encyclopedia of inland waters || bacteria, attached to surfaces

Bacteria, Attached to SurfacesJ-F Carrias and T Sime-Ngando, Universite Blaise Pascal, Clermont-Ferrand, France

ã 2009 Elsevier Inc. All rights reserved.

Introduction

Bacteria attached to surfaces can be found in alltypes of inland water habitats, including freshwaterand saline lakes, rivers, streams, ponds, ground-waters, springs, cave waters, floodplains, and alsoextreme environments such as hypersaline lakes andhot springs. Microenvironments for attached bacteriain these ecosystems consist of physical interfaces inboth the pelagic and benthic areas. They include sub-merged plants, rocks, stones, sediments, suspendedparticles and planktonic cells, and also the air–waterinterface.In general, attached bacteria form microbial consor-

tia with a diversity of other micoorganisms such asarchaea, cyanobacteria, protists (algae and protozoa),fungi aswell asmicrometazoa. Biofilms, organic aggre-gates, and microbial mats represent commons formsof these different associations of attached microbialcommunities that occur in aquatic ecosystems. Thesecomplex assemblages contain numerous species ofbacteria and play key roles in driving biogeochemicalcycles and controlling the quality of inland waters.Attached bacteria usually predominate among themicroorganisms in these consortia. They display abroad taxonomic and metabolic diversity, enablingthem to carry out a wide range of biological processes.Living inside a biofilm, a microbial mat, or an

organic aggregate undoubtedly offers bacteria numer-ous advantages. First, nutrient availability is enhancedinside the consortia and helps sustain high levelsof productivity. Second, the structure provides aprotected habitat for bacteria against harmful con-ditions such as desiccation, toxic agents, and UVradiation, as well as against various predators. Third,enhanced interactions and related processes (genetransfer, nutrient exchange, quorum sensing) are pro-moted with other bacteria, partly owing to codepen-dence on space.

Different Microhabitats for AttachedBacteria in Inland Waters

Biofilms consist of organized, complex communitiesencased in extracellular polymeric substances (EPSs)that have been synthesized by the associated bacteria.The composition of the polymeric matrix may dif-fer among and within biofilms under different lifeconditions, but exopolysaccharides are an essential

182

component of the matrix. The bacterial microcolonyis the basic living structure unit of a biofilm. In vitro,the initial unit can be produced from a single speciesor from several species. In natural conditions, micro-colonies within biofilms (Figure 1) are multispeciescommunities, and the architecture of the biofilm isthe result of complex interactions among bacteria,physicochemical factors, and the activity of protistanand metazoan grazers. Naturally occurring biofilmsare diverse and complex three-dimensional structureswith biotic and abiotic constituents. Bacteria formingbiofilms on rocks or stones (epilithic), on aquaticplants (epiphytic), and in the sediment of streams,rivers, and shallow lakes (epipelic) are known toplay an important role not only in the degradationof organic matter, but also in the degradation andtransformation of contaminants and metals. Biofilmsare also found at the air–water interface. This micro-layer is considered an extreme environment andstudies on the thriving microbial communities (e.g.,the neuston) have revealed that these biofilms presentcharacteristics similar to those found on solid surfaces.

Microbial mats are a particular type of biofilm,also known as photoautotrophic biofilms or cyanobac-terial mats. They are present in hot springs, sulfursprings, on the sediments of hypersaline lakes, and inAntarctic lakes. They aremade up of several laminated,colored layers of microbes (Figure 2) growing at thesediment–water interface of their extreme environ-ments. Light, oxygen, and hydrogen sulfide, the levelsof which vary with depth, determine the vertical dis-tribution of the associated communities. Oxygeniccyanobacteria generally dominate the surface layers,whereas anaerobic bacteria, such as anoxygenic photo-autotrophs and sulfur-reducing bacteria, are found inthe deepest zone of the mat matrix, which can rangein thickness from several millimeters to a few cen-timeters. Prokaryotic communities developing alongvertical microgradients are metabolically interdepen-dent. Unlike biofilms, naturally occurring microbialmats generally lack eukaryotic organisms, most likelybecause of the extreme conditions prevailing in theseecological niches.

Organic aggregates in pelagic environments areusually colonized by various planktonic microbes,especially bacteria. These suspended organic parti-cles and their associated microorganisms generallyoccur in the size range of <1mm to several centi-meters in plankton. The largest macroscopic aggregates

0

Yellow: Diatoms, cyanobacteria

Green: Cyanobacteria

Green: Green sulphur bacteria

Black: Permanently sulphidic zone

Purple: Purple sulphur bacteria

2

4

6

8

App

roxi

mat

e de

pth

(mm

)

Figure 2 The vertical colour banding of typical cyanobacterial

mats. Reproduced from Figure 6.3 in Fenchel T, King GM, and

Blackburn TH (2000) Bacterial Biogeochemistry – The

Ecophysiology of Mineral Cycling. Part 6: Microbial Mats andStratified Water Columns. London: Academic Press. Copyright

(1998), with permission from Elsevier.

