Haplogroup Assignment Research Articles

Insight into the genetic diversity of Selle Français horse populations based on whole D-loop sequencing of mitochondrial DNA

The Selle Français (SF) breed originated from native France horses crossed with Thoroughbreds, Arabs, Anglo-Arabs and French Trotters, which allowed for individuals to be obtained with excellent predispositions and a versatile usage for equestrian sports. The performance of the SF horse is widely accepted in the fields of show jumping and eventing; however, they have successfully started to compete in other areas such as dressage, vaulting, carriage driving and TREC competitions. To date, information about the genetic structure of the SF breed has been very limited. In the present report, Sanger sequencing was applied to analyse the whole D-loop of mitochondrial DNA of 98 samples representing Selle Français horses originating from two populations: Algerian (n=47) and French (n=51). The result obtained showed the presence of 19 SNPs, which allowed us to divide the samples into 41 haplotypes (OR909723‐OR909773). A higher number of haplotypes was detected in the Algerian population (n=24) compared to the French population (n=14), but both populations did not represent unique clusters and showed a high degree of genetic similarities. The result showed that the identified haplotypes of the SF belonged to four clades: L, A, B and G. The present report gives strong evidence that the Selle Français breed has been established based on a great influence of the Thoroughbred, whose close connection is more visible in the French population. The haplogroup assignment to L clades proved that the SF breed has European origin as ancestral haplogroup L was assigned to West and South Europe while A to West Asia and G and B to Middle East. Our results show SF as an example of a contemporary breed of sport horses with European roots further influenced by local breeds.

Ancient mitochondrial genome diversity in South America: Contributions from Quebrada del Toro, Northwestern Argentina.

The objective of this study was to enhance our understanding of the population history in South America, specifically Northwestern Argentina, by analyzing complete ancient mitogenomes of individuals from the Ojo de Agua archeological site (970 BP) in Quebrada del Toro (Salta, Argentina). We analyzed teeth from four individuals from the site Ojo de Agua (970 ± 60 BP), located in Quebrada del Toro (Andean region of Northwestern Argentina). DNA extracts were converted to double-stranded DNA libraries and indexed using unique dual-indexing primer combinations. DNA libraries were then enriched for the complete mitochondrial genome, pooled at equimolar concentrations, and sequenced on an Illumina® MiSeq™ platform. Reads from high quality libraries were trimmed, merged, and then mapped to the revised Cambridge Reference Sequence. The aDNA damage patterns were assessed and contamination estimated. Finally, variants were called, filtered, and the consensus mitogenome was constructed and used for haplogroup assignment. We also compiled available mitogenome sequences from ancient and present-day populations from the Southcentral Andes and other surrounding regions in Argentina. Maximum Likelihood and Bayesian phylogenetic reconstructions were obtained using the generated dataset. We successfully obtained the complete mitogenome sequence from one individual with an average depth coverage of 102X. We discovered a novel haplotype that was assigned to haplogroup D1. Phylogenetic reconstructions suggests that this haplotype falls within the sister branches of the D1j lineage, forming a well-supported clade. The estimate TMRCA of this clade that includes D1j and its sister branches ranged between 12,535 and 18,669 ya. The sequence analyzed in this study represents the first ancient mitogenome from within the valley region in Northwestern Argentina. We found that a representative of a lineage highly associated with D1j was already present approximately 1000 BP in the region. Our results agree with the proposed origin of D1j in other regions north of Patagonia and independent of the Pacific coast fast migratory route, contrary to what was originally hypothesized. This study highlights the lack of information regarding pre-Hispanic genetic diversity and contributes to the knowledge about the peopling process in South America.

HaploCart: Human mtDNA haplogroup classification using a pangenomic reference graph.

Current mitochondrial DNA (mtDNA) haplogroup classification tools map reads to a single reference genome and perform inference based on the detected mutations to this reference. This approach biases haplogroup assignments towards the reference and prohibits accurate calculations of the uncertainty in assignment. We present HaploCart, a probabilistic mtDNA haplogroup classifier which uses a pangenomic reference graph framework together with principles of Bayesian inference. We demonstrate that our approach significantly outperforms available tools by being more robust to lower coverage or incomplete consensus sequences and producing phylogenetically-aware confidence scores that are unbiased towards any haplogroup. HaploCart is available both as a command-line tool and through a user-friendly web interface. The C++ program accepts as input consensus FASTA, FASTQ, or GAM files, and outputs a text file with the haplogroup assignments of the samples along with the level of confidence in the assignments. Our work considerably reduces the amount of data required to obtain a confident mitochondrial haplogroup assignment.

