Regional heritability mapping method helps explain missing heritability of blood lipid traits in isolated populations.

Single single-nucleotide polymorphism (SNP) genome-wide association studies (SSGWAS) may fail to identify loci with modest effects on a trait. The recently developed regional heritability mapping (RHM) method can potentially identify such loci. In this study, RHM was compared with the SSGWAS for blood lipid traits (high-density lipoprotein (HDL), low-density lipoprotein (LDL), plasma concentrations of total cholesterol (TC) and triglycerides (TG)). Data comprised 2246 adults from isolated populations genotyped using ∼300 000 SNP arrays. The results were compared with large meta-analyses of these traits for validation. Using RHM, two significant regions affecting HDL on chromosomes 15 and 16 and one affecting LDL on chromosome 19 were identified. These regions covered the most significant SNPs associated with HDL and LDL from the meta-analysis. The chromosome 19 region was identified in our data despite the fact that the most significant SNP in the meta-analysis (or any SNP tagging it) was not genotyped in our SNP array. The SSGWAS identified one SNP associated with HDL on chromosome 16 (the top meta-analysis SNP) and one on chromosome 10 (not reported by RHM or in the meta-analysis and hence possibly a false positive association). The results further confirm that RHM can have better power than SSGWAS in detecting causal regions including regions containing crucial ungenotyped variants. This study suggests that RHM can be a useful tool to explain some of the 'missing heritability' of complex trait variation.

Application of high-dimensional feature selection: evaluation for genomic prediction in man.

In this study, we investigated the effect of five feature selection approaches on the performance of a mixed model (G-BLUP) and a Bayesian (Bayes C) prediction method. We predicted height, high density lipoprotein cholesterol (HDL) and body mass index (BMI) within 2,186 Croatian and into 810 UK individuals using genome-wide SNP data. Using all SNP information Bayes C and G-BLUP had similar predictive performance across all traits within the Croatian data, and for the highly polygenic traits height and BMI when predicting into the UK data. Bayes C outperformed G-BLUP in the prediction of HDL, which is influenced by loci of moderate size, in the UK data. Supervised feature selection of a SNP subset in the G-BLUP framework provided a flexible, generalisable and computationally efficient alternative to Bayes C; but careful evaluation of predictive performance is required when supervised feature selection has been used.

A K(ATP) channel gene effect on sleep duration: from genome-wide association studies to function in Drosophila.

Humans sleep approximately a third of their lifetime. The observation that individuals with either long or short sleep duration show associations with metabolic syndrome and psychiatric disorders suggests that the length of sleep is adaptive. Although sleep duration can be influenced by photoperiod (season) and phase of entrainment (chronotype), human familial sleep disorders indicate that there is a strong genetic modulation of sleep. Therefore, we conducted high-density genome-wide association studies for sleep duration in seven European populations (N=4251). We identified an intronic variant (rs11046205; P=3.99 × 10(-8)) in the ABCC9 gene that explains ≈5% of the variation in sleep duration. An influence of season and chronotype on sleep duration was solely observed in the replication sample (N=5949). Meta-analysis of the associations found in a subgroup of the replication sample, chosen for season of entry and chronotype, together with the discovery results showed genome-wide significance. RNA interference knockdown experiments of the conserved ABCC9 homologue in Drosophila neurons renders flies sleepless during the first 3 h of the night. ABCC9 encodes an ATP-sensitive potassium channel subunit (SUR2), serving as a sensor of intracellular energy metabolism.

Global health research priorities: mobilizing the developing world.

