April 14, 2004 Access past issues

Reviewed by: Benjamin J. Silk Rollins School of Public Health Department of Epidemiology Emory University

The Health Outcome

Neisseria meningitidis is a Gram-negative diplococcus with multiple serogroups (e.g., A, B, C, Y, W-135). Humans are the only reservoir for the bacteria and during non-epidemics, 5-10% of adults carry the pathogen in the nasopharynx without symptoms.(1) Transmission occurs by direct contact with secretions or inhalation of large droplet nuclei. (2) While most upper respiratory colonization does not result in clinical disease, invasive meningococcal disease (IMD) is rapid and often causes severe disease, physical and neurologic sequelae, and death (case fatality rate = 10-20%). (3) Though IMD typically causes septicemia, meningitis, or both, there are many other manifestations of the disease. Meningococcal disease is both sporadic and epidemic. Worldwide, approximately 500,000 cases of IMD occur every year, (4) many of which are associated with large outbreaks in the 'meningitis belt' of sub-Saharan Africa . Children are disproportionately affected by endemic disease, particularly during infancy when immunity from maternal antibodies begins to wane. Among adults, sporadic disease is often associated with complement deficiencies, and persons with chronic medical conditions and the elderly also have heightened susceptibility to invasive meningococcal disease.

Cytokines (small, soluble protein hormones that modulate cell-mediated immunity) are key regulators of the inflammation response to N. meningitidis endotoxin, or lipooligosaccharide (LOS). Release of LOS into serum via blebbing stimulates an array of inflammatory cytokines, chemokines, and other effector molecules.(5) Thus, the overall pathophysiology of invasive meningococcal disease (IMD) directly relates to the cytokine network and its role in inflammation, coagulation, and other molecular changes.

The study examines the associations between seven cytokine polymorphisms and susceptibility, severity, and outcome of IMD. These polymorphisms are:

1. Interleukin 6 (IL-6) G-174C (alleles IL-6 - 174 G/G, IL-6 - 174 G/C, and IL-6 - 174 C/C), which is associated with interleukin 6, a multifunctional cytokine with proinflammation and procoagulation effects;
2. Interleukin 10 (IL-10) - 1082 (alleles IL-10- 1082 G/G, IL-10 - 1082 G/A, and IL-10 - 1082 A/A), associated with interleukin 10, an anti-inflammatory cytokine that downregulates proinflammatory cytokines and upregulates IL-1Ra (a receptor antagonist of IL-1ß);
3. Interleukin 1 receptor antagonist (IL-1RN) VNTR (alleles IL-1RN 1/1, IL-1RN 1/2, IL-1RN 2/2, IL-1RN 1/3, and IL-1RN 2/3), which is associated with the interleukin 1 Ra, which binds to IL1 receptors to inhibit IL1- and IL1- ß;
4. Interleukin 1ß (IL-1B) +3953 (alleles IL-1B +3953 C/C, IL-1B +3953 C/T, and IL-1B +3953 T/T), associated with variations in interleukin 1B levels, which act synergistically with and similar to TNF, causing severe disease via polymorphonulear activity, upregulation of endothelial adhesion molecules, procoagulation, prostaglandin production, cytokine activity, and other proinflammation properties;
5. Interleukin 10 (IL-10) -592 (alleles IL-10 -592 C/C, IL-10 -592 C/A, and IL-10 -592 A/A), which is associated with interleukin 10, an anti-inflammatory cytokine that downregulates proinflammatory cytokines and upregulates IL-1Ra (a receptor antagonist of IL-1ß).
6. Tumor necrosis factor (TNF) -308 (alleles TNF -308 G/G, TNF -308 G/A, and TNF -308 A/A), associated with variations in TNF levels, which act synergistically with and similar to IL-1B , causing severe disease via polymorphonulear activity, upregulation of endothelial adhesion molecules, procoagulation, prostaglandin production, cytokine activity, and other proinflammation properties; and
7. Lymphotoxin (LTA) +252 (alleles LTA +252 G/G, LTA +252 G/A, and LTA +252 A/A), which is associated with lymphotoxin alpha (LTA), a proinflammatory cytokine.

The Finding

The study has two design components. First, by comparing genotype and allele frequencies in 183 cases and 389 controls, gene-disease susceptibility is investigated for the seven cytokine polymorphisms. Cases of invasive meningococcal disease (IMD) included patients who met four criteria: blood, cerebrospinal fluid, or other sterile sites that were positive for the meningococcal citrA gene by polymerase chain reaction (PCR) analysis; presence in the Irish Meningococcal and Meningitis Reference Laboratory (IMMRL) database; availability of specimens for subsequent genotyping; and either care-seeking at one of two tertiary referral pediatric hospitals or known death at another hospital. Controls samples were obtained from blood donors via the Northern Ireland Blood Transfusion Service (15%) and the Irish Blood Transfusion Service (85%).

