A comprehensive overview of the cystic fibrosis on the island of São Miguel (Azores, Portugal).

BACKGROUND: Early diagnosis and treatment are improving significantly the quality of life of patients with cystic fibrosis (CF). This recessive disease is caused by a great variability of mutations in the CF transmembrane conductance (CFTR) gene, whose spectrum and frequency can be different across populations. METHODS: We performed a retrospective cross-sectional study of CF patients from the island of São Miguel (Azores, Portugal) through a clinical, genealogical, genetic and epidemiological investigation. The clinical course of patients was analyzed as a whole and according to their genotype. RESULTS: We identified 14 CF patients within a 23-year period, corresponding to a cumulative incidence of 1:3012 births, being three of them born from consanguineous unions. Genetic analysis revealed three CFTR genotypes: p.[Ser4Ter];[Gln1100Pro] was present in one patient with a less severe phenotype (1/14); c.[120del23];p.[Phe508del], a very rare one (2/14); and p.[Phe508del];[Phe508del] in the remaining patients (11/14). Clinically, respiratory infections (8/14) and growth failure (6/14) were the most common initial manifestations. All patients presented pancreatic dysfunction, with 21.4 and 100% of them showing endocrine and exocrine insufficiency, respectively. As expected, patients with severe phenotype were homozygous for p.Phe508del and had the lowest value of body mass index. CONCLUSIONS: The present study demonstrated that São Miguel Island has an increased incidence of CF when compared to recent Portuguese data (1:7500 live births). It also allowed a comprehensive overview of CF in São Miguel, improving medical practice along with genetic counselling and creating opportunities for genotype-targeted therapies.

Diagnosis of Human Leptospirosis in a Clinical Setting: Real-Time PCR High Resolution Melting Analysis for Detection of Leptospira at the Onset of Disease.

Currently, direct detection of Leptospira can be done in clinical laboratories by conventional and by real-time PCR (qRT-PCR). We tested a biobank of paired samples of serum and urine from the same patient (202 patients) presenting at the hospital in an area endemic for leptospirosis using qRT-PCR followed by high resolution melting (HRM) analysis. The results were compared with those obtained by conventional nested PCR and with the serologic gold standard microscopic agglutination test (MAT). Differences were resolved by sequencing. qRT-PCR-HRM was positive for 46 of the 202 patients (22.7%, accuracy 100%) which is consistent with known prevalence of leptospirosis in the Azores. MAT results were positive for 3 of the 46 patients (6.5%). Analysis of paired samples allowed us to identify the illness point at which patients presented at the hospital: onset, dissemination or excretion. The melting curve analysis of Leptospira species revealed that 60.9% (28/46) of patients were infected with L. interrogans and 39.1% (18/46) were infected with L. borgpetersenii, both endemic to the Azores. We validated the use of qRT-PCR-HRM for diagnosis of leptospirosis and for identification of the Leptospira species at the earliest onset of infection in a clinical setting, in less than 2 hours.

A novel PLEC nonsense homozygous mutation (c.7159G > T; p.Glu2387*) causes epidermolysis bullosa simplex with muscular dystrophy and diffuse alopecia: a case report.

BACKGROUND: Epidermolysis bullosa simplex with muscular dystrophy (EBS-MD; OMIM #226670) is an autosomal recessive disease, characterized mainly by skin blistering at birth or shortly thereafter, progressive muscle weakness, and rarely by alopecia. EBS-MD is caused by mutations in the PLEC gene (OMIM *601282), which encodes plectin, a structural protein expressed in several tissues, including epithelia and muscle. We describe a patient affected with EBS-MD and diffuse alopecia in which we identified a novel pathogenic mutation by PCR amplification of all coding exons and exon-intron boundaries of PLEC gene, followed by bidirectional Sanger sequencing. CASE PRESENTATION: The patient, a 28-year-old female and only child of consanguineous healthy parents, was born after uneventful pregnancy. At 2 days of age, she developed skin and oral mucosal blistering, accompanied by voice hoarseness. On physical examination as an adult, we observed diffuse non-scarring alopecia on the scalp, onychodystrophy (pachyonychia) in all 20 nails, dental decay, mild dysphonia, and severe muscle atrophy mainly affecting the extremities. Neurological examination showed profoundly diminished reflexes. Mutation analysis revealed the patient to be homozygous for the novel PLEC nonsense mutation - c.7159G > T (p.Glu2387*) - located in exon 31. Thismutation predicts the lack of expression of the full-length plectin isoform. CONCLUSION: The present case appears to be the second association of EBS-MD with diffuse alopecia, both cases having different mutations involving PLEC exon 31. It remains to be elucidated whether diffuse alopecia results from PLEC mutations and/or from environmental factors.

