The Raphidiopsis (= Cylindrospermopsis) raciborskii pangenome updated: Two new metagenome-assembled genomes from the South American clade.

Two Raphidiopsis (=Cylindrospermopsis) raciborskii metagenome-assembled genomes (MAGs) were recovered from two freshwater metagenomic datasets sampled in 2011 and 2012 in Pampulha Lake, a hypereutrophic, artificial, shallow reservoir, located in the city of Belo Horizonte (MG), Brazil. Since the late 1970s, the lake has undergone increasing eutrophication pressure, due to wastewater input, leading to the occurrence of frequent cyanobacterial blooms. The major difference observed between PAMP2011 and PAMP2012 MAGs was the lack of the saxitoxin gene cluster in PAMP2012, which also presented a smaller genome, while PAMP2011 presented the complete sxt cluster and all essential proteins and clusters. The pangenome analysis was performed with all Raphidiopsis/Cylindrospermopsis genomes available at NCBI to date, with the addition of PAMP2011 and PAMP2012 MAGs (All33 subset), but also without the South American strains (noSA subset), and only among the South American strains (SA10 and SA8 subsets). We observed a substantial increase in the core genome size for the 'noSA' subset, in comparison to 'All33' subset, and since the core genome reflects the closeness among the pangenome members, the results strongly suggest that the conservation level of the essential gene repertoire seems to be affected by the geographic origin of the strains being analyzed, supporting the existence of a distinct SA clade. The Raphidiopsis pangenome comprised a total of 7943 orthologous protein clusters, and the two new MAGs increased the pangenome size by 11%. The pangenome based phylogenetic relationships among the 33 analyzed genomes showed that the SA genomes clustered together with 99% bootstrap support, reinforcing the metabolic particularity of the Raphidiopsis South American clade, related to its saxitoxin producing unique ability, while also indicating a different evolutionary history due to its geographic isolation.

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Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.