Figure 1 Thin section of a bacterial biofilm showing distinct

microcolonies (arrowheads) scattered throughout a dominant

family of Gram-negative bacteria. The extracellularpolysaccharide that adheres to the cell is also visible as the clear

material surrounding bacteria. The inert substratum, to which the

biofilm is attached, can be seen at the bottom right-hand corner.Bar¼ 1 mm. Reproduced from Figure 1 in Beveridge TJ,

Makin SA, Kadurugamuwa JL, and Li Z (1997) Interactions

between biofilms and the environment. FEMS Microbiology

Reviews 20: 291–303. Copyright (1997), with permission fromBlackwell publishers.

Protists, Bacteria and Fungi: Planktonic and Attached _ Bacteria, Attached to Surfaces 183

(>500 mm in size), known as lake snow or river snow,are generally composed of algae (mainly diatoms),fecal pellets, bacteria, and a variety of organic detri-tus. Their abundance ranges from <1 to 50–100 l�1

and they form a significant fraction of the sinkingparticulate organic matter in pelagic environments.The abundance of microscopic aggregates (<500 mmin size), is higher, ranging from 10 to 108 l�1, depend-ing on the productivity of the ecosystem. Based on thedyes used to stain and visualize these microscopicaggregates, different types of particles have beendescribed: transparent exopolymeric particles (TEP)stainable with an acid solution of Alcian Blue, pro-teinaceous particles stainable by Coomassie BrillantBlue (Coomassie Stained Particles, CSP), and DAPIYellow Particles (DYP) stainable with 40,6-diami-dino-2-phenyl-indole (DAPI). TEP, which consistmainly of polysaccharides, are those that have beenmost closely studied (Figure 3). They play an impor-tant role in the aggregation of phytoplankton cells,primarily during bloom events, and in the formationof macroscopic aggregates. The bulk of suspendedaggregates contain attached bacteria. Their numbers

increase with particle size, and most studies havefound that attached bacteria make up about 10% ofthe total bacterial numbers within the water column.This relative abundance is certainly an underestimatebecause the bacterial counting method includes apressure filtration step that can detach some cellsfrom aggregates. Bacteria associated with aggregatesare usually larger than free-living cells in the sur-rounding water and they frequently exhibit higherenzymatic activities per cell. This implies that at-tached cells are more important in terms of activitythan in terms of biomass in the water column, relativeto free-living communities.

Epibiotic bacteria are found in association withother aquatic organisms (mainly cyanobacteria) andare generally fixed to the cell surface or live inside themucilage around the host cell. The phycosphere con-sists of the microenvironment around the surface ofhost algae or cyanobacteria. In this zone, organiccompounds, mainly polysaccharides, are at elevatedconcentrations relative to the external medium andprovide a resource to the associated bacteria insidethe mucilage. Inversely, phytoplanktonic cells benefitfrom the nutrients released by the activity of asso-ciated bacteria. Phycospheres produced by the colo-nial cyanobacteria Anabaena and Microcystis haveclose similarities with organic aggregates. Occurrenceof epibiotic bacteria is not a general feature of phyto-plankton populations because most phytoplankton

Figure 3 A Transparent Exopolymer Particle (TEP) stained with Alcian blue, a polysaccharide-specific dye, (a) visualized under direct

light and (b) DAPI-stained bacteria (small brilliant-blue spots) free-living and associated to the particle under UV light excitation. Bars

represent 10mm. Photographs from JF Carrias.

184 Protists, Bacteria and Fungi: Planktonic and Attached _ Bacteria, Attached to Surfaces

have developed defense mechanisms to prevent colo-nization by sessile microorganisms.

Adhesion of Bacteria to Surfaces, BiofilmFormation, and Structure

Because attached bacteria are important in drinkingwater distribution systems, waste treatment plants,and various industrial settings, and also because theyare implicated in numerous infections, the mechan-isms of bacterial adhesion to a substratum are wellstudied. Many of these studies concern biofilms, ty-pically focusing on one or a small number of speciesgrown under laboratory conditions. Consequently,most of the findings cannot be reliably extrapolatedto natural environments in which interactions bet-ween bacteria and surfaces involve a great diversityof colonization strategies, bacterial surface composi-tions, and solid surface characteristics.However, recentstudies have revealed that distantly related bacterialspecies show common features during the attachmentprocedure. Adhesion of a bacterium to surfaces in nat-ural environments is considered to be nonspecific, andthe analysis of a large number of both in vitro andnatural biofilms, and several studies of microbial matsand organic aggregates, clearly indicate the importanceof exopolysaccharides in the formation of the matrixaround the bacterial microcolonies. This suggests thatcommon characteristics are probably used by differentbacteria to produce the useful elements for an attachedlifestyle under different environmental conditions.Figure 4 sets out current knowledge of the differ-

ent steps involved in the colonization of surfaces bybacteria. Initially transport of free-living bacteriato the substratum is required before colonization.Three different modes of transport, which are not

