Pathogens and non-specific defences

What counts as a pathogen

A pathogen is a biological agent that causes disease in a host. Bacteria, fungi and protoctists are cellular pathogens; viruses are acellular pathogens. Pathogens can damage tissue directly, release toxins or provoke damaging host responses. The body first opposes them with non-specific physical, chemical, biological and cellular defences. The four groups are defined by their effect on a host and are not closely related to one another.

Describe the drawing

A four-row table of pathogen groups. Bacteria cause tuberculosis by invading lung tissue. HIV, a virus that leads to AIDS, kills T-helper cells. Fungi cause athlete's foot by digesting skin keratin. Protoctista cause malaria by bursting red blood cells. A note underneath says antibiotics attack bacterial walls, ribosomes and enzymes, none of which a virus has.

Bacteria are prokaryotes: no nucleus, a peptidoglycan cell wall, 70S ribosomes, and a metabolism of their own. Mycobacterium tuberculosis causes tuberculosis, Vibrio cholerae causes cholera, and various staphylococci turn an ordinary wound into an infected one. Because bacteria have structures and enzymes that your cells do not, they can be attacked selectively, which is what an antibiotic does.

Viruses have no cytoplasm, no ribosomes and no metabolism. A virus is nucleic acid in a protein coat, and it can do nothing at all until it is inside a host cell whose machinery it can use. Influenza, measles and HIV are viruses. Be careful with the last one: HIV is the virus, and AIDS is the condition that develops years later when the infection has destroyed enough T-helper cells that the specific immune response can no longer be co-ordinated.

Fungi that infect humans are usually superficial. Athlete's foot and ringworm are both forms of tinea, caused by dermatophyte fungi such as Trichophyton that grow in the keratin of skin and nails, digesting tissue outside their own cells and absorbing the products. In plants the fungal diseases are far more destructive: black sigatoka in banana crops and stem rust in wheat, described below, are the standard examples.

Protoctista are eukaryotic and mostly single-celled. Plasmodium, carried between people by female Anopheles mosquitoes, causes malaria; Phytophthora infestans caused the potato blight of the 1840s. Both are eukaryotes, which is precisely why drugs against them are harder to design than antibiotics: their cells work much the way yours do.

Pathogen

A biological agent that causes disease in a host, including cellular organisms such as bacteria, fungi and protoctists and acellular agents such as viruses.

Antigen

A molecule, usually a surface protein or glycoprotein, that is recognised as self or non-self and can trigger an immune response.

Non-specific defence

A defence that acts against any pathogen, without recognising which one it is, and acts at the same speed every time.

Most microorganisms are not pathogens. Resident microbiota provide a biological, competitive defence by occupying space and using resources that might otherwise be available to pathogens, which is why the bacteria living on your skin and in your large intestine count as part of your defences rather than a threat to them. A course of broad-spectrum antibiotics kills a good many of those too, and the digestive upset that follows is the consequence.

How a pathogen causes disease

Two mechanisms of damage account for most of the examples at A level.

The first is toxins. Many bacteria release proteins that interfere with a host process. Cholera toxin binds to the cells lining the small intestine and hijacks their own signalling: the level of a messenger molecule, cyclic AMP, rises and holds open a chloride channel called CFTR, so chloride ions pour into the gut lumen. That lowers the water potential of the lumen below that of the cells, water follows by osmosis, and the resulting diarrhoea can remove several litres a day. Death in cholera results from dehydration rather than from the bacterium directly, which is why oral rehydration salts are so effective a treatment. Tetanus toxin does something different again, blocking the signals that would normally stop a motor neurone firing, so muscles contract and cannot relax.

The second is cell lysis. A virus enters a host cell, its nucleic acid is transcribed and translated using the host's ribosomes, enzymes and nucleotides, and hundreds of new virus particles are assembled. Many viruses are released by bursting the cell open; others, HIV among them, bud out through the membrane, and the infected cell declines more gradually, with much of the loss caused by the immune response eliminating infected cells. Repeat the damage across an epithelium and you have a sore throat; repeat it in the T-helper cells of an untreated HIV infection and, over years, you have AIDS. Plasmodium lyses red blood cells outright, and the bursting of infected cells in synchrony is why the fever of malaria comes in waves rather than continuously.

