Streptomyces polyrhachii sp. nov., a novel actinomycete isolated from an edible Chinese black ant (Polyrhachis vicina Roger)

A novel actinomycete, designated strain NEAU-ycm1^T, was isolated from an edible Chinese black ant (Polyrhachis vicina Roger) and characterized with a polyphasic approach. The organism was found to have morphological and chemotaxonomic characteristics typical of streptomycetes. Phylogenetic analysis based on the almost complete 16S rRNA gene sequence show that the novel isolate belongs to the genus Streptomyces and forms a separate subclade. The closest phylogenetic relatives were identified as the type strains of Streptomyces intermedius NBRC 13049^T (97.74 %), Streptomyces aureoverticillatus NRRL B-3326^T (97.69 %), Streptomyces rutgersensis NBRC 12819^T (97.68 %), Streptomyces gougerotii NBRC 3198^T (97.68 %) and Streptomyces diastaticus subsp. diastaticus NBRC 3714^T (97.68 %). Similarities to other type strains of the genus Streptomyces were lower than 97.55 %. A comparison between strain NEAU-ycm1^T and the closest related Streptomyces type strains revealed that it is different from them in morphological, physiological and biochemical characteristics. Therefore, it is proposed that NEAU-ycm1^T (=CGMCC 4.7094^T = DSM 42102^T) represents a novel species of the genus of Streptomyces, for which the name Streptomyces polyrhachii sp. nov. is proposed.     

Endophytic actinomycetes from spontaneous plants of Algerian Sahara: indole-3-acetic acid production and tomato plants growth promoting activity

Twenty-seven endophytic actinomycete strains were isolated from five spontaneous plants well adapted to the poor sandy soil and arid climatic conditions of the Algerian Sahara. Morphological and chemotaxonomical analysis indicated that twenty-two isolates belonged to the Streptomyces genus and the remaining five were non-Streptomyces. All endophytic strains were screened for their ability to produce indole-3-acetic acid (IAA) in vitro on a chemically defined medium. Eighteen strains were able to produce IAA and the maximum production occurred with the Streptomyces sp. PT2 strain. The IAA produced was further extracted, partially purified and confirmed by thin layer chromatography (TLC) analysis. The 16S rDNA sequence analysis and phylogenetic studies indicated that strain PT2 was closely related to Streptomyces enissocaecilis NRRL B 16365^T, Streptomyces rochei NBRC 12908^T and Streptomyces plicatus NBRC 13071^T, with 99.52 % similarity. The production of IAA was affected by cultural conditions such as temperature, pH, incubation period and l-tryptophan concentration. The highest level of IAA production (127 μg/ml) was obtained by cultivating the Streptomyces sp. PT2 strain in yeast extract-tryptone broth supplemented with 5 mg l-tryptophan/ml at pH 7 and incubated on a rotary shaker (200 rpm) at 30 °C for 5 days. Twenty-four-hour treatment of tomato cv. Marmande seeds with the supernatant culture of Streptomyces sp. PT2 that contained the crude IAA showed the maximum effect in promoting seed germination and root elongation.     

Genome Sequence of Kitasatospora setae NBRC 14216T: An Evolutionary Snapshot of the Family Streptomycetaceae

Kitasatospora setae NBRC 14216^T (=KM-6054^T) is known to produce setamycin (bafilomycin B1) possessing antitrichomonal activity. The genus Kitasatospora is morphologically similar to the genus Streptomyces, although they are distinguishable from each other on the basis of cell wall composition and the 16S rDNA sequence. We have determined the complete genome sequence of K. setae NBRC 14216^T as the first Streptomycetaceae genome other than Streptomyces. The genome is a single linear chromosome of 8 783 278 bp with terminal inverted repeats of 127 148 bp, predicted to encode 7569 protein-coding genes, 9 rRNA operons, 1 tmRNA and 74 tRNA genes. Although these features resemble those of Streptomyces, genome-wide comparison of orthologous genes between K. setae and Streptomyces revealed smaller extent of synteny. Multilocus phylogenetic analysis based on amino acid sequences unequivocally placed K. setae outside the Streptomyces genus. Although many of the genes related to morphological differentiation identified in Streptomyces were highly conserved in K. setae, there were some differences such as the apparent absence of the AmfS (SapB) class of surfactant protein and differences in the copy number and variation of paralogous components involved in cell wall synthesis.     

