Therapeutic Phage Candidates for Targeting Prevalent Sequence Types of Carbapenem-Resistant Escherichia coli.

Carbapenem-resistant Escherichia coli (CREC) is a global threat to public health; therefore, alternative treatment options are urgently needed. Bacteriophages have emerged as promising candidates for combating CREC infections. This study aimed to investigate the genetic basis of phage sensitivity in CREC by evaluating carbapenem resistance among multidrug-resistant (MDR) E. coli isolated in Daegu, South Korea and analyzing their sequence types (STs) with phage susceptibility spectra. Among the 60 MDR E. coli isolates, 80.4% were identified as CREC, with 77.0% demonstrating resistance to imipenem and 66.6% to meropenem. Moreover, 70 lytic E. coli bacteriophages were isolated from hospital sewage water and evaluated against those 60 E. coli isolates. The phages exhibited lytic activity of 33%-60%, with average titers ranging from 5.6 × 1012 to 2.4 × 1013 PFU/mL (Plaque-Forming Unit). Furthermore, multilocus sequence typing (MLST) analysis of the bacterial isolates revealed 14 distinct STs, mostly belonging to ST131, ST410, and ST648. Notably, the phage susceptibility spectra of ST73, ST13003, ST648, ST2311, ST167, ST405, ST607, ST7962, and ST131 were significantly different. Thus, the isolated phages can effectively lyse CREC isolates, particularly those with clinically dominant STs. Conversely, ST410 exhibited a 14.2%-87.14% susceptibility spectrum, whereas ST1139, ST1487, ST10, and ST206 did not lyse, suggesting the presence of more resistant STs. Future studies are warranted to identify the reasons behind this resistance and address it. Ultimately, this study will aid in developing focused treatments to address these pressing global health issues.

Burden of human metapneumovirus infections among children with acute respiratory tract infections attending a Tertiary Care Hospital, Kathmandu.

BACKGROUND: Acute respiratory infections (ARIs) are one of the most common causes of mortality and morbidity worldwide. Every year millions of children suffer from viral respiratory tract infections (RTIs) ranging from mild to severe illnesses. Human Metapneumovirus (HMPV) is among the most frequent viruses responsible for RTIs. However, HMPV infections and their severity among children have not been explored yet in Nepal. PURPOSE: Therefore, the study was focused on HMPV infections and other potential viral etiologies or co-infections using multiplex PCR among children attending Kanti Children's Hospital and assessed the clinical characteristics of the infections as well as found the co-infections. A hospital-based cross-sectional study was designed and a convenience sampling method was used to enroll children of less than 15 years with flu-like symptoms from both outpatients and inpatients departments over three months of the study period. RESULTS: HMPV infection (13.3%) was the most predominant infection among the different viral infections in children with ARIs in Kanti Children's Hospital. The HMPV was more prevalent in the age group less than three years (21.8%). Cough and fever were the most common clinical features present in all children infected with HMPV followed by rhinorrhea, sore throat, and wheezing. HMPV-positive children were diagnosed with pneumonia (42.9%), bronchiolitis (28.5%), upper respiratory tract infections (14.3%), and asthma (14.3%). The prevalence of HMPV was high in late winter (14.3%) followed by early spring (13.5%). CONCLUSIONS: This study provides the baseline information on HMPV and associated co-infection with other respiratory viruses for the differential diagnosis based on molecular methods and also the comparison of clinical presentations among the different respiratory syndromes.

Multipurpose cellulases of Promicromonospora sp. VP111, with broad substrate specificity and tolerance properties.

