In nature, most bacteria live attached to or in close association with surfaces. It has been estimated that at least 90% of all bacteria in the environment reside attached to a surface, and many of these form sessile communities called biofilms (34). Biofilm formation can be divided up in different stages, as follows. (i) Attachment of the bacteria to the surface in question. After the initial attachment, several possibilities are open. If the bacterium is attached to a eukaryotic cell it can sometimes initiate (ii) invasion of that cell. Alternatively, it can follow the path that ultimately leads to (iii) biofilmformation. This usually takes place via an initial expansion step resulting in microcolony formation and subsequently formation of a young biofilm. The final step is maturation into a mature biofilm, i.e., a complex three-dimensional structure. In this chapter we are primarily concerned about the first step in these developments, namely, bacterial attachment to surfaces. Bacterial attachment is a prelude to both invasion and biofilm formation. Without this crucial initial step the other developments will simply not happen.
Bacterial attachment to surfaces is mostly mediated by specific bacterial adhesins, surface structures that typically recognize specific molecular motifs in a lock-and-key fashion. This enables the bacterium to target to a specific surface, for example, a specific tissue like the bladder. This phenomenon is often referred to as tissue tropism. Escherichia coli and Salmonella strains can express a plethora of adhesins with different receptor specificities (80, 143). Individual strains are generally capable of expressing several different adhesins that can target to different receptor molecules. This provides the bacterium with a choice of potential target surfaces. Emerging evidence suggests that the expression of adhesins is often coordinated through inter system cross talk, offering flexibility in niche selection (133, 136). Here we will describe three examples of bacterial adhesins, each of which belongs to a different subgroup and follows different strategies for surface presentation and adhesin exposure. These are type 1 fimbriae, very long stiff rod-like organelles; curli, amorphous fluffy coat structures; and finally antigen 43, short outer membrane structures with a simple assembly system.
Type 1 fimbriae were originally characterized and defined by virtue of their ability to cause mannose-sensitive agglutination of a variety of eukaryotic cells (40, 41, 42). Type 1 fimbriae constitute the most widespread adhesive organelle among members of the Enterobacteriaceae. Closely related variants of these organelles have been reported to be present on E. coli, Klebsiella spp., and Shigella spp., whereas type 1 fimbriae of Salmonella and Citrobacter spp. constitute a second serological group (3). In E. coli∼80% of all strains have the capacity to express type 1 fimbriae. Arguably, the fact that these adhesive organelles are so common is indicative of important role(s) in the ecology of these bacteria.
A typical type 1-fimbriated bacterial cell has 100 to 500 fimbriae arranged peritrichously on the surface, each with a width of 7 nm and a length varying between 0.2 and 2 μm (Fig. 1). A type 1 fimbria has a tubular structure with a ∼2-nm-diameter hollow core (23, 56). The bulk of the organelle consists of a major building component, FimA (23, 73, 82). FimA monomers are noncovalently associated head-to-tail and organized in a right-helical structure with 3.1 subunits per revolution and a subunit pitch distance of 2.3 nm (23, 27, 56). Adjacent turns of the helix are connected via three binding sites, making the organelle rod rather stiff (56). At the tip of the structure a short, ~16-nm-thin, 2-nm-wide fibrillum is located (69). According to current knowledge the tip fibrillum is made of minor fimbrial components, namely, the FimF, FimG, and FimH proteins (56, 69). FimH is the type 1 fimbrial adhesin (81), and FimF and FimG act as adaptors for integration of the adhesin into the organelle structure (2, 69, 77, 82, 124). The subunits FimG and FimF connect FimH to the FimA rod, the sequential orientation being FimA–FimF–FimG–FimH (56). In addition to a tip fibril position, FimH has been reported to be interspersed along the fimbrial shaft (2, 3, 81). The detailed three-dimensional structure of FimH has been elucidated by X-ray crystallography (27). According to this the protein is folded into two domains, an N-terminal adhesive domain (residues 1 through 156) and a C-terminal organelle-integration domain (residues 160 through 279) linked by a tetra-peptide loop. An isolated truncated version of the N-terminal domain of FimH retains its adhesive faculty (129).
Type 1 fimbriae confer binding to a variety of eukaryotic cells by virtue of their capacity to recognize mannosides (42, 91, 161). The FimH adhesin is responsible for the adhesive properties of type 1 fimbriae and confers a lectin-like binding to various mannosides (81). Although FimH-mediated binding in general is sensitive to the presence of d-mannose, it has become increasingly clear that there exists significant heterogeneity among type 1 fimbriae. The FimH adhesins from different genera and species vary in their binding affinity towards defined oligomannose motifs. Striking differences were reported between the specificity of type 1 fimbriae from E. coli and Salmonella (44). Work by Sokurenko and others has clearly demonstrated that binding differences are due to alterations in the primary structures of different FimH vaiants (139, 140, 141). Other reports, although not contradicting this tenet, indicate that the affinity of FimH can be modified by the fimbrial filament on which it is presented (90, 149). It appears that changes in FimH specificity, whether caused by sequence variation in FimH or by influence from filament presentation, strongly affect target specificity and thereby tissue tropism. FimH variants from commensal isolates have weak affinity for monomannose residues but have strong affinity for complex mannose structures such as terminally exposed trimannose units. On the other hand, most uropathogenic isolates express FimH variants that have strong affinity for both types of targets (139). Minor amino acid sequence alterations in FimH can affect receptor recognition profile profoundly (114, 135, 139, 141, 142). The extent of the FimH target range can also be gathered from the fact that variants of this versatile adhesin are able to bind protein targets such as laminin and collagen (83, 114). The adaptable nature of the FimH receptor affinity was also investigated by creation of FimH libraries based on random mutagenesis of the FimH lectin domain. These studies revealed that FimH can exhibit a vast range of affinities for various targets and that even single amino acid alterations can change receptor specificity (135). The recent discovery of shear-force-induced binding enhancement of FimH further underlines the surprising flexibility of this adhesin. Computer simulations suggest that force-induced conformational changes are responsible for the phenomenon (150). Furthermore, regions in FimH distant from the receptor binding site seem to be involved. It is conceivable that force-induced conformational changes and binding enhancement can actually explain why the same FimH adhesin presented by different fimbrial filaments can adapt different receptor affinities.
Early studies of type 1 fimbriae revealed a connection between expression of these organelles and the ability to form a pellicle during growth in aerobic static broth, i.e., a bacterial surface film on the air-water interface (42). Pellicle formation was observed to be sensitive to d-mannose derivatives indicative of FimH involvement (100), which was subsequently verified by fimH-knockout mutations (59). Under static liquid conditions, type 1 fimbriae-assisted pellicle formation provides a dramatic selective advantage (in the order of 1:106) when comparing Fim+ and Fim– cells, arguably because of facilitated access to atmospheric oxygen (100, 144). In this connection it is important to note that members of the Enterobacteriaceae do have a life outside a mammalian host. Arguably, pellicle formation can provide a selective advantage in vivo under static liquid conditions, i.e., ponds, pools, etc. However, it is also conceivable that pellicle formation provides a means of host-to-host transfer: when mammals defecate in ponds and other stagnant water bodies, the excreted bacteria can subsequently form a pellicle on the surface by virtue of type 1 fimbriae expression. In turn the pellicle will provide a rich opportunity for transfer to a new host when other animals come to quench their thirst and drink from the surface.
A pellicle is essentially what we would now call an air-water biofilm. Seen in this light, it was hardly a surprise that manipulation of the function or expression of type 1 fimbriae turned out to interfere with biofilm formation on various abiotic surfaces such as plastics and glass (115, 131, 132). In line with the lessons learned from fimbriae involvement in pellicle formation, lesions that affected the fim genes or auxiliary functions such as periplasmic disulfide bridge formation were observed to reduce biofilm formation of E. coli on abiotic surfaces (49, 115). Again, the FimH adhesin seemed to play a prominent role since methyl mannoside was reported to inhibit biofilm formation (115).
In nature, bacteria often face brutal hydrodynamic flow-shear forces, for example in the urinary tract. We recently isolated FimH variants from a random mutant library that had the capacity to promote biofilm formation under hydrodynamic flow conditions (132). Some FimH variants were capable of biofilm enhancement even in the presence of methyl mannoside, and some promoted biofilm formation but were incapable of binding to mannoside targets. This suggests that mannose binding and biofilm enhancement can be independent properties of FimH. Several of the biofilm-promoting FimH variants were subsequently shown to confer autoaggregation of cells (127). Interestingly, similar phenotypes were also observed in wild-type variants of FimH originating from uropathogenic E. coli (UPEC) strains; i.e., they conferred biofilm formation and autoaggregation. However, these variants were always capable of binding to mannosides and all of them were sensitive to methyl mannoside (127, 132). Apparently there is strong selection in natural isolates for conservation of mannose recognition (to provide binding to host targets) and the additional ability to promote biofilm formation. Autoaggregation seems to be a trait acquired in, for example, some UPEC strains. Again, these observations underline the highly adaptive nature of the FimH adhesin.
As previously mentioned, the ability to express type 1 fimbriae is widespread and a conserved trait throughout the Enterobacteriaceae and underlines the importance of this adhesin in the ecology of these bacteria. In animal studies, type 1 fimbriae seem to play an important role in bacterial dissemination among litter mates. It was found that lesions affecting expression of the fim genes resulted in a dramatic decrease in transmission of E. coli among rats (19). This indicated a critical role for type 1 fimbriae-assisted oropharyngeal colonization in the fecal/oral cycle of enteric bacteria.
