Molecular phylogeny of black fungus gnats (Diptera: Sciaroidea: Sciaridae) and the evolution of larval habitats

The phylogeny of the family Sciaridae is reconstructed, based on maximum likelihood, maximum parsimony, and Bayesian analyses of 4809 bp from two mitochondrial (COI and 16S) and two nuclear (18S and 28S) genes for 100 taxa including the outgroup taxa. According to the present phylogenetic analyses, Sciaridae comprise three subfamilies and two genus groups: Sciarinae, Chaetosciara group, Cratyninae, and Pseudolycoriella group + Megalosphyinae. Our molecular results are largely congruent with one of the former hypotheses based on morphological data with respect to the monophyly of genera and subfamilies (Sciarinae, Megalosphyinae, and part of postulated “new subfamily”); however, the subfamily Cratyninae is shown to be polyphyletic, and the genera Bradysia, Corynoptera, Leptosciarella, Lycoriella, and Phytosciara are also recognized as non-monophyletic groups. While the ancestral larval habitat state of the family Sciaridae, based on Bayesian inference, is dead plant material (plant litter + rotten wood), the common ancestors of Phytosciara and Bradysia are inferred to living plants habitat. Therefore, shifts in larval habitats from dead plant material to living plants may have occurred within the Sciaridae at least once. Based on the results, we discuss phylogenetic relationships within the family, and present an evolutionary scenario of development of larval habitats.     

Parendozoicomonas haliclonae gen. nov. sp. nov. isolated from a marine sponge of the genus Haliclona and description of the family Endozoicomonadaceae fam. nov. comprising the genera Endozoicomonas,Parendozoicomonas, and Kistimonas

Two Gram-stain-negative, facultative anaerobic, motile, rod-shaped strains, S-B4-1U^T and JOB-63a, forming small whitish transparent colonies on marine agar, were isolated from a sponge of the genus Haliclona. The strains shared 99.7% 16S rRNA gene sequence identity and a DNA-DNA hybridization value of 100%, but were differentiated by genomic fingerprinting using rep-PCRs. 16S rRNA gene sequence phylogeny placed the strains as a sister branch to the monophyletic genus Endozoicomonas (Oceanospirillales; Gammaproteobacteria) with 92.3–94.3% 16S rRNA gene sequence similarity to Endozoicomonas spp., 91.9 and 92.1% to Candidatus Endonucleobacter bathymodiolin, and 91.9 to 92.1% to the type strains of Kistimonas spp. Core genome based phylogeny of strain S-B4-1U^T confirmed the phylogenetic placement. Major fatty acids were summed feature 3 (C_16:1 ω7c/C_16:1 ω6c) and 8 (C_18:1 ω7c/C_18:1 ω6c) followed by C_10:0 3-OH, C_16:0, and C_18:0. The G + C content was 50.1–51.4 mol%. The peptidoglycan diamino acid of strain S-B4-1U^T was meso-diaminopimelic acid, the predominant polyamine spermidine, the major respiratory quinone ubiquinone Q-9; phosphatidylethanolamine, phosphatidylglycerol and phosphatidylserine were major polar lipids. Based on the clear phylogenetic distinction, the genus Parendozoicomonas gen. nov. is proposed, with Parendozoicomonas haliclonae sp. nov. as type species and strain S-B4-1U^T (= CCM 8713^T = DSM 103671^T = LMG 29769^T) as type strain and JOB-63a as a second strain of the species. Based on the 16S rRNA gene sequence phylogeny of the Oceanospirillales within the Gammaproteobacteria, the Endozoicomonaceae fam. nov. is proposed including the genera Endozoicomonas, Parendozoicomonas, and Kistimonas as well as the Candidatus genus Endonucleobacter.     

