MicrobiologyBytes: Infection & Immunity: Complement Updated: October 21, 2004 Search

COMPLEMENT DEFICIENCIES

Both inherited and acquired disorders involving the complement system are associated with increased susceptibility to infections. Deficiencies in any defence mechanism of the host can lead to severe microbial infections; these are of clinical relevance. Although patients with these deficiencies are very uncommon, their identification and detailed study has proved enormously worthwhile. Deficiencies of complement proteins are frequently associated with an immunodeficiency state similar to that associated with immunoglobulin deficiency. These patients suffer with recurrent bacterial infections from organisms that are normally susceptible to opsonisation or lysis by complement.

The first descriptions of definite complement deficiency states during the early 1960s did not suggest a protective role of the complement system. On the contrary, deficiency of C1 inactivator function was found to be the cause of hereditary angio-edema (299), emphasising the importance of complement activation for development of inflammatory symptoms, while C2-deficient humans (300), C6-deficient rabbits (301), and C5-deficient mice (302) were found to be healthy. Even among workers in the complement field, the possibility was considered that a functional complement system might have "no more survival value to man than mid-digital hair" (303).

From the early 1970s a variety of hereditary and acquired complement deficiency states have been reported in conjunction with severe bacterial infections and, perhaps more surprisingly, in patients with disease conditions associated with autoimmunity and immune complex formation. Recognition of complement as a protective system of clinical importance was inaugurated by descriptions of C3 deficiency syndromes in patients susceptible to infection, with findings resembling those of hypogammaglobulinemia (304,305), the demonstration of hereditary C2 deficiency in patients with SLE (systemic lupus erythematosus)-like disease (306), and studies of acquired C3 deficiency due to C3 nephritic factors in relation to glomerulonephritis (307).

SURVEY OF COMPLEMENT DEFICIENCY STATES

Properdin deficiency states are X-linked, while other genetic defects within the complement system appear to be transmitted as autosomal recessive traits. Chromosome 1 contains the genes for C1q, C8, C4bp, factor H, CR1, CR2, and DAF, and chromosome 12 contains the genes for C1r and C1s. The gene for C3 has been localised to chromosome 19, factor I to chromosome 4, and C1 (IA) to chromosome 11. Four complement proteins, the C4A isotype, the C4B isotype, C2 and factor B, are encoded by genes within the major histocompatibility complex (MHC) on chromosome 6 (307).

Individuals with partial defects are not strictly complement-deficient. However, hereditary angio-edema is due to heterozygous deficiency or dysfunction of the C1 inhibitor protein (308). In addition, homozygous C4A and C4B deficiencies may have pathogenetic importance. Table 1). shows the principal deficiencies of circulating complement proteins. The prevalence of these conditions almost certainly varies in different populations, but has been estimated to be in the order of 0.03% (308).

Because complement deficiencies are relatively uncommon, much of the information about them has been derived through careful analysis of accumulated case reports. This approach introduces numerous opportunities for bias. For example, the ethnic background of the study population is a major determinant for both the prevalence of complement deficiency states as well as their associated diseases. Specifically, C2 deficiency occurs predominantly in Caucasian populations; in the U.S. its estimated frequency is 0.01%, whereas it has never been reported in the Japanese population (310,311). Similarly the yearly incidence of meningococcal disease differs among various ethnic populations (310).

Although the study of patients can introduce bias associated with a retrospective literature review, such an analysis can provide important clues to the contribution of individual complement components to overall complement functions and host defence as well as to the pathogenesis of neisserial infections.

A brief outline of the major complement deficiencies follows, although some will be discussed in more detailed later. Each of the five groups discussed differ in their functional defects as well as in their spectrum and prevalence of associated diseases. To date, individuals have been described with a complete deficiency of each plasma complement component except factor B (310).

