Variants in the promoter region of IKBL/NFKBIL1 gene may mark susceptibility to the development of chronic Chagas' cardiomyopathy among Trypanosoma cruzi-infected individuals.

Chagas' disease, caused by Trypanosoma cruzi, is an inflammatory disorder leading to chronic Chagas cardiomyopathy (CCC). Only one third of T. cruzi-infected individuals progress to CCC while the others are considered asymptomatic (ASY). The human inhibitory kappaB-like gene (IKBL/NFKBIL1), homologous to the IkappaB family of proteins that regulate the NFkappaB family of transcription factors, is suggested as a putative inhibitor of NFkappaB. We investigated two functional polymorphisms, -62A/T and -262A/G, in the promoter of IKBL by PCR-RFLP analysis in 169 patients with CCC and 76 ASY. Genotype distributions for both -62A/T and -262A/G differed between the CCC and ASY (chi2=7.3; P=0.025 and chi2=6.8; P=0.03, respectively). Subjects, homozygous for the -62A allele, had three-fold risk of developing CCC compared with those carrying the TT genotype (P=0.0095; Odds Ratio [OR]=2.9; [95% CI 1.2-7.3]). Similar trend was observed for the -262A homozygotes (P=0.005; OR=2.7 [95% CI 1.3-6.0]. The haplotype -262A -62A was prevalent in patients with CCC (40% versus 24%; OR 2.1 [95% CI 1.4-3.3]; Pc=0.0014). The IKBL locus itself or another critical gene in this region may confer susceptibility to the development of CCC.

Association of polymorphisms within the promoter region of the tumor necrosis factor-alpha with clinical outcomes of rheumatic fever.

Rheumatic fever (RF)/rheumatic heart disease (RHD) is an inflammatory disease with a complex etiology in which Group A streptococci within a genetically susceptible host untreated for strep-throat may deviate the innate and adaptive arms of the immune system towards recognition of autoantigens. The TNFA gene has been associated with a number of autoimmune diseases, including RF. We investigated whether the G-308A and G-238A polymorphisms of the TNFA gene are associated with clinical outcomes of RF in a cohort of 318 patients and 281 healthy controls (HC). Both polymorphisms showed borderline associations with RF (TNFA -308G/A, OR=1.4 [1-2.2], P=0.026; TNFA -238G/A, OR=1.9 [1-3.3], P=0.015). The presence of either one of the minor alleles (-308A and -238A) was more common among patients with RF/RHD than controls (P=0.0006). Stratification of patients according to clinical phenotype also showed significant associations between presence of either one of the minor alleles and RHD (Pc=0.0006) when compared with controls. This association was stronger with the development of aortic valve lesions. In contrast, there was no association between genotype and Sydenham's chorea or RF patients with mild carditis. In conclusion, we show that the TNFA is a susceptibility locus for RF. The ability to predict which RF patients will develop valve lesion may have therapeutic, economic and social implications.

T cell response in rheumatic fever: crossreactivity between streptococcal M protein peptides and heart tissue proteins.

Molecular mimicry between streptococcal and human proteins has been proposed as the triggering factor leading to autoimmunity in rheumatic fever (RF) and rheumatic heart disease (RHD). In this review we focus on the studies on genetic susceptibility markers involved in the development of RF/RHD and molecular mimicry mediated by T cell responses of RHD patients against streptococcal antigens and human tissue proteins. We identified several M protein epitopes recognized by peripheral T cells of RF/RHD patients and by heart tissue infiltrating T cell clones of severe RHD patients. The regions of the M protein preferentially recognized by human T cells were also recognized by murine T cells. By analyzing the T cell receptor (TCR) we observed that some Vbeta families detected on the periphery were oligoclonal expanded in the heart lesions. These results allowed us to confirm the major role of T cells in the development of RHD lesions.

TNF blockade aggravates experimental chronic Chagas disease cardiomyopathy.

