7T contained summed feature 3 (comprising C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0 as the predominant fatty acids. The major hydroxyl fatty acids were C12 : 0 3-OH and C16 : 0 3-OH. The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an uncharacterized aminophospholipid, an uncharacterized glycolipid and an uncharacterized phospholipid. The major isoprenoid quinone was Q-8. The DNA G+C content of the genomic DNA was 52.4 mol%. The DNA-DNA hybridization value for strain STM-7T with Chitinibacter tainanensis BCRC 17254T was less than 47 %. On the basis of the phylogenetic inference and phenotypic data, strain STM-7T should be classified as a representative of a novel species, for which the name Chitinibacter fontanus sp. nov. is proposed. The type strain is STM-7T (=BCRC 80923T=LMG 29289T=KCTC 42982T).

Paludibacterium purpuratum sp. nov., isolated from wetland soil.

A novel bacterium, designated KJ031T, was isolated from a wetland soil sample taken from Jeju island, Republic of Korea. Cells were Gram-stain-negative, curved rod shaped, oxidase- and catalase- positive, motile and facultatively anaerobic. Growth was observed at pH 6.0-8.0 and at 20-37 °C on R2A agar. Comparative analysis of 16S rRNA gene sequences revealed that strain KJ031T is a member of the genus Paludibacterium, sharing highest sequence similarities with Paludibacterium paludis KBP-21T (96.2 %) and Paludibacterium. yongneupense 5YN8-15T (96.0 %). The major fatty acids were summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C16 : 0 and summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c). The predominant respiratory quinone was Q-8. The major polar lipids of strain KJ031T were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, one unidentified aminophospholipid, two unidentified phospholipids and one unidentified polar lipid. The DNA G+C content was 59.2 mol%. On the basis of the evidence presented in this study, strain KJ031T represents a novel species of the genus Paludibacterium, for which the name Paludibacterium purpuratum sp. nov. is proposed. The type strain is KJ031T (=KCTC 42852T =CECT 8976T).

Vogesella facilis sp. nov., isolated from a freshwater river, and emended description of the genus Vogesella.

A bacterial strain, designated TTM-24T, was isolated from a freshwater river in Taiwan and characterized using a polyphasic taxonomic approach. Cells of strain TTM-24T were Gram-stain-negative, facultatively anaerobic, poly-ß-hydroxybutyrate-accumulating, motile by a single polar flagellum, rod-shaped, with rods surrounded by a thick capsule and forming white-coloured colonies. Growth occurred at 15-37 °C (optimum, 25 °C), at pH 6.0-8.0 (optimum, pH 7.0) and with 0-1 % NaCl (optimum, 0.5 %). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain TTM-24T belonged to the genus Vogesella and was most closely related to 'Vogesella amnigena' Npb-02 with sequence similarity of 97.1 %. Strain TTM-24T contained summed feature 3 (comprising C16 : 1 ω7c and/or C16 : 1 ω6c) and C16 : 0 as the major fatty acids. The major respiratory quinone was Q-8. The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an uncharacterized aminophospholipid and two uncharacterized phospholipids. The genomic DNA G+C content of strain TTM-24T was 67.4 mol%. The DNA-DNA hybridization value for strain TTM-24T with 'V. amnigena' Npb-02 was less than 45 %. On the basis of the phylogenetic inference and phenotypic data, strain TTM-24T should be classified as a novel species, for which the name Vogesella facilis sp. nov. is proposed. The type strain is TTM-24T ( = BCRC 80912T = KCTC 42742T = LMG 29003T).

Vogesella amnigena sp. nov., isolated from a freshwater river.

