Diversity of Bacterial Microbiota of Coastal Halophyte Limonium sinense and Amelioration of Salinity Stress Damage by Symbiotic Plant Growth-Promoting Actinobacterium Glutamicibacter halophytocola KLBMP 5180.

Plant-associated microorganisms are considered a key determinant of plant health and growth. However, little information is available regarding the composition and ecological function of the roots' and leaves' bacterial microbiota of halophytes. Here, using both culture-dependent and culture-independent techniques, we characterized the bacterial communities of the roots and leaves as well as the rhizosphere and bulk soils of the coastal halophyte Limonium sinense in Jiangsu Province, China. We identified 49 representative bacterial strains belonging to 17 genera across all samples, with Glutamicibacter as the most dominant genus. All Glutamicibacter isolates showed multiple potential plant growth promotion traits and tolerated a high concentration of NaCl and a wide pH range. Interestingly, further inoculation experiments showed that the Glutamicibacter halophytocola strain KLBMP 5180 isolated from root tissue significantly promoted host growth under NaCl stress. Indeed, KLBMP 5180 inoculation increased the concentrations of total chlorophyll, proline, antioxidative enzymes, flavonoids, K+, and Ca2+ in the leaves; the concentrations of malondialdehyde (MDA) and Na+ were reduced. A transcriptome analysis identified 1,359 and 328 differentially expressed genes (DEGs) in inoculated seedlings treated with 0 and 250 mM NaCl, respectively. We found that pathways related to phenylpropanoid and flavonoid biosynthesis and ion transport and metabolism might play more important roles in host salt stress tolerance induced by KLBMP 5180 inoculation compared to that in noninoculated leaves. Our results provide novel insights into the complex composition and function of the bacterial microbiota of the coastal halophyte L. sinense and suggest that halophytes might recruit specific bacteria to enhance their tolerance of harsh environments.IMPORTANCE Halophytes are important coastal plants often used for the remediation of saline coastal soils. Limonium sinense is well known for its medical properties and phytoremediation of saline soils. However, excessive exploitation and utilization have made the wild resource endangered. The use of endophytic and rhizosphere bacteria may be one of the suitable ways to solve the problem. This study was undertaken to develop approaches to improve the growth of L. sinense using endophytes. The application of actinobacterial endophytes ameliorated salt stress damage of the host via complex physiological and molecular mechanisms. The results also highlight the potential of using habitat-adapted, symbiotic, indigenous endophytic bacteria to enhance the growth and ameliorate abiotic stress damage of host plants growing in special habitats.

Synthesis of biocontrol macromolecules by derivative of chitosan with surfactin and antifungal evaluation.

A derivative of chitosan was prepared with chitosan and ß-cyclodextrins, which was synthesized by the immobilization reaction, as a carrier to adsorb surfactin produced from Bacillus amyloliquefaciens and got biological macromolecules. The antifungal activity against three sapstain fungi by a combination of macromolecules was tested. The results showed that the macromolecules inhibited the mycelium growth of sapstain fungi Lasiodiplodia rubropurpurea, L. crassispora, and L. theobromae by about 73.22%, 76.72%, and 70.22%, respectively. The macromolecules were relatively thermally stable with more than 50% of the antifungal activity even after being held at 121°C for 30 min. Meanwhile, the activity of the macromolecules remained more than 55% at a pH value ranging from 4 to 12. The macromolecules were resistant to hydrolysis by most protein-denaturing detergents and other enzymes. The results indicated the macromolecules might provide an alternative bioresource for the bio-control of sapstain.

Saccharopolyspora dendranthemae sp. nov. a halotolerant endophytic actinomycete isolated from a coastal salt marsh plant in Jiangsu, China.

A halotolerant actinomycete strain, designated strain KLBMP 1305(T), was isolated from a salt marsh plant Dendranthema indicum (Linn.) Des Moul collected from the coastal region of Nantong, Jiangsu Province, in east China and was studied in detail for its taxonomic position. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain KLBMP 1305(T) is a member of the genus Saccharopolyspora. The 16S rRNA gene sequence similarity indicated that strain KLBMP 1305(T) was most closely related to 'Saccharopolyspora pathumthaniensis' S582(T) (99.31 %), 'Saccharopolyspora endophytica' YIM 61095(T) (99.17 %) and Saccharopolyspora tripterygii YIM 65359(T) (99.15 %); similarity to other type strains of the genus Saccharopolyspora was <97.2 %. The organism had chemical and morphological features consistent with its classification in the genus Saccharopolyspora such as meso-diaminopimelic acid as the diagnostic diamino acid in the cell wall peptidoglycan and arabinose and galactose as the diagnostic sugars. The predominant menaquinone was MK-9(H4). The polar lipids detected were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, an unknown glycolipid and an unknown lipid. The major fatty acids were iso-C16:0, iso-C15:0, anteiso-C15:0, anteiso-C17:0 and sum in feature 8 (18:1ω7c/18:1ω6c). The G+C content of the genomic DNA of the type strain was 68.7 mol%. DNA-DNA relatedness data, together with phenotypic differences, clearly distinguished the isolate from its closest relatives. On the basis of these phenotypic and genotypic data, the isolate represents a novel species, for which the name Saccharopolyspora dendranthemae sp. nov. is proposed. The type strain is KLBMP 1305(T) (=KCTC 19889(T) = NBRC 108675(T)).

