Exploring statistical weight estimates for mitochondrial DNA matches involving heteroplasmy.

Massively parallel sequencing (MPS) of mitochondrial (mt) DNA allows forensic laboratories to report heteroplasmy on a routine basis. Statistical approaches will be needed to determine the relative frequency of observing an mtDNA haplotype when including the presence of a heteroplasmic site. Here, we examined 1301 control region (CR) sequences, collected from individuals in four major population groups (European, African, Asian, and Latino), and covering 24 geographically distributed haplogroups, to assess the rates of point heteroplasmy (PHP) on an individual and nucleotide position (np) basis. With a minor allele frequency (MAF) threshold of 2%, the data was similar across population groups, with an overall PHP rate of 37.7%, and the majority of heteroplasmic individuals (77.3%) having only one site of heteroplasmy. The majority (75.2%) of identified PHPs had an MAF of 2-10%, and were observed at 12.6% of the nps across the CR. Both the broad and phylogenetic testing suggested that in many cases the low number of observations of heteroplasmy at any one np results in a lack of statistical association. The posterior frequency estimates, which skew conservative to a degree depending on the sample size in a given haplogroup, had a mean of 0.152 (SD 0.134) and ranged from 0.031 to 0.83. As expected, posterior frequency estimates decreased in accordance with 1/n as the sample size (n) increased. This provides a proposed conservative statistical framework for assessing haplotype/heteroplasmy matches when applying an MPS technique in forensic cases and will allow for continual refinement as more data is generated, both within the CR and across the mitochondrial genome.

Bacterial Septic Arthritis of the Adult Native Knee Joint: A Review.

¼ Acute bacterial septic arthritis of the knee is an orthopaedic emergency and, if left untreated, can result in substantial joint degradation. ¼ Important risk factors for development of septic arthritis include age of >60 years, recent bacteremia, diabetes, cancer, cirrhosis, renal disease, drug or alcohol abuse, a history of corticosteroid injection, a recent injury or surgical procedure, a prosthetic joint, and a history of rheumatoid arthritis. ¼ The diagnosis is primarily based on history and clinical presentation of a red, warm, swollen, and painful joint with limited range of motion. Laboratory values and inflammatory markers from serum and joint fluid may serve as adjuncts when there is clinical suspicion of septic arthritis. ¼ The initial and general antibiotic regimen should cover methicillin-resistant Staphylococcus aureus and gram-negative and gram-positive organisms. The antibiotic regimen should be specified following the culture results of the infected joint. ¼ Operative management involves either arthrotomy or arthroscopy of the knee with thorough irrigation and debridement of all infected tissue. The Gächter classification is useful in establishing a prognosis or in determining the need for an extensive debridement.

External fixator-augmented flexible intramedullary nailing of an unstable pediatric femoral shaft fracture model: a biomechanical study.

The objective of this study was to test the compressive strength and torsional stiffness provided by the addition of a two-pin external fixator to an unstable pediatric femoral shaft fracture model after being instrumented with flexible intramedullary nailing (FIMN), and to compare this to bridge plating and FIMN alone. A length-unstable oblique diaphyseal fracture was created in 15 pediatric sized small femur models. Fracture stabilization was achieved by three constructs: standard retrograde FIMN with two 3.5-mm titanium (Ti) nails (Group 1), FIMN augmented with a two-pin external fixator (Group 2), and a 4.5-mm bridge plate (Group 3). Groups I and II were tested in 10 cycles of axial rotation to 10° in both directions at 0.1 Hz under 36 kg of compression. Torsional stiffness was calculated. Compressive strength was calculated by applying an axial load of 5 mm/min until failure was encountered. Failure was defined as the force required to achieve 10° varus at the fracture site or shortening of 2 cm. Group II demonstrated a greater compressive strength compared to Group I (1067.32 N vs 453.49 N, P < 0.001). No significant difference in torsional stiffness was found between Groups I and II (0.45 vs 0.38 Nm/deg, P = 0.18). Group III showed superior compressive strength and rotational stiffness compared to Groups I and II. In an unstable pediatric femoral shaft fracture model, augmenting FIMN with a two-pin external fixator increased the compressive strength by 147%, but did not increase torsional stiffness. Bridge plating with a 4.5-mm plate provided superior compressive strength and torsional stiffness.