CLASSICAL PATHWAY: Activation of the classical pathway promotes opsonophagocytic killing, serum bactericidal activity, and removal of immune complexes. Deficiencies of the classical pathway components have mainly been reported in association with immunological diseases such as SLE, glomerulonephritis and anaphylactoid purpura. SLE syndromes tend to be atypical with cutaneous manifestations being the most consistent feature (310). The frequency of homozygous C2 deficiency in patients with rheumatologic disease is about 0.2% or 20-fold more common than that in the general U.S. population (312). These individuals most frequently exhibit a syndrome consistent with systemic lupus erythematosus. Although homozygous C1 and C4 deficiencies are quite rare, reported individuals with these deficiencies also suffer principally from rheumatologic diseases (310,313). A review of all reported persons with a classical component deficiency reveals that about 66% have manifestations of a collagen vascular disease (314).

These individuals also exhibit increased susceptibility to infection. Recurrent sinopulmonary infections and meningitis caused by encapsulated organisms, especially Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae, predominate. Overall, 19 of individuals with classical component deficiencies have experienced at least one documented infection with encapsulated organisms; 4% of individuals have had both connective tissue disease and infection with encapsulated organisms (310,313). Since most reported patients come from surveys of rheumatologic disease, these frequencies of infection are subject to ascertainment bias and probably represent an underestimate of the true frequency of infection in a population.

Some probands, mainly children, have demonstrated pronounced susceptibility to bacterial infection as the only symptom. It should be noted that Sjoholm et al. (309) found that out of 17 C2-deficient probands diagnosed, 11 presented with severe bacterial infections, while only six had SLE or cutaneous vasculitis. Seven of the patients with bacterial infection were children. Homozygous C1r deficiency is combined with a partial defect of C1s (315). Some patients with hereditary angio-edema develop immune complex disease (316).

PARTIAL C4 DEFICIENCY: An increased prevalence of homozygous C4A deficiency has been found in SLE and some other immunological diseases (317). One study on C4B deficiency showed that it may be associated with bacterial meningitis caused by Haemophilus influenzae in small children (318). These findings seem to suggest that C4A could mostly be involved in immune complex handling, while C4B binds more efficiently to polysaccharides (319) such as are present on bacterial surfaces.

C3 DEFICIENCY SYNDROMES: As the focal point in the complement system, C3 participates in numerous effector mechanisms responsible for inflammation and host defence. Consequently it is not surprising that C3 deficiency leads to multiple, severe derangements including immune complex disease, impaired immune responses, impaired chemotaxis, reduced opsonophagocytosis and abnormal serum bactericidal activity. As with classical component deficiencies, these individuals are at increased risk for rheumatologic disease, 79% demonstrating a lupus-like syndrome or systemic vasculitis (310,313). These individuals have a striking disposition to severe and recurrent infections caused primarily by encapsulated organisms, which first manifests itself at an early age. Of all patients reported with this deficiency, 79% have had at least one episode of bacterial infection, particularly sinopulmonary infection, bacteremia or meningitis (310,313).

Most patients with C3 deficiency have pronounced susceptibility to bacterial infection (320). Cutaneous vasculitis, SLE-like symptoms and glomerulonephritis have also been observed. A dysfunctional form of C3 has recently been identified in a patient with SLE-like disease (321). Deficiency of the alternative pathway control proteins, factor I or factor H, leads to C3 hypercatabolism and secondary C3 deficiency (322,323). Factors H and I are primarily responsible for the down regulation of the fluid phase, alternative pathway C3 convertase. In the absence of either of these proteins the spontaneous formation of the C3 convertase goes unchecked, leading to C3 consumption and depletion. Although C3 consumption predisposes to immune complex syndromes, the small amount of residual C3 in the serum of persons lacking these factors seems to lessen their risk of these disorders.