Agrobacterium rosae sp. nov., isolated from galls on different agricultural crops

The plant tumorigenic strain NCPPB 1650^T isolated from Rosa × hybrida, and four nonpathogenic strains isolated from tumors on grapevine (strain 384), raspberry (strain 839) and blueberry (strains B20.3 and B25.3) were characterized by using polyphasic taxonomic methods. Based on 16S rRNA gene phylogeny, strains were clustered within the genus Agrobacterium. Furthermore, multilocus sequence analysis (MLSA) based on the partial sequences of atpD, recA and rpoB housekeeping genes indicated that five strains studied form a novel Agrobacterium species. Their closest relatives were Agrobacterium sp. R89-1, Agrobacterium rubi and Agrobacterium skierniewicense. Authenticity of the novel species was confirmed by average nucleotide identity (ANI) and in silico DNA–DNA hybridization (DDH) comparisons between strains NCPPB 1650^T and B20.3, and their closest relatives, since obtained values were considerably below the proposed thresholds for the species delineation. Whole-genome-based phylogeny further supported distinctiveness of the novel species, that forms together with A. rubi, A. skierniewicense and Agrobacterium sp. R89-1 a well-delineated sub-clade of Agrobacterium spp. named “rubi”. As for other species of the genus Agrobacterium, the major fatty acid of the strains studied was 18:1 w7c (73.42–78.12%). The five strains studied were phenotypically distinguishable from other species of the genus Agrobacterium. Overall, polyphasic characterization showed that the five strains studied represent a novel species of the genus Agrobacterium, for which the name Agrobacterium rosae sp. nov. is proposed. The type strain of A. rosae is NCPPB 1650^T (=DSM 30203^T = LMG 230^T = CFBP 4470^T = IAM 13558^T = JCM 20915^T).     

Morphology and phylogeny of Bryophryoides ocellatus n. g., n. sp. (Ciliophora,Colpodea) from in situ soil percolates of Idaho,U.S.A.

We describe the morphology and 18S rDNA phylogeny of Bryophryoides ocellatus n. g., n. sp., a bryophryid ciliate inhabiting in situ soil percolates from Idaho, U.S.A. The new genus is distinguished from other bryophryid genera by a combination of the following features: (1) kreyellid (irregularly meshed) silverline pattern, (2) polymorphic adoral organelles in the preoral suture, (3) absence of vestibular kineties. In phylogenetic analyses, Bryophryoides ocellatus is most closely related to Bryophrya gemmea. The 18S rDNA sequence pairwise distance of 2% between these genera, while similar to that between many colpodidan species, exceeds that between some colpodidan genera (e.g. Mykophagophrys and Pseudoplatyophrya, 1.1%), further supporting establishment of the new genus. Topology hypothesis testing strongly supports the monophyly of the Colpodida including the bryophryids. Despite weak nodal support, tests of topology constraints narrowly reject the non-monophyly of the sequenced Bryophryidae (Bryophrya + Bryophryoides + Notoxoma). Likewise, the monophyletic origin of the sequenced Bryophryidae is indicated in the phylogenetic networks though with low support.     

Salinivibrio kushneri sp. nov., a moderately halophilic bacterium isolated from salterns

Ten Gram-strain-negative, facultatively anaerobic, moderately halophilic bacterial strains, designated AL184^T, IB560, IB563, IC202, IC317, MA421, ML277, ML318, ML328A and ML331, were isolated from water ponds of five salterns located in Spain. The cells were motile, curved rods and oxidase and catalase positive. All of them grew optimally at 37 °C, at pH 7.2–7.4 and in the presence of 7.5% (w/v) NaCl. Based on phylogenetic analyses of the 16S rRNA, the isolates were most closely related to Salinivibrio sharmensis BAG^T (99.6–98.2% 16S rRNA gene sequence similarity) and Salinivibrio costicola subsp. costicola ATCC 35508^T (99.0–98.1%). According to the MLSA analyses based on four (gyrB, recA, rpoA and rpoD) and eight (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA and topA) concatenated gene sequences, the most closely relatives were S. siamensis JCM 14472^T (96.8–95.4% and 94.9–94.7%, respectively) and S. sharmensis DSM 18182^T (94.0–92.6% and 92.9–92.7%, respectively). In silico DNA–DNA hybridization (GGDC) and average nucleotide identity (ANI) showed values of 23.3–44.8% and 80.2–91.8%, respectively with the related species demonstrating that the ten isolates constituted a single novel species of the genus Salinivibrio. Its pangenome and core genome consist of 6041 and 1230 genes, respectively. The phylogeny based on the concatenated orthologous core genes revealed that the ten strains form a coherent phylogroup well separated from the rest of the species of the genus Salinivibrio. The major cellular fatty acids of strain AL184^T were C_16:0 and C_18:1. The DNA G + C content range was 51.9–52.5 mol% (T_m) and 50.2–50.9 mol% (genome). Based on the phylogenetic-phylogenomic, phenotypic and chemotaxonomic data, the ten isolates represent a novel species of the genus Salinivibrio, for which the name Salinivibrio kushneri sp. nov. is proposed. The type strain is AL184^T (= CECT 9177^T = LMG 29817^T).     