Chronic Chagas disease cardiomyopathy (CCC), caused by Trypanosoma cruzi, is an inflammatory dilated cardiomyopathy associated with increased circulating levels of TNF-alpha. We investigate whether TNF blockade with Etanercept during the chronic phase of T. cruzi infection could attenuate experimental CCC development. The effect of Etanercept was evaluated after 11 months of T. cruzi infection on survival, parasitism, left ventricular function, intensity of myocarditis, fibrosis, and left ventricular mRNA expression of cytokines and TNF-alpha-induced genes. Left ventricular function was significantly reduced in treated animals as compared to infected untreated animals. Blood and cardiac parasitism as well as survival rate were not altered with Etanercept treatment. Inflammatory infiltrates were located predominantly in the subendocardic region in treated animals, whereas in untreated animals inflammation was scattered throughout the myocardium. Left ventricular mRNA IL-10 expression was significantly higher, and iNOS, significantly lower in treated than in untreated animals. mRNA expression of TNF-alpha, IFN-gamma, TGF-beta, A20 and ANP was similar in both groups. Our results suggest that TNF-alpha/LT-alpha blockade with Etanercept enhances left ventricular dysfunction in T. cruzi-induced chronic cardiomyopathy and the absence of TNF signaling may be deleterious to the failing heart in Chagas disease cardiomyopathy.

Lack of association of tumor necrosis factor-alpha polymorphisms with Chagas disease in Brazilian patients.

One third of Trypanosoma cruzi-infected individuals develop chronic Chagas disease cardiomyopathy (CCC) while the majority remains asymptomatic (ASY). About 30% of CCC patients develop heart failure due to end-stage inflammatory dilated cardiomyopathy. Increased production of tumor necrosis factor (TNF)-alpha has been described in all clinical forms of Chagas disease, and the highest levels are detected in CCC patients with severe ventricular dysfunction. Genetic susceptibility may play a role in the clinical outcome of Chagas disease. We investigated TNF as a candidate gene for susceptibility to development and/or progression of CCC. We analyzed the TNFa microsatellite and the -308 TNF promoter polymorphisms, in 166 CCC compared to 80 ASY geographically and age-matched patients in an association study. To analyze the association of TNF polymorphisms with progression of the cardiomyopathy, CCC patients were also grouped in three categories according to degree of left ventricular (LV) dysfunction into severe (n=57), mild to moderate (n=21) and absent (n=88). Our results show no significant differences either between CCC and ASY patients, or among CCC patients according to severity of cardiomyopathy with respect to TNFa or -308 TNF promoter polymorphisms. These results indicate that TNF polymorphisms are associated neither to CCC development nor to progression to more severe forms of cardiomyopathy in Brazilian Chagas disease patients.

Rheumatic heart disease: 15 years of clinical and immunological follow-up.

Rheumatic fever (RF) is a sequel of group A streptococcal throat infection and occurs in untreated susceptible children. Rheumatic heart disease (RHD), the major sequel of RF, occurs in 30%-45% of RF patients. RF is still considered endemic in some regions of Brazil and is responsible for approximately 90% of early childhood valvular surgery in the country. In this study, we present a 15-year clinical follow-up of 25 children who underwent surgical valvular repair. Histopathological and immunological features of heart tissue lesions of RHD patients were also evaluated. The patients presented severe forms of RHD with congestive symptoms at a very young age. Many of them had surgery at the acute phase of RF. Histological analysis showed the presence of dense valvular inflammatory infiltrates and Aschoff nodules in the myocardium of 21% of acute RHD patients. Infiltrating T-cells were mainly CD4+ in heart tissue biopsies of patients with rheumatic activity. In addition, CD4+ and CD8+ infiltrating T-cell clones recognized streptococcal M peptides and cardiac tissue proteins. These findings may open the possibilities of new ways of immunotherapy. In addition, we demonstrated that the surgical procedure during acute phase of the disease improved the quality of life of young RHD patients.

The monocyte chemoattractant protein-1 gene polymorphism is associated with cardiomyopathy in human chagas disease.

BACKGROUND: Only a subset of individuals infected with Trypanosoma cruzi develop chronic Chagas cardiomyopathy (CCC). Familial aggregation of CCC in areas of endemicity indicates that susceptibility may be genetic, which may be a plausible explanation for why only one-third of T. cruzi-infected individuals develop CCC. The monocyte chemoattractant protein-1 (CCL2/MCP-1) has been shown to enhance the uptake of T. cruzi in murine macrophages and to up-regulate the inducible nitric oxide synthase/nitric oxide system, with a consequent increased production of nitric oxide that controls the replication of the parasite. METHODS: We assessed CCL2 variants at position -2518A/G, which are known to influence transcriptional activity, by polymerase chain reaction and restriction fragment-length polymorphism in 245 individuals, all of whom were infected with T. cruzi. One hundred sixty-nine patients had CCC, and 76 were asymptomatic. RESULTS: Genotype distributions differed between the CCC and asymptomatic groups (chi2 = 9.4; P = .009), with an excess of genotypes with the A allele (AA + AG) in the CCC group. Among patients with CCC, 5% were homozygous for the G allele, compared with 16% of the asymptomatic subjects (odds ratio [OR], 4.1; 95% confidence interval [CI], 1.7-11; P = .001). A similar trend was observed when individuals heterozygous for the G allele were compared with individuals homozygous for the G allele between the CCC and asymptomatic groups (OR, 2.7; 95% CI, 0.97-7.2; P = .026). The A allele seems to confer susceptibility to CCC (OR, 1.9; 95% CI, 1.3-2.9; P = .001). CONCLUSIONS: The CCL2 variant correlated with a low transcriptional level behaves as a genetic modifier of clinical outcome for T. cruzi infection, and subjects with the CCL2 -2518AA genotype have a 4-fold greater risk of developing CCC than do those without this genotype.

