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Int. J. Mol. Sci. 2013, 14(8), 15838-15859; doi:10.3390/ijms140815838
Published: 30 July 2013
Abstract: The role of bacterial surface components in combination with bacterial functional signals in the process of biofilm formation has been increasingly studied in recent years. Plants support a diverse array of bacteria on or in their roots, transport vessels, stems, and leaves. These plant-associated bacteria have important effects on plant health and productivity. Biofilm formation on plants is associated with symbiotic and pathogenic responses, but how plants regulate such associations is unclear. Certain bacteria in biofilm matrices have been found to induce plant growth and to protect plants from phytopathogens (a process termed biocontrol), whereas others are involved in pathogenesis. In this review, we systematically describe the various components and mechanisms involved in bacterial biofilm formation and attachment to plant surfaces and the relationships of these mechanisms to bacterial activity and survival.
Biofilms are bacterial communities in which cells are embedded in a matrix of extracellular polymeric compounds attached to a surface . Living in biofilms helps protect bacteria from deleterious conditions  and the formation of biofilms appears to be an important factor in the disease cycle of bacterial pathogens in both animals and plants.
Bacterial surface components and extracellular compounds [primarily flagella, lipopolysaccharides (LPSs), and exopolysaccharides (EPSs)], in combination with environmental and quorum-sensing signals, are crucial for autoaggregation and biofilm development in most bacterial species studied to date [3,4]. In the generally accepted model of biofilm formation, environmental signals trigger the process, and flagella are required for the biofilm community to approach and move across the surface. The initial steps of attachment are mediated by outer membrane proteins (e.g., calcium-binding proteins), pili, or LPSs. After the formation of microcolonies, the production of quorum-sensing signals is required for the formation of a mature biofilm . EPSs provide the architectural form of biofilms and stabilize their 3-dimensional structure. Biofilms are often permeated by channels that act as a circulatory system, allowing the bacteria to exchange water, nutrients, enzymes, and signals, dispose of potentially toxic metabolites, and display enhanced metabolic cooperativity [4,6]. The dispersal of biofilms allows bacteria to colonize other surfaces or substrates, thus completing a sequential developmental process.
The composition of biofilms varies depending on the system. The major components are typically water and the bacterial cells, followed by the EPSs of the matrix , which provides (i) a physical barrier against the diffusion of antibiotics, defense substances, or other important compounds from the host; and (ii) protection against environmental stress factors, such as UV radiation, pH changes, osmotic stress, and desiccation [8,9]. In Agrobacterium tumefaciens, a plant pathogen that persists as surface-associated populations on plants or soil particles, cellulose overproduction resulted in increased biofilm formation on roots . Minor biofilm components include macromolecules such as proteins, DNA, and various lysis products , which affect the overall properties of the biofilm.
Bacterial biofilms are widely distributed and play important roles in many environments. The environments occupied by soil bacteria range from rhizospheres rich in nutrients and root exudates to bulk soil deficient in nitrogen, phosphates, water, and other nutrients. The size of bacterial aggregates varies from small to large as a function of the nutrient availability at a given site . A hypothetical model of various 3-dimensional shapes of root-biofilm structures determined by nutrient availability has been presented .
Many species of beneficial soil bacteria, including rhizobia, form microcolonies or biofilms when they colonize roots. We recently summarized data on surface attachment and/or biofilm formation by rhizobacteria . Biofilm development also contributes to the virulence of phytopathogenic bacteria through various mechanisms, including blockage of xylem vessels, increased resistance to plant antimicrobial compounds, and/or enhanced colonization of specific habitats . The processes of autoaggregation and biofilm development are relevant to both bacterial survival and host plant colonization (Figure 1). A variety of environmental, genetic, and structural factors affect bacterial adhesion, cell-cell interactions, and plant colonization, and ultimately plant-bacterial interactions in general. In this article, we review recent findings on the mechanisms involved in attachment, cell aggregation, and biofilm formation on plant surfaces by bacteria.
2. Cell–Cell Adhesive Interactions: Bacterial Autoaggregation
Bacteria were studied for many years as isolated cell entities. However, like many microorganisms, they have a strong tendency to congregate or aggregate. A common phenotypic manifestation of this behavior is autoaggregation, which is based on adhesive interactions among bacteria. Autoaggregation can be visualized macroscopically by the typical clumping or “fluffing” of cells in liquid cultures, followed by sedimentation of the clumps under static conditions [3,14,15].
Unfavorable growth conditions or low metabolic activity have been found to induce aggregative behavior in bacteria that normally grow in a dispersed, non-aggregated manner. In this context, autoaggregation may reflect a survival strategy that is triggered under hostile environmental conditions [16–19].
The autoaggregative characteristic of bacteria has important implications for the production of bacteria-based inoculants for agriculture. Bacterial aggregates can be produced on a large scale and then separated more easily from the culture medium as compared with dispersed bacteria. The biomass of aggregated bacterial cells in bioreactors remains more constant, and the survival of such bacteria during the inoculant storage period is enhanced, in comparison with non-aggregated cells [17,18,20,21].
2.1. Surface Factors Involved in Bacterial Autoaggregation
Because of their strategic location on the cell surface, LPSs, outer membrane proteins, and proteinaceous structures such as pili have been reported to affect adhesion among bacteria and consequently the autoaggregation phenotype.
LPS is an important surface structural component of Gram-negative bacteria and covers ~75% of the surface area of the outer membrane. It is a tripartite molecule consisting of lipid A, core oligosaccharides, and O-antigen, structurally formed by amphiphilic glycoconjugates whose composition varies within and between species. LPS molecules are positioned among the proteins and phospholipids of the outer bacterial membrane and contribute to the structural properties of the membrane; e.g., they act as a permeability barrier against various types of molecules. The structural heterogeneity of the O-antigen, the most external portion of the LPS molecule, confers versatility and adaptability to bacteria that are exposed to variable environmental conditions . Changes of LPS structure usually affect adhesive forces among bacteria, possibly through alteration of cell surface hydrophobicity. For example, in rhizobacteria such as Rhizobium leguminosarum and R. etli, LPS modifications typically alter the autoaggregation phenotype [23–25]. Ensifer lpsB mutants, which have a truncated LPS core, display a more strongly autoaggregative phenotype as compared with wild-type parental strains (Sorroche et al., unpubl. data). The rhamnose-rich O-antigen in the outermost part of the LPS of the xylem-limited phytopathogen Xylella fastidiosa is involved in cell-cell aggregation . The autoaggregative ability of X. fastidiosa appears to be an important virulence mechanism because the bacterial clusters block the passage of water and nutrients from the roots to the leaves of the host plant [27–29].
Bioassay studies of the impact of biological factors on Azospirillum brasilense autoaggregation indicated that outer membrane proteins promote bacterial flocculation . A 67-kDa outer membrane lectin on the bacterial surface specifically recognized an EPS synthesized by the aggregated cells. The interaction between the lectin and the EPS may be responsible in part for cell-cell interactions leading to autoaggregation of this species .
Pili, fimbriae, and flagella are proteinaceous polymeric appendages acting as bacterial surface organelles. Their numerous functions include mediation of motility, interbacterial interactions, bacterial-host interactions, and surface colonization . Pili are associated with autoaggregation in X. fastidiosa. This process depends on the presence of polarly located type I and type IV pili, each of which plays a specific role in the structural dynamics of the bacterial aggregates .
2.2. Extracellular Factors Involved in Bacterial Autoaggregation
Some bacteria secrete molecules that promote autoaggregation. Extracellular polymeric materials have been shown to act as “molecular glue” that initiates and maintains contact between cells, causing flocculation. The main extracellular compounds are EPSs, which are linear or branched molecules formed by one repeated sugar (homopolysaccharides) or by a mixture of different sugars (heteropolysaccharides). An example is galactoglucan (EPS II) from the symbiotic rhizobacterium Ensifer meliloti. This extracellular EPS is secreted in two major fractions, low molecular weight (LMW) and high molecular weight (HMW), according to the degree of polymerization . Mutant strains that are unable to synthesize EPS II fail to autoaggregate under static conditions. A mucR mutant secreting almost exclusively the HMW fraction of EPS II showed a weak aggregative phenotype, suggesting that the LMW fraction plays the active role in autoaggregation. Aggregation of the non-EPS-producing strains and the mucR strain was restored by resuspending the cells in culture medium containing EPS II .
Hostile environmental conditions or low metabolic activity can induce an autoaggregative phenotype via the synthesis of EPSs that have agglutinating activity. For example, the aggregative phenotype of the rhizobacterium A. brasilense depends on the production of an arabinose-rich extracellular polysaccharide that is synthesized in cultures in stationary and programmed cell death phases. Aggregative behavior in Pseudomonas aeruginosa is triggered by the presence of the toxic detergent SDS. This behavior depends on production of the EPS PsI. Inactivation of either the psi gene or the c-di-GMP-mediated signaling system that activates the gene results in reduced autoaggregation [19,38,39].
