Single-Nucleotide Polymorphism and Copy Number Variation of the Multidrug Resistance-1 Locus of Plasmodium vivax: Local and Global Patterns

Emerging resistance to chloroquine (CQ) poses a major challenge for Plasmodium vivax malaria control, and nucleotide substitutions and copy number variation in the P. vivax multidrug resistance 1 (pvmdr-1) locus, which encodes a digestive vacuole membrane transporter, may modulate this phenotype. We describe patterns of genetic variation in pvmdr-1 alleles from Acre and Amazonas in northwestern Brazil, and compare then with those reported in other malaria-endemic regions. The pvmdr-1 mutation Y976F, which is associated with CQ resistance in Southeast Asia and Oceania, remains rare in northwestern Brazil (1.8%) and its prevalence mirrors that of CO resistance worldwide. Gene amplification of pvmdr-1, which is associated with mefloquine resistance but increased susceptibility to CO, remains relatively rare in northwestern Brazil (0.9%) and globally (< 4%), but became common (> 10%) in Tak Province, Thailand, possibly because of drug-mediated selection. The global database we have assembled provides a baseline for further studies of genetic variation in pvmdr-1 and drug resistance in P. vivax malaria.

Frequency of LCT -13910C>T single nucleotide polymorphism associated with adult-type hypolactasia/lactase persistence among Brazilians of different ethnic groups

Abstract Background Adult-type hypolactasia, the physiological decline of lactase some time after weaning, was previously associated with the LCT -13910C>T polymorphism worldwide except in Africa. Lactase non-persistence is the most common phenotype in humans, except in northwestern Europe with its long history of pastoralism and milking. We had previously shown association of LCT -13910C>T polymorphism with adult-type hypolactasia in Brazilians; thus, we assessed its frequency among different Brazilian ethnic groups. Methods We investigated the ethnicity-related frequency of this polymorphism in 567 Brazilians [mean age, 42.1 ± 16.8 years; 157 (27.7%) men]; 399 (70.4%) White, 50 (8.8%) Black, 65 (11.5%) Brown, and 53 (9.3%) Japanese-Brazilian. DNA was extracted from leukocytes; LCT -13910C>T polymorphism was analyzed by PCR-restriction fragment length polymorphism. Results Prevalence of the CC genotype associated with hypolactasia was similar (57%) among White and Brown groups; however, prevalence was higher among Blacks (80%) and those of Japanese descent (100%). Only 2 (4%) Blacks had TT genotype, and 8 (16%) had the CT genotype. Assuming an association between CC genotype and hypolactasia, and CT and TT genotypes with lactase persistence, 356 (62.8%) individuals had hypolactasia and 211 (37.2%) had lactase persistence. The White and Brown groups had the same hypolactasia prevalence (~57%); nevertheless, was 80% among Black individuals and 100% among Japanese-Brazilians (P < 0.01). Conclusion The lactase persistence allele, LCT -13910T, was found in about 43% of both White and Brown and 20% of the Black Brazilians, but was absent among all Japanese Brazilians studied.

Toll-like receptor 1 N248S single-nucleotide polymorphism is associated with leprosy risk and regulates immune activation during mycobacterial infection

Conflicting findings about the association between leprosy and TLR1 variants N248S and I602S have been reported. Here, we performed case-control and family based studies, followed by replication in 2 case-control populations from Brazil, involving 3162 individuals. Results indicated an association between TLR1 248S and leprosy in the case-control study (SS genotype odds ratio [OR], 1.81; P = .004) and the family based study (z = 2.02; P = .05). This association was consistently replicated in other populations (combined OR, 1.51; P < .001), corroborating the finding that 248S is a susceptibility factor for leprosy. Additionally, we demonstrated that peripheral blood mononuclear cells (PBMCs) carrying 248S produce a lower tumor necrosis factor/interleukin-10 ratio when stimulated with Mycobacterium leprae but not with lipopolysaccharide or PAM3cysK4. The same effect was observed after infection of PBMCs with the Moreau strain of bacillus Calmette-Guerin but not after infection with other strains. Finally, molecular dynamics simulations indicated that the Toll-like receptor 1 structure containing 248S amino acid is different from the structure containing 248N. Our results suggest that TLR1 248S is associated with an increased risk for leprosy, consistent with its hypoimmune regulatory function.

