II. BACTERIAL PATHOGENESIS
A. AN OVERVIEW AND QUORUM SENSING
The overall purpose of this Learning Object is to:
1) Introduce the concept bacteria virulence and pathogenicity; and
2) Introduce how quorum sensing, pathogenicity islands, and secretion systems in bacteria promotes pathogenicity.
LEARNING OBJECTIVES FOR THIS SECTION
Bacteria are found in almost every environment. Only a relatively few bacteria cause human disease and many benefit humans. For example, many are important decomposers that assure the flow and recycling of nutrients through ecosystems. Others have important industrial and pharmaceutical uses. However, in this section we are going to concentrate on bacteria that are potentially harmful to humans and try to understand what factors influence their ability to cause disease.
Pathogenicity and virulence are terms that refer to an organism's ability to cause disease. Technically, pathogenicity (def) is used with respect to differences between microbial species whereas virulence (def) denotes differences between strains of the same species. In practice they are often used interchangeably.
To cause disease, an organism must:
1. Maintain a reservoir before and after infection (humans, animals, environment, etc.),
2. Leave the reservoir and gain access to the new host,
3. Colonize the body, and
4. Harm the body.
Anything the bacterium does to aid in the above will influence its ability to cause disease and they are able to do these things primarily as a result of their structures and their metabolic products. We must keep in mind, however, that whether or not a person actually contracts an infectious disease after exposure to a particular potentially pathogenic bacterium depends not only on the microorganism, but also on the number of bacteria that enter the body and the quality of the person's innate and adaptive immune defenses. (Innate and adaptive immunity will be discussed in detail later in Units 4 and 5.)
For example, if relatively few bacteria enter the body then the body's natural defenses against infection have a much better chance of removing them before they can colonoize, multiply, and cause harm. On the other hand, if a large number of bacteria enter then the body's defenses may not be as successful.
Likewise, a person with good innate and adaptive immune defenses will be much more successful in removing potentially harmful bacteria than a person that is immunocompromized. In fact a person highly immunosuppressed, such as a person taking immunosuppressive drugs to suppress transplant rejection, or a person with advancing HIV infection, or a person with other immunosuppressive disorders, becomes very susceptible to infections by microorganisms generally considered not very harmful to a healthy person with normal defenses.
However, in this section we are going to look at bacterial virulence factors that can influence its ability to cause infectious disease. These virulence factors will be divided into two categories:
A. Virulence factors that promote bacterial colonization of the host.
There are 6 factors we will eventually discuss in this unit that will promote bacterial colonization of humans:
1. The ability to use motility and other means to contact host cells and disseminate within a host
2. The ability to adhere to host cells and resist physical removal.
3. The ability to invade host cells
4. The ability to compete for iron and other nutrients
5. The ability to resist innate immune defenses such as phagocytosis and complement
6. The ability to evade adaptive immune defenses
B. Virulence factors that damage the host.
There are 3 factors we will eventually discuss in this unit that can result in harm to humans:
1. The ability to produce cell wall components (pathogen-associated molecular patterns or PAMPS) that bind to host cells causing them to synthesize and secrete inflammatory cytokines and chemokines
2. The ability to produce harmful exotoxins
3. The ability to induce autoimmune responses
Most of the virulence factors we will discuss in this unit that enable bacteria to colonize the body and/or harm the body are the products of quorum sensing genes. Many bacteria are able to sense their own population density, communicate with each other by way of secreted chemical factors, and behave as a population rather than as individual bacteria . This plays an important role in pathogenicity and survival for many bacteria.
Bacteria can behave either as individual single-celled organisms or as multicellular populations. Bacteria exhibit these behaviors by chemically "talking" to one another through a process called quorum sensing. Quorum sensing (def) involves the production, release, and community-wide sensing of molecules called autoinducers (def) that modulate gene expression, and ultimately bacterial behavior, in response to the density of a bacterial population.
To initiate the process of quorum sensing, bacterial genes code for the production of signaling molecules called autoinducers (def) that are released into the bacterium's surrounding environment.These signaling molecules then bind to signaling receptors either on the bacterial surface or in the cytoplasm. When these autoinducers reach a critical, threshold level, they activate bacterial quorum sensing genes that enable the bacteria to behave as a multicellular population rather than as individual single-celled organisms (see Fig. 1). The autoinducer/receptor complex is able to bind to DNA promoters and activate the transcription of quorum sensing-controlled genes in the bacterium. In this way, individual bacteria within a group are able to benefit from the activity of the entire group.