3. Cell–Cell and Cell–Surface Interactions: Bacterial Biofilm Formation
Surface and extracellular bacterial components have been extensively studied because they involve molecules that play crucial roles during the process of infection of the host plant, independently of the development of a beneficial or pathogenic relationship. Such bacterial components are also key molecules in the establishment, maturation, and dispersal of biofilms. We will summarize in this section the structure and function of bacterial compounds that play a role in the development of biofilms by beneficial or pathogenic bacteria associated with the surface or interior of plant tissues.
3.1. Structural and Functional Components Involved in Biofilm Formation
All bacteria live as a multicellular conglomerate encased in a protective matrix of polymeric substances produced by the bacteria themselves. The highly organized and dynamic social structure of bacteria requires intercellular communication via quorum sensing . Bacterial aggregates frequently adhere at a solid-liquid interface prior to adsorption on a thin film of organic molecules that constitutes the adhesion site. The transport of cells to this interface may be mediated by passive mechanisms or by the intrinsic motility of planktonic bacteria. The accumulation of bacterial cells at the interface is biphasic, consisting of (i) a non-specific reversible stage mediated by hydrophobic and electrostatic interactions between cells and adjacent surfaces; and (ii) an irreversible stage in which the adhesion process is completed and a bacterial microcolony is established . Surface bacterial components such as flagella, pili, fimbriae, and LPSs play a crucial role in physical processes during the initial stages of biofilm formation on surfaces. The growth, maturation, and disassembly phases of biofilms depend primarily on the biosynthesis of extracellular biopolymers such as EPSs, proteins, and extracellular DNA (eDNA). These polymers promote or provide immobilization of bacterial cells into the matrix, mechanical stability of the biofilm structure, cohesive interaction with the interface, and the architecture and functionality of the encased microbial community .
3.1.1. Surface Bacterial Factors
Because of their exposure to the external environment and their chemical properties, LPSs on the outer membrane of Gram-negative bacteria are capable of undergoing adhesive interactions with both biotic and abiotic surfaces.
Structural changes of LPSs have been shown to alter biofilm formation or structure in beneficial plant-associated bacteria, including Pseudomonas fluorescens . Mutant strains of various beneficial rhizobacteria with altered LPS structure display changes in the biofilm formation process. For example, a Bradyrhizobium japonicum O-antigen mutant showed enhanced adhesion to plastic supports . A R. leguminosarum lipid a mutant showed increased lateral interactions with an abiotic surface; this effect had no effect on the ability of the bacteria to form a biofilm on the surface . A mutant of the alfalfa symbiont E. meliloti that synthesized a structurally modified LPS because of mutations of the lpsB and bacA genes showed reduced biofilm formation ability [46,47]. E. meliloti lpsB mutants showed reduced nodulation abilities because of delays in the invasion steps; however, their nitrogen-fixing capacity was similar to that of wild-type [48,49], and the modified LPS molecule increased bacterial adsorption to alfalfa roots .
In phytopathogenic bacteria, as in beneficial bacteria, LPSs play crucial roles during the early stages of interaction with the host and development of virulence. These phenomena coincide with the normal stages of biofilm formation. Various mutations related to LPS synthesis in phytopathogenic bacteria such as P. aeruginosa , Pseudomonas syringae , Xanthomonas axonopodis , and Xanthomonas citri  caused reductions in both biofilm formation ability and virulence.
Because of their surface cell location and physico-chemical properties, LPSs play a key role during the initial steps of biofilm formation (e.g., adherence to surfaces) and the development of mature biofilm through interactions of cells with other cells and with matrix components.
Proteinaceous appendages (pili and flagella) are bacterial virulence factors that lead to pathogenesis in plant, animal, and human hosts and play a key role during colonization steps . The initial stages of biofilm formation are dependent on bacterial motility mediated by the polar flagellum and multiple type IV pili (TfP), which enable the free-swimming phenotype to reach a suitable surface and the surface-motile phenotype to adhere to and move on the surface . These appendages thus have a dual role as motile machines and adhesins that move and fix bacteria to surfaces and among surfaces . Various types of movement (e.g., crawling, pulling, walking) have been associated with TfP . The coordinated TfP pulling associated with release and with flagellar rotation-translation has been presented as a model whereby bacteria are able to travel through the biofilm matrix . Motility-defective mutants of P. aeruginosa and E. coli were unable to attach to surfaces or to develop a normal biofilm [59,60].
Pili structure is crucial for adhesion and biofilm formation in certain phytopathogenic bacteria, including Acidovorax citrulli, the causal agent of bacterial fruit blotch in cucurbits , and X. fastidiosa, the causal agent of Pierce’s disease in grapes . A. citrulli uses TfP and X. fastidiosa uses type I pili to colonize and move upstream against sap flow in xylem vessels while oriented parallel to the surface, prior to the complete development of biofilm and plant disease. TfP is also important for the pathogenesis of Ralstonia solanacearum and Xanthomonas oryzae pv. oryzicola [63,64]. Pili are well-established virulence factors for phytopathogenic bacteria because of their important roles in adherence and plant colonization.
Motility mediated by flagella is manifested as either “swimming” of free cells in aqueous environments and coordinated “swarming” of bacterial populations on solid moist surfaces . Both swimming and swarming are essential for various stages of biofilm development, e.g., the search for a favorable habitat, attachment to a surface, architectural assembly, structural disassembly, and release from the biofilm matrix . The complex association between motility and biofilm formation involves the use of a particular structure for different functions at different stages and requires the precise integration of environmental and cellular signals .
Flagella-mediated motility in rhizobacteria is essential for biofilm establishment and therefore for plant colonization. E. meliloti, a paradigm of beneficial symbiotic interaction between rhizobacteria and legume plants, showed reduced biofilm formation ability on abiotic surfaces as a result of mutations on genes related to flagella synthesis . An association between flagella-mediated motility and biofilm formation and an effect of quorum-sensing signals on both of these processes were demonstrated in peanut-nodulating Bradyrhizobium sp. strains . Both aflagellate and flagellated but nonmotile mutants of the well-studied pathogen A. tumefaciens, the causal agent of crown gall, showed reduced biofilm formation ability under static conditions because of defects in surface attachment. However, the aflagellate mutants were able to quickly develop an unusually dense and tall biofilm under flow conditions . In contrast to results of other studies that suggest a role of flagella as an adhesin in Aeromonas spp. , these results for A. tumefaciens are consistent with those for other bacterial species , which suggest that flagella do not function as an adhesin and that other surface structures can be involved in attachment and subsequent biofilm formation .
X. axonopodis pv. citri, a phytopathogenic bacterium that establishes itself on the leaves (phyllosphere) of citrus species and produces citrus canker disease , has the ability to form biofilms on abiotic and biotic surfaces, including the cankers of diseased plants . Flagella-mediated motility plays a key role in several stages of biofilm formation, including surface adherence, maturation, and dispersal . Sliding motility (not mediated by flagella) and the regulation of both swimming and sliding motility through diffusible signal factor (DSF) were reported for X. axonopodis . Similar findings were reported for Xanthomonas campestris pv. campestris, although flagellin mutants did not show altered virulence . Swimming motility was found to be essential for biofilm formation and colonization of plant tissues in vascular pathogens such as R. solanacearum, Pantoea stewartii, and Dickeya dadantii [77–79].
In contrast to findings in Gram-negative bacteria, it appears that motility is not essential for biofilm development in various non-motile Gram-positive bacteria. Surface proteins (e.g., Bap, Esp) have been reported to be involved in initial adherence to surfaces in these non-motile species [80,81].
Several surface structures that function as adhesins have been found to play important roles during surface attachment. Rhicadhesins and Raps (Rhizobium-adhering proteins) are important for root attachment in various Rhizobiaceae species [82,83]. Glucomannan, a surface polysaccharide of R. leguminosarum bv. trifolli, binds to pea and vetch lectins . Two surface-associated proteins were found to be involved in biofilm formation in Pseudomonas putida. LapA plays a key role in the early stages of biofilm formation by mediating bacterial adherence to various surfaces (including seeds and roots), while LapF is crucial in later stages by mediating cell-cell interactions during sessile growth [85,86].
3.1.2. Extracellular Factors
The highly developed level of bacterial organization reflected by biofilm formation ability and multicellularity requires dynamic and functional microorganisms that are embedded into a complex mixture of extracellular polymeric components collectively termed the “biofilm matrix” . These components form bridges, channels, avenues, and pores and support an impressively elaborate 3-dimensional architectural structure within which cellular arrangements are transiently constructed. In addition to its structural protective function, the biofilm matrix plays a key role in bacterial physiology and ecology, including cellular interactions, nutrient utilization, horizontal gene transfer, and environmental fitness of the bacterial population . The composition of the biofilm matrix is highly variable, depending on the type of bacteria and biofilm interface. Water is the main component of a mature biofilm. In terms of dry mass, bacteria account for <10% of a biofilm, and the matrix accounts for >90%. The biofilm matrix consists primarily of EPSs and contains smaller proportions of other biopolymers such as proteins, nucleic acids, and lipids . We will summarize here the roles of the major components of the biofilm matrix; i.e., extracellular factors.