The paper focuses on two questions: (i) how to set research priorities in a transparent, systematic, fair and legitimate way?; and (ii) how to mobilize low and middle-income countries to take more ownership in defining their own research policies, rather than merely being passive recipients of international aid for research and development? I propose that the recently developed Child Health and Nutrition Research initiative (CHNRI) methodology is becoming widely accepted as a feasible answer to both those questions. In this paper, I review its numerous applications to date and show how it evolved into a practical and systematic tool that can assist priority setting in health research investments in diverse contexts. The CHNRI methodology also addresses support for different instruments of health research to achieve better balance between fundamental research, translation research and implementation research. The wide application of CHNRI methodology is expected to maximise the potential of health research to reduce disease burden and gradually reduce inequities that exist between support for research on the health problems of the rich and the poor. I believe that this tool will find application within many low and middle-income countries and assist them to pull together their own experts and actively define their priorities for research and development in the coming years.

Leprosy epidemics during history increased protective allele frequency of PARK2/PACRG genes in the population of the Mljet Island, Croatia.

INTRODUCTION: Two regulatory polymorphisms (rs1040079 and rs9356058) shared by PARK2 and PACRG genes were identified as major risk variants for leprosy susceptibility. The aim of this study was to investigate if allele frequencies of these polymorphisms in the isolated population of the island of Mljet, which served as a quarantine for leprosy patients during past centuries, were different to allele frequencies in two control populations with no history of leprosy. SUBJECTS AND METHODS: This study included 88 unrelated Caucasian individuals from the island of Mljet while two control groups included 93 individuals from the island of Rab and 160 individuals from the region of Split. Genotyping for rs1040079 and rs9356058 was performed by "real-time" PCR analysis. We also compared the allele frequency of the rs9356058 polymorphism from the population of Mljet with allele frequencies derived from the existing genome wide association scans in two additional island populations, Vis (924 subjects) and Korcula (909 subjects). RESULTS: We found a significant increase in the frequency of rs9356058 allele C in the population of Mljet when compared to both control groups. We also observed a significant increase in the frequency of rs1040079 allele A in the population of Mljet when compared with the population of Rab, however this increase was not significant when compared with the population of Split. Allele frequencies of both examined polymorphisms did not differ between the two control populations. Protective haplotype rs9356058-rs1040079 CA was also more frequent in the population of Mljet compared with the Rab and Split populations. In addition, an increase of frequency of rs9356058 allele C was also observed in the population of Mljet when compared with the frequency in the Korcula population. CONCLUSION: The results of our study show the association of polymorphisms rs9356058 and rs1040079 in gene PARK2/PACRG with leprosy. The results of our study indicate that exposure to leprosy and mortality in the population caused by leprosy on Mljet resulted in the selection of rs9356058 "protective" C allele in the PARK2 gene, while this was not observed in the two control groups. This is the first study to assess the genetic susceptibility to leprosy in a European population.

Comparison of the growth hormone, IGF-1 and insulin in cerebrospinal fluid and serum between patients with motor neuron disease and healthy controls.

Neurotrophic effects of the growth hormone (GH), insulin-like growth factor-1 (IGF-1) and insulin on the central nervous system have become more apparent in the past decade. In this study, we measured serum and cerebrospinal fluid (CSF) concentrations of GH, IGF-1 and insulin in 35 patients with motor neuron disease (MND) [24 patients with definite amyotrophic lateral sclerosis (ALS) and 11 patients with progressive bulbar palsy] and in 40 healthy controls. Levels of serum concentrations of GH and IGF-1 did not significantly differ between the MND patient group and the healthy controls, while the level of insulin was significantly decreased (P = 0.0033) in the MND patient group. However, levels of all three examined parameters in CSF were significantly lower in the MND group than in the healthy controls with the statistical significance for IGF-1 and insulin of P < 0.001. This finding has not been reported previously, and further investigations into its association with ALS should establish whether it can be used as an early marker of the disease, or whether it merely represents a consequence of ALS development.

Estimating human inbreeding coefficients: comparison of genealogical and marker heterozygosity approaches.