The excellent qualities of FimH in assisting bacterial adhesion and biofilm formation also hinted at a role for type 1 fimbriae in bacterial pathogenesis. Several studies in the 1990s strongly implicated type 1 fimbriae as virulence factors in urinary tract infections. Urinary tract infections by UPEC strains affect humans (and other mammals) and account for several million cases annually. Type 1 fimbriae are expressed by more than 90% of all UPEC strains, and they bind to the uroepithelial surface (5, 32, 33). It was demonstrated in a mouse model that type 1-fimbriated strains generally caused more severe infections than their Fim-negative counterparts and that inactivation of the fimH gene in a highly virulent UPEC strain rendered it virtually avirulent (33). While P fimbriae recognize kidney glycolipid receptors, type 1-fimbriated E. coli recognize uroplakins Ia and Ib, two major high-mannose-type glycoproteins of urothelial cells (161). Arguably, such binding would facilitate bacterial colonization of the bladder and cystitis. Further evidence implicating Fim involvement in cystitis included the observations that FimH-mediated binding to bladder cells conferred invasion of these cells, probably via FimH recognition of CD48 with ensuing internalization (10, 91, 96), and ultimately led to formation of intracellular pod-like bacterial aggregates (5). This phenomenon may account for the high amount of disease recurrence in many patients despite antibiotic treatment. Also, systemic vaccination with FimH component vaccines significantly reduces UPEC infection of the bladder in mice and monkeys (85, 86).
The fact that variants of FimH differ, often dramatically, in their affinity for target motifs, with accompanying change in tissue tropism, indicates that some variants have evolved and adapted to suit the need of specific pathogenic strains (e.g., UPEC strains). The first step in the colonization of the urinary tract by pathogenic E. coli is FimH-mediated binding to bladder epithelium. FimH variants from UPEC strains tend to exhibit a high affinity for monomannose targets, and this faculty enables binding to host cells in the urinary tract even in the presence of soluble molecular decoys such as Tamm-Horsfall glycoprotein (111). The affinity for trimannose units allows E. coli to bind, for example, buccal epithelial cells even in the presence of soluble mannose-containing molecules of saliva (139). This could be important in the transient colonization of the oropharyngeal mucosa that is thought to be a critical step for host-to-host transfer of E. coli (19). Type 1 fimbriae seem to be involved primarily in bladder infection and less in kidney infection, judging from adhesion affinity as well as in vivo expression (54, 68).
In summary, the FimH protein presented by type 1 fimbriae seems to be a highly versatile adhesin fulfilling a diverse spectrum of roles ranging from pellicle and biofilm formation to being a bona fide virulence factor in UPEC strains, where it plays important roles in the manifestation of cystitis.
Curli are thin, coiled, aggregative, amyloid-like fibers first identified on the surface of E. coli (103) and also produced by Salmonella (30) as well as other Enterobacteriaceae including Shigella, Citrobacter, and Enterobacter spp. (125, 164). In Salmonella these organelles are referred to as thin aggregative fimbriae (Tafi) (29, 30, 120). Other designations previously used include GVVPQ and SEF17 fimbriae (30, 31). In this review we will refer to these structures collectively as curli.
Expression of curli has been reported for most enterohemorrhagic, enterotoxigenic, and sepsis strains of E. coli; strains of enteroinvasive E. coli and enteropathogenic E. coli (EPEC) do not appear to produce these fibers (12, 101). Curli are also produced by avian pathogenic E. coli (118). Some E. coli K-12 laboratory strains such as MC4100 and MG1655 do not express curli despite the fact that they contain functional copies of the relevant genes (30, 57, 101). This has typically been attributed to the presence of mutations that affect the regulation of curli gene expression. Most salmonellae produce curli, the best studied being Salmonella enterica serovar Enteritidis (29) and S. enterica serovar Typhimurium (123). Curli genes in Shigella spp. are typically nonfunctional (125).
Curli organelles are approximately 6- to 12-nm-wide fibers of various lengths and appear as a tangled and amorphous matrix extending 0.5 to 1 μm from the cell surface (26) (Fig. 2). Curli characteristically bind Congo red dye, allowingfor an initial rapidscreen for their presence on the cell surface (57). These fibers also bind to a variety of human serum and tissue proteins (6, 12, 57, 103, 104, 137).
Curli fibers are composed primarily of subunits of a major component, CsgA (also referred to as curlins). CsgB is the only additional structural component that has been identified within these fibers and is present along the filament in minor amounts (16). These two proteins are nearly identical in size and share 49% amino acid similarity (58). Both proteins are required for fiber polymerization, yet possess very different biochemical properties. The assembly of curli fibers occurs outside the cell via a process referred to as the extracellular nucleation pathway (143). Curli growth proceeds by the addition of subunits principally to the distal end of the growing fiber (58). The CsgA major subunit is secreted from the cell and polymerized on the surface, assisted by the nucleator protein CsgB (16, 58). Evidence for this observation was obtained through the construction and analysis of mutant derivatives. When colonies of CsgB– CsgA+ and CsgB+ CsgA– mutants are grown in close proximity to one another, precipitation of curli subunits occurs on the side of the CsgB+ CsgA– colony facing the CsgB– CsgA+ colony (58). The CsgEFG proteins appear to be involved in export of the curli subunits and are required for fiber formation and assembly (57). CsgG is an outer membrane-located lipoprotein that protects the CsgA and CsgB subunit proteins from proteolysis and serves as a curli assembly platform. Deletion of csgG results in abrogation of curli fiber formation (88). The precise roles of CsgE and CsgF are less well understood. Recent work suggests a chaperone-like function for CsgE and a nucleation function for CsgF in the fiber assembly process (26). Cells containing a mutation in the csgE gene are more susceptible to CsgA degradation, are affected in Congo red binding, and are partially defective in their ability to nucleate extracellular CsgA subunits. Likewise, a csgF mutant possesses a nucleation defect that indicates a role for CsgF in the extracellular polymerization of CsgA (26).
Curli possess a broad protein-binding capacity that contributes to enhanced virulence. A large number of human proteins interact with curli including laminin (101), fibronectin (103), plasminogen (137), human contact phase proteins (12), and MHC class I molecules (104). It is generally assumed that the CsgA subunit is responsible for mediating these interactions.
Two protein-binding regions of the CsgA subunit capable of activating the contact system have been identified, one in the NH2-terminal region and one in the COOH-terminal region (102). These regions possess a β-sheet conformation that is characteristic of other amyloid fibrils and may also participate in the polymerization of CsgA subunits.
Curli formation promotes two fundamental processes associated with biofilm formation: initial adhesion and cell-to-cell aggregation. A role for curli in the colonization of inert surfaces has been demonstrated (9, 156). In E. coli, mutations in the ompR gene (such as the ompR234 mutation) stimulate biofilm growth by activating the transcription of csgD (156). This mutation can even stimulate curli production in K-12 strains such as MG1655 that do not normally express these fibers (116). The mutated OmpR234 protein contains a leucine-to-arginine shift at position 43 and exerts a positive effect on curli synthesis (156). A model postulating how curli production is controlled during biofilm growth has been proposed (116). According to this,increase in curli synthesis (e.g., during conditions of low osmolarity) occurs via transcriptional activation of the csgD promoter by OmpR (or, more efficiently, by OmpR234). Alternatively, repression of curli synthesis occurs in response to Cpx pathway activation (via RpoS) during conditions including high osmolarity, curlin accumulation, or a combination of both. The csgDEFG-csgBA promoter region contains sequences with high similarity to the proposed binding site for CpxR (113). Direct binding of CpxR to both the csgD and csgB promoters results in the down-regulation of curli expression (116).
Several properties inherent in the production of curli fibers suggest they contribute to bacterial virulence. The expression of curli is frequently observed on virulent isolates of E. coli and Salmonella. As already mentioned, curli mediate binding to a large repertoire of human proteins and are associated with biofilm formation. Curli production enhances adherence to various intestinal epithelial cell lines and contributes to virulence in mice (155). Curliated E. coli O157 strains contain promoter alterations that enhance curli expression and are associated with increased pathogenesis in mice and invasion of Hep-2 cells (154). Curli also contribute to virulence in avian pathogenic E. coli by promoting adherence to intestinal cells (84).
Severe sepsis and septic shock are frequently caused by gram-negative bacteria, and several factors suggest a significant role for curli during E. coli sepsis. First, E. coli strains isolated from patients with sepsis frequently express curli and these patients often also possess antibodies to CsgA (12, 15). Second, the assembly of components of the contact phase system on the surface of curliated bacteria triggers proinflammatory and procoagulatory cascades (such as the induction of proinflammatory cytokines) that may contribute to these disease states (67). Third, curliated E. coli sepsis bacteria are highly pathogenic when injected into mice or rats and induce symptoms often associated with severe sepsis, such as bleeding, lung disorders, and fall in blood pressure (17, 67, 112). Finally, high-affinity binding of curli to fibronectin is associated with invasion of eukaryotic cells (53). Curli genes from virulent septicemic E. coli isolates mediate enhanced internalization in comparison to their K-12 counterparts, suggesting differences in structure or regulation of expression between the loci originating from the two strain types (53).
