Mitochondrial DNA Sequence Variation Research Articles

Mitochondrial DNA sequence variation is largely conserved at birth with rare de novo mutations in neonates

Mitochondrial DNA (mtDNA) encodes the proteins of the electron transfer chain to produce adenosine triphosphate through oxidative phosphorylation, and is essential to sustain life. mtDNA is unique from the nuclear genome in so much as it is solely maternally inherited (non-mendelian patterning), and shows a relatively high rate of mutation due to the absence of error checking capacity. While it is generally assumed that most new mutations accumulate through the process of heteroplasmy, it is unknown whether mutations initiated in the mother are inherited, occur in utero, or occur and accumulate early in life. The purpose of this study is to examine the maternally heritable and de novo mutation rate in the fetal mtDNA through high-fidelity sequencing from a large population-based cohort. Samples were obtained from 90 matched maternal (blood) and fetal (placental) pairs. In addition, a smaller cohort (n= 5) of maternal (blood), fetal (placental), and neonatal (cord blood) trios were subjected to DNA extraction and shotgun sequencing. The whole genome was sequenced on the Illumina HiSeq platform (Illumina Inc., San Diego, CA), and haplogroups and mtDNA variants were identified through mapping to reference mitochondrial genomes (NC_012920). We observed 665 single nucleotide polymorphisms and 82 insertions-deletions variants identified in the cohort at large. We achieved high sequencing depth of the mtDNA to an average depth of 65X (range, 20-171X) coverage. The proportions of haplogroups identified in the cohort are consistent with the patient's self-identified ethnicity (>90% Hispanic), and all maternal-fetal pairs mapped to the identical haplogroup. Only variants from samples with average depth >20X and allele frequency >1% were included for further analysis. While the majority of the maternal-fetal pairs (>90%) demonstrated identical variants at the single nucleotide level, we observed rare mitochondrial single nucleotide polymorphism discordance between maternal and fetal mitochondrial genomes. In this first in-depth sequencing analysis of mtDNA from maternal-fetal pairs at the time of birth, a low rate of de novo mutations appears in the fetal mitochondrial genome. This implies that these mutations likely arise from the maternal heteroplasmic pool (eg, in the oocyte), and accumulate later in the offspring's life. These findings have key implications for both the occurrence and screening for mitochondrial disorders.

Evidence of pre-introduction hybridization of Formosan sika deer (Cervus nippon taiouanus) on Okinoshima, Wakayama Prefecture, Japan, based on mitochondrial and nuclear DNA sequences

To examine the origin of the Formosan sika deer (Cervus nippon taiouanus) introduced to the island of Okinoshima (Wakayama Prefecture, Japan), sixteen tissue samples from Okinoshima population were analyzed to determine sequence variation in maternal mitochondrial DNA (mtDNA) cytochrome b and nuclear DNA α-lactalbumin (αLAlb). Three cytochrome b haplotypes were found in the Okinoshima population, two Formosan sika deer haplotypes and one Formosan sambar (C. unicolor swinhoei) haplotype. Three nuclear DNA αLAlb sequences, i.e. Formosan sika deer, red deer (C. elaphus), and additional one unknown sequence were detected in the population. Cytonuclear disequilibrium was observed in five individuals, indicating that the Okinoshima population have occurrence of hybrid among the three deer species. The deer introduced on Okinoshima might have originated from a private deer-breeding facility in Taiwan. To avoid native Japanese sika deer (e.g. C. n. centralis) genetic introgression, the Okinoshima population should be isolated from native Japanese sika deer populations.

Mitochondrial DNA Sequence Variation Associated With Peripheral Nerve Function in the Elderly.

