Harmful Algae News

No. 59 - February 2018 · www.ioc-unesco.org/hab An IOC Newsletter on Toxic Algae and Algal Blooms

Content

25 years of HAN and IPHAB

Invited contributions ...... 1

DM Anderson A retrospective look at HAB cyst researchG Hallegraeff ( ) ...... 4 How do algal blooms kill finfish ( ) ...... 9 TheØ Moestrup IOC Taxonomic Reference List of Harmful Microalgae 25 YEARS ( P Hess) ...... 12 Algal toxins over the last 25 yearsT Wyatt ( ) ...... 15 Butterflies in Brazil (HAB events) ...... and monitoring 17 Harmful Algae News Alexandrium

was first published need for a communicationHarmful Algae channel News to Red tides of in early 1992 in response to requests strengthen networks and co-operation - - Gambierdiscusaffecting salmon in from the participants at a number ad of was obvious and Kamchatka, Russia ...... 20 hocIOC meetings and workshops on harm becameHarmful a core Algae element News in the IOC Harm in the Balearic ful algae, in particular the IOC-SCOR - ful Algal Bloom Programme. In- Islands ...... 22 Meeting in Rhode Island (USA) 2-3 ternational Marine Science started as an Light microscopy and qPCR November 1991, hosted by Ted Smay ­annex to the IOC UNESCO newsletter ‘ for ichthyotoxic microalgae da where the final draft of the Harmful ’ (IMS), with enumeration in Denmark ...... 24 Algal Bloom Programme was prepared. the enthusiastic supportHarmful of Algae IMS ­editorNews The Cawthron Institute At this time, e-mail was in its infancy, Gary Wright and assistant ­Michelle CultureISSHA’s CollectionCorner (CICCM) ...... 28 web sites still very simple but there was Turner. However, Forthcoming Events...... 29 demand for a channel to disseminate became independentHarmful Algae and eventuallyNews information about harmful algal events survived the IMS which ceased some GlobalHAB ...... 23, 30 and research, as well as to disseminate years later. has information on management/research been in publication for longer than any ...... 31 programmes, conferences, meetings other IOC newsletter and its viability etc. The field of HAB science was new,- reflects the sustained focus on harmful the community multidisciplinary and algae in the IOC programme, as well as- at a national level, often scattered be the continued interest by governments, tween many different institutions. The institutions, scientists and those im

Vladimir“The Decade Ryabinin, of Ocean Exec. Science Secretary, will be IOC a unique ten-year, global, large scale cooperative programme to seek urgently-needed scientific solution to support effective ocean management, stewardship and sustainable develop- ment” (continue next page) Harmful Algae News - pacted by harmful algal events. Since agement communityHarmful will Algae continue News to turned 20 years submit their news items and announce- old in 2012, it has been a web based- ments and that can e-newsletter which meant longer issues continue to serve all its readers includ were possible and back issuesHarmful easily Algae ac ing the International Society for the Newscessible. We are currently working on a Study of Harmful Algae. searchable index Harmful for all Algae News To subscribe to HAN, send an e-mail- issues. to “[email protected]”name with empty The start of subject line and in the text body: sub would not have been possible without scribe han e.g.: for Peter Black a dedicated professional editor, Tim —> “subscribe han Peter Black” or just Wyatt. Tim wasHarmful already Algae an experienced News - Henrik“subscribe Enevoldsen, han” Intergovernmental journal editor and author, and a main Oceanographic Commission of UNESCO, contributor to him Head, IOC Science and Communication self. In the early years he was assisted Centre on Harmful Algae by Yolanda Pazos, also at the Institute for Marine Investigations (IIM) in Vigo, Spain. Later, staff at the IOC Science and - Communication Ángeles Centre at the Spanish- Dear Readers Institute of Oceanography (IEO) in Vigo- The fact thatHarmful this year Algae we Newsare celebrat (Jorge Diogéne, Aguilera, Móni ing the 25th anniversary of the IOC- ca Lion and Cristina Sexto) provided in newsletter , makes- valuable support in the compilation and- me, as Executive Secretary of the Inter layout. Since 2012 Leif Bolding at the governmental Oceanographic Commis- Department of Biology at the Universi- sion of UNESCO, proud of the strategic ty of Copenhagen has been producing foresight of the Commission. Continu- the layout andHarmful providing Algae the News appropri ous publication of this newsletter shows ate server facilities. Tim Wyatt edited the interest in the subject by the coast 48 issues of plus al zone management community; is- some special issues over his 22 years evidence of successful delivery of new as Editor. When Tim decided to retire knowledge by the interdisciplinary re as Editor we were lucky to have Beatriz search community; and is the result of- Reguera (IEO Vigo) and Eileen Bresnan effective prioritization of IOC work by (Marine Scotland) willing to take over- its Member States. The harmful algal re and continue the same high standard.Harmful search domain is an example of ocean AlgaeNo doubt News these standards have stimu science which is of primary importance- lated submissions and made - for sustainable development. Indeed, what it is today. We hope people’s health, nutrition, availabil the international HAB science and man ity of fresh water, and many economic interests are directly affected by the emergence of harmful algae. Successful research on and predictions of harmful algae demonstrate that oceanography is becoming more and more capable of responding to people’s needs. To make decisive and major improvements in making ocean observations, research,- sustaining services and responding to growing societal needs is the main ob- jective of the United Nations Decade of Ocean Science for Sustainable Devel opment (2021-2030). The Decade is going to shape much of the activity of Key people in the early days of the HAB programme. From top to bottom: Ted Smayda (USA), the IOC and its partners in the years to host of the IOC-SCOR Workshop, Newport, October 1991, where the HAB programme was out- come. Let me invite you all to engage lined (photo DM Anderson); Tom Osborne, IOC Technical Secretary when the programme was Vladimirand contribute. Ryabinin, Executive Secretary, launched; Bernd Dybern (Sweden), 1st IPHAB Chair, 1992, and last, Adriana Zingone (Italy), Intergovernmental Oceanographic Commis- 2nd IPHAB Chair during the III IPHAB meeting in June 1997, Vigo, in the middle with Henrik sion, UNESCO, Paris Enevoldsen (IOC) on the left and Vice-Chair Rhodora Azanza (Philippines) on the right.

HARMFUL ALGAE NEWS NO. 59 / 2018

2 Intergovernmental Panel on Harmful Algal Blooms also turned 25!

Harmful Al- galDuring News 1992, the same Year as theInter IOC- governmentalpublished the firstPanel issue on Harmfulof Algal Blooms , it also established an

(IPHAB) which has met every second year since it was formed. The Panel is composed by IOC Member State- representatives and representatives of organizations working on different as pects of HABs. The Panel identifies and decides on priorities for international cooperation on HABs and also works to identify resources to facilitate the diverse work schedule to solve some of the real problems caused by harmful- algae. - Over the last 25 years the IOC Inter governmental Panel on HAB has initi- ated a large number of initiatives and activities leading to training opportu nities, manuals,‘Oceanography’ guides, projects and cooperative research. A special issue of the journal includes a paper on the history and role of the IOC HAB Programme in the developmenthttp://dx.doi. of org/10.5670/oceanog.2010.25international HAB science. The paper can be downloaded at . We here wish to pay a tribute to those who have chaired the Panel, and their Vice-chairs, the past 25 years and- their contribution to developing the HAB research and management com munity at a global scale. From top to bottom: Beatriz Reguera (Spain) and Phil Busby (New Zealand), elected as IPHAB Chair and Vice-Chair in 2002; Flowers to the outgoing Chair (Adriana Zingone, Italy) (left) and Vice- Chair (Rhodora Azanza, Philippines) (right); Leonardo Guzmán (Chile) (mid- dle), Chair from 2007 to -2011 welcomes the newly elected Chair, Rob Magnien (USA) (left) and Vice-Chair, Gyres Usup (Malaysia) (right) for the next term; Bottom: Results from last elections, at the 12th IPHAB Session, Paris, April 2015

HARMFUL ALGAE NEWS NO. 59 / 2018

3 A retrospective look at the early days of HAB cyst research, and a look to former used to disaggregate sediments and the latter to separate size fractions the future containing cysts. In his paper in the 1974 conference proceedings and in - other papers published near that time- [1-3], Wall highlighted the important Harmfulth Al- - - gaeOn thisNews occasion of the 25 anniver that came shortly before and after. roles that cysts likely played in dinoflag sary of the publication of The conference was very small com ellate bloom dynamics, including deter- , several of us were asked to pared to the current ISSHA meetings, mining where and when blooms might look backwards in time to some of the with approximately 100 participants originate, allowing survival through en- earlier days of HAB science. One area from four countries versus 500 or more vironmental extremes,Despite these and facilitatingseemingly of study that has been a major part of participants representing 50 or 60- importantgenetic recombination considerations, through the sexualencyst- my own research programme and that countries at our currentAlexandrium meetings. catenella The ment-excystmentity. He told us “ cycle in dinoflagellates of many others in our community is the focusKarenia of that brevismeeting was almost entire has not been expressly studied in rela- role of cysts and resting stages in the ly on bloomsGymnodinium of breve - tion to any specific red tides or related bloom dynamics of HAB species. Here, and (the species was then- instances of paralytic shellfish poison- I offer a personal perspective on the called Dinophysis), as the dis. ing. Details of the cyst cycle have been early stages of development in that field coveries of diarrhetic shellfish poison accumulated independently of red tide and a brief look towards the future as Pseudo-nitzschiaing (and its linkage to spp ) research well. My apologies at the outset if there and amnesic shellfish poisoning (and - are omissions of people or findings that ) were 10-15 years in- …..” [1]. This contribution and should have been included – this is not the future.Gambierdiscus The ciguatera toxicusfish poisoning other papers at the time show consid meant to be a scholarly synthesis, but- syndrome was known, but the identifi erable foresight that was fully validated rather a personal retrospective. cation of as the through a sharp increase in the cyst- harmfulTo set algal the stage,bloom let’s go back near causative organism was also several based HAB research of the subsequent ly fifty years to a time when the term- years away. Among the contributions at- decades. did not exist and that meeting, several are noteworthy in- Another talk at that conference was- when researchers worked individu the context of the theme of this narra equally insightful, this one by Karen ally or in very small teams in scattered- tive. The first was by David Wall, a geol Steidinger (Fig. 3) (“Basic factors influ locations without any major research ogist at the Woods Hole Oceanographic- encing red tides [4]”). In her talk and programmes or international initia Institution (Fig. 2). In the late 1960’s paper, Karen outlined the fundamental tives. In 1972 when I was just starting and early 70’s, Wall (working with Bar stages of blooms, focusing on initiation,- graduate school, there wasAlexandrium a massive rie Dale at the time) discovered that- transport, and“The development. recent investigations Like Dave catenellaNew England red tide caused by the the fossilized cell walls of organisms ofWall, Dr. sheDavid also Wall highlighted ……lend credence the impor to dinoflagellate we nowGonyaulax call tamaren- thought to be extinct (called hystri thetance speculation of cysts: that pelagic, toxic dino- sis G., excavatabut which then had multiple chospheres) were in fact dinoflagellate- flagellate blooms might originate from names, including cysts, and that many of the forms that and . This outbreak dealt they were using in paleontological re a devastating and unexpected blow to constructions were- the New England region of the United- still living in the States, causing shellfish closures along ocean and produc the coasts of multiple states. Consider ing cysts that could- able attention by the press and public be germinated to covered this new and worrisome First threat Inter- establish taxonom nationalto public Conference health and onfisheries Toxic Dinoflagel resources- ic affinities. (This lateand ultimatelyBlooms motivated the same line of work was concurrently- in Boston, Massachusetts in being pursued by 1974. Bill Evitt and col- At the time of that conference, I was leagues at Stanford a long-haired graduate student in the University). In or Civil and Environmental Engineering der to isolate and Department at MIT (Fig. 1), searching- germinate living for a thesis topic. People often ask me cysts from modern how someone in an engineering depart sediments, Wall ment could end up a biologist working and Dale developed- on HABs, and the answer to that lies in a technique that is Fig. 1. Don Anderson as a graduate student in the Civil and Environ- mental Engineering ­Department at the Massachusetts Institute of part with the 1972 New England red used to this day in Technology (MIT), 1975 tide, but also with the 1974 conference volving sonication and a series of events and discoveries and sieving – the HARMFUL ALGAE NEWS NO. 59 / 2018 4 dormant stages and that these stages might be associated with certain bottom sediments. This then brings up the ques- ultimatelyAlexandrium divided tion, if benthic resting stages of certain catenellato produce cells- dinoflagellates actually “seed” coastal of red tides, are there localized areas of ac- . I was al cumulation, or what we could call “seed- ready aware of the beds”?...... The possibility of benthic seed importance of this populations and seedbeds for at least discovery after the some Gonyaulax (=Alexandrium) and talks and papers of Pyrodinium has a much higher probabil- Wall and Steidinger, ity than for Gymnodinium breve.……. It and now I had proof would seem that this avenue of research that at least some of should have a high priority among phy- their speculations toplankton systematists and ecologists were valid. The door was suddenly Fig. 2. David Wall, an early pioneer in dinoflagellate cysts, collecting .” open to some very cysts from settling trays in Woods Hole in 1974 As a graduate student sitting in the exciting and new audience, I was motivated by these talks work with livingAlexandrium - and papers and immediately started to- cysts, and this new look at my own research from a new perspective on bloom dy decreased enough to be favorable for- perspective. At the time, I was in a wa namics totally changed the directions of germination remainedAlexandrium a mystery. Only ter chemistryAlexandrium laboratory doing trace- my thesis research. Furthermore, I had in recent years have we begun to under metal (copper) sensitivity experiments- found perfect study locations for my stand that mature cysts- with my cultures, reason work – the Cape Cod salt ponds where can re-enter dormancy – one cause may Alexandriuming that there was some chemical pro these blooms occurred regularly every be unfavorably warm summer tempera- cess that could explain the link between- year, and in which they were localized tures. But we have also learned that- blooms, river runoff, and due to restricted tidal exchange with exit from that dormant state into quies low salinity coastal waters (I still be nearby coastal waters. We continue to cence (a state in which cysts will germi lieve that this is the case, but I dropped work in these systems today – they are nate if conditions are favorable) occurs this line of inquiry and it has not been- essentially natural mesocosms. Several in response to a quantifiableA. catenella amount pursued to any significant extent since). publications ensued [5,6], accompanied- of chilling. By quantifying the duration- Many of my cultures contained round by a paper by Barrie Dale [7] who had and severity of cold, tracks ed, non-motile cells,A. whichcatenella I thought - left Woods Hole and was working inde the passage of winter, delaying germi could be cysts, but no cyst stage had yet pendently on this same species using nation until spring when the outlook is been described for . Fortu sediment from Norway. more favorable for bloom success. This nately for me, Woods Hole was a short AlexandriumIn one of these papers [6], I was able groundbreaking work by one of my- drive away for me, so I was able to bring- to document the extreme seasonality of students, Alexis Fischer, demonstrates my cultures to Dave Wall. After a quick blooms in the salt pond,- that dinoflagellate cysts possess physi examination, he gave me the disap raising fundamental questions about ological behavior that is similar to that pointing news that these were not what the mechanisms underlying the pat observed in terrestrial plants where a he would term true cysts, but rather a terns of excystment and encystment. I period of chilling is needed for optimal temporarypellicle cysts stage with much thinner characterized the mandatory dormancy Alexandriumseed germination, bud flowering, or walls (he and I eventually named these interval, which begins at cyst formation bulb emergence [9]. We now know that “ ” after the resistant layer- and can last several weeks to months cysts can cycle between that surrounds them [5]). Next, one of until maturation is completed. Cysts quiescence and dormancy multiple- those pivotal moments in a career oc Alexandriumwere unable to germinate during this times as seasons and years progress, curred in which luck or good fortune interval, thus explaining the absence of and equally importantly, that the fre played a role. Instead of just sending me- cells in the water column quency and timing of the cycling may back to my lab with this negative news, in the months followingAlexandrium each spring well be determinedó by thePyrodinium history of theba- Dave suggested that we sample a near bloom. What remained a mystery was hamensechilling those cysts experience. Recent- by salt pond where PSP toxicity had why the bulk of the cysts work by Cary L pez on been recurrent in the years after the in bottom sediment did not germinate- is revealing a similar environ big 1972 bloom. We collected surface later in the summer or fall or even early mentally-induced cycling behavior in- sediments with an old plankton net that winter after they had matured, but in tropical dinoflagellate cysts. was dragged across the bottom, and stead remained in a resting state until Those early days as a graduate stu Dave demonstrated the sonication and the next spring. One part of the answer- dent and then as a postdoc were timesAlex- sieving method. In short order, we were was found some years later when we- andriumof rapid discoveries, as nothing was looking at a number of unknown cysts- showed that high temperatures can in- known about the distribution of types that I set about trying to isolate hibit cyst germination [8]. Yet, the ab cysts in the region, or of the- and germinate. One produced an elon sence of a major bloom in the late sum role of temperature, nutrients, light, gateHARMFUL cell ALGAE (Fig. NEWS NO.4) 59 that / 2018 after germinating, mer and fall when temperatures had oxygen, and other environmental pa 5 -