ALTERNATIVE PATHWAY: Of components with functions strictly related to the alternative pathway, inherited defects have been described for properdin and for factor D. Through a noncovalent association, properdin stabilises the alternative pathway C3 convertase by a factor of 5-10 (324). Although this effect is relatively modest in vitro, individuals with properdin deficiency exhibit a marked propensity for infections with encapsulated organisms, especially N.meningitidis. Patients with properdin deficiency do not appear to have the same propensity for rheumatologic diseases as do individuals with either classical component or C3 deficiency states.

Factor D cleaves factor B to generate the alternative pathway C3 convertase, C3bBb. Complement factor D deficiency has been reported in a male patient with recurrent neisserial infections (325). In this study, an X-linked mode of inheritance could not be excluded. Incomplete factor D deficiency has previously been found in monozygous twins who were adult females with a record of repeated upper and lower respiratory tract infections from childhood (326).

TERMINAL COMPONENTS: As a major effector mechanism of the complement cascade, the membrane attack complex is responsible for direct complement-dependent serum bactericidal activity. Recent evidence suggests that it may also participate in tissue injury in a wide range of diseases (327,328). Perhaps for this reason, a small percentage (approx. 5%) of reported individuals with late complement component deficiency (LCCD) have evidence of immune complex or rheumatologic disease (310,313).

Recurrent systemic infection caused by N.meningitidis or N.gonorrhoeae is the only clinical manifestation clearly associated with homozygous deficiency of C5, C6, C7, or C8 (323). Compared with the frequency in the general U.S. population, LCCD increases the risk of meningococcal disease 8000-fold. As with properdin deficiency, the median age at the first neisserial infection is in the second decade of life; a significant departure from that in the general population and in persons with classical component deficiencies (314).

Although cleavage of C5 leads to the generation of potent chemotactic activity, as well as the formation of the membrane attack complex, the clinical features of C5 deficiency do not differ markedly from those of other terminal component deficiencies, suggesting that the absence of C5a does not contribute significantly to the clinical picture in these individuals. The association between C9 deficiency and neisserial infection is not as strong as in C5-C8 deficiency. Individuals with total C9 deficiency exhibit delayed, but present, serum bactericidal and haemolytic activities in vitro (329). The C8 molecule is composed of two subunits, the alpha-gamma subunit and the beta subunit, and hereditary deficiencies may involve either subunit. C8 alpha-gamma deficiency is found among patients of black or Hispanic origin, while all well documented cases of C8 beta deficiency have had a Caucasian background (323). C9 deficiency is remarkably common in the Japanese population, and various findings suggest the deficiency to be a susceptibility factor for development of meningococcal disease (330).

COMPLEMENT RECEPTORS AND MEMBRANE PROTEINS: Decreased expression of CR1 on erythrocytes and other cell types has been described in patients with SLE, and has stimulated extensive investigation of CR1 function in relation to pathogenic events in immune complex disease (331). Decreased CR1 has also been found in other disease conditions, including reactive arthritis caused by Yersinia enterocolitica (332). Patients with SLE also show decreased expression of CR2 on B lymphocytes. The weight of evidence suggests that these deficiencies are acquired through disease-related mechanisms (331).

In patients with paroxysmal nocturnal hemoglobinuria, a proportion of erythrocytes are susceptible to complement mediated lysis due to deficiency of DAF and other phospholipid anchored membrane proteins. The defect is caused by somatic mutation of blood-forming cells (333).

C2 DEFICIENCY AND NEUTROPENIA

Deficiency of the second component of complement (C2) is not a very rare genetic disease among Caucasians (334,335). C2 deficiency is often complicated by immunological disorders such as systemic lupus erythematosis (SLE), rheumatoid arthritis and vasculitis (334,335). C2 deficiency seen in Caucasians is linked to human lymphocyte antigen (HLA) B18, suggesting that the genetic change has occurred at some stage in the pedigree (336). Since HLA B18 is not seen in the Japanese population, it is considered that C2 deficiency does not exist in this race. Although one Japanese case of C2 deficiency complicated by vasculitis has been reported, the existence of C2 deficiency in Japan is generally ignored (337). A case report (338) has recently shown that a Japanese male has been found with homozygous C2 deficiency, heterozygous deficiency of the ninth component of complement (C9) and chronic idiopathic neutropenia. This kind of association between complement deficiency and chronic idiopathic neutropenia is believed to be the first case. This case report also claims to have confirmed the existence of C2 deficiency in the Japanese population.

