Exam 2: Innate Immunity: the First Lines of Defense

Pseudomonas aeruginosa is a Gram-negative bacterium that causes severe, and often life-threatening infections in immunocompromised individuals. In susceptible individuals, P. aeruginosa can establish infections in a wide range of tissues, including the lung, the GI tract, the eye, the ear, the urinary tract, the skin, and the blood. This bacterium is common in the environment, and is found on the skin of approximately 5% of healthy individuals. It is often found on hospital equipment, such as ventilators and catheters, and as a consequence, P. aeruginosa accounts for ~10% of hospital-acquired infections. To study the role of complement in the early innate immune response to P. aeruginosa, the following studies in mice were performed. Mice deficient in complement C3 or C5 (C3-/- or C5-/-, respectively) were infected by intratracheal inoculation with 10⁵ colony forming units (CFU) of P. aeruginosa, and survival was monitored over the first 72 hrs post-infection. The data from these studies are shown in Figure . Genetic data from human population studies also indicate that Individuals with genetic deficiencies in one of the collectins or ficolins show increased susceptibility to P. aeruginosa infections. a) Based on these data, evaluate the importance of complement in protection against P. aeruginosa infection, and describe the most likely complement pathway(s) involved in pathogen recognition and in pathogen destruction. Another group of individuals that are highly susceptible to P. aeruginosa infections are patients with the disease cystic fibrosis. These individuals suffer from the production of a thick mucus secretion in their lungs, which clogs the bronchial tubes. A similar increase in viscosity of bodily secretions is seen in these patients' sweat, digestive fluid, and gastrointestinal mucus. In these patients, the most common form of lung infection is that of P. aeruginosa. b) What is the most likely explanation for the increased susceptibility of cystic fibrosis patients to P. aeruginosa and other infections?

Our environment contains masses of microorganisms, many of which reside as commensal organisms on our body's mucosal and epithelial surfaces without causing disease. What two features distinguish a pathogenic microbe from these commensal microbes?

Multiple pathways for regulating complement activation limit the potential damage caused by complement deposition on host cells or caused by the spontaneous activation of complement proteins in the plasma. Genetic deficiencies in these mechanisms often lead to chronic inflammatory diseases, but in some cases can paradoxically lead to increased susceptibility to bacterial infections. This latter outcome may occur because:

One form of anemia results when individuals have a deficiency in the enzyme phosphatidylinositol glycan A (PIGA). This enzyme is required for the membrane attachment of proteins anchored by glycolipids to the plasma membrane, using what is called a 'GPI-linkage.' Included in the group of GPI-linked cell surface proteins is DAF/CD55. These individuals become anemic because:

Infants and young children with deficiencies in specific complement components present with recurrent respiratory infections caused by extracellular bacteria. The peak age of susceptibility is between 6 and 12 months after birth. At this time, as shown in Figure , maternal antibodies acquired by the child during fetal gestation are nearly gone, but the child is not yet generating robust antibody responses to new infections, as indicated by the low circulating levels of IgG and IgA. As children with this immunodeficiency get older, they outgrow this disease and show no further evidence of these recurrent infections. Based on this information, name one likely gene deficiency (in the complement system) that could cause this primary immunodeficiency, and the specific complement pathway likely to be affected. Explain your answer.

The formation of the C3 convertase is a key step in complement activation that occurs in all three complement pathways. This enzyme cleaves C3 in blood plasma, leading to a conformational change in the C3b fragment that exposes its reactive thioester group. The activated C3b is potentially harmful to the host, if it becomes covalently attached to a host cell, rather than to the surface of a pathogen. This deleterious outcome is largely avoided by:

Several pathogens produce proteins, either membrane-bound or secreted, that inactivate C3b that might be deposited on the pathogen surface. C3b is specifically targeted due to its central position in all three complement pathways.

The classical complement pathway is initiated by C1q binding to the surface of a pathogen. In some cases, C1q can directly bind the pathogen, for instance by recognizing proteins of bacterial cell walls, but in most cases C1q binds to IgM antibodies that are bound to the pathogen surface. How does this IgM-binding feature of C1q contribute to rapid, innate immune responses rather than to slow, adaptive responses?

The terminal components of the complement pathway assemble to form a membrane attack complex that can induce pathogen lysis and death. Yet, evidence indicates that this feature of complement is less important than the earlier steps that promote pathogen opsonization and induce inflammation. This conclusion is based on:

Four different clinical isolates of the Gram-positive bacterium, Staphylococcus aureus, are tested for their abilities to resist innate immune defense mechanisms. For these experiments, each bacterial strain is first grown in culture to achieve log-phase replication, and then cultures are supplemented with dilutions of human serum containing normal serum proteins as well as antibodies capable of binding to S. aureus. One hour later, the cultures are analyzed and the numbers of live bacteria are quantitated. The data from this experiment are shown in Figure . a) From these data, what general conclusions can be reached about the four strains of S. aureus? To identify the bactericidal mechanisms killing each strain of S. aureus, the serum is depleted of complement C3 by running it over an anti-complement C3 antibody affinity column. The experiment above is then repeated and the data are shown in Figure . b) What is the most likely mechanism accounting for the killing of strain D in this experiment? To determine whether strains A and C are susceptible to the same microbicidal pathway, the serum is depleted of antibody by running over an anti-human immunoglobulin affinity column. Following this treatment, it is found that strain A, but not strain C is still killed by incubation with the serum. c) From these data, what is the most likely mechanism killing S. aureus strain A? What about strain C?

The alternative pathway of complement activation has an important role in innate immunity, due to its ability to greatly amplify the amount of C3b deposited onto the pathogen surface. This amplification occurs because:

Recent studies using mouse models of pulmonary inflammation (a model for human asthma) have found that mice deficient in the C3a receptor have greatly reduced disease symptoms when challenged with inhaled preparations containing extracts of the fungal pathogen Aspergillus fumigatus. Specifically, the C3a receptor-deficient mice showed reduced influx of granulocytes and lymphocytes into the lung and reduced fluid in the lung after challenge. What is the explanation for these findings?