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Mires are specific freshwater habitats with great importance for global biodiversity. The present study investigated diatoms in different habitats of a Đon Močvar mire and their relationship with environmental parameters, as well as the use of diatom indicator values and the German Red List for conservation purposes. The study was conducted monthly from January to November 2012. Diatoms were sampled simultaneously with water for physical and chemical properties in three different habitats. A total of 50 diatom taxa were identified. Based on their relative abundance and diversity, habitat-related environmental factors shaped the diatom community when seasonal dynamics were less important. Multivariate analysis showed that chemical oxygen demand (COD) and conductivity were the most significant environmental variables describing diatom communities. The indicator values characterized the diatoms as a typical mire community and proved to be a reliable starting point for future monitoring and management. Diatom classification by German Red List revealed a dominance of species with priority for conservation, suggesting that the Đon Močvar should have conservation status. This study brings a better understanding of the response of diatoms to living in specific habitats and the basis for future studies on ecological characteristics of new or rare taxa.

Phytoplankton are considered to be one of the most sensitive indicators of the ecological status of lakes. Nowadays, it is essential to recognize the prospects of the molecular approach (eDNA metabarcoding) in phytoplankton community assessments and combine them with the existing traditional microscopy-based morphological approach before its standardization. In this study, the aim was to characterize the phytoplankton community of a natural karstic lake by combining and comparing the morphological and molecular approach to check the applicability of eDNA metabarcoding as a biomonitoring tool. A total of 51 phytoplankton taxa were found using the morphological approach, whilst the molecular approach discovered 97 ASVs that corresponded to the algal community. The comparability of both approaches in describing phytoplankton communities is evident in the designation of centric diatoms, dinoflagellates and cryptophytes as descriptive taxa. Furthermore, both approaches proved reliable in detecting functional groups (Lo, C, X2, X3) with similar ecological demands. Moreover, the results have shown that euphotic zone samples can be reliably exchanged by composite samples to provide an accurate characterization of phytoplankton communities in the euphotic zone. It was confirmed that eDNA metabarcoding is an applicable tool for biodiversity monitoring of a natural karst lake and should be used as a feasible supplement to traditional microscopy in the phytoplankton community assessments, with regards to the drawbacks of each method.

Phytoplankton represents one of the most important biological components of primary production, trophic interactions and circulation of organic matter in lakes and reservoirs. To contribute to the understanding of eutrophication processes and ecological status of the small, shallow Butoniga reservoir, a machine learning tool for induction of models in form of decision trees and rule-based models was applied on a data set comprising physical, chemical and biological variables measured at four stations. Two types of models were successfully elaborated, i.e. (1) model describing phytoplankton Phylum which describes and connects phytoplankton Phylum with phytoplankton abundance and biomass, and (2) model simulating phytoplankton biomass according to environmental variables which could be used in management purposes. Such models and their presentation contribute to a better understanding of the Butoniga reservoir ecosystem functioning.

A new marine strain of picoplanktonic algae, PMPFPPE4, was isolated from a mixed net-phytoplankton sample taken from the upper euphotic layer of the southeastern Adriatic Sea. Evaluation of the new strain included morphological investigation (by light and electron microscopy), phylogenetic analysis (utilizing plastid 16S rRNA and nuclear 18S rRNA genes), and physiological characterization (screening of pigment/lipid composition and capturing photosynthesis measurements). The new strain was proven to belong to the genusPicochlorumand the lipid composition revealed an unexpected accumulation of triacylglycerols, indicating an evolutionary adaptation for growth under unfavourable conditions. In addition, lipid remodelling in the exponential to stationary growth phase was characterized by an increased share of membrane-forming digalactosyldiacylglycerols and phosphatidylcholines. Maximum photosynthetic activity measured was at 30 degrees C, but the most rapid increase of photosynthetic activity was at lower temperatures (15-20 degrees C). Moreover, the thermotolerant strain did not exhibit photoinhibition below 40 degrees C and survived a one-month cultivation period in complete darkness. The strain's survival in low light and dark conditions suggests a potential shift from autotrophy to mixotrophy under unfavourable growth conditions. Thus, the unique physiological attributes represented by a high growth rate, thermotolerance, phototolerance and high triacylglycerol synthesis may render the strain highly attractive for biofuel production and growth in large outdoor systems.

According to the Water Framework Directive (WFD), biological quality elements constitute the basis for assessing the ecological status of surface waters. Biological communities are good indicators of water quality because they reflect environmental conditions over time and do not require frequent sampling. The main aims of this study were to assess the trophic and ecological status of the Lake Visovac (Krka River hydrosystem, South Croatia) using phytoplankton together with supportive physico-chemical parameters, eutrophication status and impact indicators. We also tested the viability of chlorophyll-a (Chl-a) and Hungarian Lake Phytoplankton Index (HLPI) as proposed metrics in the standard regulatory monitoring procedure. The phytoplankton samples were taken and Chl-a and physical- chemical parameters measured on a monthly basis from April to September in 2016 and 2019. In 2016, the dominant species was Pantocsekiella ocellata (Pantocsek) K.T.Kiss & Ács, while in 2019 it was accompanied by Ceratium hirundinella (O.F.Müller) Dujardin. According to Chl-a, Lake Visovac was within the limits of oligo- to mesotrophic status. Based on the HLPI index, the Lake ecological status was assessed as Good. Chl-a showed a statistically significant positive correlation with temperature, while HLPI was positively correlated with oxygen and Secchi depth. We propose the use of Chl-a for rapid bioassessment on a weekly basis, whilst the more complex HLPI index should be applied monthly. Further improvement of the confidence level of the metrics used to assess the ecological status and a comprehensive revision of boundaries for included indicators is of fundamental importance.

