Contemporary population genetic structure and phylogeography of pink salmon

Thesis (Master's)--University of Washington, 2016-06

Understanding the genetic basis of phenotype variability in individuals with neurocognitive disorders

Thesis (Ph.D.)--University of Washington, 2016-06

Angler Perception and Population Dynamics of the Northern Snakehead (Channa argus) in the Potomac River & Tributaries

Our research sought to address the extent to which the northern snakehead (Channa argus), an invasive fish species, represents a threat to the Potomac River ecosystem. The first goal of our research was to survey the perceptions and opinions of recreational anglers on the effects of the snakehead population in the Potomac River ecosystem. To determine angler perceptions, we created and administered 113 surveys from June – September 2014 at recreational boat ramps along the Potomac River. Our surveys were designed to expand information collected during previous surveys conducted by the U.S. Fish and Wildlife Service. Our results indicated recreational anglers perceive that abundances and catch rates of target species, specifically largemouth bass, have declined since snakehead became established in the river. The second goal of our research was to determine the genetic diversity and potential of the snakehead population to expand in the Potomac River. We hypothesized that the effective genetic population size would be much less than the census size of the snakehead population in the Potomac River. We collected tissue samples (fin clippings) from 79 snakehead collected in a recreational tournament held between Fort Washington and Wilson’s Landing, MD on the Potomac River and from electrofishing sampling conducted by the Maryland Department of Natural Resources in Pomonkey Creek, a tributary of the Potomac River. DNA was extracted from the tissue samples and scored for 12 microsatellite markers, which had previously been identified for Potomac River snakehead. Microsatellite allele frequency data were recorded and analyzed in the software programs GenAlEx and NeEstimator to estimate heterozygosity and effective genetic population size. Resampling simulations indicated that the number of microsatellites and the number of fish analyzed provided sufficient precision. Simulations indicated that the effective population size estimate would expect to stabilize for samples > 70 individual snakehead. Based on a sample of 79 fish scored for 12 microsatellites, we calculated an Ne of 15.3 individuals. This is substantially smaller than both the sample size and estimated population size. We conclude that genetic diversity in the snakehead population in the Potomac River is low because the population has yet to recover from a genetic bottleneck associated with a founder effect due to their recent introduction into the system.

A Novel Noonan Syndrome RAF1 Mutation: Lethal Course in a Preterm Infant

Noonan syndrome is a relatively common and heterogeneous genetic disorder, associated with congenital heart defect in about 50% of the cases. If the defect is not severe, life expectancy is normal. We report a case of Noonan syndrome in a preterm infant with hypertrophic cardiomyopathy and lethal outcome associated to acute respiratory distress syndrome caused by Adenovirus pneumonia. A novel mutation in the RAF1 gene was identified: c.782C>G (p.Pro261Arg) in heterozygosity, not described previously in the literature. Consequently, the common clinical course in this mutation and its respective contribution to the early fatal outcome is unknown. No conclusion can be established regarding genotype/phenotype correlation.

Development, characterization and use of genomic SSR markers for assessment of genetic diversity in some Saudi date palm ( Phoenix dactylifera L.) cultivars

Background: The present study was undertaken towards the development of SSR markers and assessing genetic relationships among 32 date palm ( Phoenix dactylifera L.) representing common cultivars grown in different geographical regions in Saudi Arabia. Results: Ninety-three novel simple sequence repeat markers were developed and screened for their ability to detect polymorphism in date palm. Around 71% of genomic SSRs were dinucleotide, 25% tri, 3% tetra and 1% penta nucleotide motives. Twenty-two primers generated a total of 91 alleles with a mean of 4.14 alleles per locus and 100% polymorphism percentage. A 0.595 average polymorphic information content and 0.662 primer discrimination power values were recorded. The expected and observed heterozygosities were 0.676 and 0.763 respectively. Pair-wise similarity values ranged from 0.06 to 0.89 and the overall cultivars averaged 0.41. The UPGMA cluster analysis recovered by principal coordinate analysis illustrated that cultivars tend to group according to their class of maturity, region of cultivation, and fruit color. Analysis of molecular variations (AMOVA) revealed that genetic variation among and within cultivars were 27% and 73%, respectively according to geographical distribution of cultivars. Conclusions: The developed microsatellite markers are additional values to date palm characterization tools that can be used by researchers in population genetics, cultivar identification as well as genetic resource exploration and management. The tested cultivars exhibited a significant amount of genetic diversity and could be suitable for successful breeding program. Genomic sequences generated from this study are available at the National Center for Biotechnology Information (NCBI), Sequence Read Archive (Accession numbers. LIBGSS_039019).