Mitochondrial dysfunction is a prominent hallmark of many sensory neuropathies. The purpose of this study was to assess the influence of mitochondrial DNA sequence variation on peripheral nerve function in the population-based Health, Aging, and Body Composition Study. We investigated the role of common mitochondrial DNA variation (n = 1,580) and complete mitochondrial DNA sequences (n = 138) on peroneal motor nerve conduction velocity and amplitude, average vibration detection threshold, and monofilament sensitivity. Nominal associations among common mitochondrial DNA variants and haplogroups were identified but were not statistically significant after adjustment for multiple comparisons. Sequence-based approaches were used to identify aggregate variant associations across the 16S rRNA (weighted-sum, p = 2E-05 and variable threshold, p = 9E-06) for nerve conduction velocity. Several of these rare 16S variants occurred at or near sites with earlier disease associations and are also in close proximity to the peptidyl transferase center, which is the catalytic center of the 16S rRNA CONCLUSIONS: These results suggest that sequence variation related to mitochondrial protein synthesis/assembly is associated with peripheral nerve function and may provide insight into targets for intervention or new clinical strategies to preserve nerve function in late life.

Genetic Divergence of Turkish Apis mellifera Subspecies Based on Sequencing of ND5 Mitochondrial Segment

Mitochondrial DNA sequence variation can be used to infer honey bee evolutionary relationships. In this study, DNA sequence diversity in the ND5 region of the mitochondrial genome was investigated in 93 samples of Apis mellifera from 15 different populations in Turkey. Five novel haplotypes were revealed for the ND5 gene segment of Turkish honeybees. The number of variable sites found was 6 for this region while 2 were parsimony informative sites. The average pairwise genetic distances were 0.3% for ND5 gene. In this study, the NJ tree of ND5 gene segment were constructed with the published sequences of Apis mellifera haplotypes. This study expands the knowledge about the mitochondrial ND5 region in Apis mellifera and it is also the first comprehensive sequencing analysis of ND5 region in Turkish honeybees.

Mitochondrial DNA haplogroup B5 confers genetic susceptibility to Alzheimer's disease in Han Chinese

Mitochondrial dysfunction has been widely reported in psychiatric and neurodegenerative diseases. We aimed to investigate the association between matrilineal structures of Han Chinese populations and Alzheimer's disease (AD) by a 2-stage case-control study: A total of 341 AD patients and 435 normal individuals from Southwest China were analyzed for mitochondrial DNA sequence variations and were classified into respective haplogroups. A total of 371 AD patients and 470 normal individuals from East China, as validation samples, were genotyped for the variants defining the risk haplogroup. Haplogroup B5 had a significantly higher frequency in AD patients (7.33%) than in control subjects (3.68%) from Southwest China, and we found a similar pattern of higher frequency of B5 in patients in the case-control sample from East China. In the combined population, association of haplogroup B5 with AD risk was strengthened (p = 0.02; odds ratio = 1.74; 95% confidence interval = 1.10–2.76). In lymphoblastoid cell lines belonging to haplogroup B5a, we observed significantly increased reactive oxygen species and decreased mitochondrial mass. Hela cells with stable expression of the MT-ATP6 gene with B5-defining variant m.8584G>A also showed a significantly decreased mitochondrial function. Taken together, our results indicated that haplogroup B5 conferred genetic susceptibility to AD in Han Chinese, and this effect was most likely mediated by ancient variant m.8584G>A. The predisposing effect of B5 to AD is consistent with the ancestral-susceptibility model of complex diseases.

Quantitative pedigree analysis and mitochondrial DNA sequence variants in adults with cyclic vomiting syndrome.

BackgroundChildren with cyclic vomiting syndrome (CVS) have a high degree of maternal inheritance of functional gastrointestinal and neurological disorders. CVS in children is also associated with an increased prevalence of mitochondrial DNA single-nucleotide polymorphisms (mtDNA SNPs) 16519 T and 3010A. Preliminary data suggests that age of onset of symptoms (pediatric vs. adult) may be a determinant of the presence of such mtDNA SNP’s. We sought to examine the degree of maternal inheritance pattern of functional disorders and the prevalence of mtDNA SNP’s16519T and 3010A in adults with CVS and correlate this with age of onset of disease.MethodsA Quantitative Pedigree Analysis (QPA) was performed in 195 of a total of 216 patients and all were genotyped using Restriction Fragment Length Polymorphism (RFLP) or sequencing.ResultsAdults with CVS had a higher degree of probable maternal inheritance (PMI) of functional disorders than controls (12% vs. 1%, p < 0.001). However, the prevalence of mitochondrial SNP’s 16519 T, 3010A and the AT genotype were similar in Haplogroup H CVS patients compared to historical controls. There was no correlation between age of onset of disease and prevalence of these mtDNA SNP’s.ConclusionsA subset of adults with CVS has a significantly higher degree of maternal inheritance pattern of functional disorders than controls. There was no association with mtDNA SNP’s 16519 T and 3010A as seen in children and future studies sequencing the entire mitochondrial and nuclear genome to identify potential causes for this maternal inheritance pattern in adults are warranted.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-230X-14-181) contains supplementary material, which is available to authorized users.

