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Calcification of Blennothrix ganeshii in tropical streams
17
Vol. 23 No. 1 • 2013
Hidrobiológica
2013, 23 (1): 17-27
Calcification of the filamentous cyanobacterium
Blennothrix ganeshii
in calcareous tropical
streams of central Mexico region
Calcificación de la cianobacteria filamentosa
Blennothrix ganeshii
en ríos calcáreos tropicales
de la región central de México
Yenny Beltrán-Magos,
1
Javier Carmona,
2
Gloria Vilaclara
3
and Miriam Bojorge-García
2
1
Posgrado en Ciencias Biológicas, Universidad Nacional Autónoma de México, Coyoacán, Ciudad de México, D.F., 04510. México
2
Facultad de Ciencias, Departamento de Ecología y Recursos Naturales, Universidad Nacional Autónoma
de México, Circuito exterior, Ciudad Universitaria, Coyoacán, D.F., 04510. México
3
FES-Iztacala, Universidad Nacional Autónoma de México, Estado de México, 54000. México
e-mail: jcj@fciencias.unam.mx
Beltrán-Magos Y., J. Carmona, G. Vilaclara and M. Bojorge-García.
2013. Calcification of the filamentous cyanobacterium Blennothrix ganeshii in calcareous
tropical streams of central Mexico region. Hidrobiológica 23 (1): 17-27.
ABSTRACT
Geochemical, mineralogical and microbiological data from four freshwater streams in central region of Mexico indicate
the importance of Blennothrix ganeshii mats (Cyanobacteria, Oscillatoriales) in promoting the formation of calcium car-
bonate crystals. The streams were characterized by alkaline waters and relative physicochemical stability during three
seasons (cold dry, warm dry and warm rainy). Calcification took the form of a thick, dense layer of calcium carbonate
crystals surrounding the extracellular polymeric substances produced by B. ganeshii filaments, giving an appearance
of micritic tubes (structures formed by crystallization in the spaces between filaments) along the sheath surfaces. The
precipitate was analyzed using X-ray diffraction and energy dispersal X-ray spectrometry, and the calcite crystal habit
was determined. The photosynthetic activity of cyanobacterial growth and the presence of abundant extracellular
polymeric substances and epiphytic species promote the absorption of ions and mineral nucleation on the surface of
the sediment and contribute to the formation of travertine in tropical regions.
Key words: Blennothrix ganeshii, calcification, cyanobacteria, micritic tubes, tropical streams.
RESUMEN
Los datos geoquímicos, mineralógicos y microbiológicos en cuatro ríos de agua dulce de la región central de México,
ponen de manifiesto la importancia de las matas de Blennothrix ganeshii (Cianobacteria, Oscillatoriales) como promo-
toras de la formación de cristales de carbonato de calcio. Los ríos se caracterizaron por presentar aguas alcalinas y
una relativa estabilidad fisicoquímica durante tres estaciones del año (seca fría, seca templada y lluviosa templada). La
calcificación estuvo caracterizada como una densa y gruesa capa de cristales de carbonato de calcio que rodean el
mucílago extracelular producido por los filamentos de B. ganeshii, dando la apariencia de tubos micríticos (estructuras
formadas por un material cristalizado en las hendiduras existentes entre filamentos) a lo largo de la superficie de la
vaina. El precipitado fue identificado como calcita por su hábito cristalino típico y por análisis de difracción de rayos-X
y espectrometría de dispersión de energía de rayos X. La actividad fotosintética de los crecimientos de la cianobac-
teria y la presencia de abundantes sustancias poliméricas extracelulares y especies epífitas promueven la absorción
de iones y nucleación de minerales en la superficie del sedimento y contribuyen a la construcción de travertino en
corrientes de regiones tropicales.
Key words: Blennothrix ganeshii, calcificación, cianobacteria, ríos tropicales, tubos micríticos.
18
Beltrán-Magos Y. et al.
