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Sinorhizobium fredii HH103 is a fast-growing rhizobial strain infecting a broad range of legumes including both American and Asiatic soybeans. In this work, we present the sequencing and annotation of the HH103 genome (7.25 Mb), consisting of one chromosome and six plasmids and representing the structurally most complex sinorhizobial genome sequenced so far. Comparative genomic analyses of S. fredii HH103 with strains USDA257 and NGR234 showed that the core genome of these three strains contains 4,212 genes (61.7% of the HH103 genes). Synteny plot analysis revealed that the much larger chromosome of USDA257 (6.48 Mb) is colinear to the HH103 (4.3 Mb) and NGR324 chromosomes (3.9 Mb). An additional region of the USDA257 chromosome of about 2 Mb displays similarity to plasmid pSfHH103e. Remarkable differences exist between HH103 and NGR234 concerning nod genes, flavonoid effect on surface polysaccharide production, and quorum-sensing systems. Furthermore a number of protein secretion systems have been found. Two genes coding for putative type III-secreted effectors not previously described in S. fredii, nopI and gunA, have been located on the HH103 genome. These differences could be important to understand the different symbiotic behavior of S. fredii strains HH103, USDA257, and NGR234 with soybean.

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Corynebacterium terpenotabidum Takeuchi et. al 1999 is a member of the genus Corynebacterium, which contains Gram-positive and non-spore forming bacteria with a high G+C content. C. terpenotabidum was isolated from soil based on its ability to degrade squalene and belongs to the aerobic and non-hemolytic Corynebacteria. It displays tolerance to salts (up to 8%) and is related to Corynebacterium variabile involved in cheese ripening. As this is a type strain of Corynebacterium, this project describing the 2.75 Mbp long chromosome with its 2,369 protein-coding and 72 RNA genes will aid the G enomic E ncyclopedia of Bacteria and Archaea project.

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Actinoplanes friuliensis HAG 010964 (DSM 7358) was isolated from a soil sample from the Friuli region in Italy and characterized as a producer of the antibiotic friulimycin. The complete genome sequence includes genomic information of secondary metabolite biosynthesis and of its lifestyle. Genbank/EMBL/DDBJ Accession Nr: CP006272 (chromosome).

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Pseudomonas sp. VLB120 was isolated in Stuttgart, Germany, as a styrene degrading organism. The complete genome sequence includes genomic information of solvent tolerance mechanisms, metabolic pathways for various organic compounds, and the megaplasmid pSTY.

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Streptomyces collinus Tü 365 (DSMZ 40733), isolated from Kouroussa (Guinea), is the producer of the elfamycin family antibiotic kirromycin, which inhibits bacterial protein biosynthesis by interfering with elongation factor EF-Tu. Here, we report on the Streptomyces collinus Tü 365 complete genome sequence of the 8.27 MB chromosome and the two plasmids SCO1 and SCO2.

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Sinorhizobium meliloti Rm41 nodulates alfalfa plants, forming indeterminate type nodules. It is characterized by a strain-specific K-antigen able to replace exopolysaccharides in promotion of nodule invasion. We present the Rm41 genome, composed of one chromosome, the chromid pSymB, the megaplasmid pSymA, and the nonsymbiotic plasmid pRme41a.

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Pseudomonas pseudoalcaligenes CECT5344 is a Gram-negative bacterium able to tolerate cyanide and to use it as the sole nitrogen source. We report here the first draft of the whole genome sequence of a P. pseudoalcaligenes strain that assimilates cyanide. Three aspects are specially emphasized in this manuscript. First, some generalities of the genome are shown and discussed in the context of other Pseudomonadaceae genomes, including genome size, G + C content, core genome and singletons among other features. Second, the genome is analysed in the context of cyanide metabolism, describing genes probably involved in cyanide assimilation, like those encoding nitrilases, and genes related to cyanide resistance, like the cio genes encoding the cyanide insensitive oxidases. Finally, the presence of genes probably involved in other processes with a great biotechnological potential like production of bioplastics and biodegradation of pollutants also is discussed.