A biofilm has been defined as a multicellular bacterial conglomerate adhered to a surface and immersed into a polymeric matrix formed primarily by EPSs . It is therefore not surprising that various bacterial strains, even of the same species, have the ability to synthesize, export, and modify their own characteristic EPS. The function and chemical composition of each EPS is different, depending on the bacterial species or strain. A particular strain may even have the ability to produce different EPSs depending on the environmental conditions, as demonstrated for P. aeruginosa  and Streptococcus thermophilus . Most EPSs are polyanionic molecules because of the presence of uronic acids and sugar having substituents such as pyruvate, sulfate, or phosphate. Polycationic EPSs have also been described . The presence of β-1,4 (or β-1,3) and α-1,2 (or α-1,6) linkages confers greater rigidity or flexibility, respectively, to the matrix structure. The stability of the biofilm structure is thus dependent on the physico-chemical and biological properties of EPSs and on their interactions with ions, low molecular weight solutes, and other macromolecules such as proteins and eDNA . EPSs are generally required not for initial adhesion but for later architectural development of the biofilm matrix . The EPS network confers mechanical stability, allows for temporary immobilization of cells, and plays a crucial role in most matrix functions, including water retention, protection from environmental stresses, adsorption of compounds, and nutrient availability .
The production of EPSs on plant surfaces or tissues allows bacterial colonization and biofilm formation. The biological roles (beneficial or pathogenic) of bacteria on plants are related to these abilities. The compositions and biological roles of selected EPSs produced by well-known plant-associated bacteria are summarized in Table 1.
The importance of cellulose (a neutral homopolysaccharide) as a key component of the polysaccharidic matrix has been demonstrated in several Enterobacteriaceae species , the plant-associated bacterium A. tumefaciens , and Rhizobium species [36,120]. Cellulose plays a key role in adherence to plant tissues, biofilm formation, and the support of matrix architecture.
Polysaccharides from Arabidopsis roots were recently found to serve as both signals for biofilm formation and a source of sugars for the synthesis of matrix EPSs in the beneficial Gram-positive bacterium Bacillus subtilis . Future studies on the triggering of bacterial biofilm formation by plant root exudates will be useful.
Extracellular proteins are major constituents of the biofilm matrix, but have received relatively little study in comparison with other components such as EPSs. Proteins in the biofilm matrix have both structural and physiological functions. Some matrix proteins function as extracellular enzymes and are associated with activities such as the degradation and recycling of biopolymers for nutrient availability and the modification of other exopolymers for shaping or releasing of cells from the biofilm structure. Enzymes that play such roles in the biofilm matrix include lipases, hydrolases, lyases, and glycanases [83,122,123]. Certain enzymes released by pathogenic bacteria may act as virulence factors [124,125], but such a function has not been evaluated in the context of biofilms on plant surfaces.
Some proteins in the biofilm matrix have structural functions, e.g., as lectins that bind bacterial cells to the polymeric matrix. Examples of such extracellular carbohydrate-binding proteins include a glucan-binding protein in Streptococcus mutans , LecA and LecB in P. aeruginosa [127,128], TasA in Bacillus subtilis , and lectins in A. brasilense . In P. aeruginosa, a large quantity of matrix proteins was found in outer membrane vesicles, a typical matrix biofilm component in this species . Amyloids are another common type of matrix protein with extracellular adhesin function .
Biofilms provide an ideal location for the exchange of genetic material. Higher levels of conjugation have been demonstrated for bacterial populations in biofilms as compared with planktonic bacteria . eDNA is an important constituent of the biofilm matrix  and plays a role in biofilm formation in various bacterial species, including P. aeruginosa  and Bacillus cereus . The quantity, localization, and origin of eDNA vary depending on the bacterial species. Some studies found that eDNA is arranged in certain patterns  and that its release is based on the lysis of certain types of bacteria , suggesting the occurrence of programmed cell death in biofilms . In Gram-positive bacteria, eDNA is involved in adhesion to hydrophobic surfaces and in bacterial autoaggregation .
Lipids are also components of the biofilm matrix , although they have received little study in the context of plant-bacterial associations. Lipids in biofilms generally act as biosurfactants with functions such as surface activity, dispersal and bioavailability of hydrophobic substances, antibacterial or antifungal properties, and bacterial attachment and detachment . Such properties have been well characterized for the rhamnolipids of P. aeruginosa, which play important roles at several stages of biofilm development, including surface interaction, microcolony formation, structural maintenance, and biofilm dispersal .
Our knowledge of the identification and functions of extracellular proteins, eDNA, and lipids in the biofilm matrix of plant-associated bacteria remains limited and fragmentary. Further studies along this line will greatly enhance our understanding of the process of biofilm formation.
4. Relationship between Biofilm Formation and Bacterial Autoaggregation
Bacteria have the unique ability to form complex cellular assemblies on biotic surfaces (animal tissues, plant tissues, detritus) and abiotic surfaces (sediments, soil particles, medical or laboratory instruments). Depending on the quantities of cells and extracellular components, such assemblies range from random cell aggregations observed on surfaces or in liquid suspension to complex, highly developed assemblies of cells encased in a exopolymeric matrix and attached to a surface (i.e., biofilms) . There are several bacterial models that are useful for studying the relationships between cell aggregations and biofilms. The adhesin AIDA, a surface glycoprotein of E. coli, has been shown to promote strong bacterial autoaggregation, biofilm formation, and in vitro adhesion to human and mammalian cells . Strains of the opportunistic bacterium Myroides odoratus that showed strong adhesion to inert supports also showed enhanced autoaggregative ability . Changes in the cell surface of Porphyromonas gingivalis (a pathogenic bacterium involved in periodontal disease) led to increased cell-cell interactions and consequent increases in autoaggregation and biofilm formation .
Variants of the phytopathogen X. fastidiosa with mutation of a gene involved in signal transduction showed a modified transcriptional profile of genes related to biofilm formation and bacterial aggregation traits such as surface attachment, EPS synthesis, and virulence .
Autoaggregative behavior has also been correlated with biofilm formation ability in beneficial plant-associated bacteria. Strongly autoaggregative mutants of the rhizobacterium A. brasilense showed a high tendency to form biofilms on inert supports . In the rhizobacterium E. meliloti, autoaggregation  and biofilm formation  depend on a combination of bacterial signals, surface components, and EPSs. The development of cell interactions in both sessile populations and planktonic aggregations of various native strains of E. meliloti showed a positive correlation with the above processes and a requirement for EPS II, indicating the involvement of the same physical adhesive forces in autoaggregation and biofilm formation .
Surface and extracellular factors are involved in both cell aggregation and biofilm formation, processes that depend in part on physical interactions among bacteria. It is therefore reasonable to presume that alterations in one of these processes lead to changes in the other. Cell aggregation on surfaces most likely represents a transitional state that precedes the development of a structured biofilm. We postulate that the linkage of or transition from bacterial aggregation to biofilm formation is crucial for the establishment of beneficial or pathogenic relationships between bacteria and plants. Such relationships are developed through (i) interactions among bacteria in or near the plant microenvironment (e.g., rhizosphere or phyllosphere); (ii) interactions between bacteria and plant surfaces (e.g., leaf or root epidermis, root hairs, transport vessels); and (iii) biofilm formation and their biological effects.
5. Intergeneric Adhesive Interactions: Coaggregation
The coaggregation process has been defined as adhesion among genetically different microorganisms . The phenomenon of coaggregation was initially described for bacteria that inhabit the human oral cavity and subsequently extended to bacteria found in other habitats, including aquatic environments and sludge . Coaggregation processes and factors that affect them have been described for plant growth-promoting rhizobacteria (PGPR) . These findings are relevant to our understanding of plant-bacterial interactions and the development of commercial inoculants.
Coaggregation is an integral process in the formation of mixed biofilms and is therefore ecologically important. Two models have been proposed to explain how a mixed biofilm can support different types of planktonic phase-derived bacteria. According to the first model, planktonic free cells in suspension specifically recognize and adhere to genetically different bacteria in a biofilm. The second model involves initial coaggregation of planktonic bacteria and subsequent adhesion and integration of the coaggregate during biofilm formation . Regardless of these models, the coaggregation phenomenon was found to be dependent on cell surface hydrophobicity and the partner strains that participated in the interaction .
