Origin of saxitoxin biosynthetic genes in cyanobacteria.

BACKGROUND: Paralytic shellfish poisoning (PSP) is a potentially fatal syndrome associated with the consumption of shellfish that have accumulated saxitoxin (STX). STX is produced by microscopic marine dinoflagellate algae. Little is known about the origin and spread of saxitoxin genes in these under-studied eukaryotes. Fortuitously, some freshwater cyanobacteria also produce STX, providing an ideal model for studying its biosynthesis. Here we focus on saxitoxin-producing cyanobacteria and their non-toxic sisters to elucidate the origin of genes involved in the putative STX biosynthetic pathway. METHODOLOGY/PRINCIPAL FINDINGS: We generated a draft genome assembly of the saxitoxin-producing (STX+) cyanobacterium Anabaena circinalis ACBU02 and searched for 26 candidate saxitoxin-genes (named sxtA to sxtZ) that were recently identified in the toxic strain Cylindrospermopsis raciborskii T3. We also generated a draft assembly of the non-toxic (STX-) sister Anabaena circinalis ACFR02 to aid the identification of saxitoxin-specific genes. Comparative phylogenomic analyses revealed that nine putative STX genes were horizontally transferred from non-cyanobacterial sources, whereas one key gene (sxtA) originated in STX+ cyanobacteria via two independent horizontal transfers followed by fusion. In total, of the 26 candidate saxitoxin-genes, 13 are of cyanobacterial provenance and are monophyletic among the STX+ taxa, four are shared amongst STX+ and STX-cyanobacteria, and the remaining nine genes are specific to STX+ cyanobacteria. CONCLUSIONS/SIGNIFICANCE: Our results provide evidence that the assembly of STX genes in ACBU02 involved multiple HGT events from different sources followed presumably by coordination of the expression of foreign and native genes in the common ancestor of STX+ cyanobacteria. The ability to produce saxitoxin was subsequently lost multiple independent times resulting in a nested relationship of STX+ and STX- strains among Anabaena circinalis strains.

Molecular quantification of symbiotic dinoflagellate algae of the genus Symbiodinium.

The dinoflagellate microalga Symbiodinium is the dominant algal symbiont in corals and related marine animals. To explore the incidence of mixed infections, methods employing real-time quantitative polymerase chain reaction (QPCR) and fluorescence in situ hybridization (FISH) were developed. In experiments focusing on Symbiodinium clades A and B, QPCR and FISH results were well correlated and generally more precise and sensitive than those from the endpoint PCR-restriction fragment length polymorphism analysis (PCR-RFLP) traditionally used for this application, thus increasing the detected incidence of mixed infections. For example, the prevalence of mixed infections in the sea anemone Condylactis gigantea was 40% by PCR-RFLP and 80%-90% by QPCR and FISH. However, the use of QPCR and FISH was limited by inter-host variation in the rRNA gene copy number per Symbiodinium cell, precluding any single conversion factor between QPCR signal and Symbiodinium cell number; and one FISH probe that gave excellent hybridization efficiency with cultured Symbiodinium yielded variable results with Symbiodinium from symbioses. After controlling for these caveats, QPCR studies revealed that field-collected hosts previously described as universally unialgal bore up to 1.6% of the alternative clade. Further research is required to establish the contribution that algal cells at low density in symbiosis and external to the symbiosis make to the minor clade.