Pronounced somatic bottleneck in mitochondrial DNA of human hair.

Heteroplasmy is the presence of variable mitochondrial DNA (mtDNA) within the same individual. The dynamics of heteroplasmy allele frequency among tissues of the human body is not well understood. Here, we measured allele frequency at heteroplasmic sites in two to eight hairs from each of 11 humans using next-generation sequencing. We observed a high variance in heteroplasmic allele frequency among separate hairs from the same individual-much higher than that for blood and cheek tissues. Our population genetic modelling estimated the somatic bottleneck during embryonic follicle development of separate hairs to be only 11.06 (95% confidence interval 0.6-34.0) mtDNA segregating units. This bottleneck is much more drastic than somatic bottlenecks for blood and cheek tissues (136 and 458 units, respectively), as well as more drastic than, or comparable to, the germline bottleneck (equal to 25-32 or 7-10 units, depending on the study). We demonstrated that hair undergoes additional genetic drift before and after the divergence of mtDNA lineages of individual hair follicles. Additionally, we showed a positive correlation between donor's age and variance in heteroplasmy allele frequency in hair. These findings have important implications for forensics and for our understanding of mtDNA dynamics in the human body. This article is part of the theme issue 'Linking the mitochondrial genotype to phenotype: a complex endeavour'.

Bottleneck and selection in the germline and maternal age influence transmission of mitochondrial DNA in human pedigrees.

Heteroplasmy-the presence of multiple mitochondrial DNA (mtDNA) haplotypes in an individual-can lead to numerous mitochondrial diseases. The presentation of such diseases depends on the frequency of the heteroplasmic variant in tissues, which, in turn, depends on the dynamics of mtDNA transmissions during germline and somatic development. Thus, understanding and predicting these dynamics between generations and within individuals is medically relevant. Here, we study patterns of heteroplasmy in 2 tissues from each of 345 humans in 96 multigenerational families, each with, at least, 2 siblings (a total of 249 mother-child transmissions). This experimental design has allowed us to estimate the timing of mtDNA mutations, drift, and selection with unprecedented precision. Our results are remarkably concordant between 2 complementary population-genetic approaches. We find evidence for a severe germline bottleneck (7-10 mtDNA segregating units) that occurs independently in different oocyte lineages from the same mother, while somatic bottlenecks are less severe. We demonstrate that divergence between mother and offspring increases with the mother's age at childbirth, likely due to continued drift of heteroplasmy frequencies in oocytes under meiotic arrest. We show that this period is also accompanied by mutation accumulation leading to more de novo mutations in children born to older mothers. We show that heteroplasmic variants at intermediate frequencies can segregate for many generations in the human population, despite the strong germline bottleneck. We show that selection acts during germline development to keep the frequency of putatively deleterious variants from rising. Our findings have important applications for clinical genetics and genetic counseling.

An approximate full-likelihood method for inferring selection and allele frequency trajectories from DNA sequence data.

Most current methods for detecting natural selection from DNA sequence data are limited in that they are either based on summary statistics or a composite likelihood, and as a consequence, do not make full use of the information available in DNA sequence data. We here present a new importance sampling approach for approximating the full likelihood function for the selection coefficient. Our method CLUES treats the ancestral recombination graph (ARG) as a latent variable that is integrated out using previously published Markov Chain Monte Carlo (MCMC) methods. The method can be used for detecting selection, estimating selection coefficients, testing models of changes in the strength of selection, estimating the time of the start of a selective sweep, and for inferring the allele frequency trajectory of a selected or neutral allele. We perform extensive simulations to evaluate the method and show that it uniformly improves power to detect selection compared to current popular methods such as nSL and SDS, and can provide reliable inferences of allele frequency trajectories under many conditions. We also explore the potential of our method to detect extremely recent changes in the strength of selection. We use the method to infer the past allele frequency trajectory for a lactase persistence SNP (MCM6) in Europeans. We also infer the trajectory of a SNP (EDAR) in Han Chinese, finding evidence that this allele's age is much older than previously claimed. We also study a set of 11 pigmentation-associated variants. Several genes show evidence of strong selection particularly within the last 5,000 years, including ASIP, KITLG, and TYR. However, selection on OCA2/HERC2 seems to be much older and, in contrast to previous claims, we find no evidence of selection on TYRP1.

