Molecular mechanisms of macrolide and fluoroquinolone resistance among Korean isolates of Mycoplasma genitalium over a period of five years 2014–2019

2021 ◽  
Vol 70 (11) ◽  
Author(s):  
Yumi Seo ◽  
Heeyoon Park ◽  
Gilho Lee
Keyword(s):  
Genetic Diversity ◽  
Antimicrobial Resistance ◽  
Type Species ◽  
Mycoplasma Genitalium ◽  
Quinolone Resistance ◽  
23S Rrna ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
23S Rrna Gene

Antimicrobial resistance in Mycoplasma genitalium has become a global issue, and certain groups have a higher probability of acquiring resistant strains. Little is known about the genetic diversity and characteristics of the antimicrobial resistance-determining sites (ARDSs) of M. genitalium in the Korean population. Therefore, we examined the genetic diversity of the ARDSs of M. genitalium-positive urogenital samples obtained from Korean females (G1) and males (G2) visiting primary care clinics and DNA samples from referred males (G3) with persistent urethritis. From 2014 to 2019, 54 patients from G1, 86 patients from G2, and 68 patients from G3 were included in the study. Sanger sequencing was performed on the 2058/2059 sites in the 23S rRNA gene and quinolone resistance-determining regions (QRDRs) of M. genitalium . The rates of mutation in G1, G2, and G3 were 1.85, 5.81, and 48.53 %, respectively, for A2059G in the 23S rRNA gene (P<0.001); 1.85, 0, and 17.78 %, respectively, for M95R or I in gyrA (P<0.001); 0, 0, and 31.11 %, respectively, for D99N or G in gyrA (P<0.001); and 7.41, 16.28, and 30 %, respectively, for S83R or N or I in parC (P=0.015). A2059G significantly increased the risk of mutations at the gyrA95, gyrA99, and parC83 sites (all P<0.01). In conclusion, although the genetic diversity of the ARDSs of M. genitalium was variable among the groups, it was generally lower in isolates with macrolide resistance and higher in isolates with quinolone resistance in Korea compared with the isolates in other countries. The G3 group demonstrated increased genetic diversity at the A2059G, gyrA95, gyrA99, and parC83 sites.

Streptococcus dentisani sp. nov., a novel member of the mitis group

2014 ◽  
Vol 64 (Pt_1) ◽  
pp. 60-65 ◽  
Author(s):  
Anny Camelo-Castillo ◽  
Alfonso Benítez-Páez ◽  
Pedro Belda-Ferre ◽  
Raúl Cabrera-Rubio ◽  
Alex Mira
Keyword(s):  
Type Species ◽  
Spherical Shape ◽  
Culture Collection ◽  
23S Rrna ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
23S Rrna Gene ◽  
Tooth Surfaces ◽  
Streptococcal Species

Genomic, taxonomic and biochemical studies were performed on two strains of α-haemolytic streptococci that showed them to be clustered with major members of the Streptococcus mitis group. These Gram-stain-positive strains were isolated from tooth surfaces of caries-free humans and showed the classical spherical shape of streptococcal species growing in chains. Sequence analysis from concatenated 16S and 23S rRNA gene and sodA genes showed that these strains belonged to the mitis group, but both of them clustered into a new phylogenetic branch. The genomes of these two isolates were sequenced, and whole-genome average nucleotide identity (ANI) demonstrated that these strains significantly differed from any streptococcal species, showing ANI values under 91 % even when compared with the phylogenetically closest species such as Streptococcus oralis and S. mitis . Biochemically, the two isolates also showed distinct metabolic features relative to closely related species, like α-galactosidase activity. From the results of the present study, the name Streptococcus dentisani sp. nov. is proposed to accommodate these novel strains, which have been deposited in open collections at the Spanish type Culture Collection (CECT) and Leibniz Institute DSMZ–German Collection of Microorganisms and Cell Cultures (DSMZ), being respectively identified as Streptococcus dentisani Str. 7746 ( = CECT 8313 = DSM 27089) and Streptococcus dentisani Str. 7747T ( = CECT 8312T = DSM 27088T).

