Mitochondrial DNA control region polymorphism in the population of Alagoas state, north-eastern Brazil

The sequences of the two hypervariable (HV) segments of the mitochondrial DNA (mtDNA) control region were determined in 167 randomly selected, unrelated individuals living in the state of Alagoas, north-eastern Brazil. One hundred and forty-five different haplotypes, associated with 139 variable positions, were determined. More than 95% of the mtDNA sequences could be allocated to specific mtDNA haplogroups according to the mutational motifs. Length heteroplasmy in the C-stretch HV1 and HV2 regions was observed in 22 and 11%, respectively, of the population sample. The genetic diversity was estimated to be 0.9975 and the probability of two random individuals presenting identical mtDNA haplotypes was 0.0084. The most frequent haplotype was shared by six individuals. All sequences showed high-quality values and phantom mutations were not detected. The diversity revealed in the mitochondrial control region indicates the importance of this locus for forensic casework and population studies within Alagoas, Brazil.     

Analysis of 16 Y STR loci in the Finnish population reveals a local reduction in the diversity of male lineages

We analysed samples of 400 Finnish males using nine Y-chromosomal short tandem repeat (STR) loci (minimal haplotype); for 200 of these subjects an additional seven Y-chromosomal STR loci were used. The geographical distribution of the observed haplotypes was determined from 200 individuals of known paternal origin within Finland. The observed number of alleles varied from 2 to 13 alleles per locus. A total of 146 minimal haplotypes were identified in our population sample. Interestingly, 90 (22.5%) individuals shared an identical haplotype. This haplotype was extremely frequent in the northern and eastern subpopulations of Savo, Pohjanmaa and Karjala (53, 42 and 37%, respectively). With the seven additional loci analysed in the sample of 200 individuals, 120 haplotypes were identified, and individuals sharing the most common haplotype decreased to 13.0%. However, in comparison to other European populations, the Finnish population showed decreased genetic diversity (GD) when the number of different minimal haplotypes in the population was divided by the sample size (36.5% in Finns versus 83.7% on average). Our results strongly support the earlier hypothesis of individual isolated Y-chromosomal lineages and population substructuring in Finland. For paternity testing, power of exclusion was 92% using minimal haplotype data, but including the seven additional loci this value increased to 97%.     

Forensic genetic analysis of mitochondrial DNA hypervariable region I/II sequences: an expanded Korean population database

We have analyzed variation of the mitochondrial DNA (mtDNA) hypervariable segments I and II (HVS-I and HVS-II) in 185 randomly chosen individuals from Korea to provide an expanded and reliable Korean database. Combined sequence comparison of HVS-I and HVS-II led to the identification of 167 different haplotypes characterized by 154 variable sites. One hundred and fifty-one of the haplotypes were individual-specific, 14 were found in two individuals and 2 were found in three individuals. A pairwise comparison of the 185 HVS-I/II sequences found an average of 10.11 +/- 4.63 differences between individuals. The random match probability and gene diversity for the combined hypervariable regions were estimated at 0.66% and 0.9988, respectively. Analyzing the expanded database including three previously reported data sets and the present data using haplogroup-based comparisons and comparison with closely related sequences allowed errors to be detected and eliminated, thus considerably improving data quality. Sample division comparisons based on PhiST genetic distance measures revealed no significant population differentiation in the distribution of mtDNA sequence variations between the present data set and a database in The Scientific Working Group on DNA Analysis Methods (SWGDAM), but did indicate differences from other sets of data. Based on the results of mtDNA profiles, almost all of the mtDNA types studied here could be classified into subsets of haplogroups common in east Asia, and show that the Koreans possess lineages from both the southern and the northern haplogroup complexes of east Asian populations. The new data, combined with other mtDNA sequences, demonstrate how useful comparison with closely related mtDNA sequences can be for improving database quality, as well as providing haplotype information for forensic and population genetic analyses in the Korean population.     

Mitochondrial diversity of a northeast German population sample

Mitochondrial DNA sequences of the hypervariable regions HV I and HV II were analyzed in 300 unrelated individuals born and living in the northeast corner of Germany (Western Pomerania) to generate a database for forensic identification purposes in this region. Sequence polymorphism were detected using PCR and direct sequencing analysis. A total of 242 different haplotypes were found as determined by 147 variable positions. The most frequent haplotype (263G, 315.1C) was found in 10 individuals and is also the most common sequence in Europe. Three other haplotypes were shared by 5 individuals, 2 sequences by 4, 8 haplotypes by 3, 15 sequences by 2 persons, and 213 sequences were unique. The genetic diversity was estimated to be 0.99 and the probability of two random individuals showing identical mitochondrial DNA (mtDNA) haplotypes is 0.6%. A comparison with other studies from Germany showed only little differences in the distribution of haplogroups. Nevertheless, one frequent haplotype in northeast Germany (five unrelated individuals) could only rarely be found in other German and European regions. Our results may indicate that despite a high admixture proportion in the German population some regions could demonstrate certain characteristic features.     