Intrapelvic Femoral Head Dislocation without Associated Proximal Femur Fracture: A Case Report and Description of Closed Reduction Technique.

Traumatic hip dislocations are potentially devastating injuries, especially in young patients, and require emergent orthopedic treatment. Given the significant amount of energy required to cause these injuries, a high index of suspicion is necessary to identify related injuries. The associated injuries, direction of dislocation, and time between injury and reduction represent the known prognostic factors, based on limited available research. Intrapelvic hip dislocations represent an uncommon variant of the traumatic hip dislocation, with all previously reported cases involving ipsilateral proximal femur fractures. We present a case of intrapelvic femoral head dislocation without an associated proximal femur fracture, as well as the maneuvers used to treat the patient via a closed reduction.

Pleistocene land bridges act as semipermeable agents of avian gene flow in Wallacea.

Cyclical periods of global cooling have been important drivers of biotic differentiation throughout the Quaternary. Ice age-induced sea level fluctuations can lead to changing patterns of land connections, both facilitating and disrupting gene flow. In this study, we test if species with differing life histories are differentially affected by Quaternary land connections. We used genome-wide SNPs in combination with mitochondrial gene sequences to analyse levels of divergence and gene flow between two songbird complexes across two Wallacean islands that have been repeatedly connected during glaciations. Although the two bird complexes are similar in ecological attributes, the forest and edge-inhabiting golden whistler Pachycephala pectoralis is comparatively flexible in its diet and niche requirements as compared to the henna-tailed jungle-flycatcher Cyornis colonus, which is largely restricted to the forest interior. Using population-genomic and coalescent approaches, we estimated levels of gene flow, population differentiation and divergence time between the two island populations. We observed higher levels of differentiation, an approximately two to four times deeper divergence time and near-zero levels of gene flow between the two island populations of the more forest-dependent henna-tailed jungle-flycatcher as compared to the more generalist golden whistler. Our results suggest that Quaternary land bridges act as semipermeable agents of gene flow in Wallacea, allowing only certain taxa to connect between islands while others remain isolated. Quaternary land bridges do not accommodate all terrestrial species equally, differing in suitability according to life history and species biology. More generalist species are likely to use Quaternary land connections as a conduit for gene flow between islands whereas island populations of more specialist species may continue to be reproductively isolated even during periods of Quaternary land bridges.

A Population Phylogenetic View of Mitochondrial Heteroplasmy.

The mitochondrion has recently emerged as an active player in myriad cellular processes. Additionally, it was recently shown that >200 diseases are known to be linked to variants in mitochondrial DNA or in nuclear genes interacting with mitochondria. This has reinvigorated interest in its biology and population genetics. Mitochondrial heteroplasmy, or genotypic variation of mitochondria within an individual, is now understood to be common in humans and important in human health. However, it is still not possible to make quantitative predictions about the inheritance of heteroplasmy and its proliferation within the body, partly due to the lack of an appropriate model. Here, we present a population-genetic framework for modeling mitochondrial heteroplasmy as a process that occurs on an ontogenetic phylogeny, with genetic drift and mutation changing heteroplasmy frequencies during the various developmental processes represented in the phylogeny. Using this framework, we develop a Bayesian inference method for inferring rates of mitochondrial genetic drift and mutation at different stages of human life. Applying the method to previously published heteroplasmy frequency data, we demonstrate a severe effective germline bottleneck comprised of the cumulative genetic drift occurring between the divergence of germline and somatic cells in the mother, and the separation of germ layers in the offspring. Additionally, we find that the two somatic tissues we analyze here undergo tissue-specific bottlenecks during embryogenesis, less severe than the effective germline bottleneck, and that these somatic tissues experience little additional genetic drift during adulthood. We conclude with a discussion of possible extensions of the ontogenetic phylogeny framework and its possible applications to other ontogenetic processes in addition to mitochondrial heteroplasmy.

The effects of Pleistocene climate change on biotic differentiation in a montane songbird clade from Wallacea.