Not every disease fits the two boxes. Mycobacterium tuberculosis survives inside the very macrophages that engulf it, and much of the lung damage in tuberculosis is done by the host's own sustained inflammatory response, which walls infected cells into masses called granulomas, rather than by a released toxin. The two exam routes are the common cases, not a complete classification.

Fungi work another way again, secreting hydrolytic enzymes onto the tissue they grow in and absorbing what dissolves, which is why athlete's foot itches and flakes. Several symptoms of an infection are produced by the host rather than by the pathogen: fever, swelling, aching and tiredness are effects of the immune response itself.

Exotoxins, endotoxins and invasion

Bacterial toxins come in two kinds, and the difference is where the toxin comes from. An exotoxin is a protein that a living bacterium secretes into its surroundings. Cholera toxin and tetanus toxin are both exotoxins. An endotoxin is part of the bacterium itself: the lipopolysaccharide (LPS) of the outer membrane that covers the thin peptidoglycan wall of Gram-negative bacteria. Small amounts are shed as the bacteria grow, and a large amount is released when they die and their walls break down.

Exotoxin

A toxic protein secreted by a living bacterium, acting on a specific target in the host; Staphylococcus species are the standard example.

Endotoxin

The lipopolysaccharide of the outer membrane of a Gram-negative bacterium, released mainly when the bacterium dies; it causes fever and inflammation. Salmonella species are the standard example.

Toxoid

An exotoxin treated so that it is no longer toxic but keeps its antigens, used as a vaccine against diseases such as tetanus and diphtheria.

Staphylococcus species are the standard exotoxin producers. Staphylococcus aureus lives harmlessly on the skin and in the nose of many people, but in a wound or in food it secretes several exotoxins with different targets. Some make pores in host cell-surface membranes, so the cells lyse. Others are enterotoxins: a strain growing in food left at room temperature secretes toxin into the food, and eating it causes vomiting within a few hours, because the toxin is already made and nothing has to grow in the gut first. The toxin behind toxic shock syndrome overstimulates T cells, which release cytokines in such quantity that blood pressure falls dangerously. The staphylococcal food toxins are unusually stable to heat for proteins, so reheating food can kill the bacteria and leave the toxin active.

Salmonella species are the standard example of endotoxin. They are Gram-negative rods; most cases of Salmonella food poisoning come from undercooked poultry and eggs, and one serotype, Salmonella Typhi, causes typhoid. The bacteria swallowed in food have to multiply and invade the cells lining the intestine before anything happens, so symptoms usually begin between 12 hours and 3 days after eating. As the bacteria are killed by the host's defences, lipopolysaccharide is released. Its lipid part binds to receptors on macrophages and other cells, which respond by releasing cytokines. The result is inflammation of the gut lining, with diarrhoea and abdominal cramps, and fever. If large amounts of endotoxin reach the blood, widespread dilation of blood vessels can lead to septic shock. The damage is done by the host's response to a part of the bacterium.

The third route is invasion of host tissue, and Mycobacterium tuberculosis is the example: it releases no major toxin, but grows inside the macrophages of the lung, and the granulomas and tissue destruction of tuberculosis are the result of that invasion and of the host's long inflammatory response to it, as described above.

Stem rust: a fungal disease of wheat

Puccinia graminis, the stem rust fungus, infects wheat, and related forms attack barley, oats and rye. It has been one of the most damaging crop diseases in history, because a field that looks healthy can be ruined in the few weeks before harvest. Its full life cycle passes through a second host plant, barberry, but at A level only transmission, infection and effect are needed.

Transmission. On an infected plant the fungus forms pustules that burst through the epidermis of the stem and leaves and release masses of rust-red spores. The spores are dry and light and are carried by the wind, to the next plant, the next field, and on air currents for hundreds of kilometres. In warm weather a new generation of spores is produced within one to two weeks of infection, so the number of spores in a region can rise very fast once an epidemic starts.

Mode of infection. A spore landing on a wheat stem or leaf germinates only in a film of water, which is why outbreaks follow warm weather with dew or rain. The germ tube grows across the surface to a stoma and enters through it, so the fungus never has to break the waxy cuticle. Inside, hyphae grow through the spaces between the cells and push branches called haustoria through the cell walls. The host cell membrane folds around each haustorium rather than breaking, so the host cell stays alive, and sugars and amino acids are absorbed from it. The fungus can feed only on living tissue.