Hidrobiológica
INTRODUCTION
The deposition of calcium salts, generally called calcification, is
a common phenomenon associated with many freshwater bac-
teria (particularly cyanobacteria), small algae, fungi and bryo-
phytes that contribute to the growth of microbial biofilms and
mats (Riding, 2000; Pentecost, 2005; Turner & Jones, 2005). Micro-
bial carbonates are most common in the geologic record in seas
and lakes but at present they are also important in spring, cave
and soil environments. Minerals originated through biologically
induced mineralization (BIM) generally nucleate and grow both
extracellularly and intercellularly as a result of various metabolic
processes, such as photosynthetic uptake of CO
2
and/or HCO
3
-
by cyanobacteria, and ammonification, denitrification and sulfate
reduction by other bacteria (Riding, 2000; Couradeau et al., 2012).
Extracellular polymeric substances (EPS), widely produced by mi-
crobes, are important in providing nucleation sites and facilitating
sediment trapping (Riding, 2000). These exopolymers have diverse
compositions, depending on the biology of the individual organ-
isms and environmental conditions, but are commonly dominated
by negatively charged polysaccharides (Decho, 1990; Turner &
Jones, 2005). Although many cyanobacteria have metabolic pro-
cesses that stimulate calcium carbonate formation, favorable
environmental conditions generally appear to be necessary for
precipitation to occur (Merz-Preiß & Riding, 1999). This facultative
and environmentally dependent calcification reflects the satura-
tion state of the ambient water as well as the metabolic activities
of the cyanobacteria.
Blennothrix ganeshii Kützing ex Anagnostidis et Komárek is
often a major component of attached lotic communities in sub-
merged (littoral and benthic) habitats and occurs in calcareous
regions of Mexico (Montejano et al., 2000; Beltrán-Magos et al.,
2005). This perennial cyanobacterium grows in large mats and is
an important biotic component in its microhabitat (Carmona et al.,
2005). Studies of B. ganeshii have concerned taxonomic and eco-
logical information (Anagnostidis & Komárek, 1988; Valadez-Cruz
et al., 1996; Komárek, 1998; Watanabe & Komárek, 1989; Monteja-
no et al., 2000; Cantoral & Aboal, 2001; Beltrán-Magos et al., 2005;
Carmona et al., 2005) but studies of calcification processes are
sparse. The purpose of this study is to evaluate the mechanisms
as well as environmental and biological factors involved in B. ga-
neshii calcification in four tropical streams in central Mexico.
MATERIALS AND METHODS
Differences in water chemistry of four freshwater streams with B.
ganeshii populations (Manantiales, Tambaque, Micos and Puente
de Dios) were evaluated by sampling three times over a period
of ten months, from January to November 2004 (Fig. 1). The cli-
matic conditions include an intense summer rainy season (García,
2004). The study period covers three main seasons during the year
in Mexico for regions below 800 m.a.s.l. at latitudes of 18-22° N:
the cold dry (February), warm dry (May) and warm rainy season
(November). Various physical and chemical parameters were re-
corded in situ at each site. Water temperature, pH and specific
conductivity (standardized to 25 °C, K
25
) were measured with a
PC-18 conductivity meter (Conductronic, Puebla, Mexico); dis-
solved oxygen was measured with an oxygen meter YSI-85 (YSI
Incorporated, Ohio, USA); and current velocity and photosyntheti-
cally active radiation (PAR) were measured as close as possible
to the algal growth using a 2100 current velocity meter (Swoffer
Instruments, Washington, USA) and a LI-1000 quantum meter (LI-
COR Biosciences, Nebraska, USA) with a flat subaquatic PAR
sensor. Depth and type of substrate were also recorded for each
site. Oxygen saturation percentage was calculated from dissolved
oxygen data, taking into account altitude and water temperature
(Wetzel & Likens, 1991).
Water samples for nutrient determination were collected in
duplicate. Each sample replicate was filtered in situ with 0.22 µm
pore size membranes (Millipore, Massachusetts, USA), preserved
with a few drops of chloroform and frozen for subsequent labo-
ratory analysis with a San plus segmented-flow analyzer (Skalar
Inc., Georgia, USA), following standard titration. Soluble reactive
phosphorous (theoretically, the majority being orthophosphates,
P-PO
4
3-
), N-NO
2
-
, N-NO
3
-
and N-NH
4
+
were analyzed following
the techniques described by ASTM (1989) and APHA (1995). Wa-
ter samples for determination of anions (HCO
3
-
, CO
3
2-
, Cl
-
, SO
4
2-
),
total dissolved solids (TDS) and pH were frozen (-20 °C) and pre-
served in dark conditions, whereas samples for cations (Ca
2+
,
Mg
2+
, Na
+
, K
+
) were preserved with 40% nitric acid (down to pH 2-
3). Determination of carbonates was carried out using the titration
method, chlorides using the selective electrode method, sulfates
Figure 1. Sampling sites of B. ganeshii populations in central
Mexico. Site 1: Manantiales, Site 2: Tambaque, Site 3: Micos
and Site 4: Puente de Dios.
