Taxonomic & functional description of macro-algae associated microbiome

28 Oct 2022

SIMBA project partners, Erik-Jan Malta (CTAQUA), Henk Bolhuis (NIOZ), Riccardo Rosselli (NIOZ), Edwin Heeren (NIOZ) and Alexandra Klonowski (MATIS), describe the work carried out in the characterisation of the microbiome of the seaweed Ulva ohnoi, a promising and sustainable source for human food and animal feed.

Benefits of seaweed cultivation

Our growing human population will require a considerable increase in the production of animal and plant biomass for food. Terrestrial crops are struggling to meet the demand, and require significant space, fresh water and nutrients for growth, the scarcities of which are further exacerbated by climate change. As an alternative, people are now looking towards the seas as a sustainable source of food and other valuable substances.

Seaweeds are considered a food of high nutritional value. They have also shown potential as nutritive ingredients in aquaculture feeds, both as a substitute for fish meal or plant ingredients of terrestrial origin, or as functional compounds which can improve marine animals’ resistance to disease. In addition to their use as food or animal feed, new seaweed applications are being discovered and exploited continuously. These include their use in fertiliser and plant growth promoting products, biodegradable packaging, cosmetic applications and climate-related and other ecosystem services.

Seaweeds and the microbiome

Large scale cultivation of high-quality seaweed biomass is necessary to keep pace with demand. Therefore, increasing our fundamental knowledge on the life-cycle history, genetics and ecology of seaweeds is essential. The role of bacteria in seaweed development and growth is still largely unknown. Although we’ve known since the late 1970s that bacteria play an essential role in the early development of the green seaweed Ulva sp. and other species, it is only quite recently that researchers are beginning to unravel the mechanisms and identify the bacterial species involved in this process.

The microbial community associated with seaweeds form the so-called “thallisphere”. Collectively, the alga (seaweed) and its bacteria are known as the “holobiont”. At present, most studies focus on the differences in the thallisphere composition in seaweeds from different sites. Little, if any, work has been done on changes in the microbiome community under different cultivation conditions, nor on potential correlations microbe communities may have with seaweed growth rates and biochemical composition.

In SIMBA, we have characterised the effect of the seaweed-associated microbiome on seaweed growth and composition, their variations in time and potential relation with cultivation conditions.

Our ambition is to identify a minimal microbiome needed to improve cultivation. Cultivation experiments and sampling were carried out by CTAQUA in the south of Spain, while partners NIOZ and MATIS worked on DNA extraction and analyses and data analyses.

Research in progress

As a model seaweed, we work with the green seaweed species, Ulva ohnoi. This seaweed is highly versatile, has many applications, widespread distribution, can be cultured in either land- or sea-based facilities and has an enormous production potential. The species has been cultured in CTAQUA since 2017.

To gain a better understanding of the seaweed microbiome, we evaluated development under varying cultivation conditions.  Increasing the level of control on cultivation conditions, we moved from outdoor culture in floating cages in earthen ponds in natural seawater (only control is biomass density) to the same outdoor tanks (controlling biomass density and nutrient levels) and indoor photobioreactors (control of biomass density and ambient parameters such as nutrients, light and temperature).

Cultivation of the green seaweed Ulva ohnoi in the three systems studied. From left to right: photobioreactors, outdoor tanks and floating cages in earthen ponds (Photo: CTAQUA).

We followed growth and biochemical composition of the seaweeds during a full year. In addition, data were collected on environmental variables such as water temperature, salinity and light to be able to analyse their potential role of the changes in the microbial communities. Each month we took samples of the algal microbiome using sterile sampling swabs and we filtered the water they grew in to analyse the water microbiome.

Ulva ohnoi blades on petri dishes with sampling swabs (Photo: CTAQUA).

First results of seaweed growth and microbiome

As expected, seaweed growth rate patterns were very different between cultivation systems. In the earthen ponds, growth rates were highest in late spring, whereas biomass disappeared completely in winter. The system experienced large difference in values of environmental variables such as temperature, light and salinity, with extremely high values for all three in late summer. Growth rates in the outdoor tanks were lowest in late summer and from December to January, and highest in early spring. Most surprising was the strong week to week variation we found in the growth rates in the photobioreactors. During the year we found periods of weeks of more or less constant or increasing growth rates, followed by “crash” episodes, sometimes even with decay of biomass resulting in negative growth rates.

In a first over-all analysis, we compared the microbial communities growing on the seaweed blades with that of the surrounding water. The differences between species diversity and species richness of the community on the earthen pond blades and the earthen pond water were striking. This indicates that the seaweed host might select potentially rare bacterial species that are below the detection limit of the water samples. Advanced comparative statistical analyses of the Ulva thallisphere in all cultivation systems and the microbial communities in the water clearly showed that the community composition of the algae is different from that in the surrounding water.

Results of statistical analyses (nonmetric multidimensional scaling) of the microbial communities of the seaweeds (grouped on the left) and those of the water (right), indicating the clear difference of the species composition between them. (Image: CTAQUA).

A closer look at the bacterial species growing on the algae and in the water confirmed this analysis. Furthermore, we found that the composition of the bacterial community is not static in time, not even when the seaweeds are growing under constant conditions, as in the photobioreactors. Under outdoor conditions, these changes are probably related to naturally changing environmental conditions such as temperature and light, whereas in the photobioreactors these changes might be the consequence of competition between bacteria and complex interactions with the seaweed host. Interestingly, in the earthen ponds, when the temperature was high and seaweed growth rates decreased dramatically, bacteria known to be associated with seaweed degradation tended to increase. Dominance of these species, in combination with the stress the seaweeds are suffering from the high temperatures, might further trigger degradation of the Ulva biomass. This might provide important clues on the bacteria species that are beneficial for Ulva growth and those that are detrimental.

Next steps

For now, we will continue to analyse the data to more detailed levels, combining them with data on environmental variables, seaweed growth rates and biochemical composition to see if we can detect other patterns. The idea is that these detailed analyses will render clues on the bacterial species that might optimise Ulva growth. This would be the first step towards developing a microbiome inoculum that has the potential to stimulate Ulva growth or to protect the seaweeds from detrimental bacteria.