The homozygous C2 deficiency in this case is associated with heterozygous C9 deficiency and chronic neutropenia. C9 is deficient in just 0.086% of Japanese, who are only incidentally associated with a variety of immune, viral, and infectious diseases (339), and the possibility that a genetic change of C2 has coincidentally occurred in a case with C9 deficiency is expected not to be high. In Japan, severe chronic idiopathic neutropenia is very rare (340). In addition, an association between chronic idiopathic neutropenia and C2 deficiency has not been reported as far as is known. Therefore, simultaneous appearance of C2 deficiency, C9 deficiency and chronic idiopathic neutropenia in an individual may not be incidental and suggests a possible relationship among them (338).

C2 deficiency in Caucasians is frequently associated with manifestations such as SLE, vasculitis, ankylosing spondylitis, rheumatoid arthritis, and type I diabetes (334,341,337). However, the mechanism by which these immunological disorders are brought about in C2 deficient individuals is not known. The gene of C2 is located in the MHC class III region (336). In this region, open reading frames exist whose protein products are supposed to be involved in the development and regulation of immune defence mechanisms, but are not yet characterised (342). Therefore, it can be speculated that a possible genetic change of MHC class III region linked with C2 deficiency may alter immune regulation mechanisms and thereby cause autoimmune diseases. This supposition may be supported by the data showing that IgA deficiency and common variable immunodeficiency are related with polymorphisms of MHC class III (342,343). Thus, it cannot be completely ruled out that C2 deficiency is linked with some kinds of genetic changes in the MHC class III region which may be related with chronic idiopathic neutropenia. However, the origin of chronic idiopathic neutropenia is not always clear, and whether genes controlling neutrophil leukocyte maturation exist in the MHC class III region is not known either. Further studies are necessary to clarify the relationship between C2 deficiency and chronic idiopathic neutropenia.

C3 DEFICIENCY

Genetically controlled deficiencies of certain complement components occur relatively frequently, for example, C2 and C9. Hereditary deficiency of the third component of complement, C3, however, is found very seldomly in humans (approximately 15 times up to 1990). C3 deficiency is associated with severe bacterial infections revealing the central role of C3 in complement activation via the classical or alternative pathway.

C3 and its split products have a central role not only in the effector phase of the immune response but also in its initiation. C3 fragments stimulate or inhibit certain functions of lymphoid and nonlymphoid cell populations and contribute as mediators or regulatory elements to the immune response despite occasional controversial reports. Analysis of the role of C3 in the afferent limb of the immune response would be facilitated by an animal model of C3 deficiency. In 1919 a complement deficiency of guinea pigs was reported which, in retrospect, may have been a C3 deficiency. The strain died out in the thirties. In the 1980s, an inherited C3 deficiency of dogs was reported; although the exact nature of this defect is unclear. Up to 1986 there were no other C3 defects known in animals, and for this reason, a genetically controlled C3 deficiency in inbred guinea pigs is used for experimental purposes (344).

The C3 defect is not linked to the MHC and, in addition, is not linked to a C3a receptor deficiency. Macrophages and hepatocytes of the C3 deficient animals have an unimpaired capacity for synthesis and secretion of C3 as measured by enzyme-linked immunosorbent assays. Possibly the fault resides in an enhanced susceptibility of their own C3 to proteolysis. However, C3 partially purified from the plasma of the C3 deficient animals, or secreted by hepatocytes, exhibits no obvious structural differences to purified normal C3. The C3 deficient serum showed a reduced bactericidal activity compared to normal or to C4 deficient serum. Nevertheless, the animals are apparently healthy without an indication for increased frequency of bacterial infections (344), a markedly different result compared with C3 deficient humans.