day, we still do not ate when one is studying vertical pro know if there are files of dead, empty, or fossilized cysts- other factors at in sediments dating back hundreds or work – perhaps a thousands of years, this type of param density-dependent eterization is not useful for biological or quorum-sensing studies in which one wants to map out type of response, the distribution and abundance of cysts or even a response- and estimate the number of germinated to the presence of cells inoculating the overlying water grazers or para column. To address the latter topic, we sites. Exploration needed to know how many cysts were of this response has present per square meter of sediment, long been limited so we started doing experiments to see- by the constraints how reproducibly we could quantify associated with- cysts per cubic centimeter or ml of sedi laboratory cultures,- ment. Our results were consistent and but now we are for scientifically appealing so we published- tunate to be enter the method [13] and began to use it in ing a new era of in- the salt ponds and other nearshore lo situ investigations- cations. This approach was not well- thanks to biosen received, however, as I was repeatedly sors like the Imag criticized at conferences when these re ing FlowCytobot sults were presented. Geologists argued (IFCB), that allow- that the water content and lithology of us to observe the surface sediments differs sufficiently- encystment pro- from site to site, or from layer to layer,- Fig. 3. Karen Steidinger, Penrose Conference on Fossil and Modern cess underwater in- and that normalization of cyst abun Dinoflagellates, 1978 the field with ex dance to a volume of sediment intro traordinary resolu- duces substantial errors compared to- tion. Among many the dry weight approach. I remember rameters on the formation, deposition, other recent findings, IFCB observa being told at an international confer and germination of cysts. One of several tions by Mike Brosnahan [11,12]A. catenel have- ence that my results and the population- challenges at the time was that we were ladocumented mass gametogenesis (75 – dynamics models based on them were- unable to produce true hypnozygotic- 90% of all cells) at the end of essentially meaningless! This disagreefuture resting cysts in our cultures. There was blooms under conditions that do not- studiesment went use standardized even further, methods as one publibased good reason to believe that sexual appear to be nutrient-limited. I believe oncation measurements [14] recommended of cyst concentrations that “ ity was involved, so compatible mating- that the tools are now in hand to re per gram of dry sediment - types needed to be combined. Further, solve the longstanding mystery of what nutrient limitation seemed to be a trig- induces cyst formation in the field and ”. This recom- ger to initiate the process. We quickly suspect that the final story will be much mendation was repeated in training learned that dinoflagellate species var more complicated than simple nutrient manuals for those working with liv ied considerably in the Scrippsiellaease with which tro- limitation. ing dinoflagellate cysts [15]. Despite choideathey could be induced to form cysts in Not long after our early successes this strong and vocal opposition, I was- cultures. Species like with sexual induction and cultures, still firmly convinced that cysts could producedAlexandrium prolific numbers my attention shifted to studies of cyst be quantified per unit volume of sedi of cysts without special precautions, distribution and dynamics in the field. ment, and so I initiated a study over whereas our cultures were We were documenting the abundance multiple years in which we quantified far more recalcitrant. Eventually, with and dynamics of vegetative cells in the cysts both ways (i.e., cysts per gram dry deliberate modifications to the content plankton during blooms, so I wanted to weight and cysts per ml). We did this in- of the culture medium as well as the- quantitatively link those observations the Gulf of Maine across a wide range- manner in which it was prepared (i.e., to cyst abundance and distribution in of sediment types and cyst concentra minimizingAlexandrium contaminants and precipi the sediments before and after those tions, and found a strong and statisti tates) we were finally able to produce- blooms. At the time, however, studies cally significant relationship between cysts in cultures [10], of dinoflagellate cysts were still heavily- the two parameters, for the top cm of opening yet another door to new stud influenced by geologists working with sediment (Fig. 5). The relationship for ies and discoveries. - sediment cores. Typically, those inves the top three cm of sediment is even It is clear that nutrientA. catenella limitation tigations enumerate cysts at different stronger. This of course does not mean- can induce sexuality and cyst forma depths in cores, expressing results in- that this relationship will apply in every tion in cultures of and terms of cysts per gram of dry sediment. location, but it does show that the hy other dinoflagellate species, but to this Although this approach is appropri pothetical concernsHARMFUL raised ALGAE NEWS about NO. 59 this/ 2018 6 approach did not stand up to data at my study locations. I raise this issue in this narrative because I want to correct what I feel are unjustified recommendations that may prevent those working on cysts- from obtaining the type of biological data that can advance our understand ing of certain types of HABs. With the- proper precautions, we can map out cyst distributions in a way that is com plementary to the manner in which we study the vegetative stages in the water column. There’s no question that both populations are patchy and difficult to adequately represent with limited sampling, but it can be done. Knowing Fig. 4. Germinating Alexandrium catenella cyst. (Photo: Don Anderson) the cyst abundance per ml of surface- sediments allows the distribution to - be mapped in a way that has biologi cal meaning, as has now been seen in for cyst-forming species. - ment layer, as this would allow a sim many studies. If one assumes that the My guidance to those working on- ple calculation that would also provide cyst germination flux comes from a- cysts in natural sediments is to demon- cysts per unit volume data. - thin layer at the sediment surface of a strate that you can sample surface sedi Armed with the knowledge that was defined thickness, it is possible to cal- ment reproducibly, and that the distri possible to map out cysts over large ar culate germling emergence per square butional data obtained makes sense in- eas, my team set about to do this in salt meter, and thereby quantify the inocu the context of bathymetry, currents, and ponds, bays, and open coastal waters. lum to the overlying water column. This- sediment type. I hope that those work- These efforts confirmed Dave Wall and is simply not possible if the cysts are ing on dinoflagellate cysts recognize Karen Steidinger’s original speculation expressed per gram of sediment, un the validity and value of the two differ – that areas of accumulation (seedbeds) less additional measurements are made- ent approaches for cyst enumeration existed and were a critical element in that determine the density (grams per – cyst number per gram dry weight for bloom dynamics. Some seedbeds were ml of the sample). Once the cyst distri paleontological reconstructions of past small, and others huge – like the ones bution is expressed in terms of area, it- populations and sediment transport we documented in the Gulf of Maine is then possible to use laboratory- or modeling, and cysts per unit area or (see figure 5) that covered2 an area as field-derived germination rates to cal- unit volume for studies of the dynamics large as 22,000 km , varying 3–4-fold culate the germination input to a body of living cysts. Another option would be- in area and 10-fold in cyst abundance of water, an approach that is fundamen to measure cysts per gram dry weight among years [16]. Those studies also tal to bloom dynamics modeling efforts and determine the density of each sedi demonstrated that the Gulf of Maine system has only two seedbeds with the bathymetry, sediment characteristics,- currents, biology, and environmental- conditions necessary to persist for dec ades or longer. The value of these map ping studies was even more evident when strong positive correlations were confirmed between the abundance of cysts in surface sediments and the size of the blooms and resulting toxicity in the subsequent year [16,17]. Our next challenge was to measure or estimate the flux of emerging cells that inoculates blooms. Efforts by my lab but in particular those of Ishikawa, Naksuike, and others [18,19] have yielded some promising devices and approaches that consistently indicate Fig. 5. Significant relationship (p < 0.0001) between cysts per cm3 and cysts per gram dry weight in the top cm of sediment at 85 stations throughout the Gulf of Maine. From [16] that germination is occurring in only a thin veneer a millimeter or so thick at HARMFUL ALGAE NEWS NO. 59 / 2018 the sediment surface, leaving the large 7 - - number of cysts in subsurface layers the methods and approaches to study than 60 years, the study of living dino unable to germinate or emerge, pre- cyst dynamics in a manner analogous flagellate cysts remains a vibrant and sumably due to lack of oxygen or to to the way we study bloom dynamics important element in HAB research,- the tortuous pathway posed by sedi in surface waters. It was not easy, as and I am grateful for the foresight and ment grains and detritus. Many might there was strong opposition to some guidance of David Wall, Karen Steiding think that major storms and waves can of the methods that were used, but the er and others, and to have been part of erode significant layers of sediment and scientific process and good data have that evolution myself. transport cysts long distances, but here overcome the speculation. There are- References again, our studies in the Gulf of Maine certainly challenges and uncertainties 1. Wall D 1975. In: VR LoCicero (ed). Toxic have shown that other than in very ahead in the study of cysts and life his Dinoflagellate Blooms (Proceedings of shallow waters, even major storms only tory dynamics, but significant progress the International Conference, Massachu- erode a millimeter or less of sediment has been made and much knowledge setts Science and Technology Foundation, - Wakefield, MA), pp 249-256 [20] and that the cysts and other eroded gained. The future looks bright for- 2. Wall D 1971. Geosci Man 3: 1-15 material don’t travel very far before set future discoveries, facilitated by our 3. Wall D & B Dale 1968. Micropaleontol- tling again [21]. Specifically, the depth growing recognition that marine dino ogy 14: 265-304 of sediment eroded ranged from about flagellate cysts have much in common 4. Steidinger KA 1975. In: VR LoCicero (ed). st 0.05 mm at a sandy 70 m deep station, with terrestrial plants and that their Proc 1 Intern Conf on Toxic Dinoflagel- late Blooms (Mass Sci Technol Foundn, to about 1.2 mm in clayey-silt sediment distribution and abundance can be Wakefield, Mass), pp 153-162 at 250 m [20], with the majority of the quantitatively mapped – this valuable 5. Anderson DM & D Wall 1978. J Phycol 14: resuspended material remaining within information will enable us to estimate 224-234 20 km of the source locations [21]. deposition and germination fluxes. We 6. Anderson DM & FMM Morel 1979. Estuar I could go on, but lack of time and clearly need continued study to help Coast Mar Sci 8: 279-293 - 7. Dale B 1977. Sarsia 63: 29-34 space argues that I bring this narrative elucidate the mechanisms that trigger 8. Anderson DM & K Rengefors 2006. Lim- to a close here. As I look back, I am for sexuality and cyst formation, and that nol Oceanogr 51(2): 860-873 tunate and grateful to have been part of will likely involve observations using 9. Fischer AD et al In rev. Protist. an era of discovery that has developed in situ sensors like the IFCB. After more 10. Anderson DM et al 1984. J Phycol 20: 418-425 11. Brosnahan ML et al 2014. Deep-Sea Res Pt II 103: 185-198 12. Brosnahan ML et al 2015. Limnol Ocean- ogr 60(6): 2059-2078 13. Anderson DM et al 1982. Limnol Ocean- ogr 27: 757-765 14. Dale B 2000. The Science of the Total Environment 264: 221-233 15. Matsuoka K &Y Fukuyo 2000. Technical Guide for Modern Dinoflagellate Cyst Study (WESTPAC-HAB/WESTPAC/IOC, Tokyo), 30 pp + 17Figs + 7 tables 16. Anderson DM et al 2014. Deep-Sea Res Pt II 103: 6-26 17. McGillicuddy Jr DJ et al 2011. Limnol Oceanogr 56(6): 2411-2426 18. Ishikawa A et al 2014. J Plankton Res 36(5):1333-1343 19. Natsuike M et al 2017. Harmful Algae 62: 52-59 20. Butman B et al 2014. Deep-Sea Res Pt II 103:79–95 21. Aretxabaleta AL et al 2014. Deep-Sea Res Pt II 103: 96–111

Author Donald M Anderson, Cooperative Institute for the North Atlantic Region (CINAR), Woods Hole Oceanographic Institution, Mail Stop 32, Redfield 332, Woods Hole MA 02543-1049 USA

E-mail: [email protected] Fig. 6. Map of Alexandrium catenella cyst abundance in the Gulf of Maine. This image depicts a multi-year (2004–2011) arithmetic mean (cysts cm3) of the surface (0–1 cm) sediment- layer. Two seedbeds are clearly visible -one at the mouth of the Bay of Fundy in the north, and the other in mid-coast Maine to the south. From [16]

