Variety of Cell Size of Cosmarium spp. and Euastrum spp. (Desmidiaceae, Charophyte) from the Aquatic Environment around Palangka Raya, Central Kalimantan, Indonesia

Key word: Microalgae Cosmarium Euastrum Chlorophyceae Desmidiaceae Desmids (Desmidiaceae) is the family of green algae that characterized by cells that are shaped of two symmetrical semicells and it is estimated that there up to 12,000 species of Desmids. Cosmarium and Euastrum are two genera of Desmids that considered as the most representative taxa of this family. They are vary widely in size and shape of the cells. Based on the results of preliminary observations, in the aquatic enviroment around University of Palangka Raya is rich in microalgal diversity including Desmids. Cell size and shape are the most important points to be known to study the microalgae. This research is aimed to study the variety of cell size of Cosmarium spp. and Euastrum spp. from the aquatic environment around Palangka Raya. The results of this study show that the Cosmarium cells have the dimension of 17-80 μm long and 14-57 μm wide; Euastrum cells have the dimension of 24-89 μm long and 17-80 μm wide. Article history: Received: 05/03/2021 Revised: 15/07/2021 Accepted: 17/07/2021


Introduction
Desmids (Desmidiaceae) is the family of green algae that characterized by cells that are shaped of two symmetrical semi-cells (Meesters & Coesel, 2007) and it is estimated that there up to 12,000 species of Desmids (Coesel & Krienitz, 2008). Cosmarium and Euastrum are two genera of Desmids that considered as the most representative taxa of this family. Cosmarium is one of the oldest genera of the family Desmidiaceae with approximately 1,026 species and Euastrum has 304 species and infraspecies which are accepted taxonomically (Guiry & Guiry, 2021). Based on the results of preliminary observations, in the aquatic enviroment around University of Palangka Raya is rich in microalgal diversity including Desmids. They are vary widely in size and shape of the cells. Cell size and shape are the most important points to be known to study the diversity of microalgae, because each microalgae species has its own distinctiveness in terms of the size and shape of its cells.
The study of the variety of cell size and shape of microalgae is necessary as there no official data to be used as a reference for further research and as a learning material for the courses related to biology, especially microalgae, at University of Palangka Raya. This research is aimed to study the variety of cell size of Cosmarium spp. and Euastrum spp. from the aquatic environment around Palangka Raya, Central Kalimantan, Indonesia.

Materials and Methods Study Sites and Sample Collection
This research is a descriptive quantitative research with direct observation method. The samples were collected from the water canals around University of Palangka Raya, which was divided, into 3 sampling sites (figure 1). It was conducted between November 2020 and January 2021. Sample identification was carried out in the Laboratory of Biology Education Program.

Species Identification
The cell morphology was observed under the microscope at 400× (10× ocular and 40× objective) magnification using Olympus CX21 light microscope and photographed using 64-megapixel Redmi Note 9 Pro camera for further identification. The identification of Cosmarium and Euastrum species is based on the observed morphological characteristics using the following published works: (1) A Beginner's Guide to Freshwater Algae (Belcher & Swale, 1978); and (2) How to Know the Freshwater Algae (Prescott, 1970). At this stage the database from the algaebase.org site is also used as a reference for identification of microalgae species.

Cell Size Measurement
The estimation of Cosmarium and Euastrum cell size (length and width) was done by calculating the ratio of the object to the diameter of field of view at certain magnification ( Table 1). The simulation of the cell size measurement is shown in Figure  2 in details.  Based on Figure 2, position the object horizontally to the diameter of the microscope field of view to measure the cell width. The measured images of Cosmarium and Euastrum are then processed to add a scale bar using ImageJ software version 1.53g (Rasband, 2020).