Streptomyces halophytocola sp. nov., an endophytic actinomycete isolated from the surface-sterilized stems of a coastal halophyte Tamarix chinensis Lour.

A novel actinomycete, designated KLBMP 1284(T), was isolated from the surface-sterilized stems of a coastal halophyte Tamarix chinensis Lour. collected from the city of Nantong, Jiangsu Province, east China. The strain was found to have morphological and chemotaxonomic characteristics typical of members of the genus Streptomyces. Analysis of the 16S rRNA gene sequence of strain KLBMP 1284(T) revealed that the strain formed a distinct clade within the phylogenetic tree based on 16S rRNA gene sequences and the highest sequence similarity (99.43 %) was to Streptomyces sulphureus NRRL B-1627(T). 16S rRNA gene sequence similarity to other species of the genus Streptomyces was lower than 97 %. Based on DNA-DNA hybridization values and comparison of morphological and phenotypic data, KLBMP 1284(T) could be distinguished from the closest phylogenetically related species, Streptomyces sulphureus NRRL B-1627(T). Thus, based on these data, it is evident that strain KLBMP 1284(T) represents a novel species of the genus Streptomyces, for which the name Streptomyces halophytocola sp. nov. is proposed. The type strain is KLBMP 1284(T) (= KCTC 19890(T) = NBRC 108770(T)).

Modestobacter roseus sp. nov., an endophytic actinomycete isolated from the coastal halophyte Salicornia europaea Linn., and emended description of the genus Modestobacter.

A novel actinomycete, designated strain KLBMP 1279(T), was isolated from surface-sterilized roots of a coastal halophyte, Salicornia europaea Linn., collected from Jiangsu Province, in the east of China. The taxonomic status of this organism was established using a polyphasic approach. 16S rRNA gene sequence analysis indicated that strain KLBMP 1279(T) was closely related to Modestobacter marinus 42H12-1(T) (99.5% 16S rRNA gene sequence similarity), Modestobacter versicolor CP153-2(T) (98.4%) and Modestobacter multiseptatus AA-826(T) (97.5%). Chemotaxonomic characteristics were consistent with its assignment to the genus Modestobacter in that the isolate had meso-diaminopimelic acid as the diagnostic diamino acid in the cell wall, MK-9(H4) as major menaquinone and a polar lipid profile containing diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannosides, two unknown aminophospholipids and an unknown phospholipid. The predominant fatty acids were iso-C16:0, iso-C15:0 and C17:1ω8c. The DNA G+C content was 71.7 mol%. However, DNA-DNA hybridization assays as well as physiological and biochemical analyses differentiated strain KLBMP 1279(T) from its closest phylogenetic relatives. On the basis of phenotypic, chemotaxonomic and phylogenetic evidence, the isolate KLBMP 1279(T) represents a novel species of the genus Modestobacter, for which the name Modestobacter roseus sp. nov. is proposed; the type strain is KLBMP 1279(T) (=KCTC 19887(T)=NBRC 108673(T)=DSM 45764(T)). An emended description of the genus Modestobacter is also proposed.

Amycolatopsis jiangsuensis sp. nov., a novel endophytic actinomycete isolated from a coastal plant in Jiangsu, China.

A novel actinomycete, designated strain KLBMP 1262(T), was isolated from a coastal plant Dendranthema indicum (Linn.) Des Moul collected from the coastal region of Nantong, Jiangsu Province, in east China and was studied in detail for its taxonomic position. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain KLBMP 1262(T) is a member of the genus Amycolatopsis. The 16S rRNA gene sequence similarity indicated that strain KLBMP 1262(T) is most closely related to Amycolatopsis sulphurea DSM 46092(T) (97.96 %), Amycolatopsis ultiminotia RP-AC36(T) (97.50 %) and Amycolatopsis jejuensis N7-3(T) (97.44 %); similarity to other type strains of the genus Amycolatopsis was less than 97.0 %. The organism was determined to have chemical and morphological features consistent with its classification in the genus Amycolatopsis such as meso-diaminopimelic acid as the diagnostic diamino acid in the cell wall peptidoglycan and arabinose and galactose as the diagnostic sugars. The predominant menaquinone was determined to be MK-9 (H(4)). The polar lipids detected were phosphatidylmethylethanolamine, phosphatidylethanolamine, an unknown aminophospholipid, two unknown glycolipids and several unknown lipids. The major fatty acids were found to be C(16:0), iso-C(16:0) and iso-C(15:0). DNA-DNA relatedness data, together with phenotypic differences, clearly distinguished the isolate from its closest relatives. On the basis of these phenotypic and genotypic data, the isolate is considered to represent a novel species, for which the name A. jiangsuensis sp. nov. is proposed. The type strain is KLBMP 1262(T) (=KCTC 19885 (T) = NBRC 108679(T)).

