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 Fig. 1. Lagoon used for production of European flounder at Fishlab, Denmark. Evaluation of phytoplankton communities is frequently used to determine the ecological status of water bodies. Hence, species diversity of phytoplankton has become an integral component of national assessment programs. The most commonly used technique for counting phytoplankton is the Utermöhl method [1] and the precision of the enumeration is evaluated with standard statistical analyses techniques [2]. Despite longterm reliance on the Utermöhl method, new techniques for algal identification and enumeration are continuously being explored. Particularly, the recent explosion in molecular tools has resulted in an influx of alternative methods (e.g. real-time qPCR, microarrays and FISHFC). While many of these methods are promising, their results often differ from those of the conventional method that they were intended to supplement (or replace). Given the importance of algal community assessments, significant efforts have been put into the quality assurance of phytoplankton counts [3]. This has led to the development of standardised procedures. This study aimed to compare cell counts of seawater samples by light microscopy (LM) and qPCR from onshore production lagoons of European Flounder (Fig. 1). A total of six lagoons had been filled with untreated seawater from the nearby Limfjorden. Sam24 ples for phytoplankton analyses were taken twice weekly from 7 March to 18 May, 2017. A total of 55 samples were examined by LM and qPCR. Potentially ichthyotoxic species (Table 1) were identified either quantitatively or qualitatively by qPCR using species-specific primer sets and hydrolysis (Taqman) probes. Results from qPCR were compared to microscopic cell counts performed by Fishlab. Examples of two representative lagoons are provided in Fig. 2A-D. Data gathered from all six lagoons revealed ca. 20 groups or species by LM (not shown) and the qPCR assays avail- Species Alexandrium tamarense Alexandrium ostenfeldii Karenia mikimotoi Prymnesium parvum Pseudochattonella farcimen Pseudochattonella verrculosa Karlodinium veneficum Pfiesteria shumwayae Pfiesteria piscicida Luciella masanensis Dictyocha speculum 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 methods. LM failed to detect seven of the potentially ichthyotoxic species and Dictyocha was the only microalga detected by both methods. Despite this, the cell densities differed markedly. The qPCR assay only detected Dictyocha speculum cells in lagoon six at a density of 2,600 cell L-1, whereas LM detected much higher numbers of 56,560 and 36,210 Dictyocha sp. in lagoons 1 and 6, respectively. Pseudochattonella farcimen and P. verruculosa were detected in all lagoons by qPCR and a succession pattern for P. farcimen to P. verruculosa was evident. When temperatures were low (8-9 C) P. farcimen was dominant but as temperatures increased above 9 C a switch occurred and a decline in P. farcimen concentration coincided with an increase in P. verruculosa cell numbers (Fig. 2A, C). During the production period, lagoons were replenished with an addition of ca. 10% of newly collected seawater. The additional water appeared to re-inoculate each lagoon with additional Pseudochattonella cells. Fish survival rates in the lagoons were between 0.5 and 13 %. In terms of monitoring, the discrep- 1 Lagoon number 2 3 4 5 + + + + + + + + + + + + + + + + + + + + + + + + + + + 6 + + + + + + + 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. + = presence of a species in the lagoons used for production of European flounder. HARMFUL ALGAE NEWS NO. 59 / 2018 Harmful Algae News An IOC Newsletter on Toxic Algae and Algal Blooms No. 59 - February 2018 www.ioc-unesco.org/hab Content 25 years of HAN and IPHAB...... 1 25 YEARS Harmful Algae News was first published in early 1992 in response to requests from the participants at a number of IOC meetings and pacted by harmful algal events. Since Harmful Algae News turned 20 years old in 2012, it has been a web based e-newsletter which meant longer issues were possible and back issues easily accessible. We are currently working on a searchable index for all Harmful Algae News issues. The start of Harmful Intergovernmental Panel on Harmful Algal Blooms also turned 25! During 1992, the same Year as the IOC published the first issue of Harmful Algal News, it also established an Intergovernmental Panel on Harmful Algal Blooms (IPHAB) which has met every second year since it was formed. The Panel is com A retrospective look at the early days of HAB cyst research, and a look to the future On this occasion of the 25th anniversary of the publication of Harmful Algae News, several of us were asked to look backwards in time to some of the earlier days of HAB science. One area of study that has been a ma dormant stages and that these stages might be associated with certain bottom sediments. This then brings up the question, if benthic resting stages of certain dinoflagellates actually seed coastal red tides, are there localized areas of accumulation, or what we could call seedbeds?..........The poss day, we still do not know if there are other factors at work perhaps a density-dependent or quorum-sensing type of response, or even a response to the presence of grazers or parasites. Exploration of this response has long been limited by the constraints associated with laboratory cultures, but now 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 understanding of certain types o number of cysts in subsurface layers unable to germinate or emerge, presumably due to lack of oxygen or to the tortuous pathway posed by sediment grains and detritus. Many might think that major storms and waves can erode significant layers of sediment and transport cysts long distances, but here ag How do algal blooms kill finfish and how can we mitigate their impacts? Algal blooms, water discolorations and their association with fish kills have been recorded since historic times, such as the description in the Bible (1000 years BC) all the waters that were in the river were turned to blood. A Table. 