International Journal of Systematic and Evolutionary Microbiology (2012), 62, 2870–2877
DOI 10.1099/ijs.0.038679-0
Toxopsis calypsus gen. nov., sp. nov.
(Cyanobacteria, Nostocales) from cave ‘Francthi’,
Peloponnese, Greece: a morphological and
molecular evaluation
V. Lamprinou,1 K. Skaraki,2 G. Kotoulas,2 A. Economou-Amilli1
and A. Pantazidou1
Correspondence
A. Pantazidou
[email protected]
1
University of Athens, Faculty of Biology, Department of Ecology and Systematics,
Panepistimiopolis, 15784 Athens, Greece
2
Hellenic Centre for Marine Research, Institute of Marine Biology and Genetics, Gournes Pediados,
PO Box 2214, 71003 Iraklio, Crete, Greece
Representatives of a new cyanobacterial genus, Toxopsis Lamprinou & Pantazidou gen. nov., were
found in fresh material from Cave ‘Francthi’ (Peloponnese, Greece) and isolated in cultures.
Ecological data relating to the environmental parameters of the sampling sites are provided, such
as the photosynthetically active radiation (PAR), temperature and relative humidity. Morphological
characteristics and the life cycle of the type species Toxopsis calypsus Lamprinou & Pantazidou
sp. nov. were studied using light microscopy and scanning and transmission microscopy.
Molecular analysis based on the 16S rRNA gene sequence was also conducted. Toxopsis
calypsus sp. nov. is a false-branched nostocalean cyanobacterium with both isopolar and
heteropolar filaments bearing mono-pored and bi-pored heterocysts, and also hormogonia and
akinetes. Isopolar filaments adhere by the centre to the substrate and are found mainly in fresh
material and in young cultures; heteropolar filaments bearing a basic mono-pore heterocyst are
dominant in aged (more than one-year-old) cultures. According to the revised taxonomic
classification system of Komárek & Anagnostidis (1989) [Komárek, J. & Anagnostidis, K. (1989).
Algol Stud, 56, 247–345] based mainly on morphological data, the new genus described here
shares morphological characters with both nostocalean families Scytonemataceae and
Microchaetaceae, showing similarities in particular to Scytonematopsis contorta [Vaccarino, M. A.
& Johansen, J. R. (2011). Fottea 11, 149–161], Microchaetaceae. Molecular data from the 16S
rRNA sequence determined in this paper showed that Toxopsis calypsus sp. nov. is more related
to the family Microchaetaceae, and the five phylotypes analysed by PCR showed that the closest
nostocalean relatives are Tolypothrix distorta SAG 93.79 (GenBank accession no. GQ287651)
and Coleodesmium sp. ANT.L52B.5 (AY493596) with 95–96 % and 96 % similarity, respectively.
In contrast, the five phylotypes showed a distant similarity to Scytonematopsis contorta (,91 %).
The phenotypic and genetic traits strongly supported the classification of the five phylotypes as a
new taxon for which the name Toxopsis calypsus Lamprinou & Pantazidou gen. nov., sp. nov. is
proposed.
INTRODUCTION
Caves represent a unique, stable and oligotrophic environment where light is the stressful environmental factor
Abbreviations: LM, light microscopy; PAR, photosynthetically active
radiation; SEM, scanning electron microscopy; TEM, transmission
electron microscopy.
The GenBank/EMBL/DDBJ accession numbers for the 16S rRNA gene
sequences for the five phylotypes of Toxopsis calypsus gen. nov., sp. nov.
are JN695681, JN695682, JN695683, JN695684 and JN695685.
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that controls phototrophic growth, and cyanobacteria
constitute the dominant group of the cave photosynthetic
microflora (Hernández-Mariné & Canals, 1994; Asencio
et al., 1996; Roldán et al., 2004; Lamprinou et al., 2009,
2011). Due to the unique environmental conditions
prevailing in the caves, several new genera and species
have already been established.
