Finding Rare, Disease-Associated Variants in Isolated Groups: Potential Advantages of Mennonite Populations

Large-scale genotyping and next-generation sequencing techniques have allowed great advances in the field of molecular genetics. Numerous common variants of low impact have been associated with many complex human traits and diseases, such as bipolar disorder and schizophrenia. Although they may exert a greater impact on risk, few rare disease variants have been found, owing to the greatly increased sample sizes that are typically necessary to demonstrate association with rarer variants. One alternative strategy is to study isolated populations, where historical bottlenecks reduce genetic diversity and some otherwise rare variants may drift to higher frequencies. Here we describe the Mennonite population settlements, considering their history of multiple bottlenecks followed by demographic expansion and a currently widespread geographical distribution. We argue that Mennonite populations are valuable partners for studies seeking genetic variants that exert a high impact on risk for a variety of common disorders, including mental illnesses

Association of MASP-2 levels and MASP2 gene polymorphisms with rheumatoid arthritis in patients and their relatives.

BACKGROUND: Mannan-binding lectin-associated serine protease 2 (MASP-2) is a key protein of the lectin pathway of complement. MASP-2 levels have been associated with different polymorphisms within MASP2 gene as well as with the risk for inflammatory disorders and infections. Despite its clinical importance, MASP-2 remains poorly investigated in rheumatoid arthritis (RA). METHODS: In this case-control study, we measured MASP-2 serum levels in 156 RA patients, 44 patient relatives, and 100 controls from Southern Brazil, associating the results with nine MASP2 polymorphisms in all patients, 111 relatives, and 230 controls genotyped with multiplex SSP-PCR. RESULTS: MASP-2 levels were lower in patients than in controls and relatives (medians 181 vs. 340 or 285 ng/ml, respectively, P<0.0001). Conversely, high MASP-2 levels were associated with lower susceptibility to RA and to articular symptoms independently of age, gender, ethnicity, smoking habit, anti-CCP and rheumatoid factor positivity (OR = 0.05 [95%CI = 0.019-0.13], P<0.0001 between patients and controls; OR = 0.12, [95%CI = 0.03-0.45], P = 0.002 between patients and relatives; OR = 0.06, [95%CI = 0.004-0.73], P = 0.03 between relatives with and without articular symptoms). MASP2 haplotypes *2A1 and *2B1-i were associated with increased susceptibility to RA (OR = 3.32 [95%CI = 1.48-7.45], P = 0.004). Deficiency-causing p.120G and p.439H substitutions were associated with five times increased susceptibility to articular symptoms in relatives (OR = 5.13 [95%CI = 1.36-20.84], P = 0.02). There was no association of MASP-2 levels or MASP2 polymorphisms with autoantibodies, Sjögren's syndrome, nodules and functional class. CONCLUSIONS: In this study, we found the first evidence that MASP-2 deficiency might play an important role in the development of RA and articular symptoms among relatives of RA patients

Association of L-ficolin levels and FCN2 genotypes with chronic Chagas disease.

BACKGROUND: L-ficolin (encoded by FCN2) binds to acetylated sugar moieties of many pathogens, including Trypanosoma cruzi, promoting their phagocytosis and lysis by the complement system. METHODS: We investigated L-ficolin levels in 160 T. cruzi infected patients with chronic Chagas disease and 71 healthy individuals, and FCN2 polymorphisms (-986 G>A, -602 G>A, and -4 A>G in the promoter and A258S in exon 8) in 243 patients, being 88 indeterminate (asymptomatic), 96 with cardiac, 23 with digestive and 33 with cardiodigestive manifestations (two were unspecified) and 305 controls (135 for A258S). RESULTS: Patients presented lower L-ficolin plasma levels than controls (p<0.0001). Among the different groups of cardiac commitment, individuals with moderate forms had higher L-ficolin levels than the severe forms (P = 0.039). Lower L-ficolin levels were found associated with the 258S variant in the patients (P = 0.034). We found less -4A/G heterozygotes in the cardiac patients, than in the controls (OR = 0.56 [95% CI = 0.33-0.94], P = 0.034). Heterozygote -4A/G genotypes with the 258S variant and 258SS homozygotes were nevertheless more frequent among cardiodigestive patients than in controls (OR = 14.1 [95% CI = 3.5-56.8], P = 0.0001) and in indeterminate patients (OR = 3.2 [95% CI = 1.1-9.4], P = 0.037). We also found an association of the allelic frequency of the 258S variant with cardiodigestive Chagas disease compared to controls (OR = 2.24 [95% CI = 1.1-4.5], P = 0.037). Thus, decreased patient levels of L-ficolin reflect not only protein consumption due to the disease process, but also the higher frequency of the 258S variant in patients with cardiodigestive symptoms. CONCLUSION: The very first study on Brazilian cohort associates both L-ficolin plasma levels and FCN2 variants to Chagas disease and subsequent disease progression. The prognostic value of L-ficolin levels and the FCN2*A258S polymorphism should be further evaluated in other settings