mutually exclusive, are generally recognized: (1) dif-fusive transport related to the Brownian motion ofthe bacteria that can be observed under a microscope,(2) liquid flow by convection, which is a faster modeof transport of cells, and is undoubtedly an importantfactor enhancing bacterial adhesion in natural waters,and (3) active transport, whereby bacteria are motileby their flagella and encounter surfaces; this activetransport may also involve chemotaxis. The next stepis the initial attachment of the cells to the substratum(Figure 4). Physicochemical processes are involved inthe initial adhesion and are related to the surfaceproperties of either the substratum or the cells. Thisshort period has been the subject of different theoret-ical attachment models, based mainly on colloid andsurface chemistry, designed to gain insight into thecontrol mechanisms of bacterial adhesion to surfaces.During the attachment process, bacteria move on thesubstratum by twitching motility and can detach andbe free-living plankton, or synthesize EPSs to forma cell monolayer and develop in a biofilm. At thispoint, the attachment becomes irreversible. Geneticstudies of the adhesion of Pseudomonas aeruginosahave revealed that cells change phenotype when theybecome sessile and that this change is achievedthrough gene expression.

The development of the biofilm involves thegrowth and multiplication of attached cells withinbacterial colonies and also the colonization of newbacterial species. Structural features on the cell sur-face (fimbriae, flagella) are involved in the firmattachment of bacteria, while EPSs, which consist ofpolysaccharides, extracellular DNA, and proteins,play an important role in the coaggregation of cells.During its maturation, the biofilm is greatly influ-enced by abiotic factors, which partly determine itsstructure. A mature biofilm consists of a complex

Timesscale

Seconds

Mins-hrs

Hrs-days

Days-weeks

Biofilm detachment

Erosion or sloughing of cells

Solute transfer to or from bulk liquid

Active bioconversion and or biodegration

Horizontal gene transfer

Spreading of biomass

Chemo taxis

Motility

Mature biofilm

Structure, surface shape,spatial distribution of biomass

Hydrodynamics and/ormechanical stress

Active biofilm

Cell monolayer

Initial attachment Formation of new biofilm

Figure 4 Schematic representation of steps involved in the formation of a biofilm and main factors influencing its evolution.Reproduced from Figure 1 in Singh R, Paul D, and Jain RK (2006) Biofilms: Implications in bioremediation. Trends in Microbiology

14: 389–397. Copyright (2006), with permission from Elsevier.



three-dimensional organization with multispeciesmicrocolonies embedded in a matrix with intercon-necting channels and water flows. Bacteria can move,allowing chemotaxis processes considered to be animportant feature of bacterial biodegradation withinthe biofilm. In addition, the proximity of bacterialcells enhances the possibility of gene transfer and ofcoordinated behavior due to signaling molecules(quorum sensing). All of these biological factors areimportant in the maturation and maintenance of bio-films. Finally, other microorganisms (algae, fungi,protozoa) colonize the surface to form amixedmaturebiofilm as found in many freshwater ecosystems,

Secretion

Losrecru

of

Matrix

DOC

Photosynthesis

Light

Spadhe

mecha

Inorganicnutrients

Populat

Quorumsensing

Solid subs

Water flow

Gelatinous matrix

Bacteria Algae Pro

Organicsolute

Externalwatermedium

Figure 5 Biological interactions within a mixed biofilm: a small-sca

stones in rivers and the littoral zone of lakes, the thickness of the biofilm

Different microorganisms within the biofilm (not draw to scale) are indi

typically highly structured, occurring as columns with interstitial spacacross the surface of the biofilm. Solid arrows simply relate organism

(2005) Freshwater Microbiology – Biodiversity and Dynamic Interactio

Wiley. Copyright (2005), with permission from Wiley.

in which complex interactions take place (Figure 5).Some of the bacteria regularly detach from the consor-tia allowing the colonization of new niches to createanother biofilm.

Composition and Community Structureof Attached Bacteria

Early investigations of the composition and diversityof bacterial communities were based on microscopicobservations and the isolation and cultivation of cells.Detailed studies of the morphology, physiology, and

Genetransfer

s anditment

cells

ecificsionnisms

ion increase

Current

tratum

Conditioning film

Fungitozoa

Ingestion

le microbial ecosystem found as a surface layer on rocks and

ranges from a fewmicrometers (mm) to several millimetres (mm).

cated by the symbols. In a mature biofilm, the gelatinous matrix is

es through which water can percolate. Water flow also occurss to particular activities. Reproduced from figure 1.5 in Sigee DC

ns of Microorganisms in the Aquatic Environments. Chichester:

Table 1 Some commonly occurring prokaryotes which are known to attach to surfaces in aquatic environments

Method of analysis Examples and modifications Use(s) Refs

Direct observation Epifluorecent microscopy, Confocal Scanning Laser

Microscopy

Morphological observation,

enumeration

1

Molecular methods In situ hybridization, comparative sequence analysis Community analysis, taxonomy 1, 3

Fluorescent labeling Fluorescence in situ hybridization in combination with

microautoradiography and microsensors

Characterization of biofilms

communities

2, 4

Detection of gene

expression

Reporter protein assay, in vivo expression technology (IVET),

recombination based IVET (RIVET)

Investigation of gene activity 2, 5

PCR Direct in situ PCR Characterization of genetic and

phylogenetic properties

6

Reproduced from Table 1 in Singh R, Paul D, and Jain RK (2006) Biofilms: Implications in bioremediation. Trends in Microbiology 14: 389–397. Copyright

(2006), with permission from Elsevier.

