Ecology of the azaspiracid producer dinoflagellate Azadinium poporum from Shiwha bay, Korea: - taxonomy, population dynamics, azaspiracid content, predation, and growth

Abstract

The azaspiracids (AZAs) are the most recently discovered group of lipophilic marine biotoxins of microalgal origin associated with cases of shellfish poisoning in humans. Since the discovery of Azadinium spinosum as a primary producer of AZA, there has been an urgent need to know more about closely related species and their distribution, the variety of AZA analogues they are producing, their population dynamics in the field, their ecophysiology, and their ecological niche within the planktonic food web.

At the initiation of that work, species of the genus Azadinium were only known from northern European waters. Following the isolation of another strain of the genus Azadinium in 2010 from Shiwha bay, a highly eutrophic area from Korea, works were undertaken in order to explore the taxonomic identity, content of AZA, population dynamics in the field, variety of grazers, and physiology of the new isolate.

The morphology based on optical and scanning electron microscopy as well as the ITS rDNA region and LSU rRNA gene phylogenies demonstrated that the strain was closely related to Azadinium poporum, but minor morphological dissimilarities and an instable phylogenetic position led us to designate it as Azadinium cf. poporum at the time. Later work revealed that the strain was conspecific with Azadinium poporum. This work extended the known distribution of the genus Azadinium further south.

The strain isolated from Shiwha bay preliminarily showed not to contain any known AZA. However, further analyses of the Korean strain by triple quadrupole mass spectrometry on the precursor and product ion mode revealed a new compound, or analogue, with high similarity to AZAs. The structure of the new compound was proposed by interpretation of fragmentation patterns and high resolution mass measurements using Fourier transform ion cyclotron resonance-mass spectrometry. This work increased the known molecular diversity of AZA produced by species in the genus Azadinium.

The long term temporal dynamics of species of the genus Azadinium in the field was completely unknown. Therefore, the population dynamics of A. poporum from Shiwha bay was investigated by qPCR (quantitative polymerase chain reaction). A. poporum revealed to occur always in relatively low concentration from 2009 to 2011 in comparison to common species found in Shiwha bay.

The reduced knowledge on interactions implicating Azadinium within the planktonic food web, the relevance of the Korean isolate of A. poporum as an AZA producer, and the possible role of predation in its field dynamics led us to determine its protistan and metazoan grazers. Furthermore, grazing and growth rates as well as gross growth efficiencies were established for some grazers. Many protistan grazers and copepods were able to feed on A. poporum. However, only two species, the heterotrophic dinoflagellate Oxyrrhis marina and the ciliate Strobilidium sp., were able to achieve sustained growth on A. poporum as the sole prey. Furthermore, for these predators, the maximum ingestion rates and maximum growth rates were the highest and lowest when compared with other prey species, respectively. This suggests that A. poporum was a low quality prey. In addition, the field concentrations of A. poporum assessed by qPCR were generally too low to affect the dynamics of the predators found in this study. Therefore, predation reveals unlikely to be a driving force in the dynamics of A. poporum in the field.

In order to further determine the causes explaining the dynamics of A. poporum in Shiwha bay, we assessed the tendency and seasonality of some environmental parameters through time such as the temperature, the salinity, the pH, the dissolved oxygen, the Secchi depth, the concentration of nutrients, and the concentration of chlorophyll a. We also determined if the previous parameters had an effect on A. poporum in the field as well as the effects of temperature, salinity, and light on the growth of A. poporum in laboratory. The species revealed to grow on a wide range of temperature and salinity and was therefore generally well adapted to the highly variable field conditions observed in Shiwha bay. Furthermore, the species revealed to be well adapted to low irradiance. The growth rates of A. poporum estimated in laboratory were also relatively high compared with other photosynthetic and mixotrophic dinoflagellates under photosynthetic growth. Such features do not explain the low abundances obtained from the field samples. However, A. poporum revealed to be more represented in the field when the concentrations of nitrite and nitrate, silicate, as well as chlorophyll a were lower and the transparency was higher. This suggests that A. poporum use reduced trophic state as windows of opportunity. Considering the low concentrations of A. poporum obtained from field samples during the survey of three years, these opportunities might represent a survival strategy in eutrophic environment.

The multifaceted studies cumulated in this thesis strengthen previous area of research related to the genus Azadinium and established new foundations in unexplored areas from which further studies can rise.