The mitochondrial DNA HVI and HVII sequences and haplogroup distribution in a population sample from Vietnam

Background Mitochondrial DNA (mtDNA) analysis has been used in forensics and requires well-established population databases for statistical interpretations. However, high-quality mtDNA data from Vietnamese population samples have been limited. Aim To examine the mtDNA sequences and haplogroup compositions of a Vietnamese population to provide an mtDNA dataset that can further be used to construct a Vietnamese-specific reference database. Subjects and methods A total of 173 Vietnamese individuals were analysed for two hypervariable regions (HVI and HVII) of mtDNA. Forensic parameters were calculated and haplogroup assignment was performed based on the resulting mtDNA haplotypes. Genetic relationships between the Vietnamese and other Asian populations were investigated through principal component analysis (PCA) and pairwise Fst. Results The Vietnamese population sample consisted of 145 different haplotypes with a random match probability of 0.96%, a power of discrimination of 0.9904, and a haplotype diversity of 0.9962. The samples were assigned to 83 haplogroups that were commonly reported in Asia. PCA and pairwise Fst revealed close relationships of the Vietnamese population with other Asian populations, especially with populations in proximity. Conclusion The results from this study can contribute to the current genetic information content as a supplementary mtDNA reference dataset for forensic investigations and phylogenetic research.

The whole mitochondrial genome signature of Teressa goat, an indigenous goat germplasm of Andaman and Nicobar Islands, India

The domestic goat is a livestock species of global importance due to its ability to adapt to different agro-climatic conditions. The present study deals with the mitogenome signature of Teressa goat, a registered goat germplasm of Andaman and Nicobar Islands. Teressa goat mitogenome was 16,640 bp in length with 13 protein coding genes (PCGs), 22 tRNAs, 2 rRNAs and a non-coding region. Among the protein coding genes, only ND6 was present on light strand (L), other genes were encoded by heavy strand (H). All the PCGs had either ATG or ATA start codon whereas incomplete stop codons were detected in ND1, ND2, COX3, ND3 and ND4. Haplogroup assignment indicated that Teressa goat falls under Haplogroup A, most prevalent goat haplogroup. Mitogenome based phylogenetic analysis revealed that Teressa goat shared close genetic affinity with Xiangdong black goat, Longdong black goat, and Jintang black goat of China and Jamnapari goat of Bangladesh. The results of the study will be helpful to understand the origin and migration route of the goat breed and for sketching conservation plan of the indigenous breed.

Assessing the factors influencing the performance of machine learning for classifying haplogroups from Y-STR haplotypes

Two distinct genetic markers, single nucleotide polymorphisms (Y-SNPs) and short tandem repeats (Y-STRs), exist simultaneously in the non-recombining portion of the Y chromosome. Because of their different rates of mutation, Y-STRs and Y-SNPs play distinct roles in forensic and evolutionary genetics. Current approaches to infer haplogroup status rely on genotyping lots of Y-SNP loci. Given the relationship between haplotype and haplogroup of a Y chromosome, a cost-effective strategy of Y-STRs typing had an advantage in haplogroup prediction. Many machine learning algorithms have sprung up for assigning a Y-STR haplotype to a haplogroup. However, a series of issues must be solved before the using of machine learning method in practice. Thus, the k-nearest neighbor (kNN) classifier was built respectively based on different situations in this study. We assessed different factors which may influence the performance of the kNN prediction model for classifying haplogroups. The training set was based on a diverse ground-truth data set comprising Y-STR haplotypes and corresponding Y-SNP haplogroups. Our results showed that combining different levels of haplogroups into the observations or transracial prediction was impractical. Moreover, using more slow mutation Y-STR loci in the category is good for promoting classification accuracy. The preconditions for an effective and accurate haplogroup assignment by the kNN classifier were revealed.

Placing Ancient DNA Sequences into Reference Phylogenies.

Joint phylogenetic analysis of ancient DNA (aDNA) with modern phylogenies is hampered by low sequence coverage and post-mortem deamination, often resulting in overconservative or incorrect assignment. We provide a new efficient likelihood-based workflow, pathPhynder, that takes advantage of all the polymorphic sites in the target sequence. This effectively evaluates the number of ancestral and derived alleles present on each branch and reports the most likely placement of an ancient sample in the phylogeny and a haplogroup assignment, together with alternatives and supporting evidence. To illustrate the application of pathPhynder, we show improved Y chromosome assignments for published aDNA sequences, using a newly compiled Y variation data set (120,908 markers from 2,014 samples) that significantly enhances Y haplogroup assignment for low coverage samples. We apply the method to all published male aDNA samples from Africa, giving new insights into ancient migrations and the relationships between ancient and modern populations. The same software can be used to place samples with large amounts of missing data into other large non-recombining phylogenies such as the mitochondrial tree.