Ag43 is a member of the family of autotransporter proteins. These are characterized by the fact that all information required for traverse of the bacterial membrane system and final routing to the surface resides in the protein itself. Autotransporter proteins possess some general features: an N-terminal signal sequence, the passenger (α) domain that is secreted to the cell surface, and a transporter (β) domain that forms a β-barrel pore, which assists the passenger domain in gaining access to the surface. Many autotransporters are virulence factors in gram-negative bacteria (63, 64). Ag43 is produced as a 1,039-amino-acid preprotein. An N-terminal signal peptide (52 amino acids) directs translocation across the cytoplasmic membrane to the periplasm. Ag43 consists of two subunits, α and β, comprising 499 and 488 amino acids, respectively. The α subunit (Ag43α) or passenger domain is presented on the cell surface via β-subunit-assisted crossing of the outer membrane. Subsequently, or during this transfer, the peptide bond linking the two domains is cleaved; all available evidence indicates that this happens by autocatalytic action (65). A sequence, LADSGAAVSGT, resembling a consensus sequence of an aspartyl protease active site is located in Ag43α (65). However, we recently mutagenized this sequence and correct processing still occurred, ruling out the potential role of this sequence in Ag43 processing (our unpublished results). Meanwhile, this observation does not in any way exclude Ag43 autocatalytic cleavage, and several other possible candidates for internal catalytic sites exist. The β subunit has all the hallmarks of a β-barrel porin and, in line with other autotransporters, presumably forms a pore through which Ag43α gains access to the surface (65). The α subunit remains attached to the cell surface via noncovalent interactions with the β subunit, but can easily be detached by brief heat treatment (65). The simplicity of the autotransporter mode of secretion is also reflected in the amenability by which Ag43 can be expressed in a wide range of gram-negative bacteria (70, 71). In all the different gram-negative bacteria in which we have expressed Ag43, we have observed correct processing into the α and β moieties; this also supports the notion that Ag43 processing is autocatalytic.
Autotransporter proteins are generally highly similar with respect to the structure of the β module, whereas they differ substantially in their α module (63). Generally, the β modules of the autotransporter proteins have between 10 and 18 (an even number) membrane-spanning amphiphatic β sheets (64). In accordance with these general features, the β module of Ag43 was modeled to consist of an 18-stranded β-barrel module (65).
Diderichsen reported that cells that were able to fluff were nonfimbriated and vice versa (38). This initial observation was later followed up by Hasman et al. (60), who demonstrated that the presence of fimbriae sterically blocks Ag43-mediated cell aggregation. It was subsequently shown that Ag43 and the type 1 class of fimbriae interact not only at the physical level but also at the transcriptional level through molecular cross-talk. Synthesis of type 1 and related fimbriae abrogates Ag43 expression; this effect is not observed in an oxyR mutant (133). The major fimbrial subunit, FimA, constitutes the most abundant phase variable surface protein in E. coli with up to 5 × 105 copies. In contrast to many other surface proteins including Ag43, FimA contains a cysteine bridge. During fimbria biosynthesis and export to the surface, disulfide bridge formation takes place at a very high level. Conceivably, this could affect the cell's thiol-disulfide status; thus fimbriae expression per se constitutes a signal transduction mechanism that affects expression of the flu gene, arguably via the oxidoreductive state of OxyR (133). Also, addition of a simple reducing agent, dithiothreitol, to cells negatively affects Ag43 expression (130, 133).
Interestingly, expression of P and type 1 fimbriae also seems to be coordinated through intersystem cross-talk via PapB, in which P fimbriation dominates (162). Thus the expressions of P and type 1 fimbriae and Ag43 are highly coordinated processes with a hierarchic structure. P fimbration is dominant to type 1 fimbriation, and fimbriation in general is dominant to Ag43 expression. The fimbrial phase "on" allows bacterial attachment to various epithelial targets, and the differential expression of type 1 and P fimbriae through cross-talk allows differential target and niche selection and cell invasion. The fimbrial phase "off" allows expression of Ag43, which promotes enhanced microcolony and biofilm formation (see later).
Ag43 expression was early on observed to correlate with cell-to-cell aggregation, fluffing, and settling of cells from static liquid suspensions. This faculty was later demonstrated to be based on intercellular Ag43-to-Ag43 recognition (60, 71). Thus, Ag43 is exceptional in being a self-recognizing adhesin; i.e., both receptor recognition and receptor target are provided in the same polypeptide. As previously mentioned, E. coli is capable of autoaggregation, and several different systems other than Ag43 are also able to independently produce this phenotype. As previously discussed, curli-mediated autoaggregation occurs via intercellular fiber precipitation. Bundle-forming pili (BFP) are a type IV class of fimbriae, produced by EPEC strains, that emanate from the cell surface and align along their longitudinal axes to form bundles of filaments (52). Expression of BFP mediates two phenotypes thought to play a role in colonization: autoaggregation in liquid cultures and localized adherence on tissue culture cell monolayers (18). In enteroaggregative E. coli strains, two flexible 2- to 3-nm-wide fimbrial types, designated aggregative adherence fimbriae I and II (AAF/I and AAF/II), have been identified (35, 97). The aggregative adherence phenotype is distinguished by prominent autoagglutination of bacterial cells to each other (98). Variants of the FimH adhesin of type 1 fimbriae have also been shown to promote cell aggregation in laboratory and wild-type background strains (127, 132). In contrast to other aggregating systems, the self-recognizing Ag43 adhesin is anchored directly to the outer membrane. Thus Ag43-mediated aggregation results in a more intimate cell-cell contact than seen with systems in which the intercellular interactions are based on polymeric structures that reach far out from the bacterial surface, i.e., fimbriae and curli.
Several lines of circumstantial evidence implicate the α subunit of Ag43 to be involved in autoaggregation. First, cells depleted in α but not β subunit do not autoaggregate. Second, addition of crude preparations of Ag43α to Ag43-expressing cells reduces autoaggregation significantly. Finally, cells treated with antiserum specific for the α subunit do not autoaggregate (our unpublished data). In a recent study on wild-type variants of Ag43 that we expressed in a defined K-12 background, three out of nine investigated Ag43 variants did not confer aggregation (78). However, these versions were expressed in normal quantities on the surface and gave rise to frizzy colonies, indicating that cell aggregation is not involved in the characteristic frizzy phenotype associated with Ag43. By employing a combination of linker insertion mutagenesis and domain swapping between aggregating and nonaggregating versions of Ag43, we pinpointed the region responsible for autoaggregation to be located within the N-terminal one-third of the passenger domain (78). Ag43 aggregation was strongly affected by pH. Aggregation was optimal at neutral and weakly acidic pH and abolished at pH values of <3 and >10. Also, high NaCl concentration inhibited autoaggregation. Taken together, the data indicate that ionic interactions between charged basic and acidic side chains play a role in Ag43-Ag43 self-recognition. The N-terminal segment of Ag43α contains numerous basic and acidic amino acids.
In addition to self-recognition, Ag43 has also been reported to confer weak binding to certain human cell line cells such as HEp-2 cells (66, 110). In this context it should be mentioned that a recent investigation could not confirm any affinity towards HeLa cells when the aggregation-positive Ag43 variant from the EDL933 EHEC strain was investigated (152). Interestingly, the same Ag43 variant, also called Cah (short for calcium-binding Ag43 homologue) was found to be capable of binding Ca2+ (152).
A biofilm mode of growth offers many advantages to a bacterial population compared to planktonic growth, such as a high level of resistance against predation and various antimicrobial agents. It has been hypothesized that phase-variable regulation of bacterial adhesins may be an important theme in virulence and biofilm formation (66, 115). It can be argued that it is in the interest of a bacterial species to maintain a fraction of the population that are primed to initiate biofilm formation under distinct conditions. In that way bacteria are always capable of immediately initiating biofilm formation when appropriate conditions are met, but they do not need to spend excessive amounts of energy by priming the cells of the entire population for such tasks. The phase-variable expression of Ag43 fits neatly into this picture.
The attachment of bacteria to a surface often results in proliferation into more complex microcolony structures. Indeed, bacterial aggregation and microcolony formation can be seen as a prelude to biofilm formation. In line with type 1 fimbriae and curli, Ag43 is implicated in aggregation and microcolony formation (61). It was therefore not too surprising that several groups have shown that Ag43 expression enhances biofilm formation of E. coli on various abiotic surfaces (Fig. 3) (36, 71, 72, 152). Recently, in a microarray-based study on global gene expression in E. coli biofilms, it was demonstrated that Ag43 expression is specifically upregulated during sessile growth when compared to both exponential and stationary planktonic cultures (131). It is also interesting to note that a survey of the biofilm-forming faculty of a diverse group of Ag43 variants showed that all of them enhanced biofilm growth, albeit with different efficacy (78). The ability of Ag43 to augment biofilm formation was recently shown to be sterically blocked by bulky surface structures such as capsules (128). It remains to be determined whether or not capsule expression is coordinated with Ag43 expression.
Many bacteria, notably pathogens, are known to have the ability to form aggregates in vitro and in vivo. Examples include a diverse range of bacteria of both gram-positive and -negative origin such as Bordetella pertussis (94), Mycobacterium tuberculosis (95), Staphylococcus aureus (93), and Streptococcus pyogenes (47). Such aggregates are known to be able to resist various host defenses, e.g., complement attack and phagocytosis, more efficiently than solitary bacteria (14, 99). In a study on S. pyogenes it was reported that mice infected with coaggregated cells more frequently developed abscesses, indicating that virulence is enhanced by aggregation (99). These observations lend strong support to the notion that aggregation is an important virulence mechanism. The formation of aggregates usually takes place through autoaggregation of cells. In a few cases the underlying molecular mechanism is known, and often self-recognizing surface proteins are responsible. A well-studied example of this phenomenon is the autoaggregation of S. pyogenes through intercellular interactions between pairs of protein H (47). On this background it can be argued with some justification that the ability of Ag43 to enhance bacterial aggregation and biofilm formation in itself constitutes a virulence trait since these phenotypes are often related to enhanced virulence. Recently, however, Ag43 was shown to be expressed in vivo during formation of intracellular bacterial aggregates or pods in bladder cells (5). It was suggested that the bacteria were in a quiescent state in these aggregates and were out of reach for killer cells and the immune defenses of the host. This interesting observation implicates Ag43 as a virulence factor in UPEC strains.