Genetic variants involved in oxidative stress, base excision repair, DNA methylation, and folate metabolism pathways influence myeloid neoplasias susceptibility and prognosis.

Myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) share common features: elevated oxidative stress, DNA repair deficiency, and aberrant DNA methylation. We performed a hospital-based case-control study to evaluate the association in variants of genes involved in oxidative stress, folate metabolism, DNA repair, and DNA methylation with susceptibility and prognosis of these malignancies. To that end, 16 SNPs (one per gene: CAT, CYBA, DNMT1, DNMT3A, DNMT3B, GPX1, KEAP1, MPO, MTRR, NEIL1, NFE2F2, OGG1, SLC19A1, SOD1, SOD2, and XRCC1) were genotyped in 191 patients (101 MDS and 90 AML) and 261 controls. We also measured oxidative stress (reactive oxygen species/total antioxidant status ratio), DNA damage (8-hydroxy-2'-deoxyguanosine), and DNA methylation (5-methylcytosine) in 50 subjects (40 MDS and 10 controls). Results showed that five genes (GPX1, NEIL1, NFE2L2, OGG1, and SOD2) were associated with MDS, two (DNMT3B and SLC19A1) with AML, and two (CYBA and DNMT1) with both diseases. We observed a correlation of CYBA TT, GPX1 TT, and SOD2 CC genotypes with increased oxidative stress levels, as well as NEIL1 TT and OGG1 GG genotypes with higher DNA damage. The 5-methylcytosine levels were negatively associated with DNMT1 CC, DNMT3A CC, and MTRR AA genotypes, and positively with DNMT3B CC genotype. Furthermore, DNMT3A, MTRR, NEIL1, and OGG1 variants modulated AML transformation in MDS patients. Additionally, DNMT3A, OGG1, GPX1, and KEAP1 variants influenced survival of MDS and AML patients. Altogether, data suggest that genetic variability influence predisposition and prognosis of MDS and AML patients, as well AML transformation rate in MDS patients. © 2016 Wiley Periodicals, Inc.

Molecular diagnosis of infectious diseases in São Miguel Island (Azores, Portugal): A hospital-based descriptive study.

INTRODUCTION: We performed a descriptive analysis of molecular diagnosis of infectious agents in the São Miguel Island population, in order to address questions like what is the frequency of clinical requests, is it observable seasonality of pathogens, and what is the positive rate for the clinical diagnosis. METHODOLOGY: This was a retrospective and descriptive study based on 878 individuals suspected of harboring infectious diseases during two consecutive years, 2012-2013. More than 25 different pathogens were investigated by polymerase chain reaction (PCR)-based methods. The individuals were stratified into gender, occupation, and age groups. RESULTS: The pathogen with more clinical requests was hepatitis C virus, investigated in 225 individuals (30.0%), followed by Leptospira spp., in 187 (24.9%). Overall, data demonstrated a gender distribution bias, where 72.9% of cases were males. The age group of 25 to 44 years was the class with more clinical requests. Regarding occupation, a predominance of construction workers (12.0%) was observed, followed by retired workers (11.0%). Patient distribution per year showed a higher number of patients in the fall months. Diagnoses of leptospirosis and respiratory virus infections presented seasonality. CONCLUSIONS: The present study provides a valid contribution to the knowledge of the epidemiology of infectious diseases in the São Miguel Island (Azores, Portugal) population.

