基于16S rRNA 初步探讨福建华溪蟹与修水华溪蟹的遗传分化

采用PCR结合DNA测序技术,测定两种华溪蟹Sinopotamon.fukienense 与 S.xiusuiense 线粒体16S rRNA基因序列的组成和变异。经比对获得513 bp的核苷酸序列,并分析S.fukienense与S.xiusuiense 间的的遗传分化。结果显示S.fukienense和S.xiusuiense两个群体之间的遗传距离为0.032±0.006,遗传分化指数（Fst）为0.70336,基因流(Nm)为0.11。这表明两群体间高度分化,基因交流很少。重建的系统发生显示它们各自形成独立的分支,这与形态学鉴定结果基本一致。     

Origin of the beta S-globin gene in blacks: the contribution of recurrent mutation or gene conversion or both.

In order to investigate the origin(s) of the mutation(s) leading to the beta S-globin gene in North American populations of African ancestry, we analyzed DNA polymorphisms in the beta-globin gene cluster in a large number of both beta A- and beta S-globin gene-bearing chromosomes in U.S. and Jamaican Blacks. We found 16 different haplotypes of polymorphic sites associated with 170 beta S-globin gene-bearing chromosomes. The three most common beta S haplotypes, which account for 151/170 of the beta S-globin gene-bearing chromosomes, are only rarely seen in the chromosomes bearing the beta A-globin gene in these populations (6/47). Two observations suggest multiple origins or interallelic gene conversion, or both, of the beta S mutation. First, the mutation is present in all three beta-globin gene frameworks. Second, the beta S haplotypes can be divided into four groups, each of which cannot be derived from any other by less than two crossing-over events. In summary, our observation of the beta S mutation on 16 different haplotypes in African populations can be best explained by (i) a number of simple recombination events 5' to the beta-globin gene and (ii) up to four independent mutations and/or interallelic gene conversions.     

pfcrt Polymorphism and the spread of chloroquine resistance in Plasmodium falciparum populations across the Amazon Basin

The widespread occurrence of drug-resistant malaria parasites in South America presents a formidable obstacle to disease control in this region. To characterize parasite populations and the chloroquine-resistance profile of Plasmodium falciparum in the Amazon Basin, we analyzed a DNA segment of the pfcrt gene, spanning codons 72-76, and genotyped 15 microsatellite (MS) markers in 98 isolates from 6 areas of Brazil, Peru, and Colombia where malaria is endemic. The K76T mutation, which is critical for chloroquine resistance, was found in all isolates. Five pfcrt haplotypes (S[tct]MNT, S[agt]MNT, CMNT, CMET, and CIET) were observed, including 1 previously found in Asian/African isolates. MS genotyping showed relatively homogeneous genetic backgrounds among the isolates, with an average of 3.8 alleles per marker. Isolates with identical 15-loci MS haplotypes were found in different locations, suggesting relatively free gene flow across the Amazon Basin. Allopatric isolates carrying SMNT and CMNT haplotypes have similar genetic backgrounds, although parasites carrying the CIET haplotype have some exclusive MS alleles, suggesting that parasites with CIET alleles were likely to have been introduced into Brazil from Asia or Africa. This study provides the first evidence of the Asian pfcrt allele in Brazil and a detailed analysis of P. falciparum populations, with respect to pfcrt haplotypes, in the Amazon Basin.     

Hybrid Tamarix widespread in U.S. invasion and undetected in native Asian range

Biological invasions are drastically altering natural habitats and threatening biodiversity on both local and global levels. In one of the United States' worst invasions, Eurasian Tamarix plant species have spread rapidly to dominate over 600,000 riparian and wetland hectares. The largest Tamarix invasion consists of Tamarix chinensis and Tamarix ramosissima, two morphologically similar species. To clarify the identity, origins, and population structuring of this invasion, we analyzed DNA sequence data from an intron of a nuclear gene, phosphoenolpyruvate carboxylase (PepC). This intron proved to be highly variable at the population level, and the 269 native and invasive specimens yielded 58 haplotypes, from which we constructed a gene genealogy. Only four of these haplotypes were common to both the U.S. and Eurasia. Surprisingly, we found that the most common plant in this U.S. invasion is a hybrid combination of two species-specific genotypes that were geographically isolated in their native Eurasian range. Less extensive hybrids exist in the invasion, involving combinations of T. ramosissima and T. chinensis with Tamarix parviflora and Tamarix gallica. The presence of potentially novel hybrids in the U.S. illustrates how importation of exotics can alter population structures of species and contribute to invasions.     