The protection provided by Ag43-mediated aggregation was also underlined in another series of experiments addressing the role of Ag43 in protection against oxidizing agents. It was speculated that the tight packing of cells in cellular aggregates connected with Ag43 expression could provide a mechanism for reducing local oxygen concentrations, thereby protecting the cells from damage caused by oxidizing agents. We found strong support for this hypothesis in the fact that Ag43 expression and ensuing cell-to-cell aggregation was observed to be concomitant with a high degree of protection against H2O2 killing (130).
A picture of the Ag43 family of autotransporters is slowly emerging. Several lines of evidence implicate this family in bacterial virulence, as follows. (i) Many, but not all, Ag43s are self-recognizing adhesins conferring cell aggregation and biofilm formation, both of which are defensive phenotypic traits related to increased virulence. (ii) Some Ag43s seem to confer a low level of adhesion to mammalian cells. (iii) Ag43 expression undergoes phase variation controlled by OxyR and Dam, not only in K-12 but also in wild-type strains (119). Phase variation has notoriously been linked to many bacterial virulence factors (66); additionally, Dam is known to regulate the expression of numerous virulence factors (89). It is hoped that future research will unravel more aspects of this group of autotransporter proteins.
Type 1 fimbriae, curli, and Ag43 are structurally different bacterial surface structures and follow completely different strategies for surface display and assembly. However, despite these differences they have several functional traits in common: (i) they are adhesins; (ii) members of all three families promote bacterial aggregation; (iii) they confer biofilm formation; (iv) members of all three families are linked with enhanced virulence; and (v) their expression promotes distinct colony morphology types. In effect, in spite of their physical differences, these surface structures are able to fulfil many of the same functions and are prime examples of convergent functional evolution. Meanwhile, despite these similarities each system has its own unique properties. For example, only type 1 fimbriae can bind to mannosides, and while Ag43 function is shielded by capsule type 1, fimbriae are not. Future work will undoubtedly identify additional virulence properties associated with these surface structures.
1. Abraham, J. M., C. S. Freitag, J. R. Clements, and B. I. Eisenstein. 1985. An invertible element of DNA controls phase variation of type 1 fimbriae of Escherichia coli. Proc. Natl. Acad. Sci. USA 82:5724-5727.[PubMed] [CrossRef]
2. Abraham, S. N., J. D. Goguen, D. Sun, P. Klemm, and E. H. Beachey. 1987. Identification of two ancillary subunits of Escherichia coli type 1 fimbriae by using antibodies against synthetic oligopeptides of fim gene products. J. Bacteriol. 169:5530-5536.[PubMed]
3. Abraham, S. N., D. Sun, J. B. Dale, and E. H. Beachey. 1988. Conservation of the d-mannose-adhesion protein among type 1 fimbriated members of the family Enterobacteriaceae. Nature 336:682-684.[PubMed] [CrossRef]
4. Allen-Vercoe, E., R. Collighan, and M. J. Woodward. 1998. The variant rpoS allele of S. enteritidis strain 27655R does not affect virulence in a chick model nor constitutive curliation but does generate a cold-sensitive phenotype. FEMS Microbiol. Lett. 167:245-253.[PubMed] [CrossRef]
5. Anderson, G. G., J. J. Palermo, J. D. Schilling, R. Roth, J. Heuser, and S. J. Hultgren. 2003. Intracellular bacterial biofilm-like pods in urinary tract infections. Science 301:105-107.[PubMed] [CrossRef]
6. Arnqvist, A., A. Olsen, and S. Normark. 1994. Sigma S-dependent growth-phase induction of the csgBA promoter in Escherichia coli can be achieved in vivo by sigma 70 in the absence of the nucleoid-associated protein H-NS. Mol. Microbiol. 13:1021-1032.[PubMed] [CrossRef]
7. Arnqvist, A., A. Olsen, J. Pfeifer, D. G. Russell, and S. Normark. 1992. The Crl protein activates cryptic genes for curli formation and fibronectin binding in Escherichia coli HB101. Mol. Microbiol. 6:2443-2452.[PubMed]
8. Aslund, F., M. Zheng, J. Beckwith, and G. Storz. 1999. Regulation of the OxyR transcription factor by hydrogen peroxide and the cellular thiol-disulfide status. Proc. Natl. Acad. Sci. USA 96:6161-6165.[PubMed] [CrossRef]
9. Austin, J. W., G. Sanders, W. W. Kay, and S. K. Collinson. 1998. Thin aggregative fimbriae enhance Salmonella enteritidis biofilm formation. FEMS Microbiol. Lett. 162:295-301.[PubMed] [CrossRef]
10. Baorto, D. M., Z. Gao, R. Malaviya, M. L. Dustin, A. van der Merwe, D. M. Lublin, and S. N. Abraham. 1997. Survival of FimH-expressing enterobacteria in macrophages relies on glycolipid traffic. Nature 389:636-639.[PubMed] [CrossRef]
11. Barnhart, M. M., J. S. Pinkner, G. E. Soto, F. G. Sauer, S. Langermann, G. Waksman, C. Frieden, and S. J. Hultgren. 2000. PapD-like chaperones provide the missing information for folding of pilin proteins. Proc. Natl. Acad. Sci. USA 97:7709-7714.[PubMed] [CrossRef]
12. Ben Nasr, A., A. Olsen, U. Sjobring, W. Muller-Esterl, and L. Bjorck. 1996. Assembly of human contact phase proteins and release of bradykinin at the surface of curli-expressing Escherichia coli. Mol. Microbiol. 20:927-935.[PubMed] [CrossRef]
13. Benz, I., and M. A. Schmidt. 1989. Cloning and expression of an adhesin (AIDA-I) involved in diffuse adherence of enteropathogenic Escherichia coli. Infect. Immun. 57:1506-1511.[PubMed]
14. Berge, A., B. M. Kihlberg, A. G. Sjoholm, and L. Bjorck. 1997. Streptococcal protein H forms soluble complement-activating complexes with IgG, but inhibits complement activation by IgG-coated targets. J. Biol. Chem. 272:20774-20781.[PubMed] [CrossRef]
15. Bian, Z., A. Brauner, Y. Li, and S. Normark. 2000. Expression of and cytokine activation by Escherichia coli curli fibers in human sepsis. J. Infect. Dis. 181:602-612.[PubMed] [CrossRef]
16. Bian, Z., and S. Normark. 1997. Nucleator function of CsgB for the assembly of adhesive surface organelles in Escherichia coli. EMBO J. 16:5827-5836.[PubMed] [CrossRef]
17. Bian, Z., Z. Q. Yan, G. K. Hansson, P. Thoren, and S. Normark. 2001. Activation of inducible nitric oxide synthase/nitric oxide by curli fibers leads to a fall in blood pressure during systemic Escherichia coli infection in mice. J. Infect. Dis. 183:612-619.[PubMed] [CrossRef]
18. Bieber, D., S. W. Ramer, C. Y. Wu, W. J. Murray, T. Tobe, R. Fernandez, and G. K. Schoolnik. 1998. Type IV pili, transient bacterial aggregates, and virulence of enteropathogenic Escherichia coli. Science 280:2114-2118.[PubMed] [CrossRef]
19. Bloch, C. A., B. A. Stocker, and P. E. Orndorff. 1992. A key role for type 1 pili in enterobacterial communicability. Mol. Microbiol. 6:697-701.[PubMed] [CrossRef]
20. Blomfield, I. C. 2001. The regulation of pap and type 1 fimbriation in Escherichia coli. Adv. Microb. Physiol. 45:1-49.[PubMed] [CrossRef]
21. Blomfield, I. C., P. J. Calie, K. J. Eberhardt, M. S. McClain, and B. I. Eisenstein. 1993. Lrp stimulates phase variation of type 1 fimbriation in Escherichia coli K-12. J. Bacteriol. 175:27-36.[PubMed]
22. Blomfield, I. C., D. H. Kulasekara, and B. I. Eisenstein. 1997. Integration host factor stimulates both FimB- and FimE-mediated site-specific DNA inversion that controls phase variation of type 1 fimbriae expression in Escherichia coli. Mol. Microbiol. 23:705-717.[PubMed] [CrossRef]
23. Brinton, C. C., Jr. 1965. The structure, function, synthesis and genetic control of bacterial pili and a molecular model for DNA and RNA transport in gram negative bacteria. Trans. NY Acad. Sci. 27:1003-1054.