Genealogical and molecular analysis of a family-based cohort of congenital heart disease patients from the São Miguel Island (Azores, Portugal).

BACKGROUND: Congenital heart disease (CHD) is one common birth malformation, accounting for ∼30% of total congenital abnormalities. AIM: Considering the unknown role of consanguinity in causing CHD, this study hypothesised that consanguineous unions and/or familial aggregation may be frequent in the Azorean Island of São Miguel (Portugal). To that end, a retrospective observational study was performed based on genealogical and molecular analyses. SUBJECTS AND METHODS: The study enrolled 112 CHD patients from São Miguel Island, which allowed the assessment of type of family (simplex or multiplex), parental consanguinity and grandparental endogamy. Based on 15 STR markers, inbreeding coefficients (FIS) in the CHD cohort and healthy control group (n = 114) were estimated. RESULTS: Multiplex families were 37.6% (n = 41/109), a rate considerably higher than previously described in the literature (< 15%). Moreover, 9.2% (n = 10/109) of the CHD families were consanguineous, mostly derived from third cousin unions, and 20.2% (n = 22/109) presented full grandparental endogamy. Higher FIS values were found in patients with parental consanguinity (0.0371) and patent ductus arteriosus (0.0277). CONCLUSION: This study analysed several genealogical and genetic features related with CHD, revealing the presence of parental consanguinity and extensive familial aggregation in the CHD patients from São Miguel Island.

Carriers of the Complex Allele HFE c.[187C>G;340+4T>C] Have Increased Risk of Iron Overload in São Miguel Island Population (Azores, Portugal).

Iron overload is associated with acquired and genetic conditions, the most common being hereditary hemochromatosis (HH) type-I, caused by HFE mutations. Here, we conducted a hospital-based case-control study of 41 patients from the São Miguel Island (Azores, Portugal), six belonging to a family with HH type-I pseudodominant inheritance, and 35 unrelated individuals fulfilling the biochemical criteria of iron overload compatible with HH type-I. For this purpose, we analyzed the most common HFE mutations- c.845G>A [p.Cys282Tyr], c.187C>G [p.His63Asp], and c.193A>T [p.Ser65Cys]. Results revealed that the family's HH pseudodominant pattern is due to consanguineous marriage of HFE-c.845G>A carriers, and to marriage with a genetically unrelated spouse that is a -c.187G carrier. Regarding unrelated patients, six were homozygous for c.845A, and three were c.845A/c.187G compound heterozygous. We then performed sequencing of HFE exons 2, 4, 5 and their intron-flanking regions. No other mutations were observed, but we identified the -c.340+4C [IVS2+4C] splice variant in 26 (74.3%) patients. Functionally, the c.340+4C may generate alternative splicing by HFE exon 2 skipping and consequently, a protein missing the α1-domain essential for HFE/ transferrin receptor-1 interactions. Finally, we investigated HFE mutations configuration with iron overload by determining haplotypes and genotypic profiles. Results evidenced that carriers of HFE-c.187G allele also carry -c.340+4C, suggesting in-cis configuration. This data is corroborated by the association analysis where carriers of the complex allele HFE-c.[187C>G;340+4T>C] have an increased iron overload risk (RR = 2.08, 95% CI = 1.40-2.94, p<0.001). Therefore, homozygous for this complex allele are at risk of having iron overload because they will produce two altered proteins--the p.63Asp [c.187G], and the protein lacking 88 amino acids encoded by exon 2. In summary, we provide evidence that the complex allele HFE-c.[187C>G;340+4T>C] has a role, as genetic predisposition factor, on iron overload in the São Miguel population. Independent replication studies in other populations are needed to confirm this association.

Genetic variation in key genes associated with statin therapy in the Azores Islands (Portugal) healthy population.