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The IncF antibiotic resistance and virulence plasmid pRSB225, isolated from an unknown bacterium released with the purified wastewater from a municipal sewage treatment plant into the environment has been analysed at the genomic level by pyrosequencing. The 164,550bp plasmid comprises 210 coding sequences (cds). It is composed of three replicons (RepFIA, RepFIB, and RepFII) and encodes further plasmid-specific functions for stable maintenance and inheritance and conjugative plasmid transfer. The plasmid is self-transmissible and shows a narrow host range limited to the family Enterobacteriaceae. The accessory modules of the plasmid mainly comprise genes conferring resistance to ampicillin (bla(TEM-1b)), chloramphenicol (catA1), erythromycin (mphA), kanamycin and neomycin (aphA1), streptomycin (strAB), sulphonamides (sul2), tetracycline (tetA(B)) and trimethoprim (dfrA14), as well as mercuric ions (mer genes). In addition, putative virulence-associated genes coding for iron uptake (iutA/iucABCD, sitABCD, and a putative high-affinity Fe²âº uptake system) and for a toxin/antitoxin system (vagCD) were identified on the plasmid. All antibiotic and heavy metal resistance genes are located either on class 1 (Tn10-remnant, Tn4352B) and class 2 transposons (Tn2-remnant, Tn21, Tn402-remnant) or a class 1 integron, whereas almost all putative virulence genes are associated with IS elements (IS1, IS26), indicating that transposition and/or recombination events were responsible for acquisition of the accessory pRSB225 modules. Particular modules of plasmid pRSB225 are related to corresponding segments of different virulence plasmids harboured by pathogenic Escherichia coli strains. Moreover, pRSB225 modules were also detected in entero-aggregative-haemorrhagic E. coli (EAHEC) draft genome sequences suggesting that IncF plasmids related to pRSB225 mediated gene transfer into pathogenic E. coli derivatives.

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Crop production may benefit from plant growth-promoting bacteria. The knowledge on bacterial communities is indispensable in agricultural systems that intend to apply beneficial bacteria to improve plant health and production of crops such as canola. In this work, the diversity of root bacterial communities associated to two different developmental phases of canola (Brassica napus L.) plants was assessed through the application of new generation sequencing technology. Total bacterial DNA was extracted from root samples from two different growth states of canola (rosette and flowering). It could be shown how bacterial communities inside the roots changed with the growing stage of the canola plants. There were differences in the abundance of the genera, family, and even the phyla identified for each sample. While in both root samples Proteobacteria was the most common phylum, at the rosette stage, the most common bacteria belonged to the family Pseudomonadaceae and the genus Pseudomonas, and in the flowering stage, the Xanthomonadaceae family and the genus Xanthomonas dominated the community. This implies in a switch in the predominant bacteria in the different developmental stages of the plant, suggesting that the plant itself interferes with the associated microbial community.

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Wickerhamomyces ciferrii is a microorganism characterized by the production and secretion of large amounts of acetylated sphingoid bases, in particular tetraacetyl phytosphingosine. Here, we present the 15.90-Mbp draft genome sequence of W. ciferrii NRRL Y-1031 F-60-10 generated by pyrosequencing and de novo assembly. The draft genome sequence comprising 364 contigs in 150 scaffolds was annotated and covered 6,702 protein-coding sequences. This information will contribute to the metabolic engineering of this yeast to improve the yield and spectrum of acetylated sphingoid bases in biotechnological production.

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Streptomyces davawensis JCM 4913 synthesizes the antibiotic roseoflavin, a structural riboflavin (vitamin B(2)) analog. Here, we report the 9,466,619-bp linear chromosome of S. davawensis JCM 4913 and a 89,331-bp linear plasmid. The sequence has an average G+C content of 70.58% and contains six rRNA operons (16S-23S-5S) and 69 tRNA genes. The 8,616 predicted protein-coding sequences include 32 clusters coding for secondary metabolites, several of which are unique to S. davawensis. The chromosome contains long terminal inverted repeats of 33,255 bp each and atypical telomeres. Sequence analysis with regard to riboflavin biosynthesis revealed three different patterns of gene organization in Streptomyces species. Heterologous expression of a set of genes present on a subgenomic fragment of S. davawensis resulted in the production of roseoflavin by the host Streptomyces coelicolor M1152. Phylogenetic analysis revealed that S. davawensis is a close relative of Streptomyces cinnabarinus, and much to our surprise, we found that the latter bacterium is a roseoflavin producer as well.

