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Background: The CFTR gene (Cystic Fibrosis conductance Transmembrane Regulator) is the gene responsible for Cystic Fibrosis, the most common severe autosomal recessive disease in Europeans. It has been extensively explored in several European and European-derived populations, but poorly studied in the other major human groups., Aim: To characterize the variability of the CFTR gene in an African population., Subjects and Methods: Using DGGE, all 27 exons (4443 bp) and 2184 bp of the flanking intronic regions of the CFTR gene were studied in a random sample of 45 Mossì from Burkina Faso (Western sub-Saharan Africa)., Results: Sixteen variable sites were found: 13 SNPs (one in the promoter region, four non-synonymous and five synonymous in the exons and three in the introns) and three intronic STRs. Only the promoter site ( - 94 G/T), slightly polymorphic in the present survey, was not variable in different European populations. Comparison between Western Africans, Eastern Africans, Europeans and Eastern Asians showed that alleles at two intronic STRs (T(n) and (TG)(m) in intron 8), four exonic (M470V, 2694 T/G, 4002 A/G and 4521 G/A) and one intronic (875+40 A/G) SNPs have very different frequencies among at least two major human groups. Moreover, the overall degree of non-synonymous variability in Mossì is much lower than that in Europeans. A possible interpretation of this finding is proposed., Conclusions: The CFTR gene has been since long hypothesized to have undergone selection in Europeans. The present study by comparing Africans and Europeans for the overall variability of the gene supports this hypothesis.

Background: Central Asia and the Indian subcontinent represent an area considered as a source and a reservoir for human genetic diversity, with many markers taking root here, most of which are the ancestral state of eastern and western haplogroups, while others are local. Between these two regions, Terai (Nepal) is a pivotal passageway allowing, in different times, multiple population interactions, although because of its highly malarial environment, it was scarcely inhabited until a few decades ago, when malaria was eradicated. One of the oldest and the largest indigenous people of Terai is represented by the malaria resistant Tharus, whose gene pool could still retain traces of ancient complex interactions. Until now, however, investigations on their genetic structure have been scarce mainly identifying East Asian signatures., Results: High-resolution analyses of mitochondrial-DNA (including 34 complete sequences) and Y-chromosome (67 SNPs and 12 STRs) variations carried out in 173 Tharus (two groups from Central and one from Eastern Terai), and 104 Indians (Hindus from Terai and New Delhi and tribals from Andhra Pradesh) allowed the identification of three principal components: East Asian, West Eurasian and Indian, the last including both local and inter-regional sub-components, at least for the Y chromosome., Conclusion: Although remarkable quantitative and qualitative differences appear among the various population groups and also between sexes within the same group, many mitochondrial-DNA and Y-chromosome lineages are shared or derived from ancient Indian haplogroups, thus revealing a deep shared ancestry between Tharus and Indians. Interestingly, the local Y-chromosome Indian component observed in the Andhra-Pradesh tribals is present in all Tharu groups, whereas the inter-regional component strongly prevails in the two Hindu samples and other Nepalese populations.The complete sequencing of mtDNAs from unresolved haplogroups also provided informative markers that greatly improved the mtDNA phylogeny and allowed the identification of ancient relationships between Tharus and Malaysia, the Andaman Islands and Japan as well as between India and North and East Africa. Overall, this study gives a paradigmatic example of the importance of genetic isolates in revealing variants not easily detectable in the general population.

An average of about 1700 CFTR (cystic fibrosis transmembrane conductance regulator) alleles from normal individuals from different European populations were extensively screened for DNA sequence variation. A total of 80 variants were observed: 61 coding SNSs (results already published), 13 noncoding SNSs, three STRs, two short deletions, and one nucleotide insertion. Eight DNA variants were classified as non-CF causing due to their high frequency of occurrence. Through this survey the CFTR has become the most exhaustively studied gene for its coding sequence variability and, though to a lesser extent, for its noncoding sequence variability as well. Interestingly, most variation was associated with the M470 allele, while the V470 allele showed an 'extended haplotype homozygosity' (EHH). These findings make us suggest a role for selection acting either on the M470V itself or through an hitchhiking mechanism involving a second site. The possible ancient origin of the V allele in an 'out of Africa' time frame is discussed.

Coding single nucleotide substitutions (cSNSs) have been studied on hundreds of genes using small samples (n(g) approximately 100-150 genes). In the present investigation, a large random European population sample (average n(g) approximately 1500) was studied for a single gene, the CFTR (Cystic Fibrosis Transmembrane conductance Regulator). The nonsynonymous (NS) substitutions exhibited, in accordance with previous reports, a mean probability of being polymorphic (q > 0.005), much lower than that of the synonymous (S) substitutions, but they showed a similar rate of subpolymorphic (q < 0.005) variability. This indicates that, in autosomal genes that may have harmful recessive alleles (nonduplicated genes with important functions), genetic drift overwhelms selection in the subpolymorphic range of variability, making disadvantageous alleles behave as neutral. These results imply that the majority of the subpolymorphic nonsynonymous alleles of these genes are selectively negative or even pathogenic.

