Seaweed-microbial interactions: key functions of seaweed-associated bacteria

M IN I R E V I EW

Seaweed–microbial interactions: key functions ofseaweed-associated bacteria

Ravindra Pal Singh1,2 & C.R.K. Reddy1

1Discipline of Marine Biotechnology and Ecology, CSIR-Central Salt and Marine Chemicals Research Institute, Bhavnagar, Gujarat, India; and2Department of Zoology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Ramat Aviv, Israel

Correspondence: C.R.K. Reddy, Discipline of

Marine Biotechnology and Ecology, CSIR-

Central Salt and Marine Chemicals Research

Institute, Bhavnagar, Gujarat 364002, India.

Tel.:+91 278 256 5801;

fax: +91 278 256 6970;

e-mail: [email protected]

Received 2 December 2013; revised 20

January 2014; accepted 4 February 2014.

DOI: 10.1111/1574-6941.12297

Editor: Gerard Muyzer

Keywords

bacterial communities; extracellular polymeric

substances; quorum sensing signalling

molecules; seaweeds; zoospores.

Abstract

Seaweed-associated bacteria play a crucial role in morphogenesis and growth of

seaweeds (macroalgae) in direct and/or indirect ways. Bacterial communities

belonging to the phyla Proteobacteria and Firmicutes are generally the most

abundant on seaweed surfaces. Associated bacterial communities produce plant

growth-promoting substances, quorum sensing signalling molecules, bioactive

compounds and other effective molecules that are responsible for normal mor-

phology, development and growth of seaweeds. Also, bioactive molecules of

associated bacteria determine the presence of other bacterial strains on sea-

weeds and protect the host from harmful entities present in the pelagic realm.

The ecological functions of cross-domain signalling between seaweeds and bac-

teria have been reported as liberation of carpospores in the red seaweeds and

settlement of zoospores in the green seaweeds. In the present review, the role

of extracellular polymeric substances in growth and settlement of seaweeds

spores is also highlighted. To elucidate the functional roles of associated bacte-

ria and the molecular mechanisms underlying reported ecological phenomena

in seaweeds requires a combined ecological, microbiological and biochemical

approach.

Introduction

The seaweed surface provides a suitable substratum for

the settlement of microorgansims and also secretes vari-

ous organic substances that function as nutrients for mul-

tiplication of bacteria and the formation of microbial

biofilms (Steinberg et al., 2002; Staufenberger et al., 2008;

Singh, 2013). Microbial communities living on the sea-

weed surface are highly complex, dynamic and consist of

a consortium of microorganisms including bacteria, fungi,

diatoms, protozoa, spores and larvae of marine inverte-

brates (Lachnit et al., 2009, 2011; Goecke et al., 2010;

Burke et al., 2011a, b). Among them, bacteria are ubiqui-

tous and occur either on the seaweed surface or in the

cytosol of living host cells (Herbaspirillum sp. in Caulerpa

taxifolia) and determine different stages of the life cycle

of eukaryotic organisms including macroalgae (Delbridge

et al., 2004; Burke et al., 2011a; Singh et al., 2011a, b, c).

Quorum sensing (QS) signalling molecules produced by

Gram-negative bacterial strains determine zoospores

settlement in Ulva species (Joint et al., 2002) and spores

liberation in Acrochaetium (Weinberger et al., 2007) and

Gracilaria species (Singh, 2013). Thallusin, a bacterial

metabolite, and nitrogen-fixing bacteria associated with

seaweeds have also been found to be responsible for

induction of morphogenesis and growth in marine mac-

roalgae, respectively (Chisholm et al., 1996; Matsuo et al.,

2005; Singh et al., 2011b). Macroalgae (as a host), also

known to be ecosystem engineers, play critical roles in

structuring of intertidal communities (Jones et al., 1994).

Some water-soluble monosaccharides such as rhamnose,

xylose, glucose, mannose and galactose are part of algal

polysaccharides that constitute part of the cell wall (Pop-

per et al., 2011) and the rest storage material (Lahaye &

Axelos, 1993; Michel et al., 2010a, b). These algal polysac-

charides are a potential source of carbon and energy for

numerous marine bacteria (Hehemann et al., 2012) that

produce specific molecules, which in turn facilitate sea-

weed–bacterial associations (Steinberg et al., 2002; Lach-

nit et al., 2013). Therefore, these interactions between

seaweeds and bacteria have fascinated and attracted the

attention of many researchers worldwide.

FEMS Microbiol Ecol && (2014) 1–18 ª 2014 Federation of European Microbiological Societies.Published by John Wiley & Sons Ltd. All rights reserved

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ROBI

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The main aim of the present review is to provide latest

insights into (1) seasonal diversity of the bacterial com-

munities associated with seaweeds, (2) the role of QS sig-

nalling molecules in the life cycle of macroalgae and (3)

the impact of bacterial biofilm and extracellular polymeric

substances (EPS) on thallus development and growth of

seaweeds. We also point to areas of new research aimed

at advancing the current knowledge of seaweed–bacterialassociations.

Comparison of seaweed-associated andplanktonic bacterial communities

In the recent decades, a great deal of research effort has

been directed to study the bacterial communities associ-

ated with seaweeds in order to understand the structure,

succession and dynamics of these communities in relation

to the ecology of bacterial–seaweed interactions. Studies

dealing with comprehensive assessments of total bacterial

communities on algal surfaces are relatively scarce. How-

ever, the available data, based on 16S rRNA gene

sequencing and denaturing gradient gel electrophoresis

(DGGE) fingerprinting, have revealed that algal-associated

bacterial communities differ from those of planktonic

communities (Burke et al., 2011b; Goecke et al., 2013).

Members of the Alphaproteobacteria and Gammaproteo-

bacteria have a global distribution in oceanic and coastal

waters (Venter et al., 2004; Rusch et al., 2007) while

other frequently encountered marine taxa include the

Bacteroidetes, Actinobacteria, Planctomycetes and Chloro-

flexi (Giovannoni & Stingl, 2005; Rusch et al., 2007;

Burke et al., 2011a).

In contrast, seaweed-associated bacterial communities

not only vary from species to species but also display

temporal variations (Table 1). Meusnier et al. (2001)

found that Alphaproteobacteria, Betaproteobacteria, Delta-

proteobacteria and Gammaproteobacteria and representa-

tives of the Bacteroidetes and Planctomycetes were

associated with the green alga C. taxifolia. The 16S

rRNA gene sequences retrieved from epiphytic bacteria

associated with the green alga Enteromorpha sp. showed

a predominance of Gammaproteobacteria and representa-

tives of the Bacteroidetes (Patel et al., 2003) whereas the

green alga Ulva sp. showed predominantly members of

Alphaproteobacteria and Bacteroidetes (Tait et al., 2009).

