ViralICE: Viral diversity and Interactions in a Changing Environment on Kelp.

Project summary

Being the most abundant and diverse entities in planet Earth, marine viruses have been proved to control their host populations, acting as driving force for the inter- and intra-species competition and succession. Their outstanding roles in food webs, competitive interactions, biodiversity patterns, and the regulation of keystone species, manifest therefore, their contribution structuring ecological communities and impacting ecosystem functioning. However, we still do not completely understand how viruses interact with their hosts.

 

Kelp forest (brown algae from the order Laminariales) impacts local oceanography and ecology, and constitute enormous energy sources for coastal benthic secondary production supporting fisheries worldwide; from temperate, to polar rocky ecosystems. Kelp deforestation has been reported globally and, after the recent discovery of Phaeoviruses on kelp, viral diseases should be taken into consideration. Evidence posses that host-pathogen interactions will become more frequent and intense in the future, leading to higher virus multiplication rates, increased transmission and host species jumps. Wild seaweed domestication is making crops more susceptible to abiotic stressors, disease and parasites, and these may act as a disease reservoir which could impact natural populations. Therefore, viral infection could not only affect natural kelp forest, but also impact the Norwegian blue bioeconomy.

 

ViralICE has been designed to throw some light into viral diseases that could potentially affect kelp communities and relevant species to the Norwegian industry. This project intends to contribute with Phaeoviral infection knowledge in brown algae, and we will: 1) study Phaeoviral diversity, 2) study the Phaeoviral host-range, 3) study the potential ecological consequences of climate change on algal-pathogen interactions, and 4) build relevant networks between research communities, industry and government.

 

Method

The project is divided in 2 working packages (WP):

• WP1: Phaeoviral diversity and phylogeny on relevant kelp species:

We will work with natural and cultured samples provided by UIB and our industry collaborators, Ocean Forest and DuPont Health & Nutrition.  

All samples will be screened for viral detection through DNA extraction following Maeda et al. 2013 and Mckeown et al. 2018, and targeting the phaeovirus-encoded major capsid protein (MCP). PCRs, Real-Time PCR (qPCR) and High Resolution Melt (HRM) analysis will be performed following to McKeown and colleagues. The raw melting temperatures will be calibrated by applying the correction factor from reference clones and genomic DNA. Each corrected melting temperature will be assigned to a viral subgroup using posterior group probabilities. All PCR products will be sequenced by Sanger sequencing.

Different parts from the samples will be analysed, meaning reproductive (sporophytes and gametophytes) and vegetative tissue.. This will enable us to test the hypothesis that phaeoviruses on kelp are expressed in gametophytes and vegetative cells, in contrast to in ectocarpoids, where viral expression occurs in both reproductive structures, but not in vegetative cells. Note that gametophyte cultures will be obtained from sporophyte spore release in the laboratory (methodology described under WP2). DAPI staining, along with light and epifluorescence microscopy, will be used to support the results obtained by molecular techniques, and further characterize the nature of viral infections on these species in cases a new viral line has been found.

For the phylogeny study, we will work with the obtained phaeoviral positive samples. BLAST research will be performed to investigate the homology between our MCP sequences and the so far described sequences in the GenBank database.

Distance analysis will be performed using the software MEGA7, bayesian inference analysis will be performed using MrBayes v3.2, and trees will be visualized using Inkscape 0.92 and Dendroscope 3. Host-virus infection analysis will be performed using the BiMat package for Matlab. 

Virus-host Phylogenetic affiliation of the phaeoviruses in our samples, along with the infection network analysis will tell us if these are similar or different from the described cultured or uncultured phaeoviruses, potentially providing general information about their evolutionary history and life strategies.

• WP2: Potential ecological consequences of climate change on algal-pathogen interactions.

During the first year of sampling, diverse physicochemical parameters of the water will be measured (e.g. temperature, salinity and hydrodynamics) at the same time the samples are collected. Correlations between these variables and the presence of viruses in the samples from all locations will be done in order to determinate if temperature gradients, in this case, increase or decrease viral presence.

Experimental tests will be also done with reproductive kelp, previously scanned for viruses.

 

In order to integrate WP1 and WP2, statistical analyses will be performed in the R statistical computing environment using different community ecology and graphic packages such as vegan v2.4.5, ggplot2 v3.0.0, and gplots v3.0.1. In this way, we will look after significance of our data and possible variable correlations (using ordination methods like PCA analysis), to properly explain and interpret our results.