PRIMARY RESEARCH PAPER

Selectivity and detrimental effects of epiphyticPseudanabaena on Microcystis colonies

Ramsy Agha . Marıa del Mar Labrador .

Asuncion de los Rıos . Antonio Quesada

Received: 4 February 2016 / Revised: 5 April 2016 / Accepted: 9 April 2016 / Published online: 26 April 2016

� Springer International Publishing Switzerland 2016

Abstract The cyanobacterium Microcystis aggre-

gates into colonies with a mucilaginous sheath that

constitutes a special microhabitat for many microor-

ganisms that associate to it. Here, we examine the

notorious, yet scarcely studied case of epiphytic asso-

ciation by the cyanobacterium Pseudanabaena sp. to

colonialMicrocystis. Co-cultivation of Pseudanabaena

with different Microcystis strains evidenced strong

specificity in the interaction, with dramatically different

outcomes in each case, including (1) inability of

Pseudanabaena to access the slime of Microcystis, (2)

neutral co-existence of epiphytic Pseudanabaena and

Microcystis, and (3) rapid epiphytic proliferation of

Pseudanabaena, followed by lysis and rapid decay of

Microcystis cells. Whereas strain-specific oligopeptide

production could not explain the observed specificity,

differences in slime microstructures amongMicrocystis

strains revealed by low-temperature scanning electron

microscopy suggest that slime structural features might

initially determine the ability of Pseudanabaena to

colonizeMicrocystis, subsequently driving the outcome

of the interaction. Furthermore, even under ‘‘neutral’’

co-existence, Pseudanabaena proliferation results in an

increase indensity that leads tocolony settling, implying

potential selective losses under natural conditions. Both

the selective and antagonistic characters of the interac-

tion indicate that epiphytic Pseudanabaena have the

potential to contribute to the dynamics of strains in

naturalMicrocystis communities.

Keywords Phycosphere � Epiphytic interaction �Microcystis � Pseudanabaena

Introduction

Cyanobacteria are the dominant component of phyto-

plankton in many freshwater and marine environments

where they may form nuisance blooms. Cyanobacte-

rial blooms disrupt ecosystem functioning by increas-

ing turbidity and inducing hypolimnetic anoxia (e.g.,

Bartram & Chorus, 1999). Among bloom-forming

cyanobacteria, genusMicrocystis represents one of the

Handling editor: Judit Padisak

Electronic supplementary material The online version ofthis article (doi:10.1007/s10750-016-2773-z) contains supple-mentary material, which is available to authorized users.

R. Agha

Department of Ecosystem Research, Leibniz Institute of

Freshwater Ecology and Inland Fisheries, Berlin,

Germany

M. del Mar Labrador � A. Quesada (&)

Department of Biology, Universidad Autonoma de

Madrid, C. Darwin 2, 28049 Cantoblanco, Spain

e-mail: antonio.quesada@uam.es

A. de los Rıos

National Museum of Natural Sciences, Spanish Council

for Scientific Research (CSIC), Serrano 115,

28006 Madrid, Spain

123

Hydrobiologia (2016) 777:139–148

DOI 10.1007/s10750-016-2773-z

most successful taxa worldwide (Sivonen & Jones,

1999), whose blooms raise public health concerns due

to its ability to produce microcystins, a group of

hepatotoxic metabolites with tumor promoting activ-

ity (Falconer & Humpage, 2005). Morphologically,

genus Microcystis is characterized by forming macro-

scopic colonies of coccoid cells that are able to

regulate their buoyancy with the aid of gas vesicles.

Colonies are embedded in a mucilaginous matrix

mainly consisting of complex heteropolysaccharides

(Plude et al., 1991; Forni et al., 1997; Pereira et al.,

2009). This matrix commonly constitutes a microhab-

itat that harbors heterotrophic bacteria, archaea,

flagellates, and other microorganisms (e.g., Shia

et al., 2010; Dziallas & Grossart, 2011). Association

to the so-called phycosphere provides colonizing

microorganisms with shelter against grazing, along

with a rich source of nutrients and organic carbon

(Worm & Søndergaard, 1998; Jiang et al., 2007).