Alphaproteobacteria and Gammaproteobacteria were also

isolated from the Australian red alga Amphiroa anceps

while Bacteroidetes and Gammaproteobacteria were iso-

lated from another red alga, Corallina officinalis (Hugg-

ett et al., 2006). Longford et al. (2007) reported only

two taxa (Deltaproteobacteria and Actinobacteria) with

high abundance on Ulva australis. Furthermore, investi-

gations carried out with Ulva intestinalis (Lachnit et al.,

2011) and U. australis (Tujula et al., 2010; Burke et al.,

2011b) revealed an abundance of Alphaproteobacteria

followed by the phyla Bacteroidetes and Gammaproteo-

bacteria. Burke et al. (2011a) employed metagenomic

analysis of U. australis-associated bacterial communities

and found that it predominantly consisted of sequences

from Proteobacteria (64.0%), Bacteroidetes (27.6%) and

Planctomycetes (3.4%). Therefore, differences between

seaweed-associated communities imply selective mecha-

nisms of assembly of the bacteria. Proteobacteria and

Actinobacteria were also dominantly present on the sur-

face of the brown alga Laminaria digitata (Sala€un et al.,

2010) while Planctomycetes seemed to dominate on

Laminaria hyperborea (Bengtsson & Øvre�as, 2010; Ben-

gtsson et al., 2012) and were more predominant on

Mastocarpus stellatus and Porphyra dioica (Bondoso

et al., 2013). Additionally, Lachnit et al. (2011) studied

bacterial communities associated with Fucus vesiculosus,

Gracilaria vermiculophylla and U. intestinalis in different

seasons and reported that seaweeds harbour species-spe-

cific (7–16% of sequences of total associated bacteria)

and temporally adapted epiphytic bacterial communities

on their surfaces.

It has also been reported that different species of mar-

ine macroalgae growing in the same ecological niche

comprised specific bacterial communities on Delesseria

sanguinea, F. vesiculosus, Saccharina latissima and Ulva

compressa (Lachnit et al., 2009) and Chondrus crispus,

Fucus spiralis, M. stellatus, P. dioica, Sargassum muticum

and Ulva sp. (Bondoso et al., 2013). In contrast, macroal-

gae belonging to the same species but occurring in differ-

ent geographical locations had similar bacterial

communities to those from different species in the same

ecological niche (Lachnit et al., 2009; Nylund et al.,

2010). For example, the core genus Granulosicoccus was

frequently found associated with the red alga Delisea pul-

chra, the green alga U. australis (Longford et al., 2007),

the brown alga F. vesiculosus (Lachnit et al., 2011) and

the brown alga S. latissima (Staufenberger et al., 2008)

when isolated from the same species belonging to differ-

ent geographical locations. Further investigation on

F. vesiculosus, G. vermiculophylla and U. intestinalis

revealed that bacterial communities also differed among

replicates of the same species sampled at the same time

(Lachnit et al., 2011). In addition, other factors such as

different seasons and life cycle of the host may affect the

composition of the associated bacterial communities

(Laycock, 1974; Sakami, 1996; Singh, 2013). Staufenberger

et al. (2008) characterized bacterial communities associ-

ated with rhizoid, cauloid, meristem and phyloid parts of

the brown alga S. latissima and found that the bacterial

communities of cauloid and meristem parts were quite

similar to each other as compared with ageing phyloid

FEMS Microbiol Ecol && (2014) 1–18ª 2014 Federation of European Microbiological Societies.Published by John Wiley & Sons Ltd. All rights reserved

2 R.P. Singh & C.R.K. Reddy

Table

1.List

ofstudiespertainingto

bacterial

communitiesassociated

withthesurfaceofdifferentmacroalgae

Macroalgae

Methodology

Location

Bacteria

Referen

ce(s)

Chlorophyta

Bryopsishypnoides

EMSanFran

ciscoBay,California

Notspecified

Burr

&West(1970)

Cau

lerpaprolifera

EM,STE

TampaBay,USA

Notspecified

Daw

es&

Lohr(1978)

Ulvarigida

CUD

LasSalinas

Beach,Sp

ain

Flavobacterium

group

Bolinches

etal.(1988)

C.taxifolia

RFLP

Med

iterranean,Tahiti,Ph

ilippines,

Australia

Alpha-,Beta-,Delta-,

Gam

map

roteobacteria,

Bacteroidetes

andPlan

ctomycetes

Meu

snieret

al.(2001)

Enteromorpha

16SrRNA

gen

esequen

cing;DGGE

Wem

bury

beach,Devon,UK

Gam

map

roteobacteriaan

d

Bacteroidetes

Patelet

al.(2003)

Monostromaoxyspermum

16SrRNA

gen

esequen

cing

Okinaw

a,Ishigakian

dIriomote

Islands,

Japan

Flavobacterium

andBacteroidetes

Matsuoet

al.(2005)

U.au

stralis

16SrRNA

gen

esequen

cing;DGGE

BotanyBay

Sydney

Deltaproteobacteriaan

d

Actinobacteria

Longford

etal.(2007)

C.cupressiodes,C.Mexican

a,

C.proliferaan

dC.taxifolia

DGGE,

SEM

TampaBay,USA

Herbaspirillum

speciesbelongingto

Alphap

roteobacteria

Delbridgeet

al.(2004)

Ulvasp.

16SrRNA

gen

esequen

cing;DGGE

Wem

bury

beach,Devon,UK

Alphap

roteobacteriaan

d

Bacteroidetes

Taitet

al.(2009)

U.compressa

16SrRNA

gen

esequen

cing;DGGE

Baltic&

NorthSea,

German

yNotspecified

Lachnitet

al.(2009)

U.au

stralis

16SrRNA

gen

esequen

cing;DGGE;

CARD-FISH

SharkPo

int,Clovelly,NSW

,Australia

Alphap

roteobacteria,

Gam

map

roteobacteriaan

d

Bacteroidetes

Tujula

etal.(2010)

U.au

stralis

16SrRNA

gen

esequen

cing,

Metag

enomic

approached

,

SharkPo

int,Clovelly,NSW

,Australia

Alphap

roteobacteria,

Gam

map

roteobacteria,

Bacteroidetes

andPlan

ctomycetes

Burkeet

al.(2011a,

b)

U.intestinalis

DGGE,

16SrRNAgen

esequen

cing

Baltic,

German

yAlphap

roteobacteria,

Gam

map

roteobacteriaan

d

Bacteroidetes

Lachnitet

al.(2011)

B.hypnoides

16SrRNA

gen

esequen

cingDGGE,

CLO

;FISH

Oaxaca,

south-w

estMexicoan

d

Nayarit,central

Mexico

Bacteroidetes,

Gam

map

roteobacteria,

Alphap

roteobacteriaorTenericutes

Hollants

etal.(2011a,

b,

2013)

B.pen

nata

16SrRNA

gen

esequen

cingDGGE,

CLO

;FISH

Oaxaca,

southwestMexicoan

d

Nayarit,central

Mexico

Bacteroidetes

and

Gam

map

roteobacteria

Hollants

etal.(2011a,

b,

2013)

Ulvasp.