Cellulolytic actinobacterium, Promicromonospora sp. VP111 concomitantly produced cellulases (CELs), xylanase and pectinase when grown on commercial cellulose and untreated agricultural lignocellulosic residues (wheat straw and sugarcane bagasse). Secreted CELs hydrolyzed (enhanced with Co2+ ion) multiple cellulosic substrates, including sodium carboxymethyl cellulose (Na-CMC), Whatman filter paper no. 1, microcrystalline cellulose (avicel), p-nitrophenyl-ß-D-glucopyranoside (pNPG), laminarin, and cellulose powder. The CELs showed stabilities in the presence of various chemicals, including glucose (0.2 M), detergents (1%, w/v or v/v), denaturants (1%, w/v or v/v), and sodium chloride (NaCl, 30%, w/v). The CELs were fractionated using ammonium sulfate precipitation and dialysis. Activities (%) of fractionated CELs were retained at 60°C for endoglucanase/carboxymethyl cellulase (CMCase) (88.38), filter paper cellulase (FPase) (77.55), and ß-glucosidase (90.52), which indicated of thermo-stability. Similarly, the activities (%) for CMCase (85.79), FPase (82.48), and ß-glucosidase (85.92) at pH 8.5 indicated of alkaline-stability. Kinetic factors, Km and Vmax for endoglucanase component of fractionated CELs were 0.014 g/l and 158.23 µM glucose/min/mL, respectively. Fractionated CELs yielded activation energies (kJ/mol) of 17.933, 6.294, and 4.207 for CMCase, FPase, and ß-glucosidase activities, respectively in linear thermostable Arrhenius plots. Thus, this study reports on the multipurpose CELs from an untreated agricultural residue utilizing Promicromonospora in relation to broad substrate specificity, halo-tolerance, alkaline-tolerance, detergent-tolerance, thermo-tolerance, organic solvent-tolerance, and end product-tolerance.

Kaistella soli sp. nov., isolated from oil-contaminated experimental soil.

A light yellow-coloured, non-motile, aerobic, Gram-stain-negative, and rod-shaped bacterial strain DKR-2T was isolated from oil-contaminated experimental soil. The strain was catalase and oxidase positive, and grew at 0-1.5% (w/v) NaCl concentration, at temperature 10-35 °C, and at pH 6.0-9.5. The phylogenetic analysis suggested that the strain DKR-2T was affiliated to the genus Kaistella, with the closest species being Kaistella haifensis DSM 19056T (97.6% 16S rRNA gene sequence similarity). The principle fatty acids were iso-C15:0, summed feature 9 (iso-C17:1 ω9c and/or C16:0 10-methyl), and antiso-C15:0. The sole menaquinone was MK-6 and major polar lipid was phosphatidylethanolamin. The DNA G+C content was 39.5%. The dDDH (in silico DNA-DNA hybridization) and ANI (average nucleotide identity) values between strain DKR-2T and K. haifensis DSM 19056T were 22.4% and 79.3%, respectively. In addition, both dDDH and ANI values between strain DKR-2T and other phylogenetically related neighbours were < 25.0% and < 77.0%, respectively. In overall, the polyphasic taxonomic data presented in this study clearly indicated that strain DKR-2T represents a novel species in the genus Kaistella, for which the name Kaistella soli sp. nov. is proposed. The type strain is DKR-2T (=KACC 22070T=NBRC 114725T).

Cellulomonas fulva sp. nov., isolated from oil-contaminated soil.

A yellow-coloured, Gram-stain-positive, motile, aerobic and rod-shaped bacteria, designated DKR-3T, was isolated from oil-contaminated experimental soil. Strain DKR-3T could grow at pH 5.0-10.5 (optimum, pH 7.0-8.5), at 10-40 °C (optimum, 25-32 °C) and tolerated 3.5 % of NaCl. Phylogenetic analyses based on its 16S rRNA gene sequence indicated that strain DKR-3T formed a lineage within the family Cellulomonadaceae and was clustered with members of the genus Cellulomonas. Strain DKR-3T had highest 16S rRNA gene sequence similarities to Cellulomonas gelida DSM 20111T (98.3 %), Cellulomonas persica JCM 18111T (98.2 %) and Cellulomonas uda DSM 20107T (97.8 %). The predominant respiratory quinone was tetrahydrogenated menaquinone with nine isoprene units [MK-9(H4)]. The principal cellular fatty acids were anteiso-C15 : 0, C16 : 0 and anteiso-C17 : 0. The major polar lipids were diphosphatidylglycerol and phosphatidylglycerol. The cell-wall diamino acid was l-ornithine whereas rhamnose and glucose were the cell-wall sugars. The DNA G+C content was 74.2mol %. The genome of strain DKR-3T was 3.74 Mb and contained three putative biosynthetic gene clusters. The average nucleotide identity and digital DNA-DNA hybridization relatedness values between strain DKR-3T and its phylogenetically related members were below the species threshold values. Based on a polyphasic study, strain DKR-3T represents a novel species belonging to the genus Cellulomonas, for which the name Cellulomonas fulva sp. nov. is proposed. The type strain is DKR-3T (=KACC 22071T=NBRC 114730T).