The role of mtDNA haplogroups on metabolic features in narcolepsy type 1

Narcolepsy type 1 (NT1) is due to selective loss of hypocretin (hcrt)-producing-neurons. Hcrt is a neuropeptide regulating the sleep/wake cycle, as well as feeding behavior. A subset of NT1 patients become overweight/obese, with a dysmetabolic phenotype. We hypothesized that mitochondrial DNA (mtDNA) sequence variation might contribute to the metabolic features in NT1 and we undertook an exploratory survey of mtDNA haplogroups in a cohort of well-characterized patients.We studied 246 NT1 Italian patients, fully defined for their metabolic features, including obesity, hypertension, low HDL, hypertriglyceridemia and hyperglycemia. For haplogroup assignment, the mtDNA control region was sequenced in combination with an assessment of diagnostic markers in the coding region.NT1 patients displayed the same mtDNA haplogroups (H, HV, J, K, T, U) frequency as those reported in the general Italian population. The majority of NT1 patients (64%) were overweight: amongst these, 35% were obese, 48% had low HDL cholesterol levels, and 31% had hypertriglyceridemia.We identified an association between haplogroups J, K and hypertriglyceridemia (P = 0.03, 61.5% and 61.5%, respectively vs. 31.3% of the whole sample) and after correction for age and sex, we observed a reduction of these associations (OR = 3.65, 95%CI = 0.76–17.5, p = 0.106 and 1.73, 0.52–5.69, p = 0.368, respectively). The low HDL level showed a trend for association with haplogroup J (P = 0.09, 83.3% vs. 47.4% of the whole sample) and after correction we observed an OR = 6.73, 95%CI = 0.65–69.9, p = 0.110.Our study provides the first indication that mtDNA haplogroups J and K can modulate metabolic features of NT1 patients, linking mtDNA variation to the dysmetabolic phenotype in NT1.

ФИЛОГЕНЕТИЧЕСКИЙ АНАЛИЗ ПОЛНЫХ ПОСЛЕДОВАТЕЛЬНОСТЕЙ МИТОХОНДРИАЛЬНОГО ГЕНОМА ОВЕЦ КУЙБЫШЕВСКОЙ ПОРОДЫ

Here, we present the results of a phylogenetic analysis of sheep of the Kuibyshev breed. Complete mitochondrial genome sequences of Russian sheep breed were obtained and then analyzed considering separate mitochondrial markers. The level of polymorphism in separate mitochondrial markers was established. Genetic diversity indicators were calculated for separate mitochondrial genes and for the complete mitochondrial genome. Based on the analysis, a phylogenetic tree was constructed, and the haplogroup assignment of the studied sheep was determined. Regular genetic monitoring will contribute to the accumulation of knowledge on the polymorphisms of all mitochondrial markers. Our study resulted in gaining new knowledge, which deepen the understanding of the role of mitochondrial DNA in the life functions support of sheep.

The performance of common SNP arrays in assigning African mitochondrial haplogroups

BackgroundMitochondrial haplogroup assignment is an important tool for forensics and evolutionary genetics. African populations are known to display a high diversity of mitochondrial haplogroups. In this research we explored mitochondrial haplogroup assignment in African populations using commonly used genome-wide SNP arrays.ResultsWe show that, from eight commonly used SNP arrays, two SNP arrays outperform the other arrays when it comes to the correct assignment of African mitochondrial haplogroups. One array enables the recognition of 81% of the African mitochondrial haplogroups from our compiled dataset of full mitochondrial sequences. Other SNP arrays were able to assign 4–62% of the African mitochondrial haplogroups present in our dataset. We also assessed the performance of available software for assigning mitochondrial haplogroups from SNP array data.ConclusionsThese results provide the first cross-checked quantification of mitochondrial haplogroup assignment performance from SNP array data. Mitochondrial haplogroup frequencies inferred from most common SNP arrays used for human population analysis should be considered with caution.