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BACKGROUND: The genus Saccharothrix is a representative of the family Pseudonocardiaceae, known to include producer strains of a wide variety of potent antibiotics. Saccharothrix espanaensis produces both saccharomicins A and B of the promising new class of heptadecaglycoside antibiotics, active against both bacteria and yeast. RESULTS: To better assess its capabilities, the complete genome sequence of S. espanaensis was established. With a size of 9,360,653 bp, coding for 8,501 genes, it stands alongside other Pseudonocardiaceae with large genomes. Besides a predicted core genome of 810 genes shared in the family, S. espanaensis has a large number of accessory genes: 2,967 singletons when compared to the family, of which 1,292 have no clear orthologs in the RefSeq database. The genome analysis revealed the presence of 26 biosynthetic gene clusters potentially encoding secondary metabolites. Among them, the cluster coding for the saccharomicins could be identified. CONCLUSION: S. espanaensis is the first completely sequenced species of the genus Saccharothrix. The genome discloses the cluster responsible for the biosynthesis of the saccharomicins, the largest oligosaccharide antibiotic currently identified. Moreover, the genome revealed 25 additional putative secondary metabolite gene clusters further suggesting the strain's potential for natural product synthesis.

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Methanoculleus bourgensis, of the order Methanomicrobiales, is a dominant methanogenic archaeon in many biogas-producing reactor systems fed with renewable primary products. It is capable of synthesizing methane via the hydrogenotrophic pathway utilizing hydrogen and carbon dioxide or formate as the substrates. Here we report the complete and finished genome sequence of M. bourgensis strain MS2(T), isolated from a sewage sludge digester.

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Periodontitis, one of the most common diseases in the world, is caused by a mixture of pathogenic bacteria and inflammatory host responses and often treated by antimicrobials as an adjunct to scaling and root planing (SRP). Our study aims to elucidate explorative and descriptive temporal shifts in bacterial communities between patients treated by SRP alone versus SRP plus antibiotics. This is the first metagenomic study using an Ion Torrent Personal Genome Machine (PGM). Eight subgingival plaque samples from four patients with chronic periodontitis, taken before and two months after intervention were analyzed. Amplicons from the V6 hypervariable region of the 16S rRNA gene were generated and sequenced each on a 314 chip. Sequencing reads were clustered into operational taxonomic units (OTUs, 3% distance), described by community metrics, and taxonomically classified. Reads ranging from 599,933 to 650,416 per sample were clustered into 1,648 to 2,659 non-singleton OTUs, respectively. Increased diversity (Shannon and Simpson) in all samples after therapy was observed regardless of the treatment type whereas richness (ACE) showed no correlation. Taxonomic analysis revealed different microbial shifts between both therapy approaches at all taxonomic levels. Most remarkably, the genera Porphyromonas, Tannerella, Treponema, and Filifactor all harboring periodontal pathogenic species were removed almost only in the group treated with SPR and antibiotics. For the species T. forsythia and P. gingivalis results were corroborated by real-time PCR analysis. In the future, hypothesis free metagenomic analysis could be the key in understanding polymicrobial diseases and be used for therapy monitoring. Therefore, as read length continues to increase and cost to decrease, rapid benchtop sequencers like the PGM might finally be used in routine diagnostic.