Analysis of betaS and betaA genes in a Mexican population with African roots

To investigate the origin of the beta(A) and beta(S) genes in a Mexican population with African roots and a high frequency of hemoglobin S, we analyzed 467 individuals (288 unrelated) from different towns in the states of Guerrero and Oaxaca in the Costa Chica region. The frequency of the sickle-cell trait was 12.8%, which may represent a public health problem. The frequencies of the beta-haplotypes were determined from 350 nonrelated chromosomes (313 beta(A) and 37 beta(S)). We observed 15 different beta(A) haplotypes, the most common of which were haplotypes 1 (48.9%), 2 (13.4%), and 3 (13.4%). The calculation of pairwise distributions and Nei's genetic distance analysis using 32 worldwide populations showed that the beta(A) genes are more closely related to those of Mexican Mestizos and North Africans. Bantu and Benin haplotypes and haplotype 9 were related to the beta(S) genes, with frequencies of 78.8, 18.2, and 3.0%, respectively. Comparison of these haplotypes with 17 other populations revealed a high similitude with the population of the Central African Republic. These data suggest distinct origins for the beta(A) and beta(S) genes in Mexican individuals from the Costa Chica region.     

Genetic variation of Anopheles dirus A and D (Diptera:Culicidae) in China: inferred by mtDNA-CO I gene sequences]

OBJECTIVE: To interpret genetic variation and population structure of Anopheles dirus A and D from China by molecular marker. METHODS: Samples included An. dirus A of Hainan laboratory colony (n=13), and field specimen from Mengla (n=17) and Jiangcheng (n=17) in Yunnan Province. The specimens were identified by PCR assay before study, mtDNA-CO I region was amplified and sequenced. Genetic variation and population structure was estimated according to sequence data. RESULTS: The mtDNA-CO I gene with a length of 959 bp was analyzed. There were three haplotypes in An. dirus A and six haplotypes in An. dirus D. The above haplotypes distributed in three populations uniformly. The average number of pairwise differences within Mengla population (7.4412) was greater than that of Jiangcheng (1.2794) and Hainan (1.0513) populations, which suggested that the level of genetic divergence was the highest within Mengla population. The result of hierarchical AMOVA estimation showed a limited geneflow (Fst=0.799 9), therefore the variation level in a population (20.01%) was smaller than among the populations (79.99%). CONCLUSION: The inter-specific genetic variation between An. dirus A and D in China was small and the level of divergence among individuals was high.     

不同地理种群白纹伊蚊线粒体基因COI的遗传多样性分析

目的探讨白纹伊蚊不同地理种群间的进化关系和遗传分化情况,为白纹伊蚊防制和蚊媒病防控提供基础资料。方法在广泛采集样品的基础上,通过PCR扩增、测序获取线粒体基因COI片段,并从GenBank下载了部分序列,比对、剪切后的598bp用于后续分析。结果系统发育分析的结果表明所有白纹伊蚊的COI序列聚成一支,没有明显的遗传差异;60条COI序列分属于19个单倍型,其中4个为共享单倍型;单倍型多样性(Hd)为0.737,核苷酸多样性(π)为0.20%;海南的白纹伊蚊种群与绝大部分地理种群间出现了明显的分化(P0.05);H1和H6形成了2个辐射中心,是较为原始的单倍型。结论我国的白纹伊蚊种群正处于扩张的趋势,海南的白纹伊蚊种群与其它地理种群间出现了明显分化。     

Hypercontrols in genotype-phenotype analysis reveal ancestral haplotypes associated with essential hypertension

The angiotensinogen gene locus has been associated with essential hypertension in most populations analyzed to date. Increased plasma angiotensinogen levels have been proposed as an underlying cause of essential hypertension in whites; however, differences in the genetic regulation of plasma angiotensinogen levels have also been reported for other populations. The aim of this study was to analyze the relationship between angiotensinogen gene polymorphisms and haplotypes with plasma angiotensinogen levels and the risk of essential hypertension in the Mexican population. We genotyped 9 angiotensinogen gene polymorphisms in 706 individuals. Four polymorphisms, A-6, C4072, C6309, and G12775, were associated with increased risk, and the strongest association was found for the C6309 allele (χ(2)=23.9; P=0.0000009), which resulted in an odds ratio of 3.0 (95% CI: 1.8-4.9; P=0.000006) in the recessive model. Two polymorphisms, A-20C (P=0.003) and C3389T (P=0.0001), were associated with increased plasma angiotensinogen levels but did not show association with essential hypertension. The haplotypes H1 (χ(2)=8.1; P=0.004) and H5 (χ(2)=5.1; P=0.02) were associated with essential hypertension. Using phylogenetic analysis, we found that haplotypes 1 and 5 are the human ancestral haplotypes. Our results suggest that the positive association between angiotensinogen gene polymorphisms and haplotypes with essential hypertension is not simply explained by an increase in plasma angiotensinogen concentration. Complex interactions between risk alleles suggest that these haplotypes act as "superalleles."     