In lake ecosystems, changes in eukaryotic and prokaryotic microbes and the concentration and availability of dissolved organic matter (DOM) produced within or supplied to the system by allochthonous sources are components that characterize complex processes in the microbial loop. We address seasonal changes of microbial communities and DOM in the largest Croatian lake, Vrana. This shallow lake is connected to the Adriatic Sea and is impacted by agricultural activity. Microbial community and DOM structure were driven by several environmental stressors, including drought, seawater intrusion, and heavy precipitation events. Bacterial composition of different lifestyles (free-living and particle- associated) differed and only a part of the particle-associated bacteria correlated with microbial eukaryotes. Oscillations of cyanobacterial relative abundance along with chlorophyll a revealed a high primary production season characterized by increased levels of autochthonous DOM that promoted bacterial processes of organic matter degradation. From our results, we infer that in coastal freshwater lakes dependent on precipitation-evaporation balance, prolonged dry season coupled with heavy irrigation impact microbial communities at different trophic levels even if salinity increases only slightly and allochthonous DOM inputs decrease. These pressures, if applied more frequently or at higher concentrations, could have the potential to overturn the trophic state of the lake. This article is protected by copyright. All rights reserved.

The Krka River is located at Balkan Peninsula in the Dinaric region of Croatia and it is a typical groundwater-fed karst river characterized by an unusual phenomenon-tufa. Tufa designates porous CaCO3 deposits forming under specific physical and chemical conditions and hosting very diverse freshwater biota. Karst springs are considered one of the most diverse habitats for diatoms and are usually inhabited by specific diatom assemblages, which include the Krka spring zone. However, such habitats are particularly impacted by human activities, so previous studies have recently been conducted to raise awareness of the importance of karstic spring zones, especially because they are inhabited by specific and rare species. During the research at Krka River, samples were collected in the spring zone in May 2017, and the observations revealed the existence of a one Cymbopleura species characterized by slightly convex to almost linear valves, which are slightly protracted, broadly rounded and weakly dorsally bent at the apices. Based on detailed morphological analyses on light and scanning electron microscopy it appears that this population resembles two previously described taxa: C. rupicola var. minor Krammer and C. rupicola var. korana Krammer. Here we provide a more detailed description of the mentioned population under the new name Cymbopleura amicula stat. nov. et nom. nov. due to the significant differences from C. rupicola, together with a comparison with similar taxa in Cymbopleura.

Due to its geographical position, relief features, specific geological past and great heterogeneity of space in a relatively small area, Bosnia and Herzegovina has specific hydro-morphological features, with numerous spring habitats and unique aquatic biodiversity, mainly composed by stenothermic and relict taxa. Spring habitats, as well as the flora and fauna that inhabit them are considered particularly vulnerable to climate change and anthropogenic pressures, due to their isolation and fragmentation. According to forecasts, these ecosystems will be exposed to multiple pressures caused by changes in water regime, rising water temperatures and a continuous decline in water quality, which will lead to the disappearance of more sensitive crenobiont species and a reduction in biodiversity. Therefore, in this chapter we present the ecological characteristics and specifics of spring habitats in Bosnia and Herzegovina, with a preliminary list of species that inhabit them and special focus on endemic, sensitive and rare species, which are potentially threatened by predicted changes in habitat quality.

Phytoplankton is one of the key Biological Quality Elements within the Water Framework Directive, used to assess the ecological status of surface water bodies. Water samples for phytoplankton identification were collected from April to September at a total of eight sampling sites in all six Croatian natural karstic lakes with an area greater than 0.5 km2. The main objective was to show the comparability of environmental DNA metabarcoding (Illumina sequencing using the hypervariable V9 region of the eukaryotic SSU rRNA gene) with morphologically based assessment and its applicability in assessing the ecological status of lakes. The value of Hungarian lake phytoplankton index (HLPI) indicating the final ecological status was calculated for both datasets using biomass and composition metrics. Chlorophyll a concentration measured using Ultra-High-Performance Liquid Chromatography and Spectrophotometer giving two biomass metrics along with the functional group approach (FG) as the composition metric for the complete taxa/operational taxonomic units (OTUs) lists as well as for the taxa/OTUs that contributed more than 5% to the total biomass/number of amplicons gave four to four HLPI values per sample. HLPI values from both approaches were highly correlated (Pearson's r > 0.92) and classified into good or high ecological status, although different compositions and proportions of FGs were recorded, thus giving the important role to the equal or similar factors assignment to different FGs with similar ecological demands and favourable habitats. In 89% of the samples, HLPI values indicate an equal range of ecological status and most differences were found due to different methods of Chlorophyll a measurement. Different composition metrics between approaches showed significant differences (p < 0.05) only in lakes Prošće and Crniševo. This study showed the applicability of the V9 region of 18S rRNA in ecological status assessment for oligotrophic and mesotrophic lakes due to the comparable results between approaches, but further development and standardization of eDNA metabarcoding are needed for the implementation in routine monitoring programs.

