Strain 101^T (DSM 43827 = ATCC 29888 = NBRC 14064, and other culture collections) is the type strain of Actinosynnema mirum, which is the type species of the genus Actinosynnema [1] (Figure 1). A. mirum was described by Hasegawa et al. in 1978 [1] as an aerobic actinobacterium which forms synnemata (compacted groups of erect hyphae which bear conidia) with zoospores [1]. The organism is of interest due to its position in the tree of life where the small genus Actinosynnema, currently comprising only two species, is located on a rather long branch within the rapidly growing actinobacterial suborder Pseudonocardineae [6]. We here present a summary classification and a set of features for A. mirum strain 101^T (Table 1), together with the description of the complete genomic sequencing and annotation.

Figure 1 Phylogenetic tree highlighting the position of A. mirum 101T relative to all type strains of the genus and to the type strains of the type species of all other genera within the family. The tree was inferred from 1,491 aligned characters [2,3] of the 16S rRNA gene sequence under the maximum likelihood criterion [4] and rooted in accordance with current actinobacterial taxonomy. The branches are scaled in terms of the expected number of substitutions per site. Numbers above branches are support values from 1,000 bootstrap replicates if larger than 60%. Lineages with a type strain genome-sequencing project registered in GOLD [5] are printed in blue; published genomes in bold.

No closely related cultivated strains are known from the literature that can be linked to the species A. mirum. Curiously, the 16S rRNA gene sequences of the type strains from the two subspecies within the second species of the genus Actinosynnema, A. pretiosum subsp. auranticum (AB303364) and A. pretiosum subsp. pretiosum (AB303365) [11], seem to have an equally or even higher degree of similarity to the 16S rRNA gene sequence derived from the genome sequence reported here than the previously reported gene sequences of strain 101^T (see Figure 1). None of the phylotypes reported from environmental screenings or genomic surveys could be linked to A. mirum with a convincing degree of sequence similarity (maximal observed degree of similarity 92%; status June 2009).

A. mirum strain 101^T cells are non-motile with fine hyphae which form aerial and substrate mycelia. Both the aerial and substrate mycelia are about 0.5 to 1.0 µm in diameter. Aerial mycelia are long branching hyphae, white to pale yellow in color (Figure 2). The substrate mycelia are also long, branching hyphae, white to yellowish orange, and penetrate into the agar medium and form synnemata [1]. Cells stain Gram-positive and are non-acid fast [1].

Figure 2 Scanning electron micrograph (SEM) of A. mirum 101T. More SEMs of A. mirum can be found in the CompendiumoftheActinobacteria, by Joachim M. Wink, University of Braunschweig.

A. mirum is capable of producing a yellowish-brown soluble pigment on tyrosine agar and a pale greenish pigment on oatmeal agar [1]. Capable of hydrolyzing starch, casein, tyrosine and gelatin, but not xanthine, hypoxanthine, adenine and urea [1]; produces nitrate reductase and phosphatase. Positive for utilization of tartrate, pyruvate, lactate and malate, but negative for benzoate, acetate, citrate and succinate [1]. Acid is produced aerobically from fructose, lactose, maltose, D-mannitol, L-arabinose, D-melibiose, D-mannose, L-rhamnose, xylose, dextrin, galactose, glucose, trehalose, raffinose, starch, sucrose, cellobiose, glycogen and adonitol, but not from inositol, sorbitol, D-ribose, salicin, inulin, glycerol, dulcitol, erythritol, α-methyl-D-glucoside and α-methyl-D-mannoside. A. mirum is a producer of nocardicin antibiotics [12] and inhibits the growth of several Gram-positive bacteria including: Bacillus megaterium, Sarcina lutea, Mycobacterium smegmatis; as well as the filamentous fungi, Aspergillus niger, Penicillium notatum and the yeasts, Saccharomyces cerevisiae and Candida tropicalis.

Figure 1 shows the phylogenetic neighborhood of A. mirum strain 101^T in a 16S rRNA based tree. The sequences of the five 16S rRNA genes in the A. mirum genome differ by no more than one nucleotide (nt) from each other, and by up to six nts from the previously reported reference sequences derived from NBRC 14064 (AF328679) and from DSM 43827 (X84447). The differences between the genome data and the previously reported 16S rRNA gene sequence are probably due to sequencing errors in the previously reported sequence data.

The genome is 8,248,144 bp long and comprises one circular chromosome with a 73.7% GC content (Table 3 and Figure 3). Of the 7,174 genes predicted, 7,100 were protein coding genes, and 74 RNAs. One hundred and eight four pseudogenes were also identified. The majority of genes (67.3%) of the genes were assigned a putative function while the remaining ones were annotated as hypothetical proteins. The properties and the statistics of the genome are summarized in Table 3. The distribution of genes into COGs functional categories is presented in Table 4.

Figure 3 Graphical circular map of the genome. From outside to the center: Genes on forward strand (color by COG categories), Genes on reverse strand (color by COG categories), RNA genes (tRNAs green, rRNAs red, other RNAs black), GC content, GC skew.