scholarly journals Neisseria oralis sp. nov., isolated from healthy gingival plaque and clinical samples

2013 ◽  
Vol 63 (Pt_4) ◽  
pp. 1323-1328 ◽  
Author(s):  
William J. Wolfgang ◽  
Teresa V. Passaretti ◽  
Reashma Jose ◽  
Jocelyn Cole ◽  
An Coorevits ◽  
...  
Keyword(s):  
Type Strain ◽  
Type Species ◽  
Novel Species ◽  
Sequence Similarity ◽  
23S Rrna ◽  
Clinical Samples ◽  
Rrna Gene ◽  
The Novel ◽  
Content Type ◽  
Link Type

A polyphasic analysis was undertaken of seven independent isolates of Gram-negative cocci collected from pathological clinical samples from New York, Louisiana, Florida and Illinois and healthy subgingival plaque from a patient in Virginia, USA. The 16S rRNA gene sequence similarity among these isolates was 99.7–100 %, and the closest species with a validly published name was Neisseria lactamica (96.9 % similarity to the type strain). DNA–DNA hybridization confirmed that these isolates are of the same species and are distinct from their nearest phylogenetic neighbour, N. lactamica . Phylogenetic analysis of 16S and 23S rRNA gene sequences indicated that the novel species belongs in the genus Neisseria . The predominant cellular fatty acids were C16 : 0, summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH) and C18 : 1ω7c. The cellular fatty acid profile, together with other phenotypic characters, further supports the inclusion of the novel species in the genus Neisseria . The name Neisseria oralis sp. nov. (type strain 6332T  = DSM 25276T  = LMG 26725T) is proposed.

scholarly journals Genome sequence analyses show that Neisseria oralis is the same species as ‘ Neisseria mucosa var. heidelbergensis’

2013 ◽  
Vol 63 (Pt_10) ◽  
pp. 3920-3926 ◽  
Author(s):  
Julia S. Bennett ◽  
Keith A. Jolley ◽  
Martin C. J. Maiden
Keyword(s):  
Genome Sequence ◽  
Type Species ◽  
Whole Genome Sequence ◽  
23S Rrna ◽  
Rrna Gene ◽  
Whole Genome ◽  
Content Type ◽  
Link Type ◽  
The Family ◽  
Neisseria Mucosa

Phylogenies generated from whole genome sequence (WGS) data provide definitive means of bacterial isolate characterization for typing and taxonomy. The species status of strains recently defined with conventional taxonomic approaches as representing Neisseria oralis was examined by the analysis of sequences derived from WGS data, specifically: (i) 53 Neisseria ribosomal protein subunit (rps) genes (ribosomal multi-locus sequence typing, rMLST); and (ii) 246 Neisseria core genes (core genome MLST, cgMLST). These data were compared with phylogenies derived from 16S and 23S rRNA gene sequences, demonstrating that the N. oralis strains were monophyletic with strains described previously as representing ‘ Neisseria mucosa var. heidelbergensis’ and that this group was of equivalent taxonomic status to other well-described species of the genus Neisseria . Phylogenetic analyses also indicated that Neisseria sicca and Neisseria macacae should be considered the same species as Neisseria mucosa and that Neisseria flavescens should be considered the same species as Neisseria subflava . Analyses using rMLST showed that some strains currently defined as belonging to the genus Neisseria were more closely related to species belonging to other genera within the family; however, whole genome analysis of a more comprehensive selection of strains from within the family Neisseriaceae would be necessary to confirm this. We suggest that strains previously identified as representing ‘ N. mucosa var. heidelbergensis’ and deposited in culture collections should be renamed N. oralis . Finally, one of the strains of N. oralis was able to ferment lactose, due to the presence of β-galactosidase and lactose permease genes, a characteristic previously thought to be unique to Neisseria lactamica , which therefore cannot be thought of as diagnostic for this species; however, the rMLST and cgMLST analyses confirm that N. oralis is most closely related to N. mucosa .

scholarly journals Mycoplasma anserisalpingitidis sp. nov., isolated from European domestic geese (Anser anser domesticus) with reproductive pathology