Similarly, association to Microcystis colonies grants

access to otherwise unreachable depths with increased

light and/or nutrients availability, thanks to buoyancy

regulation and daily vertical migration of colonies in

the water column. Likely due to the fact that Micro-

cystis rapidly looses colonial morphology upon labo-

ratory isolation (Reynolds et al., 1981), studies on

epiphytic interactions on Microcystis using colonial

laboratory isolates are unavailable. Instead, epiphytic

interactions with Microcystis have been generally

addressed by analyzing colonies directly from their

environment. This approach showed that microbial

epiphytic associations can exert both positive and

negative effects on Microcystis, such as mutually

beneficial nutrient exchanges (Jiang et al., 2007; Shen

et al., 2011), or, conversely, induced physiological

stress and cell lysis (Caiola et al., 1991; Gumbo &

Cloete, 2013). The notion that epiphytes can have a

profound effect on the fitness of Microcystis led to

recognize epiphytic interactions as a potential factor

driving Microcystis bloom dynamics and/or decay

(e.g., Manage et al., 2001). Beside heterotrophic

organisms, the cyanobacterium Pseudanabaena muci-

cola, formerly referred to as Phormidium mucicola

(Komarek & Kastovsky, 2003), can also often be

found associated to Microcystis colonies (Sedmak &

Kosi, 1997; Vasconcelos et al., 2011; Yarmoshenko

et al., 2013). Genus Pseudanabaena is a scarcely

studied group of filamentous, non-heterocystous

cyanobacteria characterized by simple trichomes with

a width below 4 lm that can present polar aerotopes,

complementary chromatic adaptation, gliding motil-

ity, and anaerobic N2 fixation in some strains

(Komarek, 2005; Acinas et al., 2009). The interaction

between Microcystis and Pseudanabaena embodies a

notorious and rather unique example of epiphytic

association between members of the phylum

Cyanobacteria that remains virtually unexplored in

the literature. To fill this gap, we herein examine

phycosphere colonization by Pseudanabaena sp. on

colonialMicrocystis isolates in order to shed light into

the nature of the interaction, as well as inferring its

potential contribution to the dynamics of conspecific

Microcystis strains in the field. In fact, this work was

stimulated by observations from a previous field study,

where morphologically identical Microcystis colonies

with and without the presence of epiphytic Pseudan-

abaena were found coexisting in the water column

(see Online Resource 1). Moreover, in that study,

sedimentation traps used to collected settling seston

presented abundant colony-like aggregates over the

season that were fully colonized by filaments of

Pseudanabaena (i.e., free ofMicrocystis cells). These

observations prompted the idea that epiphytic associ-

ation between Pseudanabaena and Microcystis might

operate on a strain-selective basis and can potentially

inflict selective sedimentation or lytic losses to

specific Microcystis colonies. In this regard, recent

studies have claimed that focusing on the species as

the lowest taxonomic unit may be insufficient to

understand the complex dynamics and ecology of

planktonic cyanobacteria. Instead, intraspecific poly-

morphic strains with regard, for example, to gas

vesicles properties (Beard et al., 2000), niche parti-

tioning (Johnson et al., 2006), or oligopeptide pro-

duction (Agha et al., 2014) are proposed to

encapsulate ecologically functional lineages that rep-

resent the basis on which different biological pro-

cesses operate, including loss processes (Agha &

Quesada, 2014). Considering such intraspecific

dimension, we employed different recently isolated

Microcystis spp. strains still conserving their colonial

morphology to address three main questions. First, we

seek to confirm the existence of specificity in the

interaction between Pseudanabaena and Microcystis

suggested by previous field observations or, con-

versely, show that the interaction occurs indifferently

among Microcystis strains. Second, we examine two

intraspecific polymorphic traits in Microcystis that

140 Hydrobiologia (2016) 777:139–148

123

could act as potential drivers of such specificity, in

particular, slime microstructures and cellular

oligopeptide compositions. Lastly, we seek evidence

for impacts in the buoyancy of Microcystis colonies

upon Pseudanabaena association, which may imply

Microcystis loss processes in the water column under

natural conditions.

Materials and methods

Co-cultures

In order to test for selectivity in the interaction

between Pseudanabaena and Microcystis, several

recently isolated, non-axenic strains were used

(Table 1). Strains are stored as unicellular cultures in

the collection of cyanobacterial strains of the Univer-

sidad Autonoma de Madrid. Colonies of each Micro-

cystis strain were transferred to Erlenmeyer flasks with

50-ml BG11 medium to achieve cellular concentra-

tions of 5 9 105 cells ml-1 and then inoculated with

104 filaments of Pseudanabaena strain UAM-700

(600 cells ml-1). Co-cultures were kept at 29�C under

continuous white fluorescent light at 50 lmol photons

m-2 s-1. Individual co-cultures were microscopically

inspected and Pseudanabaena proliferation within the

colonies was monitored on a daily basis using an

Olympus BH2 microscope equipped with a BH2-

RFCA epifluorescence system (Olympus). Individual

Microcystis colonies were collected before the addi-

tion of Pseudanabaena, and 4 and 8 days after

inoculation. Colonies were stored at -80�C for LT-

SEM examination.