Sources

1. Wimpenny J, Manz W, and Szewzyk U (2000) Heterogeneity in biofilms. FEMS Microbiology Reviews 24: 661–667.

2. Aoi Y (2002) In situ identification of microorganisms in biofilm communities. Journal of Bioscience Bioengineering 94: 552–556.

3. Woese C and Fox GE (1977) Phylogenic structure of the prokaryotic domain: The primary kingdoms. Proceedings of the National Academy of Sciences

of the United States of America 74: 5088–5090.

4. Ito T, Nielsen JL, Okabe S, Watanabe Y, and Nielsen PH (2002) Phylogenic identification and substrate uptake patterns of sulphate reducing bacteria

inhabiting an oxicanoxic sewer biofilm determined by combining microautoradiography and fluorescent in situ hybridization. Applied and Environmental

Microbiology 68: 356–364.

5. Chalfie M, Tu Y, Euskirchen G, Ward WW, and Prasher DC (1994) Green fluorescent protein as marker for gene expression. Science 263: 802–805.

6. Tani K, Kurokawa K, and Nasu M (1998) Development of a direct in situ PCR method for detection of specific bacteria in natural environments. Applied

and Environmental Microbiology 61: 4074–4082.


genetics of the isolated microorganisms have beencarried out in the laboratory from pure cultures.These bacteria often belong to the most commonlyoccurring prokaryotes in the field, and some of themare known to attach to surfaces or to form microbialmats (Table 1). A large number of bacterial speciesfrom microbial mats have been characterized, partic-ularly those of hot springs. These species includedifferent anoxygenic phototrophs (purple sulfur bac-teria, purple nonsulfur bacteria, green sulfur bacteria,green nonsulfur bacteria), some chemoautotrophs(colorless sulfur bacteria), sulfur and sulfate-reducingbacteria, and a variety of thermophilic and halophilicprokaryotes. Some widespread chemoorganotrophicspecies with specialized appendages or stalks used forattachment have also been examined in detail bystandard methods.However, most of the microorganisms have not

been cultivated. Less than 1% of bacterial cells fromnatural environments are amenable to standard cul-ture techniques. To open this ‘black box,’ new meth-ods to access the composition and structure of aquaticbacterial communities have been developed, mostlyusing molecular tools. Molecular techniques (Table 2)such as terminal restriction fragment length polymor-phism (T-RFLP) and denaturing gradient gel electro-phoresis (DGGE) of PCR amplified 16S rDNA havebeen applied for prokaryotic diversity and commu-nity structure analysis. In addition, fluorescent in situhybridization (FISH), using 16S rRNA-targeted oli-gonucleotide probes combined with epifluorescence

microscopy has become a useful tool for analyzingbacterial composition and counting bacterial groups inaquatic environments. These molecular approacheswere first applied to whole bacterial communities inaquatic habitats and have revealed an enormous diver-sity of microbial species in the natural environment.These studies also led to the discovery of novel unculti-vated prokaryotes.

More recently, more and more studies have focusedon the differences between attached and free-livingbacteria and on the community structure of biofilm-associated bacteria in aquatic environments. Specificprobes have been applied to the study of the microbialcolonization of biofilms and micro- and macroaggre-gates in both marine and freshwater ecosystems.These studies are few in number, but have providedimportant information on the diversity and commu-nity structure of bacteria attached to surfaces.

The majority of the studies using molecular toolsfor attached bacteria analysis in freshwater lakes andrivers indicate high abundances of the beta-subgroupof the Proteobacteria on suspended particles andin biofilms. In situ examination of the bacterial com-munity on river and lake snow aggregates as well asin river biofilms indicates that beta-proteobacteriaoften constitute 20–50% of total bacterial abun-dance. Alpha-proteobacteria and Bacteroidetes (formerCytophaga/Flavobacteria) represent a large propor-tion of the total bacteria on suspended particles,but generally do not exceed the abundance of beta-proteobacteria. Because members of the Bacteroidetes

Table 2 Techniques commonly used for analysis of attached bacterial communities

Genus Class Main characteristics Habitat/microhabitat

Caulobacter Alphaproteobacteria Gram-, rod-shaped, flagella, stalk, aerobic,

chimioorganotroph

Widespread in freshwater habitats

Hyphomicrobium Alphaproteobacteria Gram-, rod-shaped, hyphae formation,

budding reproduction, aerobic,

chimioorganotroph

Widespread in freshwater and marine

habitats

Sphaerotilus Betaproteobacteria Gram-, filamentous, microaerophilic,chimioorganotroph