CYBA C242T polymorphism is associated with obesity and diabetes mellitus in Brazilian hypertensive patients

Diabet. Med. 29, e55e61 (2012) Abstract Aims The CYBA C242T polymorphism has been associated with cardiovascular phenotypes such as hypertension and atherosclerosis, but available data are conflicting. This report investigated the impact of this variant on hypertension and metabolic determinants of cardiovascular risk in a large Brazilian sample. Methods We cross-sectionally evaluated 1856 subjects (826 normotensive subjects and 1030 hypertensive patients) by clinical history, anthropometry, laboratory analysis and genotyping of the CYBA C242T polymorphism. Results Genotype frequencies in the whole population were consistent with the HardyWeinberg equilibrium and genotype distributions were not different between hypertensive and normotensive subjects. Hypertensive patients with the CC genotype presented lower fasting plasma glucose levels (5.9 +/- 0.1 vs. 6.2 +/- 0.1 mmol/l, P = 0.020) and waist circumference (94.5 +/- 0.6 vs. 96.3 +/- 0.6 cm, P = 0.028) than CT + TT ones. Similarly, the prevalence of diabetes mellitus and obesity was also lower in hypertensive patients carrying the CC genotype (16% vs. 21%, P = 0.041; 36% vs. 43%, P = 0.029, respectively). In addition, multiple and logistic regression analysis demonstrated that the CYBA C242T polymorphism was associated with glucose levels, waist circumference, obesity and diabetes mellitus in hypertensive patients independently of potential confounders. Conversely, in normotensive subjects, no significant difference in studied variables was detected between the genotype groups. Conclusions These data suggest that the T allele of the CYBA C242T polymorphism may be used as a marker for adverse metabolic features in Brazilian subjects with systemic hypertension.

Evaluation of MLH1 I219V Polymorphism in Unrelated South American Individuals Suspected of Having Lynch Syndrome

Background: Some single-nucleotide polymorphisms are associated with higher risk of colorectal cancer development and are suggested to explain part of the genetic contribution to Lynch syndrome. Aim: To evaluate the mutL homolog 1 (MLH1) I219V polymorphism in 124 unrelated South American individuals suspected of having Lynch syndrome, based on frequency, association with pathogenic MLH1 and mutS homolog 2 (MSH2) mutation and clinical features. Materials and Methods: DNA was obtained from peripheral blood and polymerase chain reaction (PCR) was performed, followed by direct sequencing. Results: The Val allelic of the I219V polymorphism was found in 51.61% (64/124) of the individuals, with an allelic frequency of 0.3. MLH1 or MHS2 pathogenic mutations were found in 32.81% (21/64) and in 23.33% (14/60) of Val-carriers and non-carriers, respectively. Conclusion: The Val-carrying genotype was frequent in the studied population; however, it does not appear to exert any modifier effect on MLH1 or MSH2 pathogenic mutations and the development of colorectal cancer.

Geographic differentiation of polymorphism in the Plasmodium falciparum malaria vaccine candidate gene SERA5

SERA5 is regarded as a promising malaria vaccine candidate of the most virulent human malaria parasite Plasmodium falciparum. SERA5 is a 120 kDa abundantly expressed blood-stage protein containing a papain-like protease. Since substantial polymorphism in blood-stage vaccine candidates may potentially limit their efficacy, it is imperative to fully investigate polymorphism of the SERA5 gene (sera5). In this study, we performed evolutionary and population genetic analysis of sera5. The level of inter-species divergence (kS = 0.076) between P. falciparum and Plasmodium reichenowi, a closely related chimpanzee malaria parasite is comparable to that of housekeeping protein genes. A signature of purifying selection was detected in the proenzyme and enzyme domains. Analysis of 445 near full-length P. falciparum sera5 sequences from nine countries in Africa, Southeast Asia, Oceania and South America revealed extensive variations in the number of octamer repeat (OR) and serine repeat (SR) regions as well as substantial level of single nucleotide polymorphism (SNP) in non-repeat regions (2562 bp). Remarkably, a 14 amino acid sequence of SERA5 (amino acids 59-72) that is known to be the in vitro target of parasite growth inhibitory antibodies was found to be perfectly conserved in all 445 worldwide isolates of P. falciparum evaluated. Unlike other major vaccine target antigen genes such as merozoite surface protein-1, apical membrane antigen-1 or circumsporozoite protein, no strong evidence for positive selection was detected for SNPs in the non-repeat regions of sera5. A biased geographical distribution was observed in SNPs as well as in the haplotypes of the sera5 OR and SR regions. In Africa, OR- and SR-haplotypes with low frequency (<5%) and SNPs with minor allele frequency (<5%) were abundant and were mostly continent-specific. Consistently, significant genetic differentiation, assessed by the Wright's fixation index (FST) of inter-population variance in allele frequencies, was detected for SNPs and both OR- and SR-haplotypes among almost all parasite populations. The exception was parasite populations between Tanzania and Ghana, suggesting frequent gene flow in Africa. The present study points to the importance of investigating whether biased geographical distribution for SNPs and repeat variants in the OR and SR regions affect the reactivity of human serum antibodies to variants. (C) 2011 Elsevier Ltd. All rights reserved.