The floristic survey of caves worldwide shows that the
diversity of nostocalean cyanobacteria is very low, with the
most widely distributed nostocalean cyanobacterium in
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Toxopsis calypsus gen. nov., sp. nov.
cave ecosystems being the terrestrial Scytonema julianum
(Kützing) Meneghini (1849). There are no nostocalean taxa
established exclusively from caves. On the other hand, new
genera of the orders Stigonematales (Spelaeopogon, Geitleria,
Loriellopsis, Iphinoe) and Chroococcales (Asterocapsa) have
been established from cave ecosystems, as well as a number
of novel species including Spelaeopogon sommierii Borzi
(1917), Asterocapsa gloeotheceformis Chu (1952), Asterocapsa
hyalina Chu (1952), Asterocapsa trochiscioides Jao (Chu,
1952), Chroococcidiopsis kashaii Friedmann (1961), Geitleria
calcarea Friedmann (1955), Geitleria floridana Friedmann
(1979), Asterocapsa longipapilla Liu (1985), Gloeothece
filiformis Sant’Anna et al. (1991), Herpyzonema pulverulentum Hernández-Mariné & Canals (1994), Symphyonema
cavernicola Asencio et al. (1996), Loriellopsis cavernicola
Hernández-Mariné & Canals (2011) and Iphinoe spelaeobios
Lamprinou & Pantazidou (2011).
The identification process for a new nostocalean genus
from the Greek cave ‘Francthi’ accentuated the necessity of
the polyphasic approach in cyanobacterial taxonomy. It is
noted that identification in cyanobacterial taxonomy is
generally considered to be difficult and problematic
(Komárek, 2006, 2010; Lokmer, 2007; Korelusová, 2008).
The traditional classification system has been based on
morphological characters especially as observed in field
material (Gomont, 1892; Geitler, 1932; Desikachary, 1959;
Starmach, 1966), whereas bacteriologists have developed a
taxonomic scheme based on the physiological, molecular,
ultrastructural and morphological characteristics of cultured cyanobacteria (Stanier et al., 1978; Rippka et al.,
1979; Rippka, 1988). Since the last major classification
revision of cyanobacteria (Anagnostidis & Komárek, 1985,
1988, 1990; Komárek & Anagnostidis, 1986, 1989), many
new data derived from molecular evaluations and observations by electron microscopy have highlighted the need for
the revision of the existing taxonomic system (Hoffmann
et al., 2005; Johansen & Casamatta, 2005; Komárek,
2006).
Nowadays, a combination of morphological and molecular
data, as well as ultrastructural characteristics from field and
cultured material, has led to a modern classification system
enhancing the necessity of the polyphasic approach
(Hoffmann et al., 2005; Komárek, 2006), and resulting in
the establishment of several new taxa under all cyanobacterial families. The aim of the present paper is to describe a
new nostocalean genus of cyanobacteria, Toxopsis calypsus
Lamprinou & Pantazidou gen. nov., sp. nov., by the
application of the above-mentioned polyphasic approach
[field and culture material under light microscopy (LM),
scanning electron microscopy (SEM), transmission electron microscopy (TEM) and 16S rRNA gene sequence
analysis].
METHODS
Study area. Cave ‘Francthi’ (37u 259 21.010 N 23u 079 51.810 E;
altitude 12.5 m above sea level) is located in Argolida, Peloponnese,
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Greece. It is 150 m long with a maximum width of 45 m. According
to archaeologists, this cave represents a remarkable site on the coast of
south-east Greece since deposits found in the cave cover the period
from 20 000 BC up to 3000 BC.
Physical parameters. Air temperature (uC), relative humidity
(RH%) and photosynthetically active radiation (PAR; mmol
s21 m22) were measured using a LI-1400 data logger (LI-COR
Biosciences) over a whole year survey. From a collection of 52
specimens, Toxopsis calypsus gen. nov., sp. nov. was found only once
at a location nearest to the entrance sampling site. Over the full year
of the survey, the air temperature ranged from 13.15 to 26.18 uC, PAR
from 0.129 to 8.14 mmol s21 m22 and RH from 51.69 to 94.10 %.
Sampling and cultures. Sampling was conducted at different
distances from the physical cave entrance and from selected sites
inside the cave hosting various growth habits of cyanobacteria.
Sampling was conducted seasonally from 19 January 2009 to 8
November 2009. Samples were collected by scraping and treated
under sterile conditions. Part of the material was fixed with
formaldehyde solution at a final concentration of 2.5 % and another
part kept alive for culturing. Enrichment cultures were obtained in
flasks and Petri dishes with BG11 and BG110 (Stanier et al., 1971).
Cultures were maintained in an incubator (Sanyo, Gallenkamp)
under stable conditions under daylight (north facing window) at
room temperature for 2 years.
