European flora of the desmid genus Euastrum = Европейская флора десмидиевых водорослей из рода Euostrum. Специальные исследования рельефа клеточной стенки

M.V. LOMONOSOV MOSCOW STATE UNIVERSITY
Faculty of Biology
EUROPEAN FLORA 
OF THE DESMID GENUS 
EUASTRUM 
Special studies on the cell wall ornamentation
by Olga V. Anissimova
with 47 plates and 2 figures in text
*
*
KMK Scientific Press Ltd.
Moscow 2021

Olga V. Anissimova
European flora of the desmid genus Euastrum. М.: KMK Scientific Press. 2021, 109 p.
reviewers:
Dr. Marien Van Westen (Netherlands)
Dr. Elena Patova (Russia)
Анисимова О.В.
Европейская флора десмидиевых водорослей из рода Euastrum. Специальные исследования рельефа клеточной стенки. М.: Товарищество научных издавний КМК. 
2021, 109 с.
Основная идея этой книги — показать таксономическое значение топографии клеточной 
стенки на примере видов из рода Euastrum. В книге приведены результаты изучения рода 
Euastrum, одного из наиболее разнообразных родов десмидиевых водорослей. В общей 
сложности более 2000 клеток 45 видов Euastrum были исследованы с помощью сканирующего электронного микроскопа (СЭМ). Богатый видовой состав флоры Европы, большое 
количество образцов и более чем 10-летние исследования позволили увидеть и проанализировать распространение и взаимосвязь скробикул, бородавок, слизистых пор и традиционных морфологических признаков. Включение в список видов, рельеф оболочки которых 
еще не изучен, демонстрирует область будущих исследований в этом направлении. Объединение видов в группы в зависимости от наличия скробикул того или иного типа, а также 
в группы безскробикульных видов, я надеюсь, позволит коллегам в будущем более легко 
выявлять и описывать новые таксоны, а также обращать внимание на значимые признаки.
Во Введении приведено описание строения клетки Euastrum и основных структур, которые будут рассмотрены в систематической части. В Методической части описаны материалы, на основе которых проведено исследование, подробно объяснено, как изготавливать 
препараты для сканирующего электронного микроскопа, дан аннотированный перечень используемых терминов. В начале Систематической части подробно описан рельеф клеточной 
стенки, сходства и различия в строении клеток разных видов Euastrum. Впервые более 40 
видов были подвергнуты исследованию в СЭМ, кроме того, были обобщены данные по всем 
существующим в настоящее время Euastrum (74 вида) из Европы. Всего представлено 47 
таблиц с фотографиями в СЭМ структур поверхности клеточной стенки. В связи с тем, что исследование клеточных покровов проведено на образцах водорослей, собранных в водоемах 
России, приведены данные о распространении видов в регионах европейской части России. 
Распространение видов в Европе (с указанием стран) и в мире (с указанием континентов) 
приведено в описании таксонов. Список использованной литературы включает основные 
аннотации исследований видов рода Euastrum в Европе, в том числе публикации, в которых 
есть микрофотографии СЭМ.
ISBN 978-5-907533-03-5
© Anissimova О.V., 2021.
© Lomonosov MSU, 2021.
© KMK Scientific Press Ltd., 2021.

Contents
Preface.
....................................................................................................................... 4
Introduction.
............................................................................................................... 5
Methodical part.
......................................................................................................... 9
List of basic terms used in the text.
..................................................................... 14
Systematic part.
........................................................................................................ 16
Group 1. Species with scrobicules S1 and S2 or S3 and  S4 on the cell wall .
...... 19
Group 2. Species with only large scrobicules S4 on the cell wall........................ 29
Group 3. Species with only scrobicules S3 on the cell wall and 
with warts on the central part of the semicell ................................................... 31
Group 4. Species with only scrobicules S3 on the cell wall 
and without warts on the central part of the semicell .
...................................... 39
Group 5. Non scrobicular species with a pitted cell wall.
.................................... 49
Group 6. Non scrobicular (smooth-cell wall) species.......................................... 53
Group 7. Non scrobicular species with groups warts on the cell wall................. 59
Species of uncertain position ............................................................................. 58
Literature.
................................................................................................................. 59
Index.
........................................................................................................................ 61
Figures (Plate 1–47).
................................................................................................. 63
3