Flowing freshwater, sewage,wastewater treatment plants

Pseudomonas Gammaproteobacteria Gram-, rod-shaped, flagella, aerobic,

chimioorganotroph

Widespread: freshwater and saline

habitats, soil, plant and animal tissue

Beggiatoa Gammaproteobacteria Gram-, filamentous, microaerophilic,chimiolithotroph, gliding motility on

surfaces

Microbial mats in sediments offreshwater and saline lakes,

sulphur-springs, marine habitats

Thiocapsa Gammaproteobacteria Gram-, spherical shape, anaerobic,

photolithotrophic, purple sulphur bacteria

Microbial mats in sediments of

freshwater and saline lakes,sulphur-springs, marine habitats

Chlorobium Bacteroidetes Gram-, chains of spherical cells, anaerobic,

photolithotrophic, green sulfur bacteria

Microbial mats in sulphide-rich

habitats (hot springs, mud,sediments)

Chloroflexus Chloroflexi Gram-, filamentous, anaerobic,

photoorganotroph, green nonsulfur bacteria

Microbial mats in sediment of

freshwater and saline lakes, hot

springs, marine habitatsHalobacterium Halobacteria Gram-, Rod-shaped, aerobic,

chimioorganotroph, can use

photosynthesis to survive in anaerobic

conditions

Microbial mats in hypersaline

environments

Halococcus Halobacteria Gram-, cocci, aerobic, chimioorganotroph, Microbial mats in hypersaline inland

waters


are specialized in the degradation of complex macro-molecules, it is generally assumed that a high propor-tion of this group is related to the presence ofrefractory materials associated with suspended aggre-gates or biofilms. The community structure of free-living bacteria in lakes and rivers differs from that ofbacteria on suspended particles and on sediments andusually shows a lower proportion of beta-proteobac-teria. In contrast to freshwater, no clear dominance ofany single bacterial group on particles has beenreported in marine systems. Temporal changes in thebacterial composition of freshwater aggregates havealso been observed, suggesting a succession of differ-ent functional groups that may operate sequentially inthe degradation of organic matter. However, the fac-tors influencing the spatiotemporal changes inattached-bacterial community composition remainlargely unknown. The use of group-specific probesgives low phylogenetic resolution and provides onlya rough description of the bacterial composition.Investigations using a large number of probes at thegenus and species-specific levels in combination withother methods (Table 2), such as the use of microsen-sors, confocal scanning laser microscopy (CSLM), andmicroautoradiography, will make it possible to charac-terize in detail the composition, physiological activities,

and spatial organization of biofilm and aggregate-associated bacteria. The distribution of unculturednitrifying bacteria of the genera Nitrosospira andNitrospira in biofilms was recently studied under labo-ratory conditions using FISH in combination withCSLM and microsensor measurements. This approachwas also used in freshwater sediments, suggesting thatthese genera occur frequently in the environment, butthat they inhabit microniches different from thoseoccupied by the other well-known genera of nitrifyingbacteria; Nitrosomonas and Nitrobacter.

Control of Attached Bacteria

Attached bacteria in different microhabitats (e.g.,biofilms and suspended particles) are assumed to begoverned by the same controlling factors as free-livingbacteria. The major factors controlling abundance,biomass, and growth of attached bacteria includethe availability of organic substrates and nutrients,grazing pressure mainly from protozoa, and lysisfrom viral communities. However, the relative impor-tance of these factors is not well known for attachedbacteria compared with free-living bacteria. In addi-tion, physicochemical factors such as the light environ-ment (UV radiation) and oxygen concentrations can


affect bacterial biomass in aquatic systems, but theirimpact on attached cells remains largely unknown.Most studies of grazer-down control of attached

bacterial communities have focused on predation byprotozoa. Some amoeboid, flagellated, and ciliatedprotozoa are able to graze bacteria on surfaces andit has been shown that they can reduce bacterialdensity substantially and affect biofilm architecture.Based on studies conducted in marine systems, itis likely that a large fraction of bacteria attached topelagic aggregates are grazed by protozoa in inlandwaters. Accordingly, these microbial grazers arepotentially important players in the recycling of nutri-ents and the maintenance of community growthin attached bacterial microhabitats. Their presencecan enhance the degradation of organic matter,although the relative contribution of bacteria–grazerinteractions in attached communities (compared withfree-living ones) remains largely unknown. Somestudies have indicated that the aggregation of free-living bacteria and the formation of bacterial coloniesincrease when protozoan bacterivory is intense. Thisis considered as one of the known defense mechan-isms of aquatic bacteria against predation. Predatorsmight thus be viewed as stimulators of the early for-mation of aggregates and biofilms, although thishypothesis needs to be specifically addressed andexamined in the context of diverse populations ofbacteria and protozoa coexisting in aquatic systems.The impact of viral lysis on bacterial mortality was