COMPLEMENT DEFICIENCY AND MENINGITIS

Meningococcal disease is a prominent manifestation in a significant fraction of reported cases in all clinical patterns of complement deficiency. In early component deficiencies, the meningococci shares this role with the pneumococcus and H. influenzae. In contrast, it is virtually the sole clinical manifestation in properdin deficiency and LCCD. Given this singular predisposition, studies of these deficiencies provide important clues to the role of complement in the pathogenesis of meningococcal disease.

The complement system has an important function in host defence against bacteria. C3 promotes the phagocytosis of bacteria, and the membrane attack complex (C5-C9) effects serum bactericidal activity (345). So, individuals with a deficiency of a component (C1, C2, or C4) of the classical pathway of complement activation, or with a deficiency of C3 have an increased risk of acquiring infections due to various bacterial pathogens, including Neisseria meningitidis (346). Patients who have acquired complement deficiencies due to autoimmune antibodies to complement components or a chronic disease leading to an increased turnover of complement components are also at increased risk for bacterial infections (347,348). Deficiencies in the alternative pathway (factor D, factor H, or properdin) and in the terminal pathway (C5, C6, C7, C8, or C9) of the complement system are especially associated with infections (meningitis and sepsis) due to N.meningitidis (345,346).

N. meningitidis is the most frequently isolated pathogen from patients with bacterial meningitis (349). Meningococcal serogroups are defined by their capsular polysaccharides. Serogroups B and C represented 96% of the meningococcal strains isolated from patients with meningitis in the Netherlands in 1990 (350). Meningitis caused by nongroupable meningococci or the Neisseria-related Moraxella species and Acinetobacter species is a relatively rare event (351,352). Nongroupable meningococci do not produce capsular polysaccharides or only produce low amounts. Individuals with a deficiency of a complement component (C5-C9) who have no capacity for a complement mediated serum bactericidal activity lack a basic defence against the so-called serum-sensitive bacteria such as nongroupable meningococci (352,353). Whether the Neisseria-related Moraxella species and Acinetobacter species, common inhabitants of the human skin and mucosal surfaces (349,352), are sensitive to the complement-mediated bactericidal activity is unknown.

A study by Fijen et al. (354) was carried out in 1993 to investigate the prevalence of complement deficiencies among patients who had meningitis caused by nongroupable meningococci, Moraxella species, and Acinetobacter species. Analysis of the complement systems among six patients with meningitis due to nongroupable meningococci showed the presence of a complement deficiency in two of those patients. Previously, it has been described that meningococcal disease caused by nongroupable meningococci had been found in only three complement-(C6 and C7) deficient patients (in the years ranging from 1986 and 1989). Nongroupable meningococci that are unencapsulated or that produce only very small amounts of polysaccharide are generally considered to be vulnerable to the bactericidal activity of the complement system (353,355). Various findings support the observation that such meningococcal strains are invasive and that the complement system is the most important system for warding off infections due to these uncommon meningococcal strains, including the nongroupable strains.

Moraxella species and Acinetobacter species are included in the family of Neisseriaceae (352). Moraxella species are frequently unencapsulated (356). It has been questioned whether Acinetobacter species should be classified within this family. Previous reports indicate that infections due to Moraxella and Acinetobacter species generally occur in patients with defective immunity (351,357), and these organisms are generally considered as opportunistic pathogens (352). None of the six patients studied had nosocomial infections. In this study, two patients with a ventriculoatrial shunt infection due to Moraxella species or Acinetobacter species, had haematuria and other signs suggestive of shunt nephritis at the time of the onset of meningitis. Immunologically, a low C1, C4, C2 and C3 level and a positive C1q binding test supported the diagnosis of shunt nephritis and represented an acquired complement deficiency (354). Whether the low complement levels increased the susceptibility for infection due to these uncommon strains or whether the levels were due to meningitis is unknown. However the occurrence of M.osloensis meningitis in an otherwise healthy C8beta-deficient patient favours the hypothesis that low complement levels, either acquired or congenital, increase the risk for meningitis due to such organisms. For this reason, the recommendation that such patients should be tested for complement deficiency seems to be justified.