BACKGROUND: Inter-individual variation in response to statins (efficacy and toxicity) has been described and may be due to polymorphisms implicated in drug pharmacokinetics or pharmacodynamics. AIM: This study investigates clinically relevant pharmacogenes underlying statin response in 170 healthy Azoreans. METHODS: Eight SNPs in candidate genes-HMGCR (rs3846662, rs17238540, rs17244841), CETP (rs708272), APOE (rs7412, rs429358) and SLCO1B1 (rs2306283, rs4149056)-were genotyped. RESULTS: The allele frequencies were similar to those reported for European derived populations, excepting SLCO1B1 c.388A>G (rs2306283), which has a significant difference when compared with the HapMap CEU population (p = 1 × 10(-8)). The results of statin efficacy showed that 9.1% of Azoreans are APOE4 carriers. This allele has been associated with lower LDLc reduction from statin therapy and also higher LDLc levels at baseline. Regarding SLCO1B1, associated with statin toxicity, 1.8% of individuals have two reduced-function alleles (c.521CC). CONCLUSION: The results contribute to overcome the lack of knowledge regarding the frequency of pharmacogenetic SNPs and their corresponding haplotypes in targeted populations, such as Azores islands. Moreover, the present work constitutes an initial step to implementing pharmacogenomics in clinical practice where physicians could use a patient's genetic make-up to optimize statin therapy, regarding efficiency and myopathy risk.

Screening of copy number variants in the 22q11.2 region of congenital heart disease patients from the São Miguel Island, Azores, revealed the second patient with a triplication.

BACKGROUND: The rearrangements in the 22q11.2 chromosomal region, responsible for the 22q11.2 deletion and microduplication syndromes, are frequently associated with congenital heart disease (CHD). The present work aimed to identify the genetic basis of CHD in 87 patients from the São Miguel Island, Azores, through the detection of copy number variants (CNVs) in the 22q11.2 region. These structural variants were searched using multiplex ligation-dependent probe amplification (MLPA). In patients with CNVs, we additionally performed fluorescent in situ hybridization (FISH) for the assessment of the exact number of 22q11.2 copies among each chromosome, and array comparative genomic hybridization (array-CGH) for the determination of the exact length of CNVs. RESULTS: We found that four patients (4.6%; A to D) carried CNVs. Patients A and D, both affected with a ventricular septal defect, carried a de novo 2.5 Mb deletion of the 22q11.2 region, which was probably originated by inter-chromosomal (inter-chromatid) non-allelic homologous recombination (NAHR) events in the regions containing low-copy repeats (LCRs). Patient C, with an atrial septal defect, carried a de novo 2.5 Mb duplication of 22q11.2 region, which could have been probably generated during gametogenesis by NAHR or by unequal crossing-over; additionally, this patient presented a benign 288 Kb duplication, which included the TOP3B gene inherited from her healthy mother. Finally, patient B showed a 3 Mb triplication associated with dysmorphic facial features, cognitive deficit and heart defects, a clinical feature not reported in the only case described so far in the literature. The evaluation of patient B's parents revealed a 2.5 Mb duplication in her father, suggesting a paternal inheritance with an extra copy. CONCLUSIONS: This report allowed the identification of rare deletion and microduplication syndromes in Azorean CHD patients. Moreover, we report the second patient with a 22q11.2 triplication, and we suggest that patients with triplications of chromosome 22q11.2, although they share some characteristic features with the deletion and microduplication syndromes, present a more severe phenotype probably due to the major dosage of implicated genes.

Human leptospirosis: seroreactivity and genetic susceptibility in the population of São Miguel Island (Azores, Portugal).