recently studied in lakes, but differences betweenattached and free-living bacteria were not investigated.The relative contributions of viral lysis and protozoangrazing to attached-bacterial mortality are stillunknown. In freshwater benthic habitats (sedimentsand biofilms), a few studies have shown that viralinfectivity is less pronounced than in the upper watercolumn, suggesting that bacterial cells in colonies areless exposed not only to grazing but also to viralattacks. However, this inference is still essentially spec-ulation and needs to be confirmed by further work.The positive effect of nutrients on the development

of attached bacteria is confirmed by the generalincrease in the abundance of these populations withthe increasing trophic status of aquatic systems. Thebiomass and activities of bacteria attached to biofilmsor aggregates increase with primary production anddissolved organic carbon (DOC). The concentrationof DOC in the water column is often considereda limiting factor for bacterial growth in biofilms.However, in a given ecosystem, the productivity ofattached bacteria compared with that of free-livingcells and their spatial and temporal distributions arelargely unknown, although it is widely accepted thatin shallow lakes and in running waters, most bacterial

productivity is associated with surfaces. The impor-tance of resources, predation, and viral lysis in theregulation of attached bacteria in inland waters thusremain to be evaluated, in the context of within andbetween-system variability.

Activities of Attached Bacteria and theirPotential Contributions to theDegradation of Organic Matter

Bacterial communities of inland waters play key rolesin the degradation of organic matter and the cyclingof important elements such as carbon, nitrogen, phos-phorus, and sulfur. Because of the ubiquity of bacteriain natural ecosystems, it is likely that attached bacte-ria are involved in the overall bacterial processes.Although the role and metabolic diversity of bacteriaassociated with microbial mats in hypersaline aquaticenvironments is reasonably well documented, ourknowledge of the functional significance of bacteriaassociated with biofilms and suspended particles infreshwater remains incomplete. We will now focus onthe role of attached bacteria in the degradation oforganic matter and the related biogeochemical cyclesin freshwaters, and refer readers to the list of furtherreadings for information on the diversity and func-tional role of attached bacteria in microbial mats,hypersaline lakes, and other extreme environments,such as hot springs.

Bacteria on Freshwater Organic Aggregates

One of themost important roles of bacteria in freshwa-ter environments arises from their ability to respire andrecycle organic matter into nutrients and CO2. In addi-tion, bacteria utilize both dissolved and particulateorganic matter (DOM and POM) from external andinternal sources. Surfaces are often considered as amajor site for the respiration and recycling and henceaerobic heterotrophic processes on freshwater aggre-gates and biofilms are important. These processesinclude POM solubilization, DOM hydrolysis anduptake, respiration and biomass production withinthe organic matrix, and both substrate uptake fromand release into the surrounding waters. Attachedbacteria are often metabolically more active than free-living bacteria and exhibit higher cell-specific uptakerates of free amino acids and monosaccharides, andhigher ectoenzymatic hydrolysis rates of aminopepti-dase, phosphatase, and glucosidase. Proteins are gener-ally decomposedmore rapidly than polysaccharides byattached bacteria. This differential use has importantimplications for the decomposition of the organic mat-ter in microhabitats (e.g., macroaggregates, biofilms)dominated by diatoms, because dissolution of diatom

Oligo-/monomers

Oligo-/monomersPolymers

Attachedbacteria

Detritovores

Mineralization(C, N, P, Fe, Si)

Freebacteria

Polymers


silica frustules takes place onlywhen they are colonizedby bacteria, which are probably able to hydrolyze thecell-wall proteins inside the frustules.In freshwater diatom-derived aggregates, bacterial

respiration is often positively correlated with POMcontent, and increases with the age of the aggregates.This contrasts with bacterial growth efficiency, whichdecreases from fresh to aged aggregates. The turnoverof the aggregate-associated POM based on respira-tion losses is substantially higher than if calculatedonly on the basis of bacterial biomass production.Particle-associated bacterial biomass production onaggregates is usually <30% of total bacterial produc-tion in freshwaters. However, total bacterial produc-tion is much higher in samples with suspendedaggregates than in samples with no aggregates, andattached bacterial production generally increaseswith aggregate size. In addition, the rates of POMsolubilization and net release of labile substrates intothe surrounding waters generally exceed the carbondemand of aggregate-associated bacteria. As a resultof high hydrolytic activities of attached bacteria, andhigh metabolic activities of bacterial grazers, concen-trations of organicmatter (dissolved free and combinedamino acids, DOC, etc.) and of inorganic nutrients(e.g., C, N, P, Fe, and Si) are significantly higher in thepore water of the aggregates than in the surroundingwaters. Besides recycling within aggregates, the highlyenriched labile DOM and nutrients may also be partlyreleased into the surrounding waters. Hence, the activ-ity of attached bacteria in pelagic systems can have twomajor consequences: (1) stimulation of the growthrates of free-living microorganisms in the vicinity ofaggregates, resulting in the occurrence of hot spotmicroenvironments with high productive potentialsand (2) regulation of the degradation rates of aggre-gates and of the vertical sinking flux of organic matterfrom the water column to the sediments (Figure 6).

