Bacterial adherence and other virulence factors for urinary tract infection

INTRODUCTION — Bacterial adhesion onto mucosal or urothelial cells is an important phenomenon determining bacterial virulence. Infection in the urinary tract is related in part to the ability of bacteria to adhere and colonize epithelial cells in the gut, perineum, urethra, bladder, renal pelvicalyceal system, and renal interstitium [1-6]. Adhesion is particularly important when infection occurs in an anatomically normal urinary tract [3,5,6], but it is also important in recurrent cystitis and infection complicating indwelling bladder catheters [7,8].

The pathophysiology of bacterial adhesion in the urinary tract is complex [9]. Uropathogenic Enterobacteriaceae are electronegative and too small to overcome repulsion by the net negative charge of epithelial cells. As a result, bacterial adhesion cannot occur in the absence of fimbriae or other (nonfimbrial) surface adhesion systems. These systems have favorable electrical charge and also promote adhesion via hydrophobicity. Fimbriae allow irreversible attachment to the uroepithelial cell membrane via adhesins [5,6].

This topic review will primarily consider the role of bacterial adhesion as a virulence determinant of Escherichia coli and Proteus mirabilis, the two most common strains responsible for urinary tract infection (UTI). Virulence factors of E. coli are mainly responsible for promoting progression of the organism from the fecal reservoir into the bladder and occasionally the kidney. In comparison, virulence factors of P. mirabilis can induce UTI but are particularly important for the formation of infectious struvite stones.

Bacterial virulence is not related to resistance to antimicrobial drugs. As an example, the most adherent strain of E. coli in the patient's intestinal flora is usually sensitive to most antibiotics.

Multiple host-pathogen interactions are at work in urinary tract infections [10].

BACTERIAL ADHESION — Bacterial adhesion in the urinary tract results from a complex interplay of factors [11]. Uropathogenic E. coli (UPEC) display pili at their surface, which help mediate adhesion:

●UPEC attaches to the uroepithelium through type 1 pili, which bind the receptors uroplakin Ia and IIIa [9]. This binding stimulates signalling pathways that mediate invasion and apoptosis. Binding of type 1 pili to alpha 3 beta 1 integrins also mediates internalization of the bacteria into superficial facet cells to form intracellular bacterial communities or pods. Sublytic concentrations of the pore-forming hemolysin A toxin can inhibit the activation of Akt proteins and lead to host cell apoptosis and exfoliation. Exfoliation of the uroepithelium exposes the underlying transition cells for further UPEC invasion, and the bacteria can reside in these cells as quiescent intracellular reservoirs that may be involved in recurrent infections.

●The tip of pili consists of the adhesion factor PapG that binds to glycosphingolipids of the kidney epithelium [12].

●Some UPEC strains also possess type I pili at their surface, which have binding specificity to D-mannosylated receptors, such as the uroplakins of the bladder [13].

●Type IV pili form another class of polymeric adhesive surface structures expressed by different gram-negative bacteria [14]. They have the ability to retract through the bacterial cell wall while their tip remains attached to its target surface, allowing for "twitching motility," a flagella-independent mode of motility important for colonization of host surfaces [15]. These pili are composed of thousands of copies of the major pilin protein, which are synthetized in the bacterial cytoplasm, translocated across the inner membrane, and proteolytically processed. The assembled pili then pass through the outer membrane via a channel formed by the secretion protein [16,17]. These pili aggregate to form bundles.

In addition to pili, a wide range of bacterial surface adhesins recognize various classes of host molecules (including transmembrane proteins such as integrins or cadherins) or components of the extracellular matrix (such as collagen, fibronectin, laminin, or elastin) [18]. Some of these adhesins, after allowing the binding of bacteria to host cell surfaces, also trigger the internalization of bacteria inside host cells that protects them from antibiotics [12,19]. (See 'Adhesins' below.)

ESCHERICHIA COLI — Virulence factors of uropathogenic E. coli include several properties pertaining to a small group of O-serotypes, including O1, O2, O4, O6, O7, O16, O18, and O75 (figure 1) [20]. The uropathogenicity of these serotypes can be illustrated by their prevalence in different populations. They are present in approximately 28 percent of isolates from the normal fecal flora; in comparison, they are responsible for roughly 80 percent of cases of pyelonephritis, 60 percent of cystitis, and 30 percent of asymptomatic bacteriuria [21].

A study typing E. coli strains in women and their sexual partners showed that E. coli strains that had caused urinary tract infection were nine times more likely to have been passed between sexual partners than strains from women without infection [22]. Strains with P pili were also more likely to have been shared between partners.

Adhesins — There are several other factors that affect the uropathogenicity of E. coli. Of greatest importance is the presence of adhesins on the tip of bacterial fimbriae (also called pili) and on the bacterial surface itself (nonfimbrial adhesins) (table 1) [23,24]. Most adhesins are lectins that recognize binding site conformations provided by oligosaccharide sequences present on the epithelial cell surface.