A bacterial strain, designated Npb-02T, was isolated from a freshwater river in Taiwan and characterized in a taxonomic study using a polyphasic approach. Cells of strain Npb-02T were Gram-stain-negative, aerobic, poly-ß-hydroxybutyrate-accumulating, rod-shaped and non-motile. Growth occurred at 15­40 °C (optimum 25­30 °C), at pH 7.0­8.0 (optimum pH 7.0) and with 0­1 % NaCl (optimum 0.5 %). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain Npb-02T belonged to the genus Vogesella and was most closely related to Vogesella perlucida DS-28T with sequence similarity of 98.3 %. Strain Npb-02T contained summed feature 3 (comprising C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0 as the major fatty acids. The major respiratory quinone was Q-8.The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an uncharacterized aminophospholipid and an uncharacterized phospholipid. The genomic DNA G+C content of strain Npb-02T was 64.1 mol%. The DNA­DNA hybridization values for strain Npb-02T with Vogesella perlucida DS-28T, Vogesella mureinivorans 389T and Vogesella lacus GR13T were less than 25 %. On the basis of phylogenetic inference and phenotypic data, strain Npb-02T represents a novel species of the genus Vogesella, for which the name Vogesella amnigena sp. nov. is proposed. The type strain is Npb-02T ( = BCRC 80887T = LMG 28419T = KCTC 42195T).

Uruburuella testudinis sp. nov., isolated from tortoise (Testudo).

A polyphasic taxonomic analysis was carried out on 11 uncommon Gram-stain-negative, non-motile, catalase- and oxidase-positive, but indole-negative, bacterial strains isolated from tortoises. Phenotypically and genetically they represented a homogeneous group of organisms most closely related to, but distinct from, Uruburuella suis. In a reconstructed 16S rRNA gene tree they clustered on a monophyletic branch next to U. suis with gene similarities between strains of 99.5-100%, and of up to 98.2% with U. suis . DNA-DNA hybridization indicated the organisms represented a novel species with only 40% DNA-DNA similarity with U. suis . Partial sequencing of rpoB resulted in two subclusters confirming the 16S rRNA gene phylogeny; both genes allowed clear separation and identification of the novel species. Furthermore, they could be unambiguously identified by matrix-assisted laser desorption ionization time-of-flight MS, where, again, they formed a highly homogeneous cluster separate from U. suis and other members of the family Neisseriaceae . The major fatty acids were C(16 : 0) and summed feature C(16 : 1)ω7c/iso-C(15 : 0) 2-OH. The DNA G+C content was 54.4 mol%. Based on phenotypic and genetic data we propose classifying these organisms as representatives of a novel species named Uruburuella testudinis sp. nov. The type strain is 07_OD624(T) ( = DSM 26510(T) = CCUG 63373(T)).

Paludibacterium paludis sp. nov., isolated from a marsh.

A bacterial strain, designated KBP-21(T), was isolated from a water sample taken from the Banping Lake Wetland Park in Taiwan and characterized in a taxonomic study using a polyphasic approach. Cells of strain KBP-21(T) were Gram-stain-negative, facultatively anaerobic, poly-ß-hydroxybutyrate-accumulating, motile rods that formed yellow colonies. Growth occurred at 15-40 °C (optimum, 30 °C), at pH 5.0-8.0 (optimum, pH 8.0) and with 0-2% NaCl (optimum, 0%). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain KBP-21(T) belonged to the genus Paludibacterium within the family Neisseriaceae of the class Betaproteobacteria and the closest related neighbour was Paludibacterium yongneupense 5YN8-15(T) with a 16S rRNA gene sequence similarity value of 96.4%. Strain KBP-21(T) contained summed feature 3 (comprising C16 : 1ω7c and/or C16 : 1ω6c), C16 : 0 and C18 : 1ω7c as the predominant fatty acids. The major respiratory quinone was Q-8. The DNA G+C content of the genomic DNA was 62.1 mol%. The polar lipid profile consisted of a mixture of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, one uncharacterized aminophospholipid and several uncharacterized phospholipids. On the basis of the genotypic, chemotaxonomic and phenotypic data, strain KBP-21(T) represents a novel species in the genus Paludibacterium, for which the name Paludibacterium paludis sp. nov. is proposed. The type strain is KBP-21(T) ( = BCRC 80514(T)  = LMG 27230(T)  = KCTC 32182(T)).

Rivicola pingtungensis gen. nov., sp. nov., a new member of the family Neisseriaceae isolated from a freshwater river.

