There are four main types of bacterial cell wall

The four main types of bacterial cell wall (Fig. 14.1) belong to the following groups.

Fig. 14.1 Bacterial cell walls

Different immunological mechanisms have evolved to destroy the cell wall structure of the different groups of bacteria. All types have an inner cell membrane and a peptidoglycan wall. Gram-negative bacteria also have an outer lipid bilayer in which lipopolysaccharide (LPS) is embedded. Lysosomal enzymes and lysozyme are active against the peptidoglycan layer, whereas cationic proteins and complement are effective against the outer lipid bilayer of the Gram-negative bacteria. The compound cell wall of mycobacteria is extremely resistant to breakdown, and it is likely that this can be achieved only with the assistance of the bacterial enzymes working from within. Some bacteria also have fimbriae or flagellae, which can provide targets for the antibody response. Others have an outer capsule, which renders the organisms more resistant to phagocytosis or to complement. The components indicated with an asterisk (*) are recognized by the innate immune system as a non-specific ‘danger’ signal that selectively boosts some aspects of immune activity. (Gram staining is a method that exploits the fact that crystal violet and iodine form a complex that is more abundant on Gram-positive bacteria. The complex easily elutes from Gram-negative bacteria.)

The outer lipid bilayer of Gram-negative organisms is of particular importance because it is often susceptible to lysis by complement. However, killing of most bacteria usually requires uptake by phagocytes. The outer surface of the bacterium may also contain fimbriae or flagellae, or it may be covered by a protective capsule. These can impede the functions of phagocytes or complement, but they also act as targets for the antibody response, the role of which is discussed later.

Pathogenicity varies between two extreme patterns

The two extreme patterns of pathogenicity are:

Fig. 14.2 Mechanisms of immunopathogenicity

(1) Some bacteria cause disease as a result of only a single toxin (e.g. Corynebacterium diphtheriae, Clostridium tetani) or because of an ability to attach to epithelial surfaces (e.g. in group A streptococcal sore throat). Immunity to such organisms may require only antibody to neutralize this critical function. (2) At the other extreme there are organisms that are not toxic, and cause disease by invasion of tissues and sometimes cells, where damage results mostly from the bulk of organisms or from immunopathology (e.g. lepromatous leprosy). Where organisms invade cells, they must be destroyed and degraded by the cell-mediated immune response. (3) Most organisms fall between the two extremes, with some local invasiveness assisted by local toxicity and enzymes that degrade extracellular matrix (e.g. Staphylococcus aureus, Clostridium perfringens). Antibody and cell-mediated responses are both involved in resistance and the latter is a major cause of antibiotic-induced colitis and diarrhea.

However, most bacteria are intermediate between these extremes, having some invasiveness assisted by some locally acting toxins and spreading factors (tissue-degrading enzymes).

Corynebacterium diphtheriae and Vibrio cholerae are examples of organisms that are toxic, but not invasive. Because their pathogenicity depends almost entirely on toxin production, neutralizing antibody to the toxin is probably sufficient for immunity, though antibody binding to the bacteria and so blocking their adhesion to the epithelium could also be important.

In contrast, the pathogenicity of most invasive organisms does not rely so heavily on a single toxin, so immunity requires killing of the organisms themselves.

The first lines of defense do not depend on antigen recognition

The body’s first line of defense against pathogenic bacteria consists of simple barriers to the entry or establishment of the infection. Thus, the skin and exposed epithelial surfaces have non-specific or innate protective systems, which limit the entry of potentially invasive organisms.

Intact skin is impenetrable to most bacteria. Additionally, fatty acids produced by the skin are toxic to many organisms. Indeed, the pathogenicity of some strains correlates with their ability to survive on the skin. Epithelial surfaces are cleansed, for example, by ciliary action in the trachea or by flushing of the urinary tract.

Many bacteria are destroyed by pH changes in the stomach and vagina, both of which provide an acidic environment. In the vagina, the epithelium secretes glycogen, which is metabolized by particular species of commensal bacteria, producing lactic acid.

The second line of defense is mediated by recognition of bacterial components

If organisms do enter the tissues, they can be combated initially by further elements of the innate immune system. Numerous bacterial components are recognized in ways that do not rely on the antigen-specific receptors of either B cells or T cells. These types of recognition are phylogenetically ancient ‘broad-spectrum’ mechanisms that evolved before antigen-specific T cells and immunoglobulins, allowing protective responses to be triggered by common microbial components bearing so-called ‘pathogen-associated molecular patterns’ (PAMPs), recognized by ‘pattern recognition molecules’ of the innate immune system (see Chapter 6).

