Draft genome sequence of the extremely halophilic archaeon Haladaptatus cibarius type strain D43T isolated from fermented seafood

An extremely halophilic archaeon, Haladaptatus cibarius D43T, was isolated from traditional Korean salt-rich fermented seafood. Strain D43T shows the highest 16S rRNA gene sequence similarity (98.7 %) with Haladaptatus litoreus RO1-28T, is Gram-negative staining, motile, and extremely halophilic. Despite potential industrial applications of extremely halophilic archaea, their genome characteristics remain obscure. Here, we describe the whole genome sequence and annotated features of strain D43T. The 3,926,724 bp genome includes 4,092 protein-coding and 57 RNA genes (including 6 rRNA and 49 tRNA genes) with an average G + C content of 57.76 %.

H. cibarius was isolated from the traditional Korean salt-fermented seafood, which is made with shellfish [8]. D43 T (= DSM 19505 T = JCM 15962 T ) is a representative strain and designated as the type strain of the species. It can grow in 10%-30% (w/v) NaCl (optimum, 15%), with Mg 2+ required for growth. In addition, cells are not lysed in distilled water. The genome sequences of this genus are expected to provide fundamental information for the halotolerant features and biotechnological applications of the haloarchaea. Here, we describe the first whole genome sequence of H. cibarius along with its annotated features, and summarize the taxonomic classification.

Classification and features
The taxonomic position for H. cibarius D43 T was identified with type strains obtained from the EzTaxon-e server [10]. The 16S rRNA sequences of D43 T and closely related strains were aligned using the ClustalW multiple sequence alignment program [11] and were subsequently used for the phylogenetic analysis. Phylogenetic trees were constructed using the neighbor-joining [12], maximum-parsimony [13], and maximum likelihood [14] algorithms with bootstrap values of 1,000 using MEGA version 5 molecular evolutionary genetics analysis program [15]. Strain D43 T clustered with type strains of Haladaptatus species (Fig. 1), exhibiting 16S rRNA gene sequence similarities of 98.7% and 95.1% between strain D43 T (EF660747) and the type strain of H. litoreus and H. paucihalophilus, respectively. Classification and general features of H. cibarius D43 T are shown in Table 1.
Strain D43 T is a Gram-negative staining, coccus or coccobacillus, motile archaeon approximately 1.0 μm in diameter (Fig. 2). Catalase and oxidase tests yielded positive results, but reduction of nitrate to nitrite under aerobic conditions was negative. Cells contained the polar lipids phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, and two unidentified glycolipids. Strain D43 T hydrolyzed gelatin and Tween 80, utilized formate and acetate as carbon sources, and produced acid from sucrose and D-glucose. The strain was sensitive to anisomycin, aphidicolin, chloramphenicol, and rifampicin, and was resistant to ampicillin, erythromycin, kanamycin, streptomycin, and polymycin B.

Genome sequencing and annotation
Genome project history The genome project and sequence of the H. cibarius D43 T genome were deposited in the Genomes OnLine Database [16] (project ID: Gp0086819) and GenBank (accession number: JDTH00000000), respectively. The BioProject number was PRJNA236630. Sequencing and annotation were performed by Chun Lab Inc. (Seoul, Korea) and Integrated Microbial Genomes Expert Review (IMG-ER) [17].

Growth conditions and genomic DNA preparation
H. cibarius D43 T grew optimally on halophilic medium [6] supplemented with 15% (w/v) NaCl and 20 mM Mg 2+ adjusted to pH 7.0, producing colonies with a pink color after incubation at 37°C as previously described [8]. Genomic DNA was extracted and purified using a G-spin DNA extraction kit (iNtRON Biotechnology Inc., Sungnam, Korea), according to the manufacturer's instructions.   Evidence codes -TAS: traceable author statement (i.e., a direct report exists in the literature). These evidence codes are from the Gene Ontology project [30] Genome sequencing and assembly  [18] are shown in Table 2.

Genome properties
The draft genome sequence for H. cibarius D43 T contained 3,926,724 bp, with 13 scaffolds. The G + C content was 57.76 % (Fig. 3 and Table 3), and 4,092 protein-coding genes were predicted along with 57 RNA genes, including six rRNA (two 5S, three 16S, and one 23S rRNA), 49 tRNA, and two additional RNA genes. There were 2,676 protein-coding genes with predicted functions: 773 were enzymes, 98 encoded signal peptides, and 1,049 encoded transmembrane proteins. The distribution of genes in the COG functional categories is shown in

Insights from the genome sequence
The genome analysis of H. cibarius D43 T revealed genes involved in glycine betaine synthesis-including betaine aldehyde dehydrogenase, glycine betaine demethylase, and choline-glycine betaine transporter gene-that allow H.
cibarius to maintain osmotic balance in hypersaline environments. In addtion, trehalose-related genes of trehalose-6-phosphate synthase, trehalose-6-phosphatase, trehalose-6-phosphate synthase and trehalose-6-phosphate hydrolase, and trehalose-utilization protein genes were analyzed in the genome sequences of H. cibarius D43 T . The genes related with trehalose synthesis in the genome show the possibility of trehalose production that is important in food industry.