Rarely mentioned species in Hungary: Can we step into the same lake?

International and national protection strategies and directives focus mainly on macroscopic organism and attempt to maintain their endangered habitats. However, microscopic communities are also threatened by decreasing biodiversity and many species including freshwater algae can disappear without even knowing they were present in the habitat. Defining rarity of microscopic taxa is not easy. The species’ rarity is based on detailed knowledge of distribution and abundance of species. But only limited information is available about rare algal species especially in a given ecoregion. Reducing the data gaps, here, we present altogether 20 phytoplankton taxa rare in Hungary: three species of Chlorophyceae, eight species of Trebouxiophyceae, two taxa of Euglenophyceae, one-one species of Cyanobacteria, Bacillariophyceae and Mediophyceae and three species of Xanthophyceae. One of them, the Cylindrotheca gracilis is on the Hungarian Red List. Physical and ecological characteristics of standing waters where these species were found as well as their former occurrence all over the world are also reviewed.


Introduction
The quest to understand how the world can work always presents new challenges for researchers. In the twentieth century, tasks had to be solved were under significant social pressure. From the middle part of the twentieth, eutrophication has been become the one of the most important tackled tasks both in local and global scales (Reynolds 2006). The surplus nutrient supply, mainly phosphorus and nitrogen, can cause undesirable changes in water quality (Istvánovics 2009). Eutrophication can modify the dynamics of phytoplankton with increasing algal biomass and blooming cyanobacteria resulting alteration in the species composition (Reynolds 2006). Recently, the drastic changes of climate, primarily hydrological drought or flash floods, cause deterioration of the quantity and quality of surface waters (Murdoch et al. 2000, Mosley 2015. In freshwaters, the structure of communities living here is significantly changed by the decreasing water level coupled with the increasing nutrient concentration, by the increasing water temperature, by the sediment resuspension and also by the stratification. Finally, these changes result biodiversity loss and habitat degradation (Mosley 2015).
Since eukaryotic algae and cyanobacteria play pivotal role in functioning of freshwater ecosystems, any changes in their structure can finally alter the composition of the whole ecosystem (Reynolds 2006). Thus, planktic algae and cyanobacteria are used to describe the status of habitats and to predict different disturbation (Juráň and Kaštovský 2019). These acts requires detailed knowledge about local and worldwide distribution of taxa, especially in case of rare ones (Juráň and Kaštovský 2019). Defining rarity of microscopic taxa including algae, however, is not easy. The species' rarity is based on the distribution and the abundance of species, more precisely, the lack of actual distribution data (Molina 2013). Geographic range, abundance and habitat specialization were used to classify species into rarity categories and identify as rare or common (Molina 2013). But again, it is important to emphasize that the current distribution of taxa should be defined both at local and global scales to conclude whether taxa can be considered as rare or not.
Recently, the identification of planktic algae at species level can be carried out either by traditional, microscope method or by using modern, genetic tools, but both of them have still shortcomings (Bornet et al. 2004). The genetic-based methods are not effective for the whole planktic microflora yet, mainly because the databases are strongly incomplete (Bornet et al. 2004). On the other side, the microscopic-based methodology have to face difficulties such as (i) instrument side, light and/or inverted microscope with appropriate magnification is essentially needed; (ii) human side, a specialist with adequate knowledge who can determinate even more than 200 species from one lake during a season; (iii) state-of-art literature; (iv) time side, it takes many time to prepare one sample even in monitoring process; and (v) sample side, sufficient samples are also needed (Prof. F. Hindák pers. comm.). All of these criteria should be met in order to have reliable knowledge about actual distribution of a species.
In this paper, a survey of data of Hungarian lakes and ponds is presented focusing on the ecological and taxonomical knowledge of rare algae.

Study area
During the Hungarian surveillance monitoring program, almost 60 standing waters with catchment area greater than 50 ha were investigated in 2019 from April to September. In this article those standing waters are presented which contains rarely mentioned taxa according to the Hungarian national identification manuals (see later) ( Fig. 1; Table 1 and  Supplementary Table 1). The chosen lakes belong to small lakes, reservoirs, oxbow lakes and gravel pit lakes from very shallow to deep (30 water bodies; Table 1).
Trophic status of the studied standing waters was classified according to OECD (1982).