Quantitative pedigree analysis and mitochondrial DNA sequence variants in adults with cyclic vomiting syndrome

Quantitative pedigree analysis and mitochondrial DNA sequence variants in adults with cyclic vomiting syndrome

Phylogeography and evolution of three closely related species of Tsuga (hemlock) from subtropical eastern Asia: further insights into speciation of conifers

AbstractAimThe biogeography and speciation of conifers are of special interest because of the ecological importance of these trees, but the boundaries and evolutionary relationships of closely related species of conifers are difficult to disentangle. To investigate how lineage sorting and gene flow affect species boundaries in conifers, we studied the population genetic structure and phylogeographical history of the Tsuga chinensis–T. forrestii complex and compared the results with those for a closely related species, T. dumosa.LocationSubtropical eastern Asia.MethodsPhylogeographical analyses, in combination with coalescent simulations, were performed based on sequence variation of paternal chloroplast and maternal mitochondrial DNA in 1368 individuals from 52 populations of the three Tsuga (hemlock) species.ResultsMitotypes were shared extensively among the three species while the chlorotypes were relatively more species‐specific. Two genetic breaks and thus three distinct geographical groups were identified in the T. chinensis–T. forrestii species pair. Unidirectional pollen flow from T. forrestii to T. chinensis was detected.Main conclusionsTsuga forrestii originated from a pre‐existing isolate of T. chinensis, and a cryptic genetic break formed between eastern and western populations of the latter. Incomplete lineage sorting was responsible for the extensive sharing of mitotypes among the three hemlock species, and our results seem to support the idea that paternally inherited markers are more effective in delimiting species than maternally inherited markers. We also found that hemlock populations in the Himalayas were very likely to have experienced extinction, but that most populations in other areas of subtropical mainland China survived the late Quaternary glacial cycles by elevational shifts.

Islands in ice: isolated populations of Cryptopygus sverdrupi (Collembola) among nunataks in the Sør Rondane Mountains, Dronning Maud Land, Antarctica

Molecular variation among Antarctic Collembola has been well appreciated over the last decade. The majority of studies have focussed on the Transantarctic Mountains and Antarctic Peninsula, and more recently Dronning Maud Land, which now accounts for all known continental species. Within most species, mean mitochondrial DNA (mtDNA) sequence divergence has not been more than around 2%, but in 2010 far greater levels of infraspecific sequence divergence (9.2–10.9%) were discovered in three species: Friesea grisea (10.2%), Gressittacantha terranova (10.4%) and Cryptopygus antarcticus (9.2%). Here, we present the first phylogeographic study on Cryptopygus sverdrupi from Dronning Maud Land. We found that mtDNA COI and 16S haplotypes clustered into two lineages with a mean COI sequence divergence of around 7.1%. The mixing of haplotypes between nearby nunataks separated by up to 15 km revealed support for ongoing (but rare) dispersal, although a single site, the most geographically isolated (by 40–58 km) was also the most genetically divergent. These levels of sequence divergence indicate persistence of biota throughout the Miocene and Pliocene isolated in glacial refugia. Expanding sampling of C. sverdrupi from additional ice-free refugia throughout Dronning Maud Land will be important to further our understanding of the evolutionary processes that have influenced the Antarctic continent, its landscape and biota.