24. Brinton, C. C. J., P Gemski, Jr., S. Falkow, and L. S. Baron. 1961. Location of the piliation factor on the chromosome of Escherichia coli. Biochem. Biophys. Res. Commun. 5:293-298. [CrossRef]
25. Brown, P. K., C. M. Dozois, C. A. Nickerson, A. Zuppardo, J. Terlonge, and R. Curtiss III. 2001. MlrA, a novel regulator of curli (AgF) and extracellular matrix synthesis by Escherichia coli and Salmonella enterica serovar Typhimurium. Mol. Microbiol. 41:349-363.[PubMed] [CrossRef]
26. Chapman, M. R., L. S. Robinson, J. S. Pinkner, R. Roth, J. Heuser, M. Hammar, S. Normark, and S. J. Hultgren. 2002. Role of Escherichia coli curli operons in directing amyloid fiber formation. Science 295:851-855.[PubMed] [CrossRef]
27. Choudhury, D., A. Thompson, V. Stojanoff, S. Langermann, J. Pinkner, S. J. Hultgren, and S. D. Knight. 1999. X-ray structure of the FimC-FimH chaperone-adhesin complex from uropathogenic Escherichia coli. Science 285:1061-1066.[PubMed] [CrossRef]
28. Christman, M. F., G. Storz, and B. N. Ames. 1989. OxyR, a positive regulator of hydrogen peroxide-inducible genes in Escherichia coli and Salmonella typhimurium, is homologous to a family of bacterial regulatory proteins. Proc. Natl. Acad. Sci. USA 86:3484-3488.[PubMed] [CrossRef]
29. Collinson, S. K., S. C. Clouthier, J. L. Doran, P. A. Banser, and W. W. Kay. 1996. Salmonella enteritidis agfBAC operon encoding thin, aggregative fimbriae. J. Bacteriol. 178:662-667.[PubMed]
30. Collinson, S. K., L. Emody, K. H. Muller, T. J. Trust, and W. W. Kay. 1991. Purification and characterization of thin, aggregative fimbriae from Salmonella enteritidis. J. Bacteriol. 173:4773-4781.[PubMed]
31. Collinson, S. K., L. Emody, T. J. Trust, and W. W. Kay. 1992. Thin aggregative fimbriae from diarrheagenic Escherichia coli. J. Bacteriol. 174:4490-4495.[PubMed]
32. Connell, H., L. K. Poulsen, and P. Klemm. 2000. Expression of type 1 and P fimbriae in situ and localisation of a uropathogenic Escherichia coli strain in the murine bladder and kidney. Int. J. Med. Microbiol. 290:587-597.[PubMed]
33. Connell, I., W. Agace, P. Klemm, M. Schembri, S. Marild, and C. Svanborg. 1996. Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract. Proc. Natl. Acad. Sci. USA 93:9827-9832.[PubMed] [CrossRef]
34. Costerton, J. W., P. S. Stewart, and E. P. Greenberg. 1999. Bacterial biofilms: a common cause of persistent infections. Science 284:1318-1322.[PubMed] [CrossRef]
35. Czeczulin, J. R., S. Balepur, S. Hicks, A. Phillips, R. Hall, M. H. Kothary, F. Navarro-Garcia, and J. P. Nataro. 1997. Aggregative adherence fimbria II, a second fimbrial antigen mediating aggregative adherence in enteroaggregative Escherichia coli. Infect. Immun. 65:4135-4145.[PubMed]
36. Danese, P. N., L. A. Pratt, S. L. Dove, and R. Kolter. 2000. The outer membrane protein, antigen 43, mediates cell-to-cell interactions within Escherichia coli biofilms. Mol. Microbiol. 37:424-432.[PubMed] [CrossRef]
37. Dibb-Fuller, M., E. Allen-Vercoe, M. J. Woodward, and C. J. Thorns. 1997. Expression of SEF17 fimbriae by Salmonella enteritidis. Lett. Appl. Microbiol. 25:447-452.[PubMed] [CrossRef]
38. Diderichsen, B. 1980. flu, a metastable gene controlling surface properties of Escherichia coli. J. Bacteriol. 141:858-867.[PubMed]
39. Dorel, C., O. Vidal, C. Prigent-Combaret, I. Vallet, and P. Lejeune. 1999. Involvement of the Cpx signal transduction pathway of E. coli in biofilm formation. FEMS Microbiol. Lett. 178:169-175.[PubMed] [CrossRef]
40. Duguid, J. P., and R. R. Gillies. 1958. Fimbriae and haemagglutinating activity in Salmonella, Klebsiella, Proteus and Chromobacterium. J. Pathol. Bacteriol. 75:519-520.
41. Duguid, J. P., and E. S. Anderson. 1967. Terminology of bacterial fimbriae, or pili, and their types. Nature 215:89-90.[PubMed] [CrossRef]
42. Duguid, J. P., E. S. Anderson, and I. Campbell. 1966. Fimbriae and adhesive properties in Salmonellae. J. Pathol. Bacteriol. 92:107-138.[PubMed] [CrossRef]
43. Emsley, P., I. G. Charles, N. F. Fairweather, and N. W. Isaacs. 1996. Structure of Bordetella pertussis virulence factor P.69 pertactin. Nature 381:90-92.[PubMed] [CrossRef]
44. Firon, N., S. Ashkenazi, D. Mirelman, I. Ofek, and N. Sharon. 1987. Aromatic alpha-glycosides of mannose are powerful inhibitors of the adherence of type 1 fimbriated Escherichia coli to yeast and intestinal epithelial cells. Infect. Immun. 55:472-476.[PubMed]
45. Force, A., M. Lynch, F. B. Pickett, A. Amores, Y. L. Yan, and J. Postlethwait. 1999. Preservation of duplicate genes by complementary, degenerative mutations. Genetics 151:1531-1545.[PubMed]
46. Freitag, C. S., and B. I. Eisenstein. 1983. Genetic mapping and transcriptional orientation of the fimD gene. J. Bacteriol. 156:1052-1058.[PubMed]
47. Frick, I. M., M. Morgelin, and L. Bjorck. 2000. Virulent aggregates of Streptococcus pyogenes are generated by homophilic protein-protein interactions. Mol. Microbiol. 37:1232-1247.[PubMed] [CrossRef]
48. Gally, D. L., J. Leathart, and I. C. Blomfield. 1996. Interaction of FimB and FimE with the fim switch that controls the phase variation of type 1 fimbriae in Escherichia coli K-12. Mol. Microbiol. 21:725-738.[PubMed] [CrossRef]
49. Genevaux, P., P. Bauda, M. S. DuBow, and B. Oudega. 1999. Identification of Tn10 insertions in the dsbA gene affecting Escherichia coli biofilm formation. FEMS Microbiol. Lett. 173:403-409.[PubMed] [CrossRef]
50. Gerstel, U., C. Park, and U. Romling. 2003. Complex regulation of csgD promoter activity by global regulatory proteins. Mol. Microbiol. 49:639-654.[PubMed] [CrossRef]
51. Gerstel, U., and U. Romling. 2001. Oxygen tension and nutrient starvation are major signals that regulate agfD promoter activity and expression of the multicellular morphotype in Salmonella typhimurium. Environ. Microbiol. 3:638-648.[PubMed] [CrossRef]
52. Giron, J. A., A. S. Ho, and G. K. Schoolnik. 1991. An inducible bundle-forming pilus of enteropathogenic Escherichia coli. Science 254:710-713.[PubMed] [CrossRef]
53. Gophna, U., M. Barlev, R. Seijffers, T. A. Oelschlager, J. Hacker, and E. Z. Ron. 2001. Curli fibers mediate internalization of Escherichia coli by eukaryotic cells. Infect. Immun. 69:2659-2665.[PubMed] [CrossRef]
54. Gunther, N. W. T., V. Lockatell, D. E. Johnson, and H. L. Mobley. 2001. In vivo dynamics of type 1 fimbria regulation in uropathogenic Escherichia coli during experimental urinary tract infection. Infect. Immun. 69:2838-2846.[PubMed] [CrossRef]
55. Haagmans, W., and M. van der Woude. 2000. Phase variation of Ag43 in Escherichia coli: Dam-dependent methylation abrogates OxyR binding and OxyR-mediated repression of transcription. Mol. Microbiol. 35:877-887.[PubMed] [CrossRef]
56. Hahn, E., P. Wild, U. Hermanns, P. Sebbel, R. Glockshuber, M. Haner, N. Taschner, P. Burkhard, U. Aebi, and S. A. Muller. 2002. Exploring the 3D molecular architecture of Escherichia coli type 1 pili. J. Mol. Biol. 323:845-857.[PubMed] [CrossRef]
57. Hammar, M., A. Arnqvist, Z. Bian, A. Olsen, and S. Normark. 1995. Expression of two csg operons is required for production of fibronectin- and congo red-binding curli polymers in Escherichia coli K-12. Mol. Microbiol. 18:661-670.[PubMed] [CrossRef]
58. Hammar, M., Z. Bian, and S. Normark. 1996. Nucleator-dependent intercellular assembly of adhesive curli organelles in Escherichia coli. Proc. Natl. Acad. Sci. USA 93:6562-6566.[PubMed] [CrossRef]
59. Harris, S. L., D. A. Elliott, M. C. Blake, L. M. Must, M. Messenger, and P. E. Orndorff. 1990. Isolation and characterization of mutants with lesions affecting pellicle formation and erythrocyte agglutination by type 1 piliated Escherichia coli. J. Bacteriol. 172:6411-6418.[PubMed]
60. Hasman, H., T. Chakraborty, and P. Klemm. 1999. Antigen-43-mediated autoaggregation of Escherichia coli is blocked by fimbriation. J. Bacteriol. 181:4834-4841.[PubMed]