The biodiversity of algal communities and environmental conditions were investigated in the springs of Mt. Konjuh. The assemblages of 20 springs emerging from different lithologies (limestones and ophiolites, respectively) comprised 234 algal taxa. Diatoms and cyanobacteria were the most species-rich groups. The most common alkaliphilic, circumneutral, and eutraphentic diatoms were represented by the genera Gomphonema, Nitzschia, Navicula, Cymbella, and Achnanthidium, and by the cyanobacterial genus Phormidium. Hierarchical clustering and SIMPROF analysis based on relative algal abundance clustered springs into six groups, separating them mainly according to spring type and lithology. Indicator species for groups and springs on different lithological substrata were singled out, revealing 33 taxa with preferences for ophiolites, and 20 taxa with preferences for carbonates. The values of the Shannon-Wiener diversity index were moderately high per spring location, and similar for the two groups of springs on different lithologies. A higher similarity in species composition was noted between springs on ophiolites and limestones than between springs on ophiolites and other types of siliceous substrata. The present study suggests that algal assemblages in springs emerging from ophiolites, even those made up by a preponderance of silicates, should be analyzed separately from those related to springs on other siliceous substrata. The results obtained showed that most of the springs studied are affected by anthropogenic impacts and morphological alterations leading to the dominance of highly competitive meso- and eutraphentic algal species, thus emphasizing the importance of further investigation and conservation of these habitats.

Ciliates are single celled eukaryotes recognized as key players in the microbial loop of aquatic ecosystems. The present study was carried out on the Krka River (Croatia), a karst freshwater ecosystem characterized by tufa barriers, biomineralization and highly diverse aquatic communities. The main aims of the study were to investigate ciliate community structure in the biofilm (i.e. periphyton) samples collected from light- and dark-exposed lithified tufa/stones. Furthermore, by establishing links between ciliate community patterns and environmental parameters, we aimed to assess the bioindicator potential of specific ciliate taxa for environmental monitoring of freshwater habitats. The periphyton sampling was performed at four representative sites of the river source, upstream, middle and downstream river sections. Ciliate community was investigated via traditional microscopy analyses and environmental DNA (eDNA) metabarcoding (Illumina sequencing of the hypervariable V9-region of the SSU rRNA gene). The molecular approach recorded a substantially higher number of ciliate taxa, most of which taxonomically belonging to genera typically occurring in tufa barriers. The results from microscopy analyses did not show any links between ciliate community structure and sampling location. However, eDNA approach indicated significant differences among the sampling locations regarding the ciliate community structure. Thereby, hydrological parameters and saprobiological classification of the sampling sites were the main structuring factors for ciliate community. The coupling of eDNA metabarcoding with the morphological approach provides a robust, in-depth analytical system in elucidating the bioindicator potential of ciliated protists.

Ciliates (protozoa) are a very large and diverse group of microeukaryotes that occupy a central position in the trophic web of freshwater ecosystems. Ciliates exhibit high ecological sensitivity and have tremendous bioindicator potential, but they are largely ignored in routine biomonitoring, mainly due to limitations in morphological diagnosis, which is both time-consuming and costly. In this study, we combined molecular (Illumina sequencing of the hypervariable V9 region of the SSU rRNA gene) and morphological approaches to get a detailed insight into the periphytic ciliate community structure and function within a karst, tufa-precipitating hydrosystem (National Park Krka, Croatia). Periphyton (biofilm) was sampled from light- and dark-exposed lithified tufa/stones at four representative locations within the Krka River, including upstream, midstream, and downstream river sections. We identified hydrological parameters and saprobiological classification of the sampling locations as the main structuring factors for ciliate communities. The molecular approach showed a clear separation of sampling locations in beta diversity analyses, as confirmed by PERMANOVA test. In contrast, alpha diversity was mostly affected by light exposition of the lithified tufa/stones, as confirmed by morphological approach. Environmental conditions consistently affected ciliate community composition at all locations, resulting in clear separation of periphyton samples. Our results indicate that ciliates should be considered as important bioindicators in monitoring tufa-forming rivers and streams, and the molecular approach (V9 region) proved to be applicable because as it efficiently captured the diversity of ciliates in periphyton. Validation of such an approach would lead to significant progress in extending monitoring to a broader range of indicator organisms than those now included in standard monitoring.

Karst ecosystems play a unique role as exceptional natural habitats in sustaining biodiversity. This study focuses on diatoms, a diverse group of microeukaryotes in the periphytic community of a karstic river. In a multi-microhabitat study along the Krka River (Croatia), our goal was to obtain a detailed overview of diatom diversity and community structure using morphological and molecular approaches, and to assess the applicability of eDNA metabarcoding as a reliable tool for biomonitoring assessment. The results revealed a relatively low agreement in the diatom community composition between the two approaches, but also provided complementary information, with no differences in beta diversity detected between microhabitats. The SIMPER analysis underlined the importance of the molecular approach in identifying diatom community composition, due to errors in distinguishing between deposited diatom cells that occurred in the morphological analysis. In contrast, the morphological approach indicated a clear diatom community separation along the river with a strong location effect. Despite certain differences, both approaches provided a feasible assessment of the ecological status according to the relationship to environmental pressures, classifying the Krka River as High (morphological approach) or Good (molecular approach) throughout the most of its course. Moreover, diatom diversity based on both approaches provides a reliable dataset applicable in routine monitoring assessment and offers a deeper understanding of the presented ecological status. The incompleteness of a reference database presents one major drawback of the molecular approach, which needs further updating in order to improve routine diatom metabarcoding.