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Lactobacillus buchneri belongs to the group of heterofermentative lactic acid bacteria and is a common member of the silage microbiome. Here we report the completely annotated genomic sequence of L. buchneri CD034, a strain isolated from stable grass silage. The whole genome of L. buchneri CD034 was sequenced on the Roche Genome Sequencer FLX platform. It was found to consist of four replicons, a circular chromosome, and three plasmids. The circular chromosome was predicted to encode 2319 proteins and contains a genomic island and two prophages which significantly differ in G+C-content from the remaining chromosome. It possesses all genes for enzymes of a complete phosphoketolase pathway, whereas two enzymes necessary for glycolysis are lacking. This confirms the classification of L. buchneri CD034 as an obligate heterofermentative lactic acid bacterium. A set of genes considered to be involved in the lactate degradation pathway and genes putatively involved in the breakdown of plant cell wall polymers were identified. Moreover, several genes encoding putative S-layer proteins and two CRISPR systems, belonging to the subclasses I-E and II-A, are located on the chromosome. The largest plasmid pCD034-3 was predicted to encode 57 genes, including a putative polysaccharide synthesis gene cluster, whereas the functions of the two smaller plasmids, pCD034-1 and pCD034-2, remain cryptic. Phylogenetic analysis based on sequence comparison of the conserved marker gene rpoA reveals that L. buchneri CD034 is more closely related to Lactobacillus hilgardii strains than to Lactobacillus brevis and Lactobacillus plantarum strains. Comparison of the L. buchneri CD034 core genome to other fully sequenced and closely related members of the genus Lactobacillus disclosed a high degree of conservation between L. buchneri CD034 and the recently sequenced L. buchneri strain NRRL B-30929 and a more distant relationship to L. buchneri ATCC 11577 and L. brevis ssp. gravesensis ATCC 27305, which cluster together with L. hilgardii type strain ATCC 8290. L. buchneri CD034 genome information will certainly provide the basis for further postgenome studies with the objective to optimize application of the strain in silage production.

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Neisseria meningitidis causes invasive meningococcal disease in infants, toddlers, and adolescents worldwide. DNA sequence-based typing, including multilocus sequence typing, analysis of genetic determinants of antibiotic resistance, and sequence typing of vaccine antigens, has become the standard for molecular epidemiology of the organism. However, PCR of multiple targets and consecutive Sanger sequencing provide logistic constraints to reference laboratories. Taking advantage of the recent development of benchtop next-generation sequencers (NGSs) and of BIGSdb, a database accommodating and analyzing genome sequence data, we therefore explored the feasibility and accuracy of Ion Torrent Personal Genome Machine (PGM) sequencing for genomic typing of meningococci. Three strains from a previous meningococcus serogroup B community outbreak were selected to compare conventional typing results with data generated by semiconductor chip-based sequencing. In addition, sequencing of the meningococcal type strain MC58 provided information about the general performance of the technology. The PGM technology generated sequence information for all target genes addressed. The results were 100% concordant with conventional typing results, with no further editing being necessary. In addition, the amount of typing information, i.e., nucleotides and target genes analyzed, could be substantially increased by the combined use of genome sequencing and BIGSdb compared to conventional methods. In the near future, affordable and fast benchtop NGS machines like the PGM might enable reference laboratories to switch to genomic typing on a routine basis. This will reduce workloads and rapidly provide information for laboratory surveillance, outbreak investigation, assessment of vaccine preventability, and antibiotic resistance gene monitoring.

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BACKGROUND: Actinoplanes sp. SE50/110 is known as the wild type producer of the alpha-glucosidase inhibitor acarbose, a potent drug used worldwide in the treatment of type-2 diabetes mellitus. As the incidence of diabetes is rapidly rising worldwide, an ever increasing demand for diabetes drugs, such as acarbose, needs to be anticipated. Consequently, derived Actinoplanes strains with increased acarbose yields are being used in large scale industrial batch fermentation since 1990 and were continuously optimized by conventional mutagenesis and screening experiments. This strategy reached its limits and is generally superseded by modern genetic engineering approaches. As a prerequisite for targeted genetic modifications, the complete genome sequence of the organism has to be known. RESULTS: Here, we present the complete genome sequence of Actinoplanes sp. SE50/110 [GenBank:CP003170], the first publicly available genome of the genus Actinoplanes, comprising various producers of pharmaceutically and economically important secondary metabolites. The genome features a high mean G + C content of 71.32% and consists of one circular chromosome with a size of 9,239,851 bp hosting 8,270 predicted protein coding sequences. Phylogenetic analysis of the core genome revealed a rather distant relation to other sequenced species of the family Micromonosporaceae whereas Actinoplanes utahensis was found to be the closest species based on 16S rRNA gene sequence comparison. Besides the already published acarbose biosynthetic gene cluster sequence, several new non-ribosomal peptide synthetase-, polyketide synthase- and hybrid-clusters were identified on the Actinoplanes genome. Another key feature of the genome represents the discovery of a functional actinomycete integrative and conjugative element. CONCLUSIONS: The complete genome sequence of Actinoplanes sp. SE50/110 marks an important step towards the rational genetic optimization of the acarbose production. In this regard, the identified actinomycete integrative and conjugative element could play a central role by providing the basis for the development of a genetic transformation system for Actinoplanes sp. SE50/110 and other Actinoplanes spp. Furthermore, the identified non-ribosomal peptide synthetase- and polyketide synthase-clusters potentially encode new antibiotics and/or other bioactive compounds, which might be of pharmacologic interest.