16SrRNA

gen

esequen

cingDGGE

Portoan

dCarrec �o

,Po

rtugal

Plan

ctomycetes

Bondoso

etal.(2013)

C.racemosa

Pyrosequen

cingan

dmetag

enomics

Med

iterraneanSeaan

d

Southwestern

Australia

Actinobacteriaan

dBacteroidetes

Aires

etal.(2013)

Phaeophyta

Laminaria

longicruris

CUD

Nova

Scotia,

Can

ada

Notspecified

Laycock

(1974)

Ascophyllum

nodosum

SEM

Nah

ant,Massachusetts

Notspecified

Cundellet

al.(1977)

Fucusvesiculosus

CUD

LasSalinas

Beach,Sp

ain

Flavobacterium

group

Bolinches

etal.(1988)


Key functions of seaweed-associated bacteria 3

Table

1.Continued

Macroalgae

Methodology

Location

Bacteria

Referen

ce(s)

F.vesiculosus

16SrRNA

gen

esequen

cing

BalticSea,

German

yAlphap

roteobacteria,

Gam

map

roteobacteriaan

d

Bacteroidetes

Lachnitet

al.(2011)

F.spiralisan

dSargassum

muticum

16SrRNA

gen

esequen

cing;DGGE

Portoan

dCarrec �o

,Po

rtugal

Plan

ctomycetes

Bondoso

etal.(2013)

Saccharinalatissim

a16SrRNA

gen

esequen

cing;DGGE

Baltic&

NorthSea,

German

yAlphap

roteobacteria,

Gam

map

roteobacteriaan

d

Bacteroidetes

Stau

fenberger

etal.(2008)

S.latissim

a16SrRNA

gen

esequen

cing;DGGE

Baltic&

NorthSea,

German

yAlpha,

Gam

map

roteobacteria

Bacteroidetes

andPlan

ctomycetes

Wiese

etal.(2009)

L.hyperborea

Pyrosequen

cing(454-seq

uen

cing)

Tekslo,Landro

andFlatevossen

Alphap

roteobacteria,

Gam

map

roteobacteriaan

d

Plan

ctomycetes

Ben

gtssonet

al.(2012)an

d

Ben

gtsson&

Øvre�as

(2010)

L.digitata

16SrRNA

gen

esequen

cing

Bloscon,Harbor,Fran

ceProteobacteriaan

dActinobacteria

Sala€ unet

al.(2010)

F.vesiculosus

16SrRNA

gen

esequen

cehomology

Baltic&

NorthSea,

German

yAlphap

roteobacteria,Bacteroidetes,

Verrucomicrobia,Cyanobacteria

andGam

map

roteobacteria

Lachnitet

al.(2011)

Rhodophyta

Amphiroaan

ceps;

Corallina

officinalis

CUD,DGGE

SharkPo

int,Clovelly,NSW

,Australia

Alpha,

Gam

map

roteobacteriaan

d

Bacteroidetes

Huggettet

al.(2006)

Delisea

pulchra

16SrRNA

gen

esequen

cing;DGGE;

Metag

enomic

approached

SharkPo

int,Clovelly,NSW

,Australia

BareIsland,Australia

Alpha-,Delta- ,

Gam

map

roteobacteria

Plan

ctomycetes

andBacteroidetes

Longford

etal.(2007)an

d

Fernan

des

etal.(2012)

Delesseriasanguinea

16SrRNA

gen

esequen

cing;DGGE

Baltic&

NorthSea,

German

yNotspecified

Lachnitet

al.(2009)

Bonnem

aisonia

asparag

oides,

Lomen

tariaclavellosa

and

Polysiphonia

stricta

TRFLP,

EPF

Skag

errak,

Swed

enNotspecified

Nylundet

al.(2010)

Gracilariaverm

iculophylla

16SrRNA

gen

esequen

cing;DGGE

Baltic,

German

yAlphap

roteobacteria,Bacteroidetes

Lachnitet

al.(2011)

Chondruscrispus,

Mastocarpus

stellatusan

dPo

rphyradioica

16SrRNA

gen

esequen

cing;DGGE

Portoan

dCarrec �o

,Po

rtugal

Plan

ctomycetes

Bondoso

etal.(2013)

CUD,culture-dep

enden

tmethods.

Microscopic

methods:

EM,electronmicroscopy;

SEM,scan

ningelectronmicroscopy;

TEM,tran

smissionelectronmicroscopy;

EPF,

epifluorescen

cemicroscopy.

Moleculartechniques:CLO

,cloning;CARD-FISH,confocallaserscan

ningmicroscopy–fluorescen

cein

situ

hybridization;RFLP,

restrictionfrag

men

tlength

polymorphism;TR

FLP,

term

inal

restriction

frag

men

tlength

polymorphism

ofDNA.



parts of the same plantlets. Different bacterial communi-

ties present on different parts of the seaweed thallus

might be explained by a lack of vascular connections in

the algal thallus leading to inefficient resource transloca-

tion (Honkanen & Jormalainen, 2005). It was also

observed that rhizoid parts associated either with other

organisms or the surrounding substrates, for example sed-

iment, caused a differentiation in bacterial communities

from other parts (Luning, 1990). The old phyloid is long

and moves (away from the meristem) in seawater by

water currents, resulting in mechanical stress, which

could lead to damage to the algal tissue (Madsen et al.,

2001; Staufenberger et al., 2008). Thereby, old phyloid

becomes more susceptible to bacterial decomposition,

offering a niche for new bacterial communities. Recently,

it was observed that metabolites, which are secreted by

microorganisms and aged plantlets, can have hydrophobic

and chaotropic activities that preferentially promote the

growth and metabolism of certain bacteria and enhance

the competitive abilities for settlement of specific micro-

organsims on the plantlet surface (Cray et al., 2013a).

Seasonal variation of bacterial communities on seaweeds

could be due to the presence of different bacterial strains

in the surrounding water or the ability of bacterial species

to attach on the plantlet surface or pre-existing bacterial

communities on the macroalgal surface. Sale (1976) and

Burke et al. (2011b) explained a competitive lottery

hypothesis which argues that ecological niches were colo-

nized randomly from a guild of species with similar eco-

logical function that coexist in that niche. The high

variability of bacterial communities between different

samples of seaweeds (Lachnit et al., 2011), even among

the same species (Burke et al., 2011b), suggested that

functional redundancy exists intraspecifically. This con-

clusion followed the redundancy hypothesis, which

presumed that more than one species is capable of

performing a specific role within an ecosystem (Naeem,

1998).

Consistently dominant bacterial communities and their

respective habitats have been termed microbial weeds

(Cray et al., 2013a). It was suggested that microbial weed

species primarily dominate communities that develop in

open habitats of microorganisms, and that such habitats

can be typified by high levels of competition for new

microorganisms, eventually attaining a climax, stationary

or closed condition. Similar to the surface fluid film of

sphagnum mosses, seaweeds provide a highly fertile, open

habitat for different marine microorganisms (Goecke

et al., 2013). Therefore, Proteobacteria and Firmicutes that

are highly prevalent on the seaweed surface (Table 1) can

presumably be considered as microbial weed species while

Pseudomonas have been shown to be archetypal weed spe-

cies in many similar or comparable habitats (Cray et al.,

2013a). In addition, homogeneous environments such as

sugar-based solutions of biotic and abiotic origin and

intracellular metabolites enforce stresses due to ionic,

osmotic, chaotropic, hydrophobic and other activities of

solutes, which also determine the dominant microbial

weed species (Brown, 1990; Hallsworth et al., 1998, 2003,

2007; Lo Nostro et al., 2005; Bhaganna et al., 2010; Chin

et al., 2010; Cray et al., 2013a). Thus, the predominance

of Proteobacteria and Firmicutes suggested that these spe-

cies (1) are exceptionally well equipped to resist the

effects of multiple stress parameters, and (2) may possess

high-efficiency energy-generation systems. Bacterial spe-

cies that have the ability to grow rapidly and have high

potential to compete with other species are implicit to

the emergence of weed species, while species that grow

relatively slowly or may require a symbiotic partner or

show obligate interactions may be eliminated during

community development in open habitats (Cray et al.,

2013a).

