Open Access

High quality draft genome sequence of Streptomyces sp. strain AW19M42 isolated from a sea squirt in Northern Norway

  • Gro Elin Kjæreng Bjerga
  • , Erik Hjerde
  • , Concetta De Santi,
  • , Adele Kim Williamson
  • , Arne Oskar Smalås
  • , Nils Peder Willassen
  • and Bjørn Altermark

DOI: 10.4056/sigs.5038901

Received: 01 March 2014

Accepted: 01 March 2014

Published: 15 June 2014


Here we report the 8 Mb high quality draft genome of Streptomyces sp. strain AW19M42, together with specific properties of the organism and the generation, annotation and analysis of its genome sequence. The genome encodes 7,727 putative open reading frames, of which 6,400 could be assigned with COG categories. Also, 62 tRNA genes and 8 rRNA operons were identified. The genome harbors several gene clusters involved in the production of secondary metabolites. Functional screening of the isolate was positive for several enzymatic activities, and some candidate genes coding for those activities are listed in this report. We find that this isolate shows biotechnological potential and is an interesting target for bioprospecting.




The filamentous and Gram-positive genus Streptomyces, belonging to the phylum Actinobacteria [1], are attractive organisms for bioprospecting being the largest antibiotic-producing genus discovered in the microbial world so far [2]. These species have also been exploited for heterologous expression of a variety of secondary metabolites [3]. Additionally, these species harbor genes coding for enzymes that can be applicable in industry and biotechnology [4,5].

Since the first, complete Streptomyces genome was published [6], a number of strains isolated from terrestrial environments have been reported [7-11]. Genomic investigations on Streptomyces from marine sources have, however, just recently begun [12-16].

Here, we present the draft genome sequence of Streptomyces sp. strain AW19M42 isolated from a marine source, together with the description of genome properties and annotation. Results from functional enzyme screening of the bacterium are also reported.

Classification and features

The Streptomyces sp. strain AW19M42 was identified in a biota sample collected from the internal organs of a sea squirt (class Ascidiacea, subphylum Tunicate, phylum Chordata). The tunicate was isolated using an Agassiz trawl at a depth of 77m in Hellmofjorden, in the sub-Arctic region of Norway (Table 1). The trawling was done during a research cruise with R/V Jan Mayen in April 2010.

Table 1

Classification and general features of Streptomyces sp. strain AW19M42 according to the MIGS recommendations [17]




     Evidence code

     Domain Bacteria

     TAS [18]

     Phylum Actinobacteria

     TAS [1]

     Class Actinobacteria

     TAS [19]

     Subclass Actinobacteridae

     TAS [19,20]

     Current classification

     Order Actinomycetales

     TAS [19-22]

     Suborder Streptomycineae

     TAS [19,20]

     Family Streptomycetaceae

     TAS [19,20,22-24]

     Genus Streptomyces

     TAS [22,24-27]

     Species Streptomyces sp.


     Strain AW19M42


     Gram stain

     Gram positive


     Cell shape

     Branched mycelia



     Dispersion of spores





     Temperature range

     Range not determined, grows at 15°C and 28°C




     Not determined, but survives 50% natural sea water



     Oxygen requirements



     Carbon source

     Not reported

     Energy source

     Not reported



     Inner organs of sea squirt



     Biotic relationship







     Biosafety level



     Geographic location

     Hellmofjorden, Norway



     Sample collection time

     April 2010




     N67 49.24316




     E16 28.99465




     77.35 m


Evidence codes - IDA: Inferred from Direct Assay (first time in publication); TAS: Traceable Author Statement (i.e., a direct report exists in the literature); NAS: Non-traceable Author Statement (i.e., not directly observed for the living, isolated sample, but based on a generally accepted property for the species, or anecdotal evidence). These evidence codes are from of the Gene Ontology project [28]. If the evidence code is IDA, then the property was directly observed for a live isolate by one of the authors or an expert or mentioned in the acknowledgements.