HARMFUL ALGAE NEWS NO. 59 / 2018

8 How do algal blooms kill finfish and - how can we mitigate their impacts? cause cells to exude ichthyotoxins and/ or cause cell lysis are critical, and ich thyoxicity by these genera tends to be more variable. - Algal blooms, water discolorations and fore also have not been a priority for The first point of attack by all above- their association with fish kills have seafood regulators. Scientific progress algal groups are the fish gills (Fig. 2), re- been recorded“all the since waters historic that times, were in such the has been hindered by the use of widely sulting a generalized necrotizing degen riveras the were description turned to in blood. the BibleAnd the (1000 fish different bioassay systems and lack of eration of the epithelium of the second thatyears was BC) in the river died; and the river chemical analytical methods to quantify ary lamellae with associated sloughing,- stank, and the Egyptians could not drink and characterizeArtemia ichthyotoxinsDaphnia from, often accompanied by swelling of the of the water of the river” seawater medium. Assay systems have primary lamellar epithelium and con included or assays the gestion of branchial vessels [3]. Fish (Exodus 7:- use of fish or mammalian erythrocytes, gills tend to respond in a singular way 20-21). In this case, a non-toxic bloom-- and a wide range of juvenile or adult and gill pathology is remarkably similar forming alga became so densely con fish (damselfish, sheepshead minnow,- for different ichthyotoxic algae and in- centrated that it generated anoxic con mountain minnow, zebrafish, salmon, different fish species [3, 6-9]. Once the ditions resulting in indiscriminate kills sea bass) tested under different expo fish gills are compromised, algal neu of both fish and invertebrates. Similarly, sure regimes. The novel application of rotoxins if present can penetrate the- water discolorations and massive fish a standardised and highly sensitive and blood stream, cause fish behavioural kills in Florida coastal waters have been reproducible rainbow trout RTgill-W1 changes, and loss of the blood haemo reported by Spanish explorers since the cell line assay in Australian, Chilean, globin’s oxygen binding affinity [2]. th Karenia brevis - 15 century (now known to be caused- Danish, Mexican and US laboratories Several competing theories have by the dinoflagellate ). has allowed significant progress to been proposed as to the precise mecha Finfish held captive in intensive aqua be made in the past 10 years culture systems are extremely sensitive [4,5]. This assay has been to HABs, and the impact of fish-killing automated in a plate reader algal blooms on human society thus measuring cell viability dyes,- has been exacerbated by the increaseTi of- and been successfully applied lapiafinfish aquaculture. Shilo 1967 [1] first also to screen freshly collect- investigated problemsPrymnesium for the Israel parvum ed natural seawater samples pond aquaculture industry caused from fishkill events in Austral by the haptophyte , Whatian and is Korean the precise waters. but researchChattonella efforts marina to understand fish mechanism causing fish killing mechanisms intensified in 1973 gill damage? when raphidophyte blooms (Fig. 1) devastated yellowtail- aquaculture in the Seto Inland Sea [2] While all high biomass algal Karenia mikimotoi - 3 -1 and in the 1980s when the dinoflagel blooms (>100 x 10 cells L ) late started to im can cause mechanical stress pact on the fledgling salmon farming- to the sensitive gill tissues of industry in Norway [3]. fish and trigger excess mucus Fish-killing algal species are re production, of much greater sponsible for much greater economic concern for the aquaculture- impacts (summarized in Table 1) than industry are the Cochlodinium,highly potent, HAB eventsPseudochattonella leading to seafood biotoxinAlex- Karenia,taxonomically Chattonella, unrelated Pseu flag- andriumcontamination. catenella The 2016 fish kills in dochattonella,ellate groups Heterosigma Chile by and Prymnesium. - which led to losses of US$800M triggered social unrest. It and One fea is therefore surprising that progress ture that these algal groups in our understanding of how HABs kill have in common is that they fish has been so slow. Financial support are all fragile cells, which can Fig. 1. Mass mortality of (above) yellowtail fish in the Seto Inland Sea in Japan (photo courtesy Prof T.Okaichi) for and research interest in HABs by the- lyse even upon impact on the- and (below) blue-fin tuna in South Australia (photo fish farm industry has often been short- gills of fish, especially during- courtesy Barry Munday), both caused by Chattonella lived, and ranked lower than invest the end of blooms, or condi marina blooms. Higher potency of Australian blooms (kills occurred at 66,000 cells/L) compared to Japan Kareniaments in brevis fish husbandry, nutrition, and- tions of osmotic stress or durKar- disease control. With the exception of ing upwelling. For less fragile (500,000/L) is attributed to higher sensitivities of tuna lodinium Al- but also higher ichthyotoxicity by Australian high-light , so-called fish-killing tox- exandriumfish-killing algae such as adapted algal strains [5]. ins or “ichthyotoxins” are not known to or the armoured haveHARMFUL human ALGAE NEWS health NO. 59 / 2018 impacts and there the conditions that 9 Table. 1. Economic losses from algal blooms for finfish aquaculture in different parts of the - world EPA in the presence of ROS, this in HAB species Country Financial Losses creased the potency of EPA by up to 15- Chattonella USD500M, 1973 fold (Fig.3). Similarly, DHA in synergism- Japan Heterosigma USD135M,1980/90 with ROS became 9 times more ichthyo Cochlodinium Korea, China USD95M, 1995 toxic [11]. The precise nature of the li- polykrikoides Canada USD2M, 1999 pid peroxidation products generated remains poorly known, and new ana Heterosigma USD35M, British Columbia lytical methods for the detection of such Chaetoceros 1980-90 compounds in seawater during fish kills Heterosigma New Zealand NZD12M (e.g. using SPATT collectors) are much Karenia digitata Hong Kong USD32M, 1998 needed. Karenia mikimotoi Norway USD6M, 1988 Role for true phycotoxins? Karlodinium Alexandrium catenella USD60M, 2002/ Pseudochattonella/ Chile USD800M, Jan-Mar 2016 The dinoflagellate is A. catenella karlotoxins known to produce chemically well-- Chattonella South Australia AUD45M, 1996 defined linear polyketide , Karenia Tasmania AUD3M, 2003 which at ecologically realistic concen trations can account inPrymnesium full for their fish- - killing potency [17]. While prymnesins from the haptophyte have nism of how algae kill fish. These in proximately 3ppm [7,12]. This is short attracted considerable interest [18]- clude: free fatty acids (FFA), reactive- however of what dense algal blooms it remains to be shown whether these oxygen species (ROS), and phycotoxins would generate. compounds can account for theirgymnocin ich Reactive Oxygen Species such as brevetoxins, karlotoxins, gym thyotoxicKarenia potency. mikimotoi Similarly, cytotoxic (ROS). nocins, or varying combinations of all chemical compounds such as - previous compounds. from remain tobre be- Polyunsaturated fatty acids - vetoxinsshown to play a quantitativeKarenia role inbrevis ich (PUFA) The role of Reactive Oxygen Species thyotoxicity [19]. Polyether ladder - Prym- in ichthyotoxicityHeterosigma has longCochlodinium been sug from the Florida nesium gested from whole fish experiments- are well known as the cause of Neuro Shilo working with fish-killing with and , toxic Shellfish Poisoning in humans but pointed out how ”lipid micels”- where application of ROS mopping en purified brevetoxin PbTx2,3 exhibited released by the algae can impact on the- zymes such as catalase and peroxidase limited ichthyotoxicity against RTgill fish Chattonella,gill lamellae and interfere with os significantly improved fish survival [13,- cells [5], suggesting that peroxidation moregulation [1]. Okaichi, investigat 14]. JapaneseChattonella researchers led by Oda- pathways may need be invoked in fish- ing focused on free fatty- [15] focused on the fish-killing raphido kills. The production of brevetoxins by acids damaging fish gills [2]. This was phyte which is a potent pro raphidophytes is disputed [20]. Neuro Kareniapursued mikimotoiby French researchersGyrodinium includ au- ducers of ROSKarenia notably whenAlexandrium algal cells active compoundsHeterosigma (but not brevetoxin) - reoluming Patrick Gentien [10] working with catenellaare ruptured. Several other fish-killing have recently been claimed also for the (as algae suchHeterosigma, as Karlodiniumand raphidophytesaxitoxins [21]. Mar ), who identified free fatty acids- Prymnesiumare also strong ROS producers,- dones [11] found no evidence for a role such as OPA (octadecapentaenoic acid) but for and of in fish gill damage, which- and EPA (eicopentaenoic acid) as hav ROS on its own cannot ex instead could be explained by DHA and ing the highest ichthyotoxic potency.- plain ichthyotoxicity. Using a xanthine- ROS synergism. Using the fish-gill ich Mardones [11] Alexandrium also confirmed catenella. the xanthine oxidase chemical Chattonella reaction to thyotoxicity assay, karlotoxin and DHA ichthyotoxicity by DHA (docasahexae generate superoxide at concentrations- could account for fish kills in their own noic acid) from equivalent to fish-killing -, right, but not EPA, STX, PbTX, OPA nor Paradoxically these compounds are- Marshall [7] demonstrated that super OTA. Synergisms between DHA x ROS well-known to have a beneficial effect oxide on its own does not kill fish. Simi and EPA x ROS were able to explain fish for human health, notably heart dis Heterosigmalarly, Trick and co-workers [16] showed- Implicationskills, and so could for fatty mitigation acid aldehydes. of ease, when consumed in moderation- that hydrogen peroxide produced by fish-kill events and used to replace saturated fat. These did not explain ichthyo - PUFA compounds are prone to oxida toxicity. ROS also exhibited negligible tive degradation, in which free radicals Synergismimpact in the betweenRT fish gill assayROS [5].and Several different strategies are current “steal” electrons from the lipids in cell- free fatty acids ly practised in the fish farm industry membranes, resulting in cell damage. In to mitigate the ichthyotoxic effects of- whole fish experiments it has been con- HABs. These include: cessation of fish vincingly demonstrated that exposure- Pursuing the role of ichthyotoxicity by feeding, towing away of cages from af to OPA and EPA did cause fish gill dam EPA, Marshall [7] demonstrated that fected areas, perimeter skirts to protect age at concentrations in seawater of ap when damselfish were challenged with against algal surfaceHARMFUL slicks, ALGAE aerationNEWS NO. 59 / 2018or 10 - -

emergency harvest opera the proceedings of that meeting. Nu- tions. To prevent the build-- merous PhD students worked with me up of histamines, fish should on fish-killing algae over 20 years, in be kept alive as long as pos cluding Juan Dorantes-Aranda, Graeme- sible during harvesting. This- Lush, Judi Marshall, Jorge Mardones, can be achieved by diluting Ben Mooney, and Andreas Seger. I ben algal concentrations via air- efited from valuable collaboration with lift upwelling, or by targeted the late Barry Munday (fish pathology), in-pen emergency applica Peter Nichols (lipids), Prof David Waite tion of clays [23] that mop- References(ROS) and Prof Al Place (karlotoxins). up ichthyotoxins at clay 1. Shilo M 1967. Bacteriol Reviews 31: 180- loadings significantly low 193 er than those considered 2. Okaichi T 1983. J Oceanogr Soc Japan 39: harmful to benthic marine 267-278 Fig. 2. Diagrammatic representation of algal bloom medi- 3. Roberts RJ et al 1983. J Mar Biol Ass UK invertebrates. 63: 741-743 ated fish kills, involving rupture of fragile algal cells to Laboratory experiments release a cocktail of Reactive Oxygen Species (ROS), Free - 4. Dorantes-Aranda JJ et al 2011. Harmful Fatty Acids (FFA) but which rarely involves true Phyco- with fish gill cell lines or Algae 10: 366-373 toxins [modified after 7]. The chemical structures of two even whole fish tank ex 5. Dorantes-Aranda JJ et al 2015. PLOS one DOI:10.1371/journal.pone.0133549 dominant microalgal polyunsaturated fatty acids EPA and periments can only serve as- DHA are indicated 6. Shimada M et al 1983. Acta Histochem proxies for real-life fish farm- Cytochem 16: 232-244 situations. HAB research 7. Marshall JA et al 2003. Harmful Algae 2: - ers need to forge better col 273-281 8. Munday B & GM Hallegraeff 1998. Fish laborative links with thein fishsitu airlift upwelling to dilute harmful al farm industry to conduct research on Pathology 33: 343-350 - 9. Deeds JR et al 2006. J Aquat Animal gal concentrations, or clay flocculation fish kill events that involve the Health 1: 136-148 to reduce numbers of harmful algal measurement of ichthyotoxins in sea 10. Arzul G et al 1998. In: Baudimant et al cells [22]. Mass fish mortalities lead to water using new analytical methods. Marine Lipids. Proc Symp Brest, Plou- tonnes of dead fish being dumped in Progress in understanding fish killing zané, France, (IFREMER), pp 53-62 - 11. Mardones JI et al 2015. Harmful Algae land fills (Port Lincoln, Australia, tuna mechanisms by HABs will pave the way - 49: 40–49 mortality 1996) or dumped offshore towards more effective mitigating strat 12. Sola P et al 1999. J Appl Toxicol 19: 279- (Chilean red tide 2016), but this materi egies which are much needed to feed 284 al is rarelyKarenia used for brevis fish meal. As argued an ever growing world population with 13. Yang CZ et al 1995. Dis Aquat Org 23: here, with perhaps a single exception high quality seafood. 101-10 - 14. Tang YZ & CG Gobler 2009. Harmful (Florida ), none of these Acknowledgements Algae 8:454-462 ”ichthyotoxins” are of human health sig 15. Oda T et al. 1997. Biosci Biotechnol Bio- nificance, meaning that recently killed- chem 61: 1658-1662 fish are still fit for human consumption. This overview is partially based on- 16. Twiner MJ et al 2001. Limnol Oceanogr Triggered by autonomous HAB moni a keynote lecture presented at the 46: 1400-1405 17. Place AR et al 2012. Harmful Algae 14: toring systems, once fish start to die a ICHA17 Brasil Conference, a more com 179-195 decision thus could be made to instigate plete write up of which can be found in 18. Igarashi T et al 1999. J Am Chem Soc 121: 8499-8511 19. Satake M et al 2002. Tetrah Letters 33: 5829-32 20. McNabb P et al 2006. Afr J Mar Sc 28: 375-377 21. Astuya A et al 2015. Harmful Algae 47: 1-8 22. Rensel J & I Whyte 2003. In: G.Hallegraeff et al (eds) Manual on Harmful Marine Microalgae (IOC Manual and Guides 33. UNESCO Publishing), pp 693-722 23. Seger A et al 2017. Harmful Algae 61: 46-55

Author Gustaaf Hallegraeff , Institute for Marine Fig. 3. Toxicity of the free fatty acid form of eicosapentaenoic acid (EPA) to damselfish: (top and Antarctic Studies, University of Tas- line) EPA on its own, requiring 3 mg L-1 to kill fish in 300 min; (bottom line) EPA in the pres- mania, Hobart, Tasmania, Australia ence of xanthine generated superoxide, where 0.2 mg L-1 EPA could kill fish in 100 min. Histo- pathology of damselfish gill tissues under selected treatment regimes is illustrated. Modified E-mail: [email protected] after [7]