Meningococcal disease in individuals with LCCD demonstrates several important differences from that in the general population (310). First, the frequency of meningococcal disease in the U.S. population is 0.0072%, whereas it is 9-58% in C9 and C5 to C8-deficient individuals, respectively, 1000 to 10,000-fold increased risk of infection.

Second, individuals with LCCD are infected with uncommon meningococcal serogroups more often than normal persons (310,313). In conjunction with the bactericidal defect, these findings translate into an additional degree of difficulty for the LCCD person in the elimination of these strains compared with those that typically cause disease in the general population. Incidentally, this finding may be due, in part, to the propensity of these uncommon serogroups to cause disease in older individuals (358).

Third, individuals with LCCD deficiency experience their initial meningococcal infection at an older median age than do normal persons (17 years and 3 years respectively) (310,313). Thus, the majority of deficient persons pass through the period of highest risk of infection for normal individuals only to develop infection in the second decade. This paradox is only partially explained by the fact that deficient individuals are susceptible for life while normal

individuals are generally at risk only early in life. It seems likely that unidentified factors contribute to the susceptibility of deficient individuals later in life.

Fourth, mortality associated with meningococcal disease ranges between 10 and 19% in the general U.S. population but is only 1.5 to 2.4% in LCCD persons (310). Meningococcal disease is a paradigm for gram-negative sepsis. Initially, meningococcal endotoxin (lipo-oligosaccharide or LOS) stimulates numerous host responses including complement activation, cytokine release and infiltration of inflammatory cells (359,360). As the disease progresses, the prominent features become those of endothelial damage and consumptive coagulopathy. Several, as yet unsubstantiated, factors may contribute to the lower mortality in deficient patients: (1) lower organism inoculum required to cause disease; (2) lower plasma endotoxin concentrations; (3) milder disease, and (4) less tissue injury (314).

Fifth, recurrent meningococcal disease is rare in normal and properdin-deficient individuals (310,313). This finding suggests that prior infection provides specific and cross-reactive antibody to Neisseria in these persons. These antibodies can activate the classical pathway to prevent recurrence through either C3b-directed opsonophagocytosis or serum bactericidal activity. In contrast, recurrent disease is common (44%) in persons with C5-C8 deficiency suggesting that immunity does not follow initial infection in these individuals despite intact C3b-mediated phagocytosis (310). This analysis indicates that each episode of infection is an independent event; that is prior infection does not alter the risk of infection in these persons.

Although it may not prevent disease, acquired immunity might ameliorate the severity of subsequent infection. If so, this might help explain the reduced mortality in LCCD individuals. However, Platonov and Beloborodov (361) observed that prior infection had no effect on the severity of subsequent infections in LCCD individuals. Moreover, the few deaths that have occurred in LCCD individuals with meningococcal diseases have not been confined to the initial infection. Thus, the occurrence and severity of meningococcal disease in LCCD patients seems to be completely independent of prior disease.

PROPERDIN DEFICIENCY

The high prevalence of properdin deficiency and other defects of the complement system in patients with meningococcal disease caused by uncommon serogroups of Neisseria meningitidis, mainly serogroups W-135 and Y, has recently been recognised in Danish (362) and Dutch (363) populations. Properdin deficiency has also been identified in a healthy proband (364), and in probands with discoid lupus erythematosus (365) or recurrent pneumococcal infections (366).