On a molecular level, coaggregation of human intestinal or oral bacteria and of aquatic bacteria depends on the interaction of a lectin of one participant with a complementary glycosidic receptor of the other participant . Similar mechanisms are expected in bacteria that establish associations with plants, particularly in certain habitats in which bacterial populations are highly diverse, e.g., rhizospheric soil. Cell-free culture supernatants of E. meliloti containing EPS II induced the autoaggregation of several rhizospheric bacteria, including strains of Pseudomonas, Azospirillum, and Burkholderia. These findings suggest that interactions between EPS II and rhizobacteria play an important ecological role. EPS II itself may be capable of connecting different bacterial cells. According to this hypothetical model, EPS II-producing E. meliloti cells may act as “bridges” during the process of coaggregation with other rhizospheric bacteria .
6. Concluding Remarks
The bacterial cell surface plays a key role in bacterial aggregation, which in turn promotes bacterial dispersal, survival, and the ability to adhere to plant surfaces. It has been well documented that bacterial autoaggregation and biofilm development, and the relationship of these processes with plant colonization, are dependent on both surface bacterial factors and extracellular factors.
Bacteria gain several advantages from living in biofilms, including protection from predation, desiccation, and exposure to antibacterial substances, and improved acquisition of nutrients released in the plant environment. Biofilms provide survival sites for both beneficial and opportunistic pathogenic bacteria, by providing protection as above and increasing the potential of the bacteria to survive and evolve in the plant environment. Biofilms have been shown to enhance (i) the fitness of individual bacteria and (ii) more generalized plant health and productivity as a result of the cumulative selective advantage of the individual bacteria.
Detailed elucidation of the mechanisms involved in the various stages of biofilm formation will improve our understanding of microbial adaptations to this mode of life in a wide range of environments and of the interactions of bacteria with their eukaryotic hosts. Multidisciplinary studies using new approaches will clarify the ways in which bacteria move and interact in a variety of surface microenvironments during biofilm development. Such knowledge will enhance our understanding of biofilm formation on plant surfaces and of the sophisticated processes of interaction between prokaryotes and eukaryotes.
This review and our studies described herein were supported by grants from the Secretaría de Ciencia y Técnica (UNRC), Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT), and Consejo Nacional de Investigaciones Científicas y Técnicas of the República Argentina (CONICET). FGS was supported by a fellowship from the CONICET. PCB, MMO and WG are Career Members of CONICET. The authors thank S. Anderson for English editing of the manuscript.
Conflict of Interest
The authors declare no conflict of interest.
- Branda, S.S.; Vik, S.; Friedman, L.; Kolter, R. Biofilms: The matrix revisited. Trends Microbiol 2005, 13, 20–26.
- Davey, M.E.; O’Toole, G.A. Microbial biofilms: From ecology to molecular genetics. Microbiol. Mol. Biol. Rev 2000, 64, 847–867.
- Schembri, M.A.; Christianesen, G.; Klemm, P. FimH-mediated autoaggregation of E. coli. Mol. Microbiol 2001, 41, 1419–1430.
- Costerton, J.W.; Lewandowski, Z.; Caldwell, D.E.; Korber, D.R.; Lappin-Scott, H.M. Microbial biofilms. Annu. Rev. Microbiol 1995, 49, 711–745.
- Rinaudi, L.V.; Giordano, W. An integrated view of biofilm formation in rhizobia. FEMS Microbiol. Lett 2010, 304, 1–11.
- Stanley, N.R.; Lazazzera, B.A. Environmental signals and regulatory pathways that influence biofilm formation. Mol. Microbiol 2004, 52, 917–924.
- Sutherland, I.W. Biofilm exopolysaccharides: A strong and sticky framework. Microbiology 2001, 147, 3–9.
- Flemming, H.C. Biofilms and environmental protection. Water Sci. Technol 1993, 27, 1–10.
- Gilbert, P.; Das, J.; Foley, I. Biofilms susceptibility to antimicrobials. Adv. Dent. Res 1997, 11, 160–167.
- Matthysse, A.G.; Marry, M.; Krall, L.; Kaye, M.; Ramey, B.E.; Fuqua, C.; White, A.R. The effect of cellulose overproduction on binding and biofilm formation on roots by Agrobacterium tumefaciens. Mol. Plant Microbe Interact 2005, 18, 1002–1010.
- Van de Mortel, M.; Halverson, L.J. Cell envelope components contributing to biofilm growth and survival of Pseudomonas putida in low-water-content habitats. Mol. Microbiol 2004, 52, 735–750.
- Rudrappa, T.; Biedrzycki, M.L.; Bais, H.P. Causes and consequences of plant-associated biofilms. FEMS Microbiol. Ecol 2008, 64, 153–166.
- Mansfield, J.; Genin, S.; Magori, S.; Citovsky, V.; Sriariyanum, M.; Ronald, P.; Dow, M.; Verdier, V.; Beer, S.V.; Machado, M.A.; et al. Top 10 plant pathogenic bacteria in molecular plant pathology. Mol. Plant Pathol 2012, 13, 614–629.
- Sorroche, F.G.; Rinaudi, L.V.; Zorreguieta, A.; Giordano, W. EPS II-dependent autoaggregation of Sinorhizobium meliloti planktonic cells. Curr. Microbiol 2010, 61, 465–470.
- Sorroche, F.G.; Spesia, M.B.; Zorreguieta, A.; Giordano, W. A positive correlation between bacterial autoaggregation and biofilm formation in native Sinorhizobium meliloti isolates from Argentina. Appl. Environ. Microbiol 2012, 78, 4092–4101.
- Neyra, C.; Sadasivan, L. Flocculation in Azospirillum brasilense and Azospirillum lipoferum: Exopolysacharides and cyst formation. J. Bacteriol 1985, 163, 716–723.
- Nikitina, V.E.; Ponomareva, E.G.; Alen’kina, S.A.; Konnova, S.A. The role of cell-surface lectins in the aggregation of Azospirilla. Microbiology 2001, 70, 471–476.
- Bahat-Samet, E.; Castro-Sowinski, S.; Okon, Y. Arabinose content of extracellular polysaccharide plays a role in cell aggregation of Azospirillum brasilense. FEMS Microbiol. Lett 2004, 237, 195–203.
- Klebensberger, J.; Birkenmaier, A.; Geffers, R.; Kjelleberg, S.; Philipp, B. SiaA and SiaD are essential for inducing autoaggregation as a specific response to detergent stress in Pseudomonas aeruginosa. Environ. Microbiol 2009, 11, 3073–3086.
- Malik, A.; Sakamoto, M.; Hanazaki, S.; Osawa, M.; Susuki, T.; Tochigi, M.; Kakii, K. Coaggregation among nonfloculating bacteria isolated from activated sludge. Appl. Environ. Microbiol 2003, 69, 6056–6064.
- Burdman, S.; Jurkevitch, E.; Soria-Díaz, M.E.; Serrano, A.M.; Okon, Y. Extracellular polysaccharide composition of Azospirillum brasilense and its relation with cell aggregation. FEMS. Microbiol. Lett 2000, 189, 259–264.
- Lerouge, I.; Vanderlayden, J. O-antigen structural variation: Mechanisms and possible roles in animal/plant-microbe interactions. FEMS Microbiol. Rev 2001, 26, 17–47.
- Priefer, U.B. Genes involved in lipopolysaccharide production and symbiosis are clustered on the chromosome of Rhizobium leguminosarum biovar viciae VF39. J. Bacteriol 1989, 171, 6161–6168.
- Cava, J.R.; Elias, P.M.; Turowski, D.A.; Noel, K.D. Rhizobium leguminosarum CFN42 genetic region encoding lipopolysaccharide structures essential for complete nodule development on bean plants. J. Bacteriol 1989, 171, 8–15.
- García de los Santos, A.; Brom, S. Characterization of two plasmid-borne lps beta loci of Rhizobium etli required for lipopolysaccharide synthesis and for optimal interaction with plants. Mol. Plant Microbe Interact 1997, 10, 891–902.
- Clifford, J.C.; Rapicavoli, J.N.; Roper, M.C. A rhamnose-rich O-antigen mediates adhesion, virulence and host colonization for the xylem-limited phytopathogen, Xylella fastidiosa. Mol. Plant. Microbe Interact. 2013, 26, 676–685.
- Hopkins, D.L. Seasonal concentration of the Pierce’s disease bacterium in grapevine stems, petioles, and leaf veins. Phytopathology 1981, 71, 415–418.
- Tyson, G.E.; Stojanovic, B.J.; Kuklinski, R.F.; Divittorio, T.J.; Sullivan, M.L. Scanning electron microscopy of Pierce’s disease bacterium in petiolar xylem of grape leaves. Phytopathology 1985, 75, 264–269.
- Newman, K.L.; Almeida, R.P.P.; Purcell, A.H.; Lindow, S.E. Use of a green fluorescent strain for analysis of Xylella fastidiosa colonization of Vitis vinifera. Appl. Environ. Microbiol 2003, 69, 7319–7327.
- Burdman, S.; Jurkevitch, E.; Schwartsburd, B.; Okon, Y. Involvement of outer membrane proteins in the aggregation of Azospirillum brasilense. Microbiology 1999, 145, 1145–1152.