Phytoplankton
Phytoplankton samples were investigated with LEICA DMIL inverted microscope in 400-fold magnification. Species were documented by Canon EOS digital camera and by drawings. Most recent accepted name of algae was considered by AlgaeBase (Guiry and Guiry 2020). Curiosity of the microflora was based on previous occurrences of the taxa published in Hungarian identification manuals (Németh 1997, Schmidt and Fehér 1998, Grigorszky et al. 1999) and on the Hungarian Red List of Algae (Németh 2005).
The previous (from literature data) and the new occurrences (our observations) of taxa in Hungary were used to classify taxa into rarity scale categories The scale was similar to Juráň's (2017) categories, who has described three categories for photosynthetic euglenoids of Check Republic. These categories are based on the number of local distributional data: (1) common (more than 15 localities), (2) rare (5 to 15 localities) and (3) very rare (maximally 4 localities).

Results
The physical and chemical parameters of the studied lakes and ponds are presented in Electronic Supplementary Material 1. While most of them can be categorized as hypertrophic or eu,hypertrophic, Nyékládháza pit lake was the only standing water which is oligo,-mezotrophic. Additionally, Hasznosi reservoir and Kiskunlacháza-2 lake belong to eu,mezotrophic.
Description: Frustules very narrow, elongate with drawnout ends and longitudinal spiraling of the cell.
Dimensions: 100 × 6 μm. Similarity: The shape of the cell (spindle-like form, torsion of 2 and half twists and sub-capitate ends) is very unique, but the frustules can be easily collapsed.
New occurrence in Hungary: Kiskunlacháza-2 lake. Ecological data: We found this species in a gravel pit lake with a few individuals. This species is listed into the Vulnerable (VU) category based on the Hungarian Red List of Algae (Németh 2005). Distribution is worldwide both in marine coasts and in freshwaters with high conductivity (Guiry and Guiry 2020).
Dimensions: protoplasm: 3-5 × 4-6 μm. Similarity: D. indutum is similar to Desmatractum delicatissimum, of which cell wall is smooth, cell halves goes gradually from the equatorial level to the pointed ones. While D. indutum has longitudinal superficial ribs and cell suddenly pulled out into a long process.
Ecological data: In this study, it has occurred sporadically mainly in oxbow lakes with a few individuals. It has worldwide distribution. For more details, see in Supplementary  Table 2.
Cell dimensions: 3-4 μm: Similarity: It is different from the other Tetrastrum by the number and length of spines. Only the T. peterfii Hortobágyi species has 1-3 spines, but their spines are equal long and stand irregular.
New occurrence in Hungary: Boroszlókerti oxbow. Ecological data: We found this species in a hypertrophic oxbow lake with a few individuals. This species is mentioned sporadically in the literature. For more details, see in Supplementary Table 2 Komárek, M.Hübel, H.Hübel & Smarda, which is shorter (4.5-7.2 μm) then N. spumigena. All of them can be straight or curved.
New occurrence in Hungary: Soponyai Sós-tó. Ecological data: Here, the species was found in a soda lake with a few individuals. However, it is cosmopolitan and previously it was occurred in different waters with high content of minerals (Komárek 2013).
Ecological data: It has occurred with a few individuals, mostly in summer plankton of oxbow lakes and reservoirs in our samples. Generally, it was rarely mentioned from larger eutrophic lakes. For more details, see in Supplementary  Table 2.
Dimensions: 28 × 25.2 μm. Similarity: The Trachelomonas bacillifera var. minima is similar to Trachelomonas bacillifera f. sparsispina Deflandre, which has scattered and irregularly distributed spines. In addition, T. bacillifera var. minima is also similar to Trachelomonas bacillifera var. collifera Huber-Pestalozzi, which is differed from the other varieties by its apical pole with a short collar.
New occurrence in Hungary: Boroszlókerti oxbow. Ecological and worldwide distribution data: We found this species in a hypertrophic oxbow lake with few individuals. Trachelomonas bacillifera var. minima is cosmopolitan and occurs in different kind of water bodies, like lakes, ponds, ditches and peat-bogs Hindák 2004, 2005). For more details, see in Supplementary Table 2.
New occurrence in Hungary: Nagy-Morotva. Ecological data: It was found in a hypertrophic oxbow lake. In the international literature, it was mentioned as a cosmopolitan species (Wołowski and Hindák 2005). For more details, see in Supplementary Table 2.
Mediophyceae. Chaetoceros muelleri Lemmermann (Fig. 2i). Reference: Krammer and Lange-Bertalot (1991): p. 391, T 80: Figures 1 and 2. Description: It is a colonial species, but in our samples only single cell form was found. From each pole of valve 1-1 twisted setae emerged. Valves are cylindrical with elliptic base and are lightly silicified.