<Emphasis Type="Italic">Streptomyces</Emphasis> sp. strain SK68, isolated from peanut rhizosphere,promotes growth and alleviates salt stress in tomato (<Emphasis Type="Italic">Solanum lycopersicum</Emphasis> cv. Micro-Tom)

A novel actinobacterium, strain SK68, was isolated from the rhizosphere of peanut plant and its salinity stress alleviation ability was studied using tomato (Solanum lycopersicum cv. Micro-Tom) plants. Based on 16S rDNA based phylogenetic analysis, strain SK68 has been identified as a Streptomyces sp. Strain SK68 had branched substrate mycelium bearing smooth surfaced spores and the spore colour is brownish grey on ISP4 medium. It exhibited enzyme activities such as xylanase, cellulase, amylase, and pectinase and degraded hypoxanthine, casein, and L-tyrosine. The strain SK68 differed in its banding pattern in BOX-PCR and RAPD fingerprinting compared to the closely matching type strains Streptomyces erythrochromogenes NBRC 3304^T (AB184746), S. flavotricini NBRC 12770^T (AB184132), S. racemochromogenes NBRC 12906^T (AB184235), and S. polychromogenes NBRC 13072^T (NR041109). Strain SK68 was evaluated for its salinity stress-alleviating activity in tomato plants with 180 mmol/L NaCl under gnotobiotic condition. A significant increase in plant biomass was observed in strain SK68-inoculated tomato plants under salt stress compared to control and salt-stressed non-inoculated plants.     

Genome based analysis of type-I polyketide synthase and nonribosomal peptide synthetase gene clusters in seven strains of five representative Nocardia species

Background

Actinobacteria of the genus Nocardia usually live in soil or water and play saprophytic roles, but they also opportunistically infect the respiratory system, skin, and other organs of humans and animals. Primarily because of the clinical importance of the strains, some Nocardia genomes have been sequenced, and genome sequences have accumulated. Genome sizes of Nocardia strains are similar to those of Streptomyces strains, the producers of most antibiotics. In the present work, we compared secondary metabolite biosynthesis gene clusters of type-I polyketide synthase (PKS-I) and nonribosomal peptide synthetase (NRPS) among genomes of representative Nocardia species/strains based on domain organization and amino acid sequence homology.

Results

Draft genome sequences of Nocardia asteroides NBRC 15531^T, Nocardia otitidiscaviarum IFM 11049, Nocardia brasiliensis NBRC 14402^T, and N. brasiliensis IFM 10847 were read and compared with published complete genome sequences of Nocardia farcinica IFM 10152, Nocardia cyriacigeorgica GUH-2, and N. brasiliensis HUJEG-1. Genome sizes are as follows: N. farcinica, 6.0 Mb; N. cyriacigeorgica, 6.2 Mb; N. asteroides, 7.0 Mb; N. otitidiscaviarum, 7.8 Mb; and N. brasiliensis, 8.9 - 9.4 Mb. Predicted numbers of PKS-I, NRPS, and PKS-I/NRPS hybrid clusters ranged between 4–11, 7–13, and 1–6, respectively, depending on strains, and tended to increase with increasing genome size. Domain and module structures of representative or unique clusters are discussed in the text.

Conclusion

We conclude the following: 1) genomes of Nocardia strains carry as many PKS-I and NRPS gene clusters as those of Streptomyces strains, 2) the number of PKS-I and NRPS gene clusters in Nocardia strains varies substantially depending on species, and N. brasiliensis strains carry the largest numbers of clusters among the species studied, 3) the seven Nocardia strains studied in the present work have seven common PKS-I and/or NRPS clusters, some of whose products are yet to be studied, and 4) different N. brasiliensis strains have some different gene clusters of PKS-I/NRPS, although the rest of the clusters are common within the N. brasiliensis strains. Genome sequencing suggested that Nocardia strains are highly promising resources in the search of novel secondary metabolites.

Electronic supplementary material

The online version of this article (doi: 10.1186/1471-2164-15-323) contains supplementary material, which is available to authorized users.     