TNF gene polymorphisms are associated with reduced survival in severe Chagas' disease cardiomyopathy patients.

Chronic Chagas' disease cardiomyopathy (CCC) is the most important clinical outcome of infection by the parasite Trypanosoma cruzi, affecting 18 million individuals in Latin America. One-third of CCC patients develop heart failure due to end-stage dilated cardiomyopathy, and their survival is reduced by 50% compared to patients with other cardiomyopathies. Genetic susceptibility may play a role in the differential survival of severe CCC patients. Given the role of TNF-alpha in the progression of heart failure, and the increased TNF-alpha plasma and heart tissue levels observed in these patients, we chose TNF as a candidate gene for increased mortality in severe CCC patients. We typed the TNFa microsatellite and the -308 TNF promoter polymorphism and then analyzed the survival curves of 42 patients with severe ventricular dysfunction (left ventricular ejection fraction

Immunogenicity and safety of a tetravalent E. coli O-antigen bioconjugate vaccine in animal models.

BACKGROUND: Extra-intestinal pathogenic Escherichia coli (ExPEC) are major human pathogens; however, no protective vaccine is currently available. We assessed in animal models the immunogenicity and safety of a 4-valent E. coli conjugate vaccine (ExPEC-4V, serotypes O1, O2, O6 and O25 conjugated to Exotoxin A from Pseudomonas aeruginosa (EPA)) produced using a novel in vivo bioconjugation method. METHODS: Three doses of ExPEC-4V (with or without aluminum hydroxide) were administered to rabbits (2µg or 20µg per O-antigen, subcutaneously), mice (0.2µg or 2µg per O-antigen, subcutaneously) and rats (0.4µg or 4µg per O-antigen, intramuscularly). Antibody persistence and boostability were evaluated in rats using O6-EPA monovalent conjugate (0.4µg O-antigen/dose, intramuscularly). Toxicity was assessed in rats (16µg total polysaccharide, intramuscularly). Serum IgG and IgM antibodies were measured by ELISA. RESULTS: Robust antigen-specific IgG responses were observed in all animal models, with increased responses in rabbits when administered with adjuvant. O antigen-specific antibody responses persisted up to 168days post-priming. Booster immunization induced a rapid recall response. Toxicity of ExPEC-4V when administered to rats was considered to be at the no observed adverse effect level. CONCLUSIONS: ExPEC-4V conjugate vaccine showed good immunogenicity and tolerability in animal models supporting progression to clinical evaluation.

T cell epitope characterization in tandemly repetitive Trypanosoma cruzi B13 protein.

Proteins containing tandemly repetitive sequences are present in several immunodominant protein antigens in pathogenic protozoan parasites. The tandemly repetitive Trypanosoma cruzi B13 protein is recognized by IgG antibodies from 98% of Chagas' disease patients. Little is known about the molecular mechanisms that lead to the immunodominance of the repeated sequences, and there is limited information on T cell epitopes in such repetitive antigens. We finely characterized the T cell recognition of the tandemly repetitive, degenerate B13 protein by T cell lines, clones and PBMC from Chagas' disease cardiomyopathy (CCC), asymptomatic T. cruzi infected (ASY) and non-infected individuals (N). PBMC proliferative responses to recombinant B13 protein were restricted to individuals bearing HLA-DQA1*0501(DQ7), -DR1, and -DR2; B13 peptides bound to the same HLA molecules in binding assays. The HLA-DQ7-restricted minimal T cell epitope [FGQAAAG(D/E)KP] was identified with an overlapping combinatorial peptide library including all B13 sequence variants in T. cruzi Y strain B13 protein; the underlined small residues GQA were the major HLA contact residues. Among natural B13 15-mer variant peptides, molecular modeling showed that several variant positions were solvent (TCR)-exposed, and substitutions at exposed positions abolished recognition. While natural B13 variant peptide S15.9 seems to be the immunodominant epitope for Chagas' disease patients, S15.4 was preferentially recognized by CCC rather than ASY patients, which may be pathogenically relevant. This is the first thorough characterization of T cell epitopes of a tandemly repetitive protozoan antigen and may suggest a role for T cell help in the immunodominance of protozoan repetitive antigens.