Characterization of 27 microsatellite loci in the European flat oyster Ostrea edulis

The flat oyster Ostrea edulis is native to Europe and populations have been severely depleted by the parasite Bonamia ostreae since the 1980s. Additional genetic markers are required to improve population genetics study and linkage map development for selection for B. ostrea-resistance in this species. Here, we characterized 27 novel microsatellite loci for O. edulis. Number of alleles per locus ranged from 6 to 25 and observed heterozygosity between 0.375 and 1. Null alleles were suggested at a few loci but most loci were in Hardy-Weinberg agreement enabling their reliable use in further population and mapping genetics approaches.

Étude fonctionnelle d’un nouveau complexe multi-enzymatique régulant l’épigénome

L’ubiquitination, une modification post-traductionnelle importante pour le contrôle de nombreux processus cellulaires, est une réaction réversible. La réaction inverse, nommée déubiquitination est catalysée par les déubiquitinases (DUB). Nous nous sommes intéressés dans nos travaux à étudier l’ubiquitination de l’histone H2A (H2Aub), au niveau des résidus lysines 118 et 119 (K118/K119), une marque épigénétique impliquée dans la régulation de la prolifération cellulaire et la réparation de l’ADN. Le régulateur transcriptionnel BAP1, une déubiquitinase nucléaire, a été initialement identifié pour sa capacité à promouvoir la fonction suppressive de tumeurs de BRCA1. BAP1 forme un complexe multi-protéique avec plusieurs facteurs transcriptionnels et sa fonction principale est la déubiquitination de H2Aub. Plusieurs études ont démontré que BAP1 est un gène suppresseur de tumeurs majeur et qu’il est largement muté et inactivé dans une multitude de cancers. En effet, BAP1 émerge comme étant la DUB la plus mutée au niveau des cancers. Cependant, le ou les mécanismes d’action et de régulation du complexe BAP1 restent très peu connus. Dans cette étude nous nous sommes intéressés à la caractérisation moléculaire et fonctionnelle des partenaires protéiques de BAP1. De manière significative nous avons caractérisé un mécanisme unique de régulation entre deux composants majeurs du complexe BAP1 à savoir, HCF-1 et OGT. En effet, nous avons démontré que HCF-1 est requis pour maintenir le niveau protéique de OGT et que cette dernière est indispensable pour la maturation protéolytique de HCF-1 en promouvant son clivage par O-GlcNAcylation, une signalisation cellulaire nécessaire au bon fonctionnement de HCF-1. Également, nous avons découvert un nouveau mécanisme de régulation de BAP1 par l’ubiquitine ligase atypique UBE2O. En effet, UBE2O agit comme un régulateur négatif de BAP1 puisque l’ubiquitination de ce dernier induit sa séquestration dans le cytoplasme et l’inhibition de sa fonction suppressive de tumeurs. D’autre part nous nous sommes penchés sur la caractérisation de l’association de BAP1 avec deux facteurs de la famille des protéines Polycombes nommés ASXL1 et ASXL2 (ASXL1/2). Nous avons investigué le rôle de BAP1/ASXL1/2, particulièrement dans les mécanismes de déubiquitination et suppression de tumeurs. Nous avons démontré que BAP1 interagit directement iii via son domaine C-terminale avec le même domaine ASXM de ASXL1/2 formant ainsi deux complexes mutuellement exclusifs indispensables pour induire l’activité déubiquitinase de BAP1. De manière significative, ASXM s’associe avec BAP1 pour créer un nouveau domaine composite de liaison à l’ubiquitine. Ces interactions BAP1/ASXL1/2 régulent la progression harmonieuse du cycle cellulaire. De plus, la surexpression de BAP1 et de ASXL2 au niveau des fibroblastes induit la sénescence de manière dépendante de leurs interactions. D’autre part, nous avons identifié des mutations de cancers au niveau de BAP1 le rendant incapable de lier ASXL1/2, d’exercer sa fonction d’autodéubiquitination et de ce fait d’agir comme suppresseur de tumeurs. Ainsi nous avons révélé un lien étroit entre le gène suppresseur de tumeurs BAP1, son activité déubiquitinase et le contrôle de la prolifération cellulaire.

