Division of labor in honey bee gut microbiota for plant polysaccharide digestion.

Bees acquire carbohydrates from nectar and lipids; and amino acids from pollen, which also contains polysaccharides including cellulose, hemicellulose, and pectin. These potential energy sources could be degraded and fermented through microbial enzymatic activity, resulting in short chain fatty acids available to hosts. However, the contributions of individual microbiota members to polysaccharide digestion have remained unclear. Through analysis of bacterial isolate genomes and a metagenome of the honey bee gut microbiota, we identify that Bifidobacterium and Gilliamella are the principal degraders of hemicellulose and pectin. Both Bifidobacterium and Gilliamella show extensive strain-level diversity in gene repertoires linked to polysaccharide digestion. Strains from honey bees possess more such genes than strains from bumble bees. In Bifidobacterium, genes encoding carbohydrate-active enzymes are colocated within loci devoted to polysaccharide utilization, as in Bacteroides from the human gut. Carbohydrate-active enzyme-encoding gene expressions are up-regulated in response to particular hemicelluloses both in vitro and in vivo. Metabolomic analyses document that bees experimentally colonized by different strains generate distinctive gut metabolomic profiles, with enrichment for specific monosaccharides, corresponding to predictions from genomic data. The other 3 core gut species clusters (Snodgrassella and 2 Lactobacillus clusters) possess few or no genes for polysaccharide digestion. Together, these findings indicate that strain composition within individual hosts determines the metabolic capabilities and potentially affects host nutrition. Furthermore, the niche specialization revealed by our study may promote overall community stability in the gut microbiomes of bees.

A cross-species genome-wide analysis of sequences similar to those involved in DNA uptake bias in the Pastuerellaceae and Neisseriaceae families of pathogenic bacteria.

Acquiring new DNA allows the emergence of drug resistance in bacteria. Some Pasteurellaceae and Neisseriaceae species preferentially take up specific sequence tags. The study of such sequences is therefore relevant. They are over-represented in the genomes of the corresponding species. I found similar sequences to be present only in, but not in all, the genomes of the Pasteurellaceae and Neisseriaceae families. The genomic densities of these sequences are different both between species and between families. Interestingly, the family whose genomes harbor more of such sequences also shows more sequence types. A phylogenetic analysis allowed inferring the possible ancestral Neisseriacean sequence and a nucleotide-by-nucleotide analysis allowed inferring the potential ancestral Pasteurellacean sequence based on its genomic footprint. The method used for this work could be applied to other sequences, including transcription factor binding and repeated DNAs.

Emergence and genomic analysis of MDR Laribacter hongkongensis strain HLGZ1 from Guangzhou, China.

Background: Laribacter hongkongensis is a facultative anaerobic, non-fermentative, Gram-negative bacillus associated with community-acquired gastroenteritis and traveller's diarrhoea. No clinical MDR L. hongkongensis isolate has been reported yet. Methods: We performed WGS (PacBio and Illumina) on a clinical L. hongkongensis strain HLGZ1 with an MDR phenotype. Results: HLGZ1 was resistant to eight classes of commonly used antibiotics. Its complete genome was a single circular chromosome of 3 424 272 bp with a G + C content of 62.29%. In comparison with the reference strain HLHK9, HLGZ1 had a higher abundance of genes associated with DNA metabolism and recombination. Several inserts including two acquired resistance gene clusters (RC1 and RC2) were also identified. RC1 carried two resistance gene cassette arrays, aac(6')-Ib-cr-aadA2-Δqac-Δsul1-floR-tetR-tetG and arr-3-dfrA32-ereA2-Δqac-sul1, which shared significant nucleotide sequence identities with the MDR region of Salmonella Genomic Island 1 from Salmonella enterica serovar Typhimurium DT104. There was also an integron-like structure, intl1-arr3-dfrA27-Δqac-sul1-aph(3')-Ic, and a tetR-tetA operon located on RC2. MLST analysis identified HLGZ1 as ST167, a novel ST clustered with two strains previously isolated from frogs. Conclusions: This study provides insight into the genomic characteristics of MDR L. hongkongensis and highlights the possibilities of horizontal resistance gene transfer in this bacterium with other pathogens.

Antibiotics reduce genetic diversity of core species in the honeybee gut microbiome.