The role of Pleistocene Ice Age in tropical diversification is poorly understood, especially in archipelagos, in which glaciation-induced sea level fluctuations may lead to complicated changes in land distribution. To assess how Pleistocene land bridges may have facilitated gene flow in tropical archipelagos, we investigated patterns of diversification in the rarely-collected rusty-bellied fantail Rhipidura teysmanni (Passeriformes: Rhipiduridae) complex from Wallacea using a combination of bioacoustic traits and whole-genome sequencing methods (dd-RADSeq). We report a biogeographic leapfrog pattern in the vocalizations of these birds, and uncover deep genomic divergence among island populations despite the presence of intermittent land connections between some. We demonstrate how rare instances of genetic introgression have affected the evolution of this species complex, and document the presence of double introgressive mitochondrial sweeps, highlighting the dangers of using only mitochondrial DNA in evolutionary research. By applying different tree inference approaches, we demonstrate how concatenation methods can give inaccurate results when investigating divergence in closely-related taxa. Our study highlights high levels of cryptic avian diversity in poorly-explored Wallacea, elucidates complex patterns of Pleistocene climate-mediated diversification in an elusive montane songbird, and suggests that Pleistocene land bridges may have accounted for limited connectivity among montane Wallacean biota.

Identification of Highly Specific scFvs against Total Adiponectin for Diagnostic Purposes.

Adiponectin is one of the most abundant adipokines secreted from adipose tissue. It acts as an endogenous insulin sensitizer and plasma concentrations are inversely correlated with obesity and metabolic syndrome. A decrease in plasma adiponectin levels normally indicates increased hormonal activity of the visceral lipid tissue, which is associated with decreased insulin sensitivity. It may therefore be considered a valuable biomarker for elucidating the underlying deteriorations resulting in type 2 diabetes and macrovascular disease. Here we present the use of phage display technology to identify highly specific antibody fragments (scFvs) against adiponectin. The selected scFvs showed highly specific binding to globular and native adiponectin in ELISA tests. By using our phage display technology, we were able to obtain monoclonal antibodies with specific high affinity binding to the target protein in an effective and easy to upscale manner. The selected scFvs against adiponectin can be used for developing immunoassays suitable for use in metabolic syndrome diagnosis and monitoring.

Population structure and coalescence in pedigrees: Comparisons to the structured coalescent and a framework for inference.

Contrary to what is often assumed in population genetics, independently segregating loci do not have completely independent ancestries, since all loci are inherited through a single, shared population pedigree. Previous work has shown that the non-independence between gene genealogies of independently segregating loci created by the population pedigree is weak in panmictic populations, and predictions made from standard coalescent theory are accurate for populations that are at least moderately sized. Here, we investigate patterns of coalescence in pedigrees of structured populations. We find that the pedigree creates deviations away from the predictions of the structured coalescent that persist on a longer timescale than in the case of panmictic populations. Nevertheless, we find that the structured coalescent provides a reasonable approximation for the coalescent process in structured population pedigrees so long as migration events are moderately frequent and there are no migration events in the recent pedigree of the sample. When there are migration events in the recent sample pedigree, we find that distributions of coalescence in the sample can be modeled as a mixture of distributions from different initial sample configurations. We use this observation to motivate a maximum-likelihood approach for inferring migration rates and mutation rates jointly with features of the pedigree such as recent migrant ancestry and recent relatedness. Using simulation, we show that our inference framework accurately recovers long-term migration rates in the presence of recent migration events in the sample pedigree.

Effects of the population pedigree on genetic signatures of historical demographic events.

Genetic variation among loci in the genomes of diploid biparental organisms is the result of mutation and genetic transmission through the genealogy, or population pedigree, of the species. We explore the consequences of this for patterns of variation at unlinked loci for two kinds of demographic events: the occurrence of a very large family or a strong selective sweep that occurred in the recent past. The results indicate that only rather extreme versions of such events can be expected to structure population pedigrees in such a way that unlinked loci will show deviations from the standard predictions of population genetics, which average over population pedigrees. The results also suggest that large samples of individuals and loci increase the chance of picking up signatures of these events, and that very large families may have a unique signature in terms of sample distributions of mutant alleles.

Leveraging Distant Relatedness to Quantify Human Mutation and Gene-Conversion Rates.

