Effects of water levels on species diversity of silica-scaled chrysophytes in large tributaries of Lake Baikal

Large tributaries of Lake Baikal considered as a “hotspot” for silica-scaled chrysophytes diversity. Here we presented the updated species composition of silica-scaled chrysophytes and ecological parameters of their habitat in the Barguzin and Selenga River tributaries and delta in a high water level period. The number of registered taxa was significantly lower compared to the low water conditions (23 versus 66 species) and included the following genera with a given number of species: Chrysosphaerella – 1; Paraphysomonas – 2; Clathromonas – 1; Spiniferomonas – 3; Mallomonas – 9; Synura – 7. Mallomonas guttata and Synura borealis were identified in Russian waters for the first time. Thus, the corrected total list of silica-scaled chrysophytes in the Baikal Region includes 79 taxa. Though, the high water level reduced the total number of silica-scaled chrysophyte taxa, it made the water ecosystem more dynamic by enriching it with the entirely new species for this region.


Introduction
Lake Baikal is the most ancient and deepest (1637 m) lake in the world (Baikal. Atlas 1993). It has a tectonic origin and lies in a deep depression surrounded by mountain abundance and species diversity were recorded during the flood in July 2013 (Sorokovikova et al. 2017). The aim of our study was to investigate the impact of the high water level in July 2018 on the flora of silica-scaled chrysophytes in the Selenga and Barguzin River with their tributaries. We also compared the species composition in the tributaries under high and low water conditions.

Description of the area
The studied waterbodies are located in Russia, specifically in the south of East Siberia in the Republic of Buryatia (50°70'-53°82' N and 106°25'-109°90' E). They include the Selenga River, the mouth of the Kharauz Creek of the Selenga Delta, Lake Zavernyaikha, the Dzhida, the Temnik and Chikoy tributaries of the Selenga River, and the upstream portion of the Barguzin River (upward of the Ulyun River) with its mouth (Fig. 1).
The catchment area of the Selenga River (the main tributary of Lake Baikal) is primarily located in Mongolia, but its run-off is mainly formed in Russia. It increases three times in size from the Russian-Mongolian border to the mouth. The high altitude of the watershed and its significant slope formed the mountainous character of studied tributaries. The most full-flowing river is the Chikoy River (the right tributary of the Selenga River); its annual average run-off is 267 m3/s. The Dzhida and Temnik Rivers fall from the left bank; their run-off is significantly lower -67.6 and 29.9 m3/s, respectively (Sinyukovich 2005). The Selenga River forms a large delta that includes many creeks, lakes, and former riverbeds (Baikal. Atlas 1993). The Kharauz Creek is one of the largest in the delta. Lake Zavernyaikha is located in the Selenga Delta and is cut off the Kharauz Creek by a sand bar. During the floods and high water conditions, the lake is connected to the creek and thus has a good turnover. In winter and low water conditions, the lake is isolated from the creek (Popovskaya et al. 2011).
The Barguzin River is a tributary of Lake Baikal. Its run-off is the third largest by volume (after the Selenga and Upper Angara Rivers). At the upper reaches, this river is an impetuous mountain torrent that flows through a narrow gorge. When it enters into the Barguzin depression, the river flows on a broad valley and becomes a plain. Low parts of the flood plain have plenty of shallow eutrophic lakes and wetlands that are connected by a system of channels that provide the river with organic matter and other substances (Drucker et al. 1997). Specimens were deposited in the following collection: MCTP, Coleção de Aracnídeos, Porto Alegre (curator: Renato Augusto Teixeira) and the Smithsonian Museum of Natural History (SMNH), Arachnida and Myriapoda collection, Washington DC (curator: Hannah Wood). We attempted DNA extraction and amplification of DNA barcodes from legs of borrowed specimens preserved in ethanol, however this yielded no viable DNA.

Material and methods
We obtained the samples from the Selenga tributaries (Dzhida, Temnik and Chikoy), the Kharauz Creek and Lake Zavernyaikha in the Selenga delta and the Barguzin River (mouth and upward of the Ulyun River) in July 2018. Fourteen samples were used for analysis of the silica-scaled chrysophytes (Fig. 1).