Mitochondrial DNA population data of HVS-I and HVS-II sequences from a northeast German sample

Mitochondrial DNA sequences of the control region's two hypervariable regions HVS-I and HVS-II were determined for 213 unrelated west Eurasian individuals from northeast Germany (Mecklenburg). A total of 174 different mtDNA haplotypes were found, 25 of which were shared by more than 1 individual. The most frequent haplotypes were 263G-309.1C-315.1C, found in seven individuals, 263G-309.1C-309.2C-315.1C, found in six individuals and 263G-315.1C, found in five individuals. These sequences are also the most common haplotypes in other published European data sets. The sequence polymorphisms consisting of 150 polymorphic nucleotide positions were compared with other European databases. The genetic diversity and random match probability were calculated. Our results corroborate certain features which are characteristic for west Eurasian mtDNA population samples.     

Typing the 1.1 kb control region of human mitochondrial DNA in Japanese individuals

This study presents a reliable method that uses high-fidelity long-range PCR and optimized primers to assess polymorphism and to genotype human mitochondrial DNA (mtDNA). This method was used to analyze polymorphic sites in the human mtDNA control region, including hypervariable regions I, II, and III (HVI, HVII, and HVIII), from 124 unrelated Japanese individuals. In HVI, HVII, and HVIII, 80, 37, and 14 polymorphic sites were identified, respectively, excluding those in the homopolymeric cytosine stretch (C-stretch) regions. The region between HVI and HVII also contained 15 polymorphic sites. On the other hand, C-stretch length heteroplasmy in HVI or HVII was observed in 66 of 124 Japanese individuals (53%), which is much higher than in Caucasian populations. The variants in the C-stretch regions were characterized by counting the number of heteroplasmic peaks split from the single peak in homoplasmic sequences (i.e., 16244G and 16255G in HVI and 285G in HVII). Including the C-stretch length heteroplasmy, the 124 Japanese mtDNA samples were classified into 116 distinct haplotypes. The random match probability and the genetic diversity were estimated to be 0.95% and 0.998581, respectively, indicating that the method presented here has higher discrimination than the conventional method for mtDNA typing using HVI and HVII. [Correction added after publication 30 January 2007: in the preceding sentence random match probability and genetic diversity estimates were corrected from 0.95 and 0.998581%, respectively, to 0.95% and 0.998581, respectively.] The haplogroups and their frequencies observed in this study (i.e., D4; 13.7%, M7a1; 11.3%, D4a; 9.7% and M7b2; 8.9%) were similar to those observed in other studies of Japanese mtDNA polymorphism. The method described here is suitable for forensic applications, as shown by successful analysis of tissues from highly putrefied remains of an infant, which allowed maternal relationship to be determined via mtDNA haplotyping.     

A database of mitochondrial DNA hypervariable regions I and II sequences of individuals from Slovakia

In order to identify polymorphic positions and to determine their frequencies and the frequency of haplotypes in the human mitochondrial control region, two hypervariable regions (HV1 and HV2) of the mitochondrial DNA (mtDNA) of 374 unrelated individuals from Slovakia were amplified and sequenced. Sequence comparison led to the identification of 284 mitochondrial lineages as defined by 163 variable sites. Genetic diversity (GD) was estimated at 0.997 and the probability of two randomly selected individuals from population having identical mtDNA types (random match probability, RMP) for the both regions is 0.60%.     

The maternal inheritance of the Ashaninka native group from Peru

The Amazonia rainforest, in South America, harbours native populations with high ethnic diversity. The evaluation of the genetic composition of these populations represents a challenge, and only few studies are available describing its native groups. In this work, the maternal inheritance of 170 Ashaninka individuals living in the Amazonia region of Pasco department, Peru, was evaluated by mtDNA control region sequencing. As previously observed for other native groups from Amazonia, low haplotype diversity was obtained, and only Native American haplogroups were found. Strong founder effects were observed, especially for sub haplogroups A2aa, B2b+152, C1b and D1. During the European colonial period, the Ashaninka population seems to have remained relatively isolated, which can be explained by its remote location in the tropical forest. A comparison with other native South American populations from different linguistic families showed a lack of geographic or linguistic affiliations, highlighting the importance of having specific mtDNA database for the native groups in South America.     