The rate at which human genomes mutate is a central biological parameter that has many implications for our ability to understand demographic and evolutionary phenomena. We present a method for inferring mutation and gene-conversion rates by using the number of sequence differences observed in identical-by-descent (IBD) segments together with a reconstructed model of recent population-size history. This approach is robust to, and can quantify, the presence of substantial genotyping error, as validated in coalescent simulations. We applied the method to 498 trio-phased sequenced Dutch individuals and inferred a point mutation rate of 1.66 × 10(-8) per base per generation and a rate of 1.26 × 10(-9) for <20 bp indels. By quantifying how estimates varied as a function of allele frequency, we inferred the probability that a site is involved in non-crossover gene conversion as 5.99 × 10(-6). We found that recombination does not have observable mutagenic effects after gene conversion is accounted for and that local gene-conversion rates reflect recombination rates. We detected a strong enrichment of recent deleterious variation among mismatching variants found within IBD regions and observed summary statistics of local sharing of IBD segments to closely match previously proposed metrics of background selection; however, we found no significant effects of selection on our mutation-rate estimates. We detected no evidence of strong variation of mutation rates in a number of genomic annotations obtained from several recent studies. Our analysis suggests that a mutation-rate estimate higher than that reported by recent pedigree-based studies should be adopted in the context of DNA-based demographic reconstruction.

The SMC' is a highly accurate approximation to the ancestral recombination graph.

Two sequentially Markov coalescent models (SMC and SMC') are available as tractable approximations to the ancestral recombination graph (ARG). We present a Markov process describing coalescence at two fixed points along a pair of sequences evolving under the SMC'. Using our Markov process, we derive a number of new quantities related to the pairwise SMC', thereby analytically quantifying for the first time the similarity between the SMC' and the ARG. We use our process to show that the joint distribution of pairwise coalescence times at recombination sites under the SMC' is the same as it is marginally under the ARG, which demonstrates that the SMC' is, in a particular well-defined, intuitive sense, the most appropriate first-order sequentially Markov approximation to the ARG. Finally, we use these results to show that population size estimates under the pairwise SMC are asymptotically biased, while under the pairwise SMC' they are approximately asymptotically unbiased.

An Evaluation of In-Person and Online Engagement in Central Newfoundland.

BACKGROUND: This study evaluates the use of in-person focus groups and online engagement within the context of a large public engagement initiative conducted in rural Newfoundland. METHODS: Participants were surveyed about their engagement experience and demographic information. Pre and post key informant interviews were also conducted with organizers of the initiative. RESULTS: Of the 111 participants in the focus groups, 97 (87%) completed evaluation surveys; as did 23 (88%) out of 26 online engagement participants. Overall, focus group participants were positive about their involvement, with 87.4% reporting that they would participate in a similar initiative. Online participation was below expectations and these participants viewed their experience less positively than in-person participants. Organizers viewed the engagement initiative and the combined use of online and in-person engagement positively. CONCLUSIONS: This study presents a real-world example of the use of two methods of engagement. It also highlights the importance of the successful execution of whatever engagement mechanism is selected.

A renewal theory approach to IBD sharing.

A long genomic segment inherited by a pair of individuals from a single, recent common ancestor is said to be identical-by-descent (IBD). Shared IBD segments have numerous applications in genetics, from demographic inference to phasing, imputation, pedigree reconstruction, and disease mapping. Here, we provide a theoretical analysis of IBD sharing under Markovian approximations of the coalescent with recombination. We describe a general framework for the IBD process along the chromosome under the Markovian models (SMC/SMC'), as well as introduce and justify a new model, which we term the renewal approximation, under which lengths of successive segments are independent. Then, considering the infinite-chromosome limit of the IBD process, we recover previous results (for SMC) and derive new results (for SMC') for the mean number of shared segments longer than a cutoff and the fraction of the chromosome found in such segments. We then use renewal theory to derive an expression (in Laplace space) for the distribution of the number of shared segments and demonstrate implications for demographic inference. We also compute (again, in Laplace space) the distribution of the fraction of the chromosome in shared segments, from which we obtain explicit expressions for the first two moments. Finally, we generalize all results to populations with a variable effective size.

Introgression and phenotypic assimilation in Zimmerius flycatchers (Tyrannidae): population genetic and phylogenetic inferences from genome-wide SNPs.