Genome mining revealed polyhydroxybutyrate biosynthesis by Ramlibacter agri sp. nov., isolated from agriculture soil in Korea.

A white-colony-forming, facultative anaerobic, motile and Gram-stain-negative bacterium, designated G-1-2-2 T was isolated from soil of agriculture field near Kyonggi University, Republic of Korea. Strain G-1-2-2 T synthesized the polyhydroxybutyrate and could grow at 10-35 °C. The phylogenetic analysis based on 16S rRNA gene sequence showed that, strain G-1-2-2 T formed a lineage within the family Comamonadaceae and clustered as a member of the genus Ramlibacter. The 16S rRNA gene sequence of strain G-1-2-2 T showed high sequence similarities with Ramlibacter ginsenosidimutans BXN5-27 T (97.9%), Ramlibacter monticola G-3-2 T (97.9%) and Ramlibacter alkalitolerans CJ661T (97.5%). The sole respiratory quinone was ubiquinone-8 (Q-8). The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, and an unidentified phospholipid. The principal cellular fatty acids were C16:0, cyclo-C17:0, summed feature 3 (C16:1ω7c and/or C16:1ω6c) and summed feature 8 (C18:1ω7c and/or C18:1ω6c). The genome of strain G-1-2-2 T was 7,200,642 bp long with 13 contigs, 6,647 protein-coding genes, and DNA G + C content of 68.9%. The average nucleotide identity and in silico DNA-DNA hybridization values between strain G-1-2-2 T and close members were ≤ 81.2 and 24.1%, respectively. The genome of strain G-1-2-2 T showed eight putative biosynthetic gene clusters responsible for various secondary metabolites. Genome mining revealed the presence of atoB, atoB2, phaS, phbB, phbC, and bhbD genes in the genome which are responsible for polyhydroxybutyrate biosynthesis. Based on these data, strain G-1-2-2 T represents a novel species in the genus Ramlibacter, for which the name Ramlibacter agri sp. nov. is proposed. The type strain is G-1-2-2 T (= KACC 21616 T = NBRC 114389 T).

AMPK Amplifies IL2-STAT5 Signaling to Maintain Stability of Regulatory T Cells in Aged Mice.

AMP-activated protein kinase (AMPK), an important regulator of the aging process, is expressed in various immune cells. However, its role in regulatory T cell (Treg) stability during aging is poorly understood. Here, we addressed the role of AMPK in Treg function and stability during aging by generating Treg-specific AMPKα1 knockout mice. In this study, we found that AMPKα1-deficient Tregs failed to control inflammation as effectively as normal Tregs did during aging. AMPK knockout from Tregs reduces STAT5 phosphorylation in response to interleukin (IL)-2 stimulation, thereby destabilizing Tregs by decreasing CD25 expression. Thus, our study addressed the role of AMPK in Tregs in sensing IL-2 signaling to amplify STAT5 phosphorylation, which, in turn, supports Treg stability by maintaining CD25 expression and controlling inflamm-aging.

AMPK promotes antitumor immunity by downregulating PD-1 in regulatory T cells via the HMGCR/p38 signaling pathway.

