Impacts of ocean acidification on growth and toxin content of the marine diatoms Pseudo-nitzschia australis and P. fraudulenta

Highlights

• P. fraudulenta and P. australis strains were able to acclimate and maintain high growth rates at current pH (8.07) and projected pH in 2100 (7.77) compared to the lowest pH level (7.40)

• Domoic acid content was significantly higher for all P. australis toxic strains acclimated at the ambient pH level (8.07), and lowest at pH (7.77)

• Strong inter- and intra-specific variation related to the geographical area and the culturing history of Pseudo-nitzschia strains

Abstract

This paper present the effects of ocean acidification on growth and domoic acid (DA) content of several strains of the toxic Pseudo-nitzschia australis and the non-toxic P. fraudulenta. Three strains of each species (plus two subclones of P. australis) were acclimated and grown in semi-continuous cultures at three pH levels: 8.07, 7.77, and 7.40, in order to simulate changes of seawater pH from present to plausible future levels. Our results showed that lowering pH from current level (8.07) to predicted pH level in 2100 (7.77) did not affect the mean growth rates of some of the P. australis strains (FR-PAU-17 and L3-100), but affected other strains either negatively (L3-30) or positively (L3.4). However, the growth rates significantly decreased with pH lowered to 7.40 (by 13% for L3-100, 43% for L3-30 and 16% for IFR-PAU-17 compared to the rates at pH 8.07). In contrast, growth rates of the non-toxic P. fraudulenta strains were not affected by pH changing from 8.07 to 7.40.

The P. australis strains produced DA at all pH levels tested, and the highest particulate DA concentration normalized to cell abundance (pDA) was found at pH 8.07. Total DA content (pDA and dissolved DA) was significantly higher at current pH (8.07) compared to pH (7.77), exept for one strain (L 3.4) where no difference was found. At lower pH levels 7.77–7.40, total DA content was similar, except for strains IFR-PAU-17 and L3-100 which had the lowest content at the pH 7.77. The diversity in the responses in growth and DA content highlights the inter- and intra-specific variation in Pseudo-nitzschia species in response to ocean acidification. When exploring environmental responses of Pseudo-nitzschia using cultured cells, not only strain-specific variation but also culturing history should be taken into consideration, as the light levels under which the subclones were cultured, afterwards affected both maximum growth rates and DA content.

Introduction

Carbon dioxide (CO₂) concentration in the atmosphere increased from 278 ppm to 400 ppm during the pre-industrial period and is predicted to reach between 800 and 1000 ppm by the end of the 21st century (IPCC, 2014; Le Quéré et al., 2018). As atmospheric CO₂ concentrations continue to increase, the amount of dissolved CO₂ in the ocean will also increase and thus oceanic pH will decrease. Oceanic pH has already decreased by 0.1 unit (from on average 8.21 to 8.1) since the beginning of the industrial era, and atmospheric CO₂ is projected to increase and reach about 720–1020 ppm by the year 2100, which corresponds to a decrease in ocean surface pH by 0.14–0.32 pH units (IPCC, 2014). In addition, coastal and estuarine ecosystems are naturally exposed to pH fluctuations due to upwelling, water depth, tidal currents, residence time, photosynthesis and respiration of phytoplankton (Feely et al., 2008; Middelboe and Hansen, 2007). This acidification may have a significant impact on physiology, proliferation and distribution of phytoplankton species in marine ecosystems (Orr et al., 2005). In addition, changes in pH/pCO₂ can have serious consequences for the functioning of marine ecosystems, from direct effects on physiological responses of phytoplankton and zooplankton (Cornwall et al., 2013; Flynn et al., 2015; Kroeker et al., 2013), to more indirect effects on food web and species interactions (Orr et al., 2005). Ocean acidification (OA) may also affect the distribution of phytoplankton, allowing certain species, mainly diatoms, with an efficient carbon concentrating mechanism (CCM), to thrive and dominate the marine ecosystems (Rost et al., 2008; Trimborn et al., 2013). Apart from ocean acidification, enhanced nutrient release from land may also result in enhancing the development of large phytoplankton blooms (of both harmful algal species HABs and non-HABs) and their eventual bacterial decomposition and resultant release of dissolved carbon dioxide and pH decline (Doney, 2010). Climate change-induced ocean acidification and eutrophication may therefore result in a larger increase in HAB bloom biomass, as blooms will initiate at a lower average pH and thus delay the detrimental effects of high pH at bloom maximum (Flynn et al., 2015; Hansen et al., 2007; Lundholm et al., 2004). Several factors are thus in play when discussing the effect of lowered pH/increased DIC availability on HAB species, making it relevant, but not easy, to explore the effect of lowered pH/increased DIC availability on HAB species. Additional impacts by other biotic and abiotic factors like grazers and temperature will also affect the bloom biomass attained. Specific effects of ocean acidification will depend on the physiology of the individual species (Flynn et al, 2012, 2015) as well as any potential physiological variation within a species.