Highly polymorphic microsatellite markers for the Mediterranean endemic fan mussel Pinna nobilis

Pinna nobilis is an endemic bivalve of the Mediterranean Sea whose populations have decreased in the last decades due to human pressure; as a consequence, it was declared a protected species in 1992. Despite its conservation status, few genetic studies using mitochondrial markers have been published. We report on the isolation and development of 10 microsatellite loci for the fan mussel, Pinna nobilis. All loci (2 di-nucleotide, 5 tri-nucleotide, 2 tetra-nucleotide and 1 penta-nucleotide) are characterized by high levels of polymorphism in 76 individuals tested from two populations in the Balearic Islands (Spain, Western Mediterranean Sea). The number of alleles ranged from 4 to 24 and expected heterozygosity ranged from 0.4269 to 0.9400. These microsatellites could be very useful for the assessment of the genetic diversity and connectivity patterns of P. nobilis and the establishment of new conservation strategies.

Characterization of 10 new tetranucleotide microsatellite markers for the European eagle owl, Bubo bubo: Useful tools for conservation strategies

Bubo bubo is the largest owl in the world, showing a wide geographical distribution throughout the Palaearctic region. It underwent a demographic decline in many European countries during the last century and was considered “vulnerable” (Annex II of the CITES). Nowadays, it is classified as “Least Concern” according to IUCN. Despite its ecological importance and conservation status, few polymorphic molecular markers are available to study its diversity and population genetics. We report on the isolation and development of 10 new microsatellites for the Eagle owl, B. bubo. All loci (10 tetra-nucleotide) are characterized by high polymorphism levels. Number of alleles ranged from 5 to 13 and expected heterozygosity varied from 0.733 to 0.840. These microsatellites would be very useful to assess the genetic diversity, connectivity patterns and parentage of B. bubo. This information will allow to establish new conservation strategies and improve the management of the species.

Genetic considerations on the introduction of farmed fish in marine protected areas: The case of study of white seabream restocking in the Gulf of Castellammare (Southern Tyrrhenian Sea)

Human exploitation has drastically reduced the abundance and distribution of several marine fish and invertebrate populations through overfishing and habitat destruction. Restocking can potentially mitigate these impacts and help to reconstitute depleted stocks but genetic repercussions must be considered. In the present study, the degree of genetic similarity between white seabream (Diplodus sargus Linnaeus 1758) individuals reared for restocking purposes and the receiving population in the Gulf of Castellammare fishery reserve (Sicily, Italy) was assessed using microsatellites. We also inferred the spatial pattern of the genetic structure of D. sargus and connectivity along Sicilian coasts. The farmed population showed significant heterozygosity deficiency in 6 loci and an important reduction in the number of alleles, which could indicate an incipient inbreeding. Both the farmed population and the target one for restocking (Castellammare fishery reserve), showed high and significant values of genetic differentiation due to different allele frequencies, number of privative alleles and total number of alleles. These findings indicate a low degree of genetic similarity between both populations, therefore this restocking initiative is not advisable. The genetic connectivity pattern, highly consistent with oceanographic currents, identified two distinct metapopulations of white seabream around Sicily. Thus it is recommended to utilize broods from the same metapopulation for restocking purposes to provide a better genetic match to the wild populations.