Somatotroph pituitary adenoma with acromegaly and autosomal dominant polycystic kidney disease: SSTR5 polymorphism and PKD1 mutation

A 39-year-old woman with autosomal dominant polycystic kidney disease (ADPKD) presented with acromegaly and a pituitary macroadenoma. There was a family history of this renal disorder. She had undergone surgery for pituitary adenoma 6 years prior. Physical examination disclosed bitemporal hemianopsia and elevation of both basal growth hormone (GH) 106 ng/mL (normal 0-5) and insulin-like growth factor (IGF-1) 811 ng/mL (normal 48-255) blood levels. A magnetic resonance imaging scan disclosed a 3.0 cm sellar and suprasellar mass with both optic chiasm compression and left cavernous sinus invasion. Pathologic, cytogenetic, molecular and in silico analysis was undertaken. Histologic, immunohistochemical and ultrastructural studies of the lesion disclosed a sparsely granulated somatotroph adenoma. Standard chromosome analysis on the blood sample showed no abnormality. Sequence analysis of the coding regions of PKD1 and PKD2 employing DNA from both peripheral leukocytes and the tumor revealed the most common PKD1 mutation, 5014_5015delAG. Analysis of the entire SSTR5 gene disclosed the variant c.142C > A (p.L48M, rs4988483) in the heterozygous state in both blood and tumor, while no pathogenic mutations were noted in the MEN1, AIP, p27Kip1 and SSTR2 genes. To our knowledge, this is the fourth reported case of a GH-producing pituitary adenoma associated with ADPKD, but the first subjected to extensive morphological, ultrastructural, cytogenetic and molecular studies. The physical proximity of the PKD1 and SSTR5 genes on chromosome 16 suggests a causal relationship between ADPKD and somatotroph adenoma.

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 December 2011-31 January 2012

This article documents the addition of 473 microsatellite marker loci and 71 pairs of single-nucleotide polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Barteria fistulosa, Bombus morio, Galaxias platei, Hematodinium perezi, Macrocentrus cingulum Brischke (a.k.a. M.abdominalis Fab., M.grandii Goidanich or M.gifuensis Ashmead), Micropogonias furnieri, Nerita melanotragus, Nilaparvata lugens Stal, Sciaenops ocellatus, Scomber scombrus, Spodoptera frugiperda and Turdus lherminieri. These loci were cross-tested on the following species: Barteria dewevrei, Barteria nigritana, Barteria solida, Cynoscion acoupa, Cynoscion jamaicensis, Cynoscion leiarchus, Cynoscion nebulosus, Cynoscion striatus, Cynoscion virescens, Macrodon ancylodon, Menticirrhus americanus, Nilaparvata muiri and Umbrina canosai. This article also documents the addition of 116 sequencing primer pairs for Dicentrarchus labrax.

FGFR4 Profile as a Prognostic Marker in Squamous Cell Carcinoma of the Mouth and Oropharynx

Background: Fibroblast growth factor receptor 4 (FGFR4) is a member of a receptor tyrosine kinase family of enzymes involved in cell cycle control and proliferation. A common single nucleotide polymorphism (SNP) Gly388Arg variant has been associated with increased tumor cell motility and progression of breast cancer, head and neck cancer and soft tissue sarcomas. The present study evaluated the prognostic significance of FGFR4 in oral and oropharynx carcinomas, finding an association of FGFR4 expression and Gly388Arg genotype with tumor onset and prognosis. Patients and Methods: DNA from peripheral blood of 122 patients with oral and oropharyngeal squamous cell carcinomas was used to determine FGFR4 genotype by PCR-RFLP. Protein expression was assessed by immunohistochemistry (IHC) on paraffin-embedded tissue microarrays. Results: Presence of allele Arg388 was associated with lymphatic embolization and with disease related premature death. In addition, FGFR4 low expression was related with lymph node positivity and premature relapse of disease, as well as disease related death. Conclusion: Our results propose FGFR4 profile, measured by the Gly388Arg genotype and expression, as a novel marker of prognosis in squamous cell carcinoma of the mouth and oropharynx.