- Mora, P.; Rosconi, F.; Franco Fraguas, L.; Castro-Sowinski, S. Azospirillum brasilense Sp7 produces an outer-membrane lectin that specifically binds to surface-exposed extracellular polysaccharide produced by the bacterium. Arch. Microbiol 2008, 189, 519–524.
- Craig, L.; Pique, M.E.; Tainer, J.A. Type IV pilus structure and bacterial pathogenicity. Nature 2004, 2, 363–378.
- De La Fuente, L.; Burr, T.J.; Hoch, H.C. Autoaggregation of Xilella fastidiosa cells is influenced by type I and type IV pili. Appl. Environ. Microbiol 2008, 74, 5579–5581.
- González, J.E.; Reuhs, B.L.; Walker, G.C. Low molecular weight EPS II of Rhizobium meliloti allows nodule invasion in Medicago sativa. Proc. Natl. Acad. Sci. USA 1996, 93, 8636–8641.
- Ausmees, N.; Jonsson, H.; Höglund, S.; Ljunggren, H.; Lindberg, M. Structural and putative regulatory genes involved in cellulose synthesis in Rhizobium leguminosarum bv trifolii. Microbiology 1999, 145, 1253–1262.
- Laus, M.C.; van Brussel, A.A.; Kijne, J.W. Role of cellulose fibrils and exopolysaccharides of Rhizobium leguminosarum in attachment to and infection of Vicia sativa root hairs. Mol. Plant Microbe Interact 2005, 18, 533–538.
- Williams, A.; Wilkinson, A.; Krehenbrink, M.; Russo, D.M.; Zorreguieta, A.; Downie, J.A. Glucomannan-mediated attachment of Rhizobium leguminosarum to pea root hairs is required for competitive nodule infection. J. Bacteriol 2008, 190, 4706–4715.
- Klebensberger, J.; Rui, O.; Fritz, E.; Schink, B.; Philipp, B. Cell aggregation of Pseudomonas aeruginosa strain PAO1 as an energy-dependent stress response during growth with sodium dodecyl sulfate. Arch. Microbiol 2006, 185, 417–427.
- Klebensberger, J.; Lautenschlager, K.; Bressler, D.; Wingender, J.; Philipp, B. Detergent-induced cell aggregation in subpopulations of Pseudomonas aeruginosa as a preadaptive survival strategy. Environ. Microbiol 2007, 9, 2247–2259.
- Hooshangi, S.; Bentley, W.E. From unicellular properties to multicellular behavior: Bacteria quorum sensing circuitry and applications. Curr. Opin. Biotech 2008, 19, 550–555.
- Klausen, M.; Gjermansen, M.; Kreft, J.U.; Tolker-Nielsen, T. Dynamics of development and dispersal in sessile microbial communities: Examples from Pseudomonas aeruginosa and Pseudomonas putida model biofilms. FEMS Microbiol. Lett 2006, 261, 1–11.
- Flemming, H.C.; Wingender, J. The biofilm matrix. Nature Rev. Microbiol 2010, 8, 623–633.
- Spiers, A.J.; Rainey, P.B. The Pseudomonas fluorescens SBW25 wrinkly spreader biofilm requires attachment factor, cellulose fibre and LPS interactions to maintain strength and integrity. Microbiology 2005, 151, 2829–2839.
- Lee, Y.W.; Jeong, S.Y.; In, Y.H.; Kim, K.Y.; So, J.S.; Chang, W.S. Lack of O-polysaccharide enhances biofilm formation by Bradyrhizobium japonicum. Lett. Appl. Microbiol 2010, 50, 452–456.
- Vanderlinde, E.M.; Muszynski, A.; Harrison, J.J.; Koval, S.F.; Foreman, D.L.; Ceri, H.; Kannenberg, E.L.; Carlson, R.W.; Yost, C.K. Rhizobium leguminosarum biovar viciae 3841, deficient in 27-hydroxyoctacosanoate-modified lipopolysaccharide, is impaired in desiccation tolerance, biofilm formation and motility. Microbiology 2009, 155, 3055–3069.
- Ferguson, G.P.; Roop, R.M., II; Walker, G.C. Deficiency of a Sinorhizobium meliloti bacA mutant in alfalfa symbiosis correlates with alteration of the cell envelope. J. Bacteriol. 2002, 184, 5625–5632.
- Hirsch, A.M.; Lum, M.R.; Fujishige, N.A. Microbial Encounters of a Symbiotic Kind-Attaching to Roots and Other Surfaces. In Root Hairs: Plant Cell Monographs; Emons, A.M.C., Ketelaar, T., Eds.; Springer-Verlag: Berlin, Germany, 2009; Volume 12, pp. 295–314.
- Lagares, A.; Caetano-Anolles, G.; Niehaus, K.; Lorenzen, J.; Ljunggren, H.D.; Pühler, A.; Favelukes, G. A Rhizobium meliloti lipopolysaccharide mutant altered in competitiveness for nodulation of alfalfa. J. Bacteriol 1992, 174, 5941–5952.
- Campbell, G.R.O.; Reuhs, B.L.; Walker, G.C. Chronic intracelular infection of alfalfa nodules by Sinorhizobium meliloti requires correct lipopolysaccharide core. Proc. Natl. Acad. Sci. USA 2002, 99, 3938–3943.
- Sorroche, F.G. National University of Río Cuarto: Córdoba, Argentina. Unpublished work, 2011.
- Lau, P.C.Y.; Lindhout, T.; Beveridge, T.J.; Dutcher, J.R.; Lam, J.S. Differential lipopolysaccharide core capping leads to quantitative and correlated modifications of mechanical and structural properties in Pseudomonas aeruginosa biofilms. J. Bacteriol 2009, 191, 6618–6631.
- Deng, W.L.; Lin, Y.C.; Lin, R.H.; Wei, C.F.; Huang, Y.C.; Peng, H.L.; Huang, H.C. Effects of galU mutation on Pseudomonas syringae—Plant interactions. Mol. Plant Microbe Interact 2010, 23, 1184–1196.
- Li, J.; Wang, N. The wxacO gene of Xanthomonas citri ssp. citri encodes a protein with a role in lipopolysaccharide biosynthesis, biofilm formation, stress tolerance and virulence. Mol. Plant Pathol 2011, 12, 381–396.
- Li, J.; Wang, N. Genome-wide mutagenesis of Xanthomonas axonopodis pv. citri reveals novel genetic determinants and regulation mechanisms of biofilm formation. PLoS One 2011, 6, e21804.
- Dhakal, B.K.; Bower, J.M.; Mulvey, M.A.; Pili, Fimbriae. Encyclopedia of Microbiology; Moselio, S., Ed.; Academy Press: Oxford, UK, 2009; p. 470.
- Conrad, J.C. Physics of bacterial near-surface motility using flagella and type IV pili: Implications for biofilm formation. Res. Microbiol 2012, 163, 619–629.
- Petrova, O.E.; Sauer, K. Sticky situations: Key components that control bacterial surface attachment. J. Bacteriol 2012, 194, 2413–2425.
- Jin, F.; Conrad, J.C.; Gibiansky, M.L.; Wong, G.C.L. Bacteria use type-IV pili to slingshot on surfaces. Proc. Natl. Acad. Sci. USA 2011, 108, 12617–12622.
- Wood, T.K.; González Barrios, A.F.; Herzberg, M.; Lee, J. Motility influences biofilm architecture in Escherichia coli. Appl. Microbiol. Biotechnol 2006, 72, 361–367.
- Barken, K.B.; Pamp, S.J.; Yang, L.; Gjermansen, M.; Bertrand, J.J.; Klausen, M.; Givskov, M.; Whitchurch, C.B.; Engel, J.B.; Tolker-Nielsen, T. Roles of type IV pili, flagellum-mediated motility and extracellular DNA in the formation of mature multicellular structures in Pseudomonas aeruginosa biofilms. Environ. Microbiol 2008, 10, 2331–2343.
- Bahar, O.; de la Fuente, L.; Burdman, S. Assessing adhesion, biofilm formation and motility of Acidovorax citrulli using microfluidic flow chambers. FEMS Microbiol. Lett 2010, 312, 33–39.
- De la Fuente, L.; Montanes, E.; Meng, Y.; Li, Y.; Burr, T.J.; Hoch, H.C.; Wu, M. Assessing adhesion forces of type I and type IV pili of Xylella fastidiosa bacteria by use of a microfluidic flow chamber. Appl. Environ. Microbiol 2007, 73, 2690–2696.
- Kang, Y.; Liu, H.; Genin, S.; Schell, M.A.; Denny, T.P. Ralstonia solanacearum requires type 4 pili to adhere to multiple surfaces and for natural transformation and virulence. Mol. Microbiol 2002, 2, 427–437.