Nucleotide variability of HV-I in Afro-descendents populations of the Brazilian Amazon Region

The analysis of genetic variation in the nucleotide sequences of mitochondrial DNA, provides unique information about the population diversity and human identification. In this study, the mitochondrial DNA sequences of the first hypervariable region (HV-I) were analyzed in 243 unrelated individuals of seven Afro-descendents populations of the Amazon Region. Sequence polymorphisms were detected using PCR and direct sequencing analysis. A total of 133 different haplotypes were found determined by 97 variable nucleotides. Each one of the three more frequent haplotypes was shared by 9 samples and 91 sequences were unique. The genetic diversity was estimated to 0.9898+/-0.0016 and the probability of two random individuals showed identical mitochondrial DNA (mtDNA) haplotypes were 1.2%.     

Characterization of human control region sequences of the African American SWGDAM forensic mtDNA data set

The scientific working group on DNA analysis Methods (SWGDAM) mitochondrial DNA (mtDNA) population data set is used to infer the relative rarity of control region mtDNA profiles obtained from evidence samples and of profiles used for identification of missing persons. In this study, the African American haplogroup patterns in the SWGDAM data were analyzed in a phylogenetic context to determine relevant single nucleotide polymorphisms (SNPs) and to describe haplogroup distributions for Africans observed in these data sets. Over 200 SNPs (n=217) were observed in the African American data set (n=1148). These SNPs ranged from having 1-39 changes in the phylogenetic tree, with sites 152 and 16519 being the most variable. On average there were 5.8 changes for a character on the tree. The most variable sites (with 19 or more changes each) observed included 16093, 16129, 16189, 16311, 16362, 16519, 146, 150, 152, 189, and 195. These rapidly changing sites are consistent with other published analyses. Only 34 SNPs are needed to identify all clusters containing 10 or more individuals in the African American data set. The results show that the African American SWGDAM mtDNA data set contains variation consistent with that described in continental African populations. Thirteen of the 18 haplogroups previously observed in African populations were observed and include: L1a, L1b, L1c, L2a, L2b, L2c, L3b, L3d, L3e1, L3e2, L3e3, L3e4 and L3f. Haplogroup L2a is the most commonly observed cluster (18.8%) in the African American data set. The next most common haplogroups in the African American data set include the clusters L1c (11.0%), L1b (9.1%), L3e2 (9.0%) and L3b (8.1%). Approximately 8% of the haplogroups observed within African Americans were common in European Caucasians or East Asians; these were H (n=32), J (n=4), K (n=5), T (n=2), U5 (n=6), U6 (n=9 also known from North Africa), A (n=12), B (n=7), C (n=4), and M (n=16), respectively. The European Caucasian and East Asian haplogroups are expected due to admixture between individuals with recent ancestry in Western Eurasia and sub-Saharan Africa. The genetic characterization of these relevant data sets is fully consistent with other published mtDNA genetic variation. The sequence diversity observed in this data set makes it a valuable tool for forensic applications.     

Forensic utility of the mitochondrial hypervariable region 1 of domestic dogs, in conjunction with breed and geographic information

The 608-bp hypervariable region 1 (HV1) sequences from 36 local dogs were analyzed to characterize the population genetic structure of canid mitochondrial DNA (mtDNA). Sixteen haplotypes were identified. A 417-bp segment of this sequence was compared with GenBank sequences from a geographically representative sample of 201 dogs, two coyotes, and two wolves. Sixty-six haplotypes were identified including 62 found only in domestic dogs. Fourteen of these correspond to the 16 local haplotypes and were among the most frequent haplotypes. The local sample was judged to be representative of the much broader geographic sample. No correlation was observed between local haplotypes and the owner's characterization of dog breed. A 60-bp variation "hotspot" within the canid HV1 was identified as a potentially valuable molecular tool, particularly for assaying limited or degraded DNA samples.     

Characterization of the Caucasian haplogroups present in the SWGDAM forensic mtDNA dataset for 1771 human control region sequences. Scientific Working Group on DNA Analysis Methods

Currently, the Scientific Working Group on DNA Analysis Methods (SWGDAM) mtDNA dataset is used to infer the relative rarity of mtDNA profiles (i.e., haplotypes) obtained from evidence samples and for identification of missing persons. The Caucasian haplogroup patterns in this forensic dataset have been characterized using phylogenetic methods. The assessment reveals that the dataset is relevant and representative of U.S. and European Caucasians. The comparisons carried out were both the observation of variable sites within the control region (CR) and the selection of a subset of these sites, which partition the variation within human mtDNA control region sequences into clusters (i.e., haplogroups). The aligned sequence matrix was analyzed to determine both single nucleotide polymorphisms (SNPs) in a phylogenetic context, as well as to check and standardize haplogroup designations with a focus on determining the characters that define these groups. To evaluate the dataset for forensic utility, the haplogroup identifications and frequencies were compared with those reported from other published studies.     