HARMFUL ALGAE NEWS NO. 59 / 2018

11 with ALGAE-BASE by using the same names of the relevant algal species. The- names from ALGAE-Base are therefore 2000 automatically used in the List of Harm ful Microalgae. When name changes Harmful Algal Newsth - As part of the 25IOC-UNESCO anniversary Taxonomic issue of - become necessary, these are discussed Reference List of HarmfulI am providingAlgae www. an The contents of the list were discussed and agreed with ALGAE-BASE and in marinespecies.org/hab/overview of the - in detail at the HAB Conference in Ho- Whichserted in species both lists. should be ( bart in 2000, and it was decided to ­included in the list? ) and will high- aim at including additional informa - light some of the problems which have tion in the list such as the basionym of faced or Task are Team facing on the Taxonomy Intergovern each species (the first name applied to- The list was born as a result of the grow mental Panel on Harmful Algal Blooms the species), reference to the article in ing interest in harmful algae. The many (IPHAB) that which the species was described, refer name changes which regularly took is responsible for updating the list. ence to the article in which the species place for many harmful algae, following Historical timeline was given its current name, information publication of new information, was/ 1993 on the type locality, information on the is a source of confusion to ecologists,- main harmful effects of each species, toxicologists, people involved in routine - including up to three references with monitoring and others, thus construc A task team on the taxonomy of harmful information on toxicity, toxins or toxic tion of an agreed and updated list was algae was established at the Second Ses 2002effects. considered important. A decision had sion of the IOC-FAO Intergovernmental- to be made on which species to include Panel on Harmful Algal Blooms in Paris in in the list. EarlyAlexandrium on the distinguished minutum October 1993, in line with the HAB Pro The list was completed in 2002 and- member of the Task Team, Prof Y. Halim gramme Plan. The aim of the task team made available online on the homepage from Egypt, of was to make taxonomic recommendations of the Department of Biology, Univer fame, suggested that all harmful species- of harmful algal species, and membership Postsity of 2002 Copenhagen. should be included in the list, in other comprised Y. Fukuyo, Japan, M. Elbrächter, words the list should include also spe Germany, and Ø. Moestrup, Denmark, the- - cies which at some stage had formed latter serving as chairman. During the During the following years the list was blooms, causing low oxygen levels in the 1993 session it was “encouraged” to cre regularly updated, based on informa water at night, and leading to mortality ate a computerized taxonomic dataExpert base tion from committee members and of fish and/or or bottom invertebrates Centreof harmful on Taxonomic species, an Identification activity which at others. The list was also discussed at- such as certain species of diatoms and the time was planned betweenBotanical the Institute the meetings of the Intergovernmental silicoflagellates. While these were and (ETI) Panel on Harmful Algal Blooms in Par still are very real issues, to include in Amsterdam and the , is and suggestions for improvements all such species in the list would have 1997University of Copenhagen. 2008were made. made the list too long and difficult to- handle. It would have entailed including - numerous non-toxic and usually harm A major step forward was taken at the A major change took place in 2008 when less species. The Task Team eventually Fourth Session of the IOC Intergovern the list became part of WoRMS, the agreed on including only species known mental Panel on Harmful Algal Blooms, World Register of Marine Species. This to form a toxin or suspected to do so. which was held in Vigo, Spain 1997. change was decided following a visit by The lack of knowledge about toxins was- There it was decided to establish a Gert Hansen, Henrik Enevoldsen and me- particularly relevant for algae causing broader Task Team on Algal Taxonomy, to the headquarters of WoRMS at VLIZ fish kills, as many of the compounds re with the following terms of reference: in Oostende, Belgium. The merits of be sponsible for the fish kills were/are still I. to provide an agreed reference list coming part of WoRMS were discussed- not well known. The next problem was of harmful algal species, including and the technicalities of how to include Artemiato decide on the definition of the term correct author citation, date of valid and update species in WoRMS were ex toxic, did it mean toxic to humans, fish, publication of the species, and a list- plained and demonstrated. WoRMS is ? It was agreed to concentrate- of synonyms for each species - a very large database which includes on species toxic to humans or to fish. II. to provide guidelines to reduce no almost half a million species of marine When in doubt, a comment was includ menclature instability of harmful al organisms. It was decided that the HAB Nomenclatureed under the relevant problems species. gal species list should both be included in WoRMS III. to organize round table discussions and be accessible as a separate entity of on nomenclature and taxonomy of- WoRMS. The WoRMS list is harvesting One reason for the development of the harmful algal species - algal taxonomic data from ALGAE-BASE- list was that incorrect author citations- IV. to organize inter-calibration exer in Ireland. It was subsequently decided were recorded in the literature. In fact, cises to standardize the identifica that the IPHAB Task Team should coor the poor general knowledge of and in tion of harmful algal species dinate the List of Harmful Microalgae terest in nomenclatureHARMFUL is ALGAE still NEWS a problem. NO. 59 / 2018 12 Dinophyceae the authors intended the and in fact all taxonomic levels. Such new genus to be described according to- decisions have always been a problem, the botanical nomenclature. The lack of but molecular techniques have enabled a Latin diagnosis therefore made it inva discrimination of very small parts of the lid. The confusion has actually still not genome, inducing some researchers to- been resolved, and the problem needs- define only the genetically most similar to be discussed and decided upon by strains as belonging to the same spe- the International Nomenclature Com cies. Categories, subspecies or varieties Namemittee, beforechanges it can be fully resolved. are presently rarely used by phycolo- Fig. 1 Karenia mikimotoi (strain K-0260), a gists. If the genes examined are found historic strain isolated in 1977, as Gyrod- to be slightly more different, the mate inium aureolum, from Oslofjord, Norway, One of the problems facing the Task rial is considered to represent different by Karl Tangen (micrographs from Gert Team on Algal Taxonomy that maintains species, and if even more different, to Hansen) Karenia mikimotoi the List of Harmful Microalgae, is name- belong to different genera, families, and - changes (e.g. ) (Fig1). so on. This phenomenon shows no sign of dis- Is splitting or merging more correct? Thus mix-up of zoological and botani appearing, and it has recently become- There are no rules for this problem in cal nomenclature sometimes appears, an even more serious problem, follow the Code of Botanical Nomenclature; it- even in major phycological journals. ing the increased use of molecular se is up to the researcher her/himself to It may result in the creation of invalid quencing. - decide on how to define what consti- species names, or it causes uncertaintyStoeck- Invention and application of new tutes a species, a genus, a family, etc. - eriaabout the validity of names. An example techniques that allow species to be ex How to define a species has been dis is the description of the genus amined in more detail have commonly cussed for longer that I care to remem , which first appeared in 2005 in resulted in old species being split into- ber, but there is still little agreement on Journal of Eukaryotic Microbiology. The- two or more species. This happened this question. The same applies to the- description lacked the Latin diagnosis- when transmission electron microsco higher taxonomic levels. It is amazing required in botany at the time. The au py (TEM) became a routine tool in the that there is so much confusion, fol thors apparently intended this hetero 1950s, used in detailed studies of small lowing so many years of careful study.- trophic dinoflagellate to be described flagellates, morphological details of- Boenigk et al [1] in their discussion at using zoological nomenclature. That is- which are difficult or impossible to see theScientists VI European have been Congress trying of in Protistol vain for acceptable, a species which is validly using light microscopy (LM) but read hundredsogy in Berlin of years2011, toexpressed find the it ‘correct’clearly: described according to one Nomencla ily visible in the TEM. It resulted in the definition“ of species, but the simplest con- ture, automaticallyStoeckeria becomes valid in the description of numerous new flagellate clusion is that one does not exist other. However, in the title of the article- species, some harmful, others not. This in which was first described,- became a problem for ecologists and- ” (p 99 in the authors used the class name Dino others who did not use TEM in their- [1]). The species concept is a practical phyceae, which is botanical, not zoo- investigations and identification to spe- means that allows us to communicate- logical. Since the authors did not state- cies level was no longer possible. A sim about our organisms. However, thereThe specifically in the article which nomen ilar situation took place some time lat adventare currently of molecular more than data twenty … compels defini us clature they used, it caused some read er, notably for diatoms, when scanning- totions reconsider of what defineshow species a species, may andbe most“ ers to conclude that because of the word electron microscopy (SEM) became a- effectively and informatively delineated general tool. In recent years, the prob- lem has reached major proportions, fol ” lowing introduction of molecular meth (p 99 in [1]). The so-called biological ods. At the moment there is a tendency species concept is often mentioned, also to split species into ever smaller units, in phycology: if two populations can- discernible only by molecular methods. cross and produce fertile offspring, they In other cases, some species have been belong to the same species. This con merged, commonly based on studies of cept has strong limitations, however, as- one or a fewDinophysis genes, or on observations shown in both mammals and vascular of polymorphic life cycles (e.g. small plants. The two known species of buf cells of species) (Fig. 2). falo, the American and the European, Controversy arises when certain genes readily form fertile offspring. If they of morphologically different species were to be placed together, they would (morphospecies) are found to be very- most likely soon merge into a single similar. Do such species in fact belong to- species. Yet, we prefer to consider them- Fig. 2. Dinophysis acuminata from Mariager a single species? This has become a ma two different species. The same applies Fjord, Danish coastal waters (micrographs from Øjvind Moestrup ) jor problem for the Task Team on Tax to many other geographically separat onomy, as it results in discussions on ed organisms. The situation is further HARMFUL ALGAE NEWS NO. 59 / 2018 how to define species, genera, families, complicated by some populations of 13 - - geographically widespread species be will help taxonomy to retain its useful four species are very similar [3], in fact ing able to form fertile offspring, while ness in the general public. Logares etP. aciculiferum al. [4] found no P differencemalmo- other populations of the same species- A pragmatic Cochlodinium approach is currentlyCochlod- giensein ITS1, ITS2, 5.8S, SSU and partial LSU- are not. iniumnot heldpolykrikoides in high esteem, as exemplified- between and . Molecular techniques have contrib by the case of . . Yet these species are morpho uted very significantly to solving many C. polykrikoides (Fig. 3) is a very se logically and ecologically different and taxonomic problems, but they have not riousCochlodinium fish killer in South-East Asia, but- it would be meaningless to merge them resulted in the emergence of a finite is not the type species- into a single species. species concept. We have to accept that of , and evidence has ex Studies on the ribosomal genes all levels of classification are subjective isted for some years that the type spe- therefore do not always provide the ratherI mean than by objective,species, those and collectionsit has always of cies, whose morphology had not been final answer. Will these taxonomic- individuals,been that way. which To havecite Charles commonly Darwin: been studied in detail, is so different geneti problems be easier to solve when more so“ designated by naturalists cally that the two species should not genes or whole genomes become avail- Cochlodiniumbe included instrangulatum the same genus. If the able for use by the taxonomist? Some ” (cited by genus were to be split,Cochlodinium the type species, changes are bound to occur, and an ex M. Ereshefsky in [1]). - , would then ample of such a change is the case of At the generic level, splitting into- Cochlodinium,retain the name C. polykrikoides,, while the protist group Heliozoa, suggested many smaller genera is occurring rap unrelated species presently included in by Ernst Haeckel in the late 1800s. The idly at the moment. The genera con including group was found to be polyphyletic taining a large number of species are would need to be given one or more based on sequencing of the SSU gene- presently being split into many smaller new generic names. Such a change is [5], and Haeckel’s old name Radiolaria genera, and if this trend continues, boundCochlodinium to cause considerablepolykrikoides confusion was therefore in for modification. How- these diverse genera will become a- in countries like South Korea, where the ever, subsequent studies of transcribed thing of the past. In some cases, some of- name is well genes (single cell transcriptomics) re the new genera are monophyletic rath known by the general public. There are, sulted in the opposite result, Haeckel er than polyphyletic, and splitting be however, ways to solve such problems; was now right once more; the group- comes a matter of personal preference.- it is possible to change type species of was again monophyletic [6]. In other cases separation into smaller a genus if this is deemed to be critically- Taxonomy is going through a dif genera can result into a better under important.Nomenclature It is a solutionCommittee which has to ficult time at the moment. The DanishTyge standing of relationships of the species be discussed and accepted by the inter Christensenphycologist TygePrize Christensen, whose in this particularTask Team part ofon theTaxonomy old genus.- nalAcacia Cochlodinium, but it has name isInternational associated Phycological with the Soci- All these problems obviously leave the polykrikoideshappened before (e.g. the type species- ety awarded annually work of the dif of ). In the case ofC. polykrikoides by the ficult. One problem facing taxonomists it would have been prag , wanted genera to be identifiable is that results of taxonomic studies are maticCochlodinium to suggest that with the use of a hand lens or at least- used not only by academics themselves should become the new type species in a light microscope. If the present but also by non university people. With- for , as this would reduce- trend continues, most unicellular spe- this in mind, the taxonomic systems confusion. ThereNomenclature is no guarantee Committee that cies and genera will eventually become- suggested should not be overly academ the suggestion would have been ac- identifiable only by using molecular se ic; a pragmatic approach to taxonomy cepted by the , quencing. It will be a challenge to han Margalefidiniumhowever, but in the mean Referencesdle this situation in a pragmatic way. time a new genericCochlodinium name, 1. Boenigk J et al 2012. Eur J Protistol 48: polykrikoides , has been 96­102 2. Gómez F et al 2017. Harmful Algae 63: suggested for - [2]. 32-44 3. Annenkova NV et al 2015. ISME J A common problem fac- 9(8):1821-34 ing the Task Team has been- 4. Logares R et al 2007. Microb Ecol 53: that in some cases sequenc 549-561 ing of single or a few ribo 5. Polet S et al 2004. Protist 155: 53-63 somal genes have proved 6. Krabberød AK et al 2017. Mol Biol Evol 34: 1557-1573 insufficient to decide on the species level to be used.- Author This Peridinium was particularly aciculiferum clear Øjvind Moestrup, Chair of IPHAB Task Team on Taxonomy, Department of Biology, Marine in the case of the dinoflagelP. late Biological Section, University of Copenhagen, aciculiferum P. malmogiense Denmark complex,Scrippsiella which compriseshangoei P. Fig. 3. Margalefidinium polykrikoides (strain K-1292) , from Tsushima Island, ­Nagasaki, isolated by Mitsunori baicalense P. euryceps Email: [email protected] Iwataki in 2006 ­(micrographs from Gert Hansen) (as ), , and . The ribosomal genes of the HARMFUL ALGAE NEWS NO. 59 / 2018 14 Algal toxin discovery, management and regulation over the last 25 years afterwards the discovery of further compound groups that are produced by- Algal toxins in the dark ages - dinoflagellates and their metabolites in (pre-1992) th - shellfish was facilitated by the introduc- - mid-20 century the links between al tion of the MBA for lipophilic toxins in gae and toxins, or at least the toxic ef routine shellfish safety testing in Euro From a historic perspective, knowl fects of algae, are being made [11-15]. - pean legislation [52-53], as well as by- edge about algal toxins can be divided By the beginning of the 1990s, many several technological advances. One of into truly prehistoric occurrences such major algal toxins that cause acute poi- the main technological drivers in dis as known from paleontological studies- soning had already been discovered, covery was certainly the onset of the [1-2] and more recent historic records. including brevetoxins [16-19], cigua use of liquid chromatography coupled In these historic records, there are de toxins [20-27], domoic acid [28-36], to mass spectrometry (LC-MS) [54-56], scriptions of poisoning incidents that okadaic acid and analogues [37-40], which became quite widespread by the clearly point towards the occurrence prorocentrolide [41], and saxitoxins beginning of the 2000s [57-61]. of the algal toxins centuries ago, such (STXs) [42-46]. Quite a few analogues The number of toxin groups that as the description of Captain George of the main toxins had already been- were discovered over the period from Vancouver, whose crew suffered from discovered [47-50], as well as several 1966 to 1990, (fifteen), was not much paralytic shellfish poisoning during the- groups of compounds produced by di less than those discovered over the last exploration of the Pacific Northwest in noflagellates that provoke death in mice 25 years, (nineteen) (Fig. 1). However, 1793 [3]. Other examples include cigu- used for the mouse bioassay (MBA) but the number of analogues in each group atera [4] and paralyticth poisoning [5-6].- are not necessarily related to human has rapidly increased. A good example During the 19 century, modern taxon- poisoning events, such as pectenotoxins of this is the STX-group where a review- omy emerged as a science with devel Discovery[50] and yessotoxins of toxins [51]. over the last in 1990 counted nineteen observed opments in microscopy; by 1900, rath 25 years analogues with a further five predict er systematic studies of phytoplankton ed from plausible metabolization or communities are common [7-10] and chemical transformation pathways- provide the basis for the biogeography While discovery was mostly driven by [62]. In 2010 a review reported over of many toxic genera. In the early to human poisoning prior to the 1990s, 50 observed analogues [63]. A simi

Fig. 1. Discovery or description of the structure of the first analogue of 34 toxin groups (1966 – 2017). Colour code: blue: toxins involved in fish kills; red: toxins involved in human poisoning, violet: toxins causing skin irritation or respiratory problems (BTXs should be red, blue and violet), green: toxins known for 20-30 years and not proven to have nega- tive effects on humans or aquatic organisms, black: toxic compounds yet to be related to effects in humans or aquatic compounds. Nota bene: not many toxin groups relevant to human poisoning are being discovered while more and more toxins related to fish kills are (toxins of Karenia brevisulcata may be related to the Wellington Harbour syndrome)79-108

