Responses of rhizosphere bacterial communities, their functions and their network interactions to Cd stress under phytostabilization by Miscanthus spp.

Miscanthus has good tolerance to heavy metals (HMs) and has received increasing attention in studies of HM-contaminated soil remediation. In this study, four Miscanthus cultivars (M. lutarioriparius Xiangnadi NO4, M. sinensis Xiangmang NO1, M. lutarioriparius × M. sinensis hybrid Xiangzamang NO1, and M. floridulus Wujiemang NO1) that grow in China were studied. Their tolerance and enrichment abilities in soils containing 50 mg kg-1 cadmium (Cd) and the structure and function of their rhizosphere bacterial communities during the remediation process were analyzed. The results exhibiting a tolerance index (TI) higher than 75 in roots and the aboveground parts (TI > 60, indicating highly tolerant plants) indicated that all four Miscanthus cultivars were tolerant to high Cd concentrations. Moreover, Cd was mainly enriched in roots, the translocation ability from roots to aboveground parts was weak, and the four cultivars exhibited phytostabilization ability in Cd-contaminated soils. High-throughput sequencing (HTS) analysis showed that the Miscanthus rhizosphere bacterial community comprised 33 phyla and 446 genera, including plant growth-promoting rhizobacteria (PGPRs), such as Bacillus, Sphingomonas, and Mesorhizobium. The addition of Cd affected the Miscanthus rhizosphere bacterial community and reduced community diversity. Phylogenetic molecular ecological networks (pMENs) indicated that Cd addition reduced interactions between Miscanthus rhizosphere bacteria and thereby led to a simpler network structure, increased the number of negative-correlation links, enhanced the competition between rhizosphere bacterial species, reduced the number of key bacteria, and changed the composition of those bacteria. PICRUSt functional predictive analysis indicated that Cd stress reduced soil bacterial functions in the Miscanthus rhizosphere. The results of this study provide a basis for the remediation of Cd-contaminated soils by Miscanthus and provide a reference for the subsequent regulation of Miscanthus remediation efficiency by PGPRs or key bacteria.

Leucobacter triazinivorans sp. nov., a s-triazine herbicide prometryn-degrading bacterium isolated from sludge.

A Gram-stain-positive, rod-shaped, non-motile bacterial strain, designated JW-1T, was isolated from activated sludge collected from the outlet of an aeration tank in a prometryn-manufacturing plant, located in Binzhou City, Shandong province, PR China. Phylogenetic analysis, based on 16S rRNA gene sequences, indicated that strain JW-1T belongs to the genus Leucobacter and its closest neighbours are 'Leucobacter kyeonggiensis' F3-P9 (98.95 % similarity), Leucobacter celer subsp. astrifaciens CBX151T (98.62 %), Leucobacter celer subsp. celer NAL101T (98.53 %), Leucobacter chromiiresistens JG31T (97.86 %) and Leucobacter chironomi DSM 19883T (97.37 %). DNA-DNA hybridization values with the above strains were <55 %. The DNA G+C content of strain JW-1T was 72.6 mol%. The major fatty acids of strain JW-1T were iso-C16 : 0, anteiso-C15 : 0, anteiso-C17 : 0 and iso-C15 : 0. The predominant polar lipids were diphosphatidylglycerol, phosphatidylglycerol and glycolipid. The predominant menaquinone was MK-11. The cell wall amino acids were 2,4-diaminobutyric acid, alanine, glutamic acid, glycine and threonine. Based on the molecular and chemotaxonomic data, as well as the physiological and biochemical characteristics, strain JW-1T is considered to represent a novel species of the genus Leucobacter, for which the name Leucobacter triazinivorans is proposed. The type strain is JW-1T (=DSM 105188T=LMG 30083T).

Massilia buxea sp. nov., isolated from a rock surface.

A Gram-stain-negative, rod-shaped and motile bacterial strain, designated A9T, was isolated from the surface of rock collected from the shore of Nvshan lake in Mingguang, Anhui province, China. Phylogenetic analysis based on 16S rDNA sequence data showed that strain A9T was affiliated with the genus Massilia and showed the highest sequence similarities to Massilia plicata KCTC 12344T (98.8 %) and Massilia lurida CGMCC 1.10822T (97.9 %). The major fatty acids (>5 %) were summed feature 3 (C16 : 1ω7c and/or C15 : 0 iso 2-OH), C16 : 0 and C18 : 1ω7c. Strain A9T contained Q-8 as the predominant ubiquinone and diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and an unidentified aminophospholipid as the predominant polar lipids. The DNA G+C content was 69.9 mol%. Mean DNA-DNA relatedness values between strain A9T and its closest phylogenetic relatives, M. plicata KCTC 12344T and M. lurida CGMCC 1.10822T, were 38.8 % and 23.23 %, respectively. On the basis of the results obtained in this study, strain A9T is considered to represent a novel species of the genus Massilia, for which the name Massilia buxea sp. nov. is proposed. The type strain is A9T (=DSM 103547T=CGMCC 1.15931T=KCTC 52429T).

