Bacterial Adhesion to Host Cells | Microbiology

In this article we will discuss about the bacterial adhesion to host cells.

Before entering inside, bacteria adhere to host cells and secrete product(s) or structural products complementary to host. Hence, bacteria are found adhered to host’s epithelial cells due to direct adhesion to host cells or binding to secretory products that coat host cells or bacteria.

For example, teeth are rapidly colonised by bacteria. Besides, bacteria also adher to phagocyte cells of the host and trigger immune system and may or may not be phagocytosed.

A wide variety of surfaces are avail­able for adherence of microbial cells such as polymers of extracellular matrix (e.g. col­lagen, proteoglycans), bones, endothelial cells. Bacteria possess several surface mol­ecules and structures which facilitate to adhere these surfaces. Moreover, a particu­lar bacterium is able to adhere to only specific surface. It means that there exists tissue-tropism of bacteria (Table 27.9).

Some of the bacteria (e.g. Neisseria meningitidis and Salmonella spp.) encounter many types of surfaces to cause infection. But at least two facts of bacterial adhesion are very important, the physicochemical forces facilitating invasion and the specificity of process to guide specific surfaces.

Bacteria and host cells interact and affect the activity of one another by the secretion of toxins, low molecular weight metabolites, hormones, enzymes and antibacterial peptides. The behaviour of eukaryotic cells is affected by LPS, peptidoglycan and membrane protein on outer wall.

Basic Principles of Microbial Adhesion:

(i) Forces Affecting Adhesion:

There are different types of forces which operate between bacteria and host cell surface before adhesion of bacterial cells (Fig. 27.13). Van der Waals and electro­static forces apply when microbial cells are at a distance of tens of nanometers (A and B). Bacterial cell and host cell surface are mutually attracted due to Van der Waals forces. The electrostatic forces result in repulsion of these two objects.

Hydrophobic interactions (C) result in adhesion of bacterial and host cells and the former is brought closer to host cell surface for other adhesive interactions such as hydrogen bonding, cation bridg­ing and specific bonding of a molecule (ligand) on the bacterial surface to a receptor molecule present on host surface. Adhesin molecules present on bacterial surface are responsible for adhesion.

Fig. 27.13 : Various forces operating between bacteria and host surface affecting adhesion.

(ii) Role of Bacterial Structure in Adhesion:

Bacteria possess several structures which help in adhesion of cells for example fimbriae (or pilli), fibrils, flagella, capsule and S layer. All these structures consist of adhesins.

Capsule components of certain bacteria (e.g. Streptococcus, Staphylococcus, Klebsiella, Neisseria, Haemophilus) mediate adhesion to host cell surface. S layer consists of glycoprotein and self-assembling units (external to cell wall) which also help in adhe­sion. Fimbriae are present on cell surface and cause bacterial adhesion.

The epithelial surface secretes antimicrobial compounds (e.g. lysozyme and antibacterial peptides). The epithelium of respiratory tract is coated with mucin in which bacteria are trapped, brought to back of pharynx by ciliary action.

Also, several antimicrobial compounds are found in mucin (e.g. lysozyme, lactoferrin, secretory IgA, superoxide radicals and antibacterial peptides). The urinary tract and oral cavity are always flushed by secretions of respective tissues. Even then epithelium is colonised by bacteria.

Using broad-spectrum antibiotics, normal microflora is disturbed and undesirable microor­ganisms may be present such as Candida albicans, Clostridium difficile and pseudomonads which can infect the organs. It seems that normal microflora of human exerts protective effect.

How this mutualistic association between two types of bacteria got evolved? How does host cells encourage such selective adherence? Little is known about adherence of normal microflora of healthy tissues. Most of the information which we know are on microbial adhesion between pathogenic microor­ganisms and host cells. Several other facets to bacterial adhesion are given in Fig. 27.14.

Fig. 27.14 : Facets of bacterial adhesion to host cells.

Adhesin is located at the tip or along the whole length of fimbriae. Fimbriae are widely distributed among the Gram-negative bacteria such as Bordetella, Salmonella, Neisseria, Pseudomonas, Yersinia, etc.

Fimbriae have been classified into the five types:

Type 1 (rigid fimbriae that exhibit mannose-sensitive haemagglutination e.g. E. coli),

Type 2 (similar to type 1 but not induce haemagglutination e.g. Actinomyces naeslundii).

Type 3 (flexible and mannose-resistant fimbriae (they are common among the enterobacteriaceae e.g. Klebsiella pneumoniae).