Microscopy. For LM, natural and cultured material was observed
on glass slides under a high-resolution light microscope
(Photomicroscope III, Zeiss). For SEM, specimens were dehydrated
in an alcohol series (30–100 %), critical point-dried and spray-coated
in gold–palladium and observed under a JEOL JSM 35 scanning
electron microscope.
For TEM, samples were fixed in a mixture of glutaraldehyde (2.5 %)
in 0.1 M cacodylate buffer for 2–4 h, washed three times in this buffer
and post-fixed in 1 % OsO4 in the same buffer. The organisms were
dehydrated by a graded acetone series, washed in propylene oxide
twice and subsequently embedded in three mixtures of propylene
oxide and resins (1/0.5, 1/1, 0.5/1) and finally in Spurr’s resin. Surface
sections were made with LKB Bromma 2088 ultratome. Highresolution TEM (HR-TEM) images were obtained using a JEOL JEM2100 electron microscope, operating at 80 kV.
Molecular analysis. For molecular analysis, DNA was extracted
from cultures according to the method of Fiore et al. (2000). PCR for
the 16S rRNA gene was conducted using cyanobacteria-specific
primers CYA359F 59-GGGGAAT(C/T)TTCCGCAATGGG-3 and
CYA781R 59-GACTAC(A/T)GGGGTATCTAATCCC(A/T)TT-39 (Nübel
et al., 1997) in combination with universal bacterial primers 27f
59-AGAGTTTGATC(A/C)TGGCTCAG-39 and 1492r 59-TACGG(C/
T)TACCTTGTTACGACTT-39 (Lane, 1991) in order to obtain an
almost full-length 16S rRNA product (1455 bp).
Purified PCR products were cloned into PCR II-TOPO vector (TOPO
TA cloning kit, Invitrogen), and positive clones were sequenced on an
ABI 3730xl 96 capillary sequencer (DNA Analysis facility on Science
Hill, Yale University, USA) using vector primers M13f and M13r.
Sequences were checked for chimeras using the Chimera Check
software included in the Ribosomal Database Project II and compared
with GenBank entries by using basic local alignments tool (BLAST) to
obtain a preliminary phylogenetic affiliation of the clones. Sequences
were edited in CodonCode Aligner v 3.7.1 and aligned using
CLUSTAL_X v 2.
Phylogenetic analysis was based on a 1117 bp fragment of the 16S
rRNA gene. Three trees were constructed using Bayesian, maximumlikelihood and neighbour-joining algorithms. Parameters were
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V. Lamprinou and others
estimated using the Akaike Information Criterion in jModelTest
(Posada, 2008). Maximum-likelihood analysis was run in PhyML v3.0
(Guindon & Gascuel, 2003) using the TVM (transversional) model
(Rodrı́guez et al., 1990), with a gamma distribution of 0.57 and
bootstrapped with 1000 replicates. The MrBayes 3.1.2 software
(Huelsenbeck & Ronquist, 2001; Ronquist & Huelsenbeck, 2003)
was used for Bayesian inference. Two runs were run for 107
generations, using one cold chain and three heated chains and
sampling every 100 trees. The first 10 000 000 samples of each run
were discarded as a burn-in phase. The model used was GTR+G (the
TVM model was not supported in the MrBayes software and GTR+G
was selected as the second best model). Neighbour-joining analysis
was run using MEGA 4 (Tamura et al., 2007) with the Tamura-Nei
(Tamura & Nei, 1993) substitution model and bootstrapped with
1000 replicates.
(oscillatorialean type) and Tolypothrix-like (microchaetacean type). Sheath hyaline, becoming yellowish brown in
mature filaments. Heteropolar filaments (10) 12–18 mm
wide (12.3±1.2 mm, n530), isopolar filaments 10–14 mm
(10.6±1.3 mm, n530). Trichomes constricted at the cross
walls, (4) 6–10 mm wide. Cells shorter than wide, 2–5 mm
long. Heterocysts mainly mono-pored (Fig. 1c) and rarely
bi-pored (Fig. 1d), occurring only at the Tolypothrix-like
stages, usually intercalary and rarely basal, 5–10 mm
wide and 5–9 mm long. Necridia dark bluish green.
Reproduction by akinetes and hormogonia. Akinetes
commonly in series, rounded or ellipsoid, vacuolized
(Fig. 1f), often divided in two halves. Hormogonia (Fig.
1g) with a basal heterocyst.
RESULTS
Life cycle Toxopsis calypsus exhibits two stages in the
Diagnosis for Toxopsis calypsus gen. nov.,
sp. nov.