61. Hasman, H., M. A. Schembri, and P. Klemm. 2000. Antigen 43 and type 1 fimbriae determine colony morphology of Escherichia coli K-12. J. Bacteriol. 182:1089-1095.[PubMed] [CrossRef]
62. Henderson, I. R., M. Meehan, and P. Owen. 1997. A novel regulatory mechanism for a novel phase-variable outer membrane protein of Escherichia coli. Adv. Exp. Med. Biol. 412:349-355.[PubMed]
63. Henderson, I. R., and J. P. Nataro. 2001. Virulence functions of autotransporter proteins. Infect. Immun. 69:1231-1243.[PubMed] [CrossRef]
64. Henderson, I. R., F. Navarro-Garcia, and J. P. Nataro. 1998. The great escape: structure and function of the autotransporter proteins. Trends Microbiol. 6:370-378.[PubMed] [CrossRef]
65. Henderson, I. R., and P. Owen. 1999. The major phase-variable outer membrane protein of Escherichia coli structurally resembles the immunoglobulin A1 protease class of exported protein and is regulated by a novel mechanism involving Dam and OxyR. J. Bacteriol. 181:2132-2141.[PubMed]
66. Henderson, I. R., P. Owen, and J. P. Nataro. 1999. Molecular switches—the ON and OFF of bacterial phase variation. Mol. Microbiol. 33:919-932.[PubMed] [CrossRef]
67. Herwald, H., M. Morgelin, A. Olsen, M. Rhen, B. Dahlback, W. Muller-Esterl, and L. Bjorck. 1998. Activation of the contact-phase system on bacterial surfaces—a clue to serious complications in infectious diseases. Nat. Med. 4:298-302.[PubMed] [CrossRef]
68. Hung, C. S., J. Bouckaert, D. Hung, J. Pinkner, C. Widberg, A. DeFusco, C. G. Auguste, R. Strouse, S. Langermann, G. Waksman, and S. J. Hultgren. 2002. Structural basis of tropism of Escherichia coli to the bladder during urinary tract infection. Mol. Microbiol. 44:903-915.[PubMed] [CrossRef]
69. Jones, C. H., J. S. Pinkner, R. Roth, J. Heuser, A. V. Nicholes, S. N. Abraham, and S. J. Hultgren. 1995. FimH adhesin of type 1 pili is assembled into a fibrillar tip structure in the Enterobacteriaceae. Proc. Natl. Acad. Sci. USA 92:2081-2085.[PubMed] [CrossRef]
70. Kjaergaard, K., H. Hasman, M. A. Schembri, and P. Klemm. 2002. Antigen 43-mediated autotransporter display, a versatile bacterial cell surface presentation system. J. Bacteriol. 184:4197-4204.[PubMed] [CrossRef]
71. Kjaergaard, K., M. A. Schembri, H. Hasman, and P. Klemm. 2000. Antigen 43 from Escherichia coli induces inter- and intraspecies cell aggregation and changes in colony morphology of Pseudomonas fluorescens. J. Bacteriol. 182:4789-4796.[PubMed] [CrossRef]
72. Kjaergaard, K., M. A. Schembri, C. Ramos, S. Molin, and P. Klemm. 2000. Antigen 43 facilitates formation of multispecies biofilms. Environ. Microbiol. 2:695-702.[PubMed] [CrossRef]
73. Klemm, P. 1984. The fimA gene encoding the type-1 fimbrial subunit of Escherichia coli. Nucleotide sequence and primary structure of the protein. Eur. J. Biochem. 143:395-399.[PubMed] [CrossRef]
74. Klemm, P. 1992. FimC, a chaperone-like periplasmic protein of Escherichia coli involved in biogenesis of type 1 fimbriae. Res. Microbiol. 143:831-838.[PubMed] [CrossRef]
75. Klemm, P. 1986. Two regulatory fim genes, fimB and fimE, control the phase variation of type 1 fimbriae in Escherichia coli. EMBO J. 5:1389-1393.[PubMed]
76. Klemm, P., and G. Christiansen. 1990. The fimD gene required for cell surface localization of Escherichia coli type 1 fimbriae. Mol. Gen. Genet. 220:334-338.[PubMed] [CrossRef]
77. Klemm, P., and G. Christiansen. 1987. Three fim genes required for the regulation of length and mediation of adhesion of Escherichia coli type 1 fimbriae. Mol. Gen. Genet. 208:439-445.[PubMed] [CrossRef]
78. Klemm, P., L. Hjerrild, M. Gjermansen, and M. A. Schembri. 2004. Structure-function analysis of the self-recognizing Antigen 43 autotransporter protein from Escherichia coli. Mol. Microbiol. 51:283-296.[PubMed] [CrossRef]
79. Klemm, P., B. J. Jorgensen, I. van Die, H. de Ree, and H. Bergmans. 1985. The fim genes responsible for synthesis of type 1 fimbriae in Escherichia coli, cloning and genetic organization. Mol. Gen. Genet. 199:410-414.[PubMed] [CrossRef]
80. Klemm, P., and M. A. Schembri. 2000. Bacterial adhesins: function and structure. Int. J. Med. Microbiol. 290:27-35.[PubMed]
81. Krogfelt, K. A., H. Bergmans, and P. Klemm. 1990. Direct evidence that the FimH protein is the mannose-specific adhesin of Escherichia coli type 1 fimbriae. Infect. Immun. 58:1995-1998.[PubMed]
82. Krogfelt, K. A., and P. Klemm. 1988. Investigation of minor components of Escherichia coli type 1 fimbriae: protein chemical and immunological aspects. Microb. Pathog. 4:231-238.[PubMed] [CrossRef]
83. Kukkonen, M., T. Raunio, R. Virkola, K. Lahteenmaki, P. H. Makela, P. Klemm, S. Clegg, and T. K. Korhonen. 1993. Basement membrane carbohydrate as a target for bacterial adhesion: binding of type I fimbriae of Salmonella enterica and Escherichia coli to laminin. Mol. Microbiol. 7:229-237.[PubMed] [CrossRef]
84. La Ragione, R. M., W. A. Cooley, and M. J. Woodward. 2000. The role of fimbriae and flagella in the adherence of avian strains of Escherichia coli O78:K80 to tissue culture cells and tracheal and gut explants. J. Med. Microbiol. 49:327-338.[PubMed]
85. Langermann, S., R. Mollby, J. E. Burlein, S. R. Palaszynski, C. G. Auguste, A. DeFusco, R. Strouse, M. A. Schenerman, S. J. Hultgren, J. S. Pinkner, J. Winberg, L. Guldevall, M. Soderhall, K. Ishikawa, S. Normark, and S. Koenig. 2000. Vaccination with FimH adhesin protects cynomolgus monkeys from colonization and infection by uropathogenic Escherichia coli. J. Infect. Dis. 181:774-778.[PubMed] [CrossRef]
86. Langermann, S., S. Palaszynski, M. Barnhart, G. Auguste, J. S. Pinkner, J. Burlein, P. Barren, S. Koenig, S. Leath, C. H. Jones, and S. J. Hultgren. 1997. Prevention of mucosal Escherichia coli infection by FimH-adhesin-based systemic vaccination. Science 276:607-611.[PubMed] [CrossRef]
87. Lindenthal, C., and E. A. Elsinghorst. 1999. Identification of a glycoprotein produced by enterotoxigenic Escherichia coli. Infect. Immun. 67:4084-4091.[PubMed]
88. Loferer, H., M. Hammar, and S. Normark. 1997. Availability of the fibre subunit CsgA and the nucleator protein CsgB during assembly of fibronectin-binding curli is limited by the intracellular concentration of the novel lipoprotein CsgG. Mol. Microbiol. 26:11-23.[PubMed] [CrossRef]
89. Low, D. A., N. J. Weyand, and M. J. Mahan. 2001. Roles of DNA adenine methylation in regulating bacterial gene expression and virulence. Infect. Immun. 69:7197-7204.[PubMed] [CrossRef]
90. Madison, B., I. Ofek, S. Clegg, and S. N. Abraham. 1994. Type 1 fimbrial shafts of Escherichia coli and Klebsiella pneumoniae influence sugar-binding specificities of their FimH adhesins. Infect. Immun. 62:843-848.[PubMed]
91. Martinez, J. J., M. A. Mulvey, J. D. Schilling, J. S. Pinkner, and S. J. Hultgren. 2000. Type 1 pilus-mediated bacterial invasion of bladder epithelial cells. EMBO J. 19:2803-2812.[PubMed] [CrossRef]
92. Maurer, J. J., T. P. Brown, W. L. Steffens, and S. G. Thayer. 1998. The occurrence of ambient temperature-regulated adhesins, curli, and the temperature-sensitive hemagglutinin tsh among avian Escherichia coli. Avian Dis. 42:106-118.[PubMed] [CrossRef]
93. McDevitt, D., P. Francois, P. Vaudaux, and T. J. Foster. 1994. Molecular characterization of the clumping factor (fibrinogen receptor) of Staphylococcus aureus. Mol. Microbiol. 11:237-248.[PubMed] [CrossRef]
94. Menozzi, F. D., P. E. Boucher, G. Riveau, C. Gantiez, and C. Locht. 1994. Surface-associated filamentous hemagglutinin induces autoagglutination of Bordetella pertussis. Infect. Immun. 62:4261-4269.[PubMed]
95. Menozzi, F. D., J. H. Rouse, M. Alavi, M. Laude-Sharp, J. Muller, R. Bischoff, M. J. Brennan, and C. Locht. 1996. Identification of a heparin-binding hemagglutinin present in mycobacteria. J. Exp. Med. 184:993-1001.[PubMed] [CrossRef]