BACKGROUND: Leptospirosis is a worldwide zoonotic and recognized neglected infectious disease. It has been observed that only a proportion of individuals exposed to pathogenic species of Leptospira become infected and develop clinically evident disease. Moreover, little information is available in subsequent reinfections. In the present study, we determine if a first infection with leptospirosis protects against subsequent reinfection, and investigate which of the host genetic factors are involved in the susceptibility and resistance to leptospirosis. METHODOLOGY AND FINDINGS: We conducted, in 2011, a retrospective hospital-based case-control study in the São Miguel Island population (Azores archipelago). In order to determine the seropositivity against pathogenic Leptospira after the first episode of leptospirosis, we performed a serological evaluation in 97 unrelated participants diagnosed with leptospirosis between 1992 and 2011. The results revealed that 46.4% of the 97 participants have circulating anti-Leptospira antibodies, and from these participants 35.6% maintained the seroprevalence for the same serogroup. Moreover, three of them were reinfected with unrelated Leptospira serovars. The genetic study was carried out by adding a control group composed of 470 unrelated healthy blood donors, also from São Miguel Island. Twenty five SNPs among twelve innate immune genes - IL1α, IL1ß, IL6, IL10, IL12RB1, TLR2, TLR4, TLR9, CD14, CISH, LTA and TNF - were genotyped, as well as HLA class I (-A and -B) genes. Association analysis indicates that genotypes -511GG (OR=1.6, 95%CI 1.01-2.56, p=0.04) in IL1ß, +1196CG (OR=2.0, 95%CI 1.26-3.27, p=0.003) in IL12RB1, -292TA (OR=1.8, 95% CI 1.06-2.1, p=0.03) and +3415CG (OR=1.8, 95% CI 1.08-3.08, p=0.02), both in CISH confer susceptibility to pathogenic Leptospira. CONCLUSION: The present study suggests some degree of long-term protection against leptospires with an attenuation of symptoms in case of reinfection. Moreover, our data supports the genetic influence of IL1ß, IL12RB1 and CISH genes and the susceptibility to leptospirosis infection.

Genetic risk assessment for cardiovascular disease in Azoreans (Portugal): a general population-based study.

The identification of clinically validated genetic variants contributing to complex disorders raise the possibility to investigate individuals' risk. In this line of research, the present work aimed to assess the genetic risk for cardiovascular disease (CVD) in Azoreans. Genotyping of 19 SNPs - 9 on 9p21, 5 on LDLR and 5 on USF1 - was performed by TaqMan assays on 170 healthy Azorean individuals. Results demonstrate that the most frequent haplotype in 9p21, with a frequency of 41.4%, is TGGGCGCGC, which harbors all risk alleles. Considering haplotype homozygosity data show that females present higher value of homozygosity for both LDLR (13.5%) and USF1 (15.3%), whereas males present higher value for the 9p21 region (8.2%). Interestingly, genetic profile analysis revealed differences in terms of geographic and gender distribution. The Azorean Central group presented a higher risk for atherosclerosis, 2.7 times higher when compared to the Eastern group, while the Eastern group shows 1.5 times higher risk for dyslipidemias. Moreover, Azorean females demonstrated a 4 times higher risk for dyslipidemias when compared to males, whereas males have an increased risk for atherosclerosis. Although allele frequencies in Azoreans were similar to those reported for the HapMap CEU population, the differences in terms of haplotype and genetic profile distribution must be taken in consideration when assessing genetic risk. Taken together, the data here presented evidence for the need to perform biomedical research and epidemiologic analysis in Azoreans with the aim of developing strategies to CVD prevention, health promotion and population education.

HLA Class I and II profiles in São Miguel Island (Azores): genetic diversity and linkage disequilibrium.

BACKGROUND: Human leukocyte antigen (HLA) genes are characterized by high levels of polymorphism and linkage disequilibrium (LD), important characteristics to study the genetic background of human populations and their genetic structure. Here, we analyse the allele distribution and LD extent of HLA class I and II in São Miguel Island population (Azores archipelago, Portugal). FINDINGS: The sample set was composed of 106 healthy blood donors living in São Miguel Island obtained from the anonymized Azorean DNA bank. HLA class I (-A, -B and -Cw) and class II (-DRB1, -DQB1, -DPA1 and -DPB1) genotyping was performed by PCR-SSP Olerup SSP (GenoVision Inc.), according to the manufacturer's instructions.Genetic diversity values, based on the 7 loci, ranged from 0.821 both for HLA-DPA1 and -DQB1 to 0.934 for HLA-B, with a mean value of 0.846. Analysis of 5 HLA-A-Cw-B-DRB1-DQB1 haplotypes revealed that A*01-Cw*07-B*08-DRB1*03-DQB1*02 is the most frequent in São Miguel (7.9%) followed by A*24-B*08-Cw*07-DRB1*03-DQB1*02 (3.8%). In addition, even though the reports of high LD for HLA markers in worldwide populations, São Miguel islanders do not have extensive LD (average D' = 0.285). CONCLUSIONS: In summary, the results demonstrate high variability of HLA in São Miguel Island population as well as absence of genetic structure and extensive LD. The data here presented suggest that in São Miguel islanders autoimmune diseases studies will necessarily encompass a more focused analysis of HLA extended haplotypes as well as the evaluation of other non-HLA candidate genes.