Sequence variations in the 5' flanking and IVS-II regions of the G gamma- and A gamma-globin genes of beta S chromosomes with five different haplotypes

We have amplified and sequenced the 5' flanking and the second intervening sequence (IVS-II) regions of both the G gamma- and A gamma-globin genes of the beta S chromosomes from sickle cell anemia (SS) patients with homozygosities for five different haplotypes. The sequencing data, compared with previously published sequences for the normal chromosomes A and B, show many similarities to chromosome B for haplotypes 19, 20, and 17, while haplotypes 3 and 31 are remarkably similar to chromosome A and also similar to each other. Several unique mutations were found in the 5' flanking regions (G gamma and A gamma) of haplotypes 19 and 20 and in the IVS-II segments of the same genes of haplotypes 19, 20, and 17; the IVS-II of haplotypes 3 and 31 were identical to those of chromosome A. Dot-blot analyses of amplified DNA from additional SS patients with specific probes have confirmed that these mutations are unique for each haplotype. The two general patterns that have been observed among the five haplotypes have most probably arisen by gene conversion events between the A and B type chromosomes in the African population. These patterns correlate with high and low fetal hemoglobin expression, and it is speculated that these and other yet unknown gene conversions may contribute to the variations in hemoglobin F and G gamma levels observed among SS patients. In vitro expression experiments involving the approximately 1.3-kb 5' flanking regions of the G gamma- and A gamma-globin genes of the beta S chromosomes with the five different haplotypes failed to detect differences between the levels of expression, suggesting that the sequence variations observed between these segments of DNA are not the primary cause of the differences in hemoglobin F levels among the SS patients.     

Pancreatitis risk in primary hyperparathyroidism: relation to mutations in the SPINK1 trypsin inhibitor (N34S) and the cystic fibrosis gene

OBJECTIVE: Primary hyperparathyroidism (pHPT)-related hypercalcemia is considered to represent a risk factor for the development of pancreatitis. We therefore explored whether mutations in genes that were previously identified to increase the risk for pancreatitis coexist in a cohort of 826 patients with pHPT prospectively studied between 1987 and 2002. METHODS: Among 826 patients with pHPT, 38 patients were identified with pancreatitis (4.6%). DNA was available from 25 patients (13 women/12 men, 16 acute pancreatitis/9 chronic pancreatitis). These individuals and 50 patients with pHPT without pancreatitis were analyzed for mutations in the serine protease inhibitor Kazal type I (SPINK1) gene (N34S) and the cationic trypsinogen gene (PRSS1) (N29I, R122H) by melting curve analysis and DNA sequencing. Sequence analysis of the cystic fibrosis transmembrane conductance regulator (CFTR) gene was carried out for the detection of 36 mutations and the Tn polymorphism. RESULTS: Four of 25 patients with pHPT and pancreatitis carried the N34S missense mutation in the SPINK1 gene (16%), while all 50 controls (pHPT without pancreatitis) showed no mutation in SPINK1 or PRSS1 genes (P < 0.05 vs controls, P < 0.001 vs general population). CF-causing CFTR mutations were present in four patients (P < 0.05 vs general population), while one patient carried a 5T allele. One patient was transheterozygous (SPINK1: N34S/CFTR: R553X). Mean serum calcium levels in pancreatitis patients (3.1 mmol/L) did not differ significantly from the mean of the entire cohort (3.0 mmol/L) or pHPT patients without pancreatitis (3.1 mmol/L). CONCLUSION: Pancreatitis risk is approximately 10-fold elevated in pHPT, but pancreatitis occurs infrequently. This indicates an existing but minor impact of pHPT-related hypercalcemia. If pancreatitis occurs, it seems associated with genetic risk factors such as mutations in the SPINK1 and CFTR genes. In contrast, a combination of both hypercalcemia and genetic variants in SPINK1 or CFTR increases the risk to develop pancreatitis in patients with pHPT.     