- Wang, L.; Makino, S.; Subedee, A.; Bogdanove, A.J. Novel candidate virulence factors in rice pathogen Xanthomonas oryzae pv. oryzicola as revealed by mutational analysis. Appl. Environ. Microbiol 2007, 73, 8023–8027.
- Jarrell, K.F.; McBride, M.J. The surprisingly diverse ways that prokaryotes move. Nat. Rev. Microbiol 2008, 6, 466–476.
- Klausen, M.; Heydorn, A.; Ragas, P.; Lambertsen, L.; Aaes-Jørgensen, A.; Molin, S.; Tolker-Nielsen, T. Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants. Mol. Microbiol 2003, 48, 1511–1524.
- Verstraeten, N.; Braeken, K.; Debkumari, B.; Fauvart, M.; Fransaer, J.; Vermant, J.; Michiels, J. Living on a surface: Swarming and biofilm formation. Trends Microbiol 2008, 16, 496–506.
- Fujishige, N.A.; Kapadia, N.N.; de Hoff, P.L.; Hirsch, A.M. Investigations of Rhizobium biofilm formation. FEMS Microbiol. Ecol 2006, 56, 195–206.
- Nievas, F.; Bogino, P.; Sorroche, F.; Giordano, W. Detection, characterization, and biological effect of quorum-sensing signaling molecules in peanut-nodulating bradyrhizobia. Sensors 2012, 12, 2851–2873.
- Merritt, P.M.; Danhorn, T.; Fuqua, C. Motility and chemotaxis in Agrobacterium tumefaciens surface attachment and biofilm formation. J. Bacteriol 2007, 189, 8005–8014.
- Kirov, S.M.; Castrisios, M.; Shaw, J.G. Aeromonas flagella (polar and lateral) are enterocyte adhesins that contribute to biofilm formation on surfaces. Infect. Immun 2004, 72, 1939–1945.
- Lemon, K.P.; Higgins, D.E.; Kolter, R. Flagellar motility is critical for Listeria monocytogenes biofilm formation. J. Bacteriol 2007, 189, 4418–4424.
- Brunings, A.M.; Gabriel, D.W. Xanthomonas citri: Breaking the surface. Mol. Plant Pathol 2003, 4, 141–157.
- Rigano, L.A.; Siciliano, F.; Enrique, R.; Sendín, L.; Filippone, P.; Torres, P.S.; Qüesta, J.; Dow, J.M.; Castagnaro, A.P.; Vojnov, A.A.; et al. Biofilm formation, epiphytic fitness, and canker development in Xanthomonas axonopodis pv citri. Mol. Plant Microbe Interact 2007, 20, 1222–1230.
- Malamud, F.; Torres, P.S.; Roeschlin, R.; Rigano, L.A.; Enrique, R.; Bonomi, H.R.; Castagnaro, A.P.; Marano, M.R.; Vojnov, A.A. The Xanthomonas axonopodis pv. citri flagellum is required for mature biofilm and canker development. Microbiology 2011, 157, 819–829.
- Lee, M.C.; Weng, S.F.; Tseng, Y.H. Flagellin gene fliC of Xanthomonas campestris is upregulated by transcription factor Clp. Biochem. Biophys. Res. Commun 2003, 307, 647–652.
- Tans-Kersten, J.; Huang, H.; Allen, C. Ralstonia solanacearum needs motility for invasive virulence on tomato. J. Bacteriol 2001, 183, 3597–3605.
- Herrera, C.M.; Koutsoudis, M.D.; Wang, X.; von Bodman, S.B. Pantoea stewartii subsp. stewartii exhibits surface motility, which is a critical aspect of Stewart’s wilt disease development on maize. Mol. Plant Microbe Interact 2008, 21, 1359–1370.
- Jahn, C.E.; Willis, D.K.; Charkowski, A.O. The flagellar sigma factor fliA is required for Dickeya dadantii virulence. Mol. Plant Microbe Interact 2008, 21, 1431–1442.
- Cucarella, J.; Solano, C.; Valle, J.; Amorena, B.; Lasa, I.; Penadés, J.R. Bap, a Staphylococcus aureus surface protein involved in biofilm formation. J. Bacteriol 2001, 183, 2888–2896.
- Toledo-Arana, A.; Valle, J.; Solano, C.; Arrizubieta, M.J.; Cucarella, J.; Lamata, M. The enterococcal surface protein, Esp, is involved in Enterococcus faecalis biofilm formation. Appl. Environ. Microbiol 2001, 67, 4538–4545.
- Ausmees, N.; Jacobsson, K.; Lindberg, M. A unipolarly located, cell-surface-associated agglutinin, RapA, belongs to a family of Rhizobium-adhering proteins (Rap) in Rhizobium leguminosarum bv trifolii. Microbiology 2001, 147, 549–559.
- Russo, D.M.; Williams, A.; Edwards, A.; Posadas, D.M.; Finnie, C.; Dankert, M.; Downie, J.A.; Zorreguieta, A. Proteins exported via the PrsD-PrsE type I secretion system and the acidic exopolysaccharide are involved in biofilm formation by Rhizobium leguminosarum. J. Bacteriol 2006, 188, 4474–4486.
- Laus, M.C.; Logman, T.J.; Lamers, G.E.; van Brussel, A.A.; Carlson, R.W.; Kijne, J.W. A novel polar surface polysaccharide from Rhizobium leguminosarum binds host plant lectin. Mol. Microbiol 2006, 59, 1704–1713.
- Hinsa, S.M.; Espinosa-Urgel, M.; Ramos, J.L.; O’Toole, G.A. Transition from reversible to irreversible attachment during biofilm formation by Pseudomonas fluorescens WCS365 requires an ABC transporter and a large secreted protein. Mol. Microbiol 2003, 49, 905–918.
- Martínez-Gil, M.; Yousef-Coronado, F.; Espinosa-Urgel, M. LapF, the second largest Pseudomonas putida protein, contributes to plant root colonization and determines biofilm architecture. Mol. Microbiol 2010, 77, 549–561.
- Ryder, C.; Byrd, M.; Wozniak, D.J. Role of exopolysaccharides in Pseudomonas aeruginosa biofilm development. Curr. Opin. Microbiol 2007, 10, 644–648.
- Vaningelgem, F.; Zamfir, M.; Mozzi, F.; Adriany, T.; Vancanneyt, M.; Swings, J.; de Vuyst, L. Biodiversity of exopolysaccharides produced by Streptococcus thermophilus strains is reflected in their production and their molecular and functional characteristics. Appl. Environ. Microbiol 2004, 70, 900–912.
- Jefferson, K.K. Bacterial Polysaccharides. Current Innovations and Future Trends; Ullrich, M., Ed.; Caister Academic: Norfolk, UK, 2009; pp. 175–186.
- Sutherland, I.W. Biofilm Exopolysaccharides. In Microbial Extracellular Polymeric Substances; Wingender, J., Neu, T.R., Flemming, H.C., Eds.; Springer: Berlin, Germany, 1999; pp. 73–92.
- Reuber, T.L.; Walker, G.C. Biosynthesis of succinoglycan, a symbiotically important exopolysaccharide of Rhizobium meliloti. Cell 1993, 74, 269–280.
- Cheng, H.P.; Walker, G.C. Succinoglycan is required for initiation and elongation of infection threads during nodulation of alfalfa by Rhizobium meliloti. J. Bacteriol. 1998, 180, 5183–5191.
- Her, G.R.; Glazebroock, J.; Walker, G.C.; Reinhold, V.N. Structural studies of a novel exopolysaccharide produced by a mutant of Rhizobium meliloti strain Rm1021. Carbohyd. Res 1990, 198, 305–312.
- Rinaudi, L.V.; González, J.E. The low-molecular-weight fraction of exopolysaccharide II from Sinorhizobium meliloti is a crucial determinant of biofilm formation. J. Bacteriol 2009, 191, 7216–7224.
- Robertson, B.K.; Aman, P.; Darvill, A.G.; McNeil, M.; Albersheim, P. Host-symbiont interactions. The structure of acidic extracellular polysaccharides secreted by Rhizobium leguminosarum and Rhizobium trifolii. Plant Physiol 1981, 67, 389–400.
- Philip-Hollingsworth, S.; Hollingsworth, R.I.; Dazzo, F.B. Host-range related structural features of the acidic extracellular polysaccharides of Rhizobium trifolii and Rhizobium leguminosarum. J. Biol. Chem 1989, 264, 1461–1466.
- Janczarek, M.; Kutkowska, J.; Piersiak, T.; Skorupska, A. Rhizobium leguminosarum bv. trifolii rosR is required for interaction with clover, biofilm formation and adaptation to the environment. BMC Microbiol. 2010, 10, doi:10.1186/1471-2180-10-284.
- Rolfe, B.G.; Carlson, R.W.; Ridge, R.W.; Dazzo, R.W.; Mateos, F.B.; Pankhurst, C.E. Defective infection and nodulation of clovers by exopolysaccharide mutants of Rhizobium leguminosarum bv trifolii. Aust. J. Plant Physiol 1996, 23, 285–303.