●Fimbrial adhesins – Two major fimbrial adherence systems have been identified in E. coli strains associated with UTI: pap and sfa (table 1). The PAP adhesin, for example, is present on the tip of P fimbriae [23-27]. The term P fimbriae stems from the fact that the PAP adhesin recognizes the human digalactoside P blood group determinants on human erythrocytes and urothelial cells, leading to increased adhesion (picture 1) [24].

The binding of P fimbriae to blood group determinants has important implications for host susceptibility to infection. Women who are nonsecretors of histo-blood group antigens show increased in vitro binding of E. coli to urothelial cells and are more likely to have recurrent urinary tract infections [28]. Nonsecretors may have abnormal processing of gal-globoside glycolipid receptors resulting in the production of proteins that bind with more avidity to E. coli [29].

●Nonfimbrial adhesins – Nonfimbrial adhesins encompass a diverse range of proteins, including AFA (table 1). Other important adhesins include the AT (autotransporter) family of trimeric proteins [30]. Two of these proteins are highly homologous to the E. coli K-12 antigen 43 (Ag43a). This antigen promotes strong biofilm growth and its expression is associated with long-term E. coli persistence in the bladder.

The presence of adhesins in a given E. coli strain can be detected by phenotypic methods such as hemagglutination or agglutination of latex beads. However, phenotypic expression of uropathogenicity can be lost in cultured bacteria. As a result, genetic and molecular methods have been devised to identify the pap, afa, and sfa operons in the bacterial chromosomal DNA [26]. This has permitted more accurate estimate of the frequency of these adhesins in urinary tract infections.

The frequency of the pap, sfa and afa operons is approximately 75, 25, and 10 percent, respectively in pyelonephritis, 45, 20, and 12 percent in cystitis, and 24, 27, and 0 percent in asymptomatic bacteriuria [25]. We have detected at least one adhesin system in virtually all young females with primary pyelonephritis and a normal urinary tract [31].

These observations have led us and other investigators to suggest that patients with UTI who are infected with non-uropathogenic bacteria (ie, without fimbriae) should undergo a uroradiologic evaluation to detect an anatomic abnormality predisposing to infection such as intermittent reflux, neuromuscular dysfunction of the bladder or bladder neck obstruction [31,32]. The clinical relevance of this suggestion is at present limited since genotypic methods to detect adhesins are restricted to a few specialized laboratories.

Fimbriae — In addition to adhesins, which influence the likelihood of infection, fimbriae have other virulence properties. As examples:

●Type I fimbriae in E. coli serve as virulence factors by promoting persistence of the infection and by increasing the inflammatory response to the infection [29]. Children who have acute pyelonephritis due to these organisms have been noted to have a more rapid onset of infection, higher fever, longer duration of fever, and a higher white blood cell count than those infected with other organisms [33].

●In a murine model of pyelonephritis, Dr fimbriae in E. coli promoted renal tropism and prevented the clearance of infection. This effect was mediated by binding to renal tubular basement membranes and Bowman's capsule via decay accelerating factor (a complement regulatory receptor) and collagen type IV [23,25].

●Type I fimbriae appear to contribute to the ability of uropathogenic E. coli to impair ureteric contractility, which is thought to be instrumental in limiting ascension of bacteria from the bladder to kidneys [34,35].

The role of fimbriae has potentially important therapeutic implications for prevention of urinary tract infections. In a murine model, for example, animals vaccinated against the FimH adhesin developed antibodies that inhibited E. coli binding to human bladder cells in vitro and reduced colonization of the bladder mucosa by more than 99 percent [36]. The latter effect was associated with IgG anti-FimH antibodies in the urine. Passive immunization with sera from these animals also prevented colonization. In one study in monkeys, inoculation with E. coli FimH adhesin-chaperone complex mixed with MF59 adjuvant elicited a strong IgG antibody response to FimH [37]. After a pathogenic E. coli challenge, three of four vaccinated monkeys were protected from bacteriuria and pyuria whereas the four control monkeys were infected.

However, vaccination against FimH may be more complicated than expected, as use of certain fimbrial antigens may induce a conformation-specific immune response that has enhancing rather than inhibiting effects. As an example, the lectin domain of FimH can shift between a low- to a high-affinity conformation when interacting with its receptor. In an animal study, immunization with the lectin domain induced antibodies that predominantly recognized the high-affinity conformation [38]. Instead of inhibiting adhesive function, these antibodies enhanced bacterial adhesion to urothelial cells. Furthermore, bound antibodies were released when the FimH adhesin naturally converted from the high- to low-affinity state.

Two vaccine formulations thus far include a daily oral capsule containing membrane proteins from 18 E. coli strains and a vaginal suppository containing ten heat-killed uropathogenic E. coli strains; however, they require frequent administration [39]. These are both available in Switzerland. There are no vaccines against uropathogenic E. coli licensed in the United States. Research towards their development is ongoing, including evaluation of a tetravalent E. coli bioconjugate vaccine that elicited functional antibody responses against all vaccine serotypes [40] and experimental studies in mice that support the potential for adjuvanted UPEC antigen vaccine that can be administered intranasally [41].

It is likely that antibiotic prophylaxis of recurrent UTI with low (non-bactericidal) dose antibiotics reduces the frequency of relapses through diminished fimbrial adhesion to the urothelium.