We have used genealogies and genomic polymorphisms to estimate individual inbreeding coefficients (F) in 50 subjects with an expected range (based on recent genealogies) of F from 0.0 to 0.0625. The estimates were based on two approaches, using genotypes respectively from 410 microsatellite markers (410-STR panel) and from 10,000 SNPs (10K-SNP panel). The latter was performed in a sub-sample of 15 individuals. We concluded that for both marker panels measures of inbreeding based on the excess of homozygosity over Hardy-Weinberg expectation were not closely correlated with 4-5 generation genealogical F-values. For the 10K-SNP panel we found two alternative measures which correlated more closely with F, based respectively on standard errors and on paired homozygosity of nearby SNPs over distances of 2-4 cM. We propose an empirical method for estimating standard errors and hence individual F-values, based on the variation between individual autosomes. This method could provide useful estimates of average F-values for groups of individuals in population-based studies of the effects of inbreeding/homozygosity on quantitative traits.

Sex ratio at birth and war in Croatia (1991-1995).

BACKGROUND: We have investigated sex ratio at birth (expressed as the proportion of males) in Croatia before, during and after the war (1991-1995). METHODS: Data for each of 21 counties in Croatia (861 516 births) were collected and pooled into two groups: the first, consisting of the counties unaffected by the war, and the second, comprising the counties affected by war events. Odds ratios of being born as a male were calculated, with being born in a county exposed to war defined as the risk factor. RESULTS: No significant deviations from the expected ratio of 0.514 were found in pre-war, wartime or post-war period at the national level. The ratio was 0.515 during the pre-war and wartime periods, and 0.514 in the post-war period. Comparison of the ratios in the three periods in both affected and unaffected counties revealed no significant increase in the sex ratio. The only significant increase in the sex ratio was registered in two counties unaffected by the warfare. CONCLUSIONS: This study indicates that warfare did not cause a detectable increase of the sex ratio at birth in Croatia, in contrast to what might have been predicted based on earlier reports in the literature.

Haematological malignancies in childhood in Croatia: investigating the theories of depleted uranium, chemical plant damage and 'population mixing'.

Some of potential causes proposed to explain the reported increase of haematological malignancies in childhood during or after the war period in several countries include depleted uranium, chemical pollution and population mixing theory. The aim of this study was to define the population of Croatian children aged 0-14 years who were potentially exposed to each of those risks during the war and to investigate any possible association between the exposure and the incidence of haematological malignancies. The authors analyzed the data reported by the Cancer Registry of Croatia during the pre-war period (1986-1990), war period (1991-1995) and post-war period (1996-1999). In the group of 10 counties potentially exposed to depleted uranium and two counties where chemical war damage occurred, no significant difference in incidence of the studied haematological malignancies was noted in comparison to pre-war period. The incidence of lymphatic leukaemia significantly increased in four counties where population mixing had occurred during the war period, supporting the 'mixing theory'. In those counties, the incidence of Hodgkin's lymphoma decreased during and after the war. In Croatia as a whole, decreases in incidence of myeloid leukaemias during war and non-Hodgkin lymphoma after the war were noted.

Effect of inbreeding and endogamy on occlusal traits in human isolates.

OBJECTIVE: To discuss the genetic basis of occlusal traits through analysis of the effects of inbreeding in a subdivided isolated community. SUBJECTS AND METHODS: The sample comprised dental casts of 224 children, aged 7-14 years, from 15 villages of the Island of Hvar, Croatia. MAIN OUTCOME MEASURES: Studied traits were Angle class, overjet, vertical bite, overbite, and crowding/spacing. DESIGN: Children with complete grandparental endogamy (all four grandparents born in the village of residence of the examinee) were compared to children with incomplete grandparental endogamy. In addition, children resident in the group of villages with a high prevalence of inbreeding were compared to children resident in the groups of villages with moderate and low prevalence of inbreeding. RESULTS: In both designs, inbreeding seemed to increase the mean values of overjet, overbite, and vertical bite, while it had little or no effect on crowding/spacing. Angle classes were correlated to inbreeding at the individual level, but this was not supported at the population level. The effects were stronger in the subsample with bilaterally concordant Angle classes. CONCLUSION: The observed inbreeding effects imply that the genetic basis of some occlusal traits is polygenic and, in considerable part, influenced by rare and recessive genetic variants.