Occurrence in Hungary: Szelidi Lake (last mention from this lake was 45 years ago).
Ecological data: Here, we found this species in a high conductivity lake. It is cosmopolitan (Guiry and Guiry 2020). For more details, see in Supplementary Table 2.
Cell dimensions: 10 × 4 μm. Similarity: The colony of C. ellipoideum is reminded to Komarekia appendiculata (Chodat) Fott, because 4-celled coenobium with mother cell wall fragments is characteristic of both species. But the cells of K. appendiculata are eggshape and the cells are connected inside.
Previous occurrences in Hungary based on Schmidt and Fehér (1999): Balaton.
New occurrence in Hungary: L-I lake, Békéscsabai Téglagyári gravel pit lake.
Ecological data: We found this species in hypertrophic and eu-hypertrophic lakes. Previously, it was found in different lakes, pond and rivers mostly in Europe. For more details, see in Supplementary Table 2.
New occurrence in Hungary: Tiszalúci oxbow. Ecological data: In our study, the species occurred in hypertrophic oxbow lake with a few individuals. In the world, this species is cosmopolitan. For more details, see in Supplementary Table 2.
Description: Elongated cells with long pointed apices are in a two-celled coenobium. When 2 or more pairs of cells stay together, they are wedged into each other.
New occurrence in Hungary: Belső-Béda. Ecological data: Here, it has occurred in a hypertrophic oxbow lake. It was mentioned, however, from the whole part of the world. For more details, see in Supplementary Table 2.
Reference: Komárek and Fott (1983): p. 497, Fig. 146:2. Description: The cells were ellipsoid to oval, solitary and have chloroplast with pyrenoid. The formation of brown granules is occurred only in poles but granules are never forming crowns.
Dimensions: 6-8 × 4-4,5 μm. Similarity: It is similar to Granulocystopsis elegans (Fott) Hindák, but in case G. elegans granules are visible both in the polar and in the equatorial part of the cell.
Ecological data: It has sporadically occurred in hypertrophic oxbow lakes in our samples. It was found in different habitats, e.g. plankic and benthic of swamps, river, lakes in worldwide. For more details, see in Supplementary Table 2.
Dimensions: 14 × 8.4 μm. Similarity: It is similar to Granulocystopsis coronata var. coronata of which granules are never forming crowns.
New occurrence in Hungary: Nagybaracskai-Holt-Duna. Ecological data: In this study, it was sporadically found in a hypertrophic oxbow lake. It is a rarely mentioned species in the international literature. For more details, see in Supplementary Table 2.
Dimensions: 11-20 × 3-5 μm. Similarity: The cell shape of the J. javorkae is similar to Nephrochlamys and Kirchneriella species, but they have smooth cell wall.
Ecological data: It has sporadically occurred in eutrophic or hypertrophic lakes or oxbow lakes in our samples. Rarely mentioned species in the international literature. For more details, see in Supplementary Table 2.
Similarity: It is similar to Lagerheimia wratislawiensis Schröder, but it has only 2 equatorial spines beside 1-1 polar spines.
Ecological data: It has occurred in hypertrophic oxbow lakes with a few individuals in our samples. It was rarely mentioned in the international literature and found mainly in reservoirs. For more details, see in Supplementary Table 2.
Similarity: Paradoxia is similar to Ankyra. But there are several differences e.g. Paradoxia is characterized by coenobium, blunt cell pole, and presence of bristles, while Ankyra is always described by unicellular, acute cell pole and absent of bristles.
Ecological data: We found only a few individuals of this species in a meso-eutrophic reservoir and a hypertrophic oxbow lake. However, it was found worldwide. For more details, see in Supplementary Table 2. Xanthophyceae. Bumilleriopsis verrucosa (Hortobágyi ex Loeblich, III) Ettl (Fig. 3h).
Dimensions: 3 × 12-16 μm. Similarity: The cell wall of B. verrucosa is relatively rigid in contrast with Juranyiella javorkae, which has also brown warts on its surface.
Ecological data: It has occurred in hypertrophic oxbow lakes. It was very rarely mentioned in the international literature. For more details, see in Supplementary Table 2.
Ecological data: It has occurred in eutrophic lakes with a few individuals in our study. It is a rarely mentioned species both in Hungary and in the international literature. For more details, see in Supplementary Table 2.