Preface
The main idea of this work was to show the taxonomic importance of the 
topography of the cell wall using the example of species from the genus Euastrum, one of the more diversified investigated genera of desmids. This manuscript is based on more than ten years of my research on cell wall features 
Desmidiales. It combines a critical summary of the previous studies with an 
in-depth analysis of new results that we didn’t know prior. In total, more than 
2000 cells of 45 species of Euastrum were examined in a scanning electron 
microscope (SEM). The algal samples were collected from different localities 
in the European part of Russia. The rich species composition of the flora of 
Europe made it possible to analyze the distribution and relationship of scrobicules, warts, mucous pores, and traditional morphological characters. The 
inclusion in the list of species, the ornamentation of which has not yet been 
studied, demonstrates the area of future research in this direction. I hope that 
dividing species into groups depending on the presence of scrobicules of one 
type or another and groups of non-scrobiculous species will allow colleagues 
to identify and describe new taxa more easily and pay attention to significant 
characters. The methodological part describes the base research materials, 
details how to make samples for the SEM, and provides an annotated list of 
terms used. 
The Systematic part contains a summary and the reasoning behind the 
choice of species groups, descriptions of all the extrapolations, and pointers 
for future studies. 
In this part of the manuscript, I give detailed descriptions of the ornamentation of the cell wall, the similarities and differences in the structure of cells 
of different species of Euastrum. In addition to generalizing data on all currently existing Euastrum (74 species) from Europe, more than 40 species were 
researched using the Scanning electron microscope for the first time. A total 
of 47 plates with SEM pictures of the cell wall surface structure are presented. 
Considering the difficulties currently arising in the identification and systematization of multispecies genera of algae, I believe that this manuscript 
will interest many algologists. This book can be used as an atlas to identify 
species of the genus Euastrum.
I am much obliged to my colleagues Dr Aliya Luknitskaya, Dr Mikhail Voitekhov and Dr Dmitry Philippov for kindly lending me samples for study. I am 
much indebted to my colleagues Dr Marien Van Westen, Dr Elena Patova and 
Dr Dmitry Chudaev for valuable recommendations and screening the text. 
SEM studies were carried out using the Shared Research Facility “Electron Microscopy in Life Sciences” at Moscow State University. The studies were carried out within the framework of governmental assignment of the Lomonosov 
Moscow State University, Part 2. Art. 01-10 (No. 121032300103-6). 
4

Introduction
The genus Euastrum is one of the large genera in the family Desmidiaceae, 
numbering, according to different authors, approximately 265 species. Traditionally, this genus was separated from other desmids by morphological signs. 
As principal characters, Ehrenberg (Ralfs, 1848) indicates a deep constriction 
between semicells (sinus), undulating or separated into lateral and apical 
lobes at the margins of the semicells and the presence of a median incision 
or incurvation at the polar lobe. The surface of the semicells is uneven, ornamented with inflations at the centre or the margins, and covered with warts 
and pits.
At the moment, some authors offer descriptions of the genus in varying 
degrees of detail: so, Coesel & Meesters (2007) give only a brief overview, 
while the books of Kosinskaya (1960) and Růžička (1981) contain extensive 
and detailed descriptions. A wide variety of morphology and variability of 
some structures leads to a more significant number of intraspecific taxa, 
while some characters (e.g. the apical incision or lateral and basal lobe) are 
poorly expressed. It causes difficulties for the identification and description of 
new species. As pointed out by Růžička, many species of Euastrum are similar 
to Cosmarium and Micrasterias. Unfortunately, there are not many species 
of Euastrum with large cells, and in small-celled taxa, cell wall structure and 
morphological features are not always visible under a light microscope.
Molecular-genetic investigations of some taxa of desmids demonstrated that genera with high numbers of species are polyphyletic and require a 
closer taxonomic study. Thus, representatives of Euastrum belong to several well-supported clades together with Cosmarium (Gontcharov, Melkonian, 
2008, 2011). Comparison between molecular data and morphological characters traditionally used by systematics does not demonstrate clear phylogenetic relationships in desmids. Therefore, other previously thought unimportant 
characteristics should be investigated and taken into account.
In this work, I will not take such characters of species as linear cell sizes, 
outline, the shape of the lobes and the structure of chloroplasts, which are 
distinguishable under a light microscope, into consideration. Several publications (Kosinskaya, 1960; Růžička, 1981; Coesel & Meesters, 2007 et al.) 
describe these characteristics in detail. The situation is different concerning 
the cell wall’s surface structure, which is distinguishable only using scanning 
electron microscopy (SEM) and is described for a small number of species. 
5