Pseudonocardia nantongensis sp. nov., a novel endophytic actinomycete isolated from the coastal halophyte Tamarix chinensis Lour.

A novel isolate, designated strain KLBMP 1282(T) was isolated from the surface-sterilized leaves of a coastal halophyte Tamarix chinensis Lour., collected from Nantong, Jiangsu Province, east of China. Phylogenetic analysis based on 16S rRNA gene sequences revealed that this strain belongs to the genus Pseudonocardia, being most closely related to Pseudonocardia kongjuensis LM 157(T) (98.33 %), Pseudonocardia autotrophica IMSNU 20050(T) (97.77 %), Pseudonocardia endophytica YIM 56035(T) (97.63 %), Pseudonocardia ammonioxydans H9 (T) (97.62 %) and Pseudonocardia compacta IMSNU 20111(T) (97.56 %); similarity to other type strains of the genus Pseudonocardia was <97.5 %. Chemotaxonomic data confirmed the affiliation of strain KLBMP 1282(T) to the genus Pseudonocardia. Strain KLBMP 1282(T) contained MK-8(H(4)) as the predominant ubiquinone and iso-C(16:0) as the major fatty acid. The polar lipids detected in strain KLBMP 1282(T) were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannosides, one unknown phospholipid and four unknown glycolipids. The DNA G + C content of strain KLBMP 1282(T) was 73.1 mol %. The results of DNA-DNA hybridizations and the phylogenetic analysis, together with the phenotypic and biochemical tests, allowed the differentiation of strain KLBMP 1282(T) from strains of other recognized Pseudonocardia species. Therefore, strain KLBMP 1282(T) represents a novel species of the genus Pseudonocardia, for which the name Pseudonocardia nantongensis sp. nov. is proposed. The type strain is KLBMP 1282(T) (=KCTC 29053(T) = NBRC 108677(T)).

Nocardioides panzhihuaensis sp. nov., a novel endophytic actinomycete isolated from medicinal plant Jatropha curcas L.

A novel Gram-positive, aerobic, rod-shaped and mycelia-producing bacterial strain, designated KLBMP 1050(T), was isolated from the stem of the oil-seed plant Jatropha curcas L. collected from Sichuan Province, south-west China. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the isolate KLBMP 1050(T) belonged to the genus Nocardioides, with the highest sequence similarity to Nocardioides albus KCTC 9186(T) (99.38 %) and Nocardioides luteus KCTC 9575(T) (99.03 %). However, the DNA-DNA relatedness of isolate KLBMP 1050(T) to these two type strains were 37.5 ± 3.5 and 33 ± 2.3 %, respectively. Strain KLBMP 1050(T) grew at the pH range 6-11, temperature range 10-32 °C and with 0-12 % NaCl. The physiological properties of strain KLBMP 1050(T) differ from those of N. albus KCTC 9186(T) and N. luteus KCTC 9575(T). The cell-wall peptidoglycan contained LL: -diaminopimelic acid and MK-8(H(4)) was the major respiratory quinone. The predominant cellular fatty acid of strain KLBMP 1050(T) was iso-C(16:0) (23.3 %). The total DNA G+C content was 70.1 mol%. On the basis of the phenotypic, chemotaxonomic and phylogenetic data, strain KLBMP 1050(T) represents a novel species of the genus Nocardioides, for which the name Nocardioides panzhihuaensis sp. nov. is proposed. The type strain is KLBMP 1050(T) (= KCTC 19888(T) = NBRC 108680(T)).

Streptomyces phytohabitans sp. nov., a novel endophytic actinomycete isolated from medicinal plant Curcuma phaeocaulis.