Dimensions of cell: 4-8 × 5-12 μm. Similarity: It is similar to C. dubius Printz, which strait apical spine is shorter than the cell length. C. ellipsoideus is also similar to C. belonophorus Fott, but the cell wall of C. ellipsoideus was thicker than in C. belonophorus.
Previous occurrences in Hungary based on Schmidt and Fehér 2001: Buzsák fish pond.
Ecological data: It has occurred in eutrophic, mainly oxbow lakes, we found only a few individuals in the samples. It was sporadically mentioned in the international literature. For more details, see in Supplementary Table 2.

Discussion
Cultural eutrophication, when nutrient input is caused by human activities, is a widespread problem and it affects many freshwaters. According to the International Lake Environmental Committee 40-50% of lakes and reservoirs were eutrophicated in global scale (Istvánovics 2009). In Hungary 36.5% of our lakes, that are monitored as EU WFD required, are eutrophicated (2nd RBMPs). In oligo-and mesotrophic water bodies rare algae could be more expected than in more loaded ones reported by Juráň (2017). However, the most species presented in this study were found in hypertrophic and eu-or hypertrophic waters.
Only three species, Cylindrotheca gracilis, Paradoxia multiseta and Kolkwitziella acuta occurred in less loaded reservoir and gravel pit lakes (oligotrophic or mezotrophic). Among them the Cylindrotheca gracilis is the only species which was classified into vulnerable category based on Hungarian Red List (Németh 2005). C. gracilis is rarely mentioned in Hungary, however this species is cosmopolitan. This species was former called "brackish water species" (Christensen and Reimer 1968). The conductivity of the gravel pit lake where we found this taxon was high. In Hungary, Paradoxia multiseta had sporadic in the 20th century Fehér 1998, Borics et al. 2016), despite of having widespread distribution. This species occurred only in low cell number in a reservoir and an oxbow lake. The Kolkwitziella acuta was formerly known as a permanent member of phytoplankton in Balaton and found other standing waters all the country (Grigorszky et al. 1999). Nowadays, K. acuta, which has special thecal plate scheme and its cyst indicates low salinities, is a very rare species both in Hungary and in the world (Mertens et al. 2015).
As we mentioned above, most of the rare species were found mostly in eu-, or hypertrophic standing waters. This supports the fact that rare species may also be present in eu-, or hypertrophic lakes. The distribution classification (common, rare and very rare categories) on the local scale were also used for classify the worldwide distribution data in a taxa rarity scale.
Some of the mentioned species have worldwide distribution. Furthermore, five of them, namely Chaetoceros muelleri, Closteriopsis longissima, Desmatractum indutum, Dicloster acuatus, Granulocystopsis coronata var. coronata, can be considered as common species in global scale. In spite of having common distribution in Hungary, Closteriopsis longissima has not mentioned since 30 years. It was occurred mostly in oxbow lakes, lakes and rivers. Now, it was detected in pelagial zone of eu-, hypertrophic oxbow, while Buczkó and Ács (1992) found this species on periphyton of green reed in the Szigetköz section of the Danube River. In form and size, Desmatractum indutum can be highly variable (Kiss 1978). It was usually found in oxbow lakes in Hungary (Schmidt and Fehér 1998). Here, this species also occurred in eu-, and hypertrophic oxbow lakes along the Tisza River and the Körös River. Granulocystopsis coronata var. coronata was formerly detected in planktic flora of oxbow lakes and large rivers as the Duna River and the Tisza River mainly in mid-summer period (Schmidt and Fehér 1998). Here, we found this taxon in oxbow lakes with connection of the mentioned rivers. Similar to our observations, Schmidt (1994) also reported sporadic occurrence of Chaetoceros muelleri in Hungary. This species is known as a marine/brackish algae also occurring in inland waters with high conductivity (Krammer and Lange-Bertalot 1991). Now, we found it in a shallow lake with high chloride content and also conductivity. In Europe, Dicloster acuatus was firstly reported from a Hungarian reconstructed wetland (Schmidt et al. 1991) and after that it was sporadically appeared in the Danube and its side-branches . Now, it was found in an oxbow lake of the Danube. In general, D. acuatus is sporadically found worldwide. According to Hindák (2004, 2005) Trachelomonas bacillifera var. minima and Trachelomonas woycickii are occurred widespread. Trachelomonas bacillifera var. minima was formerly found in standing waters with different characters all over our country (Németh 1997). Now, we found it in shallow, hypertrophic oxbow lakes. Trachelomonas woycickii is rarely mentioned in Hungary. Previously, T. woycickii was recorded mostly in different kind of standing waters. The new locality of this species, Nagy-Morotva, is a hypertrophic, shallow oxbow lake which is used for fishing.