Many organisms, such as non-pathogenic cocci, are probably removed from the tissues as a consequence of these pathways, without the need for a specific adaptive immune reaction. Figure 14.3 shows some of the microbial components involved and the host responses that are triggered.

Fig. 14.3 Protective mechanisms not involving antigen-specific B or T cells

Several common bacterial PAMPs are recognized by molecules present in serum and by receptors on cells. These recognition pathways result in activation of the alternative complement pathway (factors C3, B, D, P), with consequent release of C3a and C5a; activation of neutrophils, macrophages, and NK cells; triggering of cytokine and chemokine release; mast cell degranulation, leading to increased blood flow in the local capillary network; and increased adhesion of cells and fibrin to endothelial cells. These mechanisms, plus tissue injury caused by the bacteria, may activate the clotting system and fibrin formation, which limit bacterial spread.

The immune system has selected these structures for recognition because they are not only characteristic of microbes, but are essential for their growth and cannot be easily mutated to evade discovery (though, as might be predicted, there are increasing examples of pathogen strategies that attempt to subvert this process).

It is interesting to note that the ‘Limulus assay’, which is used to detect contaminating lipopolysaccharide (LPS) in preparations for use in humans, is based on one such recognition pathway found in an invertebrate species. In Limulus polyphemus (the horseshoe crab), tiny quantities of LPS trigger fibrin formation, which walls off the LPS-bearing infectious agent.

LPS is the dominant activator of innate immunity in Gram-negative bacterial infection

Injection of pure LPS into mice or even humans is sufficient to mimic most of the features of acute Gram-negative infection, including massive production of proinflammatory cytokines, such as IL-1, IL-6, and tumor necrosis factor (TNF), leading to severe shock.

Q. How does release of proinflammatory cytokines cause shock?

A. These proinflammatory cytokines act directly on endothelium to increase vascular adhesiveness, and indirectly activate other plasma enzyme systems to release vasoactive peptides and amines leading to a drop in blood pressure.

Recognition of LPS is a complex process involving molecules that bind LPS and pass it on to cell membrane-associated receptors on leukocytes, and endothelial and other cells, which initiate this proinflammatory cascade (Fig. 14.4).

Fig. 14.4 Effects of lipopolysaccharide

LPS released from Gram-negative bacteria becomes bound to LPS-binding protein (LBP) which promotes transfer of LPS to either soluble CD14 (sCD14) or to a GPI-linked membrane form of the protein (mCD14) expressed on neutrophils and macrophages and to a lesser extent on epithelial and endothelial cells. LBP then dissociates and transfers LPS to the TLR4/MD2 complex, allowing TLR4 to transduce intracellular signals that increase release of many proinflammatory cytokines including TNFα and IL-6, as well as type I IFN and IL-1. These in turn activate endothelial cells, increasing adhesion molecule expression and also drive the acute phase response in the liver. One product of the acute phase response is further production of LBP and sCD14.

Binding of LPS to TLR4 is a critical event in immune activation. TLR4 knockout mice are resistant to LPS-induced shock and there is some evidence that polymorphisms in human TLR4 may influence the course of infection with these bacteria.

The LBP and CD14, which bind LPS, are also involved in recognition of lipid-containing bacterial components from mycoplasmas, mycobacteria, and spirochetes.

Complement is activated via the alternative pathway

Complement activation can result in the killing of some bacteria, particularly those with an outer lipid bilayer susceptible to the lytic complex (C5b–9).

Perhaps more importantly, complement activation releases C5a, which attracts and activates neutrophils and causes degranulation of mast cells (see Chapter 3). The consequent release of histamine and leukotriene (LTB₄) contributes to further increases in vascular permeability (see Fig. 14.3).

Opsonization of the bacteria, by attachment of cleaved derivatives of C3, is also critically important in subsequent interactions with phagocytes.

Release of proinflammatory cytokines increases the adhesive properties of the vascular endothelium

The rapid release of cytokines such as TNF and IL-1 (see Fig. 14.4) from macrophages increases the adhesive properties of the vascular endothelium and facilitates the passage of more phagocytes into inflamed tissue. Combined with the release of chemokines such as CCL2, CCL3, and CXCL8 (see Chapter 6), this directs the recruitment of different leukocyte populations.

Epithelial cells, neutrophils, and mast cells are also important sources of proinflammatory cytokines.

IL-1, TNF, and IL-6 also initiate the acute phase response, increasing the production of complement components as well as other proteins involved in scavenging material released by tissue damage and, in the case of CRP, an opsonin for improving phagocytosis of bacteria.