A novel actinomycete, designated strain KLBMP 4601(T), was isolated from the root of the medicinal plant Curcuma phaeocaulis collected from Sichuan Province, south-west China. The strain produced extensively branched substrate and aerial hyphae that carried straight to flexuous spore chains. Chemotaxonomic properties of this strain were consistent with those of members of the genus Streptomyces. The cell wall of strain KLBMP 4601(T) contained LL-diaminopimelic acid as the characteristic diamino acid. The major menaquinone was MK-9(H(4)), with minor amounts of MK-9(H(6)), MK-9(H(8)) and MK-10(H(2)). The major fatty acids were C(16:0), iso-C(16:0), C(18:1)ω9c and C(16:1), iso G. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain KLBMP 4601(T) belongs to the genus Streptomyces and is most closely related to Streptomyces armeniacus JCM 3070(T) (97.9 %), Streptomyces pharmamarensis PM267(T) (97.6 %) and Streptomyces artemisiae YIM 63135(T) (97.5 %). The 16S rRNA gene sequence similarity between strain KLBMP 4601(T) and other members of this genus were lower than 97.5 %. DNA-DNA hybridization studies of strain KLBMP 4601(T) with the three closest species showed relatedness values of 36.3 ± 4.2 %, 27.3 ± 0.6 %, and 30.9 ± 2.5 %, respectively. On the basis of chemotaxonomic, phenotypic and genotypic characteristics, it is evident that strain KLBMP 4601(T) represents a novel species of the genus Streptomyces, for which the name Streptomyces phytohabitans sp. nov. is proposed. The type strain is KLBMP 4601(T) (=KCTC 19892(T) = NBRC 108772(T)).

Kibdelosporangium phytohabitans sp. nov., a novel endophytic actinomycete isolated from oil-seed plant Jatropha curcas L. containing 1-aminocyclopropane-1-carboxylic acid deaminase.

A novel actinomycete, designated strain KLBMP 1111(T), was isolated from the root of the oil-seed plant Jatropha curcas L. collected from Sichuan Province, south-west China. Strain KLBMP 1111(T) formed a distinct branch in the 16S rRNA gene phylogenetic tree together with the type strains in the genus Kibdelosporangium, with the highest similarity to Kibdelosporangium aridum subsp. aridum DSM 43828(T) (98.8%), K. aridum subsp. largum DSM 44150(T) (98.1%) and Kibdelosporangium philippinense DSM 44226(T) (98.1%). The organism produced sporangium-like structures, the typical morphological characteristic of the genus Kibdelosporangium. The chemotaxonomic properties of this strain were also consistent with those of the genus Kibdelosporangium: the peptidoglycan contained meso-diaminopimelic acid; the predominant menaquinone was MK-9(H(4)); phospholipids were phosphatidylglycerol, phosphatidylethanolamine, phosphatidylmethylethanolamine, phosphatidylinositol and an unknown phospholipid; iso-C(16:0), C(16:0), anteiso-C(15:0) and iso-C(15:0) as the predominant cellular fatty acids and the G+C content was 67.2 mol%. DNA-DNA hybridization values between strain KLBMP 1111(T) and the three Kibdelosporangium species were less than 50%. This strain had the ability to produce a siderophore, utilized 1-aminocyclopropane-1-carboxylic acid (ACC) as sole source of nitrogen and possessed ACC deaminase enzyme. Based on genotypic and phenotypic data, strain KLBMP 1111(T) represents a novel species in the genus Kibdelosporangium. We propose the name Kibdelosporangium phytohabitans sp. nov. for this species. The type strain is the strain KLBMP 1111(T) (=KCTC 19775(T) = CCTCC AA 2010001(T)).

Amycolatopsis endophytica sp. nov., a novel endophytic actinomycete isolated from oil-seed plant Jatropha curcas L.

A novel actinomycete, designated KLBMP 1221(T), was isolated from the surface-sterilized seeds of an oil-seed plant Jatropha curcas L. collected from Sichuan Province, south-west China and was characterized taxonomically by using a polyphasic approach. Phylogenetic analyses based on 16S rRNA gene sequence showed that this strain formed a distinct phyletic line within the radiation of the genus Amycolatopsis. The 16S rRNA gene sequence similarity indicated that strain KLBMP 1221(T) was most closely related to Amycolatopsis eurytherma NT202(T) (98.9%), Amycolatopsis tucumanensis ABO(T) (98.8%), Amycolatopsis thermoflava N1165(T) (98.6%) and Amycolatopsis methanolica IMSNU 20055(T) (98.5%). Strain KLBMP 1221(T) had morphological and chemotaxonomic properties that were consistent with its classification in the genus Amycolatopsis. However, DNA-DNA relatedness data and phenotypic differences clearly distinguished the isolate from its closest relatives. Based on the combined genotypic and phenotypic evidence, it is proposed that strain KLBMP 1221(T) be classified as representative of a novel species for which the name Amycolatopsis endophytica sp. nov. is proposed. The type strain is KLBMP 1221(T) (= KCTC 19776(T) = CCTCC AA 2010003(T)).