There are several explanations for the host specificity

and temporal patterns of bacterial communities present

on seaweeds. Epibacterial communities are harboured in

different ways (temporal and spatial distribution on the

thallus) on the host surface because of diversity in the

biochemical composition of thalli of brown, red and

green algae (Longford et al., 2007) and their metabolites

(Steinberg et al., 2002; Paul et al., 2006). For example,

the red alga D. pulchra produces an analogous molecule

of bacterial N-acyl-homoserine lactones (AHLs) known to

inhibit the signal pathways in Gram-negative bacteria that

leads to selective colonization of Gram-positive bacteria

on thalli of this species (Steinberg et al., 2002). Similarly,

the surface chemistry and outer layer composition of the

host alga may determine the composition of the epibacte-

rial community on seaweeds (Collen & Davison, 2001;

Sapp et al., 2007).

Seaweed-associated bacterial communities, particularly

endophytic, have not been well investigated despite con-

certed efforts made in this regard. Studies based on cul-

ture-dependent techniques and electron microscopy have

revealed that many coenocytic green macroalgae such as

Caulerpa, Codium, Bryopsis and Penicillus spp. bear endo-

symbiotic bacteria (Burr & West, 1970; Turner & Fried-

mann, 1974; Dawes & Lohr, 1978; Rosenberg & Paerl,

1981; Aires et al., 2013). Delbridge et al. (2004) showed

using molecular approaches that bacteria related to Herb-

aspirillum species were also found in C. taxifolia as en-

dosymbionts. Recently, Hollants et al. (2011a, b, 2013)

identified endophytic bacteria belonging to Flavobacteria-

ceae, Bacteroidetes and Phyllobacteriaceae affiliated with

Bryopsis hypnoides, and Xanthomonadaceae, Gammaprote-

obacteria, Epsilonproteobacteria and a novel Arcobacter

species associated with Bryopsis pennata. Endophytic



bacterial communities and their role in the macroalgal life

cycle remain poorly known.

Role of pre-existing associated bacterialcommunities in deciding furthercolonization by arriving bacteria

Pre-existing associated bacteria communities are well

adapted to secure their position on the seaweed surface

by preventing subsequent colonization by other bacteria

(Wenzel & M€uller, 2005). The fucoidan-degrading activity

of Verrucomicrobia, a member of Flavobacteriaceae and

Gammaproteobacteria, suggested selective colonization on

species of the brown algae Fucus (Bakunina et al., 2000,

2002; Sakai et al., 2003; Colin et al., 2006; Barbeyron

et al., 2008). Similarly, fucoidanolytic, alginolytic and

polycyclic aromatic hydrocarbon (PAH) degrading activi-

ties were reported from Sphingomonodaceae whereas bac-

teria belonging to the Bacteroidetes, Sphingobacteria and

Actinobacteria displayed agarolytic and carrageenanolytic

activities (Michel et al., 2006; Hehemann et al., 2012).

These activities enhanced their colonization on the sea-

weed surface compared with other strains (Wong et al.,

2000; Chi et al., 2012; Hehemann et al., 2012). By con-

trast, epibacterial communities present on macroalga also

determined further colonization of other marine bacteria.

Epibacteria such as the Rhizobiales, Actinobacter and Ro-

seobacter on G. vermiculophylla and D. pulchra (Longford

et al., 2007) are known for their antibacterial activity and

important for maintaining specific bacterial associations

with macroalgae (Rao et al., 2007).

Additionally, seaweed-associated bacteria are known to

produce various bioactive compounds, including haliangi-

cin, violacein, pelagiomycin A, korormicin, macrolactines

G and M, and chlorophyll d, which exhibit antifungal,

antiprotozoal, antifouling and antibiotic activity against

Gram-negative and Gram-positive bacteria and photosyn-

thetic activity (Goecke et al., 2010; and references

therein). These types of competition were essential to

maintain bacterial diversity on the seaweed surface in this

ecosystem (Spoerner et al., 2012). Volatile organic com-

pounds produced by microorganisms inhibit other associ-

ated bacteria via their chaotropicity for compounds with

a log P < 1.9 (Hallsworth et al., 2003) or as chaotropic-

ity-mediated hydrophobic stressors for compounds with a

log P > 1.9 (Bhaganna et al., 2010; Cray et al., 2013a).

Thus, inhibitory activities of associated bacterial commu-

nities against arriving free epibionts are of enormous

importance in microhabitats such as the macroalgal sur-

face. However, it will be more interesting to evaluate the

mode of action of these antibacterial compounds in detail

with respect to chemical signalling.

Key functions of seaweed-associatedbacterial communities

Activities of seaweed-associated bacterial communities have

been reported as essential for normal morphological devel-

opment and growth of the macroalgal host (Fig. 1). Nitro-

gen fixation activity of some associated bacterial strains

significantly influences growth of green and red seaweeds

(Chisholm et al., 1996; Singh et al., 2011b). Associated

(a)

(b)

(c)

(d)

Fig. 1. Bacterial role in green and red

seaweeds development: (a) promoting Ulva

zoospores settlement on bacterial EPS; (b)

reverting normal morphogenesis in axenic

culture of Ulva upon putative morphology-

inducing bacterial strains; (c) reverting wild-

type cell structure of Ulva in the presence of

appropriate bacteria; and (d) regeneration of

new buds and growth from individual fronds

of Gracilaria dura with plant hormone-

producing and nitrogen-fixing bacterial strains.

[Images modified after Singh et al.

(2011a, b, c)].



bacterial communities are also known to induce spores lib-

eration in Acrochaetium and settlement of zoospores of

Ulva species on appropriate surfaces (Joint et al., 2007;

Weinberger et al., 2007). The key functions of associated

bacterial communities are described below.

Biofilms are highly complex communities in the natural

environment that consist of many hundreds of different

species, including prokaryotic and eukaryotic microorgan-

isms. Biofilms are characterized by complex community

interactions, genetic diversity, structural heterogeneity and

an extracellular matrix of polymeric substances (Rao et al.,

2005; Joint et al., 2007). Bacteria present within the biofilm

express a unique set of genes, including those involved in

adhesion, auto-aggregation and anoxic growth (Schembri

et al., 2003). It has been established that QS is an ideal pro-

cess for maintaining the attachment of bacteria to surfaces

and the biofilm mode of growth. The role of QS in multi-

species biofilms is much less well understood as compared

with single species biofilms such as those of Pseudomonas

aeruginosa, Aeromonas hydrophila and Vibrio cholerae (Pal-

mer et al., 2003; Parsek & Greenberg, 2005; Joint et al.,

2007). Biofilms modulate the host’s interaction with vari-

ous physiocochemical conditions and may control further

foulers, consumers or pathogens (Wahl et al., 2012; and

references therein). The ecological roles of epibiotic bio-

films on marine organisms, including seaweeds, has been

summarized by Wahl et al. (2012); their roles in spores lib-

eration and zoospores settlement are also summarized

below.

