Computational protocol: Draft Genome Sequence of Cylindrospermopsis raciborskii (Cyanobacteria) Strain ITEP-A1 Isolated from a Brazilian Semiarid Freshwater Body: Evidence of Saxitoxin and Cylindrospermopsin Synthetase Genes

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Protocol publication

[…] Cylindrospermopsis raciborskii (Woloszynska) Seenayya and Subba Raju populations of Brazilian freshwater bodies are potential saxitoxin (STX) producers (), and the presence of the sxt biosynthetic gene cluster has been confirmed in the T3 strain (). However, cylindrospermopsin (CYN), a cyanotoxin commonly detected in isolates from Australia and Asia, has recently been reported in Brazilian and South American water supply reservoirs containing C. raciborskii for the first time (). Both cyanotoxins pose significant risk to human and animal health, because CYN causes general cytotoxic, hepatotoxic, and neurotoxic effects, and STX has been implicated in paralytic shellfish poisoning syndrome.C. raciborskii ITEP-A1 was isolated from Arcoverde reservoir, Pernambuco, Brazil (8°33′32.5″S, 36°59′07.5″W). Genomic DNA was extracted from unicyanobacterial cells using the PowerSoil DNA isolation kit (MO BIO Laboratories, Inc., USA) and quantified using a Qubit Fluorometer (Life Technology, USA). Whole-genome sequencing was performed via the MiSeq personal sequencing system (Illumina, USA), using the 500-cycle MiSeq reagent kit version 2 (Illumina). The quality of paired-end reads was checked with FastQC version 0.11.3 (). PRINSEQ version 0.20.4 () was used to trim quality scores under Phred 25, and sequences shorter than 200 bp were removed. Cyanobacterial sequence reads were obtained using the CLARK full mode (confidence score ≥0.75 and gamma score ≥0.03) (), and published genome sequences of the Nostocaceae family as references. De novo genome assembly was carried out with SPAdes version 3.1.1 (), and annotation performed via the RAST server () for contigs equal or longer than 1,000 bp. The final draft genome assembly consisted of 195 contigs, 3,605,836 bp (G+C content of 40.1%), and an N50 value of 91,008. It comprises 3,553 coding sequences of genes, 43 predicted RNA genes (42 tRNA genes and 1 rRNA gene), and 352 subsystems, which represent 40% of assigned sequences. Genome analysis revealed for the first time that ITEP-A1 possesses fragments of gene cluster responsible for cylindrospermopsin biosynthesis, while the presence of saxitoxin genes was confirmed. Genes encoding heterocyst formation, nitrogen fixation, and stress response (oxidative stress, osmotic stress, and heat shock proteins) were found. Genomic analysis revealed that C. raciborskii is capable of evolving diverse genomic organization and adaptive mechanisms. […]

Pipeline specifications

Software tools FastQC, PRINSEQ, SPAdes, RAST
Application WGS analysis
Chemicals Saxitoxin