Mucilaginibacter rubeus sp. nov., isolated from rhizosphere soil.

A Gram-stain-negative, aerobic, non-motile, rod-shaped and non-spore-forming bacterium, designated EF23T, was isolated from rhizosphere soil of watermelon. Growth of strain EF23T was observed at 10-37 °C, at pH 5.0-9.0 and in the presence of 0-0.5 % (w/v) NaCl. Strain EF23T contained menaquinone 7 (MK-7) as the major isoprenoid quinone, and summed feature 3 (C16:1ω7c and/or iso-C15 : 0 2-OH), iso-C15 : 0, C16 : 0 and iso-C17 : 0 3-OH as the major fatty acids. Phosphatidylethanolamine was identified as the major polar lipid. The genomic DNA G+C content of strain EF23T was 43.7 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain EF23T was most closely related to Mucilaginibacter gossypii Gh-67T (98.9 % similarity) and Mucilaginibacter gossypiicola Gh-48T (97.6 %). DNA-DNA relatedness values between strain EF23T and M. gossypii KCTC 22380T and M. gossypiicola KCTC 22379T were 31.6 and 53.7 %. On the basis of the evidence presented in this polyphasic taxonomic study, strain EF23T is considered to represent a novel species of the genus Mucilaginibacter, for which the name Mucilaginibacter rubeus sp. nov. is proposed. The type strain is EF23T (=CGMCC 1.15913T=KCTC 52516T).

Caulobacter rhizosphaerae sp. nov., a stalked bacterium isolated from rhizosphere soil.

The Gram-reaction-negative, aerobic, white- to pale-yellow-coloured and rod-shaped bacterium with a single polar flagellum or a stalk, designated strain 7F14T, was isolated from rhizosphere soil of cultivated watermelon (Citrullus lanatus) collected from Hefei, China. Growth of strain 7F14T was observed at pH 6.0-9.0, 10-30 °C and in the presence of 0-1 % (w/v) NaCl. Cells were catalase-negative and oxidase-positive. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain 7F14T formed a phyletic lineage within the genus Caulobacter of the family Caulobacteraceae and showed the highest 16S rRNA gene sequence similarities to Caulobacter henricii ATCC 15253T (98.66 %), Caulobacter segnis ATCC 21756T (98.27 %), Caulobacter vibrioides CB51T (97.92 %) and Caulobacter flavus RHGG3T (97.44 %). The G+C content of the genomic DNA was 68.6 mol%. Strain 7F14T contained Q-10 as the sole ubiquinone and 11-methyl C18 : 1ω7c, C18 : 1ω7c, C16 : 0 and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH) as the major fatty acids. The polar lipids profile consisted of phosphatidylglycerol, an unknown phosphoglycolipid, five unknown glycolipids, an unknown phospholipid and three unknown lipids. DNA-DNA relatedness values to the most closely related type strains Caulobacter henricii DSM 4730T and Caulobacter segnis DSM 7131T were 26.0 and 19.7 %, respectively. Based on unique phenotypic traits, and phylogenetic, chemotaxonomic and DNA-DNA hybridization results, strain 7F14T should be classified as a representative of a novel species of the genus Caulobacter, for which the name Caulobacter rhizosphaerae sp. nov. is proposed. The type strain is 7F14T (=CGMCC 1.15915T=KCTC 52515T).

Complete genome sequence of a novel chlorpyrifos degrading bacterium, Cupriavidus nantongensis X1.

Cupriavidus nantongensis X1 is a chlorpyrifos degrading bacterium, which was isolated from sludge collected at the drain outlet of a chlorpyrifos manufacture plant. It is the first time to report the complete genome sequence of C. nantongensis species, which has been reported as a novel species of Cupriavidus genus. It could provide further pathway information in chlorpyrifos degradation.

Cupriavidus nantongensis sp. nov., a novel chlorpyrifos-degrading bacterium isolated from sludge.