Streptomyces erringtonii sp. nov. and Streptomyces kaempferi sp. nov., isolated from a hay meadow soil

Two filamentous actinomycetes isolated from a hay meadow soil were provisionally assigned to the genus Streptomyces based on morphological features. The isolates were found to have chemical and morphological properties typical of the genus Streptomyces and formed distinct phyletic lines in the 16S rRNA gene tree. Isolate I36^T was most closely related to Streptomyces glauciniger NBRC 100913^T and isolate I37^T to Streptomyces mirabilis NBRC 13450^T. Low DNA:DNA relatedness values were recorded between each of the isolates and their closest phylogenetic neighbour. The isolates were also distinguished from their nearest phylogenetic neighbour, and from one another, using a combination of phenotypic properties. These data indicate that the isolates should be recognised as new species in the genus Streptomyces. The names proposed for these new taxa are Streptomyces erringtonii sp. nov. and Streptomyces kaempferi sp. nov. with isolate I36^T (=CGMCC 4.7016^T = KACC 15424^T) and isolate I37^T (=CGMCC 4.7020^T = KACC 15428^T) as the respective type strains.     

Rapid identification of Streptomyces isolates by MALDI-TOF MS

The recent emergence of multidrug-resistant bacteria over the last decade has led to a renewal in the discovery of new antimicrobial drugs. Streptomyces members are practically unlimited sources of new antibiotics. However, the identification of Streptomyces species is difficult and time-consuming. Therefore, there is a need for alternative methods for their rapid identification. In this study, an efficient protocol of identification using Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry (MALDI-TOF MS) was developed and applied for the rapid identification of Streptomyces isolates from the El Kala lakes in northeastern Algeria. A collection of 48 Streptomyces isolates were used for this study. The optimized procedure allowed us to obtain specific and reproducible protein spectra for each Streptomyces isolate tested. The spectra generated were used to build a preliminary local database based on their initial 16S rRNA identification. The blind test used for the identification of 20 Streptomyces strains already available in our created database and 20 unknown Streptomyces isolates showed that all (100%) of the Streptomyces strains listed in the database were rapidly (<30 min) identified with high scores of up to 2.8. Here, for the first time we showed that MALDI-TOF MS could be used as a cost-effective tool for the rapid identification of Streptomyces isolates.     

Environmental DNA Fragment Conferring Early and Increased Sporulation and Antibiotic Production in Streptomyces Species

Here we describe the rep gene, isolated from an environmental DNA library, which when transformed into Streptomyces species resulted in increased production of secondary metabolites and accelerated sporulation. We show that Streptomyces lividans strains bearing rep are particularly useful as expression hosts for heterologous antibiotic production.     

Degradation of Softwood, Hardwood, and Grass Lignocelluloses by Two Streptomyces Strains

Two Streptomyces strains, S. viridosporus T7A and S. setonii 75Vi2, were grown on softwood, hardwood, and grass lignocelluloses, and lignocellulose decomposition was followed by monitoring substrate weight loss, lignin loss, and carbohydrate loss over time. Results showed that both Streptomyces strains substantially degraded both the lignin and the carbohydrate components of each lignocellulose; however, these actinomycetes were more efficient decomposers of grass lignocelluloses than of hardwood or softwood lignocelluloses. In particular, these Streptomyces strains were more efficient decomposers of grass lignins than of hardwood or softwood lignins.     

<Emphasis Type="Italic">Streptomyces shenzhenensis</Emphasis> sp. nov., a novel actinomycete isolated from mangrove sediment

An actinomycete strain, which was designated 172115^T, was isolated from mangrove soil in Shenzhen, China. Strain 172115^T fell within the genus Streptomyces in the 16S rRNA gene tree and could be grouped into this genus based on its chemotaxonomic and morphological data. The strain shared the highest 16S rRNA gene sequence similarities with Streptomyces lanatus NBRC 12787^T (AB184845) (98.29%) and Streptomyces lucensis NBRC 13056^T (AB184280) (98.26%). The DNA–DNA hybridization values between strain 172115^T and the two most closely related type strains were low enough to justify the assignment of the strain to a novel species. On the basis of these phenotypic, phylogenetic and chemotaxonomic characteristics, 172115^T represents a novel species of the genus Streptomyces, for which, the name Streptomyces shenzhenensis sp. nov. is proposed for strain 172115^T (=CCTCC AA 2011001^T=DSM 42034^T).     