HARMFUL ALGAE NEWS NO. 59 / 2018

15 lar rapid increase in known analogues the agents involved in fish kills has also New Zealand, then Europe [126] and has been observed for the azaspiracid been highlighted by a recent systematic Outlookmost recently Japan. (AZA) group, with the first analogue review of toxic and harmful algae [109],- described in 1998 [64] and a review in as well as by the Intergovernmental - 2014 reporting 30 analogues [65]. Only Oceanographic Commission of UN Several points can be raised looking three years later, over 50 analogues are- ESCO (IOC) Intergovernmental Panel foreward from the historic perspec known for this group, including novel on Harmful Algal Blooms (IOC-IPHAB), tive. Climate change is one of the most phosphate derivatives [66-71]. This dis that included the topic in its list of Task striking challenges that has been raised covery rate could have been even more Teams. with regards to prediction of harmful rapid if the causative organisms of AZAs- The systematic inventory of toxins algal blooms (HABs), and while certain had been elucidated earlier. However, has also been updated [78-108] by the trends appear to manifest themselves- the delay from the first poisoning re IOC-IPHAB Task Team on Biotoxins,- [127], much more research is needed port in 1995 [72] to the discovery of the- Management and Regulation over the- to fully anticipate the impacts of cli culprit organism [73] in this case was past few years and international data mate change on our ecosystems, HABs Protoperidiniumlikely due to: i) the crassipes initial misidentifica bases, e.g. the Harmful Algal Event Da and their impacts on society [128]. As- tion of the heterotrophic Azadiniumdinoflagellate tabase (HAEDAT) updated accordingly.- mentioned above, the need to improve (a vector of- This same panel also contributes to our understanding of the impacts of mi AZAs upon its feeding on ) as- other IPHAB activities whenever chem cro-algae on other aquatic organisms, the causative µm)agent, by andoptical ii) themicroscopy difficul ical expertise is required (e.g. fish kills, in particular those that serve as major ties in identification of such a small or- DriversHABs and of desalination change in etc.). manage- food resources, i.e. fish and shellfish, ganism (<15 ment and regulation has been recognised and requires major in water samples fixed with acidic Lu international efforts. The multiplicity of gol’s solution, the most common way to compounds in the marine environment preserve samples in routine plankton- There have been many drivers of change only emerges with the recent onset of monitoring. in management practises (e.g. detection- the omics and while recent studies have- Another phenomenon that has ap- methodology) and regulation. These shown the feasibility to explore this- peared repeatedly over the past 25 include (i) increased awareness by gov chemical diversity in the marine envi years is the discovery of slightly modi ernments of poisoning events and fish ronment with techniques such as me fied base skeletons for toxin groups. and shellfish mortalities through IPHAB- tabolomics [129-131], more systematic The ciguatoxin (CTX) or CTX1B (= P- communication with member states (ii) studies will be required to effectively CTX-1B) had been reported relatively- increased pressure from shellfish indus monitor our coastal waters to protect early on and had been isolated from try against the MBA for lipophilic toxins our resources and consumers. Finally, it- the moray eel [74]. The algal precur- due to its qualitative character, false- should be noted that one of the longest- sor CTX4A was only described in 1997 positive results and delays in reporting known groups of toxin, i.e. the ciguatox [75], yet a slightlyGambierdiscus modified base skel (iii) technological advances. The con- ins, still continue to cause the highest eton had been reported a few years flicts caused by the disadvantages of the number of seafood poisoning globally earlier from , i.e. CTX3C- animal assays (mouse and rat) for lipo [132] and thus deserves the attention of [76]. Prymnesins are another example philic toxins has been subject to much the scientific community over the next of such skeleton variation which is in debate [110] and decade-long efforts to References:few decades. deed very labour-intensive in natural produce the necessary standards and 1. http://hab.ioc-unesco.org/index. product discovery as basically the full reference materials for the validation of php?option=com_content&view=article& id=42&Itemid=0 discovery pipeline has to be completed:- alternative methods, which have been bioguided fractionation and isolation of aided by researchers in Canada (e.g. 2. http://www.phycotox.fr/decouvrir/ chimie/toxines-d-algues-et-classification. the compound, purification and struc- Michael Quilliam), Ireland, Japan (e.g. html?lng=en tural characterisation including mass- Takeshi Yasumoto), New Zealand and spectrometry, nuclear magnetic reso many other countries [111-122]. Again- nance (NMR), UV, infrared and poten the IPHAB panel played a pivotal role in Author Philipp Hess, Chair of IPHAB Task Team on tially many other studies [77-78]. pushing this issue at European and in Biotoxins, Management and Regulation, Finally, it should be noted that only a- ternational levels for several years with IFREMER, Phycotoxins Laboratory, Unit few compound groups discovered since the help of Phil Busby† (New Zealand DYNECO, Centre de Brest, BP 70, 29280 1992 have been clearly related to hu Food Safety Authority), a long battle for Plouzané, France man health issues. These include AZAs which the international community will Karenia brevisulcata Email: [email protected] (diarrhoea), ovatoxins and to a lesser remember him. extent the toxins of Monitoring systems, management (aerosol and direct contact exposure). practises and legislative changes have There is a significant increase of been recently reviewed for different compounds that appear related toK. trade blocks [123-125]. A major step brevisulcatafish kills, e.g. karlotoxins, karmitoxins, has been made with the switch from the prymnesins (A, B and C-Type) and mouse bioassay to chemical testing by toxins. The need to clarify LC-MS/MS for lipophilic toxins, first in HARMFUL ALGAE NEWS NO. 59 / 2018 16 Butterflies in Brazil

- - Abstracts are not always reliable guides which was the primary to authors’ intentions. They are not ex raison d’être for conven pected to reveal a great deal about the ing the conference. In evidence to be deployed in support of- subsequent discussions the science, evidence that may not even of this theme,frequency, we often in- exist before deadlines for writing ab- tensityfind some combinationspreading of stracts! Nevertheless, as examples of a the words minor art form, we are entitled to sub global epidemicand . Ted ject them to criticism. These comments Smayda used the phrase- were provoked by reading the abstracts [4]. - of the Florianópolis conference. Here In a widely cited state the focus is on two topics, i) climate ment of concern with cli Fig. 1. Trajectory in 3-dimensional phase space of solu- change, and ii) suggestions that we mate change and harmful tions of a simple model of atmospheric convection. Each public health point represents an instantaneous state of the system. The follow Ben Schneiderman [1] into the algae Gustaaf Hallegraeff - and economic impacts of region to which the points are confined is called a strange convoluted procedures of “Scienceclimate 2.0”. suchwrote events that appear ” to have attractor. This is Lorenz’s butterfly. A word count of the Florianópolis con increased in frequency, in- climateference change abstracts reveals that tensity and geographic distribution occurs 73 times, 33 times in the phrase - , 4 times in presentation ” [5]. each of these systems are determined titles [2]. Emerging1 evidence sug- Note here, it is not redappear tides to thathave have by many interactions within and be- gestsHere that are climate some change incantations is impacting from haveincreased, and the public health and- tween them, multitudes of signals with marinethe abstracts: and freshwater “ phytoplankton economic problems (not differing amplitudes and phase rela- communities”; “Climate change and an- ) increased. This distinction is of- tions, positive and negative feedback thropogenic activities in the coastal zone ten missed. Hallegraeff also discussed mechanisms, in short, complex dynam are increasing the risk of Harmful Algae the possibility that there is a real epi ics. Complexity really is complex! The Blooms”; “The worldwide distribution, demic (in Smayda’s sense). Although- mean climate of a region is the average frequency and duration of cyanobacte- he concluded that this too is apparent- of all the processes we call weather, rial blooms are driven by eutrophica- rather than real, he listed various puta conventionally calculated over a thirty tion and climate change”; “El Niño … is tive causes, amongst them eutrophica year time span (World Meteorological increasing in frequency and magnitude tion and climate variations. Organization) from surface records of due to climate change”; “As a result of Revisiting the theme nearly two temperature, humidity, precipitation,modes climate change and non-climatic factors, decades later, Hallegraeff [6] provided- winds, etc. But the mean deceives; it algal bloom frequency, composition and a succinct guide to what might happen- hides variations due to internal , spatio-temporal distributions are chang- under a conventional IPCC climate pro the best known of which are the El Niño- ing.” - jection; he discussed the potential im- Southern Oscillation (ENSO), and the pact of warming seas, changes in mixed North Atlantic and Pacific Decadal Os These quotations are mostly asser layer depth, wind regimes, runoff, up cillations (NAO, PDO); so on decadal tive; such formulae are not unique to welling rates and other variables – we- to secular time scales, regional climate the Florianópolis conference, but were can call them underwater weather – as fluctuations are largely due to modes. much more prominent there than in well as various feedbacks, and overfish- Model climateschanges are those we build in earlier conferences in the series. ing. These variables are standard fare- our imaginations and in our computers.- The idea that harmful blooms are a for marine ecology, appealed to rou Climate are persistent- growing problemthere was expressedis evidence in that the tinely to explain the changing popula trends identifiable in meteorologi paralyticfirst HA conference shellfish poisoning by Anand outbreaks Prakash tion dynamics of HABs as well as other cal records, usually considered as re arewho increasing wrote; “… in intensity and spreading marine species. sponses to changes in external forcing to new areas - The key ideas of climate change rather than modes; recent examples- intensity are not always examined critically, nor are the cooling from 1940 to 1970 or ” [3]. He invoked eutrophi with the natural suspicion that theircli- the warming from 1970 to 2000. “An- cation to account for increased frequentlymate change doctrinal character should thropogenic global warming” (AGW) of blooms, andspreading cited Tokyo Bay and the excite. So what do we mean bymean considers recent warming a result of in inner Oslofjord as examples. His only modal? On decadal to secularmodel dustrial growth, and many AGW models example of was that of PSP time scales, we can distinguish - focus on forcing by atmospheric carbon- 2 to Emerging the western Gulf of Maine in 1972, climates, climates, and dioxide concentrations [CO ]. As time - climates. Climates result from the in scales grow from secular to millen 1) is a fashionable prefix foremerging evidence, teractions of many different elements nial, we meet palaeoclimates with well evidenceas it is for markets, equities, roles, … A scien - cedirect.com search finds the phrase within the atmosphere, hydrosphere, known phenomena like Pleistocene Ice in nearly 30, 000 titles; time to give it cryosphere, and biosphere, including Ages and the Holocene climatic opti HARMFULa rest? ALGAE NEWS NO. 59 / 2018 phytoplankton. The future states of mum. But the perceived HA epidemic is 17 - on a decadal time scale, and identified- palaeoclimatic oscillations are not nec essarily a useful guide to its interpreta tion. An obvious obstacle to detecting climate signals in HAB data is posed by anthropogenic eutrophication. Another obstacle is the fact that phytoplankton respond directly to the increased [CO2] blamed for warming in many models.- Phytoplankton, macroalgae and sea grasses respond to raised [CO2] by in creased nitrate uptake and growth.- Thus higher [CO2] can lower the impact of eutrophication. Increased [CO2] con sumption by phytoplankton also raises pH, hence counters any tendency for acidification. - In the original examples of increased intensity of blooms (Tokyo Bay and in Fig. 2. PSP toxicity maxima identified by White [11] are centered on years 1945, ner Oslofjord), eutrophication seems a- 1961 and 1980; these lag minima of lunar declination in 1940, 1959, and 1978 by a highly likely cause since both are semi-- few years. Had the trend continued, further peaks might have been expected around enclosed water bodies with increasing 2000 and this year. The Lepreau Harbour toxicity time series shown here has the ly urbanized watersheds; but eutrophi three earlier peaks, and more recent ones in 1995 and 2009 which precede the lunar cation cannot have driven the sudden declination minima in 1997 and 2015. Clearly the nodal signal is not the whole story! spread of PSP into the western Gulf of (Figure courtesy of Jennifer Martin, St Andrews Biological Station, NB, Canada). Maine, open waters with low human population density along the coast. The- message from these examples is that we must analyse the frequency, inten then is the prototype of chaotic systems. much use of the subjunctive tense. But- sity, and spreading of HABs as separate We all know that algal growth and Kedong Yin & Jianzhang HE may have phenomena, on a species by species the likelihood of blooms are linked to distinguished eutrophication and cli basis, and not lump them all together wind regimes, water column stability, mate variation in a time series (1983 to as manifestations of a global epidemic. runoff, mixing rates, and so on, that is 2014) of Hong Kong red tides [10]. The same message emerges from more to the local weather, all modulated by In the case of toxins, PSP records recent examples, for example those regional climate variations. One might- from the Bay of Fundy (1944-1983)- summarized by Hallegraeff [6] Doesfrom thehis therefore anticipate that time series have yielded evidence of a multi-annual flapantipodean of a butterfly’s perspective. wings in Brazil set off of harmful algal events contain sig lunar signal, the nodal tide, with a pe a tornadoEdward in Lorenz Texas? Is[7] the asked, behavior “ of the nals provoked by multiannual trends riod of 18.6 years (Fig. 2) [11]. This atmosphere unstable with respect to per- in these oceanographic parameters, finding merits more attention than it turbations of small amplitude hence act as proxies for climate trends. has so far received given that the same- nonlinear But it seems to be very hard to extract signal has emerged from analyses of ?” There is Journalunambiguous of Sea signalsResearch from records of the PDO, the NAO, and many oceano- a relationship between the HABs and algal toxins. In an issue of the graphic and meteorological time series- two variables, butterflies and tornados,- [8], 14 papers- [12, 13]. Chaotic systems are concep meaning that an equation relatingdynamical them examine phytoplankton time series 1- tually remote from the clockwork uni is not of the first degree; nonlinear Sincerela to 4 decades inThe length, initial with hope special that these at verse of Isaac Newton and Pierre-Simon- wetionships do not are know the exactlyhallmark how of many but- datatention sets to do climate contain variability significant andsignals, eu Laplace. But the nodal tide and other terfliessystems there (Fig. are, 1). nor Lorenz where wrote: they are “ lo- andtrophication. that they “ can be identified and de- astronomical forcings are determinis cated, let alone which ones are flapping scribed, is not realized tic, they do run like clockwork, and can their wings at any instant, we cannot, if despite indisputable in principle provide forecasts of future the answer to our question is affirmative, and often major changes”, at inleast coastal on these wa- climate variation. …, accurately predict the occurrence of terstime caused scales; by and, human “ activities, no clear The first conference in this series tornados at a sufficiently distant future signals are detectable in phytoplank- took placeclimate in Boston in 1974. From then time - ton composition and dynamics, nor any until the Lund conference (1989), the- the which are distinguishable from changes word was not indexed in the butterfly.” A practical effect corollary of this sen in areas relatively unaffected by such im- proceedings. At the Newport confer sitivity to initial conditions, called pacts ence (1991), El Niño was mentioned , is that forecasts of the twice, and there were suggestions that future are nearly impossible. Climate .” [9]. Claims to the contrary make as a result of hypotheticalHARMFUL ALGAE NEWS climatic NO. 59 / 2018 18 Karenia brevis weather trends, Gymnodinium might catenatum appear From the abstracts alone, it is hard to- potentially fruitful. But if science is to more often in the South Atlantic Bight- know. A few clearly mean in be part of these activities, it must be of the US and the sense used above. Are others jump real science, not a debased imitation. bloom more often in northwestern Ibe- ingCrying on a wolf bandwagon? does not serve If so, our they discipline might References rian waters. There was also a warning andrecall we Gustaaf need to Hallegraeff’s refrain from admonition: making un- 1. Schneiderman B 2016. The new ABCs by Barrie Dale germane to such specu substantiated“ climate predictions of research: achieving breakthrough collaborations (Oxford University Press), lations, that large scale climate models - cannot predict local changes. !” [16]. 320 pp - 2. ICHA 2016.Book of Abstracts. http:// Little more was heard of supposed coupledScience human 2.0 was and represented natural systems at Flo icha2016.com/program/abstract-book. links between HABs and climate un rianópolis [17, 18] by Jianguoecological Liu’s pdf til two decades later in Hersonissosthis confer- public“ health ” or 3. Prakash A 1975. In: LoCicero VR(ed), Proc 1st Intern Conf on Toxic Dinoflagel- ence(2010). included The preface … a solid to body the Hersonissosof 50 papers CHANS [19] and Tim Lang’s “ proceedings claims that “… ”, EPH [20]. Both combine late Blooms (Mass Sci Technol Foundn, and posters on climate change Wakefield, Mass), pp 1-6 - ecosystem models with other complex- 4. Smayda TJ 1989. In: Cosper EM et al climate”; changeif true, systems. CHANS are built by linking- (eds), Novel Phytoplankton Blooms few of them appeared in the proceed models of economic, social, hydrologi (Springer-Verlag, Berlin, Heidelberg, New York), pp 159-187 Prymnesiumings, where theparvum phrase cal, atmospheric,more and than biological just evidence, sub appears in a single contribution about systems and interactions among them. 5. Hallegraeff GM 1993. Phycologia 32: 79- global warming and includes the open pursuit of social 99 in some Texan values,EPH combines highlighting “ the role of inter- 6. Hallegraeff GM 2010. J Phycol 46: 220- lakes, and the phrase Chattonella glo- est groups, and debate across society 235 bosaonly twice.Dinophysis There were tripos speculations - not just within restricted scientific cir- 7. Lorenz E 1964. Tellus 6: 1-11. J Sea Res that the appearance of 2009. Vol 61(1), 124 pp cles 8. J Sea Res 2009. 61(1): 1-124 and in Norwe Ostreopsis 9. Wyatt T 2010. In: Briand F (ed), Phyto- ovatagian coastal waters might be related to .” Climate change is central to both- plankton responses to Mediterranean en- global warming, similarly for schemes. The novelty then is to weld vironmental changes. CIESM Workshop in the upper Adriatic. ecosystem dynamics with climate dy Monographs N° 40 (CIESM Publisher, Changing patterns of abundance on namics, add some flavours from outside Monaco), 120 pp 10. Yin K 2016. Abstract OS1805, p 59 in [2] decadal time scales were reported at science, run some models, and see what Karenia Cochlodinium Chat- - 11. White A 1987. Rap Proces 187: 38-46 tonellathe ChangwonHeterosigma conference (2012) for might happen. 12. Yasuda I 2006. Geophys Res Lett 33 species of , Pyrodinium , Shneiderman’s Science 2.0 com L08606, 4 pp 13. Yndestad H et al 2008. Deep Sea Re- , and Gymnodinium in Korean and bines applied science, engineering, and- Japanese waters, in the design and attempts to tackle problems search I 55: 1201-1217 Prorocentrum Heterosigma - 14. Wyatt T 2013. HAN 47: 1-3 Philippines, in Portugal, like disaster response systems or envi 15. Dale B et al 2006. In: Granéli E & Turner and in Nar ronmental sustainability [1]. These are JT (eds), Ecology of Harmful Algae ragansett Bay. As pointed out at the time not purely scientific problems, and their (Springer-Verlag), pp 367-378 [14], some of these patterns may be complexity makes it necessary to break 16. Hallegraeff GM 2011. HAN 43: 11-12 linked to regional effects of phenomena them into smaller pieces for analysis. 17. Fleming L 2016. Abstract KN02, p 6 in [2] - 18. Lintott L 2016. Abstract POS0128, p 115 like the PDO or ENSO; and some almost One of Shneiderman’s five strategies for in [2] certainly, as Patricia Glibert argues, to doing this is classical reductionist sci 19. Liu J et al 2007. Ambio 36: 639–649 eutrophication. As already mentioned, ence [21]. It is therefore impossible to- 20. Lang T & G Rayner 2012. BMJ 2012 Aug disentangling the separate impacts of accept the viewpoint of one participant 21;345:e5466. doi: 10.1136/bmj.e5466 climate and nutrient loading is a major in the conference, that taxonomy, physi 21. Wyatt T 2017. Ethics Sci Envir Polit 17: 51-62, DOI: https://doi.org/10.3354/ problem. AGW was not invoked, indeed ology, biogeography, and population- esep00177 it is clear that we do not yet have an dynamics are no longer useful research 22. Wells M 2016. Abstract PS06, p 9 in [2] adequate data base to do so [15]. Also topics, that “they will not adequately in 23. Marshall A 1998. Stud Hist Philos Sci Pt C. Stud Hist Phil Biol & Biomed Sci 29: at Changwon, Barrie Dale warned that- form on purported climate change: HAB “available time series plankton data are linkages” [22]. 137-164 inadequate for establishing species’ re Even though complex models of the sponses to natural climate variations- climate or an ecosystem can exhibit Author and therefore offer no sound basis for chaotic dynamics, with all the technical Tim Wyatt, Borreiros Gondomar, Pontevedra, Spain predicting effectsclimate of climate”. The Wel challenges entailed, Science 2.0, EPH, lington conference (2014) abstracts and CHANS propose to combine several - Email: [email protected] contain the word only six times; such systems in search of solutions to- in the two contributions in which cli- economic and political problems. These mate change was addressed. directly aspirations read like recipes for “cyber there, its impact on HABs was again en- netic gigantism” [23]. We can agree that meshed with eutrophication - solutions to complex problems demand So what did participants at Flori multidisciplinary approaches, and that anópolis mean when they wrote of cli collaborative projects which combine mateHARMFUL ALGAEchange? NEWS NO. Was 59 / 2018 warming intended? expertise from diverse disciplines are 19 Red tides in Kamchatka coastal waters -