Central region
of Mexico
Gulf
of Mexico
100°
18°
22°
96°
N
4
3
2
1
180 km500
Calcification of Blennothrix ganeshii in tropical streams
19
Vol. 23 No. 1 • 2013
using the turbidimetric method and Na
+
and K
+
using the spectro-
photometric atomic absorption method (APHA, 1995).
Saturation of stream water with respect to calcite was cal-
culated according to the saturation index (SI), SI = pH - pH
s
, where
pH is the registered pH and pH
s
is the calculated pH in equilibrium
with CaCO
3
at the existing concentrations of Ca
2+
and HCO
3
-
(ASTM, 1989). Populations of B. ganeshii were collected for calci-
fication analysis. Maximum differences in carbonate precipitates
on cyanobacteria were observed and analyzed at the end of the
dry season (May) and at the end of the rainy season (November).
Carbonate content of samples was evaluated using the loss-on-
ignition method (550 °C for 4 h; Heiri et al., 2001; Boyle, 2004). Calci-
fication morphology and mineral composition was determined us-
ing a BX51 light microscope (Olympus Corporation, Tokyo, Japan),
a JSM-6380LV scanning electron microscope (SEM; Jeol, Tokio,
Japan) and by INCAx-sight energy-dispersive X-ray spectrosco-
py (EDXS; Oxford instruments, Oxfordshire, UK). Mineralogy was
confirmed by X-ray diffraction (XRD) using a powder diffraction
meter Broker D8-advance (CuK radiation, graphite monochroma-
tor; Bruker AXS, Wisconsin, USA). Samples were oven dried (105
°C for 24 h; Heiri et al., 2001; Boyle, 2004) and ground to a fine
powder; analysis was based on the Diffplus Bs software and the
International Centre for Diffraction Data (ICDD) database. Signifi-
cant differences (p < 0.05) in chemical composition between sites
and seasons were assessed using one-way analysis of variance
(ANOVA), followed by a Scheffé test. Relationships between cal-
cification and physicochemical parameters were evaluated with
redundancy analysis (RDA). Statistical analyses were performed
with the programs SPSS ver.12 (Levesque, 2006) and XLSTAT ver.
7.5 (XLSTAT, 2004).
RESULTS
Water conditions. Blennothrix ganeshii populations from central
Mexico are present under particular chemical and physical con-
ditions: alkaline waters (2.7-4.8 meql
-1
) dominated by SO
4
2-
/HCO
3
-
and Ca
2+
/Mg
2+
, warm temperature (23-29 °C), shallow depth (3-28
cm), moderate current velocity (13-27 cm s
-1
), high percentage of
oxygen saturation (96-107%) and different types of substrata (lime,
sand, clay, gravel, boulder or rock). Stream segments were shaded
or partly shaded (11-85 µmol photons m
-2
s
-1
; Tables 1 and 2).
Seasonal variations in temperature, pH, total alkalinities and
concentration of Ca
2+
are shown in Figs. 2A-D. The highest tem-
peratures were recorded at site 1 throughout the sampling period.
The pH of stream water varied over a small range of values (7.0-
7.9), generally with a slight increase during the warm dry season,
except for site 1. Total alkalinities showed no differences during
the seasons at site 1. At sites 2, 3 and 4, lower values were de-
tected during the warm season. Variations in Ca
2+
and ionic con-
centration between sampling dates were noted at sites 2, 3 and 4,
with higher concentrations during dry seasons and lower during
rainy seasons, showing a seasonal dilution process. Virtually no
variation in Ca
2+
ions was detected at site 1 during the sampling
period.
Figure 2A-E. Seasonal changes in temperature (A), pH (B), total alkalinity (C), Ca
2+
concentrations (D) and saturation index of
calcite (E) in the sampled streams in central Mexico.