Systematic revision of the avian family Cisticolidae based on a multi-locus phylogeny of all genera

The avian taxon Cisticolidae includes c. 110 species which are distributed throughout the tropical and subtropical parts of the Old World. We estimated the phylogeny of 47 species representing all genera assumed to be part of Cisticolidae based on sequence data from two mitochondrial and two nuclear markers, in total 3495 bp. Bayesian inference and maximum likelihood analyses resulted in a generally well-supported phylogeny which clarified the position of several previously poorly known taxa. The placement of Drymocichla, Malcorus, Micromacronus, Oreophilais, Phragmacia, Phyllolais, Poliolais and Urorhipis in Cisticolidae is corroborated, whereas Rhopophilus and Scotocerca are removed from Cisticolidae. Urorhipis and Heliolais are placed in the genus Prinia whereas Prinia burnesii is shown to be part of Timaliidae, and is placed in the genus Laticilla. Although not recovered by all single loci independently, four major clades were identified within Cisticolidae, and one of these is here described as a new taxon (Neomixinae).     

Pseudomonas gallaeciensis sp. nov., isolated from crude-oil-contaminated intertidal sand samples after the Prestige oil spill

Strains V113^T, V92 and V120 have been isolated from sand samples taken at the Atlantic intertidal shore in Galicia, Spain, after the Prestige oil spill. A preliminary analysis of the 16S rRNA and the partial rpoD gene sequences indicated that these strains belonged to the Pseudomonas genus, but they were distinct from any known Pseudomonas species. They were extensively characterized by a polyphasic taxonomic approach and phylogenetic data that confirmed that these strains belonged to the Pseudomonas pertucinogena group. Phylogenetic analysis of 16S rRNA, gyrB and rpoD gene sequences showed that the three strains were 99% similar and were closely related to members of the P. pertucinogena group, with less than 94% similarity to strains of established species; Pseudomonas pachastrellae was the closest relative. The Average Nucleotide Index based on blast values was 89.0% between V113^T and the P. pachastrellae type strain, below the accepted species level (95%). The predominant cellular fatty acid contents and whole cell protein profiles determined by MALDI-TOF mass spectrometry also differentiated the studied strains from known Pseudomonas species. We therefore conclude that strains V113^T, V92 and V120 represent a novel species of Pseudomonas, for which the name Pseudomonas gallaeciensis is proposed; the type strain is V113^T (= CCUG 67583^T = LMG 29038^T).     

Phylogeny and systematic revision of the karyorelictid genus Remanella (Ciliophora,Karyorelictea) with descriptions of two new species

We present here the systematic relationships and taxonomy of some newly collected karyorelictid ciliates from the genus Remanella isolated from the intertidal zone in China. Three species were investigated using observation in vivo and silver staining methods, of which two new species were identified: R. macrostoma sp. nov. and R. achroma sp. nov. Small subunit (SSU) rRNA genes were sequenced for six species and phylogenetic analyses indicated that the genus Remanella is paraphyletic, with three Loxodes species nesting within it. The monophyly of Remanella, however, could not be rejected through the approximately unbiased (AU) test (p = 0.311 > 0.05) and therefore we considered Remanella to be a valid genus. Based on a critical review of the literature, the validity of all nominal species of Remanella was discussed. Remanella unirugosa and R. multicorpusculata were regarded as junior synonyms of R. rugosa and R. granulosa respectively. We also presented here an illustrated taxonomic key based on morphologic and morphometric characteristics and containing all Remanella species considered identifiable in this revision.     