Results and Discussion
Cosmarium spp.
Morphologically, the Cosmarium cell is in the form of two symmetrical semi-cells which are divided by a narrowing in the area between the two semi-cells called isthmus. Semi-cells of Cosmarium have several forms of which are elliptical, spherical, hemispherical, angular, pyramidal or red beanshaped (Belcher & Swale, 1978;Janse van Vuuren et al., 2006). The nucleus lies in the isthmus, appearing white between the two semi-cells (Osório et al., 2018;Prescott, 1970).
The number Cosmarium species found in this study were 6 (six) species, namely Cosmarium botrytis, Cosmarium biretum, Cosmarium granatum, Cosmarium trilobulatum, Cosmarium sp.1, and Cosmarium sp.2 ( Figure 3). The results of the measurement of Cosmarium cell lengths and widths are presented in Table 2. The length of the cells measured in this study is the length of the two semi-cells from end to end.  Table 2 shows the range of Cosmarium spp. cell sizes according to cell length and width. The cell length ranges from 17-80 μm and the cell width ranges from 14-57 μm. The largest species of Cosmarium based on this study is Cosmarium botrytis with a length of 80 μm and a width of about 57 μm. The smallest species is Cosmarium sp.2 with a length of 17 μm and width of 14 μm. According to Belcher & Swale (1978) and Janse van Vuuren et al. (2006), the dimension Cosmarium cells are 10-200 μm long and 6-140 μm wide.

Cosmarium botrytis
Cosmarium botrytis is the largest Cosmarium species found in this study with 80 μm long and 57 μm wide. In this study, Cosmarium cells were also observed after binary fission with asymmetrical cell shapes with 67 μm long and 40-47 μm wide ( Figure  4A).
The semi-cells are joined by a narrow isthmus at the cell equator (Bellinger & Sigee, 2010) and appear to be pyramidal in shape with rounded margins. The nucleus is clearly visible in the area between the two semi-cells (isthmus). (Sahin & Akar, 2019) said that the semi-cells of C. botrytis are ovate-pyramidal in shape with angles broadly rounded. The sinus appears deep and linear straight towards the isthmus. C. botrytis is also noticed by the two pyrenoids that are clearly visible in each semi-cell ( Figure 4B). Cosmarium botrytis is the only scientific name of this species that is currently accepted taxonomically (Guiry & Guiry, 2021).

Figure 4. Cosmarium botrytis: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Cosmarium biretum Cosmarium biretum is the second largest Cosmarium species found in this study with 44 μm long and 42 μm wide ( Figure 5). Cosmarium biretum has dark green color and the nucleus is clearly visible in the isthmus. Each semi-cell is shaped like a kidney and looks like two kidneys joined together with a closed sinus.

Figure 5. Cosmarium biretum: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Cosmarium granatum Cosmarium granatum cell has 30 μm long and 24 μm wide ( Figure 6). Cosmarium granatum cell is small, has green coloration, and the nucleus is clearly visible in the isthmus. Both semi-cells are pyramidal in shape with a rounded and truncate apex. There is a slight notch at the center of the apex. The sinus between the two semi-cells is slightly closed. The pyrenoids appear to be spherical in shape and clearly visible in each semi-cell.

Figure 6. Cosmarium granatum: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Cosmarium trilobulatum Cosmarium trilobulatum is the smallest Cosmarium species found in this study with 17 μm long and 14 μm wide (Figure 7). Cosmarium trilobulatum has two subtrapizoidal and 3-lobed semi-cells (Felisberto & Rodrigues, 2004). The apex of each semi-cell is wide and truncate, sometimes with a slight notch at the center region (Lee, 2015). The sinus between the two semi-cells is closed. The pyrenoids appear to be spherical in shape and clearly visible in each semi-cell.
C. trilobulatum originally described by Reinsch 1866 and published in his paper in 1867 and currently is accepted taxonomically (Guiry & Guiry, 2021).

Figure 7. Cosmarium trilobulatum: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Cosmarium sp.1 Cosmarium sp.1 cell has 24 μm long and 21 μm wide (Figure 8). The semi-cells appear to be pyramidal in shape with wide and truncate apex. Pyrenoids are spherical and have a color that appears to almost blend with the cell's color. Cosmarium sp.2 Cosmarium sp.2 cell has 29 μm long and 28 μm wide (Figure 9). The semi-cells appear to be elliptical with rounded apex; there are thorn-like/spine-like structures throughout the outer membrane; the pyrenoids have a dark green color that looks very different from the green of the cell as a whole; the sinus between the two semi-cells is widely opened.
Cosmarium sp.2 is the only Cosmarium species in this study that was observed to have ornamentation on its cell walls in the form of thorn-like/spine-like structures.

