Bacterial invasion and overcoming the host’s defenses

The bacteria got to the host, they attached and multiplied. But even “good” bacteria do that, so where’s the disease?

Bacterial invasion of the host

Once pathogenic bacteria have colonised, multiplied and reached an infectious dose on the surface, they then invade the host.  Factors involved in cell invasion include the following:

1. External factors. Some bacteria in themselves are not invasive, but enter the host i.e. in the mouth parts or the saliva of biting insects and arachnids; iatrogenically via hypodermic needles; and animal bites and other forms of trauma.

2. Use of motility. Leptospira, a bacterium that has its flagellae trapped under its outer membrane is able to move through viscous substances by a “corkscrew” movement.

3. Invasins. Most pathogenic bacteria use  invasins. ​​​​​​​Invasins are extracellular molecules such as enzymes that are produced by bacteria to facilitate invasion. Another way bacteria use invasins is to transport themselves into the intracellular compartment of cells. This will be dealt with separately.

Definitions for terms used in this workbook

These definitions are aimed at assisting you in understanding the terms that you have read. Unless defined elsewhere in any the workbooks, you will not have to learn them.

You can test your knowledge on the definitions by filling in this crossword puzzle.

Actin: A protein found in eukaryotic cells that forms a microfilament increasing cell plasticity and contractibility.

Antigenic variation: This is a process by which a micro-organisms varies its surface carbohydrates or proteins, allowing it to evade the host’s immune responses.

Cystic fibrosis: a hereditary disorder in people affecting the exocrine glands. It causes the production of abnormally thick mucus, leading to the blockage of the pancreatic ducts, intestines, and bronchi and often resulting in respiratory infection. ​​​​​​​

Haematogenous spread: Usually refers to transport of pathogens or cancerous cells in the bloodstream to the rest of the body.

Iatrogenic: Adverse effects on the animal brought about by the actions of animal health workers or treatments. Amended to be animal-centric.

Immune-privileged site. These are sites in the body that tolerate antigens, such as bacteria, without eliciting an inflammatory response. It includes the eyes, the placenta and foetus, and the testicles.

Invasins – Molecules produced by bacteria that assist in their invasion of host tissue

Ischaemic necrosis: A condition where there is restriction of blood flow to cells resulting in their death.

Lymphogenous spread: Usually refers to the transport of pathogens or cancer cells via the lymphatic system to the rest of the body.

Mastitis: Inflammation of the mammary gland (udder), often caused by bacteria.

Mucociliary esculator: It consists of the ciliated epithelium lining the airways. This epithelium is rich in Goblet cells. Mucus from the Goblet cells traps foreign particles like bacteria and the cilia beat upwards moving the mucus with the trapped particles. These are then swallowed.

Opportunistic infection: These are infections by infecting agents that make use of an opportunity, not usually present, to invade and establish disease in a host. This often happens when the host is immunocompromised, the normal microbiota are disturbed or there is a breach in epithelial barriers. Opportunistic pathogens often are part of the normal host microbiota or are found  in the environment.

Polyarthritis: Inflammation of several joints. It can either be caused by an autoimmune disease or by infectious agents that have localised in several joints after haematogenous spread.

Pyoderma:  a bacterial skin inflammation marked by pus-filled lesions. Pyoderma is often classified as superficial or deep; and localised or generalised.

Septicaemia: The presence of bacteria and their toxins in the bloodstream resulting in systemic disease.

Trigger mechanism:  Bacterial effector proteins stimulate cell membrane enclosure of bacteria into an intra-cytoplasmic vacuole.

Zipper mechanism: Gradual cell membrane enclosure of adhered bacteria into a intra-cytoplasmic vacuole.

how bacteria use invasins to invade and spread in their hosts

Bacteria use a variety of ways to enter host cells, and some bacteria will use more than one type of invasin.

1. Cell invasion by enzyme production

For example, rapidly dividing Clostridium chauvoei the cause of blackquarter in cattle produces phospholipases and lecthinases that punch holes in cell membranes of muscle and blood vessels causing them to lyse.

Examples of bacterial invasins are in the Table below.

2. Invasion of cells

Intracellular invasion and survival by bacteria allow them to invade across epithelial barriers and evade the humoral immune response. Bacteria will also use survival in phagocytes as a means to disseminate throughout the body.

Bacteria contact and adhere to the host cell by the binding of a specific bacterial surface protein to an eukaryotic cell membrane receptor.  Phagocytosis is then effected using a zipper or trigger mechanism.