Although Stauridium privum (syn. Pediastrum privum (Printz) Hegewald) has circumpolar boreo-alpine distribution and it is a common species (number of localities > 15), it was just sporadically mentioned from each country (Geriš 2004). From Hungary, S. privum has not been reported yet. Now, it was found in a small, shallow and eutrophic reservoir in the Mátra Mountains.
Some of the mentioned species are also rare both in Hungary and worldwide. Three of them Bumilleriopsis verrucosa, Centritractus ellipsoideus and Coronastrum ellipsoideum had only one earlier data from Hungary, from fish pond near Buzsák and from Lake Balaton respectively (Transdanubian part of the country). Here, these taxa appeared in shallow, mainly hypertrophic standing waters in the Transtisza region. Interestingly, Centritractus ellipsoideus was found in India under the ice cover of the Alpine Lakes of Arunachal Pradesh (Das 2016). Further worldwide rare taxa, Granulocystopsis decorate and Centritractus brunneus were published a few times according to the Hungarian identification manuals. The G. decorata was earlier reported from Hungarian large rivers such as the Danube River and the Maros River, now we found this species in an oxbow lake of the Danube River.
Three other species are also rare in the world, these species are Tetrastrum triacanthum, Juranyiella javorkae and Lagerheimia marssonii. Tetrastrum triacanthum was formerly described from oxbow lakes and slow flowing watercourses in Hungary (T- Krasznai et al. 2013). Here, we found only few specimens of this taxon in an oxbow lake. Lagerheimia marssonii was previously described from a fishpond near Gödöllő in North Hungary (Hajdú 1976), and rarely found in different waterbodies. Juranyiella javorkae was previously mentioned as rare species in Lake Kacsa that is a dead arm of the Tisza River (Kiss and Ács 2002). Now, we found it in oxbow lakes and reservoirs characterized by high trophic status. Based on our observations, the distribution of the latter two species, i.e. Centritractus ellipsoideus and Juranyiella javorkae, shows an expanding trend in the last decades.
Beside eutrophication the drastic changes of climate can endangered our waters and their species in global and local scale. The lack of precipitation, outflow or flushing and the strong evaporation can cause drastic changes in the water level. The water level reduction can lead the deterioration of water quality with increasing total nitrogen and phosphorus content and/or salinity of waters (Mosley 2015). Species, indicating this circumstances, usually are found in small waters (soda pans or small watercourses; Schmidt and Fehér 2001, Kókai et al. 2015, Stenger-Kovács and Lengyel 2015, Stenger-Kovács et al. 2016. Since climate models predict increase in frequency and intensity of extreme climatic events in the continental region (IPCC 2014, Babka et al. 2018), these small ponds and streams are endangered by droughts and flash floods (Stenger-Kovács et al. 2016, B-Béres et al. 2019. In this study, the Soponyai-Sós-tó desiccated regularly. Here, one species rare in Hungary, Nodularia spumigena was found. This is a planktic species occurring mainly in marine waters, but it is also known from inland waters with high conductivity (Komárek 2013). The pond where we found this species had the highest conductivity among the studied standing waters.
Nowadays waters are under-sampled in Hungary. Unfortunately, even if there are results of more waters, they haven't been represented in public database. Our results indicated that less-mentioned species can be found with low sampling effort even during national monitoring program. To answer the question what makes rare a species, we have to clarify whether the locality is unique, our knowledge is poor about its distribution or we are not able to determine the given species with traditional methods. To solve this task, the maintenance of long term water quality monitoring programs is necessary (Mosley 2015).
Despite spreading knowledge of new species or new distribution data of rare species, the biodiversity of our waters are still less known. Hundreds of different species can live together in an average lake (Cotthingham 1996). Heraclitus said many years before "You cannot step into the same river twice". This universal phrases can be also true for lakes. Although the average succession pattern of phytoplankton is known, it can differ from year to year (Scheffer 2004). The few dominant species or group can be more or less predictable, while the accompanying species can be rather erratic (Padisák 1993). To find the appropriate frequent and number of sampling we can define the diversity and structure of phytoplankton, it gives beauty for our works. The mentioned rare species can be easy determined, if we have up-to-date literatures, light microscope and an algologist. On the whole the systematic seasonal studies of different habitats are still necessary.
Declarations The authors declare that they have no financial and personal relationships with other people or organizations that can inappropriately influence their work. All data and material can be available from the authors.
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