Enhancement of zoospore settlement

Microbial biofilm-forming communities provide a pri-

mary substratum for settlement of different prokaryotic

and eukaryotic organisms such as phytoplankton, inter-

tidal algae and larvae. Zoospores of Ulva must find a

suitable surface to settle and adhere to in a reasonable

time in order to germinate and complete their life cycle.

Zoospore settlement is one of the important events in the

life cycle of marine organisms (Walters et al., 1999).

Studies of zoospores colonization on bacterial biofilms

reported that settlement takes place in three steps, that is

contact, temporary and irreversible adhesion (Fletcher &

Callow, 1992). In a preliminary study, Christie et al.

(1970) reported that certain enzymes such as trypsin,

pronase and amylase play an important role in the zoo-

spores attachment processes. Later, Thomas & Allsopp

(1983) showed that biofilms of Pseudomonas, Alteromonas

and Coryneform groups enhanced the number of Entero-

morpha germlings. Dillon et al. (1989) confirmed that

mixed microbial biofilms enhanced settlement of Entero-

morpha zoospores. Joint et al. (2000) found a positive

correlation between the number of Enteromorpha zoosp-

ores and uncharacterized assemblages of a number of bac-

teria formed from natural seawater and attached to glass

slides. However, image analysis of Entermorpha zoospores

settlement onto bacteria or microcolonies revealed that

zoospores attached preferentially to certain bacterial

strains present in the natural biofilms (Joint et al., 2000).

Patel et al. (2003) and Shin (2008) demonstrated that the

specific strain of bacterial biofilms and biofilm age also

enhanced zoospore settlement. Additionally, zoospores of

the Ulvaceae respond to a number of physicochemical

characteristics such as negative phototaxis, thigmotaxis,

chemotaxis, surface chemistry and wettability (Table 2;

Mieszkin et al., 2013). For example, Ederth et al. (2008)

used cationic oligopeptide self-assembled monolayers

(SAMs) for a zoospores settlement assay. The Ulva linza

zoospores interacted strongly with lysine- and arginine-

rich SAMs in comparison with acid-washed glass. Argi-

nine-rich oligopeptide SAMs were more effective in

attracting zoospores to the surface. In another study,

Krishnan et al. (2006) used a hydrophobic fluorinated

and hydrophilic polyethylene glycolated block copolymer

Table 2. Effect of different parameters on zoospores settlement in Ulvaceae

Macroalgae Findings Reference(s)

Chlorophyta

Enteromorpha sp. Adhesive strength increased zoospores settlement Finlay et al. (2002)

Enteromorpha sp. Negative phototaxis, thigmotaxis, chemotaxis, surface chemistry, wettability and

surface topography

Callow & Callow (1998) and

Callow et al. (2000, 2002)

Ulva sp. Use polydimethylsiloxane elastomer and established that topography influenced

zoospores settlement

Schumacher et al. (2008)

Ulva sp. Used hydrophobic fluorinated and hydrophilic polyethylene glycolated block

copolymer surfaces and determined role of adhesion and wettability on zoospores

settlement

Krishnan et al. (2006)

U. linza Role of surface energy for spore settlement. Used cationic oligopeptide surfaces such

as with lysine- and arginine-rich SAM

Ista et al. (2004) and Ederth

et al. (2008)

U. fasciata EPS enhanced zoospores settlement Singh et al. (2013)

U. linza Hydrophobic tridecafluoroctyl–triethoxysilane-coated surfaces Heydt et al. (2012)



to show the strength of adhesion for zoospores settlement

and concluded that surface wettability of the surface

increases settlement. By contrast, Tait et al. (2005)

showed that surface topography of bacterial biofilms was

not important for zoospores settlement. In one instance,

bacterial biofilms were exposed to either UV light or trea-

ted with 100 lg mL�1 chloramphenicol for 30 min,

which significantly reduced zoospore settlement on the

biofilms. Thus, it was inferred that surface topography

was not a dominant factor in zoospores settlement on live

biofilms (Tait et al., 2005).

Cross-domain signalling betweenbacterial AHLs and zoospores

Investigations on QS began with the finding that biolumi-

nescence of Hawaiian squid Euprymna scolopes was due to

colonization of Vibrio fischeri in the light organ of the

squid. In the light organ, V. fischeri grows at high density

continuously secreting autoinducers that induce expres-

sion of the genes required for bioluminescence (Nealson

& Hastings, 1979). This is a unique type of symbiotic

association in which squid utilized this light for predation

and the bacteria benefited by the presence of food in the

light organ (Visick et al., 2000). The expression of lucifer-

ase in V. fischeri is controlled by LuxI (an autoinducer

synthase) and LuxR (a transcriptional regulator, present

either in the cytoplasm or in the cytoplasmic membrane)

protein (Engebrecht & Silverman, 1984). In V. fischeri

Luxl is produced by 3-oxo-C6-homoserine lactone (HSL).

When the concentration of 3-oxo-C6-HSL reaches a

threshold level, it enters the bacterial cell and it binds

with cognate transcription factor LuxR to activate AHL

synthase (LuxI) to induce specific gene expression

(Waters & Bassler, 2005). Similarly, several other types of

two components (LuxI and LuxR type) signalling cascades

have been reported in diverse Gram-negative bacteria

(Williams, 2007). AHLs are now known to modulate

expression of a huge diversity of genes involved in biofilm

formation, motility, antibiotic production and the

exchange of genetic material (Fig. 2). Putative AHL-pro-

ducing bacteria play an important role in the field of

plant–bacterial interactions and cystic fibrosis (Joint et al.,

2002; Williams, 2007). AHLs are classified based on the

length of the N-linked acyl chains (4–18 carbons long)

and substitution on the C3 carbon of the N-linked acyl

chain, usually with a hydroxy or oxo group (Chhabra

et al., 1993). The key importance of AHLs in the settle-

ment of zoospores of Ulva is well established (Joint et al.,

2002) and was thoroughly reviewed by Joint et al. (2007).

Table 3 summarizes previous findings and shows how a

QS role in bacterial zoospore settlement has progressed.

The effect of AHLs is not restricted to Ulvaceae, also

being found in Gracilaria and Acrochaetium species where

it has been shown to control carpospore liberation

(Weinberger et al., 2007; Singh, 2013). Weinberger et al.

(2007) reported that C4-HSL potentially influenced

carpospore liberation capacity in Acrochaetium sp. By

contrast, Singh (2013) revealed that both C4- and

C6-HSLs contributed equally to carpospore liberation

from Gracilaria dura. In addition, increasing concentra-

tion of C4- and C6-HSLs up to 10 lg mL�1 simulta-

neously enhanced carpospore liberation. Sodium dodecyl

sulphate–polyacrylamide gel electrophoresis of the

QS signal productionPromotor…..