BACKGROUND: AMP-activated protein kinase (AMPK) is a metabolic sensor that maintains energy homeostasis. AMPK functions as a tumor suppressor in different cancers; however, its role in regulating antitumor immunity, particularly the function of regulatory T cells (Tregs), is poorly defined. METHODS: AMPKα1fl/flFoxp3YFP-Cre, Foxp3YFP-Cre, Rag1-/-, and C57BL/6 J mice were used for our research. Flow cytometry and cell sorting, western blotting, immuno-precipitation, immuno-fluorescence, glycolysis assay, and qRT-PCR were used to investigate the role of AMPK in suppressing programmed cell death 1 (PD-1) expression and for mechanistic investigation. RESULTS: The deletion of the AMPKα1 subunit in Tregs accelerates tumor growth by increasing the expression of PD-1. Metabolically, loss of AMPK in Tregs promotes glycolysis and the expression of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), a key enzyme of the mevalonate pathway. Mechanistically, AMPK activates the p38 mitogen-activated protein kinase (MAPK) that phosphorylates glycogen synthase kinase-3ß (GSK-3ß), inhibiting the expression of PD-1 in Tregs. CONCLUSION: Our study identified an AMPK regulatory mechanism of PD-1 expression via the HMGCR/p38 MAPK/GSK3ß signaling pathway. We propose that the AMPK activator can display synergic antitumor effect in murine tumor models, supporting their potential clinical use when combined with anti-PD-1 antibody, anti-CTLA-4 antibody, or a HMGCR inhibitor.

Priestia veravalensis sp. nov., isolated from coastal sample.

The taxonomic position of two isolates, SGD-V-76T and SGD-M-37, isolated from sediment sample of Veraval coast, India, was examined using the polyphasic taxonomic approach. The morphological and chemotaxonomic characteristics of these two organisms are typical of the genus Priestia. The phylogenetic analyses performed using almost complete 16S rRNA gene sequences demonstrated that the isolate belongs to the Bacillaceae family, and forms a clade within the cluster containing Priestia flexus MTCC 2909T, Priestia aryabhattai B8W22T and Priestia megaterium KCTC 3007T and both strains showed highest similarity of > 98% with 3-29 nucleotide differences. The cell wall contained meso-diaminopimelic acid as the diagnostic diamino acid. The predominant isoprenoid quinone was MK-7 and the G + C content of strains was 37.5-37.7 mol%. However, the DNA-DNA hybridization and the phenotypic characteristics revealed that, the strain SGD-V-76T and strain SGD-M-37 are similar species but different from any known Priestia species with ANI values of 79.2, 79.3 and 79.2 and the dDDH values of 17.7, 17.8 and 18.0% respectively. On the basis of phenotypic characteristics, phylogenetic analysis and the results of biochemical and physiological tests, and genomic data strain SGD-V-76T was clearly distinguished from closely related members of the Priestia genus. Based on the above data analysis strain SGD-V-76T (= DSM28242T = KCTC33802T = CIP111056T = NCIM5510T) represents a novel species of the genus Priestia, and we propose the name Priestia veravalensis sp. nov.

Noviherbaspirillum pedocola sp. nov., isolated from oil-contaminated experimental soil.

An orange-coloured, rod-shaped, and aerobic bacterial strain DKR-6 T was isolated from oil-contaminated experimental soil. The strain was Gram-stain-negative, catalase and oxidase positive, and grew at temperature 10-42 °C, at pH 5.5-9.5, and at 0-3.0% (w/v) NaCl concentration. The phylogenetic analysis and 16S rRNA gene sequence analysis suggested that the strain DKR-6 T was affiliated to the genus Noviherbaspirillum, with the closest species being Noviherbaspirillum massiliense JC206T (96.3% sequence similarity). The chemotaxonomic profiles revealed the presence of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, and phosphatidylcholine as the principal polar lipids; C16:0, C17:0 cyclo, summed feature 3 (C16:1ω7c and/or C16: 1ω6c), and summed feature 8 (C18:1ω7c/or C18:1ω6c) as the main fatty acids; and Q-8 as a sole ubiquinone. The DNA G + C content was 61.6%. The polyphasic taxonomic features illustrated in this study clearly implied that strain DKR-6 T represents a novel species in the genus Noviherbaspirillum, for which the name Noviherbaspirillum pedocola sp. nov. is proposed with the type strain DKR-6 T (= KACC 22074 T = NBRC 114727 T).

Schlegelella koreensis sp. nov., isolated from evaporator core of automobile air conditioning system.