Fig. 1. Schematic drawing of types of pores — Р (From Neuhaus, Kiermayer, 1981 with changes) 
and scrobicules — S.
(Couté, Tell, 1981; Coesel, 1984; Wei, 1991; Šťastný, 2010; Anissimova, 2015 
and etc.).
The cell wall of the Euastrum has specific pits, which are called scrobicules 
(lat. scrobiculae). In the literature, there are different spellings of the word 
scrobicules and scrobicles, but they mean the same thing. Their size, shape 
and quantity vary from species to species. Most authors designate any concavity on the cell wall surface by the general term scrobiculate (lat. scrobiculatus). Thus, in the work of Couté and Tell (1981), the description of scrobicules 
is given as: “... la’ paroi cellulaire est creusée de petites fossettes qui, selon les 
cas, peuvent être plus ou moins profondes. Les pores n’apparaissent que dans 
les dépressions, un seul au centre de chacune”1. In my opinion, this definition 
is generally correct; however, the pores are not always present and may be 
located outside depressions if present. Moreover, this structure should not 
be confused with the cellularity of the wall (e.g. in Cosmarium or Actinotaenium).
In Euastrum, pits evenly cover the cell wall and correlate with the location of the pores. Our previous studies have shown that all scrobicules are 
invaginations outside the secondary cellulose layer; they are practically not 
observable from the inside (Anissimova, Staer, 2018). Many species of Euastrum have scrobicules whose positions are regular and often correlated with 
1 “...cell wall is pitted with small depressions, which can be of different deepness. The pores are present 
only in depressions, one pore in the centre of each”.
6

the groups of warts and inflations. The scrobicules can be divided into four 
groups according to shape and size (Anissimova, 2016). 
Scrobicules S1 are large scrobicules, clearly visible in a light microscope. 
S1 are presented by two levels of depressions (Fig. 1). The more significant 
depression is shallow, round or triangular, 4.9–10.1 µm in diameter, with 3–5 
mucilage pores penetrating it. Inside, there is a smaller depression (1.24–1.90 
µm in diam.), more often rounded but sometimes slightly elongated; pores 
are absent. Scrobicules S1 (in numbers from 1 to 7) are located in the central 
part of the semicells, often between inflations. This scrobicule type was called 
“central mucilage pores” (Kosinskaya, 1960; Neuhaus, Kiermayer, 1981). Neuhaus and Kiermayer (1981) described this structure as pore type 1 (P1). However, our investigations have shown that mucilage pores are present only in 
the large depression, not in the inner one. Therefore, we suggest that the 
description of this pore type need to be changed. (Fig. 1).  
Scrobicules S2  are smaller (3.4–4.4 µm at across). The several (2–4) 
deepening segments form the rosette shape structure, with the pore opening 
in the centre situated on the elevation of the cell wall (Fig. 1). Usually, S2 
can be found over the whole surface of the semicell, except for apices of the 
inflations and apices of the lobes. Neuhaus and Kiermayer (1981) described 
this structure as a variety of pore type 2 (Neuhaus, Kiermayer, 1981: р. 15, fig. 
18). Unfortunately, the authors did not present a schematic drawing, and the 
drawing does not correlate with the description. In our previous article, this 
pore type was redefined as P7 (Anissimova, Staer, 2018).
Scrobicules S3 are characterized by the absence of pore openings. They 
have various outlines: from small circular (0.4–1.5 µm) or elongated (2.5–2.7 
µm) to ring-shaped (3.0–3.5 µm). Scrobicules S3 can be present by a single 
one near the centre of the semicell or small groups between the lobes and under the apical lobe. Rarely do they cover the complete surface of the semicell.
Scrobicules S4 are smaller than others in size (1.3–1.6 µm at diam.), usually 
circular in outline, deep with even margins and mostly have only one pore 
(exceptionally, two). Accordingly to Neuhaus and Kiermayer (1981), these 
scrobicules correspond to pore types P2 and P3. Scrobicules S4 are mostly 
located on the apices of the inflations and lobes but can also cover the whole 
surface of the semicell.
As shown by our previous investigations (Anissimova, 2016; Anissimova, 
Staer, 2018), the differential feature of Desmidiaceae genera is the presence 
7