Importance of (antibody-dependent) complement-mediated serum killing in protection against Bordetella pertussis.

Pertussis is a highly contagious respiratory disease that is caused by Bordetella pertussis. Despite being vaccine preventable, pertussis rates have been rising steadily over the last decades, even in areas with high vaccine uptake. Recently, experiments with infant baboons indicated that although vaccination with acellular pertussis vaccines prevented disease, no apparent effect was observed on infection and transmission. One explanation may be that current acellular pertussis vaccines do not induce high levels of opsonophagocytic and/or bactericidal activity, implying that engineering of vaccines that promote bacterial killing may improve efficacy. Here, we discuss the importance of complement-mediated killing in vaccine-induced protection against B. pertussis. We first examine how B. pertussis may have evolved different complement evasion strategies. Second, we explore the benefits of opsonophagocytic and/or bactericidal killing in vaccine-induced protection and discuss whether or not inclusion of new opsonophagocytic or bactericidal target antigens in pertussis vaccines may benefit efficacy.

CXCL5-secreting pulmonary epithelial cells drive destructive neutrophilic inflammation in tuberculosis.

Successful host defense against numerous pulmonary infections depends on bacterial clearance by polymorphonuclear leukocytes (PMNs); however, excessive PMN accumulation can result in life-threatening lung injury. Local expression of CXC chemokines is critical for PMN recruitment. The impact of chemokine-dependent PMN recruitment during pulmonary Mycobacterium tuberculosis infection is not fully understood. Here, we analyzed expression of genes encoding CXC chemokines in M. tuberculosis-infected murine lung tissue and found that M. tuberculosis infection promotes upregulation of Cxcr2 and its ligand Cxcl5. To determine the contribution of CXCL5 in pulmonary PMN recruitment, we generated Cxcl5(-/-) mice and analyzed their immune response against M. tuberculosis. Both Cxcr2(-/-) mice and Cxcl5(-/-) mice, which are deficient for only one of numerous CXCR2 ligands, exhibited enhanced survival compared with that of WT mice following high-dose M. tuberculosis infection. The resistance of Cxcl5(-/-) mice to M. tuberculosis infection was not due to heightened M. tuberculosis clearance but was the result of impaired PMN recruitment, which reduced pulmonary inflammation. Lung epithelial cells were the main source of CXCL5 upon M. tuberculosis infection, and secretion of CXCL5 was reduced by blocking TLR2 signaling. Together, our data indicate that TLR2-induced epithelial-derived CXCL5 is critical for PMN-driven destructive inflammation in pulmonary tuberculosis.

MicroRNA-223 controls susceptibility to tuberculosis by regulating lung neutrophil recruitment.

The molecular mechanisms that control innate immune cell trafficking during chronic infection and inflammation, such as in tuberculosis (TB), are incompletely understood. During active TB, myeloid cells infiltrate the lung and sustain local inflammation. While the chemoattractants that orchestrate these processes are increasingly recognized, the posttranscriptional events that dictate their availability are unclear. We identified microRNA-223 (miR-223) as an upregulated small noncoding RNA in blood and lung parenchyma of TB patients and during murine TB. Deletion of miR-223 rendered TB-resistant mice highly susceptible to acute lung infection. The lethality of miR-223(­/­) mice was apparently not due to defects in antimycobacterial T cell responses. Exacerbated TB in miR-223(­/­) animals could be partially reversed by neutralization of CXCL2, CCL3, and IL-6, by mAb depletion of neutrophils, and by genetic deletion of Cxcr2. We found that miR-223 controlled lung recruitment of myeloid cells, and consequently, neutrophil-driven lethal inflammation. We conclude that miR-223 directly targets the chemoattractants CXCL2, CCL3, and IL-6 in myeloid cells. Our study not only reveals an essential role for a single miRNA in TB, it also identifies new targets for, and assigns biological functions to, miR-223. By regulating leukocyte chemotaxis via chemoattractants, miR-223 is critical for the control of TB and potentially other chronic inflammatory diseases.