- Van Workum, W.A.T.; van Slageren, S.; van Brussel, A.A.N.; Kijne, J.W. Role of exopolysaccharides of Rhizobium leguminosarum bv. viciae as host plant specific molecules required for infection thread formation during nodulation of Vicia sativa. Mol. Plant Microbe Interact 1998, 11, 1233–1241.
- Minamisawa, K. Comparison of extracellular polysaccharide composition, rhizobiotoxine production, and hydrogenase phenotype among various strains of Bradyrhizobium japonicum. Plant Cell Physiol 1989, 30, 877–884.
- Poveda, A.; Santamaria, M.; Bernabe, M.; Prieto, A.; Briux, M.; Corzo, J.; Jimenez-Barbero, J. Studies on the structure and the solution conformation of an acidic extracellular polysaccharide isolated from Bradyrhizobium. Carbohyd. Res 1997, 304, 209–217.
- Pérez-Giménez, J.; Mongiardini, E.J.; Althabegoiti, M.J.; Covelli, J.; Quelas, J.I.; López-García, S.L.; Lodeiro, A.R. Soybean lectin enhances biofilm formation by Bradyrhizobium japonicum in the absence of plants. Int. J. Microbiol. 2009, 2009, doi:10.1155/2009/719367.
- Wang, P.; Zhong, Z.; Zhou, J.; Cai, T.; Zhu, J. Exopolysaccharide biosynthesis is important for Mesorhizobium tianshanense: Plant host interaction. Arch. Microbiol 2008, 189, 525–530.
- Cangelosi, G.A.; Hung, L.; Puvanesarajah, V.; Stacey, G.; Ozga, D.A.; Leigh, J.A.; Nester, E.W. Common loci for Agrobacterium tumefaciens and Rhizobium meliloti exopolysaccharide synthesis and their roles in plant interactions. J. Bacteriol 1987, 169, 2086–2091.
- Tomlinson, A.D.; Ramey-Hartung, B.; Day, T.W.; Merritt, P.M.; Fuqua, C. Agrobacterium tumefaciens ExoR represses succinoglycan biosynthesis and is required for biofilm formation and motility. Microbiology 2010, 156, 2670–2681.
- Da Silva, F.R.; Vettore, A.L.; Kemper, E.L.; Leite, A.; Arruda, P. Fastidian gum: The Xylella fastidiosa exopolysaccharide possibly involved in bacterial pathogenicity. FEMS Microbiol. Lett 2001, 203, 165–171.
- Roper, M.C.; Greve, L.C.; Labavitch, J.M.; Kirkpatrick, B.C. Detection and visualization of an exopolysaccharide produced by Xylella fastidiosa in vitro and in planta. Appl. Environ. Microbiol 2007, 73, 7252–7258.
- Jansson, P.E.; Kenne, L.; Lindberg, B. Structure of the extracellular polysaccharide from Xanthomonas campestris. Carbohyd. Res 1975, 45, 274–282.
- Rigano, L.A.; Payette, C.; Brouillard, G.; Marano, M.R.; Abramowicz, L.; Torres, P.S.; Yun, M.; Castagnaro, A.P.; Oirdi, M.E.; Dufour, V.; et al. Bacterial cyclic beta-(1,2)-glucan acts in systemic suppression of plant immune responses. Plant Cell 2007, 19, 2077–2089.
- Torres, P.S.; Malamud, F.; Rigano, L.A.; Russo, D.M.; Marano, M.R.; Castagnaro, A.P.; Zorreguieta, A.; Bouarab, K.; Dow, J.M.; Vojnov, A.A. Controlled synthesis of the DSF cell-cell signal is required for biofilm formation and virulence in Xanthomonas campestris. Environ. Microbiol 2007, 9, 2101–2109.
- Nimtz, M.; Mort, A.; Wray, V.; Domke, T.; Zhang, Y.; Coplin, D.L.; Geider, K. Structure of stewartan, the capsular exopolysaccharide from the corn pathogen Erwinia stewartii. Carbohyd. Res 1996, 288, 189–201.
- Koutsoudis, M.D.; Tsaltas, D.; Minogue, T.D.; von Bodman, S.B. Quorum-sensing regulation governs bacterial adhesion, biofilm development, and host colonization in Pantoea stewartii subspecies stewartii. Proc. Natl. Acad. Sci. USA 2006, 103, 5983–5988.
- Nimtz, M.; Mort, A.; Domke, T.; Wray, V.; Zhang, Y.; Qiu, F.; Coplin, D.; Geider, K. Structure of amylovoran, the capsular exopolysaccharide from the fire blight pathogen Erwinia amylovora. Carbohyd. Res 1996, 287, 59–76.
- Koczan, J.M.; McGrath, M.J.; Zhao, Y.; Sundin, G.W. Contribution of Erwinia amylovora exopolysaccharides amylovoran and levan to biofilm formation: Implications in pathogenicity. Phytopathology 2009, 99, 1237–1244.
- Geier, G.; Geider, K. Characterization and influence on virulence of the levansucrase gene from the fireblight pathogen Erwinia amylovora. Physiol. Mol. Plant Pathol 1993, 42, 387–404.
- Cook, D.; Sequeira, L. Genetic and biochemical characterization of a gene cluster from Pseudomonas solanacearum required for extracellular polysaccharide production and for virulence. J. Bacteriol 1991, 173, 1654–1662.
- Chapman, M.R.; Kao, C.C. EpsR modulates production of extracellular polysaccharides in the bacterial wilt pathogen Ralstonia (Pseudomonas) solanacearum. J. Bacteriol 1998, 180, 27–34.
- Zogaj, X.; Nimtz, M.; Rohde, M.; Bokranz, W.; Römling, U. The multicellular morphotypes of Salmonella typhimurium and Escherichia coli produce cellulose as the second component of the extracellular matrix. Mol. Microbiol 2001, 39, 1452–1463.
- Matthysse, A.G.; White, S.; Lightfoot, R. Genes required for cellulose synthesis in Agrobacterium tumefaciens. J. Bacteriol 1995, 177, 1069–1075.
- Robledo, M.; Rivera, L.; Jiménez-Zurdo, J.I.; Rivas, R.; Dazzo, F.; Velázquez, E.; Martínez-Molina, E.; Hirsch, A.M.; Mateos, P.F. Role of Rhizobium endoglucanase CelC2 in cellulose biosynthesis and biofilm formation on plant roots and abiotic surfaces. Microb. Cell Fact 2012, 11, 125.
- Beauregard, P.B.; Chai, Y.; Vlamakis, H.; Losick, R.; Kolter, R. Bacillus subtilis biofilm induction by plant polysaccharides. Proc. Natl. Acad. Sci. USA 2013, 110, 1621–1630.
- Zhang, X.; Bishop, P. Biodegradability of biofilm extracellular polymeric substances. Chemosphere 2003, 50, 63–69.
- Laue, H.; Schenk, A.; Li, H.; Lambertsen, L.; Neu, T.R.; Molin, S.; Ullrich, M.S. Contribution of alginate and levan production to biofilm formation by Pseudomonas syringae. Microbiology 2006, 152, 2909–2918.
- Dafna, T.A.; Rosenberg, T.; Navon, N.; Burdman, S. A secreted lipolytic enzyme from Xanthomonas campestris pv. vesicatoria is expressed in planta and contributes to its virulence. Mol. Plant Pathol 2012, 13, 556–567.
- Hu, J.; Qian, W.; He, C. The Xanthomonas oryzae pv. oryzae eglXoB endoglucanase gene is required for virulence to rice. FEMS Microbiol. Lett 2007, 269, 273–279.
- Lynch, D.J.; Fountain, T.L.; Mazurkiewicz, J.E.; Banas, J.A. Glucan-binding proteins are essential for shaping Streptococcus mutans biofilm architecture. FEMS Microbiol. Lett 2007, 268, 158–165.
- Tielker, D.; Hacker, S.; Loris, R.; Strathmann, M.; Wingender, J.; Wilhelm, S.; Rosenau, F.; Jaeger, K.E. Pseudomonas aeruginosa lectin LecB is located in the outer membrane and is involved in biofilm formation. Microbiology 2005, 151, 1313–1323.
- Diggle, S.P.; Stacey, R.E.; Dodd, C.; Cámara, M.; Williams, P.; Winzer, K. The galactophilic lectin, LecA, contributes to biofilm development in Pseudomonas aeruginosa. Environ. Microbiol 2010, 8, 1095–1104.
- Branda, S.S.; Chu, F.; Kearns, D.B.; Losick, R.; Kolter, R. A major protein component of the Bacillus subtilis biofilm matrix. Mol. Microbiol 2006, 59, 1229–1238.
- Toyofuku, M.; Roschitzki, B.; Riedel, K.; Eberl, L. Identification of proteins associated with the Pseudomonas aeruginosa biofilm extracellular matrix. J. Proteome Res 2012, 11, 4906–4915.