(Bering Sea, Russia) are a barrier for harm AG, Germany. The-1 test-system de tection limit is 5 µg kg with specificity the salmon fishery and Pacific salmon to different PSP toxins of: PSP, 100%; decarbomoylAlexandrium PSP, tamarense20%; gonyautoxin II and III, 70%; neosaxitoxin, 12%. (Lebour) Balech was the dominant species (up 5 -1 Tripos fusus to 1.32 x 10 cells Protoperidinium L ) (Fig. 3). Other dinoflagellates,Alexandrium such leei, as (Ehrenb.) Dujard.,A. tamarense sp.- and сf. co-occurred. The diameterA. . leei of vegetaA. tamarensetive cells ranged from 31 to 40 µm, and that of сf was around 36 µm. was found both as vegetative cells (Fig. 3A-C) and in different stages- of cyst formation (Fig. 3F, G).

Saxitoxin concentration of the-1 sea water sampled was 0.33 µg L . The neurotoxic impact of the bloom on fish in Olyutorskiy Bay was observed by the- “slack” behavior of pink salmon passing through the bloom patches, and mortal Fig. 1. Map of Olyutorskiy Bay (Kamchatka, Bering Sea) where a red tide, reported by ity of some individuals. There are no fishermen, occurred in July 2017. The stars denote fishery sites: red, affected by the standards for saxitoxin concentration bloom; green, not affected; the arrow indicates the sampling area in sea and fresh water in Russia, but the

established-1 regulatory level is 800 µg kg for the edible tissues of mollusks - and crabs [2, 3]. In mid-July 2017 a red tideDelfin occurred- (IEA) with the help of the test-system Visual observation from the air of in Olyutorskiy Bay, Bering Sea (Fig. 1). RIDASCREEN® FAST PSP SC, R-Biop the Pacific salmon in the spawning Fishermen of the company wit nessed this phenomenon at their fishery sites (Fig. 2). The first signth of the bloom was observed on Julyth 11th , following a flood during July 8 -9 causedth by a strong cyclone. By July 15 the bloom extended along the central coastal zone of the bay, in a layer approximately 5-6 m in depth. The fishermen’sOncorhynchus attention gorbu- shawas attracted by the unusual behavior of pink salmon ( Fish) thatwere were slack, approaching looked tired the and rivers died soonto spawn in the and trap entered net the bloom area:- “ - ”. Additionally, na tives of the Pakhachi settlement report ed that nearshore waters were reddish- brown, had an unpleasant smell, and dead salmon were washed ashore. To investigate this phenomenon, a surfaceth water sample was taken on 15 July 2017 from the reddish-brown discoloured plume one mile from the- mouth of the Impuka River (Fig. 1). Fig. 3. Alexandrium tamarense in a water sample from Olyutoskiy Bay during the bloom event: Bright-field and epifluorescence mi- croscopy [1] were used to identify the A. General view of cells in the sample; B. Cells with content and empty thecae; С: Ventral view - of the theca; D-E. Calcofluor-stained epithecal plates; F. Shedding of the theca (cyst forma- phytoplankton present. Saxitoxin con tion); G. Cyst. Scale bar = 100 µm (A), 20 µm (B-G). tent in seawater samples was deter mined by an immune-enzyme assay HARMFUL ALGAE NEWS NO. 59 / 2018 20 Fig. 2. Red tide in Olyutorskiy Bay on 15th July 2017

References 1. Hallegraeff GM et al (eds) grounds of the Olyutorskiy Bay river- 1995. Manual on harmful basin performed during the second half marine microalgae. IOC of August showed an atypical distribu Manual and Guides 33 (UN- ESCO, Paris), 551 pp tion in the rivers. Maximal escapes were- 2. Sanitary-epidemiological recorded in river basins located in the Rules and Regulations (San- western and eastern parts of the Olyu- PiN 2.3.2.2401-08). Hygienic requirements for safety and torskiyOncorhynchus Bay area, whereas gorbusha in its O.central keta part, the abundance of the target spe food value of provision. 2008 O. nerka (in Russian) cies ( , 3. Technical Regulations of and (all called Pacific salmon) the Customs Union (TR CU was extremely low (Fig. 4). 021/2011). On Food Safety. A map of salmon distribution over 2011 the spawning a grounds (Fig. 4А) was- compared with that of satellite images Authors of chlorophyll- (Fig. 4B, C) which sug Ekaterina V Lepskaya, Kam- gested that the toxic bloom caused a- chatka Institute for Fisheries re-distribution of the migratory paths Research and Oceanography, for salmon, with most of them mov Laboratory of Hydrobiology, Petropavlovsk-Kamchatsky, ing to the rivers adjacent to Olutorsky Russia Bay. Changes in anadromous migratory paths of Pacific salmon caused some- Tatyana A Mogilnikova, Sakhalin problems for the fishery in the bloom Research Institute of Fisheries zone and additional unscheduled ex and Oceanography, Laboratory of Hydrobiology, Yuzhno-Sakhal- penses. insk, Russia This is the first documented event for Kamchatka, where a coastal bloom Sergey V Shubkin, Kamchatka of the toxic microalgae appeared to be Institute for Fisheries Research - and Oceanography, Laboratory a barrier both for fisheries and Pacific of Abundance Dynamics and salmon spawning migration into fresh Forecast Perfection for Salmo- Acknowledgementswater. nids, Petropavlovsk-Kamchatsky, - Russia - Oleg B Tepnin, Kamchatka We thank Yuri B. Okolodkov, Universi St- Institute for Fisheries Research ylabdad Veracruzana, Boca del Río, Verac and Oceanography, Laboratory ruz, Mexico, and Alexander V. Galkin, of Oceanography and Freshwater , Moscow, Russia, for their valuable Hydrology, Petropavlovsk-Kam- comments. Marcia M. Gowing, Seattle, chatsky, Russia WA, USA, kindly corrected the English Email: [email protected] style.

Fig. 4. Distribution of Pacific salmon in river basins (А) and bloom patches as detected from the chlorophyll-a concentration (B:15th July 2017; C: 21th July 2017; http:// marine.copernicus.eu).

HARMFUL ALGAE NEWS NO. 59 / 2018

21 First report of Gambierdiscus in the

Western Mediterranean Sea (Balearic ranged from 64.1 to 90.8 μm (meanµm of Islands) 78.6 μm). The original description [9] described a length range of 76-93 - and a cell width of 65-84 µm. Further Gambierdiscus morphological analysis will be per- - G. australes formed using electron microscopy. (Dinophyceae) species- Mediterranean Sea. The present study To facilitate molecular identifica are benthic dinoflagellates living in ma confirms the presence of in tion to species level, DNA was extracted rine littoral zones of circumtropical ar the two Balearic Islands of Majorca and from individual or a few clonal cells Gambierdiscus TM TM eas and have recently been described in- Minorca, and this constitutes the first using the Arcturus PicoPure DNA temperate waters [1]. Some species are report of genus in the Extraction Kit (Applied Biosystems, CA, producers of potent neurotoxins: cigua western Mediterranean Sea. USA). Afterwards, the domain D8-D10 toxins (CTXs) and maitotoxins (MTXs). In this study, microalgal samples- of the LSU rRNA gene was amplified by a Ciguatoxins are linked to Ciguatera Fish were collected from macroalgae and Polymerase Chain Reaction (PCR) using Poisoning (CFP). Ciguatera used to be- rocky substrates in 19 stations in Ma the pair of primers FD8 and RB [9], and restricted to tropical and subtropical- jorca and Minorca in September 2017. products were sequenced. The D8-D10 areas, but since the last decade, it ap Water temperatures ranged from 24 to sequences obtained in this study were pears to be expanding to more temper 27 °C and salinity from 36.2 to 38.0. In deposited in GenBank under accession ate latitudes. For example, outbreaks the laboratory, samples were observed numbers: MG708117- MG708130.Gambier DNA- of ciguatera have been reported in the under the microscope and individual- discussequence analysis of amplifiedG. australes rDNA Canary IslandsGambierdiscus and Madeira (eastern cells were isolated with micropipettes fragments confirmed that all - Atlantic Ocean), where several species to establish cultures for morphologi spp. corresponded toG. australes, of the genus have been- cal and molecular analysis. Calcofluor which was in accordance with the mor- identified [2]. - white stain was used for morphological phological identification. In the Mediterranean Sea, no thor identification. Cells were observed with was present in 10 out of the 19 sam ough evidence of cases of ciguatera ex a compoundGambierdiscus microscope equipped with pling stations in Majorca and Minorca ist. The only reports of CTX-like toxins epifluorescence at 630X (Leica DMLB).- (Fig. 2), indicating that this species is in fish, which are not confirmed, were The cells observed were- well established at different locations based on the Cigua-Check Fish Poison anterior-posteriorly compressed. Mor around the coasts of both islands. It will Test kit (Oceanit,Gambierdiscus Hawaii), a method phology of the epitheca and the hypoth be important to evaluateG. australes the temporal that has proved to be unreliable [3]. eca is shown in Fig. 1 in which the plate distribution of this species. - Nonetheless, species- terminology employed follows Fraga The first report of was in- have been reported in the last decade Fuku- and collaborators [8]. The epitheca has the Australes archipelago (French Poly yoain Crete (F. paulensis) and Cyprus (eastern Mediter a rectangular-shaped 2’ apical plate nesia). This species is also widely dis 0 ranen Sea) [4-6]. One species of - and the P plate is ventrally oriented; tributed in areas such as New Zealand Gambierdiscus, a genus that includes the hypotheca has a narrow 2’’’’ plate and the Canary Islands, but it had not species previously included in the ge equivalent to 1p plate in Chinain [9]. been reported yet in Gambierdiscusthe Mediterranean nus , was reported in The cell surface is smooth. The cell Sea. Some studies mentioned that the the Balearic IslandsGambierdiscus in 2015 [7]. Little length and width of 62 individuals were spatial expansion of and is known about diversity, distribution measured. Length ranged from 60.9 to CFP may be related to the increase of and toxicity of spp. in the 92.3 μm (mean of 75.6 μm) and width temperatures caused by climate change

Fig. 1. Epitheca (a) and hypotheca (b) of Gambierdiscus australes cells stained with Calcofluor White.