Transport

Diverse genePromotor…..…..

Receptor5’ 3’

QS signal molecules

mRNA

Bacterial cell

C4-HSL

C6-HSL

C10-HSL

C8-HSL

HC4-HSL

3-oxo-C12-HSLDifferent kinds of expression

Single cystocarp

Protein bands approximately 50 and 60 kDa

Putative receptor on cystocarpic cell for C4 and C6 HSLs

Spores

Fig. 2. Apparent molecular mechanism of

AHL production from seaweed-associated

bacteria and subsequent influence on

carpospore liberation from the cystocarp of

Gracilaria dura.



cystocarps of G. dura treated with C4- and C6-HSLs

revealed induction of specific polypeptide bands of

approximately 50 and 60 kDa that could be involved in

carpospore liberation (Fig. 2; Singh, 2013).

Role of marine bacteria in the life cycleof seaweeds

Associated marine bacteria have been reported to produce

plant growth regulators (PGRs) including cytokinin

(Maruyama et al., 1986, 1988, 1990; Mooney and Van,

1986), indol-3-acetic acid (IAA; Provasoli & Pintner,

1953; Singh et al., 2011b) and other PGRs and vitamins

(Provasoli & Carlucci, 1974) that also appear to be effec-

tive in regulating growth and morphogenesis in Ulva spe-

cies (Fries & Aberg, 1978; Bradley, 1991; Spoerner et al.,

2012). Provasoli (1958) reported that an axenic culture of

Ulva did not develop into normal foliose morphology

and showed a polymorphic behaviour. This polymorphic

behaviour of Ulva was entirely due to phenotypic expres-

sion (Provasoli, 1958; Bonneau, 1977). In another experi-

ment, Provasoli & Pintner (1964, 1980) found

development of abnormal plantlets having long, thin En-

teromorpha-like tubes when grown under axenic cultures.

These studies inferred that the morphology of these sea-

weeds was dependent on an association of specific groups

of bacteria and but not on the genera Caulobacter, Cy-

tophaga, Flavobacterium and Pseudomonas, which are

capable of inducing morphogenesis (Table 4). It was fur-

ther confirmed by Matsuo et al. (2003, 2005) that a spe-

cific bacterial strain, YM2-23 (NCBI accession number

MBIC04683), was responsible for the morphogenesis of

the green alga Monostroma oxyspermum. Interestingly, a

culture filtrate of the marine bacteria and extracts of the

brown and red algae were also able to restore normal

growth in M. oxyspermum (Tatewaki et al., 1983).

Finally, Matsuo et al. (2005) suggested that thallusin

was an essential factor for normal morphogenesis of

M. oxyspermum. However, the molecular mechanism of

thallusin’s action for normal growth of M. oxyspermum

is not yet clear. Marshall et al. (2006) studied zoospores

settlement and morphogenesis of U. linza and did not

find any correlation between bacterial isolates that stim-

ulated zoospores settlement and those that initiated

changes in morphology and/or growth of the cultured

alga. Bacteroidetes groups and their culture filtrates

revealed the same morphogenesis capability (Marshall

et al., 2006). Singh et al. (2011a) isolated 53 bacterial

strains from different species of Ulva and Gracilaria, and

only five were capable of inducing cell and thallus differ-

entiation and subsequent growth in Ulva fasciata cul-

tured in axenic conditions (Fig. 1). Analysis of partial

16S rRNA gene sequences from all five isolates with

morphogenesis-inducing ability led us to identify them

as Marinomonas sp. and Bacillus spp. Thus, Marshall

et al. (2006) and Singh et al. (2011a) demonstrated that

morphogenesis induction properties were not only

restricted to Bacteroidetes but also controlled by Firmi-

cutes. These studies concluded that bacteria associated

with seaweeds not only enhanced zoospores settlement

but also induced morphogenesis and growth, especially

in Ulvaceae. In addition, marine bacteria affiliated with

U. fasciata also enhanced individual cell size and struc-

ture (Singh et al., 2011a). Spoerner et al. (2012) studied

morphogenesis and growth in Ulva mutabilis and con-

cluded that Roseobacter, Sulfitobacter and Halomonas

Table 3. Role of bacterial biofilm and QS signalling molecules on zoospores settlement in green algae and spore liberation in red algae

Macroalgae Findings Reference(s)

Chlorophyta

Enteromorpha sp. Zoospores settled on submerge surface formed by bacteria Thomas & Allsopp (1983)

Enteromorpha sp. Increasing Zoospores settlement on mixed bacterial biofilm Dillon et al. (1989)

Enteromorpha sp. Positive correlation between bacteria to zoospores settlement Joint et al. (2000)

Enteromorpha sp. Vibrio anguillarum biofilms secreting AHLs Joint et al. (2002)

Enteromorpha sp.

and Ulva fasciata

Role of monospecies biofilms and effect of aged biofilm Patel et al. (2003) and

Shin (2008)

Ulva sp. Diffusion rates of AHLs, stability in seawater. Effect of AHLs with longer N-acyl

side-chains and their 3-oxo or 3-hydroxy substituent

Tait et al. (2005)

U. intestinalis Chemokinesis mechanism Wheeler et al. (2006)

Ulva sp. Role of calcium signalling Joint et al. (2007)

Ulva sp. Effect of single to polymicrobial biofilms Tait et al. (2009)

U. fasciata Zoospores released Singh et al. (2011a, b, c)

Rhodophyta

Acrochaetium sp. Spore liberation depended on AHLs Weinberger et al. (2007)

Gracilaria dura carpospores liberation through AHLs and phylogenetic identification of associated

bacteria

Singh (2013)



produced a specific regulatory factor, similar to a cytoki-

nin in higher plants, that enhanced cell division and for-

mation of an Ulva thallus. On the other hand, the genus

Maribacter produced a factor (similar to auxin) affecting

the enlargement and stretching of newly divided algal

cells (Fig. 1).

Nutrition and growth factor forseaweed growth

Mutualistic relationships between different microorgan-

isms may depend on nutrients, food transfer, oxygen

supply and settlement for survival. Seaweed–bacterialrelationships depend on the capacity of seaweeds to pro-

duce organic matter (food) and oxygen which are uti-

lized by bacteria (Goecke et al., 2010; and references

therein). Complementarily, associated bacteria provide

CO2, minerals and PGRs such as an auxin (IAA) and

cytokinin (adenine and kinetin) which enhance growth

and morphogenesis in seaweeds (Provasoli, 1958; Maruy-

ama et al., 1986, 1988; Mooney & Van, 1986). Fries

(1975) reported that bacteria living on Enteromorpha spe-

cies had the ability to convert tryptophan to IAA. Over-

production of IAA by Roseobacter associated with red

seaweed caused localized gall formation in Prionitis

lanceolata as compared with the rest of the thallus of the

same individuals (Ashen et al., 1999). Recently, Exiguo-

bacterium homiense and Bacillus spp. Were shown to

have the ability to produce IAA that determined the

number of buds and growth in G. dura (Singh et al.,

2011b). It was also reported that green, brown and red

seaweeds have endogenous capabilities to produce phyto-

hormones. For example, in Ectocarpus siliculosus (Le Bail

et al., 2010, 2011), Kappaphycus alvarezii (Prasad et al.,

2010) and Ulva species (Gupta et al., 2011; and refer-

ences therein) phytohormones determined morphogenesis

(Provasoli & Carlucci, 1974; and references therein) and

showed a relationship with bacterial auxin. The endoge-

nous auxin of E. siliculosus determined the progression of

development of branching and the reproductive phase.