A white-coloured, aerobic, and rod-shaped bacterium, designated strain ID0723T was isolated from evaporator core of automobile air conditioning system. The strain was Gram-stain-negative, catalase positive, oxidase negative, and grew at pH 5.5-9.5, at temperature 18-37 °C, and at 0-2.0% (w/v) NaCl concentration. The phylogenetic analysis and 16S rRNA gene sequence data revealed that the strain ID0723T was affiliated to the genus Schlegelella, with the closest phylogenetic member being Schlegelella brevitalea DSM 7029 T (98.1% sequence similarity). The chemotaxonomic features of strain ID0723T were diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine as the main polar lipids; Q-8 as an only ubiquinone; and summed feature 3 (C16:1ω7c and/or C16: 1ω6c), C16:0, and summed feature 8 (C18:1ω7c/or C18:1ω6c) as the major fatty acids. The average nucleotide identity (ANI) and in silico DNA-DNA hybridization values between strain ID0723T and S. brevitalea DSM 7029 T were 74.8% and 20.0%, respectively, which were below the cut-off values of 95% and 70%, respectively. The DNA G + C content was 69.9 mol%. The polyphasic taxonomic data clearly indicated that strain ID0723T represents a novel species in the genus Schlegelella for which the name Schlegelella koreensis sp. nov. is proposed, with the type strain ID0723T (= KCTC 72731 T = NBRC 114611 T).

Chryseobacterium cheonjiense sp. nov., isolated from forest soil.

A yellow-pigmented, non-motile and rod-shaped bacterium, designated RJ-7-14T was obtained from forest soil sampled at Cheonji-dong, Seogwipo-si, Jeju-do, South Korea. Cells were Gram-stain-negative and produced flexirubin type pigments. A phylogenetic analysis based on its 16S rRNA gene sequence revealed that strain RJ-7-14T formed a lineage within the family Weeksellaceae and clustered as members of the genus Chryseobacterium. The closest members were Chryseobacterium geocarposphaerae DSM 27617T (98.2% sequence similarity), Chryseobacterium hispalense DSM 25574T (98.0%) and Chryseobacterium nepalense KACC 18907T (98.0%). The sequence similarity for other members was < 98.0%. The genome was 4,276,416 bp long with 9 scaffolds and 3779 protein-coding genes. The sole respiratory quinone was MK-6. The major cellular fatty acids were iso-C15:0, summed feature 9 (iso-C17:1 ω9c and/or C16:0 10-methyl), summed feature 3 (iso-C15:0 2-OH and/or C16: 1ω7c) and iso-C17:0 3-OH. The major polar lipid was phosphatidylethanolamine (PE). The DNA G + C content of the type strain was 37.2 mol%. In addition, the average nucleotide identity (ANIu) and in silico DNA-DNA hybridization (dDDH) relatedness values between strain RJ-7-14T and phylogenetically closest members were ≤ 88.2% and ≤ 35.0%, respectively, which were below the threshold values of 95-96% (for ANI) and 70% (for dDDH), suggesting the allocation of novel strain to a new species. Based on genomic, chemotaxonomic, phenotypic and phylogenetic analyses, strain RJ-7-14T represents novel species in the genus Chryseobacterium, for which the name Chryseobacterium cheonjiense sp. nov. is proposed. The type strain is RJ-7-14T (= KACC 21625T = NBRC 114362T).

Luteolibacter luteus sp. nov., isolated from stream bank soil.

A non-motile, Gram-stain-negative, rod-shaped and yellow-colored bacterium, designated G-1-1-1T was obtained from soil sampled at Gwanggyo stream bank, Gyeonggi-do, Republic of Korea. Cells were aerobic, catalase positive, grew optimally at 25-30 °C and hydrolysed aesculin and casein. A phylogenetic analysis based on its 16S rRNA gene sequence revealed that strain G-1-1-1T formed a lineage within the genus Luteolibacter. The closest members were Luteolibacter flavescens GKXT (97.7% sequence similarity) and Luteolibacter arcticus MC 3726T (97.3%). The sequence similarities with other members of the genus Luteolibacter were ≤ 93.9%. The genome of strain G-1-1-1T was 6,412,079 bp long with 5176 protein-coding genes. The diagnostic amino acid of cell-wall peptidoglycan of strain G-1-1-1T was meso-diaminopimelic acid. The only respiratory quinone was menaquinone-9 and the principal polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol and unidentified phospholipids. The predominant cellular fatty acids were iso-C14:0, C16:1 ω9c, C16:0, C14:0 and anteiso-C15:0. The DNA G + C content was 61.0 mol%. The anti-SMASH analysis of whole genome showed eight putative biosynthetic gene clusters responsible for various secondary metabolites. Based on genomic, chemotaxonomic, phenotypic and phylogenetic analyses, strain G-1-1-1T represents a novel species in the genus Luteobacter, for which the name Luteolibacter luteus sp. nov. is proposed. The type strain is G-1-1-1T (= KACC 21614T = NBRC 114341T).