- Larsen, P.; Nielsen, J.L.; Dueholm, M.S.; Wetzel, R.; Otzen, D.; Nielsen, P.H. Amyloid adhesins are abundant in natural biofilms. Environ. Microbiol 2007, 9, 3077–3090.
- Molin, S.; Tolker-Nielsen, T. Gene transfer occurs with enhanced efficiency in biofilms and induces enhanced stabilisation of the biofilm structure. Curr. Opin. Biotechnol 2003, 14, 255–261.
- Dominiak, D.M.; Nielsen, J.L.; Nielsen, P.H. Extracellular DNA is abundant and important for microcolony strength in mixed microbial biofilms. Environ. Microbiol 2011, 13, 710–721.
- Yang, L.; Barken, K.B.; Skindersoe, M.E.; Christensen, A.B.; Givskov, M.; Tolker-Nielsen, T. Effects of iron on DNA release and biofilm development by Pseudomonas aeruginosa. Microbiology 2007, 153, 1318–1328.
- Vilain, S.; Pretorius, J.M.; Theron, J.; Broezel, V.S. DNA as an adhesion: Bacillus cereus requires extracellular DNA to form biofilms. Appl. Environ. Microbiol 2009, 75, 2861–2868.
- Böckelmann, U.; Janke, A.; Kuhn, R.; Neu, T.R.; Wecke, J.; Lawrence, J.R.; Szewzyk, U. Bacterial extracellular DNA forming a defined network-like structure. FEMS Microbiol. Lett 2006, 262, 31–38.
- Allesen-Holm, M.; Barken, K.B.; Yang, L.; Klausen, M.; Webb, J.S.; Kjelleberg, S.; Molin, S.; Givskov, M.; Tolker-Nielsen, T. A characterization of DNA release in Pseudomonas aeruginosa cultures and biofilms. Mol. Microbiol 2006, 59, 1114–1128.
- Wang, J.; Bayles, K.W. Programmed cell death in plants: Lessons from bacteria? Trends Plant Sci 2013, 18, 133–139.
- Das, T.; Prashant, S.K.; Busscher, H.J.; van der Mei, H.C.; Krom, B.P. Role of extracellular DNA in initial bacterial adhesion and surface aggregation. Appl. Environ. Microbiol 2010, 76, 3405–3408.
- Conrad, A.; Suutari, M.K.; Keinänen, M.M.; Cadoret, A.; Faure, P.; Mansuy-Huault, L.; Block, J.C. Fatty acid lipid fractions in extracellular polymeric substances of activated sludge flocs. Lipids 2003, 38, 1093–1105.
- Ron, E.Z.; Rosenberg, E. Natural role of biosurfactants. Environ. Microbiol 2001, 3, 229–236.
- Pamp, S.J.; Tolker-Nielsen, T. Multiple roles of biosurfactants in structural biofilm development by Pseudomonas aeruginosa. J. Bacteriol 2007, 189, 2531–2539.
- Stoodley, P.; Sauer, K.; Davies, D.G.; Costerton, J.W. Biofilms as complex differentiated communities. Annu. Rev. Microbiol 2002, 56, 187–209.
- Sherlock, O.; Schembri, M.A.; Reisner, A.; Klemm, P. Novel roles for the AIDA adhesin from diarrheagenic Escherichia coli: Cell aggregation and biofilm formation. J. Bacteriol 2004, 186, 8058–8065.
- Jacobs, A.; Chenia, H.Y. Biofilm-forming capacity, surface hydrophobicity and aggregation characteristics of Myroides odoratus isolated from South African Oreochromis mossambicus fish. J. Appl. Microbiol 2009, 107, 1957–1966.
- Yamaguchi, M.; Sato, K.; Yukitake, H.; Noiri, Y.; Ebisu, S.; Nakayama, K. Porphyromonas gingivalis mutant defective in a putative glycosyltransferase exhibits defect in biosynthesis of polysaccharide portions of lipopolysaccharide, decreased gingipain activities, strong autoaggregation and increased biofilm formation. Infec. Immun 2010, 78, 3801–3812.
- Voegel, T.M.; Doddapaneni, H.; Cheng, D.W.; Lin, H.; Stenger, D.C.; Kirkpatrick, B.C.; Roper, M.C. Identification of a response regulator involved in surface attachment, cell–cell aggregation, exopolysaccharide production and virulence in the plant pathogen Xylella fastidiosa. Mol. Plant Pathol 2013, 14, 256–264.
- Nievas, F. National University of Río Cuarto: Córdoba, Argentina. Unpublished work, 2012.
- Rickard, A.H.; Gilbert, P.; High, N.J.; Kolenbrander, P.E.; Handley, P.S. Bacterial Coaggregation: An integral process in the development of multi-species biofilms. Trends Microbiol 2003, 11, 94–100.
- Phuong, K.; Kakii, K.; Nikata, T. Intergeneric coaggregation of non-flocculating Acinetobacter spp. isolates with other sludge-constituting bacteria. J. Biosci. Bioeng 2009, 107, 394–400.
- Joe, M.M.; Jaleel, C.A.; Sivakumar, P.K.; Zhao, C.X.; Karthikeyan, B. Co-aggregation in Azospirillum brasilensense MTCC-125 with other PGPR strains: Effect of physical and chemical factors and stress endurance ability. J. Taiwan Inst. Chem. E 2009, 40, 491–499.
- Malik, A.; Kakii, K. Pair dependent co-aggregation behavior of nonflocculating sludge bacteria. Biotechnol Lett 2003, 25, 981–986.
- Ledder, R.G.; Timperley, A.S.; Friswell, M.K.; MacFarlane, S.; McBain, A.J. Coaggregation between and among human intestinal and oral bacteria. FEMS Microbiol. Ecol 2008, 66, 630–636.
|Table 1. Production, composition, and functional roles of exopolysaccharides (EPSs) in beneficial and pathogenic plant-associated bacteria.|
|Bacteria/plant association||Exopolysaccharide||Chemical composition||Function|
|E. meliloti symbiosis with Medicago sativa||Succinoglycan (EPS I), LMW-HMW ||Octasaccharide units (glucose:galactose 7:1, bearing succinyl, acetyl, and pyruvyl substituents)||Required for biofilm formation |
EPS I LMW symbiotically active 
|Galactoglucan (EPS II), LMW-HMW ||Disaccharide units (acetylated glucose-pyruvylated galactose)||EPS II LMW symbiotically active |
EPS II LMW controls biofilm formation 
|R. leguminosarum symbiosis with Trifolium, Pisum, Vicia and Phaseolus spp.||Acidic EPS [95,96]||Octasaccharide units (glucose:glucuronic acid:galactose 5:2:1, modified by acetyl, pyruvyl and 3-hydroxybutanoyl groups)||Development of a structured biofilm [83,97]|
Required for infection and nodulation [98,99]
|B. japonicum symbiosis with Glycine max||EPS [100,101]||Pentasaccharide units (mannose:galacturonic acid:glucose:galactose 1:1:2:1)||Biofilm formation on both inert and biotic surfaces. Roles during the early stages of interaction with the host plant (initial attachment of rhizobia to root epidermal cells) |
|M. tianshanense symbiosis with Glycyrrhiza uralensis||EPS||ND||Involved in biofilm formation and successful establishment of symbiosis |
|A. tumefaciens ubiquitous plant pathogen||Succinoglycan ||See above||Increased production of succinoglycan results in reduced attachment and biofilm formation |
|X. fastidiosa plant pathogen||Putative Fastidian gum ||Putative tetrasaccharide units (glucose-1-phosphate, glucose, mannose, and glucuronic acid)||Possibly involved in bacterial pathogenicity |
Cell attachment and overall biofilm formation 
|X. campestris X. axonopodis plant pathogens||Xanthan gum ||Pentasaccharide units (glucose:mannose:glucuronic acid 2:2:1 derivatized with acetyl and pyruvyl moieties)||Essential for microcolony formation |
Formation of structured biofilms on abiotic surfaces and in infected plants [109,110]
|P. stewartii plant pathogen||Stewartan ||Heptasaccharide units (glucose:galactose:glucuronic acid 3:3:1)||Essential for appropriate adhesion and for maturation of biofilm structure. Also a virulence factor required for effective host colonization and efficient dissemination through xylem vessels |
|E. amylovora plant pathogen||Amylovoran ||Pentasaccharide units (galactose:glucose 4:1, and pyruvate residues)||Pathogenicity factor required for biofilm formation |
|Levan ||Homopolymer of fructose||Virulence factor. Also contributes to biofilm formation |
|R. solanacearum plant pathogen||Acidic EPS I ||Putative structure composed by N-acetylgalactosamine and amino sugars (bacillosamine, galactosaminuronic acid)||Major virulence factor |
LMW: low molecular weight; HMW: high molecular weight; ND: not determined.
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