Type 4 (they consist of N-methyl-phenylalanine in the amino terminus region of the major subunits e.g. Pseudomonas aeruginosa), and

Type 5 (thinner than type 1, mannose-sensitive and a few in number).

(iii) Bacterial Adhesions:

Several different types of molecules present on bacterial cell surface act as adhesins and facilitate the attaching bacteria to host cell surface. One of the most extensively explored adhesins is the lectin (glycoproteins). Lectins are present at the end of pili, capsule of Gram- negative bacteria, etc.

Examples of bacteria comprising of lectins are: E. coli (N-acctyl-D- galactosamine), Kleb. pneumoniae (N-acetylmuraminic acid and N-acetyl-D-glucosamine), Staph, saprophyticus (N-acetyllactosamine). In Gram-negative bacteria, lipoteichoic acid (LTA) acts as an important adhesin. A glycoprotein (fibronectin) produced by many epithelial cells and other host cells act as receptor for LTA.

S. aureus produces a surface protein (210 KDa) which acts as adhesin and mediates the adhesion to fibronectin. The bacterium also attaches to the other host proteins e.g. fibrinogen, laminin. A proline-rich protein of Mycoplasma sp. also acts as adhesin.

Carbohydrates present on bacterial cell surfaces act as adhesins in certain bacteria. For example, P. aeruginosa secretes an exopolysaccharide (alginate) that acts as adhesin for attachment to tracheal cells and mucin, and binds to both.

Lipopolysaccharide (LPS) of Gram-negative bacteria play an impor­tant role in adhesion. For example, LPS of Comp. jujeni, E. coli, Ps. aeruginosa, Sal. typhi, Shig. flexneri, etc. mediate the bacterium to attach to host’s epithelial cells.

Bacterial enzymes (e.g. glyceraldehyde 3-phosphate-dehydrogenase of Strep, pyogenes, gingipain R and gingipain K of Por. gingivalis, cell surface urease of Hel. pylori, glucosyl transferase of cell surface of mutant streptococci have been found to func­tion as adhesins in attaching to epithe­lial cells of various tissues.

Molecular chaperonin 60 (from Hel. pylori and Haem. ducreyi) which is a heat-shock or stress protein are produced which mediates bacterial colonisation.

(iv) Nature of the Host Cells:

Bacteria adhere to host surfaces in three different ways:

(a) Directly to the lipid bilayer,

(b) Directly to the cell surface receptors whose normal function is to bind host molecules, and

(c) Indirectly to host molecules already bound to the host cell surface (Fig. 27.15).

The cell membrane is made up of lipid bilayer where proteins are embeded. Lipid structure is such that are recognised by bacterial adhesins. Proteins function in transport of molecules, recognition and binding of hormones, cytokines and extracellular matrix molecules, signal transduction and cell-cell interactions, carbohydrate of glycoproteins and amino acids of proteins act as receptors for bacterial adhesins.

Several receptor molecules are found on mammalian cell surfaces such as integrins, cadherins, selectins, serpentine receptors, cytokine receptors, intracellular adhesin molecules, etc.

The extracellular matrix (ECM) of host tissue consists of complex mixture of polymers such as fibronectin, fibrogen, collagen, proteoglycans and glycosaminoglycans. The ECM affects many cellular activities such as migration, proliferation and differentiation. Bacteria adhere to the ECM and arrest the activities of the host cells.

Effect of Adhesion on Host Cells:

After adhesion of bac­terial cells, a number of changes occur in host cells such as alteration in mor­phology, induction of loss of fluid, release of cytokine, apoptosis (Fig. 27.17). What would be the result of inter­action? This depends on types of host cells.

 Some of the effects of bacterial inva­sion on host cells are discussed below:

(i) Effect on Epithelial Cells:

Bacteria are able to colonise all types of epithelial cells of a healthy person because the microbes contain molecules (e.g. LPS, peptidoglycan, and lipoteichoic acid) which harm the mammalian cells. However, some adherent microorganisms pose minimal effects. Examples of some of the bacteria are given which cause effect on epithelial cells (Table 27.10).

(ii) Adhesion to Fibroblast:

Fibroblasts produce materials of extracellular matrix and maintain integrity of connective tissue. Besides, they can also secrete cytokines and other inflammatory materials. Hence, they are important in host defence and other maintenance of cytokine network. A profound immunological effect and structural consequences can be obtained after interfering with their function.

So far less attention has been paid on interaction between bacteria and fibroblast.