CXCL9/Mig mediates T cells recruitment to valvular tissue lesions of chronic rheumatic heart disease patients.

Rheumatic fever (RF) is an autoimmune disease triggered by Streptococcus pyogenes infection frequently observed in infants from developing countries. Rheumatic heart disease (RHD), the major sequel of RF, leads to chronic inflammation of the myocardium and valvular tissue. T cells are the main population infiltrating cardiac lesions; however, the chemokines that orchestrate their recruitment are not clearly defined. Here, we investigated the expression of chemokines and chemokine receptors in cardiac tissue biopsies obtained from chronic RHD patients. Our results showed that CCL3/MIP1α gene expression was upregulated in myocardium while CCL1/I-309 and CXCL9/Mig were highly expressed in valvular tissue. Auto-reactive T cells that infiltrate valvular lesions presented a memory phenotype (CD4(+)CD45RO(+)) and migrate mainly toward CXCL9/Mig gradient. Collectively, our results show that a diverse milieu of chemokines is expressed in myocardium and valvular tissue lesions and emphasize the role of CXCL9/Mig in mediating T cell recruitment to the site of inflammation in the heart.

Biomarkers of inflammation, immunosuppression and stress with active disease are revealed by metabolomic profiling of tuberculosis patients.

Although tuberculosis (TB) causes more deaths than any other pathogen, most infected individuals harbor the pathogen without signs of disease. We explored the metabolome of >400 small molecules in serum of uninfected individuals, latently infected healthy individuals and patients with active TB. We identified changes in amino acid, lipid and nucleotide metabolism pathways, providing evidence for anti-inflammatory metabolomic changes in TB. Metabolic profiles indicate increased activity of indoleamine 2,3 dioxygenase 1 (IDO1), decreased phospholipase activity, increased abundance of adenosine metabolism products, as well as indicators of fibrotic lesions in active disease as compared to latent infection. Consistent with our predictions, we experimentally demonstrate TB-induced IDO1 activity. Furthermore, we demonstrate a link between metabolic profiles and cytokine signaling. Finally, we show that 20 metabolites are sufficient for robust discrimination of TB patients from healthy individuals. Our results provide specific insights into the biology of TB and pave the way for the rational development of metabolic biomarkers for TB.

Granulysin-expressing CD4+ T cells as candidate immune marker for tuberculosis during childhood and adolescence.

BACKGROUND: Granulysin produced by cytolytic T cells directly contributes to immune defense against tuberculosis (TB). We investigated granulysin as a candidate immune marker for childhood and adolescent TB. METHODS: Peripheral blood mononuclear cells (PBMC) from children and adolescents (1-17 years) with active TB, latent TB infection (LTBI), nontuberculous mycobacteria (NTM) infection and from uninfected controls were isolated and restimulated in a 7-day restimulation assay. Intracellular staining was then performed to analyze antigen-specific induction of activation markers and cytotoxic proteins, notably, granulysin in CD4(+) CD45RO(+) memory T cells. RESULTS: CD4(+) CD45RO(+) T cells co-expressing granulysin with specificity for Mycobacterium tuberculosis (Mtb) were present in high frequency in TB-experienced children and adolescents. Proliferating memory T cells (CFSE(low)CD4(+)CD45RO(+)) were identified as main source of granulysin and these cells expressed both central and effector memory phenotype. PBMC from study participants after TB drug therapy revealed that granulysin-expressing CD4(+) T cells are long-lived, and express several activation and cytotoxicity markers with a proportion of cells being interferon-gamma-positive. In addition, granulysin-expressing T cell lines showed cytolytic activity against Mtb-infected target cells. CONCLUSIONS: Our data suggest granulysin expression by CD4(+) memory T cells as candidate immune marker for TB infection, notably, in childhood and adolescence.

Heterozygosity for the S180L variant of MAL/TIRAP, a gene expressing an adaptor protein in the Toll-like receptor pathway, is associated with lower risk of developing chronic Chagas cardiomyopathy.