Toxopsis gen. nov. [Tox.op9sis. Gr. n. toxon a bow; Gr.
suffix -opsis looking like; N.L. fem. n. Toxopsis (something)
looking like a bow].
Toxopsis calypsus sp. nov. (ca9lyp.sus. L. gen. n. calypsus of
Calypso, a nymph in Greek mythology).
Herbarium of Greece: ATHU-CY 3314.
Herbarium of Philadelphia (PH), Academy of Natural
Sciences: 1095565.
life cycle (Ammatoidea-like5isopolar, Tolypothrix-like5
heteropolar). The isopolar stage of life cycle was observed
initially in natural populations, and also in young cultures
(growth medium BG11) at least for a period of approximately one year; the heteropolar life cycle was observed in
older, 2-year-old cultures (growth medium BG11 and
BG110).
The ‘isopolar stage’ of the life cycle consists of filaments
(Ammatoidea-like) arched at the middle (Fig. 1a) and
gradually attenuated at both ends, producing hormogonia
next to necridia cells. The hormogonia germinate forming
young filaments, which gradually attenuate at both ends,
bending in the middle and bearing telescopic sheaths.
Thallus flavo-viridis ad atroviridis, formans parvas muscosas
aggregationes. Recenta fila isopolaria curva per medium.
Vetera fila heteropolaria cum pseudoramibus heterocystisque
basalibus et rariter cum monoporis et biporis intercalariis
heterocystis. Fila heteropolaria (10) 12–18 mm latitudine
(12.3±1.2 mm, n530), fila isopolaria 10–14 mm (10.6±
1.3 mm, n530). Cellula breviora quam latiora, 6–9 mm
latitudine, 2–5 mm longitudine. Multiplicatio hormogoniis et
akinetibus.
The ‘heteropolar stage’ of the life cycle is characterized by
heteropolar filaments with vegetative cells differentiating
into heterocysts and akinetes. Filaments slightly attenuated,
producing hormogonia, which germinate heteropolarly
with the formation of a basal heterocyst. Akinetes observed
exclusively in older cultures, rounded or ellipsoid,
vacuolized (Fig. 1f), often divided in two halves (Fig. 2e),
and commonly forming series.
Thallus green to dark green forming small moss-like strata.
Young filaments isopolar, bent in the middle. Older filaments
heteropolar and pseudobranched, with basal heterocysts and
rarely with intercalary heterocysts. Heteropolar filaments (10)
12–18 mm wide (12.3±1.2 mm, n530), isopolar filaments 10–
14 mm (10.6±1.3 mm, n530). Cells shorter than wide, 6–
9 mm wide, 2–5 mm long. Reproduction by hormogonia.
Akinetes present.
SEM and TEM observations SEM micrographs revealed
characteristic Ammatoidea-like filaments, which are bent in
the middle (Fig. 2a, c, f) and bear telescopic sheaths (Fig.
2b). TEM micrographs revealed the presence of cyanophycin
granules in the cytoplasm of vegetative cells (Fig. 2d) and the
thick multi-layered mucilaginous sheath of akinetes as well
as their division into two halves (Fig. 2e).
Molecular data – phylogenetic analysis (Fig. 3) Toxopsis
Morphology
LM observations Thallus yellow–green to dark green,
forming small moss-like strata. Young filaments isopolar,
gradually attenuated at both ends similar to the genus
Ammatoidea with thick telescopic sheath (Figs 1b, 2b).
Older filaments, pseudobranched (Fig. 1e), heteropolar,
beginning to coil within the sheath and forming
Tolypothrix-like branching. Presence of two morphologically distinct types of filaments, Ammatoidea-like
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calypsus gen. nov., sp. nov. was affiliated within the order
Nostocales and the closest relatives were Tolypothrix
distorta SAG 93.79 (GenBank accession no. GQ287651)
and Coleodesmium sp. ANT.L52B.5 (AY493596) with 95–
96 % and 96 % similarity, respectively. Most of the
representatives of the genus Tolypothrix (e.g. Tolypothrix
sp. IAM M-259 and UAM 334) had ,95 % similarity with
the isolated phylotypes. The topology of the Bayesian,
maximum-likelihood and distance (neighbour-joining)
analyses were very similar, in particular for the genera
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Toxopsis calypsus gen. nov., sp. nov.