Attachment of Treponema denticola to human gingival fibroblast induces several changes in the host cells:

(a) Retraction of pseudopods followed by rounding of cells and the formation of membrane blebs,

(b) Rearrangement of filamentous actin network into a perinuclear array,

(c) Detachment of cells from their substratum due to degradation of fibronectin by surface- associated proteases, and

(d) Apoptosis (cell death).

(iii) Adhesion to Vascular Endothelial Cells:

A continuous monolayer lining of the blood vessel walls is formed by vascular endothelial cells. These act as barrier between the blood and the vessel walls and regulate the tone of blood vessel, permeability and coagulation of blood, reactivity of leukocytes and platelets, angiogenesis and the source of vascular mediators (e.g. nitric oxide, prostacyclin, endothelin).

(iv) Adhesion to Phagocytes:

It is important to know the mechanism of adhesion to phagocytic cells because bacteria are disposed off through this mechanism. Some bacteria adhere directly (involving the interaction between adhesins and acceptors on phagocytes) or indirectly by interacting with host components (e.g. complement or immunoglobulins) which then bind to receptors of phagocytes. Uptake of bacterium into a phagosome is induced by adhesins.

Then the phagosome fuses with lysosome resulting in death of the bacterium. Moreover, there are some bacteria which survive within phagocytes, for example M. tuberculosis, B. purtusis, Y. enterocolitica, etc. Ad­hesion by some of the pathogens (Legionella pneumophila and S. typhimurium) causes the release of cytokines by the host cells. Increased expression of cytokines and chemokines occur after at­tachment of L. pneumophila or S. typhimurium to murine macroph­ages.

The other interesting result of bacterial adhesion to phagocytes is the apoptosis (programmed cell death). Apoptosis occurs when the enteric pathogens (e.g. Y. enterocolitica) bind to macroph­ages. It invades mucosa, resists phagocytosis and maintains extracellular life.

It does not induce macrophages but induces death of phagocytes which show all features of apoptosis such as cytoplasm shrinkage, nuclear chromatin condensation and DNA fragmentation. Cell death occurs due to secretion of one or more Yop proteins. The bacterium involves this strategy for its survival.

Bacterial Invasion of Host Cells:

Invasion of host cells by bacteria results in several diseases as mentioned earlier. How does this occur remains questionable? During the last decade much efforts have been made to unravel the mechanism that bacteria use for infection of host cells. Bacteria have evolved several invasive mechanisms.

Most of them involve the manipulation of normal host cell cytoskeletal components such as actin and tubulin resulting in the investigation of the host cell membrane to enclose the bacterium within the vacuole. This occurs by interfering the inhibition of signal transduction or both.

In addition, Rickettsia prowazekii digests the part of cytoplasmic membrane. The invasive bacteria may remain viable inside the vacuole (M. tuberculosis), escape from vacuole and colonise the cytoplasm (Listeria monocytogenes) or escape from the vacuole and the cell and spread systematically (Yersinia enterocalitica). In this section the adhesive processes leading to invasion of the host cell have been described.

(i) Mechanism of Invasion:

Bacterial invasion of host cells is broadly classified into the three groups on the basis of involvement of microfilaments or microtubules of host cells or make entry into the cells. The type of host cells to be invaded also governs the invasion process.

The ways through which bacteria invade epithelial cells, endothelial cells and macrophages are described below:

(a) Invasion of Epithelial Cells:

The outer integument of our body is constituted by the epithelial cells. A varying population of microbes contributing the normal microflora colonises the epithelial interface. Thus epithelium acts as the first physical barrier for commensal microorganisms.

Many bacteria enter epithelial cells of the host by inducing the rearrangement of microfilament of the cytoskeletin. Invasion of host cells by several bacteria such as L. monocytogenes, Salmonella, Shigella, and Yersinia have been most extensively studied.

Y. enterocolitica adhers to an epithelial cell. Adhesion is mediated by many adhesin between the Ail protein and Yad A protein. Close contact between bacterium and host cell is made at any point on the bacterium-host cell interface- a process described as zippering. This induces the uptake of the organism into an endocytic vacuole.

The bacteria sink into the membrane of the epithelial cell. The host cells show a normal appearance within a few minutes of entry.

Binding of invasion of bacterium to its integrin receptor (on host cell surface) along the zippering induces the uptake of bacterium (Fig. 27.18). Clustering of integrins induces tyrosine kinase activity which is required for invasion.

Cytochalasin inhibits endocytosis emptying the involvement of actin microfilaments. During early stages of in­ternalization, clathrin lattices are soon formed beneath the bacteria. The polymer­ized actin and other proteins (filamin and talin) surround the vacuole. The internal­ized bacteria survive inside the vacuole but not reproduce.