Nitrogen is one of the essential elements limiting growth in aquatic environments. Being primarily of anthropogenic origin, it exerts negative impacts on freshwater ecosystems. The present study was carried out at the nitrate-vulnerable zone within the alluvial aquifer of the large lowland Drava River. The main aim was to investigate the ecosystem&rsquo, s functionality by characterizing the bacterial and phytoplankton diversity of a small inactive gravel pit by using interdisciplinary approaches. The phytoplankton community was investigated via traditional microscopy analyses and environmental DNA (eDNA) metabarcoding, while the bacterial community was investigated by a molecular approach (eDNA). Variations in the algal and bacterial community structure indicated a strong correlation with nitrogen compounds. Summer samples were characterized by a high abundance of bloom-forming Cyanobacteria. Following the cyanobacterial breakdown in the colder winter period, Bacillariophyceae and Actinobacteriota became dominant groups. Changes in microbial composition indicated a strong correlation between N forms and algal and bacterial communities. According to the nitrogen dynamics in the alluvial aquifer, we emphasize the importance of small water bodies as potential buffer zones to anthropogenic nitrogen pressures and sentinels of the disturbances displayed as algal blooms within larger freshwater systems.

The authors acknowledge COST Action ES 1105 "CYANOCOST Cyanobacterial blooms and toxins in water resources: Occurrence impacts and management" and COST Action Global Change Biology ES 1201 NETLAKE -Networking Lake Observatories in Europe" for contributing to this study through networking and knowledge sharing with European experts in the field. We acknowledge the members of the Global Lake Ecological Observatory Network (GLEON) for their collaborative spirit and enthusiasm that inspired the grassroots effort of the EMLS. E.M. was supported by a grant from the Swiss State Secretariat for Education, Research and Innovation to Bas Ibelings and by supplementary funding from University of Geneva. We thank Wendy Beekman for the nutrient analysis. We thank Pieter Slot for assisting with the pigment analysis. We thank Dr. Ian Jones for valuable feedback on an earlier version of the manuscript. We thank the Leibniz Institute of Freshwater Ecology and the Aquatic Microbial Ecology Group for logistic and technical support of J. Fonvielle and H.-P. Grossart, and the Leibniz Association for financial support. H.P. was supported by the US National Science Foundation (1840715, 1831096). A.C.'s work was funded by the Spanish Agencia Estatal de Investigacion and EU funds through the project CLIMAWET (CGL2015-69557-R). The collection of data for Lough Erne and Lough Neagh were funded by the Department of Agriculture, Environment and Rural Affairs, Northern Ireland. We are grateful to Kristiina Vuorio from the Freshwater Centre of the Finnish Environment institute for her help in organizing, collecting and analysing samples by the University of Jyvaskyla and to Gerald Dorflinger from the Water Development Department of Cyprus for his assistance with the sampling in Cyprus and for granting the CUT team permission to use WDD's equipment. Finally, we would like to thank the numerous other assistants that helped realizing each local survey. Open access funding provided by Universite de Geneve., To determine the drivers of phytoplankton biomass, we collected standardized morphometric, physical, and biological data in 230 lakes across the Mediterranean, Continental, and Boreal climatic zones of the European continent. Multilinear regression models tested on this snapshot of mostly eutrophic lakes (median total phosphorus [TP] = 0.06 and total nitrogen [TN] = 0.7 mg L-1), and its subsets (2 depth types and 3 climatic zones), show that light climate and stratification strength were the most significant explanatory variables for chlorophyll a (Chl a) variance. TN was a significant predictor for phytoplankton biomass for shallow and continental lakes, while TP never appeared as an explanatory variable, suggesting that under high TP, light, which partially controls stratification strength, becomes limiting for phytoplankton development. Mediterranean lakes were the warmest yet most weakly stratified and had significantly less Chl a than Boreal lakes, where the temperature anomaly from the long-term average, during a summer heatwave was the highest (+4 degrees C) and showed a significant, exponential relationship with stratification strength. This European survey represents a summer snapshot of phytoplankton biomass and its drivers, and lends support that light and stratification metrics, which are both affected by climate change, are better predictors for phytoplankton biomass in nutrient-rich lakes than nutrient concentrations and surface temperature., European Cooperation in Science and Technology (COST) ES 1105 ES 1201, Swiss State Secretariat for Education, Research and Innovation, University of Geneva, Leibniz Association, National Science Foundation (NSF) 1840715 1831096, Spanish Agencia Estatal de Investigacion, European Commission CGL2015-69557-R, Department of Agriculture, Environment and Rural Affairs, Northern Ireland