1. Economic losses from algal blooms for finfish aquaculture in different parts of the world HAB species Chattonella Heterosigma Cochlodinium polykrikoides Heterosigma Chaetoceros Heterosigma Karenia digitata Karenia mikimotoi Country Financial Losses Japan Korea, China Canada British Col emergency harvest operations. To prevent the buildup of histamines, fish should be kept alive as long as possible during harvesting. This can be achieved by diluting algal concentrations via airlift upwelling, or by targeted in-pen emergency application of clays [23] that mop up ichthyotoxins at cla As part of the 25th anniversary issue of Harmful Algal News I am providing an overview of the IOC-UNESCO Taxonomic Reference List of Harmful Algae (www. marinespecies.org/hab/) and will highlight some of the problems which have faced or are facing the Intergovernmental Panel on Harmful Algal Blooms Dinophyceae the authors intended the new genus to be described according to the botanical nomenclature. The lack of a Latin diagnosis therefore made it invalid. The confusion has actually still not been resolved, and the problem needs to be discussed and decided upon by the International Nomenclatur geographically widespread species being able to form fertile offspring, while other populations of the same species are not. Molecular techniques have contributed very significantly to solving many taxonomic problems, but they have not resulted in the emergence of a finite species concept. We have t Algal toxin discovery, management and regulation over the last 25 years Algal toxins in the dark ages (pre-1992) From a historic perspective, knowledge about algal toxins can be divided into truly prehistoric occurrences such as known from paleontological studies [1-2] and more recent historic recor lar rapid increase in known analogues has been observed for the azaspiracid (AZA) group, with the first analogue described in 1998 [64] and a review in 2014 reporting 30 analogues [65]. Only three years later, over 50 analogues are known for this group, including novel phosphate derivatives [66-71]. Butterflies in Brazil Abstracts are not always reliable guides to authors intentions. They are not expected to reveal a great deal about the evidence to be deployed in support of the science, evidence that may not even exist before deadlines for writing abstracts! Nevertheless, as examples of a mino on a decadal time scale, and identified palaeoclimatic oscillations are not necessarily a useful guide to its interpretation. 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 trends, Karenia brevis might appear more often in the South Atlantic Bight of the US and Gymnodinium catenatum bloom more often in northwestern Iberian waters. There was also a warning by Barrie Dale germane to such speculations, that large scale climate models cannot predict local changes. Little m Red tides in Kamchatka coastal waters (Bering Sea, Russia) are a barrier for the salmon fishery and Pacific salmon Fig. 1. Map of Olyutorskiy Bay (Kamchatka, Bering Sea) where a red tide, reported by fishermen, occurred in July 2017. The stars denote fishery sites: red, affected by the bloom; green Fig. 2. Red tide in Olyutorskiy Bay on 15th July 2017 grounds of the Olyutorskiy Bay river basin performed during the second half of August showed an atypical distribution in the rivers. Maximal escapes were recorded in river basins located in the western and eastern parts of the Olyutorskiy Bay ar First report of Gambierdiscus in the Western Mediterranean Sea (Balearic Islands) Gambierdiscus (Dinophyceae) species are benthic dinoflagellates living in marine littoral zones of circumtropical areas and have recently been described in temperate waters [1]. Some species are producers of potent neu the SEASENSING (BIO2014-56024C2-2-R) project and the CERCA Programme/Generalitat de Catalunya. A. Tudó and A. Toldrà acknowledge IRTAURV-Banco Santander for their respective PhD grants (2016 PMF-PIPF-74 and 2015PMF-PIPF-67). The authors are grateful to Vanessa Castan and José Luis Costa for sampling 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 Fig. 1. Lagoon used for production of European flounder at Fishlab, Denmark. Evaluation o 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 present in a sample then qPCR would miss those probably due to lack of a developed assay. However, LM does require high levels of taxonomic skills and the precision in identification is only as accurate as the taxonomist allows. Different taxonomists trained in different ways using different identif As qPCR measures genetic material rather than viable cells an over estimation of cell numbers can occur due to the inclusion of dead or dying cells. Problems may also occur when targeting multiple copy genes where the organism carries different numbers of the target depending on nutritional status, The Cawthron Institute Culture Collection of Micro-algae (CICCM) The CICCM is designated as a nationally significant database by the New Zealand government and so receives partial funding for its continued existence. Isolates from 13 classes of micro-algae are maintained either as live cultures or c The XVIII International Conference on Harmful Algae is approaching! It is time for nominations for achievement awards (Yasumoto Life Time and Patrick Gentien Young Scientist), registration to the conference as a student if you wish to participate in the Maureen Keller Award competition and fundraisi Forthcoming Events First announcement of the 11th International Conference on Toxic Cyanobacteria (ICTC) We are pleased to disseminate the first announcement of the 11th International Conference on Toxic Cyanobacteria (ICTC) that will be held in Krakow, Poland from May 5 10, 2019. The ICTC is a per International Coordination of Research on Harmful Algal Blooms From GEOHAB to GlobalHAB International cooperation is fundamental to advance understanding of HAB dynamics and to improve our ability to predict them. Fostering this international cooperation was the mission of GEOHAB (Global Ecology and 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 Beatriz Reguera, IEO, Vigo, Spain Eilen Bresnan, MARLAB, Scotland, UK Regi