My seaweed looks weird: a community web portal to accelerate pathogen discovery in seaweeds

ABSTRACT Worldwide, novel species of pathogens are frequently reported in cultivated and wild macroalgae. Alongside the rapid growth of the seaweed industry, diseases and pests have become an area of significant concern for cultivation and conservation of wild stocks alike: yield and quality of a crop might be greatly affected and sometimes, the losses due to diseases and pests are such that they jeopardize the economic viability of cultivation. Disease management methods and biosecurity are still in their infancy, especially as disease-causing organisms are most often poorly known, with limited options for robust and rapid diagnostics. Here, we describe a community-based, multilingual initiative that aims to foster the description of disease-causing organisms relevant to the seaweed industry worldwide and to underpin the conservation of wild seaweed genetic resources.


Introduction
For several decades, seaweed aquaculture has been growing at a rapid and sustained pace worldwide, nearing an increase of 70% in production and value between 2008(Fao, 2020. Seaweed cultivation has thus become an economic, social and environmental stake in China, many countries of South East Asia, Eastern Africa and South America, with Europe and North America rapidly following suite (Buschmann et al., 2017). The number of cultivated species is also increasing steadily Duarte, Marbá, & Holmer, 2007). Alongside this rapid extension, diseases are increasingly pointed as a burden to the industry's continued expansion and sustainability (Tan et al., 2022); though their impact varies depending on the cultivated species and local conditions, it is often accepted that diseases cause for losses from 15% to 30% of the potential yield, with local collapses also being reported (Msuya et al., 2014;Vairappan et al., 2008). A study conducted in the two biggest Korean Pyropia farms over several years suggested that any point in time, a fifth to a third of the crop is infected by one of several pathogens; overall, the lost yields and disease management method amounted to half of the running cost of the farm (Kim, Moon, Kim, Shim, & Klochkova, 2014). Likewise, various incompletely characterized pests and pathogens hamper the cultivation of Saccharina japonica in China (Ahmad et al., 2021) and the eucheumatoid species Kappaphycus and Eucheuma Ward et al., 2021).
Besides the economic impact of diseases in seaweed cultivation, pests and pathogens are also of ecological significance, particularly in the context of changing environmental conditions (Ferrari et al., 2021;Kumar, Zozaya-Valdes, Kjelleberg, Thomas, & Egan, 2016).
Overall, a great diversity of microorganisms trigger diseases on seaweeds. Amongst eukaryotes, the best known pathogens are fungi, oomycetes, phytomyxids, and other algae (Gachon, Sime-Ngando, Strittmatter, Chambouvet, & Kim, 2010). Many bacterial genera, such as Vibrio, Pseudoalteromonas and Flavobacterium (e.g., Egan, Fernandes, Kumar, Gardiner, & Thomas, 2014;Largo, Fukami, & Nishijima, 1995;Ward et al., 2021) have also been implicated. Though their aetiology is very poorly characterized, a diverse range of viruses are also able to infect seaweeds, and some are suspected to lead to mortalities (Beattie, Lachnit, Dinsdale, Thomas, & Steinberg, 2018;Easton, Lewis, & Pearson, 1997). In recent years, a striking number of novel macroalgal pathogens have been described from every corner of the world. Metabarcoding evidence has shown that their diversity and biogeography are vastly underdocumented (Badis et al., 2020) Their host range is barely known, both in farmed and wild stocks, which hinders the identification of reservoirs and thus, risk assessment for biosecurity purposes.