Fig. 1. LM micrographs of Toxopsis calypsus gen. nov., sp. nov. (a) Vegetative filaments under a stereomicroscope; (b) a
filament bending in the middle with the characteristic telescopic sheath; (c) a filament with the mono-pored heterocyte; (d)
heteropolar growth with a basal mono-pored heterocyte (mp) and with an intercalary bi-pored heterocyte of the main filament
(bp); (e) false branched filaments with broad sheaths; (f) germination of the vacuolized akinetes; (g) formation of hormogonia
with basal heterocytes. Bars, 8 mm.
Spirirestis, Gloeotrichia, Scytonematopsis and Brasilonema.
The Bayesian tree with the node support from all three
analyses is presented in Fig. 3. The node containing the
Toxopsis calypsus phylotypes was well supported, but its
position in the tree was not as clear. Five phylotypes were
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analysed and the estimation of the similarities between
the sequences was 99.4–99.9 for 1455 nt positions. The
evolutionary analysis was conducted with MEGA5 (Tamura
et al., 2011). All sequences were submitted to GenBank
under accession numbers JN695681–JN695685.
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V. Lamprinou and others
Fig. 2. Micrographs of Toxopsis calypsus gen. nov., sp. nov. (a) and (c) Isopolar filaments under SEM, bending in the middle
and slightly attenuated at both ends; (b) the characteristic telescopic sheath, under SEM; (d) longitudinal section of filaments
under TEM, with typical cyanophycin granules; (e) an akinete divided into two halves, under TEM; (f) longitudinal section at the
middle of a bent filament, under TEM. Bars, 100 mm (a, c), 10 mm (b, f), 1 mm (d), 5 mm (e).
DISCUSSION
The presence of heterocysts, the formation of akinetes and
the false branching are the diacritic features classifying the
genus Toxopsis gen. nov. among the nostocalean cyanobacteria in the existing classic taxonomic systems based on
morphology. However, the new genus shows clear
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phenotypic and morphological differences from the other
nostocalean genera, thus deserving a distinct taxonomic
status. In detail:
According to the revision of the botanical system
(Anagnostidis & Komárek, 1985, 1988, 1990; Komárek
& Anagnostidis, 1986, 1989), the order Nostocales com-
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Toxopsis calypsus gen. nov., sp. nov.
0.01
Fig. 3. Bayesian analysis tree based on 16S rRNA gene sequences (1117 bp) gene. GenBank numbers in bold type were
determined in this study. Node support is indicated as Bayesian posterior probabilities/bootstrap support from maximumlikelihood analysis/bootstrap support from neighbour-joining. *, 1.0 or 100 %; –, ,0.50 or 50 %. Bar, 1 % nucleotide
substitutions per position.
prises four families (Scytonemataceae, Microchaetaceae,
Rivulariaceae, Nostocaceae). However, Hoffmann et al.
(2005) claimed that, due to the molecular and ultrastructural data that have been accumulated, this classification
system does not reflect the phylogeny of cyanobacteria, and
proposed a taxonomic system with four subclasses and six
orders. Within the latter taxonomic system, the order
Nostocales comprises ten families (Scytonemataceae,
Symphyonemataceae, Borzinemataceae, Rivulariaceae, Microchaetaceae, Nostocaceae, Chlorogloeopsidaceae, Hapalosiphonaceae, Loriellaceae and Stigonemataceae).
Comparing the phenotypic and morphological features
of Toxopsis gen. nov. with those of the other nostocalean
genera, and especially the presence of isopolarity and
heteropolarity as well as the vacuolized akinetes divided
into two halves, it is clear that there is a closer relationship with the species Scytonematopsis contorta (Vaccarino &
Johansen, 2011). However, there is a clear phylogenetic distance between the novel taxon proposed here
and members of this species (similarity was ,90 %)
according to molecular data (16S rRNA gene sequence
analysis).
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Sequence data from the 16S rRNA gene sequences of
members of Toxopsis gen. nov. showed a relationship with
genera of the family Microchaetaceae (see Fig. 3).
According to the classic revision of the botanical system
(Komárek & Anagnostidis, 1989), the main diacritic feature
of the microchaetacean genera is the heteropolarity of
filaments and also the characteristic life cycle, i.e. the
heteropolar development and growth of hormogonia leading to the heteropolar structure of filaments and thallus.