Unlike Yersinia, Salmonella spp. in­vade the epithelial cells after adhesion to microvili. Within 1 minute of contact, mi­crovilli form pseudopods extending from cell surface of host epithelium, engulf bac­teria and internalise within a vacuole (Fig. 27.19). Mannose-specific type 1 fimbriae mediates the adhesion of Salmonella spp. to epithelial cells of intestine.

A little information is available on the initial host signal transduction events after bacterial invasion. After invasion, intracellular level of Ca⁺⁺ increases which polymerises actin synthesis and inhibits bacterial inva­sion. Fimbriae-mediated ad­hesion of Salmonella takes place.

Besides, cytoskeletal proteins (actin, talin, tubu­lin, tropomyosin and ezrin) accumulate in host cell at the site of bacterial adhe­sion. Several proteins of host cell membrane form aggre­gates in the vicinity of bac­terial attachment. Conse­quently, bacterium is taken up by macropinocytosis (a process which involves in­take of large quantity of ex­tracellular fluid).

Thus bac­terium resides on a large fluid filled vacuole called spaceous phagosome surrounded by polymerised actin, talin and actinin. About 4-6 hours of invasion bacteria proliferate. It is accompanied by the formation of lysosomal fibrillar structures (Salmonella- induced filaments, Sifs) attached to phagosomes.

(b) Invasion of Epithelial Cells:

There are several pathogenic bacteria which enter at a site of host cell and spread throughout the body, for example H. influenzae, N. gonorrhoea, S. dysenteriae, S. pneumoniae, S. typhi, etc. They enter the blood stream and cross the cellular barrier i.e. the endothelium.

Invasion takes place by one of the four main routes:

i. Invasion followed by intracellular persistence without multiplication (e.g. S. aureus, P. aeruginosa),

ii. Invasion followed by intracellular replication (Rickettsia rickettsii),

iii. Traversed without cell disruption (spirochaetes), and

iv. Invasion within phagocytes (Listeria monocytogenes).

E. coli causes diarrhoeae, urinary tract infection and neonatal meningitis. A little is known how does it pass blood-brain barrier. It has been found that an outer membrane protein (Omp A) of the bacterium plays a central role in invading the host cells.

Bacterial Omp A protein mediates binding of bacteria to the receptors (N-acetylglucosamine, β1-4-N-acetylglucosamine epitope) present on BMEC (brain microvascular endothelial cells). The polymers of β1-4-linked N-acetylglucosamine prevent the entry of E. coli into the cerebrospinal fluid of neonatal rats. Hence, meningitis caused by it can be controlled by using receptor analogues.

(c) Invasion of Macrophages:

Macrophages are a part of our immune system. They engulf antibody or complement-coated (opsonised) bacteria, internalise in vacuole and kill them. Adhesion is mediated by interaction between Fc region of antibodies and Fcr receptors on macrophage surface.

Binding causes internalisation of bacteria within a vacuole. Then it fuses with lysosome to form phagosome inside which bacterium is killed by enzymes, antimicrobial peptides, reactive oxygen species and low pH.

Fimbrial protein FimH of uropathogenic E. coli acts as adhesin. It binds to CD48 (a glycosylphatidyl inositol-linked glycoprotein) receptors of macrophages and E. coli is internalised.

(ii) Consequences of Invasion:

What happens when a bacterium invades the host cell? Certainly both the host cells and bacterium are affected.

(a) Effect of Host Cells:

It is a difficult task to describe all the changes occurring in host cells due to bacterial invasion. Bacteria affect the host cells in many ways finally resulting in death. After invasion several cells respond by secreting cytokines which activate the immune system.

Over production of cytokines may adversely affect the host cells e.g. S. dysenteriae. The diarrhoeal response to infection is mediated by postglandin which is an important regulator for secretion of gastro-intestinal fluid by inducing C1^– secretion from mucosa. Infection by enterobacteria of intestinal epithelial cells results in secretion of postglandins.

Within 15 minutes on invasion of macrophages by S. typhimurium the former lost their phagocytic ability, induces apoptosis resulting in death of 50% cells. Shigella flexneri also induces induction of apoptosis in macrophages.

(b) Effect on Bacteria:

The bacteria have several options after invading the host cells. They may live within the vacuole of host cell (e.g. M. tuberculosis, S. typhimurium. Brucella spp., Burkhulderia cepacia) within the cytoplasm (e.g. Shigella spp., Rickettsia spp. Listeria monocytogenes) or may exit the cell and live extracellular life (e.g. Yersinia spp.). Bacteria living extracellularly are transported to another sites in the host.

