The draft genome sequence of Raphidiopsis (Cylindrospermopsis) raciborskii strain GIHE 2018, a filamentous nitrogen-fixing and potentially harmful cyanobacterium, is reported here. This is the first strain isolated from a shallow freshwater pond in South Korea. This information is expected to improve our understanding of the phylogeography of this species.
ABSTRACT
The draft genome sequence of Raphidiopsis (Cylindrospermopsis) raciborskii strain GIHE 2018, a filamentous nitrogen-fixing and potentially harmful cyanobacterium, is reported here. This is the first strain isolated from a shallow freshwater pond in South Korea. This information is expected to improve our understanding of the phylogeography of this species.
ANNOUNCEMENT
Raphidiopsis raciborskii (formerly Cylindrospermopsis raciborskii) is a filamentous nitrogen-fixing and potentially harmful cyanobacterium originating in the tropics (1). The geographical range of R. raciborskii has expanded worldwide because of global climate change and its adaptability to various environments (2, 3). Despite morphological differences, this species was reclassified as belonging to the Raphidiopsis genus in 2018 based on polyphasic (4) and genetic marker-based analyses (5). R. raciborskii strain GIHE 2018 was observed and isolated in a shallow freshwater pond in South Korea for the first time (6).
A shallow freshwater pond sample was collected from Incheon, South Korea, using the grab sampling method (7). Strain GIHE 2018 was isolated using the micropipetting method (8) and cultured in Jaworski’s medium (9) at 25°C under a 16/8-h light/dark cycle. After extracting the total DNA using a water DNA purification kit (Norgen, Thorold, Canada), the PacBio (Menlo Park, CA) and Illumina (San Diego, CA) sequencing libraries were prepared using a P6 PacBio DNA/polymerase binding kit, a PacBio DNA sequencing kit 4.0, 8 single-molecule real-time (SMRT) cells, and the TruSeq library kit, respectively, according to the manufacturer's protocols. The genome was sequenced using one run each of the PacBio RS II and Illumina HiSeq 2500 platforms, respectively.
After the removal of adaptor and low-quality sequences by self-correction during the preassembly step using FALCON-integrate software v2.1.4 (10; https://github.com/PacificBiosciences/FALCON-integrate), 119,828 subreads (1,046,050,466 bp) were generated from the PacBio platform and used for the assembly step using the same software. From Illumina paired-end sequencing, 14,386,820 reads (100 ± 0 bp) were retrieved. The quality-filtered sequence reads from the Illumina sequencing, where 90% of bases had a Phred score of 30 or higher, were used for error correction using Pilon v1.21 (11; https://github.com/broadinstitute/pilon/wiki). The final coverage (depth) of the sequenced genome was 63×. Then, the genome was annotated using NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v.4.8 (12, 13). Default parameters were used for all software unless otherwise specified.
From the PacBio sequencing, three uncircularized contigs were retrieved, with an N50 value of 1,278,585 bp (after assembly correction) and gaps of 33,630 bp (between contigs 1 and 2, gap = 21,674 bp; 2 and 3, gap = 11,956 bp) compared to those of the reference genome (GenBank accession number NZ_ACYA00000000.1). Alignment was performed to find matched areas between the draft genome and the reference genome using NUCmer (nucleotide MUMmer) software v4.0 (http://mummer.sourceforge.net/). After that, the gap size was calculated using Assemblytics Web-based software with the default option.
The genome information provided by PGAP showed that the genome contains 3,629,822 bp with 40.2% GC content and 3,265 coding sequences, 2,995 proteins, 270 pseudogenes, 42 tRNA genes, 9 rRNA genes, and 4 noncoding RNA (ncRNA) genes. Four CRISPR arrays were also discovered via PGAP. Moreover, four genes encoding a nonribosomal peptide synthetase complex related to hassallidin biosynthesis were identified.
Several strains of Raphidiopsis species produce toxins such as cylindrospermopsin or saxitoxin (14). However, in the GIHE 2018 genome, gene clusters associated with the biosynthesis of these cyanobacterial toxins were absent, suggesting that the isolated strain does not produce cyanobacterial toxins in its natural environment.
Data availability.
The draft genome sequence of Raphidiopsis raciborskii GIHE 2018 has been deposited in NCBI under accession numbers VHLJ01000001 through VHLJ01000003 (GenBank), PRJNA548643 (BioProject), and SRR10071356 through SRR10071359 (SRA).
ACKNOWLEDGMENT
This research was partially supported by the National Institute of Environmental Research.