UGT1A1, UGT1A6 and UGT1A7 genetic analysis: repercussion for irinotecan pharmacogenetics in the São Miguel Island Population (Azores, Portugal).

BACKGROUND: Glucuronidation reactions, catalyzed by uridine-diphosphate glucuronosyltransferase (UGT) enzymes, constitute a detoxification process that adds glucuronic acid to endogenous and exogenous compounds, aiding their excretion. UGT1A proteins have been implicated as risk factors for both the development of cancer and adverse drug effects. METHODS: Here, we assess the genome of 469 individuals from São Miguel Island (Azores, Portugal) in order to determine the frequencies of polymorphisms and haplotypes in UGT1A1, UGT1A6, and UGT1A7, the co-occurrence of reduced enzyme activity UGT1A variants related to irinotecan toxicity, and to calculate the extent of linkage disequilibrium (LD) in the genomic region encompassing these genes. RESULTS: Allelic analysis disclosed the presence of rare alleles - UGT1A1*36 and UGT1A1*37--only found in individuals of African descent, and UGT1A7*4. These alleles confirm our previous results on the São Miguel Island genetic background. We identified five different genotypes in UGT1A1 and UGT1A6 and nine in UGT1A7. Haplotype analysis showed that three haplotypes constituted approximately 80% of the allelic variants. Interestingly, haplotype 3 (UGT1A1*28-UGT1A6*2-UGT1A7*3), with a frequency of 0.235, gathers the three alleles encoding the low-function UGT isoforms. Additionally, LD indicates a strong interaction between functional polymorphisms related to the alteration of the UGT enzyme activity. CONCLUSIONS: In summary, the results demonstrate a high variability of alleles and haplotypes, which have important roles in modifying expression and activity of UGTs. The data presented here could improve the understanding of the predisposition to cancers and susceptibility to the adverse effects of irinotecan in the São Miguel Island population.

Thrombotic genetic risk factors and warfarin pharmacogenetic variants in São Miguel's healthy population (Azores).

BACKGROUND: The Azorean population presents the highest standardized mortality rate for cardiovascular diseases (CVD) when compared to mainland Portugal and other populations. Since thrombosis is a common cause of CVD, we assessed four polymorphisms in three thrombotic risk genes - F5 (G1691A), F2 (G20210A) and MTHFR (C677T, A1298C), in 469 healthy blood donors from São Miguel Island (Azores). We also analysed the CYP2C9 (C430T, A1075C) and VKORC1 (G1639A) variants in fifty-eight individuals with predisposition to thrombosis (possessing at least one variation in F5 or F2 genes and one in MTHFR) to evaluate their warfarin drug response genetic profiles. RESULTS: Among the 469 individuals, the data showed that thrombotic risk allele frequencies - 1691A (4.9%), 20210A (1.8%), 677T (41.7%) and 1298C (24.8%) - were similar to other Caucasians, but significantly different from mainland Portuguese (chi2, p < 0.001). The combined analysis of these variants identified twenty-two different genetic profiles (genotype order: F5, F2, MTHFR C677T and A1298C). Complete homozygosity for all wild-type alleles (GG GG CC AA) was present in 11.7%, being GG GG CT AA (22.4%) the most frequent profile. The results also demonstrated that 12.4% (58 out of 469) of São Miguel islanders have increased genetic predisposition to thrombosis. Subsequently, we evaluated these individuals for their warfarin response genetic profiles. The data showed that seven out of fifty-eight individuals are poor metabolizers (two with CYP2C9*2/*2 and five with CYP2C9*2/*3 genotypes). VKORC1 polymorphism analysis identified twelve individuals (20.7%) with AA genotype, who probably will require lower doses of warfarin. The joint analysis of CYP2C9 and VKORC1 revealed that 79.3% (46 out of 58) of the individuals carry at least one polymorphism in these genes. Within these, twenty-five individuals (43.1%) need intermediate and/or low doses of warfarin, if treatment is started. CONCLUSION: The present study demonstrated, for the first time, that São Miguel, and possibly the Azores population, shows significant differences on allele frequencies of thrombotic risk factors when compared to mainland Portugal. This research constitutes a primary approach for future studies on CVD, as well as for the implementation of warfarin dosing protocols using the patient's genotypic information.