Association between mannose-binding lectin and interleukin-1 receptor antagonist gene polymorphisms and recurrent vulvovaginal candidiasis

Objective The influence of functional polymorphisms in the genes coding for mannose-binding lectin (MBL) and interleukin-1 receptor antagonist (IL-1ra) on recurrent vulvovaginal candidiasis (RVVC) were examined in an urban Brazilian population. Methods DNA was isolated from buccal swabs of 100 women with RVVC and 100 control women and tested by gene amplification for a single nucleotide polymorphism in codon 54 of the MBL2 gene and for a length polymorphism in intron 2 of the IL1RN gene. Genotype and allele frequencies were compared between groups. Results The frequency of the variant MBL2 B allele, associated with reduced circulating and vaginal MBL concentrations, was 27.0% in RVVC and 8.5% in control women (p < .0001). The MBL2 B, B genotype was present in 12% of RVVC patients and 1% of controls (p = .0025). The IL1RN 2 allele frequency, associated with the highest level of unopposed IL-1 beta activity, was 24.0% in RVVC and 23.4% in controls. The IL1RN genotype distribution was also similar in both groups. Conclusion Carriage of the MBL2 codon 54 polymorphism, but not the IL1RN length polymorphism, predisposes to RVVC in Brazilian women.

Germline mutation analysis of Tpit in Poodle dogs with ACTH-dependent hyperadrenocorticism

There is a high incidence of pituitary-dependent hyperadrenocorticism (PDH) in Poodle dogs, with family members being affected by the disease, suggesting a genetic involvement. Tpit is an obligate transcription factor for the expression of pro-opiomelanocortingene and for corticotroph terminal differentiation. The aim of the present study was to screen the Tpit gene for germline mutations in Poodles with PDH. Fifty Poodle dogs (33 female, 8.71 +/- 2.8 years) with PDH and 50 healthy Poodle dogs (32 females, 9.4241 2.8 years) were studied. Genomic DNA was isolated from peripheral blood, amplified by PCR and submitted to automatic sequence. No mutation in the coding region of Tpit was found, whereas the new single nucleotide polymorphism p.S343G, in heterozygous state, was found in the same frequency in both PDH and control groups. We concluded that Tpit gain-of-function mutations are not involved in the etiology of PDH in Poodle dogs.

Association between diet and polymorphisms in individuals with statin-controlled dyslipidaemia grouped according to oxidative stress biomarkers

The objective of this study was to investigate whether differences in diet and in single-nucleotide polymorphisms (SNPs) found in paraoxonase-1 (PON-1), 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), cholesterol ester transfer protein (CETP) and apolipoprotein E (APOE) genes, are associated with oxidative stress biomarkers and consequently with susceptibility of low-density cholesterol (LDL) to oxidation. A multivariate approach was applied to a group of 55 patients according to three biomarkers: plasma antioxidant activity, malondialdehyde and oxidized LDL (oxLDL) concentrations. Individuals classified in Cluster III showed the worst prognoses in terms of antioxidant activity and oxidative status. Individuals classified in Cluster I presented the lowest oxidative status, while individuals grouped in Cluster II presented the highest levels of antioxidant activity. No difference in nutrient intake was observed among the clusters. Significantly higher gamma- and delta-tocopherol concentrations were observed in those individuals with the highest levels of antioxidant activity. No single linear regression was statistically significant, suggesting that mutant alleles of the SNPs selected did not contribute to the differences observed in oxidative stress response. Although not statistically significant, the p value of the APO E coefficient for oxLDL response was 0.096, indicating that patients who carry the TT allele of the APO E gene tend to present lower plasma oxLDL concentrations. Therefore, the differences in oxidative stress levels observed in this study could not be attributed to diet or to the variant alleles of PON-1, CETP, HMGCR or APO E. This data supports the influence of gamma-tocopherol and delta-tocopherol on antioxidant activity, and highlights the need for further studies investigating APO E alleles and LDL oxidation.

Using Prior Information from the Medical Literature in GWAS of Oral Cancer Identifies Novel Susceptibility Variant on Chromosome 4-the AdAPT Method