We used portable pH meter (IT-1101; Russia) to measure pH, water temperature, and dissolved oxygen concentrations at the sampling sites (Manual for chemical analysis of inland surface waters, 2009). We filtered the samples for chemical analyses through 0.45 μm membrane filters (Advantec, Japan) and measured the conductivity at 25°C with a conductometer DS-12 (Horiba, Japan). We also used colourimetric and dichromate oxidisability (COD -chemical oxygen demand) methods to determine the nutrient concentrations and total organic matter content, respectively (Manual for chemical analysis of inland surface waters 2009); Wetzel and Likens 2003).
We took the algal samples from the surface layer of water (0 m) with a 1 L water sampler and fixed with Lugol's solution (1% f.c.). We also took 10-15 mL samples by means of Whatman membrane filters (pore size 1 μm, Whatman, USA). We identify the scaled chrysophytes using scanning and transmission electron microscopy. The samples for SEM analysis were filtered, dried at room temperature, coated with gold and examined using a Quanta 200 (FEI Company, USA) scanning electron microscope. The samples for TEM analysis were taken with water sampler, settled by the sedimentation method (Kuzmin 1975), centrifuged (MiniSpin, Eppendorf, Germany) and washed in deionized water. The washed samples were processed with 30% H2O2 at 75°С for 2 h, than the procedure was repeated and the samples were put on 3-mm-diameter formvar coated grids, dried at room temperature and analyzed by means of a LEO 906E transmission electron microscope (Carl Zeiss, Germany). The scales identified by means of electron microscopy classified the certain species according to their fine structure described and represented by microphotographs (Siver 1988;Hällfors and Hällfors 1988;Siver 1995;Němcová et al. 2012;Škaloud et al. 2012;Scoble and Cavalier-Smith 2014;Siver 2015). We also used the Freshwater Algal Database of Škaloud et al. (2013). Data on river flow rates were retrieved from Hydrometeorological Research Center of Russian Federation (Hydrometcenter of Russia).

Physical and chemical characteristics of the studied waterbodies
In July 2018, a continuous low water period in the catchment area of Lake Baikal ended when the Selenga and Barguzin Rivers rose and flooded their flood plains. Water discharge of the Barguzin and the Selenga Rivers during the sampling was up to 358 m 3 /s and 1700 m 3 /s, respectively (Table 1). It was 1.5-2 fold higher than in 2016 (Fig. 2).
The water temperature in the Selenga and its tributaries was 18.5-20.9°С (Table  1); it varied from 10.2°С in the upper reaches of the Barguzin River (station 7) up to 20.9°С in its mouth (station 6). High pH values were recorded in Lake Zavernyaikha (station 4), the Kharauz Creek (station 5) and at the upper reaches of the Barguzin River (station 7); the lowest pH was recorded in the Chikoy River (station 2). The highest conductivity of 228 µS cm-1 was observed in the Dzhida River (station 1); the lowest conductivity of 58 µS cm-1 was registered in the Chikoy River (station 2; Table 1). The dissolved oxygen content varied over a wide range (Table 1).
The water temperature in the Selenga and its tributaries was 18.5-20.9°С (Table  1); it varied from 10.2°С in the upper reaches of the Barguzin River (station 7) up to  20.9°С in its mouth (station 6). High pH values were recorded in Lake Zavernyaikha (station 4), the Kharauz Creek (station 5) and at the upper reaches of the Barguzin River (station 7); the lowest pH was recorded in the Chikoy River (station 2). The highest conductivity of 228 µS cm -1 was observed in the Dzhida River (station 1); the lowest conductivity of 58 µS cm -1 was registered in the Chikoy River (station 2; Table 1). The dissolved oxygen content varied over a wide range (Table 1). The crossflow of water from the Kharauz Creek (station 5) to Lake Zavernyaikha (station 4) caused by the high water level of the Selenga River levelled the oxygen content, water temperature, pH, and conductivity at these stations.
The upper reaches of the Barguzin River (station 7) showed an increased oxygen concentration due to its better solubility in cold water and aeration due to the higher stream speed. The lowest oxygen concentration was registered in the river mouth (station 6). The concentration of silicon was high at all stations (Table 1).
The inundation of the flood plains enriched the rivers with a large amount of organic matter from the catchment area and increased its water concentration. The highest concentrations were recorded in the mouth of the Barguzin River (station 6), Dzhida River (station 1), and Kharauz Creek (station 5). The total phosphorous values at all stations (except 3 and 7) were typical for polluted eutrophic waters.