Inbreeding and nephrolithiasis in Croatian island isolates.

The aim of this study was to investigate a recessive genetic component in susceptibility to nephrolithiasis (NL) by comparing its prevalence in highly inbred, moderately inbred and non-inbred villages of three Croatian islands: Brac, Hvar and Korcula. The average inbreeding coefficient of each village population (F) was estimated in a random sample of 20-30% adults from 14 villages using Wright's path method (based on genealogical information), isonymy data and average deviation from Hardy-Weinberg expectations for MN, Ss and Kk serogenetic polymorphisms. The six villages with the greatest genealogical F value (0.025-0.049; current total population N=1,401), the four with intermediate value (0.012-0.015; N=998) and the four with the smallest value (0.002-0.008; N=1,500) were chosen for this study. Medical records of entire populations were reviewed and the diagnosis of NL was established according to unified criteria, based on the information from the specialists in general hospitals and on the agreement in diagnosis between the investigators and local general practitioners. The variance in environmental and socio-cultural factors between villages was shown to be minimal. Prevalence of NL in each village was standardised by sex and age to the total population of all 14 villages. The standardized prevalence of NL was 1.5% in the group of villages with low F, 2.3% in the group with moderate F (p<0.10), and 5.4% in the group with high F (p<0.001). The correlation factor between F values by villages and the standardized prevalence of NL was 0.45 (p<0.05). The study showed that a large number of predominantly recessive genetic factors might mediate the susceptibility to formation of renal stones in humans.

Mal de Meleda: genetic haplotype analysis and clinicopathological findings in cases originating from the island of Mljet (Meleda), Croatia.

BACKGROUND: Mal de Meleda (keratoderma palmoplantaris transgrediens) is an autosomal recessive disorder, first described on the island of Mljet (Meleda), Croatia. The candidate region for the gene responsible for this disorder was found on the chromosome 8qter, and the responsible mutations have recently been identified in 12 Algerian and 7 Croatian families. OBJECTIVES: To fully characterize all 12 living cases originating from the original setting of the disease, the island of Mljet, in the light of new findings and using modern diagnostic technology. PATIENTS AND METHODS: Twelve patients and 37 family members were identified over the period 1998-1999, interviewed and examined. RESULTS: The reconstruction of 8 genealogies suggests a common ancestry of all cases but one. The clinical presentation and pathologic findings of these cases are described in detail and are consistent with previous reports. Symptoms and signs were found to be milder in non-manual workers who had applied continuous symptomatic treatment. Blood samples were taken from 8 cases and 16 close relatives for genetic studies. These confirmed a shared haplotype in all cases, but in none of 17 unaffected control individuals, near the marker D8S1751 on chromosome 8. CONCLUSIONS: This review characterizes mal de Meleda in its original setting and shows that the sporadic cases found in the regions of medieval trade routes of the Republic of Dubrovnik (Middle East and Northern Africa) carry the same mutation as the patients from Mljet Island, Croatia.

Inbreeding and learning disability in Croatian island isolates.