Linkage disequilibrium and diversity for three genomic regions in Azoreans and mainland Portuguese.

Studies on linkage disequilibrium (LD) across the genome and populations have been used in recent years with the main objective of improving gene mapping of complex traits. Here, we characterize the patterns of genetic diversity of HLA loci and evaluate LD (D') extent in three genomic regions: Xq13.3, NRY and HLA. In addition, we examine the distribution of DXS1225-DXS8082 haplotype diversity in Azoreans and mainland Portuguese. Allele distribution has demonstrated that the São Miguel population is genetically very diverse; haplotype analysis revealed 100% discriminatory power for X- and Y-markers and 94.3% for HLA markers. Standardized multiallelic D' in these three genomic regions shows values lower than 0.33, thereby suggesting there is no extensive LD in the São Miguel population. Data regarding the distribution of DXS1225-DXS8082 haplotypes indicate that there are no significant differences among all the populations studied, (Azorean geographical groups, the Azores archipelago and mainland Portugal). Moreover, in these as well as in other European populations, the most frequent DXS1225-DXS8082 haplotype is 210-219. Even though São Miguel islanders and Azoreans do not constitute isolated populations and show LD for only very short physical distances, certain characteristics, such as the absence of genetic structure, the same environment and the possibility of constructing extensive pedigrees through church and civil records, offer an opportunity for dissecting the genetic background of complex diseases in these populations.

Linkage disequilibrium and diversity for three genomic regions in Azoreans and mainland Portuguese

Studies on linkage disequilibrium (LD) across the genome and populations have been used in recent years with the main objective of improving gene mapping of complex traits. Here, we characterize the patterns of genetic diversity of HLA loci and evaluate LD (D') extent in three genomic regions: Xq13.3, NRY and HLA. In addition, we examine the distribution of DXS1225-DXS8082 haplotype diversity in Azoreans and mainland Portuguese. Allele distribution has demonstrated that the São Miguel population is genetically very diverse; haplotype analysis revealed 100 percent discriminatory power for X- and Y-markers and 94.3 percent for HLA markers. Standardized multiallelic D' in these three genomic regions shows values lower than 0.33, thereby suggesting there is no extensive LD in the São Miguel population. Data regarding the distribution of DXS1225-DXS8082 haplotypes indicate that there are no significant differences among all the populations studied, (Azorean geographical groups, the Azores archipelago and mainland Portugal). Moreover, in these as well as in other European populations, the most frequent DXS1225-DXS8082 haplotype is 210-219. Even though São Miguel islanders and Azoreans do not constitute isolated populations and show LD for only very short physical distances, certain characteristics, such as the absence of genetic structure, the same environment and the possibility of constructing extensive pedigrees through church and civil records, offer an opportunity for dissecting the genetic background of complex diseases in these populations.

Evaluation of linkage disequilibrium on the Xq13.3 region: comparison between the Azores islands and mainland Portugal.