1. In Gram-negative bacteria, the autoinducers are typically molecules called acyl-homoserine lactones or AHL. AHLs diffuse readily out of and into bacterial cells where they bind to AHL receptors in the cytoplasm of the bacteria. When a critical level of AHL is reached, the cytoplasmic autoinducer/receptor complex functions as a DNA-binding transcriptional activator.
2. In Gram-positive bacteria, the autoinducers are oligopeptides, short peptides typically 8-10 amino acids long. Oligopeptides cannot diffuse in and out of bacteria like AHLs, but rather leave bacteria via specific exporters. They then bind to autoinducer receotors on the surface of the bacterium. When a critical level of oligopeptide is reached, the binding of the oligopeptide to its receptor starts a phosphorylation cascade that activates DNA-binding transcriptional regulatory proteins called response regulators.
The outcomes of bacteria-host interaction are often related to bacterial population density. Bacterial virulence, that is its ability to cause disease, is largely based on the bacterium's ability to produce gene products called virulence factors that enable that bacterium to colonize the host, resist body defenses, and harm the body.
At a low density of bacteria, the autoinducers diffuse away from the bacteria (see Fig. 2). Sufficient quantities of these molecules are unable to bind to the signaling receptors on the bacterial surface and the quorum sensing genes that enable the bacteria to act as a population are not activated. This enables the bacteria to behave as individual, single-celled organisms.
Possible advantages of individual bacterial behavior:
If a relatively small number of a specific bacterium were to enter the body and immediately start producing their virulence factors, chances are the body's immune systems would have sufficient time to recognize and counter those virulence factors and remove the bacteria before there was sufficient quanity to cause harm. The bacterium instead utilizes genes that enable it to act as an individual organism rather than as part of a multicellular population.
Acting as individual organisms may better enable that low density of bacteria to gain a better foothold in their new environment in the following ways:
1. Many bacteria are capable of motility and motility serves to keep bacteria in an optimum environment via taxis (def).
Motility and chemotaxis probably help some intestinal and urinary pathogens to move through the mucous layer so they can attach to the epithelial cells of the mucous membranes. In fact, many bacteria that can colonize the mucous membranes of the bladder and the intestines are motile. Motility probably helps these bacteria move through the mucus in places where it is less viscous.
2. One of the body's innate defenses is the ability to physically remove bacteria from the body through such means as the constant shedding of surface epithelial cells from the skin and mucous membranes, the removal of bacteria by such means as coughing, sneezing, vomiting, and diarrhea, and bacterial removal by bodily fluids such as saliva, blood, mucous, and urine. Bacteria may resist this physical removal by producing pili (def) (see Fig. 3), cell wall adhesin proteins (def) (see Fig. 4), and/or biofilm-producing capsules (def). Some pili, called type IV pili also allow some bacteria to "walk" or "crawl" along surfaces to spread out and eventually form microcolonies.
3. Many bacteria secrete an extracellular polysaccharide or polypeptide matrix called a capsule or glycocalyx that enables the bacteria to adhere to host cells, resist phagocytosis, and form microcolonies (def).
As the bacteria geometrically increase in number by binary fission, so does the amount of their secreted autoinducers, and production of high levels of autoinducers then enables the population of bacteria to communicate with one another by quorum sensing.At a high density of bacteria, large quantities of autoinducers are produced (see Fig. 5) and are able to bind to the signaling receptors on the bacterial surface in sufficient quantity so as to activate the quorum sensing genes that enable the bacteria to behave as a multicellular population (see Fig. 1).
Advantages of multicellular behavior:
1. By behaving as a multicellular population, individual bacteria within a group are able to benefit from the activity of the entire group. As the entire population of bacteria simultaneously turn on their virulence genes, the body's immune systems are much less likely to have enough time to counter those virulence factors before harm is done.
2. Virulence factors such as exoenzymes and toxins can damage host cells enabling the bacteria in the biofilm to obtain nutrients.3. As the area becomes over-populated with bacteria, quorum sensing enables some of the bacteria to escape the biofilm and return to individual single-celled organism behavior in order to find a new sight to colonize.
Pseudomonas aeruginosa is an example of a quorum sensing bacterium. P. aeruginosa causes severe hospital-acquired infections, chronic infections in people with cystic fibrosis, and potentially fatal infections in those who are immunocompromised.