Y-chromosomal STR haplotypes in a population from the Amazon region, Brazil

Haplotype and allele frequencies of the nine Y-STR (DYS19, DYS389 I, DYS389 II, DYS390, DYS391, DYS392, DYS393, DYS385 I/II) were determined in a population sample of 200 unrelated males from Belém, Brazil. The most common haplotypes are shared by 1.5% of the sample, while 186 haplotypes are unique. The haplotype diversity is 0.9995+/-0.0006. The data obtained were compared to those of other Brazilian populations. AMOVA indicates that 99.91% of all the haplotypical variation is found within geopolitical regions and only 0.09% is found among regions.     

Population structure of Y chromosome SNP haplogroups in the United States and forensic implications for constructing Y chromosome STR databases

A set of 61 Y chromosome single-nucleotide-polymorphisms (Y-SNPs) is typed in a sample of 2517 individuals from 38 populations to infer the geographic origins of Y chromosomes in the United States and to test for paternal admixture among African-, European-, Hispanic-, Asian-, and Native-Americans. All of the samples were previously typed with the 11 core U.S. Y chromosome short tandem repeats (Y-STRs) recommended by SWGDAM, which revealed high levels of among ethnic group variation and low levels of among-population-within-ethnic-group variation. Admixture estimates vary greatly among populations and ethnic groups. The frequencies of non-European (3.4%) and non-Asian (4.5%) Y chromosomes are generally low in European-American and Asian-American populations, respectively. The frequencies of European Y chromosomes in Native-American populations range widely (i.e., 7-89%) and follow a West to East gradient, whereas they are relatively consistent in African-American populations (26.4+/-8.9%) from different locations. The European (77.8+/-9.3%) and Native-American (13.7+/-7.4%) components of the Hispanic paternal gene pool are also relatively constant among geographic regions; however, the African contribution is much higher in the Northeast (10.5+/-6.4%) than in the Southwest (1.5+/-0.9%) or Midwest (0%). To test for the effects of inter-ethnic admixture on the structure of Y-STR diversity in the U.S., we perform subtraction analyses in which Y chromosomes inferred to be admixed by Y-SNP analysis are removed from the database and pairwise population differentiation tests are implemented on the remaining Y-STR haplotypes. Results show that low levels of heterogeneity previously observed between pairs of Hispanic-American populations disappear when African-derived chromosomes are removed from the analysis. This is not the case for an unusual sample of European-Americans from New York City when its African-derived chromosomes are removed, or for Native-American populations when European-derived chromosomes are removed. We infer that both inter-ethnic admixture and population structure in ancestral source populations may contribute to fine scale Y-STR heterogeneity within U.S. ethnic groups.     

Mitochondrial DNA study in the Shuar ethnic group from Ecuador

This study presents mitochondrial data from 55 unrelated individuals from two Ecuadorian Shuar communities: Kumbatza and Yukateis. Maternal linage was determined by analyzing the two mtDNA hypervariable regions: HRVI and HRVII. It was shown that the Shuar population exhibited the haplogroup B. This demonstrates that Shuar group is a conserved population with no mixing with the European and African diaspora populations.     

A new strategy for the discrimination of mitochondrial DNA haplogroups in Han population

Mitochondrial DNA (mtDNA) haplogroup discrimination is interesting not only for phylogenetic and clinical but also for forensic studies. We discriminated the mtDNA haplogroups of 570 healthy unrelated Han people from Zhejiang Province, Southeast China, by comprehensive analysis mutations of the hypervariable segments-I sequence and diagnostic polymorphisms in mtDNA coding region using real-time polymerase chain reaction (RT-PCR), which was compared with the widely used PCR and restriction fragment length polymorphism (PCR-RFLP) method. The results showed that in superhaplogroup M, haplogroup D was the most common haplotype within this assay to 24.6%, and in the other superhaplotype N, haplogroup B and F were the most common groups. Samples re-identified by PCR-RFLP showed the consistent results that were got with RT-PCR. In conclusion, the RT-PCR strategy appears to be an accurate, reproducible, and sensitive technique for the discrimination of mtDNA haplogroups, especially for mass screenings quickly and economically.     