20
Beltrán-Magos Y. et al.
Hidrobiológica
ANOVA showed significant differences in chemical com-
position between sites (F = 5.8, p = 0.001-0.02). Scheffé test
results indicated the presence of two groups (p < 0.05): one
containing sites 1, 2 and 4, and the other containing site 3. No
significant differences (p < 0.05) were found in chemical com-
position among seasons from different sampling years at all
sites.
Saturation of stream water with respect to calcite. Seasonal
changes in the saturation index of calcite (SI) are presented in
Fig. 2E. Stream water was generally supersaturated with respect
to calcite (SI > 0). SI values at site 3 were negative in the warm
rainy season when sampling took place soon after heavy rains.
Average annual supersaturation index values of calcite were be-
tween 0.25 and 0.85.
Table 1. Chemical composition measured at the study sites of the rivers of the portion central of Mexico. Values expressed in mg l
-1
, unless
otherwise stated. Values represent minimum, maximum, and average ± standard deviation (n = 5 for each site).
Site 1 Site 2 Site 3 Site 4
pH 7.2-7.8
7.5 ± 0.2
7.3-7.7
7.5 ± 0.1
7.0-7.9
7.3 ± 0.4
7.4-7.8
7.7 ± 0.2
K
25
a
[µS cm
-1
]
1703-1778
1740 ± 31
674-2360
1423 ± 643
701-948
810 ± 109
571-3000
1562 ± 937
TDS 1512-1635
1594 ± 48
464-1647
1139 ± 492
493-770
621 ± 127
508-2320
1289 ± 739
Total alkalinity
[meql
-1
]
4.7-5
4.8 ± 0.1
3.6-4.4
3.8 ± 0.4
2.4-3.3
2.7 ± 0.4
1.9-3.7
3 ± 0.9
HCO
3
-
225-305
269 ± 37
197-245
229 ± 21
148-286
196 ± 57
118-286
207 ± 64
CO
3
2-
0-32
11 ± 15
0-14
5 ± 7
0-11
3 ± 5
0-20
9 ± 9
Cl
-
12-28
16 ± 7
4-6
4 ± 1
5-10
7 ± 2
5-47
25 ± 18
SO
4
2-
735-907
859 ± 70
223-806
471 ± 234
196-361
294 ± 67
270-829
588 ± 269
Ca
2 ±
309-330
322 ± 8
121-335
231 ± 79
123-186
148 ± 26
155-313
237 ± 69
Mg
2 ±
65-73
68 ± 3
20-61
36 ± 17
22-33
26 ± 4
28-85
58 ± 25
Na
±
35-53
47 ± 7
4-15
11 ± 4
9-19
13 ± 4
22-55
41 ± 14
K
±
6-7
6 ± 0.4
0.6-2
1 ± 0.4
1-2
2 ± 0.2
2-5
4 ± 1
P-PO
4
3- a
0.01-0.03
0.02 ± 0.008
0.007-0.02
0.01 ± 0.003
0.005-0.01
0.01 ± 0.003
0.007-0.02
0.01 ± 0.005
N-NO
3
-
0.03-237
174 ± 98
69-344
178 ± 107
73-196
143 ± 49
104-195
151 ± 35
N-NO
2
-
0.001-0.006
0.004 ± 0.002
0-0.02
0.005 ± 0.008
0.001-0.02
0.007 ± 0.01
0.0005-0.007
0.003 ± 0.003
N-NH
4
±
0.03-0.5
0.1 ± 0.2
0.02-0.04
0.03 ± 0.01
0.02-0.06
0.04 ± 0.01
0.01-0.04
0.02 ± 0.008
IC
[meq l
-1
]
46-49
48 ± 1
17-44
30 ± 10
18-23
20 ± 2
23-59
36 ± 15
Ionic
dominance
SO
4
2-
> HCO
3
-
> Cl
-
> CO
3
2-
Ca
2 ±
> Mg
2 ±
> Na
±
> K
±
SO
4
2-
> HCO
3
-
> CO
3
2-
> Cl
-
Ca
2 ±
> Mg
2 ±
> Na
±
> K
±
SO
4
2-
> HCO
3
-
> CO
3
2-
> Cl
-
Ca
2 ±
> Mg
2 ±
> Na
±
> K
±
S04
2-
> HCO
3
-
> CO
3
2-
> Cl
-
Ca
2 ±
> Mg
2 ±
> Na
±
> K
±
SI 0.55-1.11
0.85 ± 0.2
0.45-0.71
0.62 ± 0.1
-0.28-0.74
0.25 ± 0.4
0.08-1.0
0.73 ± 0.4
a
Abbreviations: K
25
= specific conductance standardized at 25 °C, TDS = Total Dissolved Solids, IC = Ionic concentration, P-PO
4
3-
= Soluble Reactive Phos-
phorous, SI = Calcium Saturation Index.