1. The zipper mechanism is used by Listeria (epithelial cells) and Yersinia (enterocytes) where modest cell extensions and cytoskeletal rearrangements progressively attach to the bacterial cell surface eventually engulfing the bacterium in a vacuole. the binding of the host membrane to the bacterial surface sometimes requires additional molecules to bind to the bacterium. Fibronectin is the bridging molecule for the phagocytosis of staphylococci and streptococci. These bacteria produce fibronectin binding protein which facilitates attachment and thus  phagocytosis. Mycobacteria actually bind to fibronectin on M-cells within the Peyer’s patches allowing them to be phagocytosed.
2. The trigger mechanism is used by gram-negative bacteria such as Salmonella that use a Type 3 secretory system (T3SS) injector to inject an effector molecule into the host cell. This effector molecule induces actin polymerisation and remodeling of the cell cytoskeleton to engulf the bacterium. For example, Burkholderia pseudomallei, the agent of melioidiosis, attaches to macrophages and other cells and enters them by causing changes within the actin cytoskeleton through a T3SS effector.  Either it will exit the cells by cell lysis or it moves from cell to cell using an actin tail (much like Listeria does). The latter form of spread leads to cells fusing with the formation of multinucleate giant cells.

3. A third mechanism is used by clostridia that injects an effector protein using a porin (see later) resulting in damage of the villi and allows an extension of the actin out of the cell which then wraps itself around the bacterium and draws it into the cell.

3. Survival within the cell

The vacuole formed progressively acidifies as it develops into a mature degradative phagolysosome. Some pathogens survive in the phagosome either by preventing vacuole–lysosome fusion or by modifying the environment within the phagolysosome. Some bacteria such as Listeria monocytogenes will escape from the vacuole to survive in the cytoplasm of the host cell. Escape from the phagolysosome must happen rapidly before the bacterium is destroyed i.e. within 30 minutes of cell internalisation and is usually mediated by bacterial enzymes.

For example,  L. monocytogenes produces listeriolysin O in response to γ-interferon-inducible lysosomalthiol reductase and the acidic environment of the phagolysosome. Listeriolysin O inserts itself into the phagolysosome membrane and forms pores that delay phagosome-lysosome maturation. Broad-range phosphatidylcholine-specific phospholipase C (PC-PLC) will then allow L. monocytogens to escape from the phagolysosome.

How bacteria overcome the host’s defense mechanisms

The host is in a constant battle to ward off invasive infectious agents, including bacteria. On the whole, the host is highly successful. You would have learnt in immunology about all the strategies used by the host both in the innate and adaptive immunity.  You will require a good understanding of the host’s immune defenses to understand that for each host defence mechanism there are pathogenic bacteria that are able to overcome it.

Since mucosal immunity is a major first line of defence against bacterial pathogens, I recommend reading the following review article on mucosal immunity: Perez-Lopez A, Behnsen J, Nuccio S-P, Raffatellu M (2016)  Mucosal immunity to pathogenic intestinal bacteria. Nature Reviews: Immunology. 16: 135-148.

Surface structures bacteria use to fool or overcome host defences

Different pathogenic bacteria have a variety of structures that affect the immune system. One of the strategies is to hide from or fool the immune system, bacteria can do this in several ways:

1. Stealth

1. Hide in a site that has the least contact with the immune system = sequestration. I.e. Leptospira hides in the anterior and posterior chambers of the eye a so-called immune-privileged site.
2. Hide within an intracellular location.
3. Cover oneself with poor antigens. The capsule of the gram-negative bacteria Escherichia coli and Klebsiella pneumoniae produce a polysaccharide capsule that is weakly antigenic.
4. Mimic the surface structure of the host cells = molecular mimicry. The cell membrane of the wall-less Mycoplasmopsis (Mycoplasma) bovis, has proteins that are similar in structure to outer membrane proteins of lung cells,

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B. Surface antigenic variation

Change their surface proteins or carbohydrates = antigenic variation before immune-mediated destruction occurs. The agent of Q-fever, Coxiella burnetii is known to do this.

C. Subversion of host’s defences

Several surface structures are involved in subversion of the immune response. They are discussed in the following section.

Antiphagocytic responses by bacteria

Extracellular bacteria have developed strategies to avoid being phagocytosed.