2020 ◽  
Vol 70 (4) ◽  
pp. 2369-2381 ◽  
Author(s):  
Dmitriy V. Volokhov ◽  
Dénes Grózner ◽  
Miklós Gyuranecz ◽  
Naola Ferguson-Noel ◽  
Yamei Gao ◽  
...  
Keyword(s):  
16S Rrna ◽  
Type Strain ◽  
Type Species ◽  
Novel Species ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
23S Rrna ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type

In 1983, Mycoplasma sp. strain 1220 was isolated in Hungary from the phallus lymph of a gander with phallus inflammation. Between 1983 and 2017, Mycoplasma sp. 1220 was also identified and isolated from the respiratory tract, liver, ovary, testis, peritoneum and cloaca of diseased geese in several countries. Seventeen studied strains produced acid from glucose and fructose but did not hydrolyse arginine or urea, and all grew under aerobic, microaerophilic and anaerobic conditions at 35 to 37 ˚C in either SP4 or pleuropneumonia-like organism medium supplemented with glucose and serum. Colonies on agar showed a typical fried-egg appearance and transmission electron microscopy revealed a typical mycoplasma cellular morphology. Molecular characterization included analysis of the following genetic loci: 16S rRNA, 23S rRNA, 16S–23S rRNA ITS, rpoB, rpoC, rpoD, uvrA, parC, topA, dnaE, fusA and pyk. The genome was sequenced for type strain 1220T. The 16S rRNA gene sequences of studied strains of Mycoplasma sp. 1220 shared 99.02–99.19 % nucleotide similarity with M. anatis strains but demonstrated ≤95.00–96.70 % nucleotide similarity to the 16S rRNA genes of other species of the genus Mycoplasma . Phylogenetic, average nucleotide and amino acid identity analyses revealed that the novel species was most closely related to Mycoplasma anatis . Based on the genetic data, we propose a novel species of the genus Mycoplasma , for which the name Mycoplasma anserisalpingitidis sp. nov. is proposed with the type strain 1220T (=ATCC BAA-2147T=NCTC 13513T=DSM 23982T). The G+C content is 26.70 mol%, genome size is 959110 bp.

scholarly journals Evidence for Multidrug Resistance in NonpathogenicMycoplasmaSpecies Isolated from South African Poultry

2018 ◽  
Vol 84 (21) ◽  
Author(s):  
Amanda Beylefeld ◽  
Pamela Wambulawaye ◽  
Dauda Garba Bwala ◽  
Johannes Jacobus Gouws ◽  
Obed Mooki Lukhele ◽  
...  
Keyword(s):  
Multidrug Resistance ◽  
South African ◽  
Macrolide Resistance ◽  
Quinolone Resistance ◽  
23S Rrna ◽  
Clear Correlation ◽  
Rrna Gene ◽  
Poultry Production ◽  
Content Type ◽  
23S Rrna Gene

ABSTRACTOne hundred seventy-eight mycoplasma strains isolated from South African poultry flocks between 2003 and 2015 were identified by full-genome sequencing and phylogenetic analysis of the 16S rRNA gene and were classified as follows:Mycoplasma gallisepticum(25%),M. gallinarum(25%),M. gallinaceum, (23%),M. pullorum(14%),M. synoviae(10%), andM. iners(3%), as well as oneAcheoplasma laidlawiistrain (1%). MIC testing was performed on the axenic samples, and numerous strains of each species were resistant to either chlortetracycline or tylosin or both, with variable sensitivity to enrofloxacin. The strains of all species tested remained sensitive to tiamulin, except for oneM. gallinaceumsample that demonstrated intermediate sensitivity. The mutation of A to G at position 2059 (A2059G) in the 23S rRNA gene, which is associated with macrolide resistance, was found in the South AfricanM. gallisepticumandM. synoviaestrains, as well as a clear correlation between macrolide resistance inM. gallinarumandM. gallinaceumand mutations G354A and G748A in the L4 ribosomal protein and 23S rRNA gene, respectively. No correlation between resistance and point mutations in the genes studied could be found forM. pullorum. Only a few strains were resistant to enrofloxacin, apart from oneM. synoviaestrain with point mutation D420N, which has been associated with quinolone resistance, and no other known markers for quinolone resistance were found in this study. Proportionally more antimicrobial-resistant strains were detected inM. gallinaceum,M. gallinarum, andM. pullorumthan inM. gallisepticumandM. synoviae. Of concern, threeM. gallinaceumstrains showed multidrug resistance to chlortetracycline, tylosin, and oxytetracycline.IMPORTANCENonpathogenic poultryMycoplasmaspecies are often overlooked due to their lesser impact on poultry health and production compared to the OIE-listed pathogenic strainsM. gallisepticumandM. synoviae. The use of antimicrobials as in-feed growth promoters and for the control of mycoplasmosis is common in poultry production across the world. Here, we provide evidence that certain nonpathogenicMycoplasmaspecies are acquiring multidrug resistance traits. This would have significant implications if these species, for which no vaccines are applied, are able to transfer their antibiotic resistance genes to other mycoplasmas and bacteria that may enter the human food chain.