The monospecific genus Toxopsis gen. nov. is distinguished
from all other genera of the family Microchaetaceae by the
autapomorphic trait of both the isopolar young filaments
arched at the middle and the heteropolar older filaments.
This autapomorphic trait is the criterion necessary for
the establishment of a novel species according to the
phylogenetic species concept (Flechtner et al., 2002;
Johansen & Casamatta, 2005) and according to the monophyletic species concept sensu Mishler & Theriot (2000).
Furthermore, for defining prokaryotic species the Ad Hoc
Committee on Reconciliation of Approaches to Bacterial
Systematics (Wayne et al., 1987) recommended the use
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V. Lamprinou and others
of DNA–DNA hybridization studies. According to this
approach, the values to be considered simultaneously
are ¢70 % DNA–DNA relatedness and ¡5 uC DTm.
Stackebrandt & Goebel (1994) correlated DNA–DNA
reassociation values and 16S rRNA gene sequence similarity and found that species with ¢70 % DNA relatedness
always had .97.5 % 16S rRNA gene sequence similarity.
With gene sequence similarity of ,97.5 %, one can be
confident that the DNA–DNA reassociation is ,70 % and,
thus, the two organisms can be considered to represent
separate species. In our case, Toxopsis calypsus sp. nov.
showed 96 % (and some strains 95 %) similarity with
Tolypothrix distorta (SAG 93.79, GenBank accession no.
GQ287651) and 96 % similarity with Coleodesmium sp.
ANT.L52B.5 (AY493596). From the above-mentioned, one
can conclude that Toxopsis calypsus sp. nov. definitely
cannot be affiliated under the strain Tolypothrix distorta
and Coleodesmium sp. according to molecular data.
morphological characteristics that do not permit the
identification of our new specimens as members of the
genera Tolypothrix or Coleodesmium. Furthermore, the
establishment of this new genus is in accordance with
Anagnostidis & Komárek (1985), who have suggested that
‘small’ genera within the cyanobacteria should be accepted
and retained (see also Anagnostidis & Roussomoustakaki,
1985) rather than ‘larger’ genera subdividing into
subgenera.
The differentiation in molecular taxonomy between the
genera and the upper taxonomic levels such as families is
more difficult because of the fewer criteria that have been
proposed (Flechtner et al., 2002). The taxonomic concept
that could be used at the level of genera is the phylogenetic
taxonomy concept sensu Mishler & Theriot (2000) in
which species are the smallest monophyletic groups and, as
with all hierarchical levels of taxa in such a classification,
organisms are grouped in species when evidence of
monophyly is present. As a result, Toxopsis gen. nov., with
its morphological autapomorphy (isopolarity, heteropolarity) indicating monophyly, consists a new genus.
REFERENCES
Furthermore, in the 16S rRNA gene sequencing, 95 %
genetic similarity was proposed as the criterion for
separating generic clusters (Wayne et al., 1987; Komárek,
2006). If the similarity is close to 95 %, the presence of a
clear phenotypic difference or other criteria (biochemical,
ecophysiological) should be decisive (Komárek, 2006).
Toxopsis calypsus sp. nov. shows 95–96 % similarity with T.
distorta (SAG 93.79, GenBank accession no. GQ287651) as
well as 96 % similarity with Coleodesmium sp. ANT.L52B.5
(AY493596). Therefore, the phenotypic difference between
them is important to be able to make a taxonomic
distinction. Toxopsis calypsus sp. nov. is characterized by
both isopolar and heteropolar filaments bearing monopored and bi-pored basal (and sometimes intercalary)
heterocysts and by akinetes. In contrast, the genus
Tolypothrix (Kützing, 1843) is characterized only by
heteropolar filaments bearing basal heterocysts, whereas
akinetes are not well documented; differentiated heterocysts are mentioned as possible akinetes only in Tolypothrix
distorta (Kützing, 1843) and in Tolypothrix elenkinii
(Geitler, 1932). Moreover, the main diacritical character
of the genus Coleodesmium (Borzi ex Geitler, 1942) is the
presence of united filaments containing one to several
trichomes.
In conclusion, the complex life cycle, the isopolarity and
heteropolarity as well as the presence of akinetes are strong
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ACKNOWLEDGEMENTS
The authors thank Dr Alexandra Siakouli-Galanopoulou from
Electron Microscopy Laboratory, Department of Biology, University
of Crete, for technical assistance and Dr Sofia Papaioannou,
University of Athens, for the Latin amendment of Toxopsis
calypsus.
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