A bacterial strain, designated Npb-03(T), was isolated from a freshwater river in Taiwan and was characterized using a polyphasic taxonomic approach. The cells were Gram-reaction-negative, straight rod-shaped, non-motile, non-spore-forming and facultatively anaerobic. Growth occurred at 10-37 °C (optimum, 30-35 °C), at pH 6.0-8.0 (optimum, pH 6.0-7.0) and with 0-1.0% NaCl (optimum, 0%). The predominant fatty acids were summed feature 3 (comprising C(16 : 1)ω7c and/or C(16 : 1)ω6c) and C(16 : 0). The major isoprenoid quinone was Q-8 and the DNA G+C content was 64.1 mol%. The polar lipid profile consisted of a mixture of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an uncharacterized aminolipid and three uncharacterized phospholipids. The major polyamines were putrescine, 2-hydroxyputrescine, cadaverine and spermidine. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain Npb-03(T) forms a distinct lineage with respect to closely related genera within the family Neisseriaceae of the class Betaproteobacteria, most closely related to the genera Aquaspirillum, Laribacter, Leeia and Microvirgula, and the levels of 16S rRNA gene sequence similarity with respect to the type species of related genera are less than 93%. On the basis of the genotypic and phenotypic data, strain Npb-03(T) represents a novel genus and species of the family Neisseriaceae, for which the name Rivicola pingtungensis gen. nov., sp. nov. is proposed. The type strain is Npb-03(T) ( = BCRC 80376(T) = LMG 26668(T) = KCTC 23712(T)).

Prevalence and characterization of quinolone resistance in Laribacter hongkongensis from grass carp and Chinese tiger frog.

Laribacter hongkongensis is a food-borne bacterium associated with community-acquired gastroenteritis and diarrhoea. Quinolone resistance was recently reported in bacterial isolates from aquatic products, but the molecular mechanisms for resistance were still unknown. In this study, a total of 157 L. hongkongensis strains were isolated from grass carps (n = 443) and Chinese tiger frogs (n = 171). Twenty-one ciprofloxacin-resistant strains were analysed for mutations in quinolone resistance-determining regions (QRDR), acquired quinolone resistance (AQR) genes and the role of efflux pumps in resistance. All QRDR mutations in gyrA (codons 85 and 89) and parC (codons 83 and 231) were found to be closely associated with ciprofloxacin resistance. The AQR gene aac(6')-Ib-cr was found in 42.9% (9/21) of the resistant strains, but qnrA, qnrB, qnrC, qnrD, qnrS and qepA were not detected. No significant change of MICs to ciprofloxacin was observed in the presence of an efflux pump inhibitor, indicating the role of efflux pump was probably absent. All 21 ciprofloxacin-resistant strains showed different electrophoretic patterns, which suggested they were not genetically related. These data highlight the importance of QRDR mutations and the AQR gene aac(6')-Ib-cr during the development of quinolone resistance in a heterogeneous population of L. hongkongensis.

Application of matrix-assisted laser desorption ionization-time of flight mass spectrometry for identification of the fastidious pediatric pathogens Aggregatibacter, Eikenella, Haemophilus, and Kingella.

The accuracy of matrix-assisted laser desorption-ionization time of flight mass spectrometry (MALDI-TOF MS) in the identification of Haemophilus, Aggregatibacter, Cardiobacterium, Eikenella, and Kingella (HACEK) species was compared to that of phenotypic methods (Remel RapID and Vitek 2). Overall, Vitek MS correctly identified more isolates, incorrectly identified fewer isolates, and failed to identify fewer isolates than both phenotypic methods.

Matrix-assisted laser desorption ionisation-time of flight mass spectrometry for rapid identification of Laribacter hongkongensis.

Laribacter hongkongensis is a Gram-negative, facultative anaerobic, motile, S-shaped, urease-positive bacillus associated with invasive infections in liver cirrhosis patients and community-acquired gastroenteritis. Most cases of L hongkongensis infections occur in eastern countries. Information is lacking on the usefulness of matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) for the identification of bacteria important in eastern countries. Using the Bruker database extended with 21 L hongkongensis reference strains, all 240 L hongkongensis isolates recovered from patients, fish, frogs and water were correctly identified, with 224 (93.3%) strains having top match scores ≥2.0. Notably, the strain of Chromobacterium violaceum was not reliably identified although it is included in the database. MALDI-TOF MS is useful for the accurate routine identification of L hongkongensis after adding reference L hongkongensis main spectra to the database. The number of strains for each species in MALDI-TOF MS databases should be expanded to cover intraspecies variability.

Phylogenomics and molecular signatures for the order Neisseriales: proposal for division of the order Neisseriales into the emended family Neisseriaceae and Chromobacteriaceae fam. nov.