Pathogenic effect. Sugars drawn off by the fungus are sugars not translocated to the developing grain, so the grains are small and shrivelled. The infected tissue photosynthesises less. Pustules rupture the epidermis, so the plant loses water through the breaks whether or not its stomata are closed. Weakened stems break or fall over in wind and rain, which is called lodging, and a severe attack can destroy most of a crop.

Stem rust

A disease of wheat and other cereals caused by the fungus Puccinia graminis, spread by wind-borne spores and entering through stomata.

Haustorium

A branch of a fungal hypha that pushes into a living host plant cell, inside a fold of the host's cell-surface membrane, and absorbs nutrients from it.

Control relies mainly on resistant varieties of wheat, which carry resistance genes (called Sr genes) whose products let the plant recognise the fungus and set off its defences, with fungicides as a second line. Stem rust became uncommon in many wheat-growing regions once resistant varieties were planted widely in the second half of the twentieth century. In 1999 a new race of the fungus, named Ug99 after Uganda where it was identified, was found to overcome Sr31, a resistance gene carried by much of the world's wheat, and it has since spread through eastern Africa and into the Middle East. Widely grown resistant varieties select for any variant of the fungus that overcomes the resistance, and Ug99 is such a variant.

Physical, chemical and biological barriers

Before the specific immune response is involved, a pathogen must cross defences that act on all pathogens equally. They are classified as physical, chemical and biological.

Describe the drawing

Four stacked bands with arrows running down between them. The top band is physical barriers: skin, mucus and cilia, and clotting. The second is chemical barriers: stomach acid near pH 2 and lysozyme in tears and saliva. The third is the non-specific cellular response of inflammation and phagocytosis. The bottom band, reached only after a phagocyte presents an antigen, is the specific response.

Physical barriers keep pathogens out by being in the way. Skin is the obvious one: its outer layer is dead, dry, and packed with keratin, which almost nothing can digest. Where the body has to be moist and permeable, as in the airways, gut and reproductive tract, it uses mucous membranes instead. Goblet cells secrete sticky mucus that traps particles, and the ciliated epithelium beneath beats the mucus upwards to the throat, where it is swallowed. Damage that cilia layer, as cigarette smoke does, and the trapped material stays put.

A wound is a hole in the barrier, and blood clotting seals it within minutes. Platelets at the damaged vessel release substances that set off a cascade of reactions in the plasma, ending in the formation of insoluble fibrin. The fibrin fibres form a network that traps platelets and blood cells into a clot, which dries into a scab. Under the scab, wound repair begins: cells in the deeper layers of the skin divide to replace the lost tissue, and collagen is laid down to strengthen it. Expulsive reflexes remove what has already got in: coughing and sneezing expel irritants and pathogens from the airways, and vomiting and diarrhoea expel them from the gut.

Chemical barriers destroy pathogens rather than blocking them. Stomach acid holds the gut contents near pH 2, which denatures the enzymes and other proteins of most swallowed bacteria. Mucus cleared from the airways is swallowed, so bacteria trapped in it also reach the stomach: the ciliated epithelium removes them from the airways and the acid destroys them in the stomach. Lysozyme, an enzyme in tears, saliva and mucus, hydrolyses bonds in bacterial cell walls so the cell bursts. Sebum makes the skin surface slightly acidic and inhospitable.

If a pathogen does get through, the response at the site of damage is inflammation. Mast cells in the damaged tissue release histamine, which dilates arterioles and makes capillary walls more permeable. More blood arrives, so the area is red and warm, and more tissue fluid leaks out, so it swells. That fluid carries antibodies into the infected tissue, and cytokines released at the site attract phagocytes, which squeeze out of the capillaries between the cells of their walls. The swelling is part of the response rather than incidental damage.

A raised body temperature has two effects: most pathogens reproduce more slowly above about 37 °C, and the specific immune response operates faster at the higher temperature. Very high fevers are dangerous because human enzymes also denature.

Interferon: a warning to neighbouring cells

Inflammation, lysozyme and phagocytosis all act on pathogens outside cells. A virus that has entered a cell is out of their reach, and the infected cell's first response is a signal to the cells around it. The cell detects viral nucleic acid in its cytoplasm, in particular double-stranded RNA, which human cells do not normally contain, and responds by transcribing the genes for interferon. Interferons are small proteins, one class of cytokine, and the infected cell secretes them into the tissue fluid.