Aquabacterium terrae sp. nov., isolated from soil.

A yellow-colored bacterial strain, designated S2T was isolated from soil in South Korea. Cells of strain S2T were strictly aerobic, Gram-stain-negative, motile with single polar flagellum, rod-shaped, oxidase and catalase-negative. Growth occurs at 10-37 °C (optimum, 28 °C), pH 5.0-9.0 (optimum, pH 6.5-7.0) and 0-3% NaCl (w/v). Strain S2T consisted of summed feature 3 (iso-C15:0 2-OH and/or C16:1 ω7c), C16:0 and summed feature 8 (C18:1 ω7c and/or C18:1 ω6c) as major fatty acids. The sole respiratory quinone was Q-8. The polar lipid profile consisted of phosphatidylethanolamine and an unidentified lipid. The 16S rRNA gene sequence analysis showed that strain S2T is phylogenetically closest to Aquabacterium pictum W35T (98.4% sequence similarity). The genome of strain S2T was 8,039,486 bp with 56 scaffolds. The genome consisted of 10 putative biosynthetic gene clusters that are responsible for various secondary metabolites. Genomic DNA G + C content of strain S2T was 69.4%. The average nucleotide identity and in silico DNA-DNA hybridization values between strain S2T and phylogenetically related taxa were ≤ 77.9 and ≤ 21.4%, and respectively. The results of genotypic and phenotypic data showed that strain S2T could be distinguished from its phylogenetically related species and represents a novel species in the genus Aquabacterium, for which the name Aquabacterium terrae sp. nov. is proposed. The type strain is S2T (= KCTC 72741 T = NBRC 114609 T).

Caenimonas soli sp. nov., isolated from soil.

A non-motile, Gram-stain-negative, rod-shaped bacterium, designated strain S4T, was obtained from soil sampled at Wonju, Gyeonggi-do, Republic of Korea. Cells were white-coloured, aerobic, grew optimally at 25-32 °C on R2A agar plate. A phylogenetic analysis based on its 16S rRNA gene sequence revealed that strain S4T formed a lineage within the family Comamonadaceae. The closest members were Caenimonas terrae SGM1-15T (98.1% sequence similarity), Caenimonas koreensis EMB320T (97.5%) and Ramlibacter solisilvae 5-10T (97.8%). The sequence similarities of strain S4T with other members of the family Comamonadaceae were ≤ 97.5%. The sole respiratory quinone was ubiquinone-8 (Q-8) and the principal polar lipid was phosphatidylethanolamine. The predominant cellular fatty acids were summed feature 3 (iso-C15 :0 2-OH/C16 :1 ω7c), C16:0 and summed feature 8 (C18:1 ω7c and/or C18:1 ω6c). The DNA G + C content was 65.1 mol%. In addition, the average nucleotide identity (ANIu) and in silico DNA-DNA hybridization (dDDH) relatedness values between strain S4T and Caenimonas koreensis were 77.6 and 21%, respectively. Based on genomic, chemotaxonomic, phenotypic, and phylogenetic analyses, strain S4T represents a novel species in the genus Caenimonas, for which the name Caenimonas soli sp. nov. is proposed. The type strain is S4T (= KCTC 72742T = NBRC 114610T).

Novosphingobium olei sp. nov., with the ability to degrade diesel oil, isolated from oil-contaminated soil and proposal to reclassify Novosphingobium stygium as a later heterotypic synonym of Novosphingobium aromaticivorans.