The diatom Pseudo-nitzschia species are common and abundant members of coastal phytoplankton communities (Malviya et al., 2016; Trainer et al., 2012). Several species of Pseudo-nitzschia produce the neurotoxin, domoic acid (DA), which can cause serious ecological, economic, and health-related problems and are responsible for amnesic shellfish poisoning (ASP) in humans world-wide (Bates et al., 2018). In recent decades, numerous reports have linked mortality events of sea birds, fish and sea mammals to the presence of toxic Pseudo-nitzschia blooms and the accumulation of high DA concentrations in these organisms (Goldstein et al., 2008; Lefebvre et al., 2012; Scholin et al., 2000). In 2015, a long-lasting geographically extensive bloom of the highly toxic species, P. australis, was recorded along the North American west coast causing prolonged closures of shellfish harvesting areas and contributing to the illness and death of seabirds and marine mammals (Di Liberto, 2015; McCabe et al., 2016). As anthropogenic CO₂ emissions continue to increase, it is thus important to understand how growth as well as DA production of Pseudo-nitzschia species respond to the ongoing changes in the marine environment (i.e., acidification).

In general, acidification has been hypothesized to stimulate primary production of phytoplankton (Riebesell et al., 2007; Rost et al., 2008; Schippers et al., 2004a). Presently, relatively few laboratory studies have examined the effect of lowered pH and increased pCO₂ on the physiology of Pseudo-nitzschia species. The findings published show diverging results with respect to growth and DA production in different Pseudo-nitzschia species. Studies on P. fraudulenta and P. multiseries demonstrate significantly increased growth rates with acidification, and significant increase in DA content with increasing CO₂ concentration especially under nutrient-limiting conditions (Sun et al., 2011; Tatters et al., 2012). Moreover, Wingert (2017) showed an increase in DA cellular content in a single P. australis strain due to acidification, whereas the growth rates in contrast to previous studies were not affected until reaching pH 7.8. In addition, Tatters et al. (2018) found that warming and acidification interactions generally increased the cell normalized DA content in P. multisersies in a mixed, natural assemblage. In recent results of a mesocosm experiment conducted in Gullmar Fjord, Sweden, the authors also observed significantly higher particulate DA contents per litre with elevated pCO₂ (OA), here also associated with macronutrient limitation (Wohlrab et al., 2020). Thus, OA appears to generally increase the production of DA under nutrients limited conditions.

The development of high biomass phytoplankton blooms results in lower concentrations of dissolved CO₂ (basification) leading to reduced cellular growth rates in some Pseudo-nitzschia species (Lundholm et al., 2004). A decrease in growth rate and simultaneously an increase in DA content was seen in P. multiseries grown at elevated pH levels (pH 8.9) compared to pH 7.9 and 8.4 (Lundholm et al., 2004; Trimborn et al., 2008). In contrast, no change in the specific growth rates were observed in P. multiseries and P. pungens when maintained under a wide range of pH (from 5 to 9) (Cho et al., 2001). Overall, these apparently contradicting results might be explained by differences in pH levels, differences among Pseudo-nitzschia species, as well as different experimental protocols employed (batch and semi-continuous cultures). Most of the existing studies investigating the effect of changing pH on the physiology and DA production by Pseudo-nitzschia species used only one strain as representative of a species. Considering the genetic variability of Pseudo-nitzschia species (Bates et al., 2018), it is important to study several strains of each species in order to understand the tolerance and toxin production capacity of Pseudo-nitzschia species in response to future ocean acidification.

In the present study, we examined the effects of lowered pH on growth and DA content in acclimated strains of P. australis and P. fraudulenta under nutrient replete conditions. The aim was to evaluate the impact of acidification from present (pH 8.07) to probable future levels (pH 7.77 and 7.40) in three different strains of two different Pseudo-nitzschia species to understand the isolated effects of lowered pH and increased CO₂. P. australis and P. fraudulenta are common and globally widespread species, with P. australis being a highly toxic species, and P. fraudulenta a non-toxic or very slightly toxic species.