LT-SEM examination

Slime microstructures of the different Microcystis

strains were examined by Low-Temperature Scanning

Electron Microscopy (LT-SEM). Individual Micro-

cystis colonies of each strain were prepared for

observation in a cryotransfer system (Oxford

CT1500) following De los Rıos et al. (2015). Cry-

ofractured samples were gold sputter coated in prepa-

ration unit and observed under a DSM960 Zeiss SEM

microscope at -135�C.

Analysis of oligopeptide compositions

For each strain, individual Microcystis colonies or

Pseudanabaena filament suspensions were collected

for oligopeptide extraction after Agha et al. (2013).

Oligopeptide analysis was performed by Matrix-

Assisted Laser Desorption Ionization—Time of Flight

Mass Spectrometry (MALDI-TOFMS) using a Bruker

Reflex MALDI mass spectrometer equipped with a

TOF (Time of Flight) detector. MALDI-TOF MS data

acquisition, oligopeptide identification, and spectral

data processing are described in detail elsewhere

(Agha et al., 2012).

Buoyancy experiments

In order to evaluate losses of buoyancy upon Pseu-

danabaena epiphytic growth, colonies of the Micro-

cystis strain UAM-2C1B were transferred to sterile,

25-cm high tubes filled with liquid BG11 up to a

20 cmmark. The lower 15 cm of the tubes height were

wrapped in opaque plastic to reduce light availability

in the bottom of the tubes. Three tubes were inoculated

with 104 filaments of Pseudanabaena sp. UAM-700,

whereas three tubes containing only UAM-2C1B

colonies served as controls. Culture conditions were

identical as described above. Three days after the

addition of Pseudanabaena, differences in buoyancy

among treatment and control tubes were visually

evident. However, to unequivocally show differences

in density resulting from epiphytic inhabitation,

Table 1 Cyanobacterial strains used in this study

Strain name Species Origin Date of isolation

UAM-KIN M. aeruginosa See of Galilee (Israel) Summer 2011

UAM2C1B M. novacekii Cazalegas reservoir (Central Spain) Spring 2011

UAM2C1F M. aeruginosa Cazalegas reservoir (Central Spain) Spring 2011

UAM-700 Pseudanabaena sp. Valmayor Reservoir (Central Spain) Summer 2010

All strains were isolated and maintained as monoclonal non-axenic cultures at 28�C in liquid BG11 medium under continuous white

fluorescent light of 50 lmol photons m-2 s-1. All Microcystis strains presented colonial morphology

Hydrobiologia (2016) 777:139–148 141

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colonies from both control and treatment tubes were

carefully collected and transferred to centrifuge tubes

containing a previously generated Percoll (Life

Sciences) density gradient solution (1.23% Isotonic

Percoll solution in NaCl 1.5 M, generating an isopic-

nic layer in the middle of the tubes of 1.008 g cm-3).

The tubes were then centrifuged at 5009g during

15 min and colonies migrated to their respective

isopicnic layer.

Results

Selectivity experiments with clonal isolates

Microscopic examinations of the co-cultures at 0, 4,

and 8 days after the addition of Pseudanabaena

UAM-700 revealed different outcomes for each co-

culture combination, evidencing dissimilar suscepti-

bility of Microcystis strains to epiphytic colonization:

Pseudanabaena did not access the slime of the M.

aeruginosa strain UAM-2C1F and colonies remained

unaffected during the whole culture period (Fig. 1). In

contrast, Pseudanabaena sp. filaments rapidly colo-

nized the slime of theM. novacekii strain UAM-2C1B

and proliferated within the colony slime (Fig. 2).

However, microscopic inspection did not reveal any

evident changes in the vitality (i.e., autofluorescence)

and subsequent growth of Microcystis UAM-2C1B or

Pseudanabaena. Strikingly, Pseudanabaena filaments

rapidly colonized the slime of M. aeruginosa strain

UAM-KIN and strongly proliferated, coincident with

rapid declines in the density ofMicrocystis cells in the

colonies over time. After a period of 8 days, the slime

was completely overrun by Pseudanabaena, while

Microcystis cells were almost absent (Fig. 3). The

marked differences observed among co-cultures evi-

denced strong selectivity in the interaction among both

taxa.