The design of genetic studies of complex diseases is dependent on the extent and distribution of linkage disequilibrium (LD) across the genome in different populations. Here, we characterize the extent of LD in the Azores (Western, Central, and Eastern island groups) and mainland Portugal populations. LD was evaluated in the Xq13.3 region by genotyping eight STR markers spanning 20.9 Mb. Standardized multiallelic disequilibrium coefficient (D') analysis indicates that the Western group presents higher values when compared with the Central and Eastern groups. However, all island groups show values of D' lower than 0.5 and 0.33, suggesting no extensive LD in these populations. Taken together, the data show that the Azorean population presents a lower D' (0.142) than mainland Portugal (0.226). Although, both populations do not show extensive LD, the easy reconstruction of large pedigrees in the Azorean population is a valuable resource for the fine mapping of disease genes.

Azores Islands: genetic origin, gene flow and diversity pattern.

BACKGROUND: The Azores are an archipelago located in the North Atlantic Ocean (parallel 38) composed of nine islands, dispersed over three geographical groups: The Eastern group (São Miguel and Santa Maria), the Central group (Terceira, Graciosa, Pico, São Jorge and Faial) and the Western group (Flores and Corvo). Taking into consideration the geographical and settlement history differences of the archipelago, the genetic diversity pattern and the internal migration of the Azorean population were assessed, based on the analysis of 15 STR loci in 592 unrelated individuals. RESULTS: The results of this evaluation reveal that Terceira displays the highest value of gene diversity (0.7979) and Corvo the lowest (0.7717). Gene flow analysis indicates that Corvo has the lowest value for migration, 23.35, where as São Miguel and Terceira have the highest values for emigration, 108.14 and 87.66, respectively. Taken together, the data demonstrate that, despite settlement diversity, no genetic difference between the populations of the nine islands is observable today. This may be explained by internal migration. CONCLUSION: Overall, the Azorean population can be analysed as a homogeneous genetic group, which consequently, would present, possibly, the same drug-reaction profile. In terms of genomic medicine, these results will have a significant impact on the design of future genetic and pharmacogenomic studies in the Azorean population.

Genetic signature of the São Miguel Island population (Azores) assessed by 21 microsatellite loci.

To study the genetic diversity of São Miguel's population we compared 21 microsatellite loci in 204 individuals from São Miguel island and 103 individuals from mainland Portugal. The results show that São Miguel and mainland Portugal populations have an average gene diversity of 0.767 and 0.765, respectively. Allele frequencies of all markers are comparable to other European populations. This observation is corroborated by the genetic relationships analysis based on the NJ tree and principal component, where São Miguel is closely related to mainland Portugal. Overall, the data suggests that São Miguel does not show population structure and is outbred with high genetic diversity. Moreover, the characterization here described is crucial to predict and explain genotypes implicated in genetic diseases in the Azorean population.

Assessment of Azorean ancestry by Alu insertion polymorphisms.

Knowledge of population ancestry from genetic markers is essential, for example, to understand the history of human migration and to carry out admixture and association studies. Here we assess the genome ancestry of the Azorean population through analysis of six Alu polymorphic sites (TPA-25, ACE, APO, B65, PV92, and D1) in 65 Azoreans and 30 Portuguese unrelated blood donors and compare data for the Y-chromosome and mtDNA. Allele frequencies were calculated by direct counting. Statistical analysis was performed using Arlequin 2.0. Nei's genetic distance was calculated with DISPAN software, and trees were constructed by neighbor joining (NJ) using PHYLIP 3.63. The results show that all Alu insertions were polymorphic. APO is the closest to fixation. The less frequent insertions are PV92 and D1 in the Azores and Portugal, respectively. ACE and TPA-25 show the highest values of heterozygosity in both populations. Allele frequencies are very similar to those obtained in European populations. These results are validated by the Y-chromosome and mtDNA data, where the majority of the maternal and paternal lineages are European. Overall, these data are reflected in the phylogenetic tree, in which the Azoreans and the Portuguese branch with Catalans, Andalusians, Moroccans, and Algerians. We conclude that the population of the Azores shows no significant genetic differences from that of mainland Portugal and that it is an outbred population. Moreover, the data validate the use of Alu insertion polymorphisms to assess the origin and history of human populations.