BACKGROUND: The thalassemic syndromes originate from mutations of the globin genes that cause, besides the characteristic clinical picture, also an increased Hb F amount. It is not yet clear if there are more factors, besides the beta globin genotype, determining the Hb F production. We have tried to find out if there are relations between total Hb and Hb F, between erythropoietin (Epo) and Hb F, between Hb F and point mutations of the gamma gene promoters. MATERIALS AND METHODS: Hematologic parameters, iron status, alpha/non-alpha globin ratio, Epo level, and thalassemic defects of the alpha-, beta-, and gamma-globin genes were explored using standard methods in patients affected by thalassemic diseases. Ninety-five non thalassemic individuals have been examined as controls. RESULTS: Two clinical variants of beta-thalassemia intermedia referred to as beta-thal int sub-silent and evident are associated with distinct sets of mutations of the beta-globin gene. Silent beta thal mutations are invariably associated with sub-silent beta thal int; beta degrees or severe beta+ thal mutations are associated with evident beta thal int (88%) and almost invariably (98%) with thalassemia major. A positive correlation was observed between the severity of the disease and the Hb F level, but no correlation was found between the Hb F and erythropoietin (Epo) level. The mutation Ggamma -158 C&RightArrow;T was detected in 26.9% of patients affected by beta-thal int sub-silent and evident, respectively, but only in 2% of patients with thalassemia major. CONCLUSIONS: The severity of beta-thal int and the increased Hb F level are strictly dependent from the type of beta-globin gene mutations. No relation is found between Hb F synthesis and Epo secretion. The mutation Ggamma -158 C&RightArrow;T, common among patients affected by beta-thal int and very rare in thal major patients, does not seem, in this study, to influence the Hb F content in beta thal int patients.

Haemoglobin C (HbC; beta6Glu --> Lys) is common in malarious areas of West Africa, especially in Burkina Faso. Conclusive evidence exists on the protective role against severe malaria of haemoglobin S (HbS; beta6Glu --> Val) heterozygosity, whereas conflicting results for the HbC trait have been reported and no epidemiological data exist on the possible role of the HbCC genotype. In vitro studies suggested that HbCC erythrocytes fail to support the growth of P. falciparum but HbC homozygotes with high P. falciparum parasitaemias have been observed. Here we show, in a large case-control study performed in Burkina Faso on 4,348 Mossi subjects, that HbC is associated with a 29% reduction in risk of clinical malaria in HbAC heterozygotes (P = 0.0008) and of 93% in HbCC homozygotes (P = 0.0011). These findings, together with the limited pathology of HbAC and HbCC compared to the severely disadvantaged HbSS and HbSC genotypes and the low betaS gene frequency in the geographic epicentre of betaC, support the hypothesis that, in the long term and in the absence of malaria control, HbC would replace HbS in central West Africa.

Background and Objectives: In non-thalassaemic women serum erythropoietin (Epo) level increases during pregnancy, whereas that of soluble transferrin receptor (STFR) drops slightly in the first two trimesters to attain the original values in the third trimester. In this study the time-course of these two parameters was explored in b-thalassemic and non-b-thalassemic women, both pregnant and not., Design and Methods: Two hundred and fifty-seven women were studied: 64 non-b-thalassemic, non-pregnant women made up the reference group, 89 were non-b-thalassemic pregnant women, and 104 were b-thalassemic pregnant or non-pregnant women. The full blood count, hemoglobin levels and iron status (serum iron and serum ferritin levels) were explored by traditional methods. Serum Epo and STFR levels were measured with standard commercial kits., Results: In non-b-thalassemic women the mean non-pregnant Epo level (10.95 +/-4. 7 mU/mL) increased in the first trimester (17.12 +/-5.18 mU/mL), was stationary in the second, and increased again in the third (31. 43+/-14.13 mU/mL). STFR mean value dropped in early pregnancy from 2. 40+/-0.72 mg/L to 1.78 +/- 0.64 mg/L, and then returned to the original value (2.38 +/- 0.94 mg/L). In b-thalassemic women the mean non-pregnant Epo level (15.00 +/-6.56 mU/mL) was higher than in non-thal non-pregnant women. During pregnancy it progressively increased to 35.60+/-25.46 mU/mL. STFR (non-pregnant level 3.37+/-1. 07 mg/L) gradually increased throughout the whole gestation period and by the third trimester its level was markedly higher than that of non-b-thal women at the corresponding stage of gestation (9. 41+/-5.39 mg/L vs 2.40+/-0.72 mg/L)., Interpretation and Conclusions: The STFR level changed to different extents in non-b-thal and b-thal women during their pregnancies. In the former STFR markedly decreased in early pregnancy; in the latter it showed no decrease in the first trimester, increased in the second and reached very high values in the third. This time course is likely to be the consequence of erythroid bone marrow hyperplasia and hyperactivity, which are usually present in all b-thalassemic patients and in heterozygous carriers as well.