Exploration of factors driving selectivity

In a previous study, observations of Microcystis

colonies with epiphytic Pseudanabaena as settled

seston chronologically matched with the disappear-

ance of a particular Microcystis oligopeptide chemo-

type from the water column (Agha et al., 2014).

Therefore, we explored the possibility that the inter-

action betweenMicrocystis and Pseudanabaenamight

occur on a chemotype selective basis, i.e., driven by

differences in oligopeptide compositions among coex-

isting strains, in analogy to recently identified selec-

tive antagonistic interactions of cyanobacteria with

other organisms (Sønstebø & Rohrlack, 2011).

Oligopeptide profiles of the different strains used

were hence analyzed by MALDI-TOF MS and

compared. However, analyses did not show any

differentiating oligopeptide among strains that could

explain the observed differences in susceptibility

among Microcystis strains. Furthermore, Pseudan-

abaena sp. UAM-700 presented its own set of

intracellular oligopeptides (Table S1). Co-culture

samples presented no additional peptides compared

to those detected when analyzing the two respective

strains alone (data not shown).

In addition to oligopeptide analyses, individual

Microcystis colonies were examined by Low-Temper-

ature ScanningElectronMicroscopy (LT-SEM) inorder

to assess whether differences in slime microstructure of

colonies could determine the success ofPseudanabaena

Fig. 1 Micrographs of Microcystis aeruginosa strain

UAM2C1F before addition (a), 4 days (b), and 8 days

(c) after addition of Pseudanabaena sp. strain UAM-700. The

slime showed to be inaccessible for Pseudanabaena filaments

and colonies remained unaffected

142 Hydrobiologia (2016) 777:139–148

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to colonize the phycosphere and, eventually, influence

subsequent effects on Microcystis. Interestingly, LT-

SEM examination revealed notorious differences

among strains: Colonies of the M. aeruginosa strain

UAM-2C1F, which showed to be inaccessible to

Pseudanabaena, presented a massive, ‘‘concrete-like’’

mucilaginous slime that densely surrounded individual

cells (Fig. 4). In contrast, M. aeruginosa strain UAM-

KIN (susceptible toUAM-700 in co-culture) displayed a

slime envelope with a remarkably more diffuse struc-

ture, plenty of irregularities, and cavities. Upon colo-

nization, trichomes of Pseudanabaena could be

observed embedded within the slime (Fig. 5b, c). The

diffuse structure of the slime showed to be a genuine

feature of this strain that could be also observed in

colonies before Pseudanabaena inoculation (Fig. 5a).

Lastly, colonies of theM. novacekii strain UAM-2C1B

(seemingly unaffected by epiphytic Pseudanabaena)

showed for themost part amucilaginous envelopewith a

diffuse structure, resembling that of UAM-KIN. How-

ever, cells appeared closely packed together and, in

these areas, slime with greater consistency densely

surrounded groups of cells (Fig. 6).

Buoyancy loss assessment

Despite the possibility of neutral co-existence of

Pseudanabaena and Microcystis in co-culture (e.g.,

with strain UAM-2C1B), changes in colony density

upon Pseudanabaena colonization were evaluated.

Increases in colony density upon Pseudanabaena

colonization and growth can have ecological implica-

tions, especially if it leads to effective loss of buoyancy

and, subsequently, to selective settling of inhabited

colonies. Under natural conditions, strain-selective

sedimentation of Microcystis colonies, even when not

causing their decay, exerts a direct effect on the pelagic

composition of strains. To evaluate this, the buoyancy

ofMicrocystis strain UAM-2C1B with and without the

presence of epiphytic Pseudanabaena was compared.

Three days after the addition of Pseudanabaena sp.

UAM-700, differences in buoyancy were visually

Fig. 2 Micrographs of Microcystis aeruginosa strain

UAM2C1B before addition (a), 4 days (b), and 8 days

(c) after addition of Pseudanabaena sp. strain UAM-700.

Pseudanabaena filaments rapidly colonized the mucilage and

proliferated, but no apparent effects on Microcystis cells were

evident

Fig. 3 Micrographs of Microcystis aeruginosa strain UAM-

KIN before addition (a), 4 days (b), and 8 days (c) after additionof Pseudanabaena sp. strain UAM-700. Pseudanabaena fila-

ments rapidly accessed the slime and proliferated, while

Microcystis cell density within the colony sharply declined. At

day 8, Pseudanabaena filaments densely occupied most of the

colony and only marginal amounts ofMicrocystis cells could be

observed

Hydrobiologia (2016) 777:139–148 143

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evident. Colonies inhabited by Pseudanabaena accu-

mulated in the bottom of all replicate tubes, while

control colonies maintained positive buoyancy. The

increase in overall density upon Pseudanabaena colo-

nization was further confirmed by Percoll density

gradient centrifugation, with inhabited colonies migrat-

ing to the bottom layers (d[ 1.008 g cm-3) of the

density gradient solution (Fig. 7).