The gut microbiome plays a key role in animal health, and perturbing it can have detrimental effects. One major source of perturbation to microbiomes, in humans and human-associated animals, is exposure to antibiotics. Most studies of how antibiotics affect the microbiome have used amplicon sequencing of highly conserved 16S rRNA sequences, as in a recent study showing that antibiotic treatment severely alters the species-level composition of the honeybee gut microbiome. But because the standard 16S rRNA-based methods cannot resolve closely related strains, strain-level changes could not be evaluated. To address this gap, we used amplicon sequencing of protein-coding genes to assess effects of antibiotics on fine-scale genetic diversity of the honeybee gut microbiota. We followed the population dynamics of alleles within two dominant core species of the bee gut community, Gilliamella apicola and Snodgrassella alvi, following antibiotic perturbation. Whereas we observed a large reduction in genetic diversity in G. apicola, S. alvi diversity was mostly unaffected. The reduction in G. apicola diversity accompanied an increase in the frequency of several alleles, suggesting resistance to antibiotic treatment. We find that antibiotic perturbation can cause major shifts in diversity and that the extent of these shifts can vary substantially across species. Thus, antibiotics impact not only species composition, but also allelic diversity within species, potentially affecting hosts if variants with particular functions are reduced or eliminated. Overall, we show that amplicon sequencing of protein-coding genes, without clustering into operational taxonomic units, provides an accurate picture of the fine-scale dynamics of microbial communities over time.

Detection and Characterization of Streptomycin Resistance (strA-strB) in a Honeybee Gut Symbiont (Snodgrassella alvi) and the Associated Risk of Antibiotic Resistance Transfer.

Use of antibiotics in medicine and farming contributes to increasing numbers of antibiotic-resistant bacteria in diverse environments. The ability of antibiotic resistance genes (ARG) to transfer between bacteria genera contributes to this spread. It is difficult to directly link antibiotic exposure to the spread of ARG in a natural environment where environmental settings and study populations cannot be fully controlled. We used managed honeybees in environments with contrasting streptomycin exposure (USA: high exposure, Norway: low exposure) and mapped the prevalence and spread of transferrable streptomycin resistance genes. We found a high prevalence of strA-strB genes in the USA compared to Norway with 17/90 and 1/90 positive samples, respectively (p < 0.00007). We identified strA-strB genes on a transferrable transposon Tn5393 in the honeybee gut symbiont Snodgrassella alvi. Such transfer of resistance genes increases the risk of the spread to new environments as honeybees are moved to new pollination sites.

Geographically widespread honeybee-gut symbiont subgroups show locally distinct antibiotic-resistant patterns.

How long-term antibiotic treatment affects host bacterial associations is still largely unknown. The honeybee-gut microbiota has a simple composition, so we used this gut community to investigate how long-term antibiotic treatment affects host-associated microbiota. We investigated the phylogenetic relatedness, genomic content (GC percentage, genome size, number of genes and CRISPR) and antibiotic-resistant genes (ARG) for strains from two abundant members of the honeybee core gut microbiota (Gilliamella apicola and Snodgrassella alvi). Domesticated honeybees are subjected to geographically different management policies, so we used two research apiaries, representing different antibiotic treatment regimens in their apiculture: low antibiotic usage (Norway) and high antibiotic usage (Arizona, USA). We applied whole-genome shotgun sequencing on 48 G. apicola and 22 S. alvi. We identified three predominating subgroups of G. apicola in honeybees from both Norway and Arizona. For G. apicola, genetic content substantially varied between subgroups and distance similarity calculations showed similarity discrepancy between subgroups. Functional differences between subgroups, such as pectin-degrading enzymes (G. apicola), were also identified. In addition, we identified horizontal gene transfer (HGT) of transposon (Tn10)-associated tetracycline resistance (Tet B) across the G. apicola subgroups in the Arizonan honeybees, using interspace polymorphisms in the Tet B determinant. Our results support that honeybee-gut symbiont subgroups can resist long-term antibiotic treatment and maintain functionality through acquisition of geographically distinct antibiotic-resistant genes by HGT.

Salinarchaeum chitinilyticum sp. nov., a chitin-degrading haloarchaeon isolated from commercial salt.