The crossflow of water from the Kharauz Creek (station 5) to Lake Zavernyaikha (station 4) caused by the high water level of the Selenga River levelled the oxygen content, water temperature, pH, and conductivity at these stations. The upper reaches of the Barguzin River (station 7) showed an increased oxygen concentration due to its better solubility in cold water and aeration due to the higher stream speed. The lowest oxygen concentration was registered in the river mouth (station 6). The concentration of silicon was high at all stations ( Table  1).The inundation of the flood plains enriched the rivers with a large amount of organic matter from the catchment area and increased its water concentration. The highest concentrations were recorded in the mouth of the Barguzin River (station 6), Dzhida River (station 1), and Kharauz Creek (station 5). The total phosphorous values at all stations (except 3 and 7) were typical for polluted eutrophic waters.
Only seven species were identified in the Selenga tributaries Chikoy (station 1) and Dzhida (station 2), six species were recorded in the Temnik River (station 3). No chrysophytes were found in Lake Zavernyaikha (station 4) and the upper reaches of the Barguzin River upward of the Ulyun River (station 7; Table 2).
During the high water period, the chrysophyte flora was mainly represented by widespread and cosmopolitan species typical in the temperate and subarctic regions of Eurasia and North America. A total of 16 species were identified: Spiniferomonas cornuta, S. serrata, S. trioralis, Mallomonas acaroides, M. akrokomos, M. alpina, M. crassisquama, M. heterospina, M. guttata, M. striata, M. tonsurata, Synura echinulata, S. glabra, S. petersenii, S. spinosa, and S. uvella. Although the species composition varied in all waterbodies, two widespread species M. tonsurata and S. petersenii occurred everywhere. A single scale attributed to the genus Clathromonas was found in the mouth of the Barguzin River (station 6), but it was impossible to identify it to a species (Fig. 4e)   (mouth of the Barguzin River, station 6), f -Synura borealis (mouth of the Barguzin River, station 6), g -Synura heteropora (Chikoy River, station 2), h -Synura spinosa (Dzhida River, station 1), I -Synura echinulata (mouth of the Barguzin River, station 6), j -Synura uvella (mouth of the Barguzin River, station 6), k -Synura petersenii (Temnik River, station 3), l -Synura glabra (mouth of the Barguzin River, station 6). Micrographs were obtained with transmission electron microscopy; scale bars are 1 µm. Table 2. List of species and intraspecific taxa of the silica-scaled chrysophytes identified by electron microscopy in the Selenga tributaries and Barguzin River area in July 2018. See Fig. 1 for the location of the stations. A plus (+) indicates the presences of the species at the station. The asterisk (*) indicates the species observed in the Baikal Region for the first time.

Discussion
Biogeographical structure of silica-scaled chrysophytes of the studied area The flooding on the Selenga and Barguzin Rivers decreased the species diversity of silica-scaled chrysophytes (23 species) versus 66 species observed before this studyduring the low water period (Bessudova et al. 2018a). Although the chrysophyte species diversity decreased almost threefold, their biogeographical distribution changed proportionally (Table 3). Three species identified during this study rarely occur in Russian waters. One of them, C. baicalensis, is endemic to Lake Baikal, and two are arctoboreal species: Paraphysomonas acuminata acuminata and Paraphysomonas vulgaris.
P. acuminata acuminata was found and described in one Austrian freshwater lake (Scoble and Cavalier-Smith 2014). We identified the species in the mouth of the guzin River, the creeks of the Selenga Delta (including Lake Zavernyaikha; Bessudova et al. 2018a), Lake Baikal (Bessudova et al. 2017) and in the mouth of the Srednyaya Creek of the Angara-Kichera Delta (Bessudova et al. 2018b). This species occurs in spring, summer and autumn, while most numerous in May. During this study, the species was found in the Dzhida (station 1) and the Chikoy (station 2) Rivers.
P. vulgaris was found and described in freshwaters of England (Scoble and Cavalier-Smith 2014). To date, this species has been identified in the mouth of the Barguzin River and the Selenga tributaries (Bessudova et al. 2018a). A spine morphologically similar to those of P. vulgaris was also found in Lake Toko, Japan (see Gusev et al. 2018, fig. 39). During this study, P. vulgaris was registered in the Temnik River (station 3). Two species, Mallomonas trummensis and S. heteropora, have a limited distribution in Russian waters. M. trummensis occurs in waters of the temperate and subarctic zones of Europe (Škaloud et al. 2013).