HARMFUL ALGAE NEWS NO. 59 / 2018

22 - the SEASENSING (BIO2014-56024- C2-2-R) project and the CERCA Pro gramme/Generalitat de Catalunya. A.- Tudó and A. Toldrà acknowledge IRTA- URV-Banco Santander for their respec tive PhD grants (2016 PMF-PIPF-74 and 2015PMF-PIPF-67). The authors are grateful to Vanessa Castan and José Luis ReferencesCosta for sampling assistance. 1. Litaker R et al 2010. Toxicon 56: 711-730 2. Rodríguez F et al 2017. Harmful Algae 67: 131-143 3. Bentur Y et al 2007. Clin Toxicol 45: 695- 700 4. Aligizaki K et al 2008. J Biol Res 9: 75–82 5. Aligizaki K et al 2009. In: Lassus P (ed) 7th International Conference on Mol- luscan Shellfish Safety, Nantes, France, Fig. 2. Locations where Gambierdiscus australes was recorded in the Balearic Islands 14-19 June (IFREMER 2009), pp 1-6 (39° 30’N, 3° 00’ E), Spain. 6. Holland W et al 2013. Toxicon 65: 15–33 7. Laza-Martínez A et al 2016. J Eukaryot - Microbiol 63(4): 481-97 8. Fraga S et al 2011. Harmful Algae 11: [10]. The Mediterranean Sea, which is sary to study whether the Balearic Is 10-22 a semi-enclosed sea, seems to be one lands could be a new spot of ciguatera. 9. Chinain M et al 1999. J Phycol 35: 1282–96 of the regions strongly affected by the Improving our knowledge about 10. Friedman M et al 2017. Mar Drugs 15(3): rising of temperatures, and this makes diversity and toxicity of these benthic- 72 this region more suitableGambierdiscus for tropical - dinoflagellates will provide a better 11. Lejeusne et al 2010. Trends Ecol Evol 25: 250-60 species [11]. A recent study describes characterization of health risks tak a high diversity of spe ing into consideration climate change cies in the Canary Islands which would trends. Authors suggest that this genus is not a recently- Acknowledgements Àngels Tudó, Anna Toldrà, Karl B. Andree, introduced taxon in that area, although - María Rey, Margarita Fernández-Tejedor, Mònica Campàs & Jorge Diogène, IRTA, Ctra. climate change may contribute to in - crease the populations density [2]. The authors acknowledge financial sup Poble Nou, km 5.5, 43540 Sant Carles de la Gambierdiscus - Ràpita, Spain It will be important to understand port from the European Food Safety Au the origin of Gambierdiscus in the Medi thority (EFSA) through the EUROCIGUA E-mail: [email protected] terranean and. the effect that climate- project (GP/EFSA/AFSCO/2015/03), change may have on the Ministerio de Economía, Industria populations Moreover, it will be neces y Competitividad (MINECO) through Forthcoming Events

- A. Deliver National Reports on harmful F. HABs and the EU Marine Strategy algal events and bloom dynamics for Framework Directive (MSFD). Ap the year 2017. proaches in Europe to including B. Review progress and summary of HABs in the assessment of the Good- fish killing algae activities underway Environmental Status for the EU ICES-IOC Working Group on during the reporting period 2017– ­Marine Strategy Framework Direc Harmful Algal Bloom Dynamics 2020. tive. (WGHABD) C. Updating of the ICES-PICES-IOC G. New results about how physical, Harmful Algal Event Database (HAE- chemical and biological interactions DAT. control the dynamics of selected D. Global HAB Status Report for the harmful micro-algae. The next ICES-IOC WGHABD, hosted- North Atlantic area: report using H. Ciguatera Fish Poisoning (CFP) in the by Margarita Fernandez-Tejedor at data and products generated from ICES area: review new developments

IRTA, Sant Carles de la Rapita, Tarrard HAE-DAT and supplementary time in methodology to research the issue, gona,th Spain, will meet from the 23 series data as appropriate. modelling efforts, risk assessments – 26 April (inclusive) to work on the E. New findings in harmful algal bloom to protect human health, initiatives followingHARMFUL ALGAE NEWSterms NO. 59 of / 2018 reference: dynamics. in other bodies (IPHAB, PICES etc.). 23 Comparison by light microscopy and qPCR of potentially ­ichthyotoxic microalgae in Danish on-shore lagoons producing European flounder Platichthys( flesus): Pros and cons of microscopical and molecular methods

able detected 8 out of the 11 potentially fish killing species (Fig. 2A, C). There were very few cross overs in terms of species identification between the two Dictyochamethods. LM failed to detect seven of- the potentially ichthyotoxic species and was the only microalga de tected by both methods. DespiteDictyocha this, speculumthe cell densities differed markedly. The qPCR assay only detected

cells in-1 lagoon six at a density of 2,600Dictyocha cell L , whereas LM detected much higher numbersPseudochattonella of 56,560 farci and- Fig. 1. Lagoon used for production of European flounder at Fishlab, Denmark. men36,210 P. verruculosa‘sp.’ in lagoons 1 and 6, respectively. - and P. farcimen wereP. verruculosa detected in all lagoons by qPCR and a succession Evaluation of phytoplankton communi ples for phytoplankton analyses were pattern for° P. farcimento ties is frequently used to determine the taken twice weekly from 7 March to 18 was evident. When temperatures were ecological status of water bodies. Hence,- May, 2017. A total of 55 samples were °low (8-9 C) was dominantP. species diversity of phytoplankton has examined by LM and qPCR. Potentially- farcimenbut as temperatures increased above 9 become an integral component of na ichthyotoxic species (Table 1) were C a switch occurredP. verruculosa and a decline in - tional assessment programs. The most identified either quantitatively or quali concentration coincided with commonly used technique for counting tatively by qPCR using species-specific- an increase in cell num phytoplankton is the Utermöhl method primer sets and hydrolysis (Taqman)- bers (Fig. 2A, C). During the production- [1] and the precision of the enumeration probes. Results from qPCR were com period, lagoons were replenished with is evaluated with standard statistical pared to microscopic cell counts per an addition of ca. 10% of newly col analyses techniques [2]. Despite long- formed by Fishlab. Examples of two lected seawater.Pseudochattonella The additional water term reliance on the Utermöhl method,- representative lagoons are provided in appeared to re-inoculate each lagoon new techniques for algal identification- Fig. 2A-D. with additional cells. and enumeration are continuously be Data gathered from all six lagoons- Fish survival rates in the lagoons were- ing explored. Particularly, the recent ex revealed ca. 20 groups or species by LM between 0.5 and 13 %. plosion in molecular tools has resulted (not shown) and the qPCR assays avail In terms of monitoring, the discrep in an influx of alternative methods (e.g. Lagoon number real-time qPCR, microarrays and FISH- Species 1 2 3 4 5 6 FC). While many of these methods are Alexandrium tamarense promising, their results often differ Alexandrium ostenfeldii + + + + from those of the conventional method Karenia mikimotoi that they were intended to supplement - Prymnesium parvum + + (or replace). Given the importance of- algal community assessments, signifi Pseudochattonella farcimen + + + + + + cant efforts have been put into the qual Pseudochattonella verrculosa + + + + + + ity assurance of phytoplankton counts Karlodinium veneficum + + + + + + [3]. This has led to the development of Pfiesteria shumwayae + + + + standardised procedures. Pfiesteria piscicida + + + + + This study aimed to compare cell Luciella masanensis counts of seawater samples by light Dictyocha speculum + microscopy (LM) and qPCR from on- shore production lagoons of European Table 1. List of species for which qPCR assays are available for quantitative (cell abundance) - and qualitative (presence/absence) of potentially ichthytoxic microalgae in this study. ‘+’ = Flounder (Fig. 1). A total of six lagoons- presence of a species in the lagoons used for production of European flounder. had been filled with untreated seawa ter from the nearby Limfjorden. Sam HARMFUL ALGAE NEWS NO. 59 / 2018 24

1 Fig. 2. Comparison of phytoplankton species identification and cell densities (cells L-1) by qPCR (A and C) and light microscopy (target species and groups which potentially could comprise ichthyotoxic organisms) (B and D) in lagoon 1 (A and B) and lagoon 6 (C and D), respectively. The right Y-axis shows the water temperature during the growth period. Presence of Pfiesteria piscicida and P. shumwayae at specific dates are indicated by H. Information on the number of flounder larvae added and the survival rates in percentage at the harvest dates are provided at the top of A and C.

- - ancy between the qPCR and LM results from sampling to counting comes with costs and equipment requirements (ba- is obviously quite worrying. There are its own forms of variation, we will criti sically an inverted microscope and a clearly problems with one or even both cally analyse each method and discuss settling chamber). One of the main ad- methods and it raises many questions Microscopicpotential problems analysis and sources of error. vantages of LM over qPCR is its ability to over their accuracy. When evaluating identify at least theoretically all organ phytoplankton data, each inaccuracy- isms present in the sample in contrast- associated with sampling, sub-sampling Traditionally light microscopy has been to qPCR, which will only target species and sample preparation should be tak the gold standard for phytoplankton of interest, e.g. toxic or nuisance organ enHARMFUL into ALGAE account. NEWS NO. 59 As / 2018 each individual step identification due to its relatively low isms. If there were any new species 25 - - present in a sample then qPCR would- are taxa specific. It is not always easy to- is one of the key determinants for ob- miss those probably due to lack of a obtain reliable estimates from fixed ma taining reliable and reproducible data.- developed assay. However, LM does re terial; preservatives can alter the sam As with microscopy fixatives and stor quire high levels of taxonomic skills and- ple in various ways creating a biased age techniques play a large part in qual the precision in identification is only as measurement. Lugol’s iodine [4] has- ity of the samples. Fortunately for short accurate as the taxonomist allows. Dif- long been the fixative of choice due to term storage Lugol’s iodine is the most ferent taxonomists trained in different its relatively low toxicity and high sta ideal fixative and the same sample can ways using different identification lit bility. However, it is known to introduce be used for both microscopic and qPCR erature can cause large person-person artefacts such as changes to cell size, a analysis [7]. Before amplification DNA- differences. The ease of identification reduction in cell number and in some must be extracted from the cellular is also species dependent. For example instances it may fail to preserve certain material and commonly commercial ex highly plastic species, or those with a taxa all together [5-6]. Each alternative traction kits are applied. To get purified- variable life cycle areDictyocha harder to speculum identify fixative comes with its own issues [7]. - genomic DNA the sample must undergo- and can often be easily misidentified.- Settling chambers themselves can a number of steps to lyse the cells, re- The nakedPseudochattonella form of - be another source of variation. For re- move contaminants and purify the re can easily be confusedAlexandrium with the round liable cell counts, specimens must be sulting DNA. In cases where the purifica ed cells of . Some spe completely randomly distributed with tion step is inefficient the resulting DNA cies of the genus cannot in the chamber. If cells do not follow a- may not be representative of the sample be identified to species level due their poisson distribution then it will bias the and/or contain compound(s) that will very subtle morphological differences enumeration and any statistical analy- cause assay interference. In these cases in their thecal plates. When dealing- sis will be affected. - the performance of the reaction will- with toxic species, false positives are To prevent uneven settling, the sam be sub-optimal, causing a reduction in less problematic but can cause sub ples must be at a constant tempera the sensitivity and/or amplification ef stantial financial losses if they result in ture during the settling period. For a ficiency. Inhibition of amplification can the closure of a fishery, but when toxic- higher chance of getting a well-mixed- occur in different ways. Firstly, when or problematic organisms are missed distribution then samples must first high molecular weight compounds, e.g. completely this could have dire con be homogenised. The best way to ho humic acids or complex carbohydrates,- sequences. To reduce confusion, each mogenise a sample is the ‘Paul-Schatz’- combine with metal ions to sequester taxonomist should be provided with a figure of eight rotation method where the nucleic acids away from the poly checklist of common species with up to- samples are mixed 100 times in a rhyth merases and prevent amplification. date taxonomic names. mic pulsating motion. Even when all While some molecules block or inhibit Undertaking frequent inter-com precautions are taken, it is still almost the polymerase or alter the specificity parison exercises, e.g. the ring test or impossible for cells to be randomly- of the primers, inhibitors which block- the International Phytoplankton Inter-- distributed due to issues such as cell or delay polymerase activity are highly- comparison (IPI) exercise, provide clumping caused by polysaccharide fi problematic and they can lead to an un- feedback on how individual labora brils or inconsistent settling conditions. derestimation of material in the sam tories and taxonomists perform. This Due to radial abundance gradients cell- ple or false negatives [9]. Typical ap- forum also provides an opportunity to- abundances at the periphery can be up proaches to combat inhibition include convene a discussion on nomenclatural to 50% lower than at the center, caus alternative DNA extraction kits, dilu changes and new technological advanc- ing a settling bias. Uneven settling will tion, specialised polymerases, addition es in monitoring techniques. affect the counting strategy. For any of adjuvants and internal controls. As error can be introduced in vari counting strategy a predetermined When designing a qPCR assay it ous different forms, in order to get the number of units must be observed. The- is important to select an appropriate- most accurate and reproducible results number of units differs depending on target and to design specific primers each individual step from: collection,- the organism and the research objec with no cross reactivity with other or storage, subsampling, homogenisation, tives. Typically to reach an accuracy of ganisms. This study used previously filling the chamber, settling and count 10% at least 400 cells must be counted validated species-specific hydrolysis- ing strategies all require their own [8]. Whole chamber cell counts should probes in combination with primers to- standardised protocols. - be carried out where possible but other add an extra level of specificity. To en- All aspects of the protocol need to counting strategies are often used such sure accuracy, time is required to opti be consideredPseudochatonella from the storage contain qPCRas transects and random fields. mise the efficiency of the assay and vali ers to the type of fixative used. Many - date it multiple times. This sometimes- cells e.g. spp. are means altering the constituents of the- sticky and can adhere to plastic walls- When carefully designed, with optimi- master-mix used, e.g. nucleotides, mag and as plastic bottles is often preferred- sation and validation, qPCR assays are nesium chloride or polymerase con- over glass especially when transport highly accurate and sensitive, but with centrations. This is extremely relevant ing samples this can become problem out due care and optimisation, qPCR when tackling issues arising from mul atic for accurate cell enumeration. The can be plagued by reproducibility and tiplexing assays [9-10]. choice of preservative is important and reliability problems. Once optimised users can still face a often the optimal preservation methods The quality of the starting material number of precisionHARMFUL related ALGAE NEWSchallenges. NO. 59 / 2018 26 References - 1. Utermöhl H 1958. Limnol 9:1–38 As qPCR measures genetic material qPCR experiments in the laboratory and 2. Lund J W G et al 1958. Hydrobiologia 11: rather than viable cells an over estima are designed to improve experimental 143–170 tion of cell numbers can occur due to workflow and should be followed when 3. Rott E et al 2007. Hydrobiologia 578: - 141–146 the inclusion of dead or dying cells.- Conclusionsdesigning any qPCR assay [12]. 4. Throndsen J & A Sournia 1978. Phy- Problems may also occur when target toplankton Manual (Monographs on ing multiple copy genes where the or Oceanographic Methodology, UNESCO, ganism carries different numbers of the Clearly despite efforts to standardise Paris), 337 pp. target depending on nutritional status, procedures for both techniques there 5. Leakey R J G et al 1994. J Plankton Res - 16:375–389 stress or replication stage. This can lead- are still many problems affecting the 6. Stoecker DK et al 1994. Mar Ecol Prog to an over or under estimation of total accuracy and the quality of the re Ser 110: 293–299 cell numbers. Common problems asso sults. The comparison of enumeration 7. Eckford-Soper L K & N Daugbjerg 2015. Harmful Algae 42: 52–59 ciated with cell number enumeration- techniques that was carried out in this and copy number do not occur until late study has highlighted the difficulties in 8. Karlson et al 2010. IOC Manuals and Guides 55, UNESCO, Paris, 110 pp exponential-stationary phase mean obtaining comparative data especially- 9. Webb S 2013. Biotechniques 55: 165–168 ing that cell numbers can be accurately of small-sized, ichthyotoxic microalgae. 10. Eckford-Soper L K & N Daugbjerg 2015. quantified until this point [11]. One way Enumeration by LM missed many im Harmful Algae 48: 37-43 of potentially overcoming this issue it to portant species, which emphasizes how 11. Eckford-Soper L K & N Daugbjerg 2016. J Phycol 52: 174-183 use standards created using cells from difficult it is to identify phytoplankton 2. Bustin S A et al 2009. Clin Chem 55: all parts of the growth curve to produce from Lugol’s fixed material. We are now 611–622 an ‘average’ copy number. However, this moving into the era of ‘bio-monitoring will decease the overall accuracy of the 2.0’ and with the reduction in costs for- assay. meta-barcoding based techniques it is Authors - Lisa Eckford-Soper & Niels Daugbjerg, As with microscopic analysis qPCR still to be seen if these molecular tech Dept of Biology, University of Copenhagen, requires standardisation/normalisa niques will eventually replace LM. Yet Universitetsparken 4, 2100 Copenhagen Ø, tion for both the laboratory protocols improvements need to be made across- Denmark Minimumand statistical information analysis for strategies. publication To the board for all techniques. The low Louise Nørremark & Kirsten Engell-Sørensen, ofaid Quantitative this, in 2009 Real-time the MIQE PCR guidelines, Experi- survival rate of European flounder ob served in the 2017 production may be Fishlab, Terp Skovvej 107 b, 8270 Højbjerg, ments - Denmark. explained by the diverse assemblage of- , were published. These guide potentially ichthyotoxic microalgae in Email: [email protected] lines are designed to ‘encourage better the lagoons. In previous years the sur experimental practice’. The guidelines vival rate has been 40-50%. establishes a framework for conducting Recent Advances in the Analysis of Marine Toxins

