Complete genome sequence of Delftia tsuruhatensis strain HA60 isolated from a commercial hydroxyapatite nano-particle product (nano-hydroxyapatite)

ABSTRACT

Delftia tsuruhatensis is a Gram-negative rod-shaped aerobic bacterium with environmental remediation functions. D. tsuruhatensis strain HA60 was isolated from a commercial nano-particle product, nano-hydroxyapatite. We report that the genome of D. tsuruhatensis strain HA60 has a circular genome of 6,922,195 base pairs with a G+C content of 66.45%.

ANNOUNCEMENT

Delftia tsuruhatensis is known as a plant growth-promoting rhizobacterium, an environmental bioremediation bacterium, and an emerging opportunistic pathogen (1). Pan-genome analysis of D. tsuruhatensis reveals extensive gene diversity through horizontal gene transfer (1, 2). D. tsuruhatensis is capable of the biodegradation of environmental pollutants such as chlorobenzene, dimethylphenols, diesel fuel, naphthalene, and peptidoglycan (3, 4). D. tsuruhatensis is resistant to heavy metals and capable of the synthesis of phytohormones, siderophores, and antibiotics (1, 3). Though the pathogenic roles of D. tsuruhatensis are little known, recent reports of clinical isolates indicate likely an emerging opportunistic pathogen (5, 6). The HA60 genome report provides additional genetic information about this understudied bacterium.

D. tsuruhatensis was isolated from a commercial hydroxyapatite nano-particle product. One hundred microliters of the nano-particle solution was directly streaked on tryptic soy agar containing 5% sheep blood (Remel) and cultured for 2 days at 35°C aerobically. A Gram-negative rod bacterium was subcultured to obtain a single-colony pure culture. The bacterium was identified as Delftia acidovorans using the matrix-assisted laser desorption/ionization–time of flight Biotyper (Bruker) (7). The bacterium was reclassified by the NCBI taxonomy team as D. tsuruhatensis.

Genomic DNA was isolated from a colony grown overnight on Tryptic Soy Blood Agar (Remel) using the Qiagen Blood & Tissue Kit for Illumina short-read and Qiagen MagAttract HMW DNA Kit for Nanopore long-read sequencing with no shearing or size selection. The quality and quantity of DNA were determined using Nanodrop and Qubit (Fisher Scientific, Waltham, MA). Default parameters were used for all software unless otherwise noted. A short-read genomic library was constructed using the Nextera XT library prep kit and sequenced using the MiSeq v3 reagent kit (2 × 300 bp) on MiSeq (Illumina). For long reads, a genomic library was constructed using the ligation sequencing kit SQK-LAK114, sequenced on Nanopore MinION FLO-MIN1114 R10, and data acquisition and basecalling were done in the FAST mode using MinKNOW22.12.5 (Guppy6.4.6) on MinION Mk1C (Oxford Nanopore, Oxford, UK). Raw short reads were quality-checked and trimmed using Fastq Utilities (CutAdapt v2.2 with Python v3.7.9) in BV-BRC^3.33.16 (8), and raw long reads were quality-checked and trimmed with Porechop v0.2.1 and Galore v0.6.7 in Galaxy (9). Illumina short reads (n = 1,705,027) and Nanopore long reads (n = 39,914; N₅₀ = 2,295 bp) were assembled in one circular contig with 123× depth using Unicycler v0.4.8 (default normal bridging mode, rotation) in BV-BRC^3.33.16 (8). The genome of D. tsuruhatensis HA60 is 6,922,195 bp in length with a G+C content of 66.45%. The HA60 genome, annotated using the NCBI Prokaryotic Genome Annotation Pipeline v4.11, has 6,299 coding genes, 54 pseudogenes, 79 tRNAs, 15 rRNAs, and 2 CRISPR arrays (10). The closest reference genome is D. tsuruhatensis NBRC16741 (BCTO000000000) with an average nucleotide identity of 98.4% estimated using MinHashv2.3 (11). HA60 genome is predicted to encode the entire NRPS/PKS genes (delH, delG, and delF) necessary for the biosynthesis of delftibactin that detoxifies soluble gold and mineralizes solid gold (2). HA60 genome also encodes multiple antibiotic resistance-related genes such as rpoB (rifampicin), murA (fosfomycin), gyrB (fluoroquinolone), macB (erythromycin), and bla (β-lactam) (12).

DATA AVAILABILITY

The whole-genome sequence has been deposited at DDBJ/ENA/GenBank under the accession number CP135236. The raw reads have been deposited in the Sequence Read Archive (SRA) under the accession numbers SRR26117949 and SRR26117950.