A globally diverse reference alignment and panel for imputation of mitochondrial DNA variants

BackgroundVariation in mitochondrial DNA (mtDNA) identified by genotyping microarrays or by sequencing only the hypervariable regions of the genome may be insufficient to reliably assign mitochondrial genomes to phylogenetic lineages or haplogroups. This lack of resolution can limit functional and clinical interpretation of a substantial body of existing mtDNA data. To address this limitation, we developed and evaluated a large, curated reference alignment of complete mtDNA sequences as part of a pipeline for imputing missing mtDNA single nucleotide variants (mtSNVs). We call our reference alignment and pipeline MitoImpute.ResultsWe aligned the sequences of 36,960 complete human mitochondrial genomes downloaded from GenBank, filtered and controlled for quality. These sequences were reformatted for use in imputation software, IMPUTE2. We assessed the imputation accuracy of MitoImpute by measuring haplogroup and genotype concordance in data from the 1000 Genomes Project and the Alzheimer’s Disease Neuroimaging Initiative (ADNI). The mean improvement of haplogroup assignment in the 1000 Genomes samples was 42.7% (Matthew’s correlation coefficient = 0.64). In the ADNI cohort, we imputed missing single nucleotide variants.ConclusionThese results show that our reference alignment and panel can be used to impute missing mtSNVs in existing data obtained from using microarrays, thereby broadening the scope of functional and clinical investigation of mtDNA. This improvement may be particularly useful in studies where participants have been recruited over time and mtDNA data obtained using different methods, enabling better integration of early data collected using less accurate methods with more recent sequence data.

Haplogroup Prediction Using Y-Chromosomal Short Tandem Repeats in the General Population of Bosnia and Herzegovina.

Human Y-chromosomal haplogroups are an important tool used in population genetics and forensic genetics. A conventional method used for Y haplogroup assignment is based on a set of Y-single nucleotide polymorphism (SNP) markers deployed, which exploits the low mutation rate nature of these markers. Y chromosome haplogroups can be successfully predicted from Y-short tandem repeat (STR) markers using different software packages, and this method gained much attention recently due to its labor-, time-, and cost-effectiveness. The present study was based on the analysis of a total of 480 adult male buccal swab samples collected from different regions of Bosnia and Herzegovina. Y haplogroup prediction was performed using Whit Athey’s Haplogroup Predictor, based on haplotype data on 23 Y-STR markers contained within the PowerPlex® Y23 kit. The results revealed the existence of 14 different haplogroups, with I2a, R1a, and E1b1b being the most prevalent with frequencies of 43.13, 14.79, and 14.58%, respectively. Compared to the previously published studies on Bosnian-Herzegovinian population based on Y-SNP and Y-STR data, this study represents an upgrade of molecular genetic data with a significantly larger number of samples, thus offering more accurate results and higher probability of detecting rare haplogroups.

Identification of Ajnala skeletal remains using multiple forensic anthropological methods and techniques: A bioarchaeological report

Fragmented and badly damaged commingled human remains present a tough challenge for their identification pursuits in forensic anthropology. Thousands of unknown human remains along with items of contextual identity, reportedly belonging to 282 Indian sepoys killed in 1857, were exhumed mon-scientifically from a disused ancient well at Ajnala (Amritsar, India). In this manuscript, the non-scientific excavation of unknown human remains from the abandoned well, their forensic anthropological strategies for identification purposes, challenges being faced and future possibilities of their biological profiling have been discussed. Multiple methods and techniques like anthropological examinations, odontological profiling, radiological analyses, stable isotope and mitochondrial DNA (mtDNA) analyses were applied to few bones and teeth collected from the Ajnala skeletal assemblage. Though majority of studied bones and teeth were found belonging to adult males, very few of them had morphological, osteological and molecular features questioning the authenticity and validity of the written records. Due to certain ambiguous findings or gaps observed between the anthropological analyses of the Ajnala skeletal remains and the reported versions about their affiliations; certain advanced radiological, chemical and molecular techniques were applied to estimate their probable age, sex and populational affinity. The obtained radiological, isotopic and molecular signatures of the remains were compared with the available databases to estimate their affinity with the individuals of geographic area to whom the remains reportedly belonged to. The maternally inherited mtDNA haplogroup assignments, and stable isotope analysis of carbon and oxygen suggested that the studied human remains belonged to the individuals from West Bengal, Bihar, Odisha, Awadh (presently in Uttar Pradesh) and parts of Meghalaya and Manipur as potential regions of their geographic identity and thus, attributing the victims to be non-local to the site. However, merely on the basis of forensic anthropological examinations of very few bones and teeth (collected out of a huge collection of thousands of bones and teeth); it would be just an unqualified and sweeping conclusion to claim their identity as adult or non-adult, male or female, local or non-local, victims of 1857 mass killings or to the victims of ceremonial sacrifices or criminal activities committed in the past. A sufficient number of bones and teeth along with items of personal identity needs to be examined with multiple scientific techniques to arrive at some valid conclusions about their biological identity.