Two polymorphic sites, -107 C-->T and -100 G-->C with respect to the cap site of the human beta pseudogene of the hemoglobin gene, are described. They have been studied in five European, one Indian, two Asian, and two sub-Saharan African populations. The -107 C-->T site turned out to be polymorphic in all five European populations and the Indian population (pooled q = 0.142 +/- 0.018) and in the two Asian populations (pooled q = 0.073 +/- 0.025), but it was monomorphic in the two sub-Saharan populations. On the contrary, the -100 G-->C site was polymorphic in the two sub-Saharan samples (q = 0.093 +/- 0.024), but the variant allele was not found in any of the European, Indian, or Asian samples. Thus this only 8-bp-long stretch of DNA is informative for estimating the extent of genetic admixture in sub-Saharan Africans.

Background: alpha thalassemias are very common in all thalassemic areas; however, complete knowledge of the phenotypic, genotypic and epidemiological features of these thalassemias has not yet been achieved for a number of reasons: the frequent absence of a thalassemic hematologic picture, the lack of a specific characteristic comparable to the Hb A2 increase for beta thalassemias, and the almost complete homology between the two alpha genes., Methods and Results: A new set of PCR techniques, each based on primer(s) specific for a particular type of alpha globin gene disorder, has been devised in our laboratory. The procedures are simple, and non-radioactive. They lead to the identification of all alpha globin disorders common in the Mediterranean area [-alpha 3.7, -alpha 4.2, alpha Hphl, alpha Ncol, --MED, -(alpha)20.5, alpha alpha alpha anti3.7]. The electrophoretic patterns specific for the main alpha globin alterations as observed with this set of techniques, are presented., Conclusions: Owing to their advantageous properties, these techniques are suitable for precise molecular characterization of the numerous subjects selected through mass population screenings.

The genetic structure of Rajasthan Hindus and Punjab-Haryana Hindus and Sikhs has been studied for ABO, RH, APOC2, C6, C7, F13A, F13B, HP, ORM1, ACP1, ADA, AK1, ESD, GLO1, PGD, PGM1 subtyping, and PGP. This is the first genetic survey on Hindus of Rajasthan. Furthermore, many of these markers have never been studied on Hindus before (APOC2, C6, C7, F13A, F13B, ORM1, PGP). These data, together with those previously available for Hindus, have been utilized to analyze the within-Hindus genetic heterogeneity by RST statistic and correspondence analysis. The genetic relationships of Hindus to other Causcasoid populations were also investigated. In the first analysis, two eastern states (Orissa and Andhra Pradesh) were found to be quite separate from each other and clearly distinct from the northwestern and western states. Out of the markers which could not be utilized in this analysis, PGM1 subtyping turned out to discriminate between the Dravidian-speaking and the Indo-Aryan-speaking Hindus. The second analysis shows a clear-cut separation of Hindus from Europeans, with Near Eastern and Middle Eastern populations genetically in an intermediate position.

Sardinians, a population with many distinct anthropogenetic features, has been studied for the COLIA1 and COLIA2 genes at the DNA level for two purposes: to look for new RFLPs (restriction fragment length polymorphisms) and to study the distribution of three known COLIA2 RFLPs (EcoRI, RsaI, MspI) at both the allele and the haplotype levels. None of the eleven enzyme-probe systems examined led to the discovery of a new polymorphism. The following frequency q was found for the less common allele of the three RFLPs: EcoRI, q(+) = 0.178 +/- 0.031; RsaI, q(-) = 0.316 +/- 0.038; MspI, q(-) = 0.046 +/- 0.017. EcoRI turned out to be the most discriminant of the three polymorphisms because the frequency of the (+) allele in Sardinians was about half that estimated for a large homogeneous white sample (0.18 +/- 0.03 vs. 0.30 +/- 0.01). So far as the haplotype level is concerned, the sample is made up of triplets (parents and child). Therefore all the haplotype frequencies and delta values (degrees of disequilibrium, D) were obtained by direct counting of the unambiguously identified haplotypes rather than being based on their maximum-likelihood estimates. This together with their analytical and detailed presentation makes these data comparable with future findings, provided that the two data sets are presented in a comparable way. At this level the three RFLPs are efficient in distinguishing Sardinians from Calabrians (southern Italy) but not from the central Italian population. The present results, besides adding a further discriminative criterion between Sardinians and Italians (and whites on the whole), identify the complex COLIA2 locus as a valuable anthropogenetic marker.