REFERENCES
- 1.Padisák J, Vasas G, Borics G. 2016. Phycogeography of freshwater phytoplankton: traditional knowledge and new molecular tools. Hydrobiologia 764:3–27. doi: 10.1007/s10750-015-2259-4. [DOI] [Google Scholar]
- 2.Kokociński M, Gągała I, Jasser I, Karosienė J, Kasperovičienė J, Kobos J, Koreivienė J, Soininen J, Szczurowska A, Woszczyk M, Mankiewicz-Boczek J. 2017. Distribution of invasive Cylindrospermopsis raciborskii in the East-Central Europe is driven by climatic and local environmental variables. FEMS Microbiol Ecol 93:1–8. doi: 10.1093/femsec/fix035. [DOI] [PubMed] [Google Scholar]
- 3.Soares MCS, Lürling M, Huszar V. 2013. Growth and temperature-related phenotypic plasticity in the cyanobacterium Cylindrospermopsis raciborskii. Phycol Res 61:61–67. doi: 10.1111/pre.12001. [DOI] [Google Scholar]
- 4.Aguilera A, Gómez EB, Kaštovský J, Echenique RO, Salerno GL. 2018. The polyphasic analysis of two native Raphidiopsis isolates supports the unification of the genera Raphidiopsis and Cylindrospermopsis (Nostocales, Cyanobacteria). Phycologia 57:130–146. doi: 10.2216/17-2.1. [DOI] [Google Scholar]
- 5.Dyble J, Paerl HW, Neilan BA. 2002. Genetic characterization of Cylindrospermopsis raciborskii (cyanobacteria) isolates from diverse geographic origins based on nifH and cpcBA-IGS nucleotide sequence analysis. Appl Environ Microbiol 68:2567–2571. doi: 10.1128/aem.68.5.2567-2571.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Jeong J-Y, Lee S-H, Yun M-R, Oh S-E, Kim J-S, Moon H-C, Hwang C-W, Park H-D. 2019. First isolation report and molecular phylogenetic characteristics of Raphidiopsis raciborskii in South Korea. Korean J Microbiol 55:350–359. [Google Scholar]
- 7.Downing JA. 1984. Sampling the benthos of standing waters, p 87–130. In Downing JA, Rigler FH (ed), A manual on methods for the assessment of secondary productivity in fresh waters: IBP handbook no. 17, 2nd ed Blackwell Scientific Publications, Oxford, United Kingdom. [Google Scholar]
- 8.Andersen RA. 2005. Algal culturing techniques. Elsevier Academic Press, Burlington, MA. [Google Scholar]
- 9.Thompson AS, Rhodes JC, Pettman I. 1988. Natural Environmental Research Council Culture Collection of Algae and Protozoa—catalogue of strains, p 22 Freshwater Biology Association, Ambleside, United Kingdom. [Google Scholar]
- 10.Chin C-S, Peluso P, Sedlazeck FJ, Nattestad M, Concepcion GT, Clum A, Dunn C, O'Malley R, Figueroa-Balderas R, Morales-Cruz A, Cramer GR, Delledonne M, Luo C, Ecker JR, Cantu D, Rank DR, Schatz MC. 2016. Phased diploid genome assembly with single-molecule real-time sequencing. Nat Methods 13:1050–1054. doi: 10.1038/nmeth.4035. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Walker BJ, Abeel T, Shea T, Priest M, Abouelliel A, Sakthikumar S, Cuomo CA, Zeng Q, Wortman J, Young SK, Earl AM. 2014. Pilon: an integrated tool for comprehensive microbial variant detection and genome assembly improvement. PLoS One 9:e112963. doi: 10.1371/journal.pone.0112963. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Tatusova T, DiCuccio M, Badretdin A, Chetvernin V, Nawrocki EP, Zaslavsky L, Lomsadze A, Pruitt KD, Borodovsky M, Ostell J. 2016. NCBI Prokaryotic Genome Annotation Pipeline. Nucleic Acids Res 44:6614–6624. doi: 10.1093/nar/gkw569. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Haft DH, DiCuccio M, Badretdin A, Brover V, Chetvernin V, O'Neill K, Li W, Chitsaz F, Derbyshire MK, Gonzales NR, Gwadz M, Lu F, Marchler GH, Song JS, Thanki N, Yamashita RA, Zheng C, Thibaud-Nissen F, Geer LY, Marchler-Bauer A, Pruitt KD. 2018. RefSeq: an update on prokaryotic genome annotation and curation. Nucleic Acids Res 46:D851–D860. doi: 10.1093/nar/gkx1068. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Lagos N, Onodera H, Zagatto P, Andrinolo D, Azevedo SMFQ, Oshima Y. 1999. The first evidence of paralytic shellfish toxins in the freshwater cyanobacterium Cylindrospermopsis raciborskii, isolated from Brazil. Toxicon 37:1359–1373. doi: 10.1016/S0041-0101(99)00080-X. [DOI] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The draft genome sequence of Raphidiopsis raciborskii GIHE 2018 has been deposited in NCBI under accession numbers VHLJ01000001 through VHLJ01000003 (GenBank), PRJNA548643 (BioProject), and SRR10071356 through SRR10071359 (SRA).