Two chitin-degrading halophilic archaeal strains, MC-74T and MC-23, were isolated from commercial salt samples. Cells were motile, rod-shaped and stained Gram-negative. Colonies were vermillion-pigmented. Strains MC-74T and MC-23 were able to grow with 1.5-5.1 M NaCl (optimum, 2.6-3.1 M) at pH 6.0-10.0 (optimum, pH 7.0) and at 20-50 °C (optimum, 40 °C). The orthologous 16S rRNA gene sequence similarity between the two strains was 99.8 %, and the closest phylogenetic relative was Salinarchaeum laminariae JCM 17267T with 99.3-99.5 % similarity. The level of DNA-DNA relatedness between the two strains was 93 and 94 % (reciprocally), and those between the two strains and Salinarchaeumlaminariae JCM 17267T were 35-36 % and 38-39 % (reciprocally). The polar lipids of both strains were phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester and phosphatidylglycerol sulfate. Glycolipids were not detected. Based on the phenotypic and phylogenetic analyses, the strains represent a novel species of the genus Salinarchaeum, for which the name Salinarchaeum chitinilyticum sp. nov. is proposed. The type strain is MC-74T (=JCM 19597T=KCTC 4262T), isolated from solar salt produced in France. Strain MC-23, isolated from a commercial solar salt sample produced in China, is an additional strain of the species.

Formation of indigoidine derived-pigments contributes to the adaptation of Vogesella sp. strain EB to cold aquatic iron-oxidizing environments.

We investigated previously under explored cold aquatic environments of Andean Patagonia, Argentina. Oily sheens similar to an oil spill are frequently observed at the surface of water in creeks and small ponds in these places. Chemical analysis of a water sample revealed the occurrence of high concentrations of iron and the presence of a free insoluble indigoidine-derived pigment. A blue pigment-producing bacterium (strain EB) was isolated from the water sample and identified as Vogesella sp. by molecular analysis. The isolate was able to produce indigoidine and another derived-pigment (here called cryoindigoidine) with strong antifreeze properties. The production of the pigments depended on the cell growth at cold temperatures (below 15 °C), as well as on the attachment of cells to solid surfaces, and iron limitation in the media. The pigments produced by strain EB showed an inhibitory effect on the growth of diverse microorganisms such as Candida albicans, Escherichia coli and Staphylococcus aureus. In addition, pigmented cells were more tolerant to freezing than non-pigmented cells, suggesting a role of cryoindigoidine/indigoidine as a cold-protectant molecule. The possible roles of the pigments in strain EB physiology and its interactions with the iron-rich environment from which the isolate was obtained are discussed. Results of this study suggested an active role of strain EB in the investigated iron-oxidizing ecosystem.

Strain diversity and host specificity in a specialized gut symbiont of honeybees and bumblebees.

Host-restricted lineages of gut bacteria often include many closely related strains, but this fine-scale diversity is rarely investigated. The specialized gut symbiont Snodgrassella alvi has codiversified with honeybees (Apis mellifera) and bumblebees (Bombus) for millions of years. Snodgrassella alvi strains are nearly identical for 16S rRNA gene sequences but have distinct gene repertoires potentially affecting host biology and community interactions. We examined S. alvi strain diversity within and between hosts using deep sequencing both of a single-copy coding gene (minD) and of the V4 region of the 16S rRNA gene. We sampled workers from domestic and feral A. mellifera colonies and wild-caught Bombus representing 14 species. Conventional analyses of community profiles, based on the V4 region of the 16S rRNA gene, failed to expose most strain variation. In contrast, the minD analysis revealed extensive strain variation within and between host species and individuals. Snodgrassella alvi strain diversity is significantly higher in A. mellifera than in Bombus, supporting the hypothesis that colony founding by swarms of workers enables retention of more diversity than colony founding by a single queen. Most Bombus individuals (72%) are dominated by a single S. alvi strain, whereas most A. mellifera (86%) possess multiple strains. No S. alvi strains are shared between A. mellifera and Bombus, indicating some host specificity. Among Bombus-restricted strains, some are restricted to a single host species or subgenus, while others occur in multiple subgenera. Findings demonstrate that strains diversify both within and between host species and can be highly specific or relatively generalized in their host associations.

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).

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).

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).

Cloning, Expression and 3D Structure Prediction of Chitinase from Chitinolyticbacter meiyuanensis SYBC-H1.