The aim of this study was to investigate the prevalence of learning disability (LD) in isolate populations with different inbreeding coefficients (F). Prevalence of LD and F were determined in 10 villages from five Croatian islands: Brac, Hvar, Korcula, Lastovo and Susak. For the purpose of this study, LD was defined as the inability to attend the public school system. As the elementary schools (grade 1-8) in the place of the study are both public and compulsory, the assessment of child's inability to attend the school is performed at the age of six. This is required by all children in the country based on standard set of tests of cognitive performance defined by the Ministry of Education and Culture of the Republic of Croatia. The average inbreeding coefficients in each village population (F) were estimated in a random sample of 20-30% adults in each of the 10 villages based on 4 ancestral generations and using Wright's path method. Prevalence of LD ranged from 0.43% to 2.47%, and the inbreeding coefficients ranged from 0.8% to 4.9%. The Pearson's correlation coefficient between F and LD prevalence was 0.80 (p < 0.01). Although the relative risk per 5% inbreeding appeared very high (about 10), the absolute risk only increased from 0.18% to 1.77%. The genetic effect of inbreeding (GEI) was approximately 0.69% and the population-attributable fraction 76.6%. A review of the literature and the results of this study lead to a conclusion that a very large number of predominantly recessive genetic factors might mediate the genetic susceptibility to various forms of LD in these populations.

The evidence of mtDNA haplogroup F in a European population and its ethnohistoric implications.

Mitochondrial DNA polymorphism was analysed in a sample of 108 Croatians from the Adriatic Island isolate of Hvar. Besides typically European varieties of human maternal lineages, haplogroup F was found in a considerable frequency (8.3%). This haplogroup is most frequent in southeast Asia but has not been reported before in Europe. The genealogical analysis of haplogroup F cases from Hvar suggested founder effect. Subsequent field work was undertaken to sample and analyse 336 persons from three neighbouring islands (Brac, Korcula and Krk) and 379 more persons from all Croatian mainland counties and to determine if haplogroup F is present in the general population. Only one more case was found in one of the mainland cities, with no known ancestors from Hvar Island. The first published phylogenetic analysis of haplogroup F worldwide is presented, applying the median network method, suggesting several scenarios how this maternal lineage may have been added to the Croatian mtDNA pool.

Endogamy and variation in blood pressure levels in Croatian island isolates.

Blood pressure variation was investigated among populations inhabiting islands and peninsula of Middle Dalmatia, Croatia. The number of previous anthropological studies pointed to isolation and different genetic population structure in this environmentally fairly homogeneous area. Variation in blood pressure (systolic and diastolic) among the populations of the islands of Brac, Hvar, Korcula, and the Peljesac peninsula was assessed at three levels involving village populations, regional (western and eastern) populations and the entire island populations. The blood pressure data were collected from 3834 adult individuals inhabiting 37 rural communities and were adjusted for age and body mass index. Variation in blood pressure levels existed among regions and villages. Due to the history of differential settlement, small village sizes and high levels of reproductive isolation, the observed blood pressure variation could be attributed to founder effect, genetic drift and inbreeding. The involvement of genetic factors was tested by relating blood pressure variation among villages to degree of isolation among them. Blood pressure means and proportions of hypertensives increased with endogamy levels in males. In females, this effect could not be observed. However, in both sexes the highest proportions of hypertensives (more than 40%) were found in villages that are most reproductively closed (endogamy greater than 80%). These populations are considered particularly promising for further genetic epidemiological research.

Ancestral kinship and cancer in Lastovo Island, Croatia.

The aim of this study was to compute the average kinship coefficient of cancer cases in an extreme isolate (Lastovo Island, Croatia) and to compare it to the corresponding value for the island's unaffected population. Kinship estimates were obtained by Wright's path method (phi) based on pedigree reconstruction and Lasker's coefficient of relationship by isonymy (Ri) based on surname distribution. A total of 76 cancer cases were recorded on the island between 1971 and 1995. The unaffected control population of 1144 persons was obtained from the population census in 1971. The data on five ancestral generations were collected from parish registries preserved in the local church, and included 5484 persons born on the island between 1750 and 1970. Both Wright's path method and Lasker's isonymy method indicated significantly greater kinship of female cancer cases than of the unaffected population. Cancer cases with increased values of Ri among them and Ri among their ancestors included cancers of the ovary, colon, brain, breast, and prostate and leukemias. This study confirms the role of heredity and common ancestry in the development of cancer, providing a rough insight into their relative importance for specific cancer sites.