A look of paternal ancestry in a sample of Ecuadorian “MESTIZO” population analyzed through PowerPlex Y23

Ecuadorian population is mainly composed of Mestizos, an ethnic group whose origins date back to admixture events in post-Columbian periods, including Amerindians, Europeans, and Africans. Hence, it is important to elucidate its genetic complexity thus increasing Ecuadorian registers in local and global databases, aiding in future forensic analysis and evolutionary studies. We analyzed 101 haplotypes of self-determined Mestizo male individuals by the PowerPlex Y23 System. Haplogroup assignment was evaluated by comparing three Y-STR haplogroup in silico algorithms. Ninety-nine haplotypes were found to be unique; therefore, the haplotype diversity for this population was 0.9998 +/− 0.0014. By means of haplogroup prediction we found native lineages (41%), and non-native-American lineages (59%). The comparison between predictors showed similar haplogroup assignments; however, some samples predictions were ambiguous.

Forensic Y-SNP analysis beyond SNaPshot: High-resolution Y-chromosomal haplogrouping from low quality and quantity DNA using Ion AmpliSeq and targeted massively parallel sequencing

Y-chromosomal haplogroups assigned from male-specific Y-chromosomal single nucleotide polymorphisms (Y-SNPs) allow paternal lineage identification and paternal bio-geographic ancestry inference, both being relevant in forensic genetics. However, most previously developed forensic Y-SNP tools did not provide Y haplogroup resolution on the high level needed in forensic applications, because the limited multiplex capacity of the DNA technologies used only allowed the inclusion of a relatively small number of Y-SNPs. In a proof-of-principle study, we recently demonstrated that high-resolution Y haplogrouping is feasible via two AmpliSeq PCR analyses and simultaneous massively parallel sequencing (MPS) of 530 Y-SNPs allowing the inference of 432 Y-haplogroups. With the current study, we present a largely improved Y-SNP MPS lab tool that we specifically designed for the analysis of low quality and quantity DNA often confronted with in forensic DNA analysis. Improvements include i) Y-SNP marker selection based on the “minimal reference phylogeny for the human Y chromosome” (PhyloTree Y), ii) strong increase of the number of targeted Y-SNPs allowing many more Y haplogroups to be inferred, iii) focus on short amplicon length enabling successful analysis of degraded DNA, and iv) combination of all amplicons in a single AmpliSeq PCR and simultaneous sequencing allowing single DNA aliquot use. This new MPS tool simultaneously analyses 859 Y-SNPs and allows inferring 640 Y haplogroups. Preliminary forensic developmental validation testing revealed that this tool performs highly accurate, is sensitive and robust. We also provide a revised software tool for analysing the sequencing data produced by the new MPS lab tool including final Y haplogroup assignment. We envision the tools introduced here for high-resolution Y-chromosomal haplogrouping to determine a man’s paternal lineage and/or paternal bio-geographic ancestry to become widely used in forensic Y-chromosome DNA analysis and other applications were Y haplogroup information from low quality / quantity DNA samples is required.

Genetic structure in the paternal lineages of South East Spain revealed by the analysis of 17 Y-STRs

The genetic data of 17 Y chromosome short tandem repeats in 146 unrelated donor residents in the provinces of Granada, Málaga, and Almería (GMA) were analyzed to determine the genetic legacy of the male inhabitants of the former Kingdom of Granada. A total of 139 unique haplotypes were identified. Observed allele frequencies and haplogroup frequencies were also analyzed. By AMOVA and STRUCTURE analysis, the populations of the 3 provinces could be treated genetically as a single population. The most frequent haplogroup was R1b1b2 (58.22%). By network analysis of all individuals, we observed a distribution according to haplogroup assignment. To improve the characterization of GMA population, it was compared with those of North Africa, the Iberian Peninsula, and southern Europe. In our analysis of allele frequencies and genetic distances, the GMA population lay within the Spanish population group. Further, in the STRUCTURE analysis, there was no African component in the GMA population, confirming that, based on our genetic markers, the GMA population does not reflect any male genetic influence of the North African people. The presence of African haplogroups in the GMA population is irrelevant when their frequency is compared with those in other European populations.