Understanding the formation of ancient intertropical disjunct distributions using Asian and Neotropical hinged-teeth snakes (Sibynophis and Scaphiodontophis: Serpentes: Colubridae)

Numerous taxa show ancient intertropical disjunct distributions. Many can be explained by well-known processes of historical vicariance, such as the breakup of Gondwanaland. Others, such as Asian–Neotropical divergences are not as well understood. To clarify the phylogenetic position and understand biogeographic and temporal origins of the geographically disjunct and morphologically unique genera of hinged-teeth snakes, Scaphiodontophis (n = 1) and Sibynophis (n = 9; Colubridae), we inferred a time-calibrated phylogeny with additional 107 taxa representing the superfamily Colubroidea using four genes (c-mos, cyt-b, ND2, RAG-1; 3085 bp). We used this tree to estimate ancestral areas for the group. The results show that Scaphiodontophis is sister to Sibynophis, both originated in the late Eocene/Oligocene in Asia and likely dispersed through Beringia to the New World, but unlike other snake groups left no extant species in temperate North America. Current recognition of Scaphiodontophiinae renders Colubrinae paraphyletic, and we resurrect the previously named subfamily Sibynophiinae to encompass both genera and use the tribes Sibynophiini (Sibynophis) and Scaphiodontophiini (Scaphiodontophis) to highlight the geographically distinct areas occupied by these taxa. These results suggest that intercontinental dispersal with extinction in intermediate areas can explain puzzling patterns of ancient intertropical disjunct distributions.     

Analysis of Actinobacteria from mould-colonized water damaged building material

Mould-colonized water damaged building materials are frequently co-colonized by actinomycetes. Here, we report the results of the analyses of Actinobacteria on different wall materials from water damaged buildings obtained by both cultivation-dependent and cultivation-independent methods. Actinobacteria were detected in all but one of the investigated materials by both methods. The detected concentrations of Actinobacteria ranged between 1.8 × 10⁴ and 7.6 × 10⁷ CFU g^?1 of investigated material. A total of 265 isolates from 17 materials could be assigned to 31 different genera of the class Actinobacteria on the basis of 16S rRNA gene sequence analyses. On the basis of the cultivation-independent approach, 16S rRNA gene inserts of 800 clones (50%) were assigned to 47 different genera. Representatives of the genera Streptomyces, Amycolatopsis, Nocardiopsis, Saccharopolyspora, Promicromonospora, and Pseudonocardia were found most frequently. The results derived from both methods indicated a high abundance and variety of Actinobacteria in water damaged buildings. Four bioaerosol samples were investigated by the cultivation-based approach in order to compare the communities of Actinobacteria in building material and associated air samples. A comparison of the detected genera of bioaerosol samples with those directly obtained from material samples resulted in a congruent finding of 9 of the overall 35 detected genera (25%), whereas four genera were only detected in bioaerosol samples.     

Multilocus phylogeny and recent rapid radiation of the viviparous sea snakes (Elapidae: Hydrophiinae)

The viviparous sea snakes (Hydrophiinae: Hydrophiini) comprise a young but morphologically and ecologically diverse clade distributed throughout the Indo-Pacific. Despite presenting a very promising model for marine diversification studies, many relationships among the 62 species and 16 genera in Hydrophiini remain unresolved. Here, we extend previous taxonomic and genomic sampling for Hydrophiini using three mitochondrial fragments and five nuclear loci for multiple individuals of 39 species in 15 genera. Our results highlight many of the impediments to inferring phylogenies in recent rapid radiations, including low variation at all five nuclear markers, and conflicting relationships supported by mitochondrial and nuclear trees. However, concatenated Bayesian and likelihood analyses, and a multilocus coalescent tree, recovered concordant support for primary clades and several previously unresolved inter-specific groupings. The Aipysurus group is monophyletic, with egg-eating specialists forming separate, early-diverging lineages. All three monotypic semi-aquatic genera (Ephalophis, Parahydrophis and Hydrelaps) are robustly placed as early diverging lineages along the branch leading to the Hydrophis group, with Ephalophis recovered as sister to Parahydrophis. The molecular phylogeny implies extensive evolutionary convergence in feeding adaptations within the Hydrophis group, especially the repeated evolution of small-headed (microcephalic) forms. Microcephalophis (Hydrophis) gracilis is robustly recovered as a relatively distant sister lineage to all other sampled Hydrophis group species, here termed the ‘core Hydrophis group’. Within the ‘core Hydrophis group’, Hydrophis is recovered as broadly paraphyletic, with several other genera nested within it (Pelamis, Enhydrina, Astrotia, Thalassophina, Acalyptophis, Kerilia, Lapemis, Disteira). Instead of erecting multiple new genera, we recommend dismantling the latter (mostly monotypic) genera and recognising a single genus, Hydrophis Latreille 1802, for the core Hydrophis group. Estimated divergence times suggest that all Hydrophiini last shared a common ancestor ～6 million years ago, but that the majority of extant lineages diversified over the last ～3.5 million years. The core Hydrophis group is a young and rapidly speciating clade, with 26 sampled species and 9 genera and dated at only ～1.5–3 million years old.     