The beta-globin gene cluster haplotypes in sickle cell anemia patients from Northeast Brazil: a clinical and molecular view

The beta(S)-globin haplotypes were studied in 78 sickle cell Brazilian patients from Bahia, Northeast Brazil, that has a large population of African origin. Hemoglobin (Hb) profiles were developed by high-performance liquid chromatography (HPLC), and beta(S)-globin gene haplotypes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) techniques. We identified 44 (55.0%) patients with the CAR/Ben (Central African Republic/Benin) genotype, 16 (20.0%) Ben/Ben, 13 (16.2%) CAR/CAR and seven (8.8%) with other genotypes. Analyses of the phenotypes showed clinical differences related only to Hb F levels and blood transfusion therapy; the presence of -alpha(-3.7)-thalassemia (thal) demonstrated statistical significance when associated with hematocrit (p=0.044), MCV (p=0.0007), MCH (p=0.012) and spleen sequestration events. The haplotype diversity found in the present study can be justified by information about the origin of the slave traffic period in Bahia during the 19th century. The specific characteristics described among the Bahian sickle cell patients could be confirmed by increasing the number of patients with specific genotypes and further studies of genetic markers.     

Phenotypic Diversity of Sickle Cell Disease in Patients with a Double Heterozygosity for Hb S and Hb D-Punjab

Phenotypic heterogeneity for sickle cell disease is associated to several genetic factors such as genotype for sickle cell disease, β-globin gene cluster haplotypes and Hb F levels. The coinheritance of Hb S (HBB: c.20A?>?T) and Hb D-Punjab (HBB: c.364G?>?C) results in a double heterozygosity, which constitutes one of the genotypic causes of sickle cell disease. This study aimed to assess the phenotypic diversity of sickle cell disease presented by carriers of the Hb S/Hb D-Punjab genotype and the Bantu [–?+?– – – –] haplotype. We evaluated medical records from 12 patients with sickle cell disease whose Hb S/Hb D-Punjab genotype and Bantu haplotype were confirmed by molecular analysis. Hb S and Hb D-Punjab levels were quantified by chromatographic analysis. Mean concentrations of Hb S and Hb D-Punjab were 44.8?±?2.3% and 43.3?±?1.8%, respectively. Painful crises were present in eight (66.7%) patients evaluated, representing the most common clinical event. Acute chest syndrome (ACS) was the second most prevalent manifestation, occurring in two individuals (16.7%). Three patients were asymptomatic, while another two exhibited greater diversity of severe clinical manifestations. Medical records here analyzed reported a significant clinical diversity in sickle cell disease ranging from the absence of symptoms to wide phenotypic variety. The sickle cell disease genotype, Bantu haplotype and hemoglobin (Hb) levels did not influence the clinical diversity. Thus, we concluded that the phenotypic variation in sickle cell disease was present within a specific genotype for disease regardless of the β-globin gene cluster haplotypes.     

Genetic structure of Aedes aegypti populations in Thailand using mitochondrial DNA

A hierarchical population genetic study was conducted among 19 Aedes aegypti populations in Thailand from Chiang Mai in the north to Songkhla province in the south. Single-strand conformation polymorphism analysis was used to examine variation in a 359-basepair region of the NADH dehydrogenase subunit 4 mitochondrial DNA gene (ND4). Seven haplotypes were detected in two lineages previously identified in ND4 haplotypes from North America. Gene flow estimates and highly significant variation among populations within 25 kilometers implicated genetic drift and vector control efforts as major factors in genetic structure. Mantel regression analysis demonstrated no isolation by distance. Urban areas were relatively panmictic, while suburban/rural sites exhibited more restricted gene flow. Significant genetic structure among groups of collections > 100 kilometers apart is consistent with recent (approximately 50 year) expansion of Ae. aegypti from highly populated areas accompanied by founder effects, but could also reflect the overall low genetic diversity in ND4 in Thailand.     