scholarly journals Bradyrhizobium erythrophlei sp. nov. and Bradyrhizobium ferriligni sp. nov., isolated from effective nodules of Erythrophleum fordii

2015 ◽  
Vol 65 (Pt_6) ◽  
pp. 1831-1837 ◽  
Author(s):  
Yao Yao ◽  
Xin Hua Sui ◽  
Xiao Xia Zhang ◽  
En Tao Wang ◽  
Wen Xn Chen
Keyword(s):  
Type Strain ◽  
Type Species ◽  
Novel Species ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
Erythrophleum Fordii ◽  
Slow Growing

Six slow-growing rhizobial strains isolated from effective nodules of Erythrophleum fordii were classified into the genus Bradyrhizobium based on their 16S rRNA gene sequences. The results of multilocus sequence analysis of recA, glnII and gyrB genes and 16S–23S rRNA intergenic spacer (IGS) sequence phylogeny indicated that the six strains belonged to two novel species, represented by CCBAU 53325T and CCBAU 51502T, which were consistent with the results of DNA–DNA hybridization; CCBAU 53325T had 17.65–25.59 % relatedness and CCBAU 51502T had 22.69–44.58 % relatedness with five closely related type strains, Bradyrhizobium elkanii USDA 76T, B. pachyrhizi LMG 24246T, B. lablabi CCBAU 23086T, B. jicamae LMG 24556T and B. japonicum USDA 6T. In addition, analysis of phenotypic characteristics and fatty acid profiles also distinguished the test strains from defined species of Bradyrhizobium . Two novel species, Bradyrhizobium erythrophlei sp. nov., represented by the type strain CCBAU 53325T ( = HAMBI 3614T = CGMCC 1.13002T = LMG 28425T), and Bradyrhizobium ferriligni sp. nov., represented by the type strain CCBAU 51502T ( = HAMBI 3613T = CGMCC 1.13001T), are proposed to accommodate the strains.

scholarly journals Leuconostoc mesenteroides subsp. suionicum subsp. nov.

2012 ◽  
Vol 62 (Pt_7) ◽  
pp. 1548-1551 ◽  
Author(s):  
Chun Tao Gu ◽  
Fang Wang ◽  
Chun Yan Li ◽  
Fei Liu ◽  
Gui Cheng Huo
Keyword(s):  
Type Species ◽  
Gene Sequence ◽  
Leuconostoc Mesenteroides ◽  
Sequence Similarity ◽  
Intergenic Spacer ◽  
23S Rrna ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
Content Type ◽  
Link Type