Irrespective of above option, the bacteria adapt the new environmental conditions by expressing gene products that help their survival in new habitats. The bacterial regulatory system responds to changes of environmental factors e.g. pH, osmolarity and concentration of O₂, CO₂, micro- and macro- nutrients and antibacterial substances.

For example, Salmonella spp. after ingestion invades epithelial cells of mucosa. Its invasion depends on production of secreted invasion proteins (Sips) exported by Type III secretion system which is activated after contact with cell membrane of host cells. Genes located on Salmonella pathogenicity island (SPI-2) encode the proteins. After intemalisation, it remains within a vacuole where it replicates until its exit.

Within the vacuole, expression of iron- and magnesium-regulated genes (i.e. iro A and mgt B, respectively is increased). Low pH of vacuole upregulates the expression of lysine decarboxylase gene. It suggests that O₂ and lysine are present in the vacuole at pH 6.0.

After intemalisation, Salmonella grows and survives within the macrophage by expressing a large number of genes. The environmental conditions within the macrophage (i.e. limited nutrients, low pH and high osmolarity) force the bacteria to produce alternative sigma factor for RNA polymerase. The increased levels of sigma factor render the bacteria more resistant to adverse conditions and enable it to respond within the macrophage.

Development of molecular and genetical techniques has helped the investigation of ways in which bacteria respond within their host. These techniques include isolation of bacterial mRNA, synthesis of bacterial cDNA and probes, signature-tagged mutagenesis, differential fluorescence induction, in vivo expression technology, etc.

(iii) Bacterial Survival and Growth after Invasion:

Once the bacterium has invaded the epithelial cell, it may proliferate within the cell and come out of cell or infect deeper tissues and spread the outer sites. Thus the bacterium leads two types of life style: extracellular and intracellular life styles.

If the bacteria remains extracellular, they face secretions of blood or tissues such as complements, antibodies, antimicrobial molecules discharges by phagocytes and phagocytic cells. Bacteria have evolved various ways to deal with these antimicro­bial substances and phagocytic cells through pro­duction of toxins (see preceding section), capsule (S. aureus, K. pneumoniae, E. coli, S. pyogenes), etc.

There is a large number of bacteria which lead intracellular lifestyle surviving inside the vacu­ole, phagolysosome or cytoplasm of infected host cells.

(a) Survival in Phagosome:

It may be exem­plified by Coxiella burnetti causing Q fever. Its main habitat is macrophage inside which it grows and multiplies. It exists in two phases: I and II. Phase I bacteria have a smooth form of lipopolysaccharide and are highly virulent, whereas phase II bacteria have a rough lipopolysaccharide and have reduced virulence.

After adhesion, phase I bacteria bind to a leukocyte, response integrin (a membrane protein) and integrin-associated proteins (Fig. 27.20). The bacterium is internalised by microfilament- dependent process. Then phagosome and lysosome get fused to form apparent normal phagolysosome containing membrane proton ATPase and lysosomal enzymes.

The infected host cell is capable of asymmetric division resulting in two daughter cells, one containing the vacuole (inside which bacterium is present) and the other parasite-free cell. The vacuole containing cell is broken liberating the bacterium, whereas the other cell may be invaded by bacterium repeating the similar cycle. Little is known how the bacterium reproduces in acidic phagolysosome (pH 5.2).

Fig. 27.20 : Stages in life cycle of Coxiella burnetil.

(b) Survival within Vacuoles:

Legionella pneumophila causing legionnaire’s disease invades the macrophages and gets internalised by coiled phagocytosis (Fig. 27.21). After 15 minutes of internalisation, phagosome is surrounded by smooth vesicles and after 1 hour by mitochondria.

The phagosome does not fuse with lysosome. Hence, it lacks endosomal receptor (transferrin) and endosomal/lysosomal markers (CD63). Phagosome-lysosomal fusion is prevented by the polycationic protein (Mip) of the bacterium.

After 4 to 8 hours of internalisation, the phagosome is surrounded by ribosome and ribose-containing vesicles. Bacterium multiplies exponentially with the doubling time of 2 hours using bacterial cell organelles resulting in cell lysis.

(c) Survival within the Host Cytoplasm:

There is a number of bacteria which escape from vacuole (after invading the host cell) and remain within the cytoplasm, for example L. monocytiogenes, Shigella spp., S. aureus, streptococci, Rickettsia spp., Haemophilus ducreyi, of these only Rickettsia spp. are the obligate intracellular parasite.