Auxillary branching occurs mainly in the central part

(having round cells) of the filaments of E. siliculosus.

Subsequently, auxillary branching bodies differentiate

into erect filaments, which later carry the sporangia (Le

Bail et al., 2011). However, the relative roles of endoge-

nous and exogenous auxin in E. siliculosus have not been

determined. When the genome sequence of E. siliculosus

was compared with that of Arabidopsis, it was found that

the production of auxin in E. siliculosus followed a

Trp-dependent pathway (Le Bail et al., 2010).

Table 4. Studies on macroalgal–bacterial interaction with reference to morphogenesis

Macroalga Findings Reference(s)

Chlorophyta

Ulva lactuca Nitrate, phosphate, growth factors (IAA, adenine and

kinetin) and trace metals when cultured in synthetic

media in axenic condition

Provasoli & Pintner (1953), Provasoli &

Carlucci (1974), Maruyama et al.

(1986, 1988, 1990), Mooney & Van

(1986) and Bradley (1991)

U. lactuca Antibiotics in cultured media, forms polymorphic

morphology and determined that plant hormones

were required for morphogenesis

Provasoli (1958) and Bonneau (1977)

Enteromorpha compressa

and E. linza

Tubular-like growth Fries (1975)

U. lactuca and Monostroma

oxyspermum

Many strains of marine and associated bacteria

induced growth, such as Enteromorpha

Provasoli & Pintner (1980)

M. oxyspermum Caulobacter, Cytophaga, Flavobacterium and

Pseudomonas species were required for

morphogenesis

Provasoli et al. (1977) and Provasoli &

Pintner (1964)

E. oxyspermum Culture filtrate of bacteria and extracts of brown and

red alga were also capable of morphogenesis

Tatewaki et al. (1983) and Tatewaki &

Provasoli (1977)

U. pertusa Direct physical attachment needed for morphogenesis Nakanishi & Nishijima (1996) and

Nakanishi et al. (1999)

M. oxyspermum Specific bacterial strain YM2-23 belonging to Zobellia

sp. secreting thallusin hormone

Matsuo et al. (2003, 2005)

U. linza Bacteroidetes group bacteria and their culture filtrate

required for morphogenesis

Marshall et al. (2006)

U. fasciata Firmicutes bacteria and their culture filtrate also

induced morphogenesis

Singh et al. (2011a, b, c)

U. mutabilis Roseobacter, Sulfitobacter and Halomonas species

were capable of morphogenesis

Spoerner et al. (2012)



Seaweed-associated bacterial isolates have can fix atmo-

spheric nitrogen. For example, endosymbiotic bacteria

belonging to Agrobacterium and the Rhizobium group

have been isolated from rhizoids of the green alga C. taxi-

folia (Table 5). These strains contain the nifH gene cod-

ing for nitrogenase, which is involved in nitrogen fixation

(Chisholm et al., 1996). Azotobacter species present on

the macroalga Codium fragile ssp. tomentosoides provided

significant nitrogen fixation activity to the host (Head &

Carpenter, 1975; Fig. 3). Nitrogen fixation capabilities of

associated bacteria contribute to successful invasion of

noxious macroalgae (such as C. taxifolia or C. fragile)

into oligotrophic environments (Chisholm et al., 1996)

and induced growth in G. dura (Fig. 1d). The molecular

action of IAA production by seaweed-associated bacteria

is not well understood as compared with those bacteria

associated with higher plants as described by Pedraza

et al. (2004).

Bacteria are also involved in the production and degra-

dation of various phytohormones and biostimulants of

cell growth and development (Berland et al., 1972; Bolin-

ches et al., 1988; Meusnier et al., 2001). For example,

catalase was produced by favourable growth-promoting

Pseudoalteromonas porphyrae and had a cell growth-

regulating function on Laminaria japonica (Dimitrieva

et al., 2006). Better growth of U. fasciata in oligotrophic

and contaminated environments suggested that bacteria

played a role in the protection against toxic compounds

such as heavy metals (Goecke et al., 2010; and references

therein) and petroleum oil (Singh et al., 2011c). Recent

studies have revealed that seaweeds acquired various

genes from their associated bacteria through horizontal

gene transfer (HGT). First genome sequences of the

brown alga E. siliculosus (Cock et al., 2010) and the red

alga C. crispus (Coll�ena et al., 2013) revealed crucial HGT

from seaweed-associated bacteria. Notably, the common

ancestor of brown algae had acquired the biosynthetic

routes for D-mannitol (Michel et al., 2010a) and alginate

as well as contributing genes for hemicellulose biosynthe-

sis (Michel et al., 2010b) by HGT with an ancestral mar-

ine Actinobacterium. In the brown alga, photoassimilated

D-fructose 6-phosphate is not used to produce sucrose as

in higher plants, but it is mainly converted to D-mannitol

(Michel et al., 2010a). Similarly, the red alga also

acquired several genes from associated marine bacteria for

the biosynthetic pathway for digeneaside (mannosylgly-

cerate) from an ancestral marine bacterium Rhodothermus

marinus (Coll�ena et al., 2013). Also found was the loss of

starch metabolism in the ancestor of Stramenopiles of the

red algal endosymbiont. Similarly, the presence of auxin

Table 5. Effect of macroalgal-associated bacterial secreting compounds and biological activities on macroalgal growth

Macroalgae Growth-enhancing molecules Reference(s)

Chlorophyta

Ulva lactuca Nitrates, phosphates, growth factors and trace metals Provasoli & Pintner (1953)

U. lactuca IAA, adenine and kinetin Provasoli (1958)

Marine macroalga Cytokinin Maruyama et al. (1986, 1988)

Caulerpa taxifolia Nitrogen was supplied by endosymbiotic Agrobacterium–Rhizobium group Chisholm et al. (1996)

Codium fragile ssp. tomentosoides Nitrogenase activity of Azotobacter sp. Head & Carpenter (1975)

U. fasciata Induced cell size and growth Singh et al. (2011a, b, c)

Phaeophyta

Laminaria japonica Catalase enzyme was produced by Pseudoalteromonas porphyrae Dimitrieva et al. (2006)

Rhodophyta

Prionitis lanceolata IAA was produced by Roseobacter group Ashen et al. (1999)

Gracilaria dura Auxin (IAA) Singh et al. (2011a, b, c)

Chondracanthus chamissoi IAA, 2,4-dichlorophenoxyacetic acid and benzylaminopurine Yokoya et al. (2013)

Ferredoxin(oxidised)

Ferredoxin(reduced)

Fe reduced

Fe oxidised

MoFe(oxidised)

MoFe(reduced)

NH4 + H2

N2 8H+

e–

e– e–

e–

e–

e–

Nitrogenase I Nitrogenase II

2 ATP

2 ADP

Fig. 3. Hypothetical representation of

nitrogenase activity in nitrogen-fixing

seaweed-associated bacteria.



biosynthetic genes in the genome of E. siliculosus also

questions the origin of these algal genes through chromal-

veolate while favouring HGT from seaweed-associated

bacteria. Further experimental analyses will be needed to

confirm the biochemical function of the identified HGT

genes with regard to the chromalveolate hypothesis as

well as to establish a new hypothesis.