Strains LMG 8159 and LMG 11499 were reclassified by a polyphasic approach, including 16S rRNA gene sequence analysis, 16S–23S rRNA intergenic spacer (IGS) sequence analysis, (GTG)5-PCR fingerprinting, RAPD fingerprinting, fatty acid methyl ester analysis and an analysis of phenotypic features using API 50 CH. The two strains were closely related to the type strains of the three defined subspecies of Leuconostoc mesenteroides , showing 99.7–99.9 % 16S rRNA gene sequence similarity, 99.2 % 16S–23S rRNA gene intergenic spacer sequence similarity, 97.1–97.4 % pheS gene sequence similarity and 98.0–98.2 % rpoA gene sequence similarity. Low atpA gene sequence similarity (91.4–91.7 %), (GTG)5-PCR fingerprinting, RAPD fingerprinting, fatty acid compositions and phenotypic features allowed us to differentiate strains LMG 8159 and LMG 11499 from all established subspecies within L. mesenteroides . Based upon the data obtained in the present and previous studies, a novel subspecies is proposed within the species L. mesenteroides , Leuconostoc mesenteroides subsp. suionicum subsp. nov., with the type strain LMG 8159T ( = ATCC 9135T  = DSM 20241T  = NCIMB 6992T).

Epibacterium ulvae gen. nov., sp. nov., epibiotic bacteria isolated from the surface of a marine alga

2013 ◽  
Vol 63 (Pt_5) ◽  
pp. 1589-1596 ◽  
Author(s):  
Anahit Penesyan ◽  
Sven Breider ◽  
Peter Schumann ◽  
Brian J. Tindall ◽  
Suhelen Egan ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Type Species ◽  
Marine Alga ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
23S Rrna ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type

Two Gram-reaction-negative, rod-shaped, motile bacteria, designated strains U82 and U95T, were isolated from the marine alga Ulva australis collected at Sharks Point, Clovelly, a rocky intertidal zone near Sydney, Australia. Both strains were oxidase- and catalase-positive, formed brown- to black-pigmented colonies and required NaCl for growth. Phylogenetic analysis based on nearly complete 16S rRNA gene sequences revealed that these strains belong to the Roseobacter clade within the Alphaproteobacteria . The 16S rRNA genes of both strains were identical across the sequenced 1326 nt, but showed differences in the intergenic spacer region (ITS) between the 16S and the 23S rRNA genes. At the genomic level the DNA G+C contents of strains U82 and U95T were identical (52.6 mol%) and they had a DNA–DNA hybridization value of 83.7 %, suggesting that these strains belong to the same species. The closest described phylogenetic neighbour to strains U82 and U95T was Thalassobius aestuarii DSM 15283T with 95.8 % 16S rRNA gene sequence similarity. Other close relatives include further species of the genera Thalassobius and Shimia . Strains U82 and U95T were negative for bacteriochlorophyll a production, showed antibacterial activity towards other marine bacteria, were resistant to the antibiotics gentamicin and spectinomycin and were unable to hydrolyse starch or gelatin. The major fatty acids (>1 %) were 18 : 1ω7c, 16 : 0, 18 : 2, 10 : 0 3-OH, 12 : 0, 20 : 1 2-OH and 18 : 0. The polar lipid pattern indicated the presence of phosphatidylglycerol, phosphatidylcholine, two unidentified aminolipids and four unidentified phospholipids. Both strains produced ubiquinone 10 (Q-10) as the sole respiratory lipoquinone. Based on their phenotypic and phylogenetic characteristics, it is suggested that strains U82 and U95T are members of a novel species within a new genus for which the name Epibacterium ulvae gen. nov., sp. nov. is proposed. The type strain of the type species is U95T ( = DSM 24752T = LMG 26464T).

scholarly journals Helicobacter himalayensis sp. nov. isolated from gastric mucosa of Marmota himalayana

2015 ◽  
Vol 65 (Pt_6) ◽  
pp. 1719-1725 ◽  
Author(s):  
Shoukui Hu ◽  
Dong Jin ◽  
Shan Lu ◽  
Sha Liu ◽  
Ji Zhang ◽  
...  
Keyword(s):  
Gastric Mucosa ◽  
16S Rrna ◽  
Type Species ◽  
Gene Sequence ◽  
Novel Species ◽  
23S Rrna ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
Content Type ◽  
Link Type