The process of travertine formation and carbonate deposition in the rivers is unique, delicate, and depends on the activity of algae and mosses. Although diatoms have been used extensively in hydrobiological studies, the comparative analysis data on diatom communities of the travertine barriers in karstic rivers are still scarce. The study aimed to detect the diatom composition on travertine barriers in the Una River, the large karstic river in Bosnia and Herzegovina. An integrated classical morphological identification approach with metabarcoding was applied on eight samples across the river length profile. Morphological analyses were performed using both light and scanning electron microscopes. Subsequent DNA metabarcoding of the chloroplastic gene 312bp rbcL was done. The DADA2 pipeline was used for the bioinformatic treatment of the demultiplexed MiSeq reads to infer Amplicon Sequence Variants (ASVs). ASVs were taxonomically assigned using the Diat.barcode v7 reference database. A total of 126 species were identified using the morphological approach, while 133 ASVs were taxonomically assigned to 58 unique taxa with the molecular approach. Diatom community structures in terms of molecular and morphological approaches were congruent with 49 shared species. Species from genera Gomphonema, Navicula and Encyonema were less assigned in molecular analysis. The most abundant taxa in the Una River are alkaliphilous, belonging to the genera Gomphonema, Nitzshia and Navicula. Although specific for their extremely good chemical status, the travertine barriers of the Una River are largely inhabited with meso-eutraphentic taxa.

EnglishThe spatial and temporal patterns of phytoplankton in relation to physical and chemical properties were studied in a drinking water reservoir - the Lazberc Reservoir - located in north-east Hungary. The main objectives were to determine the role of the hydrologic regime in structuring algal growth and the effects of physical and chemical variables on the coexistence of a dominant species: cyanobacteria, Microcystis aeruginosa (Kutzing) Kutzing and an eukaryotic dinoflagellates, Ceratium hirundinella (O.F. Muller) Dujardin. The results suggest that nutrients (TP, NH4–N and NO3–N), turbidity, and the hydrologic regime play an important role in regulating the occurrence of investigated species. The rainy summer period resulting in a mixing of the water column, and the low level of nitrogen coupled with the internal release of phosphorus from the lake sediment under brief periods of anoxia promote cyanobacterial bloom. During this period, the water column was characterised by high turbidity, which created favourable conditions for developing a huge C. hirundinella biomass and the simultaneous occurrence of M. aeruginosa. This study also demonstrated - that contrary to what was previously known, deep standing waters in the temperate zone are not only dimictic (mixing twice a year during the spring and the fall), but they can also be mixed during the summer period due to continued rainfall, which equalizes the temperature in the whole water column. This can promote the occurrence of cyanobacterial and eukaryote planktonic organisms portuguesOs padroes espaciais e temporais do fitoplâncton e sua correlacao com as especificidades variaveis fisicas e quimicas, foram examinados no nordeste da Hungria, localizados em um reservatorio tipico de agua potavel no reservatorio de Lazberc. Nosso principal objetivo foi para esclarecer o papel da hidrologia, e os diferentes especificidades fisicas e quimicas na coexistencia de especies dominantes: cianobacteria, Microcystis aeruginosa (Kutzing) Kutzing e Dinoflagellata eucariotica, Ceratium hirudinella (OF Muller) Dujardin. De acordo com os resultados, os nutrientes (TP, NH4-N, NO3-N), turbidez e os condicoes hidrologicas desempenham um papel importante na regulacao da ocorrencia das especies examinadas. O periodo de verao chuvoso que resulta um equilibrio do nivel da agua, o baixo nivel de nitrogenio, juntamente com a liberacao interna de fosforo do sedimento, durante um curto periodo anoxico promove a floracao de cianobacterias. Durante este periodo, a coluna de agua foi caracterizada por alta turbidez, criando condicoes favoraveis para o surgimento de uma enorme expansao da biomassa C. hirundinella e a aparicao simultânea de M. aeruginosa. Este estudo mostrou tambem que as aguas profundas nas areas da zona moderada, nao tem apenas dimictic especialidade (misturando duas vezes por ano, na primavera e no outono), mas tambem durante a estacao do verao pode ser misturada por chuvas continuas que compensam a temperatura em toda a coluna da agua. Isso ajuda a presenca de cianobacterias e organismos eucariotas planctonicos

The type material of Achnanthes hauckiana Grunow has been investigated using light and scanning electron microscopy. The species, transferred to the genus Planothidium, was frequently reported from brackish and even marine environments. The original Grunow material was collected from a karstic spring near Trieste. Analysis of the material and of samples collected from the Croatian Krka River, resulted in a better understanding of the morphology of Planothidium hauckianum and a better characterization of the diatom flora of which it is a dominant component. The paper compares the species with similar Planothidium species from the P. delicatulum complex and revises published images of P. hauckianum in literature, making it clear that all reported and illustrated brackish populations of P. hauckianum belong to different, often unknown species. The presence of Planothidium hauckianum is limited to karstic spring areas, most likely showing a very restricted biogeographical distribution.