Alteromonas flava sp. nov. and Alteromonas facilis sp. nov., two novel copper tolerating bacteria isolated from a sea cucumber culture pond in China

Two bacterial strains, P0211^T and P0213^T, were isolated from a sea cucumber culture pond in China. The strains were able to resist high copper levels. These two strains were characterized at the phenotypic, chemotaxonomic, and genomic level. They were completely different colors, but the 16S rRNA genes showed 99.30% similarity. Phylogenetic analysis based on the sequences of the 16S rRNA gene and five housekeeping genes (dnaK, sucC, rpoB, gyrB, and rpoD) supported the inclusion of these strains within the genus Alteromonas, and the two isolated strains formed a group separated from the closest species Alteromonas aestuariivivens KCTC 52655^T. Genomic analyses, including average nucleotide identity (ANIb and ANIm), DNA–DNA hybridization (DDH), and the percentage of conserved proteins (POCP), clearly separated strains P0211^T and P0213^T from the other species within the genus Alteromonas with values below the thresholds for species delineation. The chemotaxonomic features (including fatty acid and polar lipid analysis) of strains P0211^T and P0213^T also confirmed their differentiation from the related taxa.The results demonstrated that strains P0211^T and P0213^T represented two novel species in the genus Alteromonas, for which we propose the names Alteromonas flava sp. nov., type strain P0211^T (= KCTC 62078^T = MCCC 1H00242^T), and Alteromonas facilis sp. nov., type strain P0213^T (= KCTC 62079^T = MCCC 1H00243^T).     

Mitochondrial DNA based phylogeny of the woodpecker genera Colaptes and Piculus,and implications for the history of woodpecker diversification in South America

The woodpecker genus Colaptes (flickers) has its highest diversity in South America and the closely related genus Piculus is restricted to South and Central America. Two species of flickers occur in North America, and one species is endemic to Cuba. We conducted a Bayesian phylogenetic analysis of three mitochondrial encoded genes (cyt b, COI, 12S rRNA) and confirmed that the two genera are paraphyletic. Three species historically classified as Piculus are actually flickers. We found that the Cuban endemic C. fernandinae is the most basal species within the flickers and that the Northern Flicker is the next most basal species within the Colaptes lineage. The South American clade is most derived. The age of the South American diversification is estimated to be 3.6 MY, which is synchronous with the emergence of the Isthmus of Panama. The pattern of diversification of South American flickers is common among South American woodpeckers. Although woodpeckers have their greatest diversity in South America, we hypothesize that woodpeckers (Family Picidae) originated in Eurasia, dispersed to North America via the Bering land bridge, and multiple lineages entered South America as the Isthmus approached its final closing.     

Biogeographic origin and radiation of Cuban Eleutherodactylus frogs of the auriculatus species group,inferred from mitochondrial and nuclear gene sequences

We studied phylogenetic relationships of the Eleutherodactylus auriculatus species group to infer colonization and diversification patterns in this endemic radiation of terrestrial frogs of the genus Eleutherodactylus in the largest of the Greater Antilles, Cuba. An initial screening of genetic diversity based on partial sequences of the 16S rRNA gene in almost 100 individuals of all species of the group and covering the complete known geographic range of their occurrence found most species endemic to small ranges in the eastern Cuban mountains while a single species was widespread over most of Cuba. Our molecular phylogeny, based on 3731 bp of four mitochondrial and one nuclear gene, suggests that most cladogenetic events within the group occurred among clades restricted to the eastern mountains, which acted as refugia and facilitated the diversification in this group. Our results reveal two separate colonization events of Central and Western Cuba and allow inferring the timing of the subsequent diversification events that occurred between 11 and 2 Mya. Because populations previously assigned to E. auriculatus represent four genetically strongly divergent lineages that also differ in their advertisement calls, we propose that E. auriculatus as currently recognized comprises four species. The difficulties in assigning the name auriculatus to any of these four species, and the fact that E. principalis is nested within one of them, stress the need for a thorough taxonomic revision of this group.     

