Draft Genome Sequence of Cyanobacterium sp. Strain HL-69, Isolated from a Benthic Microbial Mat from a Magnesium Sulfate-Dominated Hypersaline Lake.

The complete genome sequence of Cyanobacterium sp. strain HL-69 consists of 3,155,247 bp and contains 2,897 predicted genes comprising a chromosome and two plasmids. The genome is consistent with a halophilic nondiazotrophic phototrophic lifestyle, and this organism is able to synthesize most B vitamins and produces several secondary metabolites.

GENOME ANNOUNCEMENT

Cyanobacteria in phototrophic microbial mats are responsible for most of the primary production, provide fixed nitrogen and sulfur to the community, and contribute to the structural integrity of the mat (1, 2). We present here the complete genome sequence of the coccoid unicellular Cyanobacterium sp. strain HL-69 (CHL-69), which was derived from a microbial mat from the magnesium sulfate-dominated hypersaline Hot Lake in northern Washington (3, 4). CHL-69 was isolated from a Hot Lake mat enrichment culture by streaking until axenic on Hot Lake autotroph (HLA) medium, which is BG-11 amended to mimic Hot Lake water (5).

CHL-69 genomic DNA was extracted using a modified cetyltrimethylammonium bromide (CTAB) protocol (5) and was sequenced by the Department of Horticulture Genomics Lab at Washington State University in Pullman, WA, USA, on a PacBio RS II platform, which generated 60,773 reads with a mean length of 5,905 nucleotides (nt). De novo assembly with Hierarchical Genome Assembly Process (HGAP) (SMRT portal version 2.2.0) (Pacific Biosciences) (6) yielded 5 unique contigs. Gaps and sequence errors were resolved using assembled shotgun metagenome data (Illumina HiSeq) from the enrichment culture (https://github.com/jenmobberley/CyanobacteriumHL69). Gene prediction was performed with Prodigal (7) and through the Rapid Annotations using Subsystems Technology (RAST) server (8), and rRNAs and tRNAs were identified with Rfam (9). Genes were assigned functional annotation by use of information from the RAST server (8), BlastKOALA (10), and TIGRFAMs (11).

The genome of CHL-69 consists of a circular chromosome (3,155,247 bp) with an average G+C content of 37.8% and two plasmids, pCHL69-1 (86,432 bp) and pCHL69-2 (55,266 bp), with average G+C contents of 34.1% and 35.32%, respectively. Sequence analysis revealed 3,039 coding sequences, 9 rRNAs, and 44 tRNAs. The chromosome contained a putative prophage as well as a clustered regularly interspaced short palindromic repeat (CRISPR)-cas subtype I-D system. Each plasmid contained parA and toxin-antitoxin genes, which suggests that the plasmids are maintained at a low copy number. Average nucleotide identity (ANI) calculations showed that the HL-69 genome was 95.8% identical to that of the freshwater isolate Cyanobacterium sp. strain IPPAS B-1200 (3,410,249 bp) (GenBank accession no. LWHC00000000) (12) and 82.75% identical to that of the soda lake isolate Cyanobacterium stanieri PCC 7202 (3,163,381 bp) (GenBank accession no. CP003940) (13).

The nutritional dependencies of Cyanobacterium sp. HL-69 were revealed through metabolic reconstruction. HL-69 contains nitrate assimilation genes but lacks nitrogenase, supporting experiments showing HL-69 grows on nitrate but not dinitrogen (Y.M. and J.K.C, unpublished data). The genome of HL-69 indicates it is auxotrophic for vitamin B12 and is capable of salvage through an ABC transporter (btuBFCD). HL-69 is prototrophic for B2, B6, B7, and B9; however, the presence of genes for uptake of B7 (bioY) and B9 (folT) suggests it might be conditionally syntrophic for those vitamins (14). Consistent with CHL-69 growing under a wide range of salinity and light conditions, stress response pathways were identified, such as biosynthesis of the osmolytes glucosyl-glycerol (ggpS) and choline (glpQ), as well as the UV protectant mycosporine, which may be induced by oxidative stress due to high light levels (15).

Accession number(s).

This whole-genome shotgun project has been deposited in GenBank under the accession no. CP024912 (CHL-69), CP024913 (pCHL69-1), and CP024914 (pCHL69-2). The versions described in this paper are the first versions, CP024912.1, CP024913.1, and CP024914.1. The metagenome for the cyanobacterial enrichment culture is publically accessible in JGI’s Integrated Microbial Genomes and Microbiomes (IMG) under IMG Genome ID 3300005412.