Fetal hemoglobin levels and beta s globin haplotypes in an Indian populations with sickle cell disease

To further explore the cause for variation in hemoglobin F (Hb F) levels in sickle cell disease, the beta globin restriction-fragment length polymorphism haplotypes were determined in a total of 303 (126 SS, 141 AS, 17 S beta degrees, 7 A beta, degrees and 12 AA) Indians from the state of Orissa. The beta s globin gene was found to be linked almost exclusively to a beta S haplotype ( -++-), which is also common in Saudi Arabian patients from the Eastern Province (referred to as the Asian beta s haplotype). By contrast, the majority of beta A and beta degree thalassemia globin genes are linked to haplotypes common in all European and Asian populations (+-----[+/-]; --++-++). Family studies showed that there is a genetic factor elevating Hb F levels dominantly in homozygotes (SS). This factor appears to be related to the Asian beta s globin haplotype, and a mechanism for its action is discussed. There is also a high prevalence of an independent Swiss type hereditary persistence of fetal hemoglobin (HPFH) determinant active in both the sickle cell trait and in sickle cell disease.     

Molecular phylogenetic study of Culex quinquefasciatus mosquito from different geographical regions of India using 16S rRNA gene sequences

Culex quinquefasciatus is a major vector of filariasis and various encephalitis in India and worldwide. Vector control remains the most successful strategy for the suppression of mosquito borne diseases. The genetic structure of vector populations in terms of insecticide resistance and susceptibility or refractoriness to infection may possibly vary. To exploit the genetic variability in vector population could pave the path for the alternative strategies in vector management. The sequences of ribosomal RNA molecules have been widely used for such studies. Here, we examined the molecular phylogenetic relationship among the Cx. quinquefasciatus collected from different geographical regions of India, using 16S ribosomal RNA (16S rRNA) gene nucleotide sequences. The distances among the species were measured using Pearson correlation; the Neighbor-Joining (NJ) method was used for the clustering with appropriate bootstrap values using Data Analysis in Molecular Biology and Evolution (DAMBE) software. The results revealed that the populations are genetically diverse. Based on the distance values and the tree topology on the basis of 16S rRNA sequences reflected the clear biogeographical and geoclimatic pattern among the different geographical populations from India.     

Structural analysis of the 5'' flanking region of the beta-globin gene in African sickle cell anemia patients: further evidence for three origins of the sickle cell mutation in Africa.

Haplotype analysis of the beta-globin gene cluster shows two regions of DNA characterized by nonrandom association of restriction site polymorphisms. These regions are separated by a variable segment containing the repeated sequences (ATTTT)n and (AT)xTy, which might be involved in recombinational events. Studies of haplotypes linked to the sickle cell gene in Africa provide strong argument for three origins of the mutation: Benin, Senegal, and the Central African Republic. Nevertheless, the haplotype determination does not give any information about the variable segment and does not totally exclude the possibility of recombination leading to different haplotypes linked to the mutation. The structure of the variable segment in the three African populations was studied by S1 nuclease mapping of genomic DNA, which allows a comparison of several samples. A 1080-base-pair DNA segment was sequenced for one sample from each population. S1 nuclease mapping confirmed the homogeneity of each population with regard to both (ATTTT)n and (AT)xTy repeats. We found three additional structures for (AT)xTy correlating with the geographic origin of the patients. Ten other nucleotide positions, 5' and 3' to the (AT)xTy copies, were found to be variable when compared to homologous sequences from human and monkey DNAs. These results allow us to propose an evolutionary scheme for the polymorphisms in the 5' flanking region of the beta-globin gene. The results strongly support the hypothesis of three origins for the sickle mutation in Africa.     

Mitochondrial DNA variation among Anopheles albimanus populations.

Barriers to gene flow between Pacific and Atlantic coast populations of Anopheles albimanus were reported in an earlier study of variation in the intergenic spacer of the nuclear ribosomal DNA. We examined the distribution of mitochondrial DNA haplotypes among A. albimanus populations to test for gene flow barriers with an independent genetic marker. A region of the NADH dehydrogenase subunit 5 gene was amplified by the polymerase chain reaction (PCR) in 1,105 mosquitoes collected from 16 locations in Guatemala and in single collections from Mexico, Honduras, Nicaragua, Costa Rica, Panama, Colombia, and Venezuela. The PCR products were tested for variation using single strand conformation polymorphism analysis and 45 haplotypes were detected. Haplotype frequencies did not vary between coasts in Guatemala. Populations within approximately 200 km of one another were panmictic. However, at distances > 200 km, FST and geographic distances were correlated suggesting that populations are isolated by distance.     

Location of the Genes for 16S and 23S Ribosomal RNA in the Genetic Map of Escherichia coli

By means of RNA-DNA hybridization with DNA from different merodiploids of Escherichia coli, the genes for 16S and 23S ribosomal RNA were found near minute 74 of the genetic map.