<i>MASP2</i> haplotypes nomenclature and frequencies (% ± SD).

a<p>: phylogenetic nomenclature according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090979#pone.0090979-Boldt2" target="_blank">[26]</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090979#pone.0090979-Nebert1" target="_blank">[38]</a>, where “h”, “ï” and “l” refer to <i>MASP2</i> haplotypes associated with low (<200 ng/ml), intermediary (200–600 ng/ml) and high (≥600 ng/ml) MASP-2 levels (cut off adopted by <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090979#pone.0090979-Boldt2" target="_blank">[26]</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090979#pone.0090979-Probst1" target="_blank">[35]</a>),</p>b<p>: reference sequence NT_021937 (GenBank sequence); anti-CCP: anti-cyclic citrullinated peptide antibodies; n: number of chromosomes; SD: standard deviation. In bold: recombinant haplotype.</p

MASP-2 levels in RA patients, relatives and controls according to haplotype producing profiles.

<p>Data is shown with medians, differences were calculated with Kruskal-Wallis test A: Low MASP-2 producing haplotypes (<i>*2B2B-l/*1A</i>, <i>*2B2A-i/*2B2B-l</i>, <i>2B2B-l/2A1</i>, <i>*2B2A-i/*2A2-l</i>, <i>2B1-i/2A2-l</i>, <i>*2B1-i/*1C2-l</i>, <i>*1C2-l/*1C2-l</i>, <i>*2B2A-i/*1C2-l</i>, <i>*2B2B-l/*1C2-l</i>, <i>*2A2-l/*1C2-l</i>); B: Intermediary MASP-2 producing haplotypes (<i>*2B2A-i/*2B2A-i</i>, <i>*2B2A-i/*1A</i>, <i>*2B2A-i/*2B1-i</i>, <i>*2B1-i/*1A</i>, <i>*2B1-i/2A1</i>, <i>*2B1-i/*2B1-i</i>); C: High MASP-2 producing haplotypes (<i>*2B2A-i/*1B1-h</i>, <i>*2B2A-i/*1B2-h</i>, <i>*2B1-i/*1B1-h</i>, <i>*1B1-h/*1B1-h</i>, <i>*1A/*1B1-h</i>, <i>*2A1/1B1-h</i>, <i>*2A1/*2A1</i>, <i>*2B2A-i/*2A1</i>, <i>*2A1/*1A</i>, <i>*2B2A-i.1B1-h/1B1-h</i>).</p

Clinical and demographic data of RA patients.

a<p>: Total number of samples with available data for the extra-articular manifestation.</p

Reduced binary logistic regression models for rheumatoid arthritis.

<p>The original model included age, gender and ethnicity for all observations; smoking habit, anti-CCP and RF positivity for patients and relatives. Significance of the model is given in parentheses. MASP-2 levels were normalized as logarithms on base 10 (Log₁₀). anti-CCP: anti-cyclic citrullinated peptide antibodies; n: number of included observations; &: not independent; OR: odds ratio;</p>a<p>: with intermediary levels,</p>b<p>: with low MASP-2 levels.</p

Differences between MASP-2 levels of patients, relatives with and without articular symptoms, and controls.

<p>Data is shown in medians and interquartile ranges, differences were calculated with Mann-Whitney tests (P<0.0001 between patients and controls, and patients and asymptomatic relatives; P = 0.0339 between symptomatic relatives and controls and P = 0.0143 between symptomatic and asymptomatic relatives). AS: articular symptom.</p