Of the 53 persons with verified properdin deficiency or dysfunction, 45% had proven meningococcal disease, and 6% died from the infection (309). The number of lethal cases, though, is clearly underestimated since very few of these patients have been investigated for complement function. With the addition of 16 family members who died from fulminant infection and may be assumed to have had inherited defects of properdin yields a total number of 69 cases and a fatality rate of 28%. In 21 patients with proven meningococcal disease, the median age at the time of infection was 14 years. Infection occurred before the age of 18 in 81% of the patients. These figures emphasise that genetic defects of properdin are major determinants of susceptibility to meningococcal disease. Recurrent meningococcal infection appears to be rare in patients with defects of properdin, and has so far been reported only once (367).

The significance of other clinical findings in patients with properdin deficiency or dysfunction is less clear. As already mentioned, properdin deficiency has been reported in discoid lupus erythematosus (365), and combined with C2 deficiency in a boy with recurrent pneumococcal bacteremia (366). A 61-year old man with properdin dysfunction (368), who survived meningococcal meningitis, later developed septicaemia caused by Escherichia coli. In the same family, meningitis of unknown cause (one patient), bronchopneumonia (one patient), recurrent otitis media (one patient), epididymitis (two patients), and rapidly progressive systemic schlerosis (one patient) were also observed in affected males (309).

Family studies of inherited properdin defects strongly suggest an X-linked mode of transmission. The existence of an X-linked properdin dysfunction state implies that a structural gene for properdin is probably involved. RFLP linkage analyses and in situ hybridisation experiments have been carried out to suggest the position of such a structural gene (364,368). Analysis of DNA from a few properdin deficient persons has not revealed gross deletions or alterations of the properdin structural gene (369).

Various family data studied have been consistent with an autosomal recessive mode of inheritance without being convincing.

EFFECT OF VACCINATION ON PATIENTS WITH COMPLEMENT DEFICIENCY

Subjects with late complement component deficiency (LCCD) tend to have recurrent neisserial infections, probably due to a failure of membrane attack complex formation (370). Currently, there is no universally accepted strategy to prevent the recurrence of neisserial infections in complement-deficient persons. In some patients with LCCD, a single vaccination with tetravalent meningococcal vaccine causes a rise in antimeningococcal antibodies against all 4 strains of Neisseria meningitidis (A, C, Y, W135) (371). However, it is not clear to what extent these antibodies are protective against meningococcal infections (370). Ross et al. (372) showed that after two vaccinations with a divalent A-C meningococcal vaccine, the serum of a C8beta-deficient patient (who had four previous episodes of meningococcal infections including one with the Y strain) showed a marked opsonisation of A and C meningococci in a neutrophil killing assay.

The association of LCCD with neisserial infections is well documented. Of persons with LCCD, 57% will contract meningococcemia or meningitis at least once and approx. 40% will have more than one infection. This contrasts with the very low frequency (0.0072%) and recurrence rate (0.34%) of meningococcal infection in the general population (370,373). Since infection with meningococci from serogroups, other than group B, are relatively more common in subjects with LCCD (370,363), vaccination with the tetravalent meningococcal vaccine may be a potential prophylactic option.

Experiments carried out by Schlesinger et al. (374) have shown that the phagocytic killing of strains A, C, Y, and W markedly improved after vaccination in subjects with LCCD, as well as in normal controls.

Analysis of various data (372,375) revealed that the phagocytic activity in the presence of serum was much more efficient than was phagocytic activity in the presence of heat inactivated serum, suggesting the possible contribution of C3b to the killing process. The mechanism of the increase in phagocytic killing after vaccination is not entirely clear, although activation of the complement system via binding of meningococci with the specific antibody and formation of opsonins, especially C3b, may be a reasonable explanation. Biselli et al. (375) have suggested that antibodies to meningococcal capsular antigens deposit C3 on an exposed location of the organism's polysaccharide capsule. Deposited C3 and antibodies can promote the ingestion of the organism through interaction with specific receptors on the surface of the phagocytic cells.