When NK cells are stimulated by the phagocyte-derived cytokines IL-12 and IL-18 they rapidly release large quantities of interferon-γ (IFNγ). This response happens within the first day of infection, well before the clonal expansion of antigen-specific T cells, and provides a rapid source of IFNγ to activate macrophages. This T cell-independent pathway helps to explain the considerable resistance of mice with SCID (severe combined immune deficiency, a defect in lymphocyte maturation) to infections such as with Listeria monocytogenes. In mice, CD1d-restricted NK T cells also secrete IFNγ in response to IL-12 and IL-18 and other ligands, and help to further activate both NK cells and macrophages.

Pathogen recognition generates signals that regulate the lymphocyte-mediated response

The signals generated following the recognition of pathogens not only generate a cascade of innate immune events, but also regulate the development of the appropriate lymphocyte-mediated response.

Dendritic cells (DCs) are crucial for the initial priming of naive T cells specific for bacterial antigens. Contact with bacteria in the periphery induces immature DCs to migrate to the draining lymph nodes and augments their antigen-presenting ability by increasing their:

Some of this DC activation occurs secondary to their production of cytokines such as type I IFN.

Activated macrophages also act as antigen-presenting cells (APCs), but probably function more at the site of infection, providing further activation of effector rather than naive T cells. Following initial T cell activation by dendritic cells, B cells are also able to act as APCs during B cell–T cell cooperation and are essential for the protective action of polysaccharide-conjugate vaccines in children against encapsulated bacteria such as S. pneumoniae and H. influenzae.

Binding of bacterial components to pattern recognition receptors such as TLRs induces a local environment rich in cytokines such as IFNγ, IL-12, and IL-18, which promote T cell differentiation down the TH1 rather than TH2 pathway.

Immunologists have made use of these effects for many decades (even without knowing their true molecular basis) in the use of adjuvants in vaccination. ‘Adjuvant’ is derived from the Latin adjuvare, to help. When given experimentally, soluble antigens evoke stronger T and B cell-mediated responses if they are mixed with bacterial components that act as adjuvants. Components with this property are indicated in Figure 14.1. This effect probably reflects that the antigen-specific immune response evolved in a tissue environment that already contained these pharmacologically active bacterial components.

With the exception of proteins such as flagellin, which itself stimulates TLR5 and is also a strong T cell immunogen, the response to pure bacterial antigens, injected without adjuvant-active bacterial components, is essentially an artificial situation that does not occur in nature.

The best known adjuvant in laboratory use, complete Freund’s adjuvant, consists of killed mycobacteria suspended in oil, which is then emulsified with the aqueous antigen solution.

New-generation adjuvants based on bacterial components (and safe to use in humans, unlike Freund’s adjuvant) include synthetic TLR activators such as CpG motifs and monophosphoryl lipid A (MPL) as well as recombinant cytokines such as IL-12, IL-1, and IFNγ. Identifying the best adjuvant for inclusion in a vaccine is arguably as important as the choice of antigens and is dramatically illustrated in the RTS,S malaria vaccine – a product which was not effective until reformulated with a new MPL based adjuvant.

Pathogenic bacteria may avoid the effects of antibody

Neisseria gonorrhoeae is an example of a pathogenic bacterium that uses several immune evasion strategies (Fig. 14.8) and humans can be repeatedly infected with N. gonorrhoeae with no evidence of protective immunity.

Fig. 14.8 Mechanisms used by Neisseria gonorrhoeae to avoid the effects of antibody

N. gonorrhoeae is an example of a bacterium that uses several strategies to avoid the damaging effects of antibody. First, it fails to evoke a large antibody response, and the antibody that does form tends to block the function of damaging antibodies. Second, the organism secretes an IgA protease to destroy antibody. Third, blebs of membrane are released, and these appear to adsorb and so deplete local antibody levels. Finally, the organism uses three strategies to alter its antigenic composition: (i) the LPS may be sialylated, so that it more closely resembles mammalian oligosaccharides and promotes rapid removal of complement; (ii) the organism can undergo phase variation, so that it expresses an alternative set of surface molecules; (iii) the gene encoding pilin, the subunits of the pilus, undergoes homologous recombination to generate variants. N. gonorrhoeae also impairs T cell activation by engaging a co-inhibitory receptor CEACAM-1 on the lymphocyte surface by one of its OPA proteins.

Antibodies may also be important for effective immunity against some intracellular bacteria such as Legionella and Salmonella spp.

Pathogenic bacteria can avoid the detrimental effects of complement

Related posts:

Cell-mediated Cytotoxicity Immune Responses in Tissues Hypersensitivity (Type IV) Mechanisms of Innate Immunity Complement Antibodies

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