Molecular characterization of β-fructofuranosidases from Rhizopus delemar and Amylomyces rouxii

Of the 19 strains of Rhizopus delemar deposited as Rhizopus oryzae, seven of them, NBRC 4726, NBRC 4734, NBRC 4746, NBRC 4754, NBRC 4773, NBRC 4775, and NBRC 4801, completely hydrolyzed exogenous sucrose and fructooligosaccharides. The sucrose-hydrolyzing enzyme was purified from the culture filtrate of R. delemar NBRC 4754 and classified to β-fructofuranosidase, similar to that of Amylomyces rouxii CBS 438.76. Fragments including β-fructofuranosidase genes (sucA) of seven strains of R. delemar and A. rouxii CBS 438.76 were amplified and sequenced by PCR with degenerated primers synthesized on the basis of the internal amino acid sequences of purified enzymes and successive inverse PCR. Nucleotide sequences of the obtained fragments revealed that open reading frames of 1,569 bp have no intron and encode 522 amino acids. The presumed proteins contained the typical domain of the glycoside hydrolase 32 family, including β-fructofuranosidase, inulinase, levanase, and fructosyltransferases. Amino acid sequences of SucA proteins from the seven strains of R. delemar were identical and showed 90.0 % identity with those of A. rouxii CBS 438.76. A dendrogram constructed from these amino acid sequences showed that SucA proteins are more closely related to yeast β-fructofuranosidases than to other fungal enzymes.     

Biocontrol of Rhizoctonia solani damping-off and promotion of tomato plant growth by endophytic actinomycetes isolated from native plants of Algerian Sahara

Thirty-four endophytic actinomycetes were isolated from the roots of native plants of the Algerian Sahara. Morphological and chemical studies showed that twenty-nine isolates belonged to the Streptomyces genus and five were non-Streptomyces. All isolates were screened for their in vitro antifungal activity against Rhizoctonia solani. The six that had the greatest pathogen inhibitory capacities were subsequently tested for their in vivo biocontrol potential on R. solani damping-off in sterilized and non-sterilized soils, and for their plant-growth promoting activities on tomato seedlings. In both soils, coating tomato seeds with antagonistic isolates significantly reduced (P < 0.05) the severity of damping-off of tomato seedlings. Among the isolates tested, the strains CA-2 and AA-2 exhibited the same disease incidence reduction as thioperoxydicarbonic diamide, tetramethylthiram (TMTD) and no significant differences (P < 0.05) were observed. Furthermore, they resulted in a significant increase in the seedling fresh weight, the seedling length and the root length of the seed-treated seedlings compared to the control. The taxonomic position based on 16S rDNA sequence analysis and phylogenetic studies indicated that the strains CA-2 and AA-2 were related to Streptomyces mutabilis NBRC 12800^T (100% of similarity) and Streptomyces cyaneofuscatus JCM 4364^T (100% of similarity), respectively.     

Fatty acid production of thraustochytrids from Saudi Arabian mangroves

This is the first report of thraustochytrids from Saudi Arabia. A total of 108 isolates of thraustochytrid were cultured from Syhat mangroves, Arabian Gulf, Saudi Arabia. Isolated thraustochytrids belonged to five genera: Aplanochytrium, Aurantiochytrium, Schizochytrium, Thraustochytrium and Ulkenia. Cultured thraustochytrids isolated from decaying leaves of Avicennia marina (77 isolates), sediment (15), seawater (10) and decaying thalli of Sargassum (6). Of the 108 isolates, three strains (SY25, SY38 and SY52) were selected based on their high biomass productivity and high percentages of PUFAs. Phylogenetic analyses based on 18S rDNA placed the three strains within the Aurantiochytrium clade with high statistical support. Species of Aurantiochytrium formed six separate clades, the two strains (SY38 and SY52) formed a separate clade that is a sister clade to the one that contains the type species A. limacinum, while SY25 grouped with Aurantiochytrium sp. TA4, that is also isolated from mangroves in Iran, Arabian Gulf. The strains (SY38 and SY52) shared the phylogenetic placement, their morphology and fatty acid profile. The strain SY25 have different shape of sporangia that divide to give zoospores directly, sporogenous cells are surrounded by thick gelatinous sheath and produce high levels of Linoleic and Oleic essential unsaturated fatty acids. The three studied strain produced high levels of Palmitic acid (ranged between 31.1 and 65.3 % of total fatty acids) that can be further optimized for biofuel production.     