BACKGROUND: Chagas disease is caused by the protozoan parasite Trypanosoma cruzi. Among T. cruzi-infected individuals, only a subgroup develops severe chronic Chagas cardiomyopathy (CCC); the majority remain asymptomatic. T. cruzi displays numerous ligands for the Toll-like receptors (TLRs), which are an important component of innate immunity that lead to the transcription of proinflammatory cytokines by nuclear factor-kappaB. Because proinflammatory cytokines play an important role in CCC, we hypothesized that single-nucleotide polymorphisms (SNPs) in the genes that encode proteins in the TLR pathway could explain differential susceptibility to CCC among T. cruzi-infected individuals. METHODS: For 169 patients with CCC and 76 T. cruzi-infected, asymptomatic individuals, we analyzed SNPs by use of polymerase chain reaction-restriction fragment length polymorphism analysis for the genes TLR1, TLR2, TLR4, TLR5, TLR9, and MAL/TIRAP, which encodes an adaptor protein. RESULTS: Heterozygous carriers of the MAL/TIRAP variant S180L were more prevalent in the asymptomatic group (24 [32%] of 76 subjects) than in the CCC group (21 [12%] of 169) (chi2=12.6; P=.0004 [adjusted P (Pc)=.0084]; odds ratio [OR], 0.31 [95% confidence interval {CI}, 0.16-0.60]). Subgroup analysis showed a stronger association when asymptomatic patients were compared with patients who had severe CCC (i.e., patients with left-ventricular ejection fraction40%) (chi2=7.7; P=.005 [Pc=.11]; OR, 0.33 [95% CI, 0.15-0.73]). CONCLUSION: T. cruzi-infected individuals who are heterozygous for the MAL/TIRAP S180L variant that leads to a decrease in signal transduction upon ligation of TLR2 or TLR4 to their respective ligand may have a lower risk of developing CCC.

PDIA3, HSPA5 and vimentin, proteins identified by 2-DE in the valvular tissue, are the target antigens of peripheral and heart infiltrating T cells from chronic rheumatic heart disease patients.

Rheumatic fever (RF) is a post-infectious autoimmune disease due to sequel of group A streptococcus (GAS) pharyngitis. Rheumatic heart disease (RHD), the major manifestation of RF, is characterized by inflammation of heart valves and myocardium. Molecular mimicry between GAS antigens and host proteins has been shown at B and T cell level. However the identification of the autoantigens recognized by B and T cells within the inflammatory microenvironment of heart tissue in patients with RHD is still incompletely elucidated. In the present study, we used two-dimensional gel electrophoresis (2-DE) and mass spectrometry to identify valvular tissue proteins target of T cells from chronic RHD patients. We could identify three proteins recognized by heart infiltrating and peripheral T cells as protein disulfide isomerase ER-60 precursor (PDIA3), 78kD glucose-regulated protein precursor (HSPA5) and vimentin, with coverage of 45%, 43 and 34%, respectively. These proteins were recognized in a proliferation assay by peripheral and heart infiltrating T cells from RHD patients suggesting that they may be involved in the autoimmune reactions that leads to valve damage. We also observed that several other proteins isolated by 2-DE but not identified by mass spectrometry were also recognized by T cells. The identified cardiac proteins are likely relevant antigens involved in T cell-mediated autoimmune responses in RF/RHD that may contribute to the development of RHD.

Association of mannose-binding lectin gene polymorphism but not of mannose-binding serine protease 2 with chronic severe aortic regurgitation of rheumatic etiology.

N-Acetylglucosamine (GlcNAc) is the major immunoepitope of group A streptococcal cell wall carbohydrates. Antistreptococcal antibodies cross-reactive with anti-GlcNAc and laminin are present in sera of patients with rheumatic fever. The cross-reactivity of these antibodies with human heart valvular endothelium and the underlying basement membrane has been suggested to be a possible cause of immune-mediated valve lesion. Mannose-binding lectin (MBL) encoded by the MBL2 gene, a soluble pathogen recognition receptor, has high affinity for GlcNAc. We postulated that mutations in exon 1 of the MBL2 gene associated with a deficient serum level of MBL may contribute to chronic severe aortic regurgitation (AR) of rheumatic etiology. We studied 90 patients with severe chronic AR of rheumatic etiology and 281 healthy controls (HC) for the variants of the MBL2 gene at codons 52, 54, and 57 by using a PCR-restriction fragment length polymorphism-based method. We observed a significant difference in the prevalence of defective MBL2 alleles between patients with chronic severe AR and HC. Sixteen percent of patients with chronic severe AR were homozygotes or compound heterozygotes for defective MBL alleles in contrast to 5% for HC (P = 0.0022; odds ratio, 3.5 [95% confidence interval, 1.6 to 7.7]). No association was detected with the variant of the MASP2 gene. Our study suggests that MBL deficiency may contribute to the development of chronic severe AR of rheumatic etiology.