Thaumasiovibrio occultus gen. nov. sp. nov. and Thaumasiovibrio subtropicus sp. nov. within the family Vibrionaceae,isolated from coral reef seawater off Ishigaki Island,Japan

Two phylogenetically distinct Vibrionaceae strains C4II189^T and C4V358^T isolated from reef seawater off Ishigaki Island, Japan, in 2014 were studied with advanced genome-based taxonomy approaches. All aspects of phylogenetic (16S rRNA phylogeny, MLSA), phenotypic and genetic (ANI, DDH, AAI, and the number of core genes) cohesions between the two identified species were high enough to propose them as members of a new genus within the family Vibrionaceae. Consequently, an eighth genus Thaumasiovibrio gen. nov. is proposed that contains two new species Thaumasiovibrio occultus sp. nov. strain C4II189^T (=DSM 101554^T = JCM 31629^T) (type species) and Thaumasiovibrio subtropicus sp. nov. strain C4V358^T (=DSM 101555^T = JCM 31630^T). Thaumasiovibrio species were phylogenetically distinct from the other Vibrionaceae species based on pyrH gene sequences. The combination of catalase negative, sensitivity to vibriostatic agent O/129, and green colony formation on TCBS for the phylogenetically affiliated strains was the diagnostic features for the current tentative identification of this genus.     

Characterization of Acinetobacter chengduensis sp. nov., isolated from hospital sewage and capable of acquisition of carbapenem resistance genes

Two strains of the genus Acinetobacter, WCHAc060005^T and WCHAc060007, were isolated from hospital sewage in China. The two strains showed different patterns of resistance to clinically important antibiotics and their taxonomic positions were investigated. Cells are Gram-negative, obligate aerobic, non-motile, catalase-positive and oxidase-negative coccobacilli. A preliminary analysis based on the 16S rRNA gene sequences indicated that the two strains had the highest similarity to Acinetobacter cumulans WCHAc060092^T (99.02%). Whole-genome sequencing of the two strains and genus-wide phylogeny reconstruction based on a set of 107 Acinetobacter core genes indicated that they formed a separate and internally cohesive clade within the genus. The average nucleotide identity based on BLAST and in silico DNA–DNA hybridization values between the two new genomes were 99.77% and 98.7% respectively, whereas those between the two genomes and the known Acinetobacter species were <88.93% and <34.0%, respectively. A total of 7 different genes were found in the two genome sequences which encode resistance to five classes of antimicrobial agents, including clinically important carbapenems, oxyimino-cephalosporins, and quinolones. In addition, the combination of their ability to assimilate gentisate, but not l-glutamate and d,l-lactate could distinguish the two strains from all known Acinetobacter species. Based on these combined data, we concluded that the two strains represent a novel species of the genus Acinetobacter, for which the name Acinetobacter chengduensis sp. nov. is proposed. The type strain is WCHAc060005^T (CCTCC AB 2019139 = GDMCC 1.1622 = JCM 33509).     

Dysgonomonas hofstadii sp. nov., isolated from a human clinical source

A Gram-negative staining, facultative anaerobic, cocco-bacillus-shaped organism was isolated from a post-operative abdominal wound. Based on morphological and biochemical criteria, strain MX 1040 ( = CCUG 54731^T) was tentatively identified as Bacteroidaceae but did not correspond to any recognized species of this family. Comparative 16S rRNA gene sequencing analysis demonstrated the organism to be related to species of the genus Dysgonomonas, although sequence divergence values of >5% with the other members of this genus demonstrated the organism to represent a novel species. Phylogenetic analysis revealed the novel organism to be most closely related to Dysgonomonas gadei. The major long-chain cellular fatty acids of the novel species consisted of iso-C_14:0, anteiso-C_15:0, C_16:0, and iso-C_16:0. Based on the phenotypic criteria and phylogenetic considerations, it is proposed that strain MX 1040 from a human clinical source represents a new species of the genus Dysgonomonas, as Dysgonomonas hofstadii sp. nov. The type strain of D. hofstadii is CCUG 54731^T ( = CCM 7606^T).     