To underpin the sustainable development of the seaweed cultivation sector and inform conservation policies, it is increasingly important to understand the epidemiology and aetiology of seaweed diseases and accordingly, to develop practical tools for diagnostic. Yet, hardly any resources are available to academics, policy makers, or seaweed professionals on this topic; likewise, best practices for disease management and biosecurity policies are still very much in their infancy (Campbell et al., 2020). A difficulty towards the development of keys for diagnostic is that macroscopic symptoms of diseases are often generic, which makes them of little use to discriminate between pathogens; in some species (e.g., eucheumatoids), this issue is compounded by very plastic thallus morphology and pigmentation, so that visually similar syndromes such as ice-ice (i.e., thallus bleaching and rot) can be caused by very different pathogens, including bacteria and fungi. There is also a general lack of consensus on how to define some widely used terms, which generates confusion in the literature. Finally, it has become increasingly clear that many seaweed diseases are in fact multifactorial syndromes resulting from the combined expression of unfavourable abiotic factors with the proliferation of opportunistic pathogens; the observed macroscopic symptoms correlate with complex shifts in the microbiome, which makes the definition of a precise aetiology challenging (Dittami et al., 2021;Egan & Gardiner, 2016).
Here, we describe a web portal that aims to accelerate pathogen discovery and to produce accessible resources on seaweed diseases for scientists, professionals and policy makers alike. We hope that this will stimulate research for disease-resistant breeds, facilitate diagnostic, inform farm management methods and inform biosecurity and conservation policies.

The "my seaweed looks weird" web portal: what it is and what we do
The "My Seaweed Looks Weird" (MSLW) web portal provides a platform for reporting suspected macroalgal diseases and submitting physical samples or pictures thereof (Fig 1a; https://globalseaweed.org/?page_id= 889). Currently available in two languages (English (MSLW) and Spanish (MAVE; MI ALGA SE VE EXTRANA)) with plans to develop a Portuguese mirror, this portal signifies a collaborative effort between two laboratories (Scottish Association for Marine Science, Oban, UK and Instituto de Acuicultura, Puerto Montt, Chile). In order to document observations of unhealthylooking algae, the submitter is asked to fill in a short questionnaire with key information relevant to their sighting. No prior knowledge of seaweed diseases is required, and the user can request full confidentiality, for example if commercial sensitivities are involved. Conversely, we offer the possibility of a fully open collaboration, which is typically more suited for academic partners. Automatic notifications will be sent to the MSLW team upon each new submission; we will subsequently get in contact with the submitter in order to discuss the observation further. Typically, providing macrophotographic material of unhealthy-looking algae will first enable the MSLW team to ask a few targeted questions and establish a first preliminary assessment. Whenever relevant and feasible, our team will then explore the relevance of submitting physical a b  samples, according to the most adapted storage and fixation protocol, in full compliance with the Nagoya protocol (Smith, Da Silva, Jackson, & Lyal, 2017). The processing and analysis of unhealthy-looking algae is adapted on a case-by-case basis, and we use submitted samples to pilot novel protocols. Overall, we first aim to document macroscopic and microscopic disease symptoms (Fig 2). The microscopic analysis of samples relies on the availability of stains that bind to cell wall components or DNA to reveal pathogen structures (e.g., Gachon et al., 2017;Gerphagnon, Latour, Colombet, & Sime-Ngando, 2013). Based on this initial assessment, more targeted analyses can subsequently be envisaged, including molecular identification with barcoding and ultrastructural investigation with electron microscopy. Isolation and laboratory cultivation of candidate pathogens, albeit challenging, especially in case of obligate-biotrophs, can also be attempted, either on selective medium (bacteria, fungi; e.g., Vallet et al., 2018) or in co-incubation with healthy host algae (e.g., Strittmatter et al., 2016). This is particularly useful to capture various life cycle stages of the pathogen, to study host range and culture condition of the pathogen and examine host resistance (Gachon, Strittmatter, Müller, Kleinteich, & Küpper, 2009;Müller, Küpper, & Küpper, 1999). The results are then compared to existing literature and potentially, to other existing MSLW entries (Fig 2). A detailed report is then returned to the submitter. A few case studies are posted on the portal for the purpose of illustrating this process of sample analysis (https://globalseaweed.org/?page_id = 517). In case of doubt, we very much encourage the submission of material that documents macroscopic symptoms that can be mistaken for or combined with disease symptoms, such as grazing or abiotic stress.