Discussion

Whereas selective losses in cyanobacterial popula-

tions have been associated to grazing (Czarnecki et al.,

2006), parasitism (Sønstebø & Rohrlack, 2011), or

programed cell death (Sigee et al., 2007), epibiotic

interactions in the phycosphere have rarely been

addressed. In the case of the epiphytic interaction

between Pseudanabaena and Microcystis, the few

existing studies dating from the 1980s already

attributed negative effects to the interaction. These

were based on observations that Pseudanabaena

grows very aggressive in culture, often destroying

Microcystis cells in short times (Gorham et al., 1982),

which led to describing epiphytic Pseudanabaena as a

parasite (Chang, 1985). However, these studies did not

explore the epiphytic interaction directly, possibly due

to the problems in maintainingMicrocystis in colonial

morphology under culture conditions (Reynolds et al.,

1981; Bolch & Blackburn, 1996). Here, the interaction

Fig. 4 LT-SEM images of Microcystis aeruginosa strain

UAM2C1F before addition (a), 4 days (b), and 8 days

(c) after addition of Pseudanabaena sp. strain UAM-700, taken

in back-scattered electron mode (a, b) and secondary electron

mode (c). Cells are embedded in a dense, massive slime which

covers the whole colony. Green arrows indicate Microcystis

cells

Fig. 5 LT-SEM images in back-scattered electron mode of

Microcystis aeruginosa strain UAM2C1B before addition (a),4 days (b), and 8 days (c) after addition of Pseudanabaena sp.

strain UAM-700. Colonies present a diffuse slime structure with

numerous cavities. Epiphytic Pseudanabaena filaments are

indicated by red arrows.Green arrows indicateMicrocystis cells

144 Hydrobiologia (2016) 777:139–148

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was addressed using colonialMicrocystis isolates. Our

findings support the antagonistic nature of interaction

reported previously and evidence detrimental effects

on Microcystis upon Pseudanabaena association.

These effects manifest either directly by cell lysis or,

more subtly, by colony sedimentation. However, the

mechanisms by which Pseudanabaena induces Mi-

crocystis lysis remain unknown and deserve further

research. Possible causes include hypersensitive

response (Sigee et al., 2007) or induced lysis (Caiola

& Pellegrini, 1984) or activation of lysogenic viral

cycles (Sedmak et al., 2008).

Besides its antagonistic nature, the interaction

displayed a high degree of specificity. High specificity

is consistent with prior field observations reporting the

co-existence of colonies with and without epiphytic

Pseudanabaena (Ilhe, 2008). However, in that study,

the occurrence of epiphytic Pseudanabaena could not

be related to morphospecies affiliation, microcystin

production, or cell quota. Here, we explored a broader

intraspecific chemical polymorphism as possible

drivers of specificity, namely, the differential produc-

tion of bioactive oligopeptides (Agha & Quesada,

2014). While oligopeptides remain largely within the

producing cells, they are also localized in the mucilage

and can therefore affect the chemical environment

around the colony (Young et al., 2005). However, our

analyses did not reveal any distinctive oligopeptide

among strains that could explain the observed speci-

ficity patterns. Instead, examination of colonies by

LT-SEM revealed marked differences in colony slime

microstructure, suggesting that the topology and

consistency of the mucilage may be an important

feature driving selectivity. The characteristics of the

substratum, roughness, and microtopological features

Fig. 6 LT-SEM images in back-scattered electron mode of

Microcystis aeruginosa strain UAM-KIN before addition (a),4 days (b), and 8 days (c) after addition of Pseudanabaena sp.

strain UAM-700. Microcystis cells (green arrows) are densely

packed together in small subcolonies surrounded by dense

slime, while slime between subcolonies displays a diffuse,

loosely bound structure

Fig. 7 Partitioning of colonies of Microcystis strain UAM-

2C1B after Percoll centrifugation. (1.23% isotonic Percoll

solution in NaCl 1.5 M, generating a 1.008 g cm-3 isopicnic

plain in the middle of the tubes). Left Control colonies in the

absence of Pseudanabaena. Right Colonies inoculated with

Pseudanabaena UAM-700

Hydrobiologia (2016) 777:139–148 145

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are determinant factors for epibiotic colonization

(Donlan, 2001; Bers & Wahl, 2004) and might

significantly affect the attachment and proliferation

of epiphytic Pseudanabaena onMicrocystis slimes. In

our study, readily accessible slimes for Pseudan-

abaena (strains UAM-KIN and UAM-2C1B) showed

a diffuse structure, dominated by loosely bound EPS,

presenting numerous cavities and irregularities.