Calcification of Blennothrix ganeshii in tropical streams
21
Vol. 23 No. 1 • 2013
Calcification. All studied populations were characterized by cal-
cium carbonate precipitation around the sheaths and were found
in shallow water conditions, even when substantial parts of the
mat were outside the water column, facilitating evaporation pro-
cesses and cementation of minerals. Variability in precipitation
was found between populations at different sites. Samples from
sites 1 and 4 had the highest mean carbonate precipitation re-
gardless of sampling season (31 and 36%, respectively), whereas
populations from sites 2 and 3 showed the lowest precipitation
(13 and 16%, respectively). The eigenvalues of the first two RDA
axes were high (RDA1, 0.66 and RDA2, 0.33) when compared with
subsequent axes, and they accounted for 99% of the variance in
carbonate precipitation (Fig. 3). Both axes accounted for 99% of
the variance in the relationship between carbonate precipitation
at the four sites and environmental variables. The results of the
permutation tests revealed the statistical significance (p ≤ 0.05)
of the effects of current velocity and N-NO
2
-
at sites 2 and 3; total
alkalinity, K
25
, SO
4
2-
, Ca
2+
, Mg
2+
, temperature, P-PO
4
3-
and SI at
site 1, and pH and SI at site 4. No differences in total alkalinity
among the seasons at site 1 were associated with the direction of
the distribution pattern of carbonate precipitation.
Calcification occurred as dense, thick encrustation around
the sheaths, creating micritic (metabolically induced, but envi-
ronmentally dependent) tubes typically 10-25 µm thick enclosing
B. ganeshii filaments (Figs. 4A, C). The calcareous tubes around
the filaments were separate from each other. Distal ends of the
filaments remained uncalcified, and allowing trichomes to be vis-
ible (Fig. 4E). Calcite crystals displayed a rhombohedral form with
triangular surfaces and irregular disposition (Figs. 4C-D), forming
the “gothic arch crystals” described by Rainey and Jones (2009).
Scanning-electron-microscopic examination of the initial precipi-
tate nucleation indicates the presence of calcium carbonate par-
ticles with diameters that range from several tenths of a microm-
eter to as much as 1 micrometer on the polysaccharide sheath
(Fig. 4B). EDXS of these precipitates showed they contained a
lower Ca
2+
percentage than the fully formed calcite crystals (Fig.
5A-B). Inclusions contained Ca
2+
as a major element. Around the
sheaths, the calcite coating was enlarged (up to five times the di-
ameter of the trichome) mainly by the addition of calcite crystals,
which grouped themselves in cylindrical tubes (Fig. 4F).
X-ray diffraction showed a high number of minerals (car-
bonates, silicates and oxides) deposited on B. ganeshii sheaths
(Table 3). XRD analyses indicated that calcite was the only phase
of CaCO
3
present in readily detectable amounts at all sites. Sites
1, 3 and 4 had the highest proportion of calcite (average = 95, 81
and 88%, respectively). A temporal variation in precipitate miner-
als around the sheaths was recorded in all sampling sites. A com-
parison among seasons showed that major calcification occurred
Table 2. Climatic and physical factors at the study sites on central Mexico. Values represent minimum, maximum, and average ± standard
deviation; n = 5 at each site.