EPS and seaweed growth

EPS is a network of organic compounds (polysaccharides,

carbohydrate, proteins and nucleic acids) bound with

cations and/or anions, and either indirectly attached to

the cell surface or tightly associated with the cells of pro-

ducers (Costerton, 1999). Hydrophilic polymeric sub-

stances which make up the bulk of microbial EPS are

osmotropic substances that are well-hydrated and stabilize

macromolecular systems (Cray et al., 2013b). EPS thereby

helps to hold marine aggregates and keep bacterial

networks intact, eventually facilitating bacterial biofilm

formation (Flemming & Wingender, 2001). Microbial bio-

film-forming communities provide primarily a substratum

for settlement of different prokaryotic and eukaryotic

microorganisms. Recently, it was observed that bacterial

EPS enhanced the growth of marine eukaryotic communi-

ties (Mandal et al., 2011; Singh et al., 2011c, 2013). For

example, EPS was secreted by Bacillus pumilus, enhanced

the growth of the toxic dinoflagellate Amphidinium carte-

rae Hulburt 1957 and, similarly, B. pumilus and Bacillus

flexus enhanced the growth of U. fasciata. The organic

and inorganic contents of the EPS provide nutrients to

phytoplankton and seaweeds for their better survival (Nic-

hols et al., 2005; Mandal et al., 2011; Singh et al., 2011c).

EPS exhibits a polyanionic state in marine environments,

displaying a high binding affinity for cations and trace

metals. Thus, in a natural marine environment, nutrients

can interact with EPS to increase the rate of element

uptake and concentrate the dissolved organic compounds,

making them readily available for microbial growth and

the surrounding communities (Logan & Hunt, 1987;

Decho, 1990). How EPS promotes growth by capturing

nutrients from the surrounding environment is not yet

known. Singh et al. (2013) established that zoospores of

Ulva species moved to EPS and decreased in their mobility

and subsequently settled at the production site of EPS.

Similarly, Wieczorek & Todd (1997) reported increasing

settlement of ascidian Ciona intestinalis larvae with

increasing biofilm age, due to the combined effects of

active habitat selection and physical entrapment of larvae

onto the biofilm’s EPS. In addition, bacterial EPS also has

the ability to emulsify the organic pollutants and provide

healthy environments to support seaweed survival (Singh

et al., 2013). The role of EPS between rhizobacteria and

legume plants is well established, in that binding of plant

lectins to bacterial polysaccharide influences legume nodu-

lation. As an example in pea, a root-hair-expressed lectin

binding to glucomannan surface polysaccharide was pro-

duced by Rhizobium leguminosarum and promoted bacte-

rial binding to root hairs (Laus et al., 2006). Similarly, a

soybean lectin promoted the attachment of Bradyrhizobi-

um japonicum to root hairs (Lodeiro & Favelukes, 1999;

Lodeiro et al., 2000). Once bacteria accumulate on the

root hairs, these enhance delivery of Nod factors to the

root hairs and eventually enhanced nitrogen supply (Van

Rhijn et al., 1996). The seaweed–bacterial association is an

interested topic in ecological studies but we still have a

long way to go to understand the ecological significance of

EPS in the host life cycle.

Concluding remarks and futureperspective

In recent decades, improved microbiological techniques

have significantly helped to establish the phylogenetic

affiliation of epi- and endophytic bacterial communities

associated with seaweeds. Yet there is insufficient evidence

by which the functional relationship of the seaweed–bacterial interaction can be established and understood

properly. Epiphytic bacteria communities are fast coloniz-

ers of seaweed surfaces, are occasionally adaptive and are

capable of rapid metabolization of algal exudates. These

epiphytic bacteria communities play a key role in deter-

mining subsequent colonizers on the surface of seaweeds

by other planktonic fouling microorganisms. Epibacterial

community variations are due not only to changes in the

physicochemical environment but are also dependent on

preassociated bacterial and seaweed metabolites that

collectively regulate bacterial variability. Associated bacte-

ria secrete chemical compounds that act as antifouling

agents and provide protection to the host alga from path-

ogenic bacteria. Why these selective repelling activities are

maintained by bacteria during their interaction in an

ecological context is not yet fully understood. Based on

preliminary findings, it is inferred that competition for

space and nutrients between epiphytic and pelagic bacte-

ria adapts the repelling actions by associated bacteria.

Therefore, this aspect of seaweed–bacterial interactions,

including host specificity, nutrients and metabolite

exchange, needs further investigations.

Essential factors, PGRs and QS signals produced by

associated bacteria are important for normal morphology

and development of seaweeds. Thus far, only one com-

pound (thallusin) has been reported with a proven role

in morphogenesis in M. oxyspermum. It is presumed that

there could be several other compounds involved in mor-

phogenesis in other seaweeds and such compounds



should also be identified by further analysis. From an

evolutionary point of view, it can be assumed that AHL

signals ensure settlement of Ulva zoospores near a bacte-

rium that produces PGRs and essential factors to help in

morphology and development of seaweeds. Settlement of

zoospores of Ulva near AHLs and essential factors pro-

ducing bacteria predicted that these behaviours were evo-

lutionarily adapted to complete their normal life cycle.

After the association has developed, bacteria utilizing

organic matter excreted by developing plantlets might

enhance succession of the bacterial biofilm, accumulating

more AHLs. Except for roles of AHLs in zoospore settle-

ment and spore liberation, no role of AHLs in further

development of seaweeds has been observed (Twigg et al.,

2013). Additionally, there is no significant evidence that

these effects due to AHLs are density-dependent. In other

words, such a scenario does not provide a mechanism for

how these processes have evolved or are maintained in

different seaweeds. Recent studies by us found that bacte-

rial EPS has significant roles in zoospore settlement and

development of green alga (Singh et al., 2013). To

develop a better understanding of the role of EPSs in sea-

weeds, there is a need to explain what contents of bacte-

rial EPSs are required for seaweed development and how

they transfer to host alga. Thus, there remains much to

study and understand regarding the detailed mechanisms

and functional aspects of seaweed–bacterial interactions

that can be better illustrated with integration of ecologi-

cal, microbiological and biochemical studies.

Acknowledgements

The first author gratefully acknowledges the CSIR, New

Delhi (India), for award of a Senior Research Fellowship.

We also thank two anonymous reviewers for their critical

comments and suggestions on an earlier version of the

manuscript.

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Seaweed-microbial interactions: key functions of seaweed-associated bacteria

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