Background: Genome-wide association studies (GWAS) require large sample sizes to obtain adequate statistical power, but it may be possible to increase the power by incorporating complementary data. In this study we investigated the feasibility of automatically retrieving information from the medical literature and leveraging this information in GWAS. Methods: We developed a method that searches through PubMed abstracts for pre-assigned keywords and key concepts, and uses this information to assign prior probabilities of association for each single nucleotide polymorphism (SNP) with the phenotype of interest - the Adjusting Association Priors with Text (AdAPT) method. Association results from a GWAS can subsequently be ranked in the context of these priors using the Bayes False Discovery Probability (BFDP) framework. We initially tested AdAPT by comparing rankings of known susceptibility alleles in a previous lung cancer GWAS, and subsequently applied it in a two-phase GWAS of oral cancer. Results: Known lung cancer susceptibility SNPs were consistently ranked higher by AdAPT BFDPs than by p-values. In the oral cancer GWAS, we sought to replicate the top five SNPs as ranked by AdAPT BFDPs, of which rs991316, located in the ADH gene region of 4q23, displayed a statistically significant association with oral cancer risk in the replication phase (per-rare-allele log additive p-value [p(trend)] = 2.5 x 10(-3)). The combined OR for having one additional rare allele was 0.83 (95% CI: 0.76-0.90), and this association was independent of previously identified susceptibility SNPs that are associated with overall UADT cancer in this gene region. We also investigated if rs991316 was associated with other cancers of the upper aerodigestive tract (UADT), but no additional association signal was found. Conclusion: This study highlights the potential utility of systematically incorporating prior knowledge from the medical literature in genome-wide analyses using the AdAPT methodology. AdAPT is available online (url: http://services.gate.ac.uk/lld/gwas/service/config).

Using gene-network landscape to dissect genotype effects of TCF7L2 genetic variant on diabetes and cardiovascular risk

Vaquero AR, Ferreira NE, Omae SV, Rodrigues MV, Teixeira SK, Krieger JE, Pereira AC. Using gene-network landscape to dissect genotype effects of TCF7L2 genetic variant on diabetes and cardiovascular risk. Physiol Genomics 44: 903-914, 2012. First published August 7, 2012; doi:10.1152/physiolgenomics.00030.2012.-The single nucleotide polymorphism (SNP) within the TCF7L2 gene, rs7903146, is, to date, the most significant genetic marker associated with Type 2 diabetes mellitus (T2DM) risk. Nonetheless, its functional role in disease pathology is poorly understood. The aim of the present study was to investigate, in vascular smooth muscle cells from 92 patients undergoing aortocoronary bypass surgery, the contribution of this SNP in T2DM using expression levels and expression correlation comparison approaches, which were visually represented as gene interaction networks. Initially, the expression levels of 41 genes (seven TCF7L2 splice forms and 40 other T2DM relevant genes) were compared between rs7903146 wild-type (CC) and T2DM-risk (CT + TT) genotype groups. Next, we compared the expression correlation patterns of these 41 genes between groups to observe if the relationships between genes were different. Five TCF7L2 splice forms and nine genes showed significant expression differences between groups. RXR alpha gene was pinpointed as showing the most different expression correlation pattern with other genes. Therefore, T2DM risk alleles appear to be influencing TCF7L2 splice form's expression in vascular smooth muscle cells, and RXR alpha gene is pointed out as a treatment target candidate for risk reduction in individuals with high risk of developing T2DM, especially individuals harboring TCF7L2 risk genotypes.

Haplotypes of susceptibility to chronic periodontitis in the Interleukin 8 gene do not influence protein level in the gingival crevicular fluid

Objective: Previously, we identified that the ATC/TTC haplotype formed by polymorphisms in the Interleukin-(IL)8 gene conferred susceptibility to chronic periodontitis (CP). The aim of the study was to investigate whether the IL8 haplotype ATC/TTC was associated with the volume of gingival crevicular fluid (GCF), the concentration of interleukin IL-8 in the GCF, as well as periodontal conditions in patients with CP in comparison to controls without CP. Methods: Seventy-nine individuals (CP: n = 41, controls: n = 38) were grouped according to the presence (susceptible for CP) or absence (not susceptible for CP) of the IL8 ATC/TTC haplotype. After periodontal clinical evaluation, they were subdivided by the presence or absence of CP. GCF was collected from each patient and the IL-8 levels were determined by ELISA. The GCF volume of each subject was measured by means of a calibrated electronic device. Comparisons of means between carriers and non-carriers of the ATC/TTC haplotype were evaluated using the Mann-Whitney test. Linear regression and stepwise linear regression analysis were used to analyse the association of the GCF volume with potential covariates and their contribution for the phenotype. Results: We did not find significant differences of both periodontal conditions and IL-8 concentration in the GCF of patients with the presence or absence of the IL8 ATC/TTC haplotype. However, the GCF volume was significantly higher amongst the patients affected by CP that are absent for the IL8 ATC/TTC haplotype. In addition, linear regression analysis showed a statistically significant association between GCF volume and CP, IL8 haplotype ATC/TTC and IL-8 concentration. Conclusions: The IL8 haplotype of susceptibility to CP was neither associated with IL-8 cytokine levels nor with clinical periodontal parameters. Also, CP, IL8 haplotype and IL-8 concentration showed a positive association with the GCF volume levels in the studied patients. (c) 2012 Published by Elsevier Ltd.