POM

Big detritovores

Sinking flux

Phytoplankton

Phytodetritus

Figure 6 Loss processes and microbial decomposition

pathways of macroscopic-organic aggregates. POM: Particulateorganic matter. Reproduced from Figure 5 in Simon M, Grossart

H-P, Schweitzer B, and Ploug H (2002) Microbial ecology of

organic aggregates in aquatic ecosystems. Aquatic Microbial

Ecology 28: 175–211. Copyright (2002), with permission fromInter-Research.

Bacteria in Freshwater Biofilms

Few studies have focused on the measurements of theactivities of bacteria associated with biofilms. Mostof our knowledge on this topic comes from studies instreams where contributions by biofilm bacteria, dueto their high abundance and diverse metabolic cap-abilities, dominate the assimilation and flux of DOM,the dominant form of organic carbon in aquatic sys-tems. Bacterial activity in biofilms is dependent on thesource, quality, type, and quantity of DOM and nutri-ents. Different types of DOM and inorganic nutrientsinduce various responses in biofilm bacterial commu-nities, but this is obvious only when different taxa aredistinguished and seasonal changes are taken intoaccount. Attached bacterial activity is more directly

dependent on DOM, primarily the low molecularweight (LMW) organic substrates, than to inorganicnutrients, known to influence epilithic bacteria indi-rectly via the algal compartment. Bacterial vs. algalcompetition for inorganic nutrients seems to be lessmarked in biofilms than in plankton. Algal-releasedDOM, which comprises largely labile, easily utiliz-able LMW compounds, is thus considered as the mainsource of carbon for heterotrophic bacteria in biofilms.In the absence of significant amounts of toxins, bacte-rial productivity, biomass, biovolume, and enzymepro-duction are enhanced by algal exudates. The mutualreliance of bacteria and algae in biofilms is furtherfacilitated by their close spatial proximity. However,in canopy-covered streams, algal-derived resources areoften insufficient for the metabolic needs of biofilmbacteria, partly because of reduced light penetration.There is also evidence of photoinhibition of algae inbiofilm communities in open streams. In addition toalgal exudates, allochthonous DOM such as leachatefrom litter fall is an important source of organic mate-rial in many streams, but not all bacteria species inbiofilms can utilize leaf leachate as a carbon source.Phenolic compounds contained within leachates canactually inhibit the growth of some bacteria.


Conclusion and Future Research

Bacterial studies in inland waters have focusedmainlyon the pelagic compartment, andmost often only free-living bacteria are considered. It is only relativelyrecently that bacteria attached to surfaces have beentaken into account, even though the attached lifes-tyle is considered common in aquatic microbial ecol-ogy. However, the mechanisms that control theadhesion and detachment of natural aquatic bacteriaand their ecological significance need to be furtherinvestigated, including the relative importance of thephysical, chemical, and biological factors involved.Compared with the free-living mode, our knowledgeof the diversity and the functional roles of attachedbacteria in inlandwaters is scant, partly because of thebroad diversity of colonizable habitats for thesemicroorganisms. More sampling efforts are needed,combined with the use of modern molecular andanalytic tools, to gain insight into species-specificcommunity composition, seasonal and spatial organi-zation, and the related metabolisms of the microbialassociations. Top-down and bottom-up control ofattached populations and interactions (competition)within these populations and with other biologicalcompartments are still imperfectly understood. Inthis regard, the relative importance of grazing andviral lysis should be considered in future studies.Attached bacteria need to be considered as a bio-logical compartment in ecosystem studies, becausetheir potential role in processing organic matter andcycling carbon is probably important, possibly moreimportant than free-living bacteria. This need is espe-cially important for lotic ecosystems and shallowlakes where attached bacteria largely dominate thetotal bacterial community in terms of both biomassand productivity. In addition, the importance ofattached bacteria in degrading pollutants must beaddressed by in situ investigations, because mostof the related findings derive from experimentsconducted in controlled laboratory conditions. Over-all, there is shortage of in situ studies on the roleof attached bacteria in cycling or sequesteringmajor elements (C, N, P) in inland waters and sedi-ments, and also in the degradation of pollutants. Thisknowledge is critical for better management of inlandwater ecosystems, the major resource for life onthe earth.

Glossary

Alpha-proteobacteria – A class of proteobacteria, amajor group of bacteria.

Autoroph – Organism that produces complex organiccompounds from simple inorganic molecules

and an external source of energy, such as light(photoautotroph) or chemical reactions of inorgan-ic compounds (chemoautotroph).

Bacteroidetes – A phylum of bacteria that are widelydistributed in the environment.

Beta-proteobacteria – A class of proteobacteria, amajor group of bacteria.

Bottom-up control – A bottom-up control of a popu-lation is established when an increase of food sup-ply increases the abundance of this population.

Chemoorganotroph – Organism that obtains carbonfrom organic compounds and energy from the oxi-dation of inorganic compounds.

Chemotaxis – Phenomenon in which organisms directtheir movements according to chemical substancesin their environment.