Daily ingestion of cranberry juice, commonly prescribed in women with recurrent UTI, has no effect on fimbrial adhesion to the urothelium [42].

Other virulence factors — Several additional factors also affect the virulence of E. coli [20]. These include:

●The presence of flagellae, which are necessary for motility.

●Production of a hemolysin, which induces formation of pores in the cell membrane, and aerobactin, a siderophore necessary for iron uptake in the iron-poor environment of the urinary tract.

●Resistance to plasma bactericidal properties.

●Interaction with vaginal microbiota – Recurrent UTI can be triggered by transient exposure to vaginal bacteria, specifically Gardnerella vaginalis [43]. This observation suggests that UTI pathogenesis can be driven by short-term urinary tract exposure to vaginal microbiota, which are not themselves traditionally considered uropathogenic.

Entry into urothelial cells — Uropathogenic E. coli (UPEC) are able to invade urothelial cells to replicate within them [6]. Entry into bladder epithelial cells provides UPEC with a protected niche where the bacteria can persist, quiescent, for long periods, unperturbed by host defenses and protected from antibiotic therapy. These reservoirs likely serve as sources of seemingly recurrent infections [44,45].

Biofilms — Bacterial adhesion to cell surfaces is a key element in the formation of biofilms (ie, extracellular, matrix-enclosed, microbial assemblies that can adhere to biological or nonbiological surfaces, especially foreign material such as indwelling catheters). Biofilm formation entails a protected mode of growth that allows bacterial survival in a hostile environment. These matrix-embedded bacterial aggregates are more resistant to host defenses or antibiotic treatments. The in vivo role of biofilms during bacterial infections is an active field of research [46,47].

PROTEUS MIRABILIS — P. mirabilis is armed with various virulence factors, including the production of hemolysin and IgA protease, iron acquisition, flagella, fimbriae and, most importantly, the secretion of urease (figure 1) [48]. Urease is a high molecular weight cytoplasmic enzyme that hydrolyzes urea to ammonia and carbon dioxide. Ammonia combines with hydrogen to form ammonium; the ensuing reduction in free hydrogen ion concentration leads to alkalinization with the urine pH being well above 7.0 and sometimes as high as 9.0.

The alkaline urine promotes the precipitation of phosphate, carbonate and magnesium, leading to the formation of struvite stones that often form large staghorn calculi. These stones contain a mixture of a proteinaceous matrix, leukocytes, struvite and bacteria. This process is self-perpetuating since the Proteus-contaminated stone is a permanent source of bacteria. In addition, its presence within the urinary tract leads to urinary stasis, which promotes further bacterial multiplication, urinary alkalinization, and the deposition of new layers of struvite. (See "Kidney stones in adults: Struvite (infection) stones".)

In terms of bacterial adhesion, P. mirabilis produces at least four types of fimbriae that do not appear to be absolutely required for the induction of infection [48]. At least two fimbrial systems may contribute to urinary tract colonization: the MR/P fimbriae for bladder and kidney infection; and the PMF fimbriae for bladder infection.

The acquisition of a particular phenotype known as "swarm cell differentiation" that facilitates P. mirabilis ascent into the urinary tract has also been identified as a factor in uropathogenicity [49]. It is characterized by the formation of very long flagellae. Mice inoculated with wild type Proteus strains that are endowed with the "swarm" phenotype have a higher mortality rate than those inoculated with vegetative strains. Bladder inoculation with the "swarm" type of Proteus significantly increases the incidence of pyelonephritis. The relevance of this phenomenon in human UTI remains to be determined.

Studies of vaccination with Proteus fimbrial proteins have been conducted in animals [50]. As an example, intranasal immunization with the N-terminal domain of the tip adhesin of MR/P fimbriae was effective in preventing P. mirabilis urinary tract infection in mice.

STAPHYLOCOCCUS SAPROPHYTICUS — Staphylococcus saprophyticus is a common cause of UTI, particularly cystitis in young, sexually active women [51,52]. It is also an infrequent cause of acute pyelonephritis [31,53]. S. saprophyticus adheres strongly to the urothelium, a process that appears to be linked to a lactosamine residue [52].

SUMMARY

●Adhesion onto mucosal or urothelial cells is an important phenomenon determining bacterial virulence of uropathogenic Enterobacteriaceae and relies upon the presence fimbriae or other surface adhesion systems (figure 1). (See 'Introduction' above.)

●In Escherichia coli, adhesins that are located on the tip of bacterial fimbriae or on the bacterial surface itself bind to target sites on host cells and help mediate uropathogenicity (table 1). Virtually all cases of E. coli pyelonephritis in young women with a normal urinary tract are caused by isolates that possess at least one adhesin system. The fimbriae themselves also contribute to virulence through other mechanisms and are an attractive target for vaccination strategies. (See 'Escherichia coli' above.)

●The secretion of urease by Proteus mirabilis is an important virulence factor that leads to alkalinization of the urine and formation of stones that contain a proteinaceous matrix, leukocytes, struvite, and bacteria. These stones are a permanent source of bacteria. (See 'Proteus mirabilis' above.)