Cytonuclear discordance and historical demography of two brown frogs, Rana tagoi and R. sakuraii (Amphibia: Ranidae)

Prior studies of mitochondrial genomic variation reveal that the Japanese brown frog Rana tagoi comprises a complex of cryptic species lineages, and that R. sakuraii arose from within this complex. Neither species forms a monophyletic group on the mitochondrial haplotype tree, precluding a simple explanation for the evolutionary origins of R. sakuraii. We present a more complete sampling of mitochondrial haplotypic variation (from the ND1 and 16S genes) plus DNA sequence variation for five nuclear loci (from the genes encoding NCX1, NFIA, POMC, SLC8A3, and TYR) to resolve the evolutionary histories of these species. We test hypotheses of population assignment (STRUCTURE) and isolation-with-migration (IM) using the more slowly evolving nuclear markers. These demographic analyses of nuclear genetic variation confirm species-level distinctness and integrity of R. sakuraii despite its apparent polyphyly on the mitochondrial haplotype tree. Divergence-time estimates from both the mitochondrial haplotypes and nuclear genomic markers suggest that R. sakuraii originated approximately one million years ago, and that incomplete sorting of mitochondrial haplotype lineages best explains non-monophyly of R. sakuraii mitochondrial haplotypes. Cytonuclear discordance elsewhere in R. tagoi reveals a case of mitochondrial introgression between two species lineages on Honshu. The earliest phylogenetic divergence within this species group occurred approximately four million years ago, followed by cladogenetic events in the Pliocene and early Pleistocene yielding 10–13 extant species lineages, including R. sakuraii as one of the youngest.

A Matrilineal Genetic Legacy from the Last Glacial Maximum Confers Susceptibility to Schizophrenia in Han Chinese

Mitochondrial dysfunction has been widely reported in schizophrenia patients. To dissect the matrilineal structure of Han Chinese with or without schizophrenia and to decipher the maternal influence and evolutionary history of schizophrenia, a total of 1212 schizophrenia patients and 1005 matched healthy controls, all of Han Chinese origin, were recruited in Hunan Province, China. We classified haplogroup for each individual based on mitochondrial DNA (mtDNA) sequence variations and compared the haplogroup distribution pattern between cases and controls. Haplogroup B5a presented a higher frequency in cases than in controls (P = 0.02, OR = 1.67, 95% CI = [1.09, 2.56]), and this result could be confirmed by permutation analysis. Age estimation of haplogroup B5a in cases revealed a much younger age than that of controls, which was coincident with the Northern Hemisphere deglaciation at the end of the Last Glacial Maximum. Analysis of complete mtDNA in five patients belonging to haplogroup B5a showed that this background effect might be caused by haplogroup-defining variants m.8584G>A and m.10398A>G. Our results showed that matrilineal risk factor for schizophrenia had an ancient origin and might acquire a predisposing effect on schizophrenia due to the environment change and/or orchestration with other nuclear genetic factors appeared recently in human evolutionary history.

Conservation implications of the evolutionary history and genetic diversity hotspots of the snowshoe hare.

With climate warming, the ranges of many boreal species are expected to shift northward and to fragment in southern peripheral ranges. To understand the conservation implications of losing southern populations, we examined range-wide genetic diversity of the snowshoe hare (Lepus americanus), an important prey species that drives boreal ecosystem dynamics. We analysed microsatellite (8 loci) and mitochondrial DNA sequence (cytochrome b and control region) variation in almost 1000 snowshoe hares. A hierarchical structure analysis of the microsatellite data suggests initial subdivision in two groups, Boreal and southwestern. The southwestern group further splits into Greater Pacific Northwest and U.S. Rockies. The genealogical information retrieved from mtDNA is congruent with the three highly differentiated and divergent groups of snowshoe hares. These groups can correspond with evolutionarily significant units that might have evolved in separate refugia south and east of the Pleistocene ice sheets. Genetic diversity was highest at mid-latitudes of the species' range, and genetic uniqueness was greatest in southern populations, consistent with substructuring inferred from both mtDNA and microsatellite analyses at finer levels of analysis. Surprisingly, snowshoe hares in the Greater Pacific Northwest mtDNA lineage were more closely related to black-tailed jackrabbits (Lepus californicus) than to other snowshoe hares, which may result from secondary introgression or shared ancestral polymorphism. Given the genetic distinctiveness of southern populations and minimal gene flow with their northern neighbours, fragmentation and loss of southern boreal habitats could mean loss of many unique alleles and reduced evolutionary potential.