Consortium (Plural: consortia) – A microbial consor-tium is a group of different species of microorgan-isms with different metabolic activities. Theorganisms interacts each other and all benefit fromthe activities of others.

Cytophaga – A genus of rod-shape, gram-negativebacteria.

Epibiotic – Able to grow or to attach on the surface ofa living organism.

Epilithic – Able to grow or to attach on the surface ofstones and rocks.

Epipelic – Able to grow or to attach on the surface ofmud, sediment.

Epiphytic – Able to grow or to attach on the surfaceof plants or algae.

Fimbriae – Thin projection of the bacterial cell that isused to adhere to surfaces.

Flagella – Projection from the cell body forming afilament surrounded by the plasma membrane andusually used for locomotion.

Flavobacteria – Bacteria of the genus Flavobac-terium. They are found in soil and freshwaterenvironments.

Hot-spot – In aquatic microbial ecology, a hot-spot isa microenvironment of high microbial abundance,diversity and activities, usually formed from sus-pended detritus.

Mucilage – Various gelatinous substances (generallypolysaccharides) secreted by plants and micro-organisms (especially algae) into their surroundingenvironments.


Neuston – Organisms living at the air–water interface.

Phycosphere – Zone surrounding an algal cell withinwhich others microorganisms are influenced byalgal activity.

Quorum sensing – Ability of bacteria to communicateand coordinate their behavior via signaling molecule.

Raptorial – Organism adapted for seizing prey.

Top-down control – A top-down control of a popula-tion is demonstrated by an increase in the abun-dance of this population when the pressure of itspredators is reduced.

See also: Bacteria, Bacterioplankton; Bacteria,Distribution and Community Structure; Chemical Fluxesand Dynamics in River and Stream Ecosystems;Cyanobacteria; Microbial Food Webs; Natural OrganicMatter; Protists; Viruses.

Further Reading

An YH and Friedman R (2000)Handbook of Bacterial Adhesion –Principles,Methods, andApplications.Totowa: Humana Press Inc.

Berkeley RCW, Lynch JM, Melling J, Rutter PR, and Vincent B

(1980) Microbial Adhesion to Surfaces. London: Ellis Horwood

limited.

Cohen Y and Rosenberg E (1989) Microbial Mats: PhysiologicalEcology of Benthic Microbial Communities. ASM editions:

Washington, DC.

Fletcher M (1996) Bacterial Adhesion – Molecular and EcologicalDiversity. New York: Wiley-Liss, Inc.

Jefferson KK (2004) What drives bacteria to produce a biofilm?

FEMS Microbiology Letters 236: 161–173.

Krumbein WE, Paterson DM, and Zavarzin GA (2003) Fossil andRecent Biofilms: A Natural History of Life on Earth. KluwerAcademic Publishers: 481p.

Lawrence JR and Neu TR (2003) Microscale analyses of the for-

mation and nature of microbial biofilm communities in river

systems. Reviews in Environmental Science and Bio/Technology2: 85–97.

Marshall K (2006) Planktonic versus sessile life of prokaryotes. In:

Dworkin M, Falkow S, Rosenberg E, et al. (eds.) The Prokar-yotes – A Handbook on the Biology of Bacteria. Volume 2:Ecophysiology and Biochemistry, 3rd edn., pp. 3–15. New

York: Springer.

Ofek I, Sharon N, and Abraham SN (2006) Bacterial adhesion. In:

Dworkin M, Falkow S, Rosenberg E, et al. (eds.) The Prokar-yotes – A Handbook on the Biology of Bacteria. Volume 2:Ecophysiology and Biochemistry, 3rd edn., pp. 3–15. New

York: Springer.Rajbir S, Debarati P, and Rakesh KJ (2006) Biofilms: Implications

in bioremediation. Trends in Microbiology 14: 389–397.

Sigee DC (2005) Freshwater Microbiology – Biodiversity and Dy-namic Interactions of Microorganisms in the Aquatic Environ-ments. Chichester: Wiley.

Simon M, Grossart HP, Schweitzer B, and Ploug H (2002) Micro-

bial ecology of organic aggregates in aquatic ecosystems.AquaticMicrobial Ecology 28: 175–211.

Van Loosdrecht MCM, Lyklema J, Norde W, and Zehnder AJB

(1990) Influence of interfaces on microbial activity. Microbiolo-gical Reviews 54: 75–87.

Ward DM, Ferris MJ, Nold SC, and Bateson M (1998) A natural

view of microbial biodiversity within hot spring cyanobacterial

mat communities.Microbiology and Molecular Biology Reviews62: 1353–1370.

Wimpenny J, Manz W, and Szewzik U (2000) Heterogeneity in

biofilms. FEMS Microbiology Reviews 24: 661–671.

Relevant Websites

http://www.erc.montana.edu/.http://www.biofilmsonline.com.

http://www.microbeworld.org/.

http://www.erc.montana.edu/

http://www.biofilmsonline.com

http://www.microbeworld.org/

encyclopedia of inland waters || bacteria, attached to surfaces

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