The far-reaching effect of the production, degradation, and trophic transfer of organic arsenic species in the cycle of arsenic

Several organic compounds in aquatic environments contain arsenic, despite of its toxicity. Lipid-soluble arsenic-containing compounds, also known as arsenolipids, are the least studied. In fact, the metabolic origin, purpose and fate of arsenolipids remain shrouded in mystery. Particularly within the microbial realm their role in the arsenic cycle, and significance within trophic webs is still unknown. This thesis aims at improving our understanding about arsenolipids in terms of: 1) the factors that govern their production in microorganisms, 2) their lability, and 3) their trophic transfer from microbial primary producers to eukaryotic grazers. Considering these three approaches, this doctoral thesis follows the path of arsenic starting from cellular pathways to the aquatic environment, and ultimately through food webs, with the aim of assessing how the microbial production of arsenolipids influences arsenic cycling, and which are the factors that affect them.
Arsenic methylation is a key process in the arsenic cycle, converting inorganic arsenic into methylated organic products that can evaporate, altering arsenic mobility. Evidence shows that the subsequent methylation of the highly toxic monomethylarsine (MMA) to a less toxic dimethylarsine (DMA) is regulated by thioredoxin (TRX), which in phototrophs TRX is tightly modulated by oxygenic photosynthesis and light. In Chapter I of this thesis, we investigated how light influences arsenic methylation in phototrophic microorganisms, and what effects could this have on arsenolipid production and arsenic mobility in the environment. We hypothesized that light would drive arsenic methylation and boost DMA production, while production of MMA will predominate in darkness. Using model cyanobacterial cultures and samples from natural arsenic-rich lakes, we found that light stimulates DMA production, while dark anoxic conditions promote toxic MMA production, possibly serving as an antibiotic. Dark oxic settings had no overall influence. This finding provides the first evidence of a physical factor such as light that can influence arsenic methylation, which can also impact arsenolipid production, as DMA is a crucial precursor for their synthesis. These results provide insights as to why arsenolipids are found to be synthesized by phototrophs, with the influence of light on arsenic methylation having an important role. This can also impact arsenic cycling, as the final methylated organic products that can evaporate out of the water column and remove arsenic from the system. Ultimately, this study found that light and oxygenic photosynthesis can be important regulators in the production of methylated arsenic species, which can influence the production of arsenolipids and arsenic cycling.
So how can arsenolipids affect the arsenic cycle? And can light also have an effect on their abundance? In Chapter II, we delved deeper into these questions by characterizing how light affects the occurrence and concentration of arsenolipids within a naturally arsenic-rich microbial mat, as well as their degradation under laboratory conditions. We found arsenolipids in both the mat and water column, predominantly as novel phytyl 2-O-methyl arsenoribosides (arsenosugar phytols/AsPhy). The initial parent arsenosugar phytol (AsPhy 547) was rare in the mat, whereas its shorter chain degradation product (AsPhy 521) was abundant at the mat's surface but absent at greater depths. Given the absence of the initial parent AsPhy 547 on the light-exposed top of the mat and the high abundance of the shorter chain AsPhy 521, we hypothesized that light could degrade AsPhy 547 to AsPhy 521. Results showed light-driven degradation of AsPhy 547 within 24 hours, while AsPhy 521 was more resistant and increased in abundance, suggesting that it is a degradation product. In the dark, all AsPhy were preserved over 63 days. These results could explain the absence of the initial arsenosugar phytol in the illuminated surface layers of the mat, as well as the higher abundance of its degradation products there. AsPhy are so far only known to be made by microalgae, and since the studied mats were cyanobacteria-dominated, we suspect a microalgal-pelagic origin of these AsPhy, with their presence in sediment as detritus. The recalcitrant nature of degraded AsPhy that they can be a sink of arsenic, which eventually degrade over very long period of time with continued burial. These results show that the production of arsenolipids by microorganisms can impact arsenic cycling, as some of their degradation products can be highly refractile, and that light can be an important factor modulating their mobility.
To better understand the role of arsenolipids in the arsenic cycle, studying their transport through the trophic web, from microbial producers to eucaryotes, is crucial. In the arsenic-rich hypersaline lakes of our studies, the aquatic grazer artemia are the main link between microalgae and larger eucaryotes. In Chapter III, we examined artemia's accumulation of arsenolipids during the diel cycle, considering reproductive sex. We conducted feeding experiments on both captured and commercial artemia with arsenic-enriched or non-enriched microalgal food, or under fasting conditions. Results showed that artemia primarily contained arsenosugar phytols (AsPhy) 547 and 521, similar to those in microorganisms from the same lakes, indicating uptake through feeding. In one site, female artemia contained higher amounts of arsenolipids than males, while arsenolipid content was similar between sexes in another site, hinting that environmental factors influence uptake. Long-term fasting and feed-washing incubations showed no significant decrease in arsenolipids, suggesting they are not stored in energy reserves affected during catabolism. Fecal pellets also contained arsenolipids, demonstrating artemia’s role actively in transportation arsenic from microalgae in the water column into the sediment. Thus, we show that the transfer of arsenolipids to eukaryotic grazers can influence arsenic cycling through transport of arsenolipids from the water column to the sediment, as well as their retention in tissue.
Overall, the findings of this study show that microbial organisms, through the production of organic arsenic species, affect arsenic cycling across different scales and facets, starting from the molecular in their synthesis and degradation, to the interaction between organisms as antibiotics, and finally at ecosystem level throughout the food web.