Yleaf: Software for Human Y-Chromosomal Haplogroup Inference from Next-Generation Sequencing Data.

Next-generation sequencing (NGS) technologies offer immense possibilities given the large genomic data they simultaneously deliver. The human Y-chromosome serves as good example how NGS benefits various applications in evolution, anthropology, genealogy, and forensics. Prior to NGS, the Y-chromosome phylogenetic tree consisted of a few hundred branches, based on NGS data, it now contains many thousands. The complexity of both, Y tree and NGS data provide challenges for haplogroup assignment. For effective analysis and interpretation of Y-chromosome NGS data, we present Yleaf, a publically available, automated, user-friendly software for high-resolution Y-chromosome haplogroup inference independently of library and sequencing methods.

Female-specific association among I, J and K mitochondrial genetic haplogroups and cancer: A longitudinal cohort study

Recent studies highlighted the role of mitochondrial dysregulation in cancer, suggesting that the different mitochondrial haplogroups might play a role in tumorigenesis and risk of cancer development. Our aim is to investigate whether any mitochondrial haplogroups carried a significant higher risk of cancer development in a large prospective cohort of North American people. The haplogroup assignment was performed by a combination of sequencing and PCR-RFLP techniques. Our specific outcome of interest was the incidence of any cancer during follow-up period. Overall, 3222 participants were included in the analysis. Women having I, J, K haplogroup reported a significant higher incidence of cancer compared to people with other haplogroups (p < 0.0001), whilst in men non association was found. In the multivariate analysis, women having I, J, K mitochondrial haplogroup reported a 50% increased risk of cancer (HR = 1.50; 95%CI: 1.04–2.16; p = 0.03). This gender-linked association may be partly explained by the role of mitochondrial function in female-specific (e.g. BRCA-driven) oncogenesis, but further studies are needed to better understand this potential correlation. Our findings may have important implications for cancer epidemiology and prevention.

Yleaf: Software for Human Y-Chromosomal Haplogroup Inference from Next-Generation Sequencing Data.

Next-generation sequencing (NGS) technologies offer immense possibilities given the large genomic data they simultaneously deliver. The human Y-chromosome serves as good example how NGS benefits various applications in evolution, anthropology, genealogy, and forensics. Prior to NGS, the Y-chromosome phylogenetic tree consisted of a few hundred branches, based on NGS data, it now contains many thousands. The complexity of both, Y tree and NGS data provide challenges for haplogroup assignment. For effective analysis and interpretation of Y-chromosome NGS data, we present Yleaf, a publically available, automated, user-friendly software for high-resolution Y-chromosome haplogroup inference independently of library and sequencing methods.

Multiple endocrine neoplasia 2 in Cyprus: evidence for a founder effect

PurposeMultiple endocrine neoplasia type 2 (MEN2) affects patients with RET proto-oncogene mutations. This cohort study refers to patients who were diagnosed with familial medullary thyroid carcinoma (MTC) and underwent RET genetic testing in Cyprus between years 2002 and 2017.Methods and patientsForty patients underwent RET testing by Sanger sequencing of exons 10–11 and 13–16. Genotyping with STR genetic markers flanking the RET gene along with Y-chromosome genotyping and haplogroup assignment was also performed.ResultsRET mutations were identified in 40 patients from 11 apparently unrelated Cypriot families and two non-familial sporadic cases. Nine probands (69.2%) were heterozygous for p.Cys618Arg, one (7.7%) for p.Cys634Phe, one (7.7%) for the somatic delE632-L633 and two (15.4%) for p.Met918Thr mutations. The mean age at MTC diagnosis of patients carrying p.Cys618Arg was 36.8 ± 14.2 years. The age of pheo diagnosis ranged from 26 to 43 years and appeared simultaneously with MTC in 5/36 (13.9%) cases. The high frequency of the p.Cys618Arg mutation suggested a possible ancestral mutational event. Haplotype analysis was performed in families with and without p.Cys618Arg. Six microsatellite markers covering the RET gene and neighboring regions identified one core haplotype associated with all patients carrying p.Cys618Arg mutation.ConclusionsThe mutation p.Cys618Arg is by far the most prevalent mutation in Cyprus followed by other reported mutations of variable clinical significance. The provided molecular evidence speculates p.Cys618Arg mutation as an ancestral mutation that has spread in Cyprus due to a possible founder effect.