Site and
location
Altitude
[m.a.s.l.]
a
Climate
b
Water
temperature
[°C]
Depth
[cm]
Current
velocity
[cm s
-1
]
PAR
c
[µmol
photons
m
-2
s
-1
]
Substrate
d
Dissolved
Oxygen
[mg l
-1
]
Oxygen
Saturation
[%]
Site 1
Manantiales
18° 55´ N
96° 00´ W
800 A w
0
(w) 26-29
29 ± 1
5-10
6 ± 2
14-26
20 ± 4
18-276
85 ± 108
L, S, C 7.2-7.8
7.5 ± 0.4
107
Site 2
Tambaque
21° 41´ N
99° 02´ W
150 (A)C(m)(w) 23-24
23 ± 1
11-60
28 ± 19
11-41
27 ± 12
13-69
34 ± 25
S, G, B, R 7.6-8.4
8.1 ± 0.4
96
Site 3
Micos
22° 05´ N
99° 09´ W
120 (A)C(m)(w) 20-27
23 ± 2
7-13
10 ± 3
3-43
21 ± 16
3-14
11 ± 5
L, S, G, B, R 8.1-8.8
8.4 ± 0.3
100
Site 4
Puente de Dios
21° 55´ N
99° 24´ W
450 (A)C(m)(w) 23-27
25 ± 2
2-7
3 ± 2
0-30
13 ± 14
4-245
73 ± 116
L, S, C, G, B, R 7-8.2
7.8 ± 0.7
104
a
Meters above sea level
b
Climate: (A)C(m)(w), Semicalid humid with abundant spring rains, mean annual precipitation 1500 mm; Aw
0
(w), Calid humid with summer rains, mean annual
precipitation 800-1000 mm; Bs
1
kw, Semicalid dry with summer rains, mean annual precipitation 503 mm (INEGI, 1985, 1992; SPP, 1981).
c
Photosynthetically active radiation
d
Substrate: B = boulder, C = clay, G = gravel, L = lime, R = rock, S = sand.
22
Beltrán-Magos Y. et al.
Hidrobiológica
in the dry season, when higher SI values were found, except in
site 3, which had a negative SI value.
DISCUSSION
The formation of travertine in tropical streams of the central re-
gion of Mexico evidently is stimulated by calcite supersaturation
in the water. In particular, B. ganeshii populations occurred under
conditions of alkaline water and the presence of abundant EPS
and ephyphitic species that promote the absorption of ions and
mineral nucleation.
High SO
4
2-
concentrations occurred in the water at all sam-
pling sites, resulting from solution of underlying gypsum deposits
(Consejo de Recursos Minerales 1992, 1993, 2000). According to
Carmona et al. (2005) sulfates are an essential component of B.
ganeshii sheaths as well as of several other cyanobacteria. Our
analysis shows that Ca
2+
, HCO
3
-
and Mg
2+
are the prevailing ions
in the studied streams, as would be expected for water in con-
tact with limestone and dolomite (Consejo de Recursos Minerales
1992, 1993, 2000; Ferrusquía-Villafranca, 1998). Na
+
, K
+
and Cl
-
are
present in low concentrations at the studied localities. Similar
water composition has been described in thermal carbonate
spring deposits in Canada and Mexican maar-crater lakes (Vila-
clara et al., 1993; Armienta et al., 2008; Rainey & Jones, 2009). The
most abundant nutrient in water from all sampling sites is nitrogen
as nitrate. Phosphate concentrations at all sites are low due to
the presence of high concentrations of Ca
2+
. This condition has
been observed in similar karstic systems elsewhere (Reddy, 1988)
due to the co-precipitation of phosphates with calcium carbonate
(Kleiner, 1990).
Mineral formation in B. ganeshii could be explained by the
reduction of CO
2
in the water due to photosynthetic activity. As
the water in which these cyanobacteria live is supersaturated
with respect to calcium carbonate, CO
2
reduction induces miner-
alization and increases in pH values, alkalinity and CO
3
2-
ion con-
centration, thus principally promoting calcite precipitation. EPS
excreted by cyanobacterial cells favor calcium carbonate en-
crustation by providing an ideal surface for the adsorption of ions
and mineral nucleation (Emeis et al., 1987; Braissant et al., 2003;
Dittrich & Sibler, 2010). EPS possess the ability to concentrate
Ca
2+
cations from solution due to the net-negative surface charge
that occurs on several cyanobacteria, including Pleurocapsa sp.,
Figure 3. Redundancy analysis (RDA) biplot of the carbonate precipitation and physicochemical parameters of four sites on riv-
ers in central Mexico. CV = Current velocity. TA = Total alkalinity.