Agrobacterium bohemicum sp. nov. isolated from poppy seed wastes in central Bohemia

Two non-pathogenic strains R89-1 and R90^T isolated from poppy seed (Papaver somniferum L.) wastes were phenotypically and genotypically characterized. Multilocus sequence analysis (MLSA) was conducted with six genes (atpD, glnA, gyrB, recA, rpoB, 16S rRNA). The strains represented a new species which clustered with Agrobacterium rubi NBRC 13261^T and Agrobacterium skierniewicense Ch11^T type strains. MLSA was further accompanied by whole-genome phylogeny, in silico DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) analyses for both strains. ANI and dDDH values were deep below the species delineation threshold. Phenotypic features of the novel strains unequivocally allowed their differentiation from all other Agrobacterium species. Unlike other agrobacteria, the strains were salt sensitive and were able to biotransform morphine alkaloids. The dominant cellular fatty acids are 18:1 w7c, 16:0 and 12:0 aldehyde/16:1 iso I/14:0 3OH summed in feature 2 and the major respiratory quinine is Q-10 (87%). The DNA G + C content is 56 mol%. Microbial community analysis indicated probable association with P. somniferum plant material. Altogether, these characteristics showed that strains R90^T and R89-1 represent a new species of the genus Agrobacterium which we propose to name Agrobacterium bohemicum. The type strain of A. bohemicum is R90^T (=CCM 8736^T = DSM 104667^T).     

Pantoea endophytica sp. nov., novel endophytic bacteria isolated from maize planting in different geographic regions of northern China

Four endophytic bacterial strains were isolated from root, stem and leaf of maize planted in different regions of northern China. The four strains possessed almost identical 16S rRNA gene sequences. However, REP-PCR fingerprint patterns discriminated that they were not from one clonal origin. Furthermore, the average nucleotide identity (ANI) values among them were higher than 95%, suggesting they all belong to one species. Based on 16S rRNA gene phylogeny, the four strains were clustered together with Pantoea rodasii LMG 26273^T and Pantoea rwandensis LMG 26275^T, but on a separate branch. Multilocus sequence analysis (MLSA) indicated that the four strains form a novel Pantoea species. Authenticity of the novel species was confirmed by ANI comparisons between strain 596^T and its closest relatives, since obtained values were considerably below the proposed thresholds for the species delineation. The genome size of 596^T was 5.1Mbp, comprising 4896 predicted genes with DNA G + C content of 57.8 mol%. The respiratory quinone was ubiquinone-8 (Q-8) and the polar lipid profile consisted of phosphatidylethanolamin, diphosphatidylglycerol, phosphatidylglycerol, unidentified aminophospholipid and unidentified phospholipid. The major fatty acids of strain 596^T were C_16:0, summed feature 2 (C_12:0 aldehyde), summed feature 3 (C_16:1ω7c and/or C_16:1ω6c) and summed feature 8 (C_18:1ω7c and/or C_18:1ω6c). Based on phylogenetic, genomic, phenotypic and chemotaxonomic data, the four isolates are considered to represent a novel species of the genus Pantoea, for which the name Pantoea endophytica sp. nov., is proposed, with 596^T (= DSM 100,785^T = CGMCC 1.15280^T) as type strain.     

Morphological and molecular characterization of the name-bearing type species Rimaleptus binucleatus (Kahl, 1931), with a phylogenetic re-analysis of dileptid evolutionary history (Ciliophora: Litostomatea: Rhynchostomatia)

We collected Rimaleptus binucleatus from soil in the surroundings of the town of Ulsan, South Korea. Its morphology and 18S rRNA gene were studied using standard methods. This possibly widely distributed species is characterized by: (i) a size of about 170–400 × 20–65 μm; (ii) a narrowly to cylindrically dileptid body with proboscis occupying about 30–65% of body length; (iii) two dorsal contractile vacuoles; (iv) two size groups of rod-shaped extrusomes; and (v) about 18–29 ciliary rows, 4–6 of them anteriorly differentiated into a staggered dorsal brush. Phylogenetic analyses of five new rhynchostomatian 18S rRNA gene sequences supported monophylies of the orders Tracheliida and Dileptida, but revealed that the genera Rimaleptus and Pseudomonilicaryon are polyphyletic. Monophyly of genera with two macronuclear nodules was consistently rejected, but monophylies of dileptids with many scattered macronuclear nodules and of dileptids with moniliform macronucleus and multi-rowed dorsal brush could not be excluded by statistical topology tests. Nevertheless, phylogenetic network analyses indicated considerable conflict in the phylogenetic signal provided by the 18S rRNA gene to resolve unambiguously relationships among dileptid genera.