In Russia, the species was identified only in the mouth of the Barguzin River and the creeks of the Selenga Delta (Bessudova et al. 2018a). Here, we found it only at one station, namely the mouth of the Barguzin River (Station 6). S. heteropora occurs in waters of Europe (Škaloud et al. 2014), but in Russia it was previously observed only in Lake Baikal (Bessudova et al. 2017), the mouth of the Barguzin River and the creeks of the Selenga Delta (Bessudova et al. 2018a). During the present study, S. heteropora scales occurred only in the Chikoy River (station 2).

Ecology of silica-scaled chrysophytes
The studied area is interesting not only to study the effect of floods on species composition of silica-scaled chrysophytes, but also to evaluate the impact on species ecology. The water parameters in study area during the low and high water levels were in a sharp contrast with the optimum for high diversity of silica-scaled chrysophyte (Eloranta 1995;Siver 1995;Siver and Lott 2017).
The highest silica-scaled chrysophyte diversity has been recorded in waters with pH low or close to neutral (below 7), low mineralization, conductivity close to or slightly less than 40 µs cm -1 , low nutrient content (oligotrophic to mesotrophic), and moderate quantity of dissolved humic compounds (Eloranta 1995;Němcova et al. 2003;Siver 1995;Siver and Lott 2017). A recent study in Newfoundland Island corroborated high silica-scaled chrysophyte diversity (47 species) in waters with pH 3.9-6.7, high content of humic substances and low nutrient concentration (Siver and Lott 2017).
Numerous studies allowed Siver (2015) to divide the silica-scaled chrysophytes into four groups with congruent boundaries along a pH gradient: (1) species that inhabit waters with a low pH (below 6); (2) species that inhabit waters with modern levels of alkalinity (pH below 7 but above 5); (3) species that inhabit waters with a neutral pH (pH-indifferent species), (4) species that inhabit waters with a high pH (above 7). One species, S. echinulata despite their affiliation with the low pH group, was found in waters with high pH (7.87; station 6). S. echinulata was previously observed in the Selenga Delta and Lake Zavernyaikha at pH 8.01 and 8.03, respectively (Bessudova et al. 2018a). We also found two species characteristic to average waters: M. heterospina (station 2) observed at pH 7.39 and S. spinosa (stations 1 and 6) observed at pH 7.87 and 8.0, respectively. M. heterospina was previously identified in the Selenga River and in creeks of its delta at high pH values (7.85 and 7.98), but it was erroneously identified as Mallomonas pugio Bradley (Bessudova et al. 2018a). S. spinosa was also previously observed in the Selenga River and in delta creeks at pH 7.7 and 8.03 (Bessudova et al. 2018a). Only two species characteristic to alkaline waters, namely M. alpina and M. tonsurata, occurred in the study area at pH 7.39-8.15. Němcová et al. (2003) demonstrated that the high diversity of silica-scaled chrysophytes was typical in waters with conductivity close to or below 40 µs cm -1 , whereas conductivity above 200 µs cm -1 reduced the species diversity.
The study area had high conductivity values during low and high water levels. The minimum conductivity values of 58 and 92 µs cm-1 were recorded only at two sites, stations 2 and 3, respectively (Table 1). However, this factor did not influence the diversity of silica-scaled chrysophytes; indeed, we observed the opposite situation where it were twice as many species at high conductivity value (174 µs cm -1 ; station 6). The highest silica-scaled chrysophyte diversity, (35 species and intraspecific taxa) was registered during the previous study in the mouth of the Barguzin River at a conductivity of 151 µs cm -1 (Bessudova et al. 2018a).
Despite some evidences that silica-scaled chrysophytes prefer oligo-and mesotrophic conditions (Eloranta 1995;Němcová et al. 2003;Siver 1995;Siver and Lott, 2017), high species diversity and biomass were also recorded in eutrophic waters (Cronberg 1996;Kristiansen 1985Siver 2015;Siver and Wujek, 1993), including the mouth of the Barguzin River in the Selenga Delta (Bessudova et al. 2018a). Notably, none of the species that would prefer oligo-and mesotrophic conditions according to Siver (2015) was found in the area studied. At the same time, M. tonsurata and M. alpina, both considered to prefer eutrophic waters, were found during our study in high and low water levels. Furthermore, these species also occurred in the plankton of the oligotrophic Lake Baikal (Bessudova et al. 2017).