Calcification of Blennothrix ganeshii in tropical streams
23
Vol. 23 No. 1 • 2013
Plectonema sp. and Scytonema sp. (Golubić, 1973; Merz-Preiß &
Riding, 1999; Riding, 2000; Frankel & Bazylinski, 2003; Pentecost,
2005); possibly the same phenomenon occurs in B. ganeshii. Pen-
tecost (1978) and Konhauser (2007) proposed that cyanobacterial
species that produce sheaths or EPS generally precipitate more
calcium carbonate than those species without such structures.
Merz-Preiß and Riding (1999) report that calcium carbonate
precipitation in freshwater streams becomes conspicuous where
average annual supersaturation index values exceed 0.75. How-
ever, all B. ganeshii populations showed extracellular mineral for-
mation, although relatively lower values were measured in several
seasons, particularly in the warm rainy season. Nucleation can be
disadvantaged by the mechanical removal of minerals related to
high flow velocity.
The evaporation processes occurring on exposed surfaces
of benthic mats could explain the plentiful calcification. Accord-
ing to Schneider and Le Campion-Alsumard (1999), boundary lay-
ers are the most important environments for cyanobacterial activ-
ity during the formation of carbonates.
The Blennothrix ganeshii calcification process involves two
main phases: first, the formation of a solid nucleus from dissolved
ions and second, the addition of ions to the nucleus to form crystals
of calcium carbonate. During the first phase, ions are positioned
over the sheath or epiphytic species and resist rapid dissolution
with a posterior crystalline phase. According to Merz-Preiß and
Riding (1999), Schneider and Le Campion-Alsumard (1999), Pedley
(2009), Pedley et al. (2009) and Rainey and Jones (2009) microbes
do not contribute to the establishment of elevated supersatura-
Figure 4A-F. Calcification in Blennothrix ganeshii. (A) Light-microscopy view of an encrusted filament segment; the scale bar
represents 30 µm.(B) SEM view of nucleation of spherical particles of amorphous calcium carbonate (arrow); the scale bar
represents 1 µm. (C) SEM views of without encrustation (left side) and a cluster of calcite triangles on a heavily encrusted fila-
ment (right side). The rectangular area is enlarged in (D). The scale bar represents 20 µm. (D) SEM view of lamellar triangular
plan view of calcite crystals; the scale bar represents 10 µm. (E) SEM view of cross-section of a heavily encrusted filament with
calcite crystals; the scale bar represents 20 µm. (F) SEM view of broken encrustation showing the trichome in the interior; the
scale bar represents 50 µm.
24
Beltrán-Magos Y. et al.
Hidrobiológica
tion states but passively serve as substrates upon which calcite
precipitates. Filaments of B. ganeshii were used as substrate by
several cyanobacteria, diatoms and epiphytic species of red al-
gae present throughout the entire study, including Chamaesiphon
confervicola A. Braun, Chamaecalyx swirenkoi (Sirsov) Komárek
et Anagnostidis, Stichosiphon sansibaricus (Hieronymus) Drouet
et Daily, Xenococcus bicudoi Montejano, Gold et Komárek, X.
willei Gardner, Cocconeis placentula Ehrenberg var. placentula,
Gomphonema gracile Ehrenberg, Surirella linearis W. Smith, Syn-
edra ulna (Nitzsch) Ehrenberg var. ulna, Terpsinoë musica Ehren-
berg, Audouinella meiospora (Skuja) Garbary and Compsopogon
coeruleus (C. Agardh) Montagne. Several of them have been re-
Table 3. Minerals deposited on B. ganeshii sheaths from sites 1 through 4 of rivers of the central portion of Mexico. Values are expressed
as percentage of total minerals present.
Site 1 Site 2 Site 3 Site 4
Class Group Mineral 28.v.04
a
18.xi.04 15.v.04 5.xi.04 15.v.04 6.xi.04 14.v.04 7.xi.04
carbonates calcite calcite 97 93 49 18 71 90 90 86
silicates clays palygorskite 2
silicates clays clinochlore 2
silicates clays halloysite 25
silicates clays montmorillonite 16 15
silicates feldspars albite 2 5
silicates feldspars anorthite 3
silicates feldspars microcline 8 2
silicates quartz quartz 1 27 23 11 2 5 8
silicates quartz tridymite 1
silicates quartz cristobalite 1
silicates micas muscovite 8 3 3 6
silicates micas eastonite 8
silicates amphibole richterite 12
silicates zeolites laumontite 6
oxides hematite hematite 1
a
Sampling dates. Site 1 = Manantiales, site 2 = Tambaque, site 3 = Micos, site 4: Puente de Dios.