Second, a case-series design split cases into two sets of groups; to investigate disease severity for the seven cytokine polymorphisms, patients with mild invasive meningococcal disease (IMD) and no sequelae were compared to IMD patients with severe disease (poor prognostic indicators, sequelae, and death). To investigate disease outcome for the same polymorphisms, IMD survivors were compared to nonsurvivors.

Susceptibility to invasive meningococcal disease (IMD): In comparing cases to controls, a statistically significant difference was found in the prevalence of the IL-1RN 2/2 genotype, which was identified in 14% of cases and 8% of controls (p=0.033).

Severity of IMD: Cases with severe IMD more often carried the IL-6 - 174 G/G allele (41%) compared to cases with mild disease (26%), a difference with statistical significance (p=0.037). In addition, the IL-10 - 1082 A/A allele was more common in severe IMD cases (29%) than mild cases (13%) (p=0.0078). Severe cases were 2.7 times more likely to carry this variant than other variants of the same gene (odds ratio (OR) = 2.7, 95% confidence interval (CI) =2.3 - 3.6).

Outcome of IMD (mortality): The IL-6 - 174 G/G variant, which was associated with severe disease, was also associated with mortality (p=0.023). IMD patients who did not survive were 2.6 times more to carry this genotype (OR=2.64, CI=1.12-6.22). Similarly, in comparing mild IMD to cases of IMD where patients did not survive, there was a strong association with the homozygous G/G allele (p=0.012). The IL-1RN 1/1 and IL-1RN 1/2 genotypes also had significant associations with mortality. Fatal cases of IMD more often carried the 1/1 variant (60% vs. 43% in survivors) and survivors more often had the 1/2 variant (41% versus 20% in nonsurvivors) (p=0.043).

Multipolymorphism analysis: Among persons carrying all three deleterious genotypes ( IL-6 - 174 G/G, IL-10 - 1082 A/A, and IL-1RN 1/1), differences with respect to severity of disease or survival were not found in using logistic regression, though this combination was relatively uncommon. The two-way combinations of the same genotypes, however, were significant. The combination of IL-6 - 174 G/G and IL-10 - 1082 A/A, while also uncommon, was associated with severity of disease (p=0.016) and the combination of IL-6 - 174 G/G and IL-1RN 1/1 was strongly associated with fatality (p=0.0008)

The authors conclude that key findings in the study were, in general, not highly statistically significant - thereby necessitating further investigation. They note that severe meningococcemia is certainly multifactorial, and therefore their study should be viewed as a preliminary, exploratory study.

The data suggest roles for the IL-6 - 174 G/G and IL-10 - 1082 A/A genotypes with disease severity. In addition, the IL-1RN 1/2 and 2/2 variants may be important to disease susceptibility as well as mortality. When in combination, the IL-6 - 174 G/G and IL-10 - 1082 A/A were also related to disease severity, and the IL-6 - 174 G/G and IL-1RN 1/2 were related to mortality. Other polymorphisms related to cytokines, however, did not appear to be related to susceptibility, severity, or outcome of IMD. These included the interleukin 1 ß (IL-1B) +3953, interleukin 10 (IL-10) -592, tumor necrosis factor (TNF) -308, and lymphotoxin (LTA) +252 polymorphisms .

Public Health Implications

Given that invasive meningococcal disease is rare, genetic susceptibility testing would be unlikely to have a meaningful public health impact. The contribution of the study lies instead in advancing toward clarifying the influence of these seven genetically variant cytokine mediators in invasive meningococcal disease; elucidating the cytokine network's complex inflammatory response to meningococcal endotoxin will likely be a challenge for years to come.

While cytokine polymorphism research may prove valuable in predicting susceptibility, severity, and outcome of meningococcemia, it is unlikely to be the only useful information. Research related to host characteristics beyond the cytokine network, as well as bacterial characteristics such as meningococcal virulence factors and key genetically-defined, invasive clonal groups (e.g., ET-37 and ET-5 complexes, and the A4 cluster), (6) offer equal promise for advancing our understanding of clinical variability in meningococcemia

References

1. Greenfield S, Sheehe PR, Feldman HA. Meningococcal carriage in a population of 'normal' families. J Infect Dis 1971; 123:67-73.
2. American Public Health Association. In: Chin J, ed. Control of Communicable Diseases Manual, 17 th edition. Washington , D.C. , 2000: 341-342.
3. Tzeng YL and Stephens DS. Epidemiology and pathogenesis of Neisseria meningitidis.
   Microbes and Infection 2000: 2: 687-700.
4. Tikhomirov E, Santamaria M, Esteves K. Meningococcal disease: public health burden and control, Wld Hlth Statist Quart 1997: 50.
5. Hackett SJ, Thomson APJ, Hart CA. Cytokines, chemokines and other effector molecules involved in meningococcal disease. J Med Microbiol 2001;50:847-859.
6. Caugant DA. Population genetics and molecular epidemiology of Neisseria meningitidis.
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