1. P. aeruginosa first enters the body functioning as individual bacteria. Motility genes (coding for flagella) and adhesin genes (coding for pili and cell wall adhesins) are expressed. The flagella enable the initial bacteria to swim through mucus towards host tissues such as mucous membranes. Pili then enable the bacteria to reversibly attach to host cells in order to resist flushing and begin colonization (See Fig. 6A). Type IV pili, which enable a twitching motility in some bacteria, then enable the bacteria as they replicate to crawl along and spread out over the mucous membranes (See Fig. 6B). The pili subsequently retract and bacterial cell wall adhesins enable a more intimate attachment of the bacterium to the mucous membranes (See Fig. 6C).
2. Once P. aeruginosa is able to replicate and achieve a high population density, quorum sensing genes trigger production of an extracellular polysaccharide called alginate to form microcolonies and enable irreversible attachment to the mucous membranes (See Fig. 6D).Biofilm (def) formation begins.
3. Quorum sensing genes coding for enzymes and toxins that damage host cells are produced. These are injected into the host cells by way of an injectosome. This releases nutrients for the bacteria in the biofilm. The bacteria continue to replicate as the biofilm continues to develop, mushroom up, and mature (See Fig. 6E).
4. As the bacteria replicate, the biofilm continues to mature (See Fig. 6F). Water channels form within the biofilm to deliver water, oxygen, and nutrients to the growing population of P. aeruginosa. The high density of bacteria bacteria are now acting as a multicellular population rather than as individual bacteria.
- Electron micrograph of a biofilm of Haemophilus influenzae from Biomedcentral.com
The biofilm enables bacteria to:
- resist attack by antibiotics;
- trap nutrients for bacterial growth and remain in a favorable niche;
- adhere to environmental surfaces and resist flushing;
- live in close association and communicate with other bacteria in the biofilm; and
- resist phagocytosis and attack by the body's complement pathways.
5. When the population of P. aeruginosa begins to outgrow their local environment, quorum sensing enables them to turn off adhesin genes and turn on flagella genes that allow some of the bacteria to spread out of the biofilm to new location within that environment via motility (See Fig. 6G and Fig. 6H).
It turns out that bacteria are multilingual. They use quorum sensing not only to "talk" to members their own species (intraspecies communication), but also to "talk" to bacteria that are not of their genus and species (interspecies communication). Intraspecies autoinducers and receptors enable bacteria to communicate with others of their own species while interspecies autoinducers and receptors enable bacteria to communicate with bacteria of a different species or genus (see Fig. 7). The autoinducers for interspecies communications are referred to as AI-2 family autoinducers and are different from the intraspecies (AI-1) autoinducers. In some cases bacteria use interspeciecies communication to work cooperatively with various other bacteria in their biofilm to the benefit all involved; in other cases, bacteria may use interspecies communication in such a way that one group benefits at the expense of another.
Furthermore, bacteria are capable of interkingdom communication, communication between bacteria and their animal or plant host. Increasing numbers of bacteria are being found that have signaling receptors that recognize human hormones. For example, a number of bacteria that are pathogens of the human intestinal tract have a sensing molecule called QseC that binds the human hormones adrenaline and noradrenaline. This, in turn, activates various virulence genes of the bacteria. On the other hand, some bacterial autoinducers can enter human host cells and regulate human cellular function. For example, at low concentration some bacterial autoinducers supress host immune responses thus better enabling those bacteria to better establish themselves in the body. At high concentrations, however, they stimulate an inflammatory response in the host to help the bacteria to spread from the initial infection site. One bacterial autoinducer has been found to initiate apoptosis (cell suicide) in phagocytes such as neutrophils and macrophages.
Most genes coding for virulence factors in bacteria are located in pathogenicity islands or PAIs (def) and are usually acquired by horizontal gene transfer (def). The genomes of most pathogenic bacteria contain multiple PAIs, accounting for up to 10 - 20% of the bacterium's genome. Conjugative plasmids (def) are the most frequent means of transfer of PAIs from one bacterium to another.
Many bacteria involved in infection have the ability to co-opt the functions of host cells for the bacterium’s own benefit. This is done by way of bacterial secretions systems that enable the bacterium to directly inject bacterial effector molecules into the cytoplasm of the host cell in order to alter its cellular machinery or cellular communication.
The most common type is the type 3 secretion system (see Fig. 8). A secretion apparatus in the cytoplasmic membrane and cell wall of the bacterium polymerizes a hollow needle that is lowered to the cytoplasmic membrane of the host cell and a translocon protein is then delivered to anchor the needle to the host cell. Effector proteins in the bacterium can now be injected into the cytoplasm of the host cell. The delivery system is sometimes called an injectisome. (A type 4 secretion system can transfer effector proteins and/or DNA into the host cell because it is similar to the conjugation transfer system initiated by tra genes discussed in Unit 1 under horizontal gene transfer.)
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