Two yellow-pigmented, non-motile, Gram-stain-negative, and rod-shaped bacteria, designated TW-4T and TNP-2 were obtained from oil-contaminated soil. Both strains degrade diesel oil, hydrolyse aesculin, DNA, Tween 40 and Tween 60. A phylogenetic analysis based on its 16S rRNA gene sequence revealed that strain TW-4T formed a lineage within the family Erythrobacteraceae and clustered as members of the genus Novosphingobium. The closest members of strain TW-4T were Novosphingobium subterraneum DSM 12447T (97.9 %, sequence similarity), Novosphingobium lubricantis KSS165-70T (97.8 %), Novosphingobium taihuense T3-B9T (97.8 %), Novosphingobium aromaticivorans DSM 12444T (97.7 %), Novosphingobium flavum UCT-28T (97.7 %), and Novosphingobium bradum STM-24T (97.6 %). The sequence similarity for other members was ≤97.6 %. The genome of strain TW-4T was 4 683 467 bp long with 44 scaffolds and 4280 protein-coding genes. The sole respiratory quinone was Q-10. The major cellular fatty acids were summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C16 : 0 and C14 : 0 2-OH. The major polar lipids were phosphatidylethanolamine (PE), phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), phosphatidylcholine (PC), phosphatidyl-n-methylethanolamine (PME) and sphingoglycolipid (SGL). The DNA G+C content of the type strain was 65.0 %. The average nucleotide identity (ANIu) and in silico DNA-DNA hybridization (dDDH) relatedness values between strain TW-4T and closest members were below the threshold value for species delineation. Based on polyphasic taxonomic analyses, strain TW-4T represents novel species in the genus Novosphingobium, for which the name Novosphingobium olei sp. nov. is proposed. The type strain is TW-4T (=KACC 21628T=NBRC 114364T) and strain TNP-2 (=KACC 21629=NBRC 114365) represents an additional strain. Based on new data obtained in this study, it is also proposed to reclassify Novosphingobium stygium as a later heterotypic synonym of Novosphingobium aromaticivorans.

Chitinophaga fulva sp. nov., isolated from forest soil.

An aerobic, Gram-stain-negative, oxidase- and catalase-positive, non-motile, non-spore-forming, rod-shaped and yellow-coloured bacterium designated strain G-6-1-13T was isolated from Gwanggyo mountain forest soil. Strain G-6-1-13T could grow at 15-40 °C (optimum, 20-32 °C), pH 4.5-10.5 (optimum, pH 6.0-9.0), at 2 % (w/v) NaCl concentration, and produced flexirubin-type pigments. Phylogenetic analysis based on its 16S rRNA gene sequence showed that strain G-6-1-13T formed a lineage within the genus Chitinophaga that was distinct from other species of the genus. Closest member was Chitinophaga varians 10-7 W-9003T (98.6 % sequence similarity) followed by C. eiseniae DSM 22224T (98.4 %), C. qingshengii JN246T (97.6 %) and C. terrae KP01T (97.4%). The major cellular fatty acids were iso-C15 : 0, C16 : 1 ω5c, and summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1 ω6c). MK-7 was the sole respiratory quinone. The major polar lipids were phosphatidylethanolamine and an unidentified phospholipid. The DNA G+C content of strain G-6-1-13T was 48.7 mol%. Average nucleotide identity and in silico DNA-DNA hybridization were below the species threshold. On the basis of phenotypic, genotypic, phylogenetic and chemotaxonomic characterization, G-6-1-13T represents a novel species in the genus Chitinophaga, for which the name Chitinophaga fulva sp. nov. is proposed. The type strain is G-6-1-13T (=KACC 21624T=NBRC 114361T).

Description of antibiotic-producing novel bacteria Paraburkholderia antibiotica sp. nov. and Paraburkholderia polaris sp. nov.