- - Recent Advances in the Analysis of Ma • Immunoassays and optical rine Toxins, Volume 78, edited by J. Dio- biosensors (visual, SPR, géne and Monica Compás, the newest fluorescence) for marine- release in the Comprehensive Analyti toxins, and cal Chemistry series, provides chapters • Electrochemical biosen from well known authors in the field. sors for marine toxins. Updated sections include topics such as: Details on the table of • The importance of toxin detection- https://www.elsevier.com/content and others can and quantification: environmental books/recent-be found at: issues, public health, food safety, ani advances-in-the- mal health, bioterrorism, bioactive analysis-of-marine-toxins/­ compounds, medical approach, an diogene/ LC-MS/MS analysis of marine toxins; 978-0-444-63941-7 • Animal bioassays: identification of- toxins and mechanism of action; • Receptor binding assays for the anal ysis of marine toxins;

HARMFUL ALGAE NEWS NO. 59 / 2018

27 The Cawthron Institute Culture CAWTHRON FOUNDATION is a char- ­Collection of Micro-algae (CICCM) itable trust which raises donations, bequests and endowments for public-good science. The Founda- - tion also funds scholarships to sup- - - port talented emerging scientists The CICCM is designated as a “national- quote Kendall, “It has been a rewarding ly significant database” by the New Zea experience helping this team to con and delivers community education land government and so receives par struct the CICCM database and I have programmes: http://www.cawthn. org.nz/foundation/scholarships/ tial funding for its continued existence. been fascinated to learn how research- Isolates from 13 classes of micro-algae involving the collection has increased are maintained either as live cultures understanding of marine and freshwa- - or cryopreserved. The approximate 500 ter biotoxins in New Zealand.” isolates include benthic, epiphytic and- The database, built using Micro industry and by supporting capabil- planktonic marine harmful algal bloom- soft’s ‘Share Point Online’ platform, will- ity development in all aspects of HABs species as well as a collection of fresh meet the strategic aim for all collections through provision of cultures to post water cyanobacteria. Most of the iso- and databases in Australasia to be eas graduate students and post-doctoral lates are toxin producers and there are- ily interrogated. Initially the database Acknowledgementsresearchers. also species that are unique to New Zea- will only be accessible ‘in-house’, but - land waters. Every isolate in the collec will enable the curators to provide full tion has an associated body of informa www.cultures.cawthron.information with ordered isolates. On- The 2017/18 Kathleen Curtis scholar- tion. The collection underpins research line ordering continues via the website: ship was made possible with generous programmes, for example the Safe New- financial support from the Emery Fam Zealand Seafood programme, and is the The CICCM is distinct from larger- ily. focus of many student projects both na collections in the Northern Hemisphere Authors Lesley Rhodes, Kirsty Smith, Kendall Cawthrontionally and Foundation internationally. - and this has attracted commercial in- Morman, Krystyna Ponikla & Sarah Over Kathleen the summer Curtis of 2017/18 the terest in potential micro-algal products. (see box) pro A key user of the collection is the Caw Challenger, Cawthron Institute, Nelson, New Zealand vided a scholarship for thron Natural Compounds group which an undergraduate student to assist the aims to make a wider range of toxins - E-mail: [email protected] curators in populating a new CICCM- available to the global market through a- database with the relevant information partnership with Sigma-Aldrich to sup for each isolate, including site of isola- ply algal toxins via their renowned cata tion, culturing tips, molecular and toxin logue (see HANews 51 August 2015). information and associated publica The CICCM will continue to generate- tions. The scholarship was awarded to economic benefits to New Zealand by- Kendall Morman, a biomedical science underpinning the development of im student at the Auckland University of proved and cheaper toxin and molecu Technology, New Zealand (Figure 1). To lar tests for the New Zealand shellfish

Fig. 2. A selection of cultures held in the Fig. 1. Kendall Morman being welcomed at a powhiri (a Maori welcoming ceremony) CICCM. From top, Vulcanodinium rugosum led by Harvey Ruru, Cawthron Institute’s kaumatua (senior elder member of the (non-motile cells), Ostreopsis siamensis, and group) Phormidium sp

HARMFUL ALGAE NEWS NO. 59 / 2018

28 - The XVIII International Conference wish to include a picture of your item. A on Harmful Algae is approaching! It is- ISSHA’s CORNER list of items will bewayne.litaker@noaa. presented on the IS time for nominations for achievement- govSHA website before the conference. Dr. awards (Yasumoto Life Time and Pat Wayne Litaker ( rick Gentien Young Scientist), registra ) can provide a letter template to use tion to the conference as a student if and awardees are announced during to request donations from companies. you wish to participate in the Maureen the ICHA closing ceremony. The address where items may be sent in Keller Award competition and fund- The presentations are evaluated- advance of the meeting will be provided Nominationsraising for the forauction. the 2018 Yasumoto based on the following criteria: on the conference website. Lifetime Achievement Award and • Clearly stated hypothesis or objec the Patrick Gentien Young Scientist tive - Award • Scientific merit and originality • Coherence (consistency between in troduction, data, conclusions) ISSHA members are invited to submit • Intelligibility (selection of important nominations for the Yasumoto Lifetime points, brevity, accuracy, and clarity Achievement Award and the Patrick of expression) Gentien Young Scientist Award. Furtherwww. • Presentation (appearance, layout, issha.orginformation on the appropriate profile clarity) of the nominees can be found at - (click on “Awards”). Candidates can ask to behttps://www. considered Any ISSHA member in good stand- icha2018.com/abstracts/call-for-abfor these awards during the abstract- ing may submit nominations for either stracts/50submission) process ( ISSHA conference achievement award, which should in clude a description of the nominee’s ISSHA AUCTION https:­// contribution (not more than one page).- Donations for the ISSHA Auction www.icha2018.comPreparation of the ISSHA conference 21- Please, make sure that you and your 2018 26/10/2018,plenary Nantes, speakers France ( nominee have renewed your member - ) is progressing well. ships for the period 2017-2018.marta@ Five have already- icm.csic.esNominations should be sent by e- ISSHA auctions have been a tremen confirmed their attendance: mail to Dr. Marta Estrada ( dous success during past International • Prof. Elena Litchman (Univ. Minne ) Chair of the Committee on Conferences on Harmful Algae and are- sota, Michigan State University, USA) Achievement Awards. Deadline: May becoming established as a tradition. It- • Prof. Bill Gerwick (Univ. California- 15, 2018. is an opportunity for conference partic San Diego - Skaggs & Scripps, USA) Please write “ISSHA Achievement ipants to bid on a broad range of donat- • Prof. Erik Jeppesen (Aarhus Univer Awards 2018” in the message subject. ed items such as books, signed reprints, sity, NL) Nominations will be considered by the photos, jewelry, T-shirts, paintings, al- • Dr. Mireille Chinain (Institut Louis ISSHA Council, and the awards will be gae-related items, liquor, and scientific Malardé, French Polynesia) presented at the 18th ICHA Conference equipment. The auction items are do • Prof. Thomas Hartung (John Hopkins TheNantes, Maureen France, Keller 21-26 StudentOctober 2018.Award nated by ISSHA members from around University, USA)session topics for best student presentations at the world. Funds collected during the https://www.icha2018.com/scientific- ICHA auction are an important income for program/session-topics/35Check out the here: ISSHA. ISSHA was able to offer partial travel support to 25 students to attend abstract submission The Maureen Keller Award is given the Conference in Florianópolis, Brazil deadline to the outstanding graduate student ([email protected] October 2016). - And note the oral and poster presentation at the Please, contact Dr. Wayne Litaker (15/4/2018) ICHA2018. Conference participants are- ( ) who is re https://www.icha2018.com/abstracts/ chosen in advance to review students’ sponsible for the auction about your call-for-abstracts/50Please submit abstracts here: presentations. The evaluations are pro donations (including ”ISSHA Auction cessedHARMFUL ALGAE by NEWSthe NO.ISSHA 59 / 2018 Awards Committee 2018” in the message subject). You may 29 Forthcoming Events

First announcement of the 11th International Conference on Toxic Cyanobacteria (ICTC)

11th Internation- alWe Conference are pleased on to disseminateToxic Cyanobacteria the first announcement of the -

(ICTC) that will be held in Krakow, Po- land from May 5 – 10, 2019. The ICTC is a periodic scientific meeting that in cludes members of the international- future http:// community and focuses on the science ictc11.org/ andLearning study of fromcyanotoxins the past andto predict toxic thecy ”. Please mark your calendars and May, 2019! Conference website: anobacteria. The theme of the ICTC 11 make plans to join us in Krakow during is: “

Harmful Algal Bloom (HAB) - sessions at the upcoming 2018 - ASLO summer meeting in Victo- students and colleagues, his reach ex • Cyanobacterial ecology as a basis for ria, British Columbia tended into phytoplankton-zooplank- their mitigation and control under ton interactions and the structuring of global change (SS51). planktonic ecosystems. With his pass - ing in the spring of 2017, we invite the The first session will focus on broad Ted Talks: The career, contributions, community to join us in a celebration- HAB research and management solu and impact of Theodore J. Smayda of his career with contributions on any tions, whereas the second session will - aspect of phytoplankton or oceanogra focus specifically on cyanobacterial and- Session SS04 [email protected] by: ) Keywords:phy that were inspired or affected by algal metabolites. The third session will Tracy A. Villareal, The Universityjyoder@whoi. of Tex his work. focus on aspects of cyanobacterial ecol- eduax at) Austin ( Biogeography, Ecosystem, ogy in relation to environmentalhttps://aslo.org/ change James A. Yoder, WHOI ( Ecology, Marine, Phytoplankton - victoria2018/special-sessionsand bloom management. Session ab [email protected]) stracts are located at Edward G. Durbin, Univ. of Rhode Island There will be another three Harmful Al ( gal Bloom (HAB) related sessions: - • Crossing disciplinary boundaries If you are a HAB researcher planning Ted Smayda’s career in marine science- across the freshwater-marine contin on attending the 2018 ASLO summer spanned 60 years on topics ranging uum to advance the understanding of- meeting, please consider submitting from phytoplankton suspension, ecol HABs (SS71). an abstract to eitherhttps://aslo.org/victo session. Abstracts- ogy, succession, community structure, • Cyanobacterial and algal metabo ria2018/mainare due 16-Feb-2018; registration can growth rates, biogeography, and the lites: occurrence, ecology, prediction, be completed at problems of harmful algae. Through his and management (SS07), and HARMFUL ALGAE NEWS NO. 59 / 2018 30 Ecologi- cal Studies, Analysis and Synthesis - GEOHAB, under the sponsorship of will be published by Springer ( the Intergovernmental Oceanographic- , Vol Commission (IOC) of UNESCO and the- ume 232) in spring 2018. Scientific Committee on Oceanic Re- As GEOHAB ended, the community- search (SCOR), focused on the physi recognized the benefits ofinternational ological, behavioral, and genetic charac cooperation and encouraged continua- teristics of harmful microalgal species, tion of a new program built on legacy and the interactions between physical provided by GEOHAB, but now includ and other environmental conditions- ing freshwater systems, and addressing that promote the success of one group- the effects of HABs on human society of species over another. GEOHAB im now and in a rapidly changing world.- plementation was based on a multidis- Thanks to the invaluable support of IOC ciplinary, multi-scale and comparative- and SCOR, the new programme, Global approach, and stimulated the develop HAB, was launched in January 2016.- International Coordination of ment of new experimental, observa Since then, the GlobalHAB Scientific Research on Harmful Algal tional and modelling tools. - Steering Committee has been elabo- Blooms During the 15 years of GEOHAB’ rating the Science and Implementation- From GEOHAB to GlobalHAB activity, the international communi- Plan that provides a scientific frame ty has contributed to understanding work for the integration and coordina - mechanisms underlying HAB popula tion of research and expertise of many tion dynamics within an ecological and- individual scientists in the study of International cooperation is fundamen oceanographic context. At the end of HABs in different aquatic ecosystems.- tal to advance understanding of HAB- GEOHAB,Oceanography advances and existing chal As in GEOHAB, the direct implication of dynamics and to improve our ability lenges were summarized in six papers the international community in the pro to predict them. Fostering this interna- in http://tos.org/oceanography/ (The Official Magazine gramme is fundamental for GlobalHAB tional cooperation was the mission of issue/volume-30-issue-01of the Oceanographic Society) in March success. Informationhttp://www.globalhab.info on the GlobalHAB GEOHAB (Global Ecology and Ocean 2017, ( lobal Ecology and Oceanog- programme and how to participate can ography of Harmful Algal Blooms), the- raphy of Harmful Algal Blooms). A GEOHAB Elisabe found Berdalet, at Chair, and the GlobalHAB . first worldwide research programme synthesis, G Scientific Steering Committee focusing exclusively on harmful ma , edited rine microalgae. From 1998 to 2013, by Patricia M. Glibert and co-editors

GlobalHAB GOAL AND MISSION -

The overall Goal of GlobalHAB is to improve understand ing and prediction of HABs in aquatic ecosystems, and management and mitigation of their impacts. -

The Mission of GlobalHAB includes the following ele ments: -

• Foster international coordination and cooperative re search to address the scientific and societal challenges of HABs, including the environmental, human health and economic impacts, in a rapidly changing world. • Consolidate linkages with broader scientific fields and other regional and international initiatives relevant to The Themes integrated in GlobalHAB range from small-scale- HABs. (e.g., cellular) subjects (e.g., biodiversity, adaptive strategies) • Foster the development and adoption of advanced and- to studies at ecosystem scale and climate change-related pro cost-effective technologies. cesses. • Promote training, capacity building and communica tion of HAB research to society. • Serve as a liaison between the scientific community, stakeholders and policy makers, informing science- based decision-making.

HARMFUL ALGAE NEWS NO. 59 / 2018

31 18th International Conference on Harmful Algae www.icha2018.com

IMPORTANT DEADLINES

Abstract submission deadline: 15 April 2018 Early bird registration: 15 July 2018 Get the 17 ICHA Proceedings at www.issha.org

Eds-in-chief Compiled and edited by

Beatriz Reguera, IEO, Vigo, Spain Beatriz Reguera, Instituto Español de Oceanografía RegionalEilen Bresnan, Editors MARLAB, Scotland, UK (IEO), Subida a Radio Faro 52, 36390 Vigo, Spain Tel: [email protected] 986 492111 [email protected] Fax: +34 986 498626 • Caribbean: Ernesto Mancera Email: [email protected] and • Europe: Philip Hess Eileen Bresnan, Marine Scotland, Victoria Road, [email protected] ­Aberdeen AB1 9DB, Scotland • India: K.B. Padmakumar [email protected] Tel.: [email protected] 122 4876544 [email protected] Fax: +44 1224295511 • Western Pacific: Rhodora Azanza , Email: [email protected] Po Teen Lim • North Africa: Hamid Taleb The opinions expressed herein are those of the [email protected] authors indicated and do not necessarily reflect - • North America: Patricia Tester the views of UNESCO or its IOC. Texts may be freely [email protected] reproduced and translated (except when reproduc and Jennifer Martin tion or translation rights are indicated as reserv­ed), [email protected] provided that mention is made of the author and • South America: Luis Proenca Projectsource and Coordinator a copy sent to the Editors. [email protected] • Africa: to be [email protected] • South Pacific: Mireille Chinain Henrik Enevoldsen, IOC Science and Communication and Lesley Rhodes Centre on Harmful Algae, ­University of Copenhagen, [email protected]­parken 4, 2100 Copenhagen Ø, Denmark Please feel free to contact any of the editors if you Tel.: +45 33 13 44 46 have article, ideas for article or special issues and we Lay-outE-mail: Deadlinewill work with you!

Department of Biology, University of Copenhagen, Deadline to submit material for HAN 60: Denmark 15 May 2018 The publication of Harmful Algae News is sponsored by the Department of Biology, University of Copenhagen

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