Instead, strain UAM-2C1F presented a massive,

tightly bound slime that was inaccessible for Pseu-

danabaena. Whereas a chemical characterization of

these differences would be desirable, a rapid loss of

colonial morphology in culture prevented undertaking

this analysis. Our observations suggest that differ-

ences in structural organization of the mucilaginous

envelope in colonies of Microcystis might be crucial

for an effective association, acting as a first barrier to

epiphytic colonization by Pseudanabaena.

Our observations indicate that access to themucilage

by Pseudanabaena does not necessarily lead to direct

detrimental effects for Microcystis cells, as evidenced

by co-cultures in the case of strain UAM-2C1B.

However, we show that Pseudanabaena proliferation

causes increased density of hosting colonies, leading to

effective buoyancy loss. This phenomenon has evident

implications in the field, as epiphytic growth of

Pseudanabaena in natural systems, even when not

causing their lysis, may lead to colony sedimentation

and thereby to selective loss processes. In light of, first,

the existing selectivity in the epiphytic association and,

second, its impacts on buoyancy and viability of

inhabited colonies, epiphytic association by Pseudan-

abaena likely represents a biological process exerting

selective losses to particular Microcystis strains. Much

like in the case of other parasites (e.g., Sønstebø &

Rohrlack, 2011), strain-selective interactions poten-

tially contribute to the composition and dynamics of

coexisting strains in the water column. Hence, further

quantitative field studies addressing this interaction are

needed to uncover its relative importance to overall

Microcystis losses in the water column.

A priori, deep pelagic habitats do not seem suit-

able for Pseudanabaena spp., as access to photic depths

is strongly restricted due to its lack of gas vesicles and

inability to regulate their buoyancy. In pelagic environ-

ments, Pseudanabaena likely profits from attaching to

Microcystis colonies by accessing depths with adequate

light conditions in an otherwise aphotic environment.

Ilhe (2008) could observe that the abundance of M.

aeruginosa and M. novacekii colonies presenting epi-

phytic Pseudanabaena increased with time, being

lowest at the onset of the season, steadily increasing

during summer pelagic growth, and reaching maxima

toward autumnal sedimentation periods. These obser-

vations support the hypothesis that Pseudanabaena

associates to Microcystis colonies as a strategy to

colonize pelagic habitats in deep systems: Much like in

the case ofMicrocystis and other planktonic cyanobac-

teria (Cires et al., 2013), the recruitment phase likely

constitutes for Pseudanabaena the initial inoculum into

the water column, followed by summer pelagic growth

and maximum epiphytic development toward autumnal

sedimentation. Maximum proliferation toward the end

of the season would maximize population size to resist

benthic overwintering, which arguably represents a

stage of latency associated to substantial population

losses for Pseudanabaena. This is consistent with

previous studies showing Pseudanabaena abundances

to be lowest when re-entering the water column at the

onset of the season (Ilhe, 2008). Whereas this study

represents a first exploration of the overlooked interac-

tion between cyanobacteria of the genera Pseudan-

abaena and Microcystis, further research is definitely

needed todescribe the notorious life style and ecologyof

genus Pseudanabaena, as well as unraveling the

mechanisms underlying the selectivity and effects

resulting from its association to Microcystis, which

arguably constitutes an additional biotic interaction

contributing to the complex successional patterns of

Microcystis strains in natural populations.

Acknowledgments This article is dedicated to our colleague

Fernando Pinto, who sadly passed away during the preparation

of this manuscript and whose assistance operating the LT-SEM

made this work possible. Prof. Assaf Sukenik is acknowledged

for kindly providing bloom samples from Lake Kinneret. RA

was supported by a Postdoctoral Fellowship from the Alexander

von Humboldt Foundation during the writing process of this

manuscript. The authors also acknowledge the European Co-

Operation in Science and Technology COST Action ES1105

‘CYANOCOST’ for networking and knowledge-transfer

support. LT-SEM analyses were supported by the grant

CTM2012-3822-C02-02.

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