Genetic introgression is pervasive in nature and may lead to large-scale phenotypic assimilation and/or admixture of populations, but there is limited knowledge on whether large phenotypic changes are typically accompanied by high levels of introgression throughout the genome. Using bioacoustic, biometric, and spectrophotometric data from a flycatcher (Tyrannidae) system in the Neotropical genus Zimmerius, we document a mosaic pattern of phenotypic admixture in which a population of Zimmerius viridiflavus in northern Peru (henceforth "mosaic") is vocally and biometrically similar to conspecifics to the south but shares plumage characteristics with a different species (Zimmerius chrysops) to the north. To clarify the origins of the mosaic population, we used the RAD-seq approach to generate a data set of 37,361 genome-wide single nucleotide polymorphisms (SNPs). A range of population-genetic diagnostics shows that the genome of the mosaic population is largely indistinguishable from southern Z. viridiflavus and distinct from northern Z. chrysops, and the application of parsimony and species tree methods to the genome-wide SNP data set confirms the close affinity of the mosaic population with southern Z. viridiflavus. Even so, using a subset of 2710 SNPs found across all sampled lineages in configurations appropriate for a recently proposed statistical ("ABBA/BABA") test that distinguishes gene flow from incomplete lineage sorting, we detected low levels of gene flow from northern Z. chrysops into the mosaic population. Mapping the candidate loci for introgression from Z. chrysops into the mosaic population to the zebra finch genome reveals close linkage with genes significantly enriched in functions involving cell projection and plasma membranes. Introgression of key alleles may have led to phenotypic assimilation in the plumage of mosaic birds, suggesting that selection may have been a key factor facilitating introgression.

Characterization of transcriptomes from sexual and asexual lineages of a New Zealand snail (Potamopyrgus antipodarum).

Understanding the evolution and maintenance of sexual reproduction is one of the central challenges of evolutionary biology, yet we know very little about how sex influences molecular evolution. The New Zealand freshwater snail Potamopyrgus antipodarum is ideally suited to address this knowledge gap because obligately sexual individuals often coexist with multiple independently derived obligately asexual lineages. This unusual situation allows direct comparisons both between sexual and asexual P. antipodarum and across populations that differ in the relative frequency of sexual individuals. As such, P. antipodarum has received a great deal of attention as a model system for the maintenance of sex in nature and is also used as a model for environmental toxicology and biological invasions. Molecular genetic resources for P. antipodarum will thus be useful to investigators in a variety of biological fields. We used 454 sequencing of cDNA libraries to generate transcriptomes from two sexual and two asexual P. antipodarum lineages. A de novo assembly of 116.7 Mb of sequence reads produced 41 396 contigs, and sequence similarity-based Gene Ontology annotations were obtained for 3740 contigs. We detected 408 315 SNP loci and 7315 microsatellite loci, which together represent the first genome-scale resource available for P. antipodarum. Raw 454 read sequences, contig sequences, annotation data and polymorphism data are publicly available in a searchable online database and for download at http://www.biology.uiowa.edu/neiman/transcriptome.php.

Classification of threespine stickleback along the benthic-limnetic axis.

Many species of fish display morphological divergence between individuals feeding on macroinvertebrates associated with littoral habitats (benthic morphotypes) and individuals feeding on zooplankton in the limnetic zone (limnetic morphotypes). Threespine stickleback (Gasterosteus aculeatus L.) have diverged along the benthic-limnetic axis into allopatric morphotypes in thousands of populations and into sympatric species pairs in several lakes. However, only a few well known populations have been studied because identifying additional populations as either benthic or limnetic requires detailed dietary or observational studies. Here we develop a Fisher's linear discriminant function based on the skull morphology of known benthic and limnetic stickleback populations from the Cook Inlet Basin of Alaska and test the feasibility of using this function to identify other morphologically divergent populations. Benthic and limnetic morphotypes were separable using this technique and of 45 populations classified, three were identified as morphologically extreme (two benthic and one limnetic), nine as moderately divergent (three benthic and six limnetic) and the remaining 33 populations as morphologically intermediate. Classification scores were found to correlate with eye size, the depth profile of lakes, and the presence of invasive northern pike (Esox lucius). This type of classification function provides a means of integrating the complex morphological differences between morphotypes into a single score that reflects the position of a population along the benthic-limnetic axis and can be used to relate that position to other aspects of stickleback biology.