A review of the genus Pempheris (Perciformes, Pempheridae) of the Red Sea, with description of a new species.

Four species of the fish genus Pempheris are recognized for the Red Sea: P. adusta Bleeker, 1877; P. mangula Cuvier, 1829; P. nesogallica Cuvier in Cuvier & Valenciennes, 1831; and a new species P. tominagai. All are wide-ranging in the western Indian Ocean, and P. mangula has migrated via the Suez Canal to the eastern Mediterranean Sea. Morphological and genetic analysis of 15 species in this genus show that P. adusta, a widely distributed species, that can't be divided into different species, because of the continuity of morphologies and distribution, and lack of variance in genetics between Indian Ocean, Red Sea, and Pacific Ocean populations. This confirms that the two subspecies described by Randall et al. (2013) are both synonyms of P. adusta. Pempheris adusta is distinguished from other species by a blackish spot on pectoral fin base, pored lateral-line scales 56-64, scale rows above lateral line 4 1/2-6 1/2, distinct blackish band on outer edge of anal fin, and blackish band on posterior edge of caudal fin. Pempheris mangula was named by Cuvier (1829) in a footnote making reference to a drawing and short description in Russell (1803) of a Pempheris from southeast India, giving only the native name ''Mangula-Kutti'', and listing no specimen. The wide distribution of this species, from the Indian Ocean to the Red Sea is also demonstrated by morphological and genetic analysis. Thus, the specimen collected from southern India is herein designated as the neotype. This species is distinguished from other species by its huge eye, deep body, blackish tip of the dorsal fin, pored lateral-line scales 49-60, and scale rows above lateral line 4 1/2-5 1/2. The extant syntype of Kossmann & Räuber's P. rhomboidea is designated as the lectotype of the species; however, P. rhomboidea is a synonym of P. mangula. In addition, Kossmann & Räuber's Pempheris erythraea and P. russellii Day, 1888 are also synonyms of P. mangula. Of two existing syntypes of P. nesogallica from Mauritius, one is designated as the lectotype, the other is re-identified as P. mangula; P. nesogallica is presently known only from the southern Red Sea. This species has a similar morphology to P. mangula, but can be distinguished by a smaller eye than P. mangula, and lack irregular faint longitudinal light stripes on the body side. Pempheris tominagai are distinguished from P. schwenkii Bleeker 1855, formerly misidentified, by the form of posterior nostril, scale counts, color of caudal fin, and by a 2.1% mitochondrial DNA sequence divergence.

Mitochondrial DNA Variant in COX1 Subunit Significantly Alters Energy Metabolism of Geographically Divergent Wild Isolates in Caenorhabditis elegans

Mitochondrial DNA (mtDNA) sequence variation can influence the penetrance of complex diseases and climatic adaptation. While studies in geographically defined human populations suggest that mtDNA mutations become fixed when they have conferred metabolic capabilities optimally suited for a specific environment, it has been challenging to definitively assign adaptive functions to specific mtDNA sequence variants in mammals. We investigated whether mtDNA genome variation functionally influences Caenorhabditis elegans wild isolates of distinct mtDNA lineages and geographic origins. We found that, relative to N2 (England) wild-type nematodes, CB4856 wild isolates from a warmer native climate (Hawaii) had a unique p.A12S amino acid substitution in the mtDNA-encoded COX1 core catalytic subunit of mitochondrial complex IV (CIV). Relative to N2, CB4856 worms grown at 20°C had significantly increased CIV enzyme activity, mitochondrial matrix oxidant burden, and sensitivity to oxidative stress but had significantly reduced lifespan and mitochondrial membrane potential. Interestingly, mitochondrial membrane potential was significantly increased in CB4856 grown at its native temperature of 25°C. A transmitochondrial cybrid worm strain, chpIR (M, CB4856>N2), was bred as homoplasmic for the CB4856 mtDNA genome in the N2 nuclear background. The cybrid strain also displayed significantly increased CIV activity, demonstrating that this difference results from the mtDNA-encoded p.A12S variant. However, chpIR (M, CB4856>N2) worms had significantly reduced median and maximal lifespan relative to CB4856, which may relate to their nuclear–mtDNA genome mismatch. Overall, these data suggest that C. elegans wild isolates of varying geographic origins may adapt to environmental challenges through mtDNA variation to modulate critical aspects of mitochondrial energy metabolism.