of scrobicules S1–S3, which can be found only on the cell wall of Euastrum. S4 
scrobicules can also be found in other genera of desmids.
It should be noted that all species of Desmidiaceae have the primary wall 
produced during morphogenesis. They shed it after the formation of the secondary wall. The scrobicules, pores and warts on the cell wall will be visible 
in SEM only after this process. In addition, the mucilage pores on the cell wall 
often have small plugs (Brook, 1981), which can be confused with small warts. 
8

Methodical part
The present work is based on investigations of the cell wall of 50 species 
of Euastrum, which were found in different biotopes of lakes and bogs. In this 
work, I used samples of algae personally collected from 1998 to 2019 in different types of water bodies of the European part of Russia: Moscow, Kursk, Tver, 
Murmansk, Leningrad regions, the republics of Karelia and Komi. Some samples were kindly provided to the author by Dr A. F. Luknitskaya (samples from 
Leningrad region), Dr L. A. Philippov (Vologda and Arkhangelsk reg.) and Dr 
M. Ya. Voitekhov (Pskov reg. and republik of Komi). Species with no available 
samples for investigation were described in previously published SEM images 
(Coesel, 1984; Kowets, 1984; Couté, Tell, 1981; Wei, 1991; Gontcharov, Watanabe, 1999; Salazar, 2007; Šťastný, 2010; Van Westen, 2015; Van Westen, Coesel, 2020). 
I was able to study more than 2000 photographs of different Euastrum 
species from different regions of the European part of Russia. It suggests that 
the shell structure is a more or less stable trait of the species. In this regard, 
I consider it possible to extrapolate the obtained data to the same species 
inhabiting Western and Central Europe.
Moreover, I thought it was necessary to include the 23 species never studied by SEM technique and whose cell wall surface structure has never been 
described in the list. These species have their “counterparts”, very similar outlines, with the well-studied structure of the cell wall surface. I believe this 
allows me to assume the cell wall structure as well. Undoubtedly, such species 
require additional research and my mentioning them in the text draws the 
attention of researchers to them.
For long-term storage, collected samples were fixed with a 4% formaldehyde 
solution or complex fixative (FAA) comprising formalin, acetic acid and ethyl 
alcohol (Gough et al., 1976). The use of acetic acid and ethyl alcohol clarifies 
the cell’s contents, making it possible to study such material under a light microscope better. Previously, all the material would have been examined under 
a light microscope using the phase contrast and differential contrast methods. 
Nevertheless, a detailed study of mucus pores, scrobicules and warts is impossible without a large magnification (more than x1000). For this, I recommend 
using a scanning electron microscope (SEM). Samples for the SEM analysis were 
prepared on slides with poly-L-lysine. It helped to ensure better adhesion of the 
desmid cells and reduced the number of lost cells. For this purpose, 3 μl of an 
9

aqueous poly-L-lysine solution (concentration 1%) was applied on a clean cover 
glass 20x20 mm and dried by heating at 50°C on a heating table. The procedure 
was repeated three times. Then several drops of the fixed material were placed 
on the glass, and, when almost dried, the coverslip was transferred into ethyl 
alcohol. To dehydrate the fixed samples increasing concentrations of ethyl alcohol (30%, 50%, 70%, 90%, 96%) for 10 min and subsequent exposure to 100% 
acetone for 10 min were used. According to the literature, there are different 
methods of dehydration (for example, only using acetone) and different concentrations of substances. Rules state that after 90% ethyl alcohol, you need 
B
A
C
Fig. 2. The basic elements and structures of Euastrum cell. A — frontal view; B — lateral view; 
C — apical view.
BI — basal inflations, BL — basal lobe, LL — lateral lobe, AI — apical incision/incurvation, ALL — 
apex of lateral lobe, lateral angle, A — apex, MAI — margins of apical incision, I — istmus, CI — 
central inflations, Sc — semicell, AL — apical lobe, Si — sinus, BA — basal angle, AA — apical angle, 
S1 — “middle pore” (scrobicules S1), S2-4 — scrobicules S2-S4.
10