Figure 5A-B. Electron dispersive X-ray spectroscopy of B. ganeshii filaments: (A) nucleation and (B) calcite crystals.
Calcification of Blennothrix ganeshii in tropical streams
25
Vol. 23 No. 1 • 2013
ported in similar tropical and alkaline environments (Golubic, 1973;
Prins & Elzenga, 1989; Freytet & Verrecchia, 1998; Merz-Preiß &
Riding, 1999; Beltrán-Magos et al., 2005). According to Emeis et
al. (1987), epiphytic diatoms excrete mucilage enriched in aspar-
tic acid as a response to high Ca
2+
concentrations. The mucilage
traps micrite particles that are suspended in the water, which
then act as crystal seeds for inorganic calcite precipitation. The
second phase includes the precipitation mainly of calcium car-
bonate ions and the formation of micritic tubes of calcite miner-
als that enclose sheaths. Pentecost (1978), Tavera and Komárek
(1996), Freytet and Verrecchia (1998), Schneider and Le Campion-
Alsumard (1999) and Pentecost (2005) reported this type of calci-
fication is the most common on travertine surfaces, and similar
trends have been observed in several filamentous cyanobacteria:
Tapinothrix janthina (Bornet et Flahault) Bohunická et Johansen
(H. janthina Bornet et Flahault), Lyngbya aerugineo-caerulea Go-
mont, Microcoleous vaginatus Gomont ex Gomont, Phormidium
incrustatum (Nägeli) Gomont ex Gomont, Plectonema gloeophilus
Borzi, P. gracillimum Zopf ex Hansgirg, P. phormidioides Hansgirg
ex Forti, Rivularia haematites (De Candolle) Agardh ex Bornet et
Flahault, R. varians Obenlüneschloss, Schizothrix calcicola Go-
mont and Scytonema myochrous (Dillwyn) C.A. Agardh ex Bornet
et Flahault. The extracellular biomineralization reported in these
species and the intracellular calcification in a cyanobacterium
belonging to Gloeobacterales confirm the importance of cellular
control and physicochemical parameters on the formation and
morphology of different types of travertine (Tavera & Komárek,
1996; Couradeau et al., 2012).
Cyanobacterial mats of B. ganeshii in the central region of
Mexico create ideal conditions for biologically induced mineral-
ization of calcite and have presumably played a significant role
in the development of these natural calcium carbonate environ-
ments. This facultative but environmentally dependent calcifi-
cation reflects the saturation state of the water, as well as the
oxic sediment role of B. ganeshii sheaths. The present investiga-
tion also highlights the significance of the study of physical and
chemical seasonal variations in freshwater alkaline streams and
the calcification processes of cyanobacteria so as to understand
their functioning and prevalence in tropical streams.
ACKNOWLEDGMENTS
The authors are indebted to H. Sergio Castillo Sandoval (ICMyL-
UNAM) for nutrient analysis; to Q. F. B. Nora Elia Ceniceros B., I.
Q. Alejandra Aguayo Ríos and Q. F. B. Olivia Cruz R. (IGeof-UNAM)
for carrying out mayor ion analysis; to M. C. Leticia Baños López
(IIM-UNAM) for X-ray diffraction analysis and to Dr. Rafael Quin-
tanar (FES Iztacala-UNAM) for SEM and EDXS. Special thanks are
given to Posgrado en Ciencias Biológicas, Universidad Nacional
Autónoma de México. This paper constitutes a partial fulfillment
of the Graduate Program in Biological Sciences of the National
Autonomous University of México (UNAM). Y. P. Beltrán-Magos
acknowledges the scholarship and financial support provided
by the National Council of Science and Technology (CONACyT)
(175787), and UNAM.
J. Carmona received financial support from research grant
AECI (A/010529/07 and A/016417/08).
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Recibido: 22 de abril del 2012.
Aceptado: 4 de octubre del 2012.