Significant population genetic structuring but a lack of phylogeographic structuring in the endemic Tasmanian tree frog (Litoria burrowsae)

Conservation of frogs is of global concern, owing to declines resulting from habitat destruction, global climate change, and disease. Knowledge of genetic variation in frog species is therefore desirable for the identification of management units. Here we surveyed mitochondrial DNA sequence variation in the Tasmanian endemic hylid frog Litoria burrowsae, which is infected by chytrid fungus, Batrachochytrium dendrobatidis, and may be declining. Neither phylogeographic structure nor deep phylogenetic divergence was detected in the species, although its populations were highly differentiated with respect to haplotype frequencies. The low-haplotype diversity in L. burrowsae suggests a recent bottleneck in the species, and population genetic structuring may reflect isolation by distance as well as founder effects associated with range expansion. Three putative management units were identified that require verification based on nuclear DNA variation and adaptation to local environments.

Collembola, the biological species concept and the underestimation of global species richness

Despite its ancient origin, global distribution and abundance in nearly all habitats, the class Collembola is comprised of only 8000 described species and is estimated to number no more than 50,000. Many morphologically defined species have broad geographical ranges that span continents, and recent molecular work has revealed high genetic diversity within species. However, the evolutionary significance of this genetic diversity is unknown. In this study, we sample five morphological species of the globally distributed genus Lepidocyrtus from 14 Panamanian sampling sites to characterize genetic diversity and test morphospecies against the biological species concept. Mitochondrial and nuclear DNA sequence data were analysed and a total of 58 molecular lineages revealed. Deep lineage diversification was recovered, with 30 molecular lineages estimated to have established more than 10 million years ago, and the origin almost all contemporary lineages preceding the onset of the Pleistocene (~2 Mya). Thirty-four lineages were sampled in sympatry revealing unambiguous cosegregation of mitochondrial and nuclear DNA sequence variation, consistent with biological species. Species richness within the class Collembola and the geographical structure of this diversity are substantially misrepresented components of terrestrial animal biodiversity. We speculate that global species richness of Collembola could be at least an order of magnitude greater than a previous estimate of 50,000 species.

Germline mitochondrial DNA mutations aggravate ageing and can impair brain development

Ageing is due to an accumulation of various types of damage, and mitochondrial dysfunction has long been considered to be important in this process. There is substantial sequence variation in mammalian mitochondrial DNA (mtDNA), and the high mutation rate is counteracted by different mechanisms that decrease maternal transmission of mutated mtDNA. Despite these protective mechanisms, it is becoming increasingly clear that low-level mtDNA heteroplasmy is quite common and often inherited in humans. We designed a series of mouse mutants to investigate the extent to which inherited mtDNA mutations can contribute to ageing. Here we report that maternally transmitted mtDNA mutations can induce mild ageing phenotypes in mice with a wild-type nuclear genome. Furthermore, maternally transmitted mtDNA mutations lead to anticipation of reduced fertility in mice that are heterozygous for the mtDNA mutator allele (PolgA(wt/mut)) and aggravate premature ageing phenotypes in mtDNA mutator mice (PolgA(mut/mut)). Unexpectedly, a combination of maternally transmitted and somatic mtDNA mutations also leads to stochastic brain malformations. Our findings show that a pre-existing mutation load will not only allow somatic mutagenesis to create a critically high total mtDNA mutation load sooner but will also increase clonal expansion of mtDNA mutations to enhance the normally occurring mosaic respiratory chain deficiency in ageing tissues. Our findings suggest that maternally transmitted mtDNA mutations may have a similar role in aggravating aspects of normal human ageing.