A Gram-stain-negative, microaerophilic strain, 80(YS1)T, with a spiral-shaped morphology and 1–2 sheathed flagella at each end of the cells was isolated from the gastric mucosa of Marmota himalayana, the animal reservoir of Yersinia pestis in China, on the Qinghai-Tibet Plateau. The strain grew at 30, 35 and 42 °C, but not at 25 °C. Growth was in the form of a thinly spreading film on brain heart infusion agar containing 8 % sheep blood under microaerobic conditions. The strain did not hydrolyse urea or hippurate, and did not grow on media containing 1 % glycine. It reduced nitrate to nitrite, and was catalase- and alkaline-phosphatase-positive, susceptible to nalidixic acid and resistant to cefalotin. It was positive for genus-specific PCR for the genus Helicobacter , but could not be classified to any recognized species according biochemical tests results. Therefore, a phylogenetic study based on 16S rRNA, 23S rRNA, 60 kDa heat-shock protein (hsp60) and gyrase subunit B (gyrB) genes was conducted. The 16S rRNA gene sequence (1468 bp) analysis showed that strain 80(YS1)T was most closely related to Helicobacter marmotae (96.7 % similarity). The 23S rRNA gene sequence (2879 bp) analysis showed that the strain was most closely related to Helicobacter canis (96 % similarity). The complete gyrB gene sequence (2325 bp) analysis showed that it was related phylogenetically to Helicobacter cinaedi (79.4 % similarity) and H. marmotae (79.1 % similarity). Analysis of the partial sequence of the hsp60 gene of strain 80(YS1)T showed closest similarity to the sequences of Helicobacter equorum (82 %) and H. cinaedi (81 %), respectively. However, there was no hsp60 sequence of H. marmotae available for analysis. The data of morphological, biochemical and phylogenetic characteristics all supported that this strain represents a novel species. The name Helicobacter himalayensis sp. nov. is proposed for this novel species with the type strain 80(YS1)T ( = CGMCC 1.12864T = DSM 28742T)

scholarly journals Kushneria phosphatilytica sp. nov., a phosphate-solubilizing bacterium isolated from a solar saltern

2020 ◽  
Author(s):  
Guang-Xun Du ◽  
Ling-Yun Qu ◽  
Xu-Guang Hong ◽  
Cheng-Hua Li ◽  
De-Wen Ding ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Type Species ◽  
Sequence Similarity ◽  
23S Rrna ◽  
Phylogenetic Position ◽  
Rrna Gene ◽  
Circular Chromosome ◽  
Content Type ◽  
Link Type ◽  
A Genome

A Gram-stain-negative, motile, rod-shaped, non-endospore-forming, aerobic and halophilic bacterium, designated strain YCWA18T, was isolated from the sediment of Jimo-Daqiao saltern in China. This strain was able to grow at NaCl concentrations in the range 0.5–20 % (w/v) with optimum growth at 6 % (w/v) NaCl. Growth occurred at temperatures of 4–40 °C (optimum 28 °C) and pH 4.0–9.0 (optimum 7.0). Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YCWA18T belonged to the genus Kushneria and shared the highest sequence similarity of 98.7 % with Kushneria sinocarnis DSM 23229T. Moreover, the phylogenetic analysis based on the 23S rRNA gene sequence also confirmed the phylogenetic position of this novel strain. The predominant fatty acids were C16 : 0, C17 : 0 cyclo and C12 : 0 3-OH. The major isoprenoid quinone was Q-9 (94.2 %) and the polar lipids were diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), an unidentified aminolipid (AL), an unidentified phospholipids (PL) and two unidentified lipids (L). The complete genome of strain YCWA18T consisted of a single, circular chromosome of 3 624 619 bp, with an average G+C content of 59.1 mol%. A genome-based phylogenetic tree constructed using an up-to-date bacterial core gene set (UBCG) showed that strain YCWA18T formed a clade with K. sinocarnis DSM 23229T. However, the level of the ANI and dDDH values between strain YCWA18T and K. sinocarnis DSM 23229T were 82.3 and 24.6 %, respectively, which were low enough to distinguish strain YCWA18T from K. sinocarnis DSM 23229T. Overall, based on the phenotypic, chemotaxonomic, phylogenetic and genomic analyses, strain YCWA18T represents a novel species of genus Kushneria . The name Kushneria phosphatilytica sp. nov. is proposed, with the type strain YCWA18T (=CGMCC 1.9149T=NCCB 100306T).

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