Microsatellite analysis of pacu broodstocks used in the stocking program of Paranapanema River, Brazil

O monitoramento da diversidade genética é fundamental em um programa de repovoamento. Avaliouse a diversidade genética de pacu Piaractus mesopotamicus (Holmberg, 1887) em duas estações de piscicultura em Andirá -Paraná, Brasil, utilizadas no programa de repovoamento do Rio Paranapanema. Foram amplificados seis loci microssatélite para avaliar 60 amostras de nadadeira. O estoque de reprodutores B apresentou maior número de alelos e heterozigose (alelos: 22 e H O: 0,628) que o estoque de reprodutores A (alelos: 21 e H O: 0,600). Alelos com baixos níveis de frequência foram observados nos dois estoques. Os coeficientes positivos de endogamia no locus Pme2 (estoque A: F IS = 0,30 e estoque B: F IS = 0,20), Pme5 (estoque B: F IS = 0,15), Pme14 (estoque A: F IS = 0,07) e Pme28 (estoque A: F IS = 0,24 e estoque B: F IS = 0,20), indicaram deficiência de heterozigotos. Foi detectada a presença de um alelo nulo no lócus Pme2. As estimativas negativas nos loci Pme4 (estoque A: F IS = -0,43 e estoque B: F IS= -0,37), Pme5 (estoque A: F IS = - 0,11), Pme14 (estoque B: F IS = - 0,15) e Pme32 (estoque A: F IS = - 0,93 e estoque B: F IS = - 0,60) foram indicativas de excesso de heterozigotos. Foi evidenciado desequilíbrio de ligação e riqueza alélica baixa só no estoque A. A diversidade genética de Nei foi alta nos dois estoques. A distância (0,085) e identidade (0,918) genética mostraram similaridade entre os estoques, o qual reflete uma possível origem comum. 6,05% da variância genética total foi devida a diferenças entre os estoques. Foi observado um recente efeito gargalo nos dois estoques. Os resultados indicaram uma alta diversidade genética nos estoques de reprodutores e baixa diferenciação genética entre eles, o que foi causado pelo manejo reprodutivo das pisciculturas, redução do tamanho populacional e intercâmbio genético entre as pisciculturas.

The Role of the Transcription Factor Ets1 in Melanocyte Development

Melanocytes, pigment-producing cells, derive from the neural crest (NC), a population of pluripotent cells that arise from the dorsal aspect of the neural tube during embryogenesis. Many genes required for melanocyte development were identified using mouse pigmentation mutants. The deletion of the transcription factor Ets1 in mice results in hypopigmentation; nevertheless, the function of Ets1 in melanocyte development is unknown. The goal of the present study was to establish the temporal requirement and role of Ets1 in murine melanocyte development. In the mouse, Ets1 is widely expressed in developing organs and tissues, including the NC. In the chick cranial NC, Ets1 is required for the expression of Sox10, a transcription factor critical for the development of melanocytes, enteric ganglia, and other NC derivatives. Using a combination of immunofluorescence and cell survival assays Ets1 was found to be required between embryonic days 10 and 11, when it regulates NC cell and melanocyte precursor (melanoblast) survival. Given the requirement of Ets1 for Sox10 expression in the chick cranial NC, a potential interaction between these genes was investigated. Using genetic crosses, a synergistic genetic interaction between Ets1 and Sox10 in melanocyte development was found. Since Sox10 is essential for enteric ganglia formation, the importance of Ets1 on gut innervation was also examined. In mice, Ets1 deletion led to decreased gut innervation, which was exacerbated by Sox10 heterozygosity. At the molecular level, Ets1 was found to activate a Sox10 enhancer critical for Sox10 expression in melanoblasts. Furthermore, mutating Ets1 at a site I characterized in the spontaneous variable spotting mouse pigmentation mutant, led to a 2-fold decrease in enhancer activation. Overexpression and knockdown of Ets1 did not affect Sox10 expression; nonetheless, Ets1 knockdown led to a 6-fold upregulation of the transcription factor Sox9, a gene required for melanocyte and chondrocyte development, but which impairs melanocyte development when its expression is prolonged. Together, these results suggest that Ets1 is required early during melanocyte development for NC cell and melanoblast survival, possibly acting upstream of Sox10. The transcription factor Ets1 may also act indirectly in melanocyte fate specification by repressing Sox9 expression, and consequently cartilage fate.