Geographical heterogeneity of Y-chromosomal lineages in Norway

Y-chromosomal variation at five biallelic markers (Tat, YAP, 12f2, SRY(10831) and 92R7) and nine multiallelic short tandem repeat (STR) loci (DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS385I/II and DYS388) in a Norwegian population sample are presented. The material consists of 1766 unrelated males of Norwegian origin. The geographical distribution of the population sample reflects fairly well the population distribution around the year 1942, which is the median birth year of the index persons. Seven hundred and twenty-one different Y-STR haplotypes but 726 different lineages (Y-STRs plus biallelic markers) were encountered. We observed six known (P*(xR1a), BR(xDE, J, N3, P), R1a, N3, DE, J), and one previously undescribed haplogroup (probably a subgroup within haplogroup P*(xR1a)). Four of the haplogroups (P*(xR1a), BR(xDE, J, N3, P), R1a and N3) represented about 98% of the population sample. The analysis of population pairwise differences indicates that the Norwegian Y-chromosome distribution most closely resembles those observed in Iceland, Germany, the Netherlands and Denmark. Within Norway, geographical substructuring was observed between regions and counties. The substructuring reflects to some extent the European Y-chromosome gradients, with higher frequency of P*(xR1a) in the south-west and of R1a in the east. Heterogeneity in major founder groups, geographical isolation, severe epidemics, historical trading links and population movements may have led to population stratification and have most probably contributed to the observed regional differences in distribution of haplotypes within two of the major haplogroups.     

Mitochondrial DNA typing screens with control region and coding region SNPs

Mitochondrial DNA (mtDNA) analysis has found an important niche in forensic DNA typing. It is used with highly degraded samples or low-copy number materials such as might be found from shed hair or bones exposed to severe environmental conditions. The primary advantage of mtDNA is that it is present in high copy number within cells and therefore more likely to be recovered from highly degraded specimens. A major disadvantage to traditional forensic mtDNA analysis is that it is time-consuming and labor-intensive to generate and review the 610 nucleotides of sequence information commonly targeted in hypervariable regions I and II (HVI and HVII) of the control region. In addition, common haplotypes exist in HVI/HVII mtDNA sequences that can reduce the ability to differentiate two unrelated samples. In this report we describe the utility of two newly available screening assays for rapid exclusion of non-matching samples. The LINEAR ARRAY mtDNA HVI/HVII Region-Sequencing Typing Kit (Roche Applied Science, Indianapolis, IN) was used to type 666 individuals from U.S. Caucasian, African American, and Hispanic groups. Processing of the LINEAR ARRAY probe panels "mito strips" was automated on a ProfiBlot workstation. Observable variation in 666 individuals is reported and frequencies of the mitotypes within and between populations are presented. Samples exhibiting the most common Caucasian mitotype were subdivided with a multiplexed amplification and detection assay using eleven single nucleotide polymorphisms in the mitochondrial genome. These types of screening assays should enable more rapid evaluation of forensic casework samples such that only samples not excluded would be subjected to further characterization through full HVI/HVII mtDNA sequence analysis.     

The EDNAP mitochondrial DNA population database (EMPOP) collaborative exercises: organisation, results and perspectives

This paper presents an overview of the organisation and the results of the collaborative exercises (CE) of the European DNA Profiling (EDNAP) Group's mitochondrial DNA population database project (EMPOP). The aim of the collaborative exercises was to determine whether uniformity of mtDNA sequencing results could be achieved among different laboratories. These were asked to sequence either the complete mtDNA control region or the two hypervariable regions HVI (16024-16365) and HVII (73-340) from DNA extracts, buccal swabs or bloodstains, proceeding in accordance with the protocol and strategies used in each individual laboratory. The results of the collaborative exercises were employed to identify possible sources of errors that could arise during the analysis and interpretation of mtDNA profiles. These findings were taken as a basis to tentatively make suitable arrangements for the construction of a high quality mtDNA database. One hundred fifty mtDNA profiles were submitted to the evaluating laboratory, and disaccording profiles were classified into four groups corresponding to the source of error: clerical errors, sample mix-ups, contaminations and discrepancies with respect to the mtDNA nomenclature. Overall, 14 disaccording haplotypes (16 individual errors) were observed. The errors included 10 clerical errors, 3 interpretation problems, 2 cases of sample mix-up and 1 case of point heteroplasmic mixture, where the 2 sequencing reactions brought inconsistent base calls. This corresponds to an error rate of 10.7% in a virtual mtDNA database consisting of the collaborative exercise results. However, this estimate is still conservative compared to conclusions drawn by authors of meanwhile numerous publications critically reviewing published mtDNA population databases. Our results and earlier published concerns strongly emphasize the need for appropriate safety regulations when mtDNA profiles are compiled for database purposes in order to accomplish the high standard required for mtDNA databases that are used in the forensic context.