The bacterium was isolated during four weeks of incubation at 4-15°C on humic acid containing agar media that is selective for growth of actinomycetes [29,30]. For isolation and nucleic acid extraction the bacterium was cultivated in autoclaved media containing 0.1% (w/v) malt extract, 0.1% (v/v) glycerol, 0.1% (w/v) peptone, 0.1% (w/v) yeast extract, 2% (w/v) agar in 50% (v/v) natural sea water and 50% (v/v) distilled water, pH 8.2 [29]. The gene encoding16S rRNA was amplified by using two universal primers, 27F (5′-AGAGTTTGATCCTGGCTCAG) and 1492R (5′-GGTTACCTTGTTACGACTT) [31], in a standard Taq polymerase driven PCR (VWR) on crude genomic DNA prepared by using InstaGene Matrix (BioRad). Following PCR purification by PureLink PCR Purification (Invitrogen), sequencing was carried out with the BigDye terminator kit version 3.1 (Applied Biosystems) and a universal 515F primer (5′-GTGCCAGCMGCCGCGGTAA) [32]. Using the 16S rRNA sequence data in a homology search by BLAST [33] indicated that the isolate belonged to the Streptomyces genus, among the Streptomycetaceae family of Actinobacteria. A phylogenetic tree was reconstructed from the 16S rRNA gene sequence together with other Streptomyces homologues (Figure 1) using the MEGA 5.10 software suit [34]. The evolutionary history was inferred using the UPGMA method [35] and the evolutionary distances were computed using the Maximum Composite Likelihood method [36]. The phylogenetic analysis confirmed that the isolate AW19M42 belongs to the genus Streptomyces. The closest neighbor with a reported, complete genome sequence is Streptomyces griseus subsp. griseus [7], however, the phylogenetic tree indicates that the Streptomyces sp. strain AW19M42 isolate belongs to a closely related but separate clade. Draft genomes have not been reported for this clade previously.

Figure 1

Phylogenetic tree indicating the phylogenetic relationship of Streptomyces sp. strain AW19M42 relative to other Streptomyces species. The phylogenetic tree was made by comparing the 16S rDNA sequence of the Streptomyces sp. strain AW19M42 to the closest related sequences from both validated type strains and unidentified isolates. S. venezuelea is used as outgroup. All positions containing gaps and missing data were eliminated. There were a total of 1,389 positions in the final dataset. The bar shows the number of base substitutions per site.

Genome sequencing and annotation

The organism was selected for genome sequencing on the basis of its phylogenetic position. The genome project is part of a Norwegian bioprospecting project called Molecules for the Future (MARZymes) which aims to search Arctic and sub-Arctic regions for marine bacterial isolates that might serve as producers of novel secondary metabolites and enzymes. High quality genomic DNA for sequencing was isolated with the GenElute Bacterial Genomic DNA Kit (Sigma) according to the protocol for extraction of nucleic acids from gram positive bacteria. A 700 bp paired-end library was prepared and sequenced using the HiSeq 2000 (Illumina) paired-end technology (Table 2). This generated 13.94 million paired-end reads that were assembled into 670 contigs larger than 500 bp using the CLC Genomics Workbench 5.0 software package [37]. Gene prediction was performed using Glimmer 3 [38] and gene functions were annotated using an in-house genome annotation pipeline.

Table 2

Genome sequencing project information





    Finishing quality

     Improved high quality draft


    Libraries used

     One Illumina Paired-End library


    Sequencing platforms

     Illumina HiSeq2000


    Fold coverage




     CLC paired-end assembly


    Gene calling method

     Glimmer 3

    Genbank ID


    Genbank Date of Release

     September 11, 2013



    Project relevance


Genome properties

The total size of the genome is 8,008,851 bp and has a GC content of 70.57% (Table 3), similar to that of other sequenced Streptomyces isolates. A total of 7,727 coding DNA sequences (CDSs) were predicted (Table 3). Of these, 6,400 could be assigned to a COG number (Table 4). In addition, 62 tRNAs and 8 copies of the rRNA operons were identified.

Table 3

Genome statistics, including nucleotide content and gene count levels



     % of totala

Genome size (bp)



DNA coding region (bp)



DNA G+C content (bp)



Total genes



rRNA operons



tRNA genes



Protein-coding genes



Genes assigned to COGs



Genes with signal peptides



Genes with transmembrane helices



aThe total is based on either the size of the genome in base pairs or the total number of protein coding genes in the annotated genome.

Table 4

Number of genes associated with the 25 general COG functional categories












     RNA processing and modification








     Replication, recombination and repair




     Chromatin structure and dynamics




     Cell cycle control, mitosis and meiosis




     Nuclear structure




     Defense mechanisms




     Signal transduction mechanisms




     Cell wall/membrane biogenesis




     Cell motility








     Extracellular structures




     Intracellular trafficking and secretion




     Posttranslational modification, protein turnover, chaperones




     Energy production and conversion




     Carbohydrate transport and metabolism




     Amino acid transport and metabolism




     Nucleotide transport and metabolism




     Coenzyme transport and metabolism




     Lipid transport and metabolism




     Inorganic ion transport and metabolism




     Secondary metabolites biosynthesis, transport and catabolism




     General function prediction only




     Function unknown




     Not in COGs

aThe total is based on the total number of protein coding genes in the annotated genome.