The genus Gomphosphenia Lange-Bertalot was separated from Gomphonema Ehrenberg and when established, it was considered a species limited genus with 17 taxa including species and varieties. None of the species from the genus Gomphosphenia have yet been recorded in Croatia and for the first time was observed in lake Crveno jezero (Red lake). The lake is one of the largest karstic sinkhole lake in Europe, situated in the town Imotski in Croatia, near the border with Bosnia and Hercegovina. Diatom flora of this lake was unknown due to difficulties in collecting samples. Sampling of phytobenthos was conducted as part of systematical natural sciences research organized during the International speleodiving expedition “Crveno jezero 2017”. Observations revealed the existence of a new species, Gomphosphenia plenkoviciae Gligora Udovič & Žutinić sp. nov. Gomphosphenia plenkoviciae possesses typical morphological and ultrastructural characteristics of genus Gomphosphenia. It is characterized by having narrowly clavate to linear valves, which are slightly heteropolar in larger specimens to clavate in smaller specimens, thus differing from the closest taxon by the valve outline and shape of the central area.

Insight into how environmental change determines the production and distribution of cyanobacterial toxins is necessary for risk assessment. Management guidelines currently focus on hepatotoxins (microcystins). Increasing attention is given to other classes, such as neurotoxins (e.g., anatoxin-a) and cytotoxins (e.g., cylindrospermopsin) due to their potency. Most studies examine the relationship between individual toxin variants and environmental factors, such as nutrients, temperature and light. In summer 2015, we collected samples across Europe to investigate the effect of nutrient and temperature gradients on the variability of toxin production at a continental scale. Direct and indirect effects of temperature were the main drivers of the spatial distribution in the toxins produced by the cyanobacterial community, the toxin concentrations and toxin quota. Generalized linear models showed that a Toxin Diversity Index (TDI) increased with latitude, while it decreased with water stability. Increases in TDI were explained through a significant increase in toxin variants such as MC-YR, anatoxin and cylindrospermopsin, accompanied by a decreasing presence of MC-LR. While global warming continues, the direct and indirect effects of increased lake temperatures will drive changes in the distribution of cyanobacterial toxins in Europe, potentially promoting selection of a few highly toxic species or strains. © 2018 by the authors. Licensee MDPI, Basel, Switzerland., The authors acknowledge COST Action ES 1105 “CYANOCOST—Cyanobacterial blooms and toxins in water resources: Occurrence impacts and management” and COST Action ES 1201 “NETLAKE—Networking Lake Observatories in Europe” for contributing to this study through networking and knowledge sharing with European experts in the field. Evanthia Mantzouki was supported by a grant from the Swiss State Secretariat for Education, Research and Innovation (SERI) to Bas Ibelings and by supplementary funding from the University of Geneva. We thank Clare Ahnlund, Ena Suarez and Irene Gallego for helping out with the Swiss survey. We thank Wendy Beekman and Els J. Faassen for the nutrient and toxin analysis.

Under ongoing climate change and increasing anthropogenic activity, which continuously challenge ecosystem resilience, an in-depth understanding of ecological processes is urgently needed. Lakes, as providers of numerous ecosystem services, face multiple stressors that threaten their functioning. Harmful cyanobacterial blooms are a persistent problem resulting from nutrient pollution and climate-change induced stressors, like poor transparency, increased water temperature and enhanced stratification. Consistency in data collection and analysis methods is necessary to achieve fully comparable datasets and for statistical validity, avoiding issues linked to disparate data sources. The European Multi Lake Survey (EMLS) in summer 2015 was an initiative among scientists from 27 countries to collect and analyse lake physical, chemical and biological variables in a fully standardized manner. This database includes in-situ lake variables along with nutrient, pigment and cyanotoxin data of 369 lakes in Europe, which were centrally analysed in dedicated laboratories. Publishing the EMLS methods and dataset might inspire similar initiatives to study across large geographic areas that will contribute to better understanding lake responses in a changing environment., The authors acknowledge COST Action ES 1105 “CYANOCOST – Cyanobacterial blooms and toxins in water resources: Occurrence impacts and management” and COST Action ES 1201 “NETLAKE – Networking Lake Observatories in Europe” for contributing to this study through networking and knowledge sharing with European experts in the field. Evanthia Mantzouki was supported by a grant from the Swiss State Secretariat for Education, Research and Innovation (SERI) to Bas Ibelings and by supplementary funding from University of Geneva. We thank Wendy Beekman for the nutrient analysis and the University of Wageningen for covering the costs of this analysis from the personal funding of dr. Miquel Lürling. We thank Pieter Slot for assisting with the pigment analysis and the University of Amsterdam for covering the costs of the analysis through funding from the group of Prof. Jef Huisman and dr. Petra Visser (IBED). We would like to thank the Environmental Data Initiative for covering the cost of archiving the EMLS dataset. We would like to thank the Leibniz Institute of Freshwater Ecology and Inland Fisheries, Dept. of Experimental Limnology and the Aquatic Microbial Ecology Group for logistic and technical support of J. Fonvielle and H.-P. Grossart; and the Leibniz Association for financial support. The collection of data for Lough Erne and Lough Neagh were funded by the Department of Agriculture, Environment and Rural Affairs, Northern Ireland.

The increased amount of organic matter (OM) accompanied with the development of hypoxia/anoxia and appearance of the sulphur species are indicators of eutrophication in natural waters. To predict biogeochemical and ecological changes, specific OM components in the Lake Brljan (the karst Krka River, Croatia; station B and station B2 downstream) were studied during 2011 and in January 2012 using voltam¬metry. Surface‒active substances (eq. 0.018 to 0.150 mg dm–3 Triton‒X‒100) and Cu complexing capacity (54‒680 nmol dm–3) showed the highest values in surface water layer in May 2011 (B) and in November 2011 (B2), associated with the highest dissolved (0.505‒1.530 mg C dm–3) and particulate (0.051‒0.725 mg C dm–3) organic carbon and phytoplankton biomass. Autochthonously produced OM did not cause hypoxia/anoxia during the investigated period. Brljan's lake water could be characterised as oligotrophic with occasionally elevated OM concentration depending on season and Krka River water regime. This work is licensed under a Creative Commons Attribution 4.0 International License.