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Sinorhizobium fredii HH103 is a fast-growing rhizobial strain that is able to nodulate legumes that develop determinate nodules, e.g., soybean, and legumes that form nodules of the indeterminate type. Here we present the genome of HH103, which consists of one chromosome and five plasmids with a total size of 7.22 Mb.

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Structural composition and gene content of a biogas-producing microbial community from a production-scale biogas plant fed with renewable primary products was recently analyzed by means of a metagenome sequencing approach. To determine the transcriptionally active part of the same biogas community and to identify key transcripts for the biogas production process, the metatranscriptome of the microorganisms was sequenced for the first time. The metatranscriptome sequence dataset generated on the Genome Sequencer FLX platform is represented by 484,920 sequence reads. Taxonomic profiling of the active part of the community by classification of 16S ribosomal sequence tags revealed that members of the Euryarchaeota and Firmicutes account for the dominant phyla. Only smaller fractions of the 16S ribosomal sequence tags were assigned to the phyla Bacteroidetes, Actinobacteria and Synergistetes. Among the mRNA-derived sequence tags from the metatranscriptome dataset, transcripts encoding enzymes involved in substrate hydrolysis, acidogenesis, acetate formation and methanogenesis could be identified. Transcripts for enzymes functioning in methanogenesis are among the most abundant mRNA tags indicating that the corresponding pathway is very active in the methanogenic sub-community. As a frame of reference for evaluation of metatranscriptome sequence data, the 16S rDNA-based taxonomic profile of the community was analyzed by means of high-throughput 16S rDNA amplicon sequencing. Processing of the obtained amplicon reads resulted in 18,598 high-quality 16S rDNA sequences covering the V3-V4 hypervariable region of the 16S rRNA gene. Comparison of the taxonomic profiles deduced from 16S rDNA amplicon sequences and the metatranscriptome dataset indicates a high transcriptional activity of archaeal species. Overall, it was shown that the most abundant species dominating the community also contributed the majority of the transcripts. In the future, key transcripts for the biogas production process will provide valuable markers for evaluation of the performance of biogas-producing microbial communities with the objective to optimize the biotechnology of this process.

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The dissemination of antibiotic resistance genes among bacteria often occurs by means of plasmids. Wastewater treatment plants (WWTP) were previously recognized as hot spots for the horizontal transfer of genetic material. One of the plasmid groups that is often associated with drug resistance is the incompatibility group IncN. The aim of this study was to gain insights into the diversity and evolutionary history of IncN plasmids by determining and comparing the complete genome sequences of the four novel multi-drug resistance plasmids pRSB201, pRSB203, pRSB205 and pRSB206 that were exogenously isolated from the final effluent of a municipal WWTP. Their sizes range between 42,875 bp and 56,488 bp and they share a common set of backbone modules that encode plasmid replication initiation, conjugative transfer, and plasmid maintenance and control. All plasmids are transferable at high rates between Escherichia coli strains, but did not show a broad host range. Different genes conferring resistances to ampicillin, streptomycin, spectinomycin, sulfonamides, tetracycline and trimethoprim were identified in accessory modules inserted in these plasmids. Comparative analysis of the four WWTP IncN plasmids and IncN plasmids deposited in the NCBI database enabled the definition of a core set of backbone genes for this group. Moreover, this approach revealed a close phylogenetic relationship between the IncN plasmids isolated from environmental and clinical samples. Phylogenetic analysis also suggests the existence of host-specific IncN plasmid subgroups. In conclusion, IncN plasmids likely contribute to the dissemination of resistance determinants between environmental bacteria and clinical strains. This is of particular importance since multi-drug resistance IncN plasmids have been previously identified in members of the Enterobacteriaceae that cause severe infections in humans.

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