1. Most of the toxins produced in this group are porin toxins and will destroy a particular type of cell. Since neutrophils and macrophages are involved in protection against bacterial infections, it is these cells that are targeted by the leukocidins or leukotoxins of pathogenic bacteria. Leucocidins are produced by staphylococci, streptococci and Fusobacterium. They destroy neutrophils and contribute to pus formation. The leukotoxin of Mannheimia haemolytica, the main cause of feedlot pneumonia in cattle, at low doses stimulates alveolar macrophages to release cytokines, but at high doses will act as a porin toxin.
2. Enzymes produced by bacteria can reduce the efficacy of the immune response. For example, catalase produced by most aerobic bacteria will counteract the oxidative burst reaction of neutrophils by inactivating the peroxides.
3. Bind to and inactivate antibodies. Protein A of Staphylococcus aureus binds to IgG preventing complement mediated opsonisation.
4. Distance activation of Complement by the polysaccharide chain of the gram-negative endotoxin i.e. smooth strains of Escherichia coli which have very long O-polysaccharide chains

Bacteria that survive within the intracellular environment allow themselves to be phagocytosed, but have developed ways to survive and even thrive in the intracellular niche. They do this by using a single or many of the following strategies:

1. Don’t activate the macrophages
2. Inhibiting phagosome-lysosome fusion
3. Resist destruction in the phagosome. The murein in the outer cell membrane of streptococci is such a structure.
4. Escape using porins from the phagosome

Mycobacteria the cause of chronic granulomas in people and animals persist in its host by surviving within macrophages. Early in the disease they fail to activate the macrophages and are able to prevent phagosome-lysosome fusion, later in the disease when the macrophages are activated they are able to survive in the phagolysosomes. They are also able to escape from the phagosome.

However, cells do have means to protect themselves against intracellular bacteria, such as Salmonella and Mycobacterium.  This is usually by them producing a doubled layered vacuole known as an autophagosome that surrounds and sequester damaged phagocytic vacuoles containing bacteria or even free bacteria in the cytoplasm. Lysosomes will fuse with the autophagosomes to destroy its contents. Nevertheless, in the cat-and-mouse world of pathogens and host cells, some bacteria can use this autophagy to their own advantage. They can produce intracellular toxins like the oedema toxin of Bacillus anthracis, the agent of anthrax, that stops the initiation of autophagy in a cell; they can mask themselves from autophagy by recruiting cell proteins around themselves i.e. ActA of Listeria monocytogenes;  they stop autophagosome fusion with lysosome i.e. some intracellular Escherichia coli strains will prevent maturation of the autophagosome. Some bacteria will even block autophagosome lysosome fusion and use the autophagosome to divide in i.e. Brucella species.

Bacterial dissemination in the body

Bacteria spread in the body from their point of entry following a number of routes:

1. Direct spread

2. Lymphogenous spread

3. Haematogenous spread

Direct spread of bacteria

Bacteria invading from the outside often spread by extension. For example, most cases of bacterial pneumonia in animals are caused by bacteria from the upper respiratory tract. These bacteria descend down the airways to the lungs when the mucociliary escalator fails. This is either due to concurrent viral or bacterial infections or as a result of physiological abnormalities i.e. cystic fibrosis patients are unable to trap and expel bacteria and thus are very prone to opportunistic infections. Horses travelling in a horse box for long distances and are not allowed to keep their heads down are prone to lower respiratory tract infections.

Furthermore, the same invasins that allowed bacteria to invade can also result in their localised spread.

Lymphogenous spread

The lymphatic system and associated lymph nodes often trap bacteria closest to the site of bacterial entry. These are known as the regional lymph nodes. If they are not successful in trapping the invading bacteria the bacteria will spread along the lymphatics and eventually drain into the heart where it will enter the bloodstream and disseminate further in the body. This is typical for many of the bacteria causing sepsis and is a reason why superficial lymph nodes should always be checked for swellings.

Haematogenous spread

Many bacteria will enter the bloodstream from any point (i.e. respiratory tract; g.i.t.) causing a bacteraemia and if their toxins are present, they cause a septicaemia. This typically occurs in young animals whose immune systems have not fully matured or have not received passive maternal immunity ( i.e. were not fed colostrum). Some bacteria will enter the lymphatics from a localised site i.e. the skin and then enter the bloodstream via the thoracic duct and then spread haematogenously (via the bloodstream). Animals that are older or partially immune tend to not be very ill when the bacterium traverses the bloodstream but may develop lesions at various sites in the body. Common sites of organ localisation include the joints, where several are affected (polyarthritis), the brain and meninges, the kidneys. Other sites include the skin, liver, lungs and gravid uterus. The spleen as part of the immune system acts as a trap for bacteria within the blood and lymphatic circulation.