We searched for the East Asian mtDNA 9-bp deletion in the intergenic COII/tRNA(Lys) region in a sample of 107 Tharus (50 from central Terai and 57 from eastern Terai), a population whose anthropological origin has yet to be completely clarified. The deletion, detected by electrophoresis of the PCR-amplified nt 7392-8628 mtDNA fragment after digestion with HaeIII, was found in about 8% of both Tharu groups but was found in none of the 76 Hindus who were examined as a non-Oriental neighboring control population. A complete triplication of the 9-bp unit, the second case so far reported, was also observed in one eastern Tharu. All the mtDNAs with the deletion, and that with the triplication, were further characterized (by PCR amplification of the relevant mtDNA fragments and their digestion with the appropriate enzymes) to locate them in the Ballinger et al. phylogeny of East Asian mtDNA haplotypes. The deletion was found to be associated with four different haplotypes, two of which are reported for the first time. One of the deletions and especially the triplication could be best explained by the assumption of novel length-change events. Ballinger's classification of East Asian mtDNA haplotypes is mainly based on the phenotypes for the DdeI site at nt 10394 and the AluI site at nt 10397. Analysis of the entire Tharu sample revealed that more than 70% of the Tharus have both sites, the association of which has been suggested as an ancient East Asian peculiarity. These results conclusively indicate that the Tharus have a predominantly maternal Oriental ancestry. Moreover, they show at least one and perhaps two further distinct length mutations, and this suggests that the examined region is a hot spot of rearrangements.

About 80 Sephardim, 80 Ashkenazim and 100 Czechoslovaks were examined for the Y-specific RFLPs revealed by the probes p12f2 and p49a,f on TaqI DNA digests. The aim of the study was to investigate the origin of the Ashkenazi gene pool through the analysis of markers which, having an exclusively holoandric transmission, are useful to estimate paternal gene flow. The comparison of the two groups of Jews with each other and with Czechoslovaks (which have been taken as a representative source of foreign Y-chromosomes for Ashkenazim) shows a great similarity between Sephardim and Ashkenazim who are very different from Czechoslovaks. On the other hand both groups of Jews appear to be closely related to Lebanese. A preliminary evaluation suggests that the contribution of foreign males to the Ashkenazi gene pool has been very low (1% or less per generation).

Tharus--a population of Terai (a region with a severe malarial morbidity in the past)--can be subdivided into three main groups: Western, Central and Southern Tharus. They have usually been considered a Mongoloid population and this has been further substantiated by mtDNA findings on Central Tharus. Studies on the distribution of malaria-related genes have shown an extremely high frequency (0.8) of the alpha-thal gene among Western and Central Tharus. This frequency, however, unexpectedly turned out to be only 0.04 in a sample of Eastern Tharus. This raised doubts on the common notion that Tharus are a single anthropological entity. In the present investigation mtDNA markers were studied in the same sample of Eastern Tharus previously examined for the alpha-thal gene. The findings were: 1. the same three features which confirmed the classification of Central Tharus as Mongoloids (i.e., the common occurrence of HpaI-1/HincII-1 and HaeII-5 morphs, and the lack of BamHI polymorphism) were also present in this sample. Since the only neighbouring population accessible to Tharus, until recently, has been Hindu (Caucasoids), this result strongly supports the notion that Tharus are indeed a single anthropological entity; 2. two statistically significant differences between Eastern and Central Tharus--namely, a much higher HaeII morph 5 frequency among Central Tharus, and the absence in the same group of the mutation at 15.487 bp (very common among Eastern Tharus)--together with the results on alpha-tal gene, suggested that Tharu subgroups underwent an effective reproductive isolation.

190 Punjabis (Hindus and Sikhs) of Chandigarh and 152 Hindus of Jodhpur (Rajasthan) were examined for six genetic markers, four of which (APO C-II, C6, C7 and FXIIIA) were not studied before in Asiatic Indians. For APO C-II and C7 only the common phenotype was found in a total of 229 and 99 subjects, respectively. For the remaining four markers the two samples were pooled since the gene frequency estimates were not significantly different: FXIIIA*2 = 0.205 +/- 0.016; C6*B = 0.366 +/- 0.037; PGM1*2 = 0.247 +/- 0.017; PGD*C = 0.041 +/- 0.008. These data may contribute to evaluate the extent of the Mongoloid genetic admixture into the Caucasoid gene pool of the Punjab and Rajasthan Hindu population.