96. Mulvey, M. A., Y. S. Lopez-Boado, C. L. Wilson, R. Roth, W. C. Parks, J. Heuser, and S. J. Hultgren. 1998. Induction and evasion of host defenses by type 1-piliated uropathogenic Escherichia coli. Science 282:1494-1497.[PubMed] [CrossRef]
97. Nataro, J. P., Y. Deng, D. R. Maneval, A. L. German, W. C. Martin, and M. M. Levine. 1992. Aggregative adherence fimbriae I of enteroaggregative Escherichia coli mediate adherence to HEp-2 cells and hemagglutination of human erythrocytes. Infect. Immun. 60:2297-2304.[PubMed]
98. Nataro, J. P., J. B. Kaper, R. Robins-Browne, V. Prado, P. Vial, and M. M. Levine. 1987. Patterns of adherence of diarrheagenic Escherichia coli to HEp-2 cells. Pediatr. Infect. Dis. J. 6:829-831.[PubMed] [CrossRef]
99. Ochiai, K., T. Kurita-Ochiai, Y. Kamino, and T. Ikeda. 1993. Effect of co-aggregation on the pathogenicity of oral bacteria. J. Med. Microbiol. 39:183-190.[PubMed] [CrossRef]
100. Old, D. C., and J. P. Duguid. 1970. Selective outgrowth of fimbriate bacteria in static liquid medium. J. Bacteriol. 103:447-456.[PubMed]
101. Olsen, A., A. Arnqvist, M. Hammar, S. Sukupolvi, and S. Normark. 1993. The RpoS sigma factor relieves H-NS-mediated transcriptional repression of csgA, the subunit gene of fibronectin-binding curli in Escherichia coli. Mol. Microbiol. 7:523-536.[PubMed] [CrossRef]
102. Olsen, A., H. Herwald, M. Wikstrom, K. Persson, E. Mattsson, and L. Bjorck. 2002. Identification of two protein-binding and functional regions of curli, a surface organelle and virulence determinant of Escherichia coli. J. Biol. Chem. 277:34568-34572.[PubMed] [CrossRef]
103. Olsen, A., A. Jonsson, and S. Normark. 1989. Fibronectin binding mediated by a novel class of surface organelles on Escherichia coli. Nature 338:652-655.[PubMed] [CrossRef]
104. Olsen, A., M. J. Wick, M. Morgelin, and L. Bjorck. 1998. Curli, fibrous surface proteins of Escherichia coli, interact with major histocompatibility complex class I molecules. Infect. Immun. 66:944-949.[PubMed]
105. Olsen, P. B., and P. Klemm. 1994. Localization of promoters in the fim gene cluster and the effect of H-NS on the transcription of fimB and fimE. FEMS Microbiol. Lett. 116:95-100.[PubMed] [CrossRef]
106. Olsen, P. B., M. A. Schembri, D. L. Gally, and P. Klemm. 1998. Differential temperature modulation by H-NS of the fimB and fimE recombinase genes which control the orientation of the type 1 fimbrial phase switch. FEMS Microbiol. Lett. 162:17-23.[PubMed] [CrossRef]
107. Orndorff, P. E., and S. Falkow. 1984. Organization and expression of genes responsible for type 1 piliation in Escherichia coli. J. Bacteriol. 159:736-744.[PubMed]
108. Owen, P. 1983. Antigens of the Escherichia coli cell envelope, p. 347-373. In O. J. Bjerrum (ed.), Electroimmunochemical Analysis of Membrane Proteins. Elsevier Science Publishing, Amsterdam, The Netherlands.
109. Owen, P. 1992. The gram-negative outer membrane: structure, biochemistry and vaccine potential. Biochem. Soc. Trans. 20:1-6.[PubMed]
110. Owen, P., M. Meehan, H. de Loughry-Doherty, and I. Henderson. 1996. Phase-variable outer membrane proteins in Escherichia coli. FEMS Immunol. Med. Microbiol. 16:63-76.[PubMed] [CrossRef]
111. Pak, J., Y. Pu, Z. T. Zhang, D. L. Hasty, and X. R. Wu. 2001. Tamm-Horsfall protein binds to type 1 fimbriated Escherichia coli and prevents E. coli from binding to uroplakin Ia and Ib receptors. J. Biol. Chem. 276:9924-9930.[PubMed] [CrossRef]
112. Persson, K., W. Russell, M. Morgelin, and H. Herwald. 2003. The conversion of fibrinogen to fibrin at the surface of curliated Escherichia coli bacteria leads to the generation of proinflammatory fibrinopeptides. J. Biol. Chem. 278:31884-31890.[PubMed] [CrossRef]
113. Pogliano, J., A. S. Lynch, D. Belin, E. C. Lin, and J. Beckwith. 1997. Regulation of Escherichia coli cell envelope proteins involved in protein folding and degradation by the Cpx two-component system. Genes Dev. 11:1169-1182.[PubMed] [CrossRef]
114. Pouttu, R., T. Puustinen, R. Virkola, J. Hacker, P. Klemm, and T. K. Korhonen. 1999. Amino acid residue Ala-62 in the FimH fimbrial adhesin is critical for the adhesiveness of meningitis-associated Escherichia coli to collagens. Mol. Microbiol. 31:1747-1757.[PubMed] [CrossRef]
115. Pratt, L. A., and R. Kolter. 1998. Genetic analysis of Escherichia coli biofilm formation: roles of flagella, motility, chemotaxis and type I pili. Mol. Microbiol. 30:285-293.[PubMed] [CrossRef]
116. Prigent-Combaret, C., E. Brombacher, O. Vidal, A. Ambert, P. Lejeune, P. Landini, and C. Dorel. 2001. Complex regulatory network controls initial adhesion and biofilm formation in Escherichia coli via regulation of the csgD gene. J. Bacteriol. 183:7213-7223.[PubMed] [CrossRef]
117. Prigent-Combaret, C., G. Prensier, T. T. Le Thi, O. Vidal, P. Lejeune, and C. Dorel. 2000. Developmental pathway for biofilm formation in curli-producing Escherichia coli strains: role of flagella, curli and colanic acid. Environ. Microbiol. 2:450-464.[PubMed] [CrossRef]
118. Provence, D. L., and R. Curtiss III. 1992. Role of crl in avian pathogenic Escherichia coli: a knockout mutation of crl does not affect hemagglutination activity, fibronectin binding, or curli production. Infect. Immun. 60:4460-4467.[PubMed]
119. Roche, A., J. McFadden, and P. Owen. 2001. Antigen 43, the major phase-variable protein of the Escherichia coli outer membrane, can exist as a family of proteins encoded by multiple alleles. Microbiology 147:161-169.[PubMed]
120. Romling, U., Z. Bian, M. Hammar, W. D. Sierralta, and S. Normark. 1998. Curli fibers are highly conserved between Salmonella typhimurium and Escherichia coli with respect to operon structure and regulation. J. Bacteriol. 180:722-731.[PubMed]
121. Romling, U., W. Bokranz, W. Rabsch, X. Zogaj, M. Nimtz, and H. Tschape. 2003. Occurrence and regulation of the multicellular morphotype in Salmonella serovars important in human disease. Int. J. Med. Microbiol. 293:273-285.[PubMed] [CrossRef]
122. Romling, U., M. Rohde, A. Olsen, S. Normark, and J. Reinkoster. 2000. AgfD, the checkpoint of multicellular and aggregative behaviour in Salmonella typhimurium, regulates at least two independent pathways. Mol. Microbiol. 36:10-23.[PubMed] [CrossRef]
123. Romling, U., W. D. Sierralta, K. Eriksson, and S. Normark. 1998. Multicellular and aggregative behaviour of Salmonella typhimurium strains is controlled by mutations in the agfD promoter. Mol. Microbiol. 28:249-264.[PubMed] [CrossRef]
124. Russell, P. W., and P. E. Orndorff. 1992. Lesions in two Escherichia coli type 1 pilus genes alter pilus number and length without affecting receptor binding. J. Bacteriol. 174:5923-5935.[PubMed]
125. Sakellaris, H., N. K. Hannink, K. Rajakumar, D. Bulach, M. Hunt, C. Sasakawa, and B. Adler. 2000. Curli loci of Shigella spp. Infect. Immun. 68:3780-3783.[PubMed] [CrossRef]
126. Sanderson, K. E., A. Hessel, and K. E. Rudd. 1995. Genetic map of Salmonella typhimurium, edition VIII. Microbiol. Rev. 59:241-303.[PubMed]
127. Schembri, M. A., G. Christiansen, and P. Klemm. 2001. FimH-mediated autoaggregation of Escherichia coli. Mol. Microbiol. 41:1419-1430.[PubMed] [CrossRef]
128. Schembri, M. A., D. Dalsgaard, and P. Klemm. 2004. Capsule shields the function of short bacterial adhesins. J. Bacteriol. 186:1249-1257.[PubMed] [CrossRef]
129. Schembri, M. A., H. Hasman, and P. Klemm. 2000. Expression and purification of the mannose recognition domain of the FimH adhesin. FEMS Microbiol. Lett. 188:147-151.[PubMed] [CrossRef]
130. Schembri, M. A., L. Hjerrild, M. Gjermansen, and P. Klemm. 2003. Differential expression of the Escherichia coli autoaggregation factor antigen 43. J. Bacteriol. 185:2236-2242.[PubMed] [CrossRef]
131. Schembri, M. A., K. Kjaergaard, and P. Klemm. 2003. Global gene expression in Escherichia coli biofilms. Mol. Microbiol. 48:253-267.[PubMed] [CrossRef]
132. Schembri, M. A., and P. Klemm. 2001. Biofilm formation in a hydrodynamic environment by novel FimH variants and ramifications for virulence. Infect. Immun. 69:1322-1328.[PubMed] [CrossRef]