The infected cell gains little from this; the interferon acts on its neighbours. It diffuses to nearby cells and binds to receptors on their cell-surface membranes, which switches on a set of genes inside each one. The proteins they code for put the cell into an antiviral state. One is an enzyme that stops protein synthesis at the ribosomes as soon as viral double-stranded RNA appears; another activates a ribonuclease that hydrolyses RNA, so viral mRNA is destroyed. A virus that then enters one of those cells cannot be replicated, because it has no ribosomes or enzymes of its own and depends entirely on the host cell's. Interferon also makes cells display more antigen on their surface, which helps T-killer cells find the infected ones, and it activates macrophages and natural killer cells.

Interferon

A protein secreted by a virus-infected cell that binds to receptors on neighbouring cells and makes them resistant to viral replication, by inhibiting protein synthesis and destroying viral RNA.

Interferon is non-specific. It is released in response to many different viruses, and the antiviral state it produces works against all of them. It does not bind the virus and is not an antibody: it acts on host cells, not on the pathogen. It appears within hours of infection, days before any antibody, and holds the spread of a virus back while the specific response develops. Several symptoms of a viral infection come from it: the aching, fever and tiredness of influenza are partly the effect of interferon and other cytokines, and patients given interferon as a drug report much the same symptoms.

Interferons were discovered in 1957 by Alick Isaacs and Jean Lindenmann in London, who found that cells exposed to inactivated influenza virus released something that interfered with infection by live virus, which gave the protein its name.

Phagocytosis, stage by stage

Phagocytes are white blood cells that engulf and digest pathogens. Neutrophils are the short-lived ones that arrive first and die in large numbers, which is what pus is; macrophages are larger, live longer, and present antigen to lymphocytes. The process is the same in both, and is described as an ordered sequence.

Describe the drawing

Six numbered rows, each with a small drawing of a phagocyte on the left. The cell moves along a chemical gradient, binds the pathogen's antigens with its receptors, folds its membrane round the pathogen to form a phagosome, has a lysosome fuse with that phagosome, hydrolyses the pathogen and absorbs the products, and finally displays the pathogen's antigens on its own surface membrane.

Phagocytosis

The engulfing and hydrolysis of a pathogen or other foreign material by a phagocyte, forming a phagosome that fuses with a lysosome.

Phagosome

The vesicle formed when a phagocyte's cell-surface membrane closes around an engulfed pathogen.

Lysosome

An organelle containing hydrolytic enzymes, including lysozymes, kept separate from the cytoplasm.

Antigen-presenting cell

A cell displaying antigens from a pathogen or other non-self material on its own cell-surface membrane, such as a phagocyte after phagocytosis, an infected body cell or a B cell that has taken in the antigen.

In words: the phagocyte moves towards the pathogen along a concentration gradient of chemicals released by the pathogen itself or by damaged tissue, which is chemotaxis. Receptors on the phagocyte's membrane attach to the pathogen's surface: some recognise molecular patterns shared by whole classes of microbe, others grip antibodies or complement proteins already coating the pathogen, coatings called opsonins that make the hold much firmer. The membrane then folds around it, and the two edges fuse to enclose the pathogen in a vesicle, the phagosome. Lysosomes move to the phagosome and fuse with it, emptying hydrolytic enzymes inside. Those enzymes hydrolyse the pathogen, and the soluble products are absorbed into the cytoplasm.

Two details matter. The enzymes remain enclosed by membrane throughout, so the phagocyte is not hydrolysed. And engulfing is endocytosis: it requires ATP and a flexible membrane, which a plant cell with a rigid cell wall does not have.

Antigen presentation

After hydrolysis, the phagocyte moves some of the pathogen's antigens to its own cell-surface membrane and displays them there. That is antigen presentation, and it turns the phagocyte into an antigen-presenting cell.

Presentation is required because of how a complementary lymphocyte is found. Millions of different T cells circulate, each with a differently shaped receptor, and the probability of the complementary one encountering free pathogen in an infected tissue is low. Presented antigen, concentrated on the surface of a cell that then travels to a lymph node full of lymphocytes, makes the meeting far more likely. In the A-level account, nothing specific begins until this has happened: no clonal selection, no antibody, no memory cell. (In the fuller picture, dendritic cells are the most effective presenters, and a B cell can also bind free antigen directly with its own receptor; the macrophage route here is the examined one.)

Phagocytosis therefore has two functions. It removes pathogens immediately, at the same rate on every exposure, and it initiates the specific response, which is faster on a second exposure. Antigen presentation is the step that links the non-specific response to the specific one.