Sporadic distribution and distinctive variations of cylindrospermopsin genes in cyanobacterial strains and environmental samples from Chinese freshwater bodies

Abstract

Increasing reports of cylindrospermopsins (CYNs) in freshwater ecosystems have promoted the demand for identifying all of the potential CYN-producing cyanobacterial species. The present study explored the phylogenetic distribution and evolution of cyr genes in cyanobacterial strains and water samples from China. Four Cylindrospermopsis strains and two Raphidiopsis strains were confirmed to produce CYNs. Mutant cyrI and cyrK genes were observed in these strains. Cloned cyr gene sequences from eight water bodies were clustered with cyr genes from Cylindrospermopsis and Raphidiopsis (C/R group) in the phylogenetic trees with high similarities (99%). Four cyrI sequence types and three cyrJ sequence types were observed to have different sequence insertions and repeats. Phylogenetic analysis of the rpoC1 sequences of the C/R group revealed four conserved clades, namely, clade I, clade II, clade III, and clade V. High sequence similarities (>97%) in each clade and a divergent clade IV were observed. Therefore, CYN producers were sporadically distributed in congeneric and paraphyletic C/R group species in Chinese freshwater ecosystems. In the evolution of cyr genes, intragenomic translocations and intergenomic transfer between local Cylindrospermopsis and Raphidiopsis were emphasized and probably mediated by transposases. This research confirms the existence of CYN-producing Cylindrospermopsis in China and reveals the distinctive variations of cyr genes.

FIG 1

Schematic structure of cyr gene clusters from CYN-producing cyanobacterial strains. Gray and white bars, cyr genes; black bar, transposase sequences or vestiges thereof; open triangles, base mutation in this position; solid triangles, nucleotide deletion in this position.

FIG 2

Illustration of four sequence types of the cyrI gene. (A) Schematic structures of cyrI sequence types. White bar, cyrI sequences; black and gray bars, repeat sequences; slash and backslash bar, insertion sequences; C. raciborskii AWT205, reference strain. (B) Partial alignment of representative cyrI gene sequences. Repeat sequences and insertion sequences were italicized. Dashed line, gaps introduced into the alignment; bold line, ITRs; arrow, beginning of the repeat sequences.

FIG 3

Illustration of three sequence types of the cyrJ gene. (A) Schematic structures of cyrJ sequence types. White bar, cyrJ sequence; gray bar, repeat sequences; triangle, nucleotide deletions; C. raciborskii AWT205, reference strain. (B) Partial alignment of representative cyrJ gene sequences and deduced protein sequences. Repeat sequences were italicized. Dashed line, gaps introduced into the alignment; arrow, beginning of the repeat sequences.

FIG 4

Partial alignment of mutant cyrK gene sequences and deduced protein sequences. Rectangle, nucleotide deletion; asterisk, stop codon.

FIG 5

Phylogenetic tree of rpoC1 gene sequences from environmental samples and cyanobacterial strains (topology based on a Bayesian tree). Bootstrap values above 50% are indicated at the nodes of the tree (Bayesian/ML/NJ). Aphanizomenon gracile ANA196-A and Anabaena variabilis ATCC 29413 were used as outgroups.