Low Mitochondrial Dna Sequence Variation in the Microendemic Cuatro Ciénegas PlatyfishXiphophorus gordoni

The desert spring ecosystem of the Cuatro Ciénegas basin, Mexico, hosts an extraordinary but increasingly imperiled assemblage of native and endemic aquatic organisms. Among the more vulnerable to extinction is the microendemic Cuatro Ciénegas platyfish Xiphophorus gordoni, which is at particular risk due to its highly restricted distribution and small population size. Given the growing demands on water resources in this arid region, we conducted a survey of mitochondrial DNA sequence variation in the cytochrome b gene to establish a baseline for long-term genetic monitoring of this endangered species. Fifty-eight specimens were collected from the 2 subpopulations known to persist, Laguna Santa Tecla (27) and Teclitas (31). Three haplotypes were observed across all samples, with 3 and 2 haplotypes observed at Laguna Santa Tecla and Teclitas, respectively. Sequence divergence ranged from 0.02% to 0.03% among variants and averaged 0.317 differences between populations. Haplotype and nucleotide diversities were moderately low in Laguna Santa Tecla (0.467 and 0.0008, respectively) and severely low in Teclitas (0.065 and 0.0001, respectively). Exact tests revealed significant heterogeneity in haplotype distributions between sites (P = 0.006), as did the pairwise estimate of ΦST (0.135; P = 0.005). Tests of deviation from neutrality were nonsignificant. Microendemism of X. gordoni and marginal substitution differences between the few extant haplotypes suggest that low genetic variation is a historical characteristic of this species; however, population declines associated with recent habitat loss have likely further compromised standing levels of genetic variation. Development of a long-term population and genetic monitoring program that incorporates multilocus nuclear DNA markers is advised and should be tied to a more thorough characterization of the basic biology and life history of this species, establishment of captive or refuge populations, and further improvement of habitat.

Mitochondrial DNA sequence divergence between “Kunimasu” Oncorhynchus kawamurae and “Himemasu” O. nerka in Lake Saiko, Yamanashi Prefecture, Japan, and their identification using multiplex haplotype-specific PCR

In order to develop a simplified method for discriminating between Oncorhynchus kawamurae (Kunimasu) and sympatric Oncorhynchus nerka (Himemasu), sequence variations of mitochondrial DNA control region were examined on 71 specimens of Kunimasu and 103 of Himemasu. Sequence analyses for 530 bp revealed 3 variable sites, which defined three haplotypes in Kunimasu and one in Himemasu. Based on determined nucleotide sequences, species-specific primers were designed for the multiplex haplotype-specific polymerase chain reaction, which amplified ca. 900 and 1,040 bp fragments in Kunimasu and Himemasu, respectively. This simple rapid approach can be used to distinguish between the two species without the need to sacrifice specimens.

Mitochondrial DNA sequence variation in whiting Merlangius merlangus in the North East Atlantic

Genetic variation in whiting Merlangius merlangus was examined using a 621 base pair fragment of the cytochrome c oxidase subunit I mitochondrial gene in 138 individuals sampled from Iceland, Norway and the North Sea. In total 10 segregating sites were observed defining 12 haplotypes. Three of the haplotypes were found at high frequencies (>5 %). All but one mutations were synonymous and the nonsynonymous mutation was found as a singleton. This suggests weak or no natural selection acting on the observed polymorphism making it useful for examination of population breeding structure. The genetic variation suggests that the whiting population has undergone sudden expansion in the past, estimated to have started 70 Kyr ago, during the last glacial period. Spatial genetic analysis reveals genetic uniformity across long geographic distances suggesting high level of gene flow. The long pelagic phase at early age, allowing for high dispersal rate, may partly explain the observed pattern.