Prevalence and audiological features in carriers of GJB2 mutations, c.35delG and c.101T>C (p.M34T), in a UK population study

OBJECTIVES: To determine the carrier rate of the GJB2 mutation c.35delG and c.101T>C in a UK population study; to determine whether carriers of the mutation had worse hearing or otoacoustic emissions compared to non-carriers. DESIGN: Prospective cohort study. SETTING: University of Bristol, UK. PARTICIPANTS: Children in the Avon Longitudinal Study of Parents and Children. 9202 were successfully genotyped for the c.35delG mutation and c.101>T and classified as either carriers or non-carriers. OUTCOME MEASURES: Hearing thresholds at age 7, 9 and 11 years and otoacoustic emissions at age 9 and 11. RESULTS: The carrier frequency of the c.35delG mutation was 1.36% (95% CI 1.13 to 1.62) and c.101T>C was 2.69% (95% CI 2.37 to 3.05). Carriers of c.35delG and c.101T>C had worse hearing than non-carriers at the extra-high frequency of 16 kHz. The mean difference in hearing at age 7 for the c.35delG mutation was 8.53 dB (95% CI 2.99, 14.07) and 12.57 dB at age 9 (95% CI 8.10, 17.04). The mean difference for c.101T>C at age 7 was 3.25 dB (95% CI -0.25 to 6.75) and 7.61 dB (95% CI 4.26 to 10.96) at age 9. Otoacoustic emissions were smaller in the c.35delG mutation carrier group: at 4 kHz the mean difference was -4.95 dB (95% CI -6.70 to -3.21) at age 9 and -3.94 dB (95% CI -5.78 to -2.10) at age 11. There was weak evidence for differences in otoacoustic emissions amplitude for c.101T>C carriers. CONCLUSION: Carriers of the c.35delG mutation and c.101T>C have worse extra-high-frequency hearing than non-carriers. This may be a predictor for changes in lower-frequency hearing in adulthood. The milder effects observed in carriers of c.101T>C are in keeping with its classification as a mutation causing mild/moderate hearing loss in homozygosity or compound heterozygosity.

Genetic signature of the northward expansion of the Egyptian mongoose Herpestes ichneumon (Herpestidae) in the Iberian Peninsula

In the last three decades, the range of the Egyptian mongoose (Herpestes ichneumon) has increased in the Iberian Peninsula. A panel of microsatellites was used to confront the patterns of genetic diversity of the species with the scenario of its recent northward expansion in its Iberian range. Evidence of substructure and significant genetic differentiation within the studied population were recorded, with a central-northern subpopulation (CNorth) and a southern subpopulation (S). Northward range expansion was supported by the observed allelic frequencies, diversity parameters, and observed heterozygosity of the studied loci, with S showing a higher allelic diversity and a higher number of private alleles than CNorth. Patterns of isolation-by-distance and isolation-by-barrier as a result of the Tagus River were demonstrated, suggesting that the river acted as a semi-permeable barrier, possibly leading to genetic differentiation of the studied population. The observed individuals from CNorth in southern locations and individuals from S in central/northern areas might comprise evidence for long range dispersals across the studied range. A bottleneck event after population expansion was supported by a significant heterozygosity deficiency in CNorth, which is in agreement with a scenario of founder events occurring in recently colonized areas after the crossing of the Tagus River.

Genetic diversity of the forage peanut in the Jequitinhonha, São Francisco, and Paranã River valleys of Brazil.

Arachis pintoi and A. repens are legumes with a high forage value that are used to feed ruminants in consortium systems. Not only do they increase the persistence and quality of pastures, they are also used for ornamental and green cover. The objective of this study was to analyze microsatellite markers in order to access the genetic diversity of 65 forage peanut germplasm accessions in the section Caulorrhizae of the genus Arachis in the Jequitinhonha, São Francisco and Paranã River valleys of Brazil. Fifty-seven accessions of A. pintoi and eight of A. repens were analyzed using 17 microsatellites, and the observed heterozygosity (HO), expected heterozygosity (HE), number of alleles per locus, discriminatory power, and polymorphism information content were all estimated. Ten loci (58.8%) were polymorphic, and 125 alleles were found in total. The HE ranged from 0.30 to 0.94, and HO values ranged from 0.03 to 0.88. By using Bayesian analysis, the accessions were genetically differentiated into three gene pools. Neither the unweighted pair group method with arithmetic mean nor a neighbor-joining analysis clustered samples into species, origin, or collection area. These results reveal a very weak genetic structure that does not form defined clusters, and that there is a high degree of similarity between the two species.