Efficient production of ethanol from raw starch by a mated diploid Saccharomyces cerevisiae with integrated α-amylase and glucoamylase genes

The goal of this research was to construct a stable and efficient process for the production of ethanol from raw starch, using a recombinant Saccharomyces cerevisiae, which is productive even under conditions such as non-selection or long-term operation. Three recombinant yeast strains were used, two haploid strains (MT8-1SS and NBRC1440SS) and one diploid strain (MN8140SS). The recombinant strains were constructed by integrating the glucoamylase gene from Rhizopus oryzae fused with the 3′-half of the α-agglutinin gene as the anchor protein, and the α-amylase gene from Streptococcus bovis, respectively, into their chromosomal DNA by homologous recombination. The diploid strain MN8140SS was constructed by mating these opposite types of integrant haploid strains in order to enhance the expression of integrated amylase genes. The diploid strain had the highest ethanol productivity and reusability during fermentation from raw starch. Moreover, the ethanol production rate of the integrant diploid strain was maintained when batch fermentation was repeated three times (0.67, 0.60, and 0.67 g/l/h in each batch). These results clearly show that a diploid strain developed by mating two integrant haploid strains is useful for the establishment of an efficient ethanol production process.     

Molecular and culture dependent characterization of endolithic bacteria in two beach sand samples and description of Rhizobium endolithicum sp. nov.

The taxonomic position of a streptomycete isolated from a potato tubercle was determined by using a polyphasic approach. The organism had chemotaxonomic and morphological properties consistent with its classification in the genus Streptomyces and formed a distinct phyletic line in the Streptomyces 16S rRNA gene tree. It was found to be closely related to Streptomyces celluloflavus NRRL B-2493^T (99.4 % 16S rRNA gene similarity) and shared a 99.0 % 16S rRNA gene similarity value with Streptomyces albolongus NRRL B-3604^T and Streptomyces cavourensis subsp. cavourensis NBRC 13026^T; low levels of DNA–DNA relatedness with these organisms showed that the isolate belonged to a distinct genomic species. The isolate was distinguished readily from the type strains of these species using a combination of morphological and other phenotypic properties. On the basis of these results, it is proposed that isolate ASBV-1^T (= CBMAI 1465^T = CCMA 894^T = NRRL B-24922^T) be classified as the type strain of Streptomyces araujoniae sp. nov.     

Streptomyces jiujiangensis sp. nov., isolated from soil in South China

An actinomycete capable of lysing cyanobacteria, strain JXJ 0074^T, was isolated from a soil sample collected from Jiangxi province, south China, and characterized by using polyphasic taxonomy. The new isolate showed morphological and chemotaxonomic properties typical of members of the genus Streptomyces. Phylogenetic analysis of the near-complete 16S rRNA gene sequence indicated that strain JXJ 0074^T should be affiliated to the genus Streptomyces and exhibited highest similarities to Streptomyces shenzhenensis DSM 42034^T (98.99 %) and Streptomyces lucensis NBRC 13056^T (98.60 %), while the similarities to other members of the genus are lower than 98.22 % similarity. However, the DNA–DNA hybridization values between strain JXJ 0074^T and S. shenzhenensis DSM 42034^T or S. lucensis NBRC 13056^T were 46.2 ± 2.6 and 32.6 ± 3.1 %, respectively. Thus, on the basis of the polyphasic data, strain JXJ 0074^T represents a novel species of the genus Streptomyces, for which the name Streptomyces jiujiangensis sp. nov. is proposed. The type strain is JXJ 0074^T (= BCRC 16953^T = KCTC 29262^T).     