The diatom genus Entomoneis is known from the benthos and plankton of marine, brackish, and freshwaters. Entomoneis includes diatoms with a bilobate keel elevated above the valve surface, a sigmoid canal raphe, and numerous girdle bands. Owing mostly to the scarcity of molecular data for a diverse set of species, the phylogeny of Entomoneis has not been investigated in depth. The few previous studies that included Entomoneis were focused on broader questions and the available data were from a small number of either unidentified Entomoneis or well-known species (e.g., E. paludosa). Since the first description of new species combining both molecular and morphological characters (E. tenera), we have continued to cultivate and investigate Entomoneis in the plankton of the Adriatic Sea. Combined multigene phylogeny (SSU rDNA sequences, rbcL, and psbC genes) and morphological observations (LM, SEM and TEM) revealed six new Entomoneis species supported by phylogenetic and morphological data: E. pusilla, E. gracilis, E. vilicicii, E. infula, E. adriatica, and E. umbratica. The most important morphological features for species delineation were cell shape, the degree and mode of torsion, valve apices, the appearance and structure of the transition between keel and valve body, the ultrastructure and the shape of the girdle bands, and the arrangement and density of perforations along the valve and valvocopulae. Our results highlight the underappreciated diversity of Entomoneis and call for a more in-depth morphological and molecular investigation of this genus especially in planktonic habitats.

Benthic diatoms are widely used in Europe and worldwide to access ecological status of running waters. One of key goals of Water Framework Directive is to classify rivers and streams using biological quality elements and type specific reference conditions. According to system B which incorporates additional abiotic descriptors, there are 24 water types in Croatia. For biological analyses 92 rivers and streams with 140 sampling points were chosen and sampled for benthic diatoms and water chemistry simultaneously. Self organizing map (SOM) analysis was used to define biotypes from species composition and abundance of benthic diatoms. Grouping of samples in SOM resulted in 10 distinctive groups. Based on their geographical position and site characteristics, groups represent sites with similar properties (as waterbed, catchment size, altitude, size of stream) belonging to different ecoregions in Croatia. Analysis of variance revealed statistically significant differences (p

The genus Entomoneis Ehrenberg includes diatoms with structurally complex frustules having a bilobate keel elevated above the valve surface and numerous girdle bands. We describe here a new member of the genus, Entomoneis tenera sp. nov. , a minute species found in the plankton of the south-eastern Adriatic Sea. The description and proposed taxonomic affiliation are based on both morphological observations and molecular analyses obtained from three cultivated strains. The cultures were established from plankton net samples taken during BIOTA (Bio-tracing Adriatic Water Masses) cruise conducted in February–March 2015. In addition to characteristic Entomoneis features such as panduriform cells, often twisted around the apical axis and a raphe with simple endings positioned on the sigmoid keel, morphological characteristics of E. tenera are: (1) very small cells, 16–21 μm long and 5–20 μm wide (2) very lightly silicified, delicate frustules without valve striation discernible in light microscopy, (3) broad lanceolate valves with scalpeliform apices (4) a straight to slightly arcuate junction line. Phylogenetic analyses using SSU, rbc L and psb C supported the position of E . tenera within the Entomoneis genus with a clear separation from the other described species.

An investigation into bryophyte communities in karst Jankovac Stream (Papuk Nature Park, Croatia) was carried out once a month from July 2008 to June 2009. Samples were taken from two lotic microhabitats: (i) Jankovac Spring (JS), a hypocrenal habitat with dense bryophyte clusters (90% bottom cover) and (ii) Jankovac Waterfall (JW), with scattered bryophyte clusters (50% bottom cover). At the same time, seston samples were collected during the spring as the source site and after the JW, as the outflow site. The goals of this study were to understand the (i) algal, protozoan and metazoan bryophyte community assemblages in these two lotic microhabitats, (ii) influence of environmental factors on the structuring of the bryophyte community and (iii) structure of seston along the longitudinal profile. A total of 172 taxa were determined: 68 algae, 55 protozoa, 24 meiofauna and 25 macroinvertebrates. Statistically significant differences between two microhabitats differing in percentage of bryophyte cover were established testing 13 environmental parameters. In dense bryophyte clusters, community structure was determined by flow velocity and pH, and macroinvertebrates achieved higher diversity and abundance. On the contrary, in scattered bryophyte coverage algae, protozoa and meiofauna reached higher abundance and diversity governed by the amount of suspended organic matter and epiphyton. In contrast to previous studies, the inverse ratio of community diversity and abundanceversuspercentage of bryophyte cover was established. We assume this to be the result of an enrichment of the scattered bryophyte clusters by upstream seston. Additionally, the effects of anthropogenic hydromorphological disturbance are reflected in macroinvertebrate diversity and abundance reduction.