Figure 9. Cosmarium sp.2: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Euastrum spp. Euastrum cells are somewhat flattened and have angular lobes. Two identical semicells are formed from deep sinus constriction (Belcher & Swale, 1978). The semi-cells are oval, elliptical or pyramid-shaped and distinctly compressed when viewed from the side (Janse van Vuuren et al., 2006;Taft, 1945). The cell walls appear to be smooth or have ornamentations such as warts, granules, and spines depending on the species (Janse van Vuuren et al., 2006). Apex usually truncate with an incision. The nucleus is clearly visible and is located in the isthmus.

Figure 10. Euastrum species found in this study: (A-E) Euastrum didelta; (F) Euastrum bidentatum; (G) Euastrum denticulatum; (H) Euastrum pulchellum; (I-J) Euastrum turgidum
The result of the measurement of Euastrum cell lengths and widths is presented in Table 3 in detail. The length of the cells measured in this study is the length of the two semi-cells from end to end.  Table 3 shows the range of Euastrum spp. cell sizes according to cell length and width. The cell length ranges from 24-89 μm and the cell width ranges from 17-80 μm. The largest species of Euastrum based on this study is Euastrum sp.1 with a length of 89 μm and a width of 80 μm. The smallest species is Euastrum denticulatum with a length of 24 μm and width of 17 μm.

Euastrum didelta
Euastrum didelta is vary widely in the cell size among all Euastrum species found in this study. The cell length of E. didelta ranges from 50-71 μm and the cell width ranges from 32-42 μm (Figure 11b). The smallest E. didelta cell sizes observed in the study were 50 μm long, 32 μm wide and 14 μm wide of the apex.
The semi-cells are trapezoidal in shape (Silva & Felisberto, 2015) and divided by deep median constrictions that are almost completely enclosed with a slight opening at the end. The cell walls appear to be smooth without any ornamentations. The polar lobe with truncated apex, forming a lip; thick cell walls and adorned with punctuations (Silva & Felisberto, 2015). Aquino et al. (2017) described the semi-cells of E. didelta as pyramidal in shape with rounded lateral margin and apical margin truncate ( Figure  11a).

Figure 12. Euastrum bidentatum: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Euastrum denticulatum Euastrum denticulatum is the smallest species of Euastrum found in this study with 24 μm long and 17 μm wide ( Figure 13). The semi-cells appear to be subtrapeziform (Aquino et al., 2017) and semi-quadrangular (Silva & Felisberto, 2015) in shape, 3-lobed with tiny spine at each angles as the ornamental features of the cell walls. Apical lobes appear wide with deep V-shaped median notch. The sinus between the two semi-cells is enclosed. E. denticulatum originally described by Gay in 1884 and is currently regarded as a synonym of Euastrum amoenum F. Gay. E. denticulatum has two heterotypic synonyms that are E. denticulatum var. granulatum West 1892 and E. denticulatum var. angusticeps Grönbald 1921 (Guiry & Guiry, 2021).

Figure 13. Euastrum denticulatum: (A) Cell Dimension (length and Width) and (B) Cell Morphology
Euastrum pulchellum Euastrum pulchellum has 29 μm long and 20 μm wide ( Figure  14). Morphologically, E. pulchellum semi-cells look somewhat like E. denticulatum but with longer apical lobes. The semi-cells are quadrangular in shape with tiny spine at each angle as well; sinus between the two semicells is enclosed. The apical lobes appear wide with deep narrow median incision straight towards the pyrenoid. E. pulchellum originally described by Brébisson in 1856 and is currently accepted taxonomically (Guiry & Guiry, 2021). Euastrum turgidum Euastrum turgidum is one of the largest Euastrum species found in this study with 89 μm long and 80 μm wide. In this study, E. turgidum cells were also observed after binary fission with asymmetrical cell shapes with 80 μm long and 38-80 μm wide ( Figure 15). The semi-cells appear to be trapezoid in shape and are divided by deep closed-sinus. The apical lobe is very wide, straight and truncate (Scott & Prescott, 1960), approximately 75% of the width of the cell.
E. turgidum was first described by Wallich in 1855 and published in his paper in 1860 (Guiry & Guiry, 2021;Scott & Prescott, 1960). Euastrum turgidum is the scientific name of this species that is currently accepted taxonomically (Guiry & Guiry, 2021).