Intergeneric Conjugation in Streptomyces peucetius and Streptomyces sp. Strain C5: Chromosomal Integration and Expression of Recombinant Plasmids Carrying the chiC Gene

Intergeneric conjugal transfer of plasmid DNA from Escherichia coli to Streptomyces circumvents problems such as host-controlled restriction and instability of foreign DNA during the transformation of Streptomyces protoplasts. The anthracycline antibiotic-producing strains Streptomyces peucetius and Streptomyces sp. strain C5 were transformed using E. coli ET12567(pUZ8002) as a conjugal donor. When this donor species, carrying pSET152, was mated with Streptomyces strains, the resident plasmid was mobilized to the recipient and the transferred DNA was also integrated into the recipient chromosome. Analysis of the exconjugants showed stable integration of the plasmid at a single chromosomal site (attB) of the Streptomyces genome. The DNA sequence of the chromosomal integration site was determined and shown to be conserved. However, the core sequence, where the crossover presumably occurred in C5 and S. peucetius, is TTC. These results also showed that the C31 integrative recombination is active and the phage attP site is functional in S. peucetius as well as in C5. The efficiency and specificity of C31-mediated site-specific integration of the plasmid in the presence of a 3.7-kb homologous DNA sequence indicates that integrative recombination is preferred under these conditions. The integration of plasmid DNA did not affect antibiotic biosynthesis or biosynthesis of essential amino acids. Integration of a single copy of a mutant chiC into the wild-type S. peucetius chromosome led to the production of 30-fold more chitinase.     

Unraveling the cellulolytic and hemicellulolytic potential of two novel <Emphasis Type="Italic">Streptomyces</Emphasis> strains

The Streptomyces spp. are notorious plant biomass decomposers in soil environments, but only few strains were biochemically and genetically characterized. Here, we employed functional screening along with genomic sequencing for identification of novel lignocellulolytic Streptomyces strains. Streptomyces strains isolated from soil were functional screened based on their cellulolytic and hemicellulolytic capacities by enzymatic plate assays containing carboxymethylcellulose (CMC) and beechwood xylan as sole carbon source. Subsequently, genomes of Streptomyces strains were sequenced, annotated, and interpreted to correlate their genetic contents with biochemical properties. Among the 80 bacterial isolates that were screened for enzymatic activity, two Streptomyces strains (named as F1 and F7) exhiting higher endoglucanase and endoxylanase activities were selected for biochemical and genomic characterization. After cultivation on steam-pretreated sugarcane bagasse-based medium, the supernatant of the strains F1 and F7 exhibited enzymatic activity against different substrates, such as arabinan, rye arabinoxylan, β-glucan, starch, CMC, xylan, and chitin. Furthermore, strain F7 was able to degrade pectin, mannan, and lichenan. The genomic analysis of both strains revealed a diversity of carbohydrate-active enzymes. The F1 and F7 genomes encode 33 and 44 different types of glycosyl hydrolases families, respectively. Moreover, the genomic analysis also identified genes related to degradation of lignin-derived aromatic compounds. Collectively, the study revealed two novel Streptomyces strains and further insights on the degradation capability of lignocellulolytic bacteria, from which a number of technologies can arise, such as saccharification processes.     

Microorganisms hydrolyse amide bonds; knowledge enabling read-across of biodegradability of fatty acid amides

To get insight in the biodegradation and potential read-across of fatty acid amides, N-[3-(dimethylamino)propyl] cocoamide and N-(1-ethylpiperazine) tall oil amide were used as model compounds. Two bacteria, Pseudomonas aeruginosa PK1 and Pseudomonas putida PK2 were isolated with N-[3-(dimethylamino)propyl] cocoamide and its hydrolysis product N,N-dimethyl-1,3-propanediamine, respectively. In mixed culture, both strains accomplished complete mineralization of N-[3-(dimethylamino)propyl] cocoamide. Aeromonas hydrophila PK3 was enriched with N-(1-ethylpiperazine) tall oil amide and subsequently isolated using agar plates containing dodecanoate. N-(2-Aminoethyl)piperazine, the hydrolysis product of N-(1-ethylpiperazine) tall oil amide, was not degraded. The aerobic biodegradation pathway for primary and secondary fatty acid amides of P. aeruginosa and A. hydrophila involved initial hydrolysis of the amide bond producing ammonium, or amines, where the fatty acids formed were immediately metabolized. Complete mineralization of secondary fatty acid amides depended on the biodegradability of the released amine. Tertiary fatty acid amides were not transformed by P. aeruginosa or A. hydrophila. These strains were able to utilize all tested primary and secondary fatty acid amides independent of the amine structure and fatty acid. Read-across of previous reported ready biodegradability results of primary and secondary fatty acid amides is justified based on the broad substrate specificity and the initial hydrolytic attack of the two isolates PK1 and PK3.