Crveno jezero (Red Lake) is one of the largest karstic sinkhole lakes in the Europe. Diatom flora of this lake was unknown due to difficulties in collecting samples. Recently, samples were collected by speleodivers in the frame of systematical natural sciences research, during the cave diving expedition “Crveno jezero 2017” and observations revealed the existence of a new species, Gomphosphenia plenkoviciae Gligora Udovič & Žutinić sp. nov. The species is described herein as new to science based on light and scanning electron microscopy. Gomphosphenia plenkoviciae is characterized by having narrowly clavate to linear valves, which are slightly heteropolar in larger specimens to clavate in smaller specimens, thus differing from the closest taxon by the valve outline and shape of the central area. Smaller valves of G. plenkoviciae differ from other small-sized Gomphosphenia species by valve outline, striae morphology and the shape of axial and central areas.

In four deep karstic lakes (Lake Vransko, Lake Visovacko, Kozjak and Prosce) several functionally well-adapted phytoplankton groups interchange in dominance during the seasonal cycle from April to September and likely tolerate the constraining conditions of nutrients deficiency more successfully as an aggregated group than as taxonomic units. This paper considers the functional approach by Reynolds et al. (2002), a method capturing much of the species ecology and which is successfully used for describing phytoplankton successions. Deep karstic lakes are attributed with the early spring (April) formation of stratification, low phytoplankton biomass, and with maximum biomass accumulating in July. The co-occurrence of functional groups in those lakes is evident but typically distinct according to different biomass proportions. The functional groups A, B, C, D, P, E and LO, represented by different Cyclotella, Ulnaria/Fragilaria, Dinobryon, Ceratium/Peridinium species, co-occurred in 50% and up to 90% of phytoplankton biomass in all investigated lakes. By observing the co- occurrence across the Croatian karstic region we: (i) demonstrate the regional distribution of phytoplankton assemblages in relation to low nutrient environment, and (ii) build a simple uniform pattern which successfully describes a variety of phytoplankton in deep karstic lakes and provides information of reliable indicators for those systems.

Influence of hydrological characteristics and nutrient concentrations on phytoplankton was investigated in four large rivers (Mura, Drava, Danube and Sava) in the Pannonian ecoregion in Croatia to understand how phytoplankton of rivers can be explained by the “different functional group approach”. To gain a clearer understanding of the factors that affect river phytoplankton, the present study examined phytoplankton biomass and composition in relationship with physical and chemical parameters assessed in detail by preparing self-organising maps using functional groups and morpho-functional groups. Total nitrogen along with water residence time showed to be the best predictor to determine phytoplankton biomass and chlorophyll a. Phytoplankton diversity increased with higher water discharge, but it had the consequence of diluting algae and decreasing biomass. Bacillariophyceae and Chlorophyceae species dominated the phytoplankton assemblages in all rivers. Diatoms predominated in rivers with shorter residence time. Dominant diatom codons of functional groups were C, D and TB while morpho- functional groups were represented by only diatom group VI. As residence time increased, the proportion of chlorococcalean green algae, represented by functional group codon T and morpho-functional group IV grew in summer. Since potamoplankton is dominated by diatoms, functional groups with its fine partition of diatom codons proved to be excellent descriptor of the potamoplankton. Application of morpho-functional groups originally developed from the lake data, showed to be limiting because of the predominating presence of only one diatom group.

Species composition of the net-phytoplankton assemblage and seasonality of the dominant species were investigated in the travertine monomictic stratified riverine Lake Visovac in relation to the environmental variables. The one year investigation was based on monthly sampling from April 1995 to March 1996 at the two deepest vertical profiles, with a maximum depth of 25 m. Diatoms, especially Asterionella formosa Hass. dominated net-phytoplankton assemblages except in late summer and autumn when dinoflagellate species Ceratium hirundinella (O. F. Müller) Bergh became the dominate form. The effects of the short term dynamics of environmental factors on the algal growth in the barrage lenitic area of the calcareous river stressed several variables as important factors controlling annual net-phytoplankton distribution. Temperature was the variable that most affected species abundance, changes in dominance and species composition (Primer 5, BIO-ENV; http://www.primer-e.com/). CCA analysis performed on abundance dataset and environmental variables confirm the importance of temperature but also highlighted total phosphorus, conductivity and silica., Tijekom jednogodišnjeg razdoblja od travnja 1995. do ožujka 1996. istraživana je, u jednomjesečnim intervalima, vertikalna i sezonska distribucija mrežnog fitoplanktona u lenitičkom području Visovačkog jezera. Naglasak tijekom istraživanja bio je na sastav vrsta te povezanost dominantnih vrsta i okolišnih čimbenika. Najzastupljenija vrsta tijekom kasnog proljeća je dijatomeja Asterionella formosa Hass, dok ljeti i u jesen dominaciju preuzima dinoflagelat Ceratium hirundinella (O. F. Müller) Bergh. Od mjerenih ekoloških parametara, temperatura vode najznačajnije je utjecala na promjene u sastavu vrsta, njihovu zastupljenost i dominaciju (Primer 5, BIO-ENV). CCA analiza dobivena na osnovi zastupljenosti fitoplanktonskih stanica i praćenih ekoloških čimbenika potvrdila je znatan utjecaj temperature vode, kao i ukupnog fosfora, provodljivosti i silicija.