Two CHI genes from Chitinolyticbacter meiyuanensis SYBC-H1 encoding chitinases were identified and their protein 3D structures were predicted. According to the amino acid sequence alignment, CHI1 gene encoding 166 aa had a structural domain similar to the GH18 type II chitinase, and CHI2 gene encoding 383 aa had the same catalytic domain as the glycoside hydrolase family 19 chitinase. In this study, CHI2 chitinase were expressed in Escherichia coli BL21 cells, and this protein was purified by ammonium sulfate precipitation, DEAE-cellulose, and Sephadex G-100 chromatography. Optimal activity of CHI2 chitinase occurred at a temperature of 40 °C and a pH of 6.5. The presence of metal ions Fe(3+), Fe(2+), and Zn(2+) inhibited CHI2 chitinase activity, while Na⁺ and K⁺ promoted its activity. Furthermore, the presence of EGTA, EDTA, and ß-mercaptoethanol significantly increased the stability of CHI2 chitinase. The CHI2 chitinase was active with p-NP-GlcNAc, with the Km and Vm values of 23.0 µmol/L and 9.1 mM/min at a temperature of 37 °C, respectively. Additionally, the CHI2 chitinase was characterized as an N-acetyl glucosaminidase based on the hydrolysate from chitin. Overall, our results demonstrated CHI2 chitinase with remarkable biochemical properties is suitable for bioconversion of chitin waste.

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)).

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).

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)).

The adaptation of bumblebees to extremely high elevation associated with their gut microbiota.

Bumblebees are among the most abundant and important pollinators for sub-alpine and alpine flowering plant species in the Northern Hemisphere, but little is known about their adaptations to high elevations. In this article, we focused on two bumblebee species, Bombus friseanus and Bombus prshewalskyi, and their respective gut microbiota. The two species, distributed through the Hengduan Mountains of southwestern China, show species replacement at different elevations. We performed genome sequencing based on 20 worker bee samples of each species. Applying evolutionary population genetics and metagenomic approaches, we detected genes under selection and analyzed functional pathways between bumblebees and their gut microbes. We found clear genetic differentiation between the two host species and significant differences in their microbiota. Species replacement occurred in both hosts and their bacteria (Snodgrassella) with an increase in elevation. These extremely high-elevation bumblebees show evidence of positive selection related to diverse biological processes. Positively selected genes involved in host immune systems probably contributed to gut microbiota changes, while the butyrate generated by gut microbiota may influence both host energy metabolism and immune systems. This suggests a close association between the genomes of the host species and their microbiomes based on some degree of natural selection.IMPORTANCETwo closely related and dominant bumblebee species, distributed at different elevations through the Hengduan Mountains of southwestern China, showed a clear genomic signature of adaptation to elevation at the molecular level and significant differences in their respective microbiota. Species replacement occurred in both hosts and their bacteria (Snodgrassella) with an increase in elevation. Bumblebees' adaptations to higher elevations are closely associated with their gut microbiota through two biological processes: energy metabolism and immune response. Information allowing us to understand the adaptive mechanisms of species to extreme conditions is implicit if we are to conserve them as their environments change.

Pseudogulbenkiania gefcensis sp. nov., isolated from soil.

A novel strain, yH16, was isolated on nutrient agar from soil samples collected at KyungHee University, Suwon City, Republic of Korea. Cells of strain yH16(T) were short rods, Gram-negative-staining, motile and non-spore-forming, with a polar flagellum. Biochemical and molecular characterization revealed that this strain was most similar to Pseudogulbenkiania subflava BP-5(T). Further 16S rRNA gene sequencing studies revealed that the new strain clustered with Pseudogulbenkiania subflava BP-5(T) (95.9 % similarity), Paludibacterium yongneupense 5YN8-15(T) (95.2 % similarity), Gulbenkiania mobilis E4FC31-5(T) (94.6 % similarity) and Chromobacterium aquaticum CC-SE-YA-1(T) (93.9 % similarity). The isolate was able to grow at 25-40 °C, 0.3-2 % NaCl and pH 5.5-7. The DNA G+C content was 65.9 ± 1.0 mol%. The predominant fatty acids were summed feature 3 (C(16 : 1)ω7c and/or iso-C(15 : 0) 2-OH) and C(16:0). Ubiquinone 8 was the major respiratory quinone. It was evident from the data obtained that the strain should be classified as a novel species of the genus Pseudogulbenkiania. The name proposed for this taxon is Pseudogulbenkiania gefcensis sp. nov., and the type strain is yH16(T) (=KCCM 90100(T) = JCM 17850(T)).