A Gram-stain-negative, aerobic, coccoid to small rod-shaped bacterium, designated X1T, was isolated from sludge collected from the vicinity of a pesticide manufacturer in Nantong, Jiangsu Province, China. Based on 16S rRNA gene sequence analysis, strain X1T belonged to the genus Cupriavidus, and was most closely related to Cupriavidus taiwanensis LMG 19424T (99.1 % 16S rRNA gene sequence similarity) and Cupriavidus alkaliphilus LMG 26294T (98.9 %). Strain X1T showed 16S rRNA gene sequence similarities of 97.2-98.2 % with other species of the genus Cupriavidus. The major cellular fatty acids of strain X1T were C16 : 0, C16 : 1ω7c and/or iso-C15 : 0 2-OH (summed feature 3), C18 : 1ω7c and C17 : 0 cyclo, and the major respiratory quinone was ubiquinone Q-8. The major polar lipids of strain X1T were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, aminophospholipid, phospholipid and hydroxyphosphatidylethanolamine. The DNA G+C content was 66.6 mol%. The DNA-DNA relatedness values of strain X1T with the five reference strains C. taiwanensis LMG 19424T, C. alkaliphilus LMG 26294T, Cupriavidus necator LMG 8453T, Cupriavidus gilardii LMG 5886T and 'Cupriavidus yeoncheonense' KCTC 42053 were lower than 70 %. The results obtained from phylogenetic analysis, phenotypic characterization and DNA-DNA hybridization indicated that strain X1T should be proposed to represent a novel species of the genus Cupriavidus, for which the name Cupriavidus nantongensis sp. nov. is proposed. The type strain is X1T (=KCTC 42909T=LMG 29218T).

Caulobacter flavus sp. nov., a stalked bacterium isolated from rhizosphere soil.

A Gram-stain-negative, aerobic, yellow-pigmented and rod-shaped bacterium with a single polar flagellum or a stalk, designated strain RHGG3T, was isolated from rhizosphere soil of cultivated watermelon (Citrullus lanatus) collected from Hefei, China. Optimal growth of strain RHGG3T was observed at pH 7.0 and 28-30 °C. Cells were catalase-positive and oxidase-negative. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain RHGG3T belonged to the genus Caulobacter and showed the highest 16S rRNA gene sequence similarities to Caulobacter segnis ATCC 21756T (98.6 %), Caulobacter vibrioides CB51T (98.3 %) and Caulobacter henricii ATCC 15253T (97.2 %). The G+C content of the genomic DNA was 70 mol%. Strain RHGG3T contained Q-10 as the sole ubiquinone and the major fatty acids (>8 %) were 11-methyl C18 : 1ω7c, C18 : 1ω7c, C16 : 0, C15 : 0 and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH). The polar lipids were various unknown glycolipids, phosphatidylglycerol and phosphoglycolipids. DNA-DNA relatedness of strain RHGG3T to type strains of the most closely related species (Caulobacter segnis ATCC 21756T, Caulobacter vibrioides DSM 4738 and Caulobacter henricii ATCC 15253T) was 32.4-40.9 %. Based on polyphasic taxonomy analysis (phylogenetic, unique phenotypic traits, chemotaxonomic and DNA-DNA hybridizations), strain RHGG3T represents a novel species of the genus Caulobacter, for which the name Caulobacter flavus sp. nov. is proposed. The type strain is RHGG3T ( = CGMCC 1.15093T = KCTC 42581T = JCM 30763T).

Genetic diversity and characterization of heavy metal-resistant-endophytic bacteria from two copper-tolerant plant species on copper mine wasteland.

The diversity of endophytic bacteria from the copper-tolerant species Elsholtzia splendens and Commelina communis was evaluated by using cultivation and cultivation-independent techniques. Phylogenetic analysis based on 16S rDNA sequences showed that the Cu-resistant isolates belonged to three major phylogenetic groups: Firmicutes, Actinobacteria and Proteobacteria. Bacillus and Acinetobacter were predominant among the Cu-resistant isolates. Sequence analysis from the 16S rDNA clone libraries of the two plant roots revealed sequences of Alpha-, Beta-, Gamma-Proteobacteria, Firmicutes,Actinobacteria and Bacteroidetes. Gammaproteobacteria was predominant in the two 16S rDNA clone libraries. Thirty-two endophytic bacteria were isolated and characterized with respect to heavy metal resistance and production of plant growth-promoting factors. In experiments involving rape plants grown in vermiculite containing 4 mg kg(-1) of Cu, inoculation with the isolates was found to increase the dry weights of roots (ranging from 132% to 155%) and above-ground tissues (ranging from 71% to 83%) compared to the uninoculated control. Increase in above-ground tissue Cu content varied from 63% to 125% in inoculated-rape plants cultivated in Cu-contaminated substrate compared to the uninoculated control.