Draft genome sequence of Thermoactinomyces sp. strain AS95 isolated from a Sebkha in Thamelaht, Algeria

The members of the genus Thermoactinomyces are known for their protein degradative capacities. Thermoactinomyces sp. strain AS95 is a Gram-positive filamentous bacterium, isolated from moderately saline water in the Thamelaht region of Algeria. This isolate is a thermophilic aerobic bacterium with the capacity to produce extracellular proteolytic enzymes. This strain exhibits up to 99 % similarity with members of the genus Thermoactinomyces, based on 16S rRNA gene sequence similarity. Here we report on the phenotypic features of Thermoactinomyces sp. strain AS95 together with the draft genome sequence and its annotation. The genome of this strain is 2,558,690 bp in length (one chromosome, but no plasmid) with an average G + C content of 47.95 %, and contains 2550 protein-coding and 60 RNA genes together with 64 ORFs annotated as proteases.


Introduction
Modern metagenomic approaches have provided insights on the evolution and functional capacity of microbial communities resistant to classical culturebased methods [1]. However, these classical techniques remain crucial for understanding the molecular adaptations of microbial guilds, especially those with potential biotechnological applications [2,3]. Consequently, efforts to isolate novel taxa, particularly from environmentally extreme habitats remain widespread [4,5].
Here, we report the draft genome sequence of Thermoactinomyces sp. strain AS95, which was isolated from a sebkha (endorheic salt pan) in the Thamelaht region of Algeria. We present a summary of the classification and set of phenotypic features for Thermoactinomyces sp. strain AS95 together with the description of the non-contiguous genome sequence and its annotation with particular reference to ORFs encoding proteolytic enzymes.

Classification and features
Thermoactinomyces strain AS95 was isolated from a sebkha water sample collected in June 2013 from the Thamelaht region of Algeria (Table 1). This isolate is a Gram-positive, aerobic, thermophilic, filamentous bacterium ( Fig. 1) belonging to the order Bacillales. Based on the 16S rRNA gene sequence similarity searches by BLASTN against the NCBI-NT database, strain AS95 showed 97-99 % sequence similarity to members of the genus Thermoactinomyces. A 16S rRNA gene-based phylogenetic tree of Thermoactinomyces sp. strain AS95 was constructed (Fig. 2), based on neighbor-joining and maximum composite likelihood models with 1000 bootstrap replications using MEGA 7 [12]. The Thermoactinomyces sp. strain AS95 (KU942442) 16S rRNA gene sequence exhibited high identity (99 %) with Thermoactinomyces vulgaris RVH210302 (AY114167), the closest validly published Thermoactinomyces species.
The strain was cultivated on Thermus medium agar containing 2.0 g NaCl, 4.0 g yeast extract, 8.0 g peptone and 30.0 g agar per liter of distilled water. The bacterium grew optimally at 55°C, with a broad temperature growth  [29]. If the evidence is IDA, then the property was directly observed for a live isolate by one of the authors or an expert mentioned in the acknowledgements range of between 40 and 65°C ( Table 1). The strain grew in liquid media at pH values from 5.6 to 8.6, but optimal growth occurred at a pH of 7.2. Morphologically, the isolate forms white colonies and abundant aerial mycelia with the appearance of well-developed, branched and septate substrate mycelia. The micromorphology of the cells was examined using scanning electron microscopy (Fig. 1). The predominant menaquinone was MK-7. Major fatty acids included iso-C15:0, and significant amounts of iso-C17:0 were also present.

Genome sequencing information
Genome project history A high-quality draft genome sequence is deposited at DDBJ/EMBL/GenBank under the accession LSVF00000000 and consists of 11 scaffolds of 11 contigs. A summary of the project information and its association with MIGS version 2.0 compliance are shown in Table 2 [13].

Growth conditions and genomic DNA preparation
Thermoactinomyces sp. strain AS95 was grown aerobically on Thermus medium agar (pH 7.2) at 55°C for 24 h. Genomic DNA was extracted using a modification of a previously described protocol [14]. The quantity and quality of the genomic DNA was measured using a NanoDrop Spectrophotometer and a Qubit™ Fluorometer (Thermo Fisher Scientific Inc.).

Genome sequencing and assembly
Genomic DNA samples of Thermoactinomyces sp. strain AS95 were sequenced at MR DNA (Shallowater, TX, USA). Genome sequencing was performed on a MiSeq (Illumina, Inc.) generating 2 x 300 bp paired-end libraries.
The sequencing run produced a total of 5,085,250 reads, with a mean length of 265.58 bp. The raw paired-end sequences were subjected to the fastxtools software [15] for quality trimming using a phred quality score ≥ 20. After trimming, a total of 3,013,639 reads with a mean length of 171.11 bp were assembled using SPAdes, version 3.5.0 [16]. The final assembly resulted in a total of 11 scaffolds, which generated a genome size of 2.56 Mb.

Genome annotation
Genome annotation was carried out on the RAST server [17] and using the NCBI Prokaryotic Genome Annotation Pipeline tools [18]. This Whole Genome Shotgun sequence project has been deposited at DDBJ/EMBL/ GenBank under accession LSVF00000000. The version described in this paper is version LSVF00000000.

Genome properties
The genome is composed of 2,558,690 nucleotides with 47.95 % G + C content (Table 3) and comprised 11 scaffolds of 11 contigs. The genome contains a total of 2649 genes, 2550 of which were protein coding, 39 pseudogenes and 60 RNA coding genes. The majority of protein-coding genes (75.45 %) were assigned a putative function while the remaining genes were annotated as hypothetical. The distribution of genes in COGs functional categories is presented in Table 4. A blastp comparison was conducted against the MER-OPS database. A total of 64 protein-coding genes (2.4 %) were predicted to share homology with various categories of proteases (Table 5). Of these predictions indicated that 36 were putatively secreted in a classical pathway (SignalP), whereas the other 28 were secreted in a nonclassical pathway (SecretomeP). Only 2 of the 64 protein-coding genes share sequence similarities with The 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. ND: Not determined The total is based on the total number of protein coding genes in the annotated genome proteases of the Thermoactinomyces vulgaris and sp. E79 families of peptidases in the MEROPS database.

Conclusions
This study describes the draft genome sequence of Thermoactinomyces sp. strain AS95, which is associated with a high level of extracellular proteolytic activities. To date, only a few metabolic pathways involved in protein degradation have been characterized for the genus Thermoactinomyces [19]. The genome sequence and characteristics of strain AS95 will provide new insights into the mechanisms of protein degradation in the genus Thermoactinomycetes, and towards establishing a comprehensive genomic catalog of the metabolic diversity of the genus Thermoactinomyces.