The high rate of phagocytic killing after vaccination seen in three C8beta-deficient patients studied, despite a mild increase in antimeningococcal antibodies, may indicate that phagocytic killing is a more sensitive indicator than is the increase in antimeningococcal antibodies. Although it is not known which assay is a better predictor of protection against meningococcal infection, the phagocytic killing assay may be more relevant because it demonstrates the protective function of neutrophils, whereas the biological relevance of the antibodies may depend on various conditions, including the antigen epitopes toward which they are directed or their affinity (376,377). In addition, there is no data for determining which antibody level is of protective value.

Only the long-term follow-up of vaccinated persons with LCCD, and repeated measurements of antibody titres and phagocytic killing will provide a better understanding of the effect of vaccination against meningococcal infection and lead to a more efficient strategy for prevention of these infections. At present it seems reasonable to recommend that tetravalent meningococcal vaccine be given to all persons with LCCD.

DETECTION OF COMPLEMENT DEFICIENCY BY ELISA

Diagnostic investigations of the complement system are usually restricted to the measurement of a few complement proteins, such as C3 and C4, which implies that rare but clinically important complement deficiency states are often overlooked. Simple assays for assessment of complement function are required in diagnostic work (378). Haemolytic assay systems are commonly used for this purpose, and simplified procedures suitable for relatively large scale investigations of patients have been described (379,380). On the other hand, complement analysis tends to be restricted to a few specialised laboratories, and the development of alternative methods based on more widely employed technologies might be considered.

To be useful in diagnostic work, screening procedures for the detection of complement deficiencies, should be simple and rapid, and should clearly distinguish between defects within the functional units of the complement system, i.e. the classical activation pathway (C1, C2, C4), the alternative activation pathway (C3, factor B, factor D, properdin), and the terminal sequence (C5-C9). Furthermore the assays should not be influenced by rheumatoid factors, and the reagents should be commercially available. The ELISA procedure described by Fredrikson et al. (381) appears to meet these requirements.

The sera investigated by Fredrikson et al. (381) were incubated in microtitre plates with solid phase complement activators. Human polyclonal IgG or monoclonal IgM were used for classical activation pathway assays and Salmonella typhosa LPS for alternative activation pathway assays. This particular analysis focused on deposition of C9 and properdin as detected with enzyme-conjugated antibodies. In an attempt to avoid spurious results due to rheumatoid factors in patient sera, monoclonal mouse and chicken antibodies were unsuccessfully tested as indicator reagents in the assay with solid-phase IgG. However, the use of solid-phase IgM as an activator completely circumvented the influence of rheumatoid factors. With solid-phase IgG or IgM, properdin deposition occurred in the absence of factor D. A combination of assays is suggested for diagnostic purposes: IgM-coated plates with detection of bound C9 and properdin for the classical pathway; and LPS-coated plates with detection for bound properdin for the alternative pathway. Therefore, the procedure distinguishes between defects of the classical activation pathway, the alternative activation pathway, and the terminal complement components. This analytical approach may be useful for the detection of inherited complement deficiency and assessment of complement function in acquired complement deficiency states.

In diagnostic work, a combination of two screening assays, i. e. the IgM-ELISA with detection of C9 deposition and the LPS-ELISA with detection of properdin deposition, would probably reveal all known types of complement deficiency and provide a basis for further identification .

Several forms of acquired complement deficiency could produce difficulties of interpretation unless the analysis is combined with ordinary determinations of proteins, such as C3 and C4. An interesting question relates to the possible value of the ELISA procedure for the assessment of complement function in diseases, such as systemic lupus erythematosus (282).

While haemolytic assays will certainly remain the standard for documentation of complement function (283), the present technology shows promise for diagnostic purposes. Different kinds of erythrocytes are not readily available in all laboratories, whereas the ELISA procedure described, uses stable and commercially available reagents. Therefore, the ELISA procedure described appears to be a simple, rapid, and reliable method for the assessment of complement function, particularly the detection of complement deficiency states.

   


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