Two white colony-forming, Gram-stain-negative, non-sporulating and motile bacteria, designated G-4-1-8T and RP-4-7T, were isolated from forest soil and Arctic soil, respectively. Both strains showed antimicrobial activity against Gram-negative pathogens (Pseudomonas aeruginosa and Escherichia coli) and could grow at a pH range of pH 4.0-11.0 (optimum, pH 7.0-9.0). Phylogenetic analyses based on their 16S rRNA gene sequences indicated that strains G-4-1-8T and RP-4-7T formed a lineage within the family Burkholderiaceae and were clustered as members of the genus Paraburkholderia. Strain G-4-1-8T showed the highest 16S rRNA sequence similarity to Paraburkholderia monticola JC2948T (98.1 %), while strain RP-4-7T showed the highest similarity to Paraburkholderia metrosideri DNBP6-1T (98.8 %). The only respiratory quinone in both strains was ubiquinone Q-8. Their principal cellular fatty acids were C16 : 0, cyclo-C17 : 0, summed feature 3 (iso-C15 :0 2-OH and/or C16 :1 ω7c) and summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c). Their major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol and an unidentified aminophospholipid. The DNA G+C content of strains G-4-1-8T and RP-4-7T were 63.7 and 61.3 mol%, respectively, while their genome lengths were 7.44 and 9.67 Mb, respectively. The genomes of both strains showed at least 12 putative biosynthetic gene clusters. The average nucleotide identity and in silico DNA-DNA hybridization relatedness values between both strains and most closely related Paraburkholderia species were below the species threshold values. Based on a polyphasic study, these isolated strains represent novel species belonging to the genus Paraburkholderia, for which the names Paraburkholderia antibiotica sp. nov. (G-4-1-8T= KACC 21617T=NBRC 114603T) and Paraburkholderia polaris sp. nov. (RP-4-7T=KACC 21621T=NBRC 114605T) are proposed.

Cold-shock gene cspC in the genome of Massilia polaris sp. nov. revealed cold-adaptation.

A straw coloured, motile and Gram-stain-negative bacterium, designated RP-1-19T was isolated from soil of Arctic station, Svalbard, Norway. Based on the phylogenetic analysis of its 16S rRNA gene sequence, strain RP-1-19T formed a lineage within the family Oxalobacteraceae and clustered together within the genus Massilia. The closest members were M. violaceinigra B2T (98.6% sequence similarity), M. eurypsychrophilia JCM 30074T (98.3%) and M. atriviolacea SODT (98.1%). The only respiratory quinone was ubiquinone-8. The principal cellular fatty acids were summed feature 3 (iso-C15:0 2-OH/C16:1ω7c) and C16:0. The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol and phosphatidylglycerol. The DNA G + C content of the type strain was 63.2%. The average nucleotide identity and in silico DNA-DNA hybridization values between strain RP-1-19T and closest members were ≤ 80 and 23.2%, respectively. The genome was 4,522,469 bp long with 30 scaffolds and 4076 protein-coding genes. The genome showed eight putative biosynthetic gene clusters responsible for various secondary metabolites. Genome analysis revealed the presence of cold-shock proteins CspA and CspC. Presence of cspA and cspC genes in the genome manifest ecophysiology of strain RP-1-19T that may help in cold-adaptation. Based on these data, strain RP-1-19T represents a novel species in the genus Massilia, for which the name Massilia polaris sp. nov. is proposed. The type strain is RP-1-19T (= KACC 21619T = NBRC 114359T).

Nakamurella aerolata sp. Nov., Isolated from an Automobile Air Conditioning System.

A novel white-colored, aerobic, Gram-stain-positive, rod-shaped bacterium, designated strain DB0629T was isolated from a motor car evaporator core collected in South Korea. Strain DB0629T grew at 10-35 °C, pH 6.0-9.0, and 0-5.0% (w/v) NaCl concentration. The 16S rRNA gene sequence analysis showed that strain DB0629T belonged to the genus Nakamurella, with the nearest phylogenetic neighbor being Nakamurella lactea DSM 19367T (97.6% sequence similarity). The strain comprised diphosphatidylglycerol and phosphatidylinositol as the main polar lipids; MK-8(H4) as a sole respiratory quinone; meso-diaminopimelic acid as the diagnostic diamino acid in the cell-wall peptidoglycan and anteiso-C15:0, anteiso-C17:0, iso-C16:0, iso-C15:0, and C16:0 as the major fatty acids. The average nucleotide identity (ANI) and in silico DNA-DNA hybridization values between strain DB0629T and N. lactea DSM 19367T were 74.9% and 20.8%, respectively, which were below the threshold values of 95% and 70%, respectively. The DNA G + C content was 69.5 mol%. Based on the polyphasic taxonomic data, the novel species Nakamurella aerolata sp. nov. is proposed with the type strain DB0629T (= KCTC 72726T = NBRC 114624T).