So far, the MSLW web portal has received a rather small number of submissions which at least to some extent is attributed to the ongoing sanitary crisis. The majority of submitted cases relates to farmed seaweed, with occasional submissions concerning wild macroalgae. Beyond documenting the occurrence and aetiology The extend of analysis largely depends on the initial MSLW entry. Whereas observations in the form of GPS data and macroscopic diseaselike symptoms allow for a preliminary, putative symptom classification, the exact identification of pathogens remains largely impossible. Nevertheless, such primary symptom detection will be recorded and can be compared to existing literature and MSLW entries. The example image here shows macroscopic disease symptoms on Pythium-infected Porphyra sp. with small light to withish spots (a; scale bar 1 mm). Fixed samples allow several analyses of microscopy and molecular identification depending on the storage and fixation conditions initially applied to the sample. The example microscopy image shows the oomycete pathogen Pythium porphyrae growing hyphae (arrows) through the Porphyra host cells (b; scale bar 100 µm). Live samples in contrast can be for all sort of subsequent analysis like microscopy, molecular identification and the establishment of live cultures. The example image of the culture shows the oomycete pathogen Pythium porphyrae isolated on corn meal agar (c, scale bar 1 cm, inset 100 µm).
of known organisms, this initiative has already enabled us to initiate the characterization and taxonomic description of several pathogens previously unknown to science (e.g., Badis et al., 2019), with more in progress.

Our objectives
To the best of our knowledge, the MSLW web portal represents the first-ever online platform that offers free scientific expertise on the identification of seaweed-disease causing organisms. Whereas the microscopic observation of pathogen structures and life cycle stages provide crucial information on an initial diagnosis, the subsequent identification of pathogens with molecular methods is of high importance. Historically, a variety of diseases has been described based on morphological characteristics (e.g., Andrews, 1976), and there is evidence that re-addressing this body of knowledge with molecular barcoding is likely to identify new species and genera (e.g., Badis et al., 2020;Bernard et al., 2019). This might lead to the revision of currently accepted taxonomy, not only for the pathogen, but also for the algal host (Yang et al., 2020). The breadth of molecular methods that we use is largely dictated by the biological material at hand; it spans from Sanger-sequencing of barcode markers to metabarcoding or genome sequencing (e.g., Badis et al., 2019;Garvetto, Perrineau, Dressler-Allame, Bresnan, & Gachon, 2020;Zozaya-Valdes, Roth-Schulze, Egan, & Thomas, 2017).
As seaweed cultivation is developing in non-English speaking countries, we aim to provide this service in multiple languages, in order to increase its reach and to communicate with a global audience. In particular, our interfaces in Spanish and Portuguese aim at encouraging sample submissions from Latin America and accordingly, at supporting the development of aquaculture and conservation policies in this continent (Alemañ, Robledo, & Hayashi, 2019).
Our overarching long-term objective is to gather knowledge on seaweed diseases and pests, interpret data, and make resources freely available to scientists, seaweed professionals and policy makers in a meaningful format. All data received through the MSLW web portal are gathered in a structured manner and added to an internal database. In accordance with the guidelines of our funders, and assuming that the original submitter has granted us permission, we will endeavour to make their data and our findings available online in the form of an open-access database (Strittmatter et al.,in preparation;Fig 1b). For this reason, we only process samples in confidentiality if it allows us to gather useful intelligence, for example on relevant diseases in commercial facilities; however, our most labour-and moneyintensive investigations shall be focused on biological material for which the submitters have authorized us to release (or co-publish, if relevant) our findings.
We aim to illustrate the breadth of algal diseases, already described as well as novel pathogens, in a comprehensive database. In terms a participative effort is required in order to be able to capture the breath of macroalgal diseases in terms of pathogen diversity, biogeography and host range. As seaweed diseases are of global concern, we believe that the putting in place of such a web portal responds to the current needs for scientists, farmers and stakeholders. The accompanying open-access database will hopefully be enlarged over the coming years and serve as source of information for a large, diverse and global community.