The species from the order Neisseriales are currently distinguished from other bacteria on the basis of branching in 16S rRNA gene trees. For this order containing a single family, Neisseriaceae, no distinctive molecular, biochemical, or phenotypic characters are presently known. We report here detailed phylogenetic and comparative analyses on the 27 genome sequenced species of the order Neisseriales. Our comparative genomic analyses have identified 54 conserved signature indels (CSIs) in widely distributed proteins that are specific for either all of the sequenced Neisseriales species or a number of clades within this order that are also supported by phylogenetic analyses. Of these CSIs, 11 are specifically present in all of the sequenced species from this order, but are not found in homologous proteins from any other bacteria. These CSIs provide novel molecular markers specific for, and delimiting, this order. Twenty-one CSIs in diverse proteins are specific for a group comprised of the genera Neisseria, Eikenella, Kingella, and Simonsiella (Clade I), which are obligate host-associated organisms, lacking flagella and exhibiting varied morphology. The species from these genera also formed a strongly supported clade in phylogenetic trees based upon concatenated protein sequences; a monophyletic grouping of these genera and other genera displaying similar morphological characteristics was also observed in the 16S rRNA gene tree. A second clade (Clade II), supported by seven of the identified CSIs and phylogenetic trees based upon concatenated protein sequences, grouped together species from the genera Chromobacterium, Laribacter, and Pseudogulbenkiania that are rod-shaped bacteria, which display flagella-based motility and are capable of free living. The remainder of the CSIs were uniquely shared by smaller groups within these two main clades. Our analyses also provide novel insights into the evolutionary history of the Neisseriales and suggest that the CSIs that are specific for the Clade I species may play an important role in the evolution of obligate host-association within this order. On the basis of phylogenetic analysis, the identified CSIs, and conserved phenotypic characteristics of different Neisseriales genera, we propose a division of this order into two families: an emended family Neisseriaceae (corresponding to Clade I) containing the genera Alysiella, Bergeriella, Conchiformibius, Eikenella, Kingella, Neisseria, Simonsiella, Stenoxybacter, Uruburuella and Vitreoscilla and a new family, Chromobacteriaceae fam. nov., harboring the remainder of the genera from this order (viz. Andreprevotia, Aquaspirillum, Aquitalea, Chitinibacter, Chitinilyticum, Chitiniphilus, Chromobacterium, Deefgea, Formivibrio, Gulbenkiania, Iodobacter, Jeongeupia, Laribacter, Leeia, Microvirgula, Paludibacterium, Pseudogulbenkiania, Silvimonas, and Vogesella).

Iodobacter arcticus sp. nov., a psychrotolerant bacterium isolated from meltwater stream sediment of an Arctic glacier.

Two novel violet-pigmented, Gram-negative, rod-shaped, non-motile bacteria, designated strains M4-16(T) and M4-9, were isolated from sediment from an Arctic glacier. The predominant fatty acids of both strains were C16 : 1ω7c and/or C16 : 1ω6c (summed feature 3), C16 : 0, C14 : 0 and C18 : 1ω7c and/or C18 : 1ω6c (summed feature 8) and both strains contained ubiquinone-8 as the respiratory quinone. The polar lipids consisted of phosphatidylethanolamine, two unidentified phospholipids and one unidentified aminolipid. 16S rRNA gene sequence analysis indicated that strains M4-16(T) and M4-9 were members of the genus Iodobacter and closely related to Iodobacter fluviatilis ATCC 33051(T) with pairwise sequence similarity of 98.9 %. The DNA-DNA relatedness between strains M4-9 and M4-16(T) was 92.5 %, while strains M4-9 and M4-16(T) had DNA-DNA relatedness values of 21.5 and 18.2 %, respectively, with Iodobacter fluviatilis JCM 9044(T). The RAPD-PCR banding patterns of strains M4-9 and M4-16(T) were similar but differed from that of Iodobacter fluviatilis JCM 9044(T). Based on data from the current polyphasic study, strains M4-16(T) and M4-9 represent a novel species of the genus Iodobacter, for which the name Iodobacter arcticus sp. nov. is proposed. The type strain of Iodobacter arcticus is M4-16(T) ( = CIP 1103011(T) = MTCC 11351(T)).