All putative protein coding sequences were assigned KEGG orthology [39], and mapped onto pathways using the KEGG Automatic Annotation Server (KAAS) server [40]. The analysis revealed that Streptomyces sp. strain AW19M42 harbors several genes related to biosynthesis of secondary metabolites. We have identified genes that map to the streptomycin biosynthesis pathway (glucose-1-phosphate thymidylyltransferase (EC, dTDP-glucose 4,6-dehydratase (EC and dTDP-4-dehydrorhamnose reductase (EC Also, several genes map to the pathways for biosynthesis of siderophore group nonribosomal peptides, biosynthesis of type II polyketide product pathway and polyketide sugar unit biosynthesis. Interestingly, two clusters, comprising five genes, both mapped to the biosynthesis of type II polyketide backbone pathway. These genes clusters comprise genes STREP_3146-3150 and STREP_4370-4374. This suite of genes may contribute to a distinct profile of secondary metabolites production.

Insights from the Genome Sequence

The isolate was successfully screened for lipase, caseinase, gelatinase, chitinase, amylase and DNase activities (Figure 2), by using marine broth (Difco) agar plates incubated at 20°C [41-46]. The plates were supplemented with 1% (v/v) tributyrin, 1% (w/v) skim milk, 0.4% (w/v) gelatin, 0.5% (w/v) chitin or 2% (w/v) starch, respectively (all substrates from Sigma), whereas DNase test agar (Merck) was supplemented with 0.3M NaCl, representing sea water salt concentration, before screening for DNase activity. Putative genes coding for these activities were identified in the genome based on annotation or by homology search (Table 5).

Figure 2

Degradation halos around colonies of Streptomyces sp. AW19M42 growing on agar plates supplemented with A, skim milk, B, gelatin, C, tributyrin, D, DNA, E, chitin and F, starch.

Table 5

Candidate genes coding for putative lipase, caseinase, gelatinase and DNase activities identified in Streptomyces sp. strain AW19M42 draft genome.

Putative gene


    Size (aa)






     Triacylglycerol lipase



     G-D-S-L family lipolytic protein



     Lipase class 2



     Triacylglycerol lipase



     Secreted hydrolase



     Lipase or acylhydrolase family protein









     Triacylglycerol lipase




     Peptidase M4 thermolysin



     Peptidase M4 thermolysin



     Peptidase M4 thermolysin




     Putative secreted serine protease









     Serine protease



     M6 family metalloprotease domain-containing protein



     M6 family metalloprotease domain protein



     Putative secreted subtilisin-like serine protease






     Metalloprotease domain protein



     ATP-dependent protease La



     ATP-dependent metalloprotease FtsH



     Streptogrisin-B - Pronase enzyme B SGPB/Serine protease B



     ATP-dependent Clp protease proteolytic subunit



     ATP-dependent Clp protease, ATP-binding subunit ClpX















     Putative protease



     Serine protease







     Exodeoxyribonuclease VII, large subunit



     Exodeoxyribonuclease VII small subunit



     Exodeoxyribonuclease III Xth



     TatD-related deoxyribonuclease



     Deoxyribonuclease V



     Deoxyribonuclease/rho motif-related TRAM






     Probable endonuclease 4 - Endodeoxyribonuclease




     Chitinase, glycosyl hydrolase 18 family



     Chitinase, glycosyl hydrolase 18 family



     Carbohydrate-binding CenC domain protein



     Glycoside hydrolase family protein



     Putative endochitinase



     Chitinase, glycosyl hydrolase 19 family



     Chitinase, glycosyl hydrolase 19 family




     Glycoside hydrolase starch-binding protein



     Secreted alpha-amylase



     Malto-oligosyltrehalose synthase



     Alpha-1,6-glucosidase, pullulanase-type



The 8 Mb draft genome belonging to Streptomyces sp. strain AW19M42, originally isolated from a marine sea squirt in the sub-Arctic region of Norway has been deposited at ENA/DDBJ/GenBank under accession number CBRG000000000. The isolate was successfully screened for several enzymatic activities that are applicable in biotechnology and candidate genes coding for the enzyme activities were identified in the genome. Streptomyces sp. strain AW19M42 will serve as a source of functional enzymes and other bioactive chemicals in future bioprospecting projects.



This work was supported by the Research Council of Norway (Grant no. 192123). We would like to acknowledge Kristin E. Hansen and Seila Pandur for technical assistance during bacterial isolation and nucleic acid extraction. The sequencing service was provided by the Norwegian Sequencing Centre (Web Site), a national technology platform hosted by the University of Oslo and supported by the "Functional Genomics" and "Infrastructure" programs of the Research Council of Norway and the Southeastern Regional Health Authorities.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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