133. Schembri, M. A., and P. Klemm. 2001. Coordinate gene regulation by fimbriae-induced signal transduction. EMBO J. 20:3074-3081.[PubMed] [CrossRef]
134. Schembri, M. A., P. B. Olsen, and P. Klemm. 1998. Orientation-dependent enhancement by H-NS of the activity of the type 1 fimbrial phase switch promoter in Escherichia coli. Mol. Gen. Genet. 259:336-344.[PubMed] [CrossRef]
135. Schembri, M. A., E. V. Sokurenko, and P. Klemm. 2000. Functional flexibility of the FimH adhesin: insights from a random mutant library. Infect. Immun. 68:2638-2646.[PubMed] [CrossRef]
136. Schembri, M. A., D. W. Ussery, C. Workman, H. Hasman, and P. Klemm. 2002. DNA microarray analysis of fim mutations in Escherichia coli. Mol. Genet. Genomics 267:721-729.[PubMed] [CrossRef]
137. Sjobring, U., G. Pohl, and A. Olsen. 1994. Plasminogen, absorbed by Escherichia coli expressing curli or by Salmonella enteritidis expressing thin aggregative fimbriae, can be activated by simultaneously captured tissue-type plasminogen activator (t-PA). Mol. Microbiol. 14:443-452.[PubMed] [CrossRef]
138. Smith, S. G., and C. J. Dorman. 1999. Functional analysis of the FimE integrase of Escherichia coli K-12: isolation of mutant derivatives with altered DNA inversion preferences. Mol. Microbiol. 34:965-979.[PubMed] [CrossRef]
139. Sokurenko, E. V., V. Chesnokova, D. E. Dykhuizen, I. Ofek, X. R. Wu, K. A. Krogfelt, C. Struve, M. A. Schembri, and D. L. Hasty. 1998. Pathogenic adaptation of Escherichia coli by natural variation of the FimH adhesin. Proc. Natl. Acad. Sci. USA 95:8922-8926.[PubMed] [CrossRef]
140. Sokurenko, E. V., H. S. Courtney, S. N. Abraham, P. Klemm, and D. L. Hasty. 1992. Functional heterogeneity of type 1 fimbriae of Escherichia coli. Infect. Immun. 60:4709-4719.[PubMed]
141. Sokurenko, E. V., H. S. Courtney, D. E. Ohman, P. Klemm, and D. L. Hasty. 1994. FimH family of type 1 fimbrial adhesins: functional heterogeneity due to minor sequence variations among fimH genes. J. Bacteriol. 176:748-755.[PubMed]
142. Sokurenko, E. V., M. A. Schembri, E. Trintchina, K. Kjaergaard, D. L. Hasty, and P. Klemm. 2001. Valency conversion in the type 1 fimbrial adhesin of Escherichia coli. Mol. Microbiol. 41:675-686.[PubMed] [CrossRef]
143. Soto, G. E., and S. J. Hultgren. 1999. Bacterial adhesins: common themes and variations in architecture and assembly. J. Bacteriol. 181:1059-1071.[PubMed]
144. Stentebjerg-Olesen, B., T. Chakraborty, and P. Klemm. 2000. FimE-catalyzed off-to-on inversion of the type 1 fimbrial phase switch and insertion sequence recruitment in an Escherichia coli K-12 fimB strain. FEMS Microbiol. Lett. 182:319-325.[PubMed] [CrossRef]
145. Stentebjerg-Olesen, B., T. Chakraborty, and P. Klemm. 1999. Type 1 fimbriation and phase switching in a natural Escherichia coli fimB null strain, Nissle 1917. J. Bacteriol. 181:7470-7478.[PubMed]
146. Sukupolvi, S., R. G. Lorenz, J. I. Gordon, Z. Bian, J. D. Pfeifer, S. J. Normark, and M. Rhen. 1997. Expression of thin aggregative fimbriae promotes interaction of Salmonella typhimurium SR-11 with mouse small intestinal epithelial cells. Infect. Immun. 65:5320-5325.[PubMed]
147. Thanassi, D. G., E. T. Saulino, and S. J. Hultgren. 1998. The chaperone/usher pathway: a major terminal branch of the general secretory pathway. Curr. Opin. Microbiol. 1:223-231.[PubMed] [CrossRef]
148. Thanassi, D. G., E. T. Saulino, M. J. Lombardo, R. Roth, J. Heuser, and S. J. Hultgren. 1998. The PapC usher forms an oligomeric channel: implications for pilus biogenesis across the outer membrane. Proc. Natl. Acad. Sci. USA 95:3146-3151.[PubMed] [CrossRef]
149. Thankavel, K., A. H. Shah, M. S. Cohen, T. Ikeda, R. G. Lorenz, R. Curtiss, III, and S. N. Abraham. 1999. Molecular basis for the enterocyte tropism exhibited by Salmonella typhimurium type 1 fimbriae. J. Biol. Chem. 274:5797-5809.[PubMed] [CrossRef]
150. Thomas, W. E., E. Trintchina, M. Forero, V. Vogel, and E. V. Sokurenko. 2002. Bacterial adhesion to target cells enhanced by shear force. Cell 109:913-923.[PubMed] [CrossRef]
151. Toledano, M. B., I. Kullik, F. Trinh, P. T. Baird, T. D. Schneider, and G. Storz. 1994. Redox-dependent shift of OxyR-DNA contacts along an extended DNA-binding site: a mechanism for differential promoter selection. Cell 78:897-909.[PubMed] [CrossRef]
152. Torres, A. G., N. T. Perna, V. Burland, A. Ruknudin, F. R. Blattner, and J. B. Kaper. 2002. Characterization of Cah, a calcium-binding and heat-extractable autotransporter protein of enterohaemorrhagic Escherichia coli. Mol. Microbiol. 45:951-966.[PubMed] [CrossRef]
153. Uhlich, G. A., J. E. Keen, and R. O. Elder. 2001. Mutations in the csgD promoter associated with variations in curli expression in certain strains of Escherichia coli O157:H7. Appl. Environ. Microbiol. 67:2367-2370.[PubMed] [CrossRef]
154. Uhlich, G. A., J. E. Keen, and R. O. Elder. 2002. Variations in the csgD promoter of Escherichia coli O157:H7 associated with increased virulence in mice and increased invasion of HEp-2 cells. Infect. Immun. 70:395-399.[PubMed] [CrossRef]
155. van der Velden, A. W., A. J. Baumler, R. M. Tsolis, and F. Heffron. 1998. Multiple fimbrial adhesins are required for full virulence of Salmonella typhimurium in mice. Infect. Immun. 66:2803-2808.[PubMed]
156. Vidal, O., R. Longin, C. Prigent-Combaret, C. Dorel, M. Hooreman, and P. Lejeune. 1998. Isolation of an Escherichia coli K-12 mutant strain able to form biofilms on inert surfaces: involvement of a new ompR allele that increases curli expression. J. Bacteriol. 180:2442-2449.[PubMed]
157. Waldron, D. E., P. Owen, and C. J. Dorman. 2002. Competitive interaction of the OxyR DNA-binding protein and the Dam methylase at the antigen 43 gene regulatory region in Escherichia coli. Mol. Microbiol. 44:509-520.[PubMed] [CrossRef]
158. Warne, S. R., J. M. Varley, G. J. Boulnois, and M. G. Norton. 1990. Identification and characterization of a gene that controls colony morphology and auto-aggregation in Escherichia coli K12. J. Gen. Microbiol. 136(Part 3):455-62.[PubMed]
159. Welch, R. A., V. Burland, G. Plunkett, 3rd, P. Redford, P. Roesch, D. Rasko, E. L. Buckles, S. R. Liou, A. Boutin, J. Hackett, D. Stroud, G. F. Mayhew, D. J. Rose, S. Zhou, D. C. Schwartz, N. T. Perna, H. L. Mobley, M. S. Donnenberg, and F. R. Blattner. 2002. Extensive mosaic structure revealed by the complete genome sequence of uropathogenic Escherichia coli. Proc. Natl. Acad. Sci. USA 99:17020-17024.[PubMed] [CrossRef]
160. White, A. P., D. L. Gibson, S. K. Collinson, P. A. Banser, and W. W. Kay. 2003. Extracellular polysaccharides associated with thin aggregative fimbriae of Salmonella enterica serovar enteritidis. J. Bacteriol. 185:5398-5407.[PubMed] [CrossRef]
161. Wu, X. R., T. T. Sun, and J. J. Medina. 1996. In vitro binding of type 1-fimbriated Escherichia coli to uroplakins Ia and Ib: relation to urinary tract infections. Proc. Natl. Acad. Sci. USA 93:9630-9635.[PubMed] [CrossRef]
162. Xia, Y., D. Gally, K. Forsman-Semb, and B. E. Uhlin. 2000. Regulatory cross-talk between adhesin operons in Escherichia coli: inhibition of type 1 fimbriae expression by the PapB protein. EMBO J. 19:1450-1457.[PubMed] [CrossRef]
163. Zheng, M., F. Aslund, and G. Storz. 1998. Activation of the OxyR transcription factor by reversible disulfide bond formation. Science 279:1718-1721.[PubMed] [CrossRef]
164. Zogaj, X., W. Bokranz, M. Nimtz, and U. Romling. 2003. Production of cellulose and curli fimbriae by members of the family Enterobacteriaceae isolated from the human gastrointestinal tract. Infect. Immun. 71:4151-4158.[PubMed] [CrossRef]
165. Zogaj, X., M. Nimtz, M. Rohde, W. Bokranz, and U. Romling. 2001. The multicellular morphotypes of Salmonella typhimurium and Escherichia coli produce cellulose as the second component of the extracellular matrix. Mol. Microbiol. 39:1452-1463.[PubMed] [CrossRef]