Overall, the Baikal Region significantly expands the optimal conditions to develop and maintain high diversity of silica-scaled chrysophytes.
In large rivers, the flow rates increase proportionally to the rise of water discharge (Fig. 5). Hence, the conditions for phytoplankton growth in 2018 were less favorable compared to 2016.
The flood pulse concept elaborated by Junk et al. (1989) stated that a seasonal flood is useful for river ecosystems and could affect their biotic composition, nutrient transport, and sediment distribution. However, violent floods can be destructive for aquatic organisms (Talbot et al. 2018).
The seasonal flood in the study area was one of the factors influenced the species composition of silica-scaled chrysophytes in the mouths of the Selenga and Barguzin Rivers. During the flood, we observed a significant depauperisation in the chrysophyte species composition compared to the previous data. Thus, recent studies demonstrated that the beginning of a flood was accompanied by species impoverishment in plankton communities, even if the silicon and nitrogen concentrations were sufficient for their development (Talbot et al. 2018). We suggested that the concentrations of chemical components, including oxygen, silicon, nitrogen, and phosphorous could not limit the development of silica-scaled chrysophytes in studied area. These concentrations were similar or sometimes higher than in low water period (Table 1). However, most species previously observed in the creeks of the Selenga Delta and the mouth of the Barguzin River were absent during the flood. Nevertheless, the number of registered species were proportionally distributed among the genera (Fig. 6).  In previous studies, 15 and 20 species of silica-scaled chrysophytes were identified in Lake Zavernyaikha and in the mouth of the Barguzin River, respectively, under the low water level. However, chrysophytes were absent from the mouth of the Kharauz Creek in May, July andSeptember 2016 (Bessudova et al. 2018a). The increased water content in the main tributaries of Lake Baikal favoured significant changes in chrysophyte diversity in these areas. Thus, only 14 species were identified in the mouth of the Barguzin River (station 6) and five species in the Kharauz Creek (station 5), while they were absent in Lake Zavernyaikha (station 4). The high water level connected the waterbodies (stations 4 and 5), which levelled the difference in hydrochemical conditions (oxygen content, water temperature, pH, and conductivity). This phenomenon limited the development of some chrysophytes but did not favour similarity in their species composition.
In previous studies, 15 and 20 species of silica-scaled chrysophytes were identified in Lake Zavernyaikha and in the mouth of the Barguzin River, respectively, under the low water level. However, chrysophytes were absent from the mouth of the Kharauz Creek in May, July and September 2016 (Bessudova et al. 2018a). The increased water content in the main tributaries of Lake Baikal favoured significant changes in chrysophyte diversity in these areas. Thus, only 14 species were identified in the mouth of the Barguzin River (station 6) and five species in the Kharauz Creek (station 5), while they were absent in Lake Zavernyaikha (station 4). The high water level connected the waterbodies (stations 4 and 5), which levelled the difference in hydrochemical conditions (oxygen content, water temperature, pH, and conductivity). This phenomenon limited the development of some chrysophytes but did not favour similarity in their species composition.
The high diversity of silica-scaled chrysophytes in the mouths of the main tributaries of Lake Baikal, the Selenga, Upper Angara River and Barguzin Rivers in low water conditions could be caused by anterior floods. The inundation of the flood plains leads to integration of small creeks and lakes that enrich their flora due to dissemination of a broad spectrum of species (Fernandes et al. 2014;Junk et al. 1989). Water retreats control the bloom of phytoplankton, inter alia chrysophytes, in the warm and shallow waterbodies with high level of biological production, which favours the diversity of silica-scaled chrysophytes in the Baikal Region. Additionally, the climatic changes recorded worldwide during the previous decades, including the Baikal Region, may also underscore the quantitative and qualitative development of silica-scaled chrysophytes (Shimaraev et al. 2002;Sinyukovich et al. 2010;Sorokovikova et al. 2015). Global climate change influences the hydrology of waterbodies (Mushet et al. 2017) that will significantly impact the development of aquatic organisms. The interchange of floods and low water levels created various environmental conditions (Table 1) and stimulated dynamics of the ecosystem allow the formation of a "hotspot" for silica-scaled chrysophytes diversity.