Species and Hybrids: Chemical and Physical Foliar Attributes and Implications for Herbivory.

Hybridization is an important biological phenomenon that can be used to understand the evolutionary process of speciation of plants and their associated pests and diseases. Interactions between hybrid plants and the herbivores of the parental taxa may be used to elucidate the various cues being used by the pests for host location or other processes. The chemical composition of plants, and their physical foliar attributes, including leaf thickness, trichome density, moisture content and specific leaf weight were compared between allopatric pure and commercial hybrid species of Corymbia, an important subtropical hardwood taxon. The leaf-eating beetle Paropsis atomaria, to which the pure taxa represented host (C. citriodora subsp. variegata) and non-host (C. torelliana) plants, was used to examine patterns of herbivory in relation to these traits. Hybrid physical foliar traits, chemical profiles, and field and laboratory beetle feeding preference, while showing some variability, were generally intermediate to those exhibited by parent taxa, thus suggesting an additive inheritance pattern. The hybrid susceptibility hypothesis was not supported by our field or laboratory studies, and there was no strong relationship between adult preference and larval performance. The most-preferred adult host was the sympatric taxon, although this species supported the lowest larval survival, while the hybrid produced significantly smaller pupae than the pure species. The results are discussed in relation to plant chemistry and physical characteristics. The findings suggest a chemical basis for host selection behavior and indicate that it may be possible to select for resistance to this insect pest in these commercially important hardwood trees.

Identification of intersectional Corymbia hybrids based on seedling morphology improves with parental divergence

Differences in morphology have provided a basis for detecting natural interspecific hybridisation in forest trees for decades but have come to prominence again more recently as a means for directly measuring gene flow from planted forests. Here we examined the utility of seedling morphology for hybrid discrimination in three hybrid groups relevant to the monitoring of gene flow from plantings of Corymbia (L.D. Pryor & L.A.S. Johnson ex Brooker) taxa in subtropical Australia. Thirty leaf and stem characters were assessed on 907 8-month old seedlings from four parental and six hybrid taxa grown in a common garden. Outbred F1 hybrids between spotted gums (Corymbia citriodora subspecies variegata, C. citriodora subspecies citriodora and Corymbia henryi) tended to more closely resemble their maternal Corymbia torelliana parent and the most discriminating characters were the ratio of blade length to maximum perpendicular width, the presence or absence of a lignotuber, and specific leaf weight. Assignment of individuals into genealogical classes based on a multivariate model limited to a set of the more discriminating and independent characters was highest in the hybrid group, where parental taxa were genetically most divergent. Overall power to resolve among outbred F1 hybrids from both parental taxa was low to moderate, but this may not be a limitation to its likely major application of identifying hybrids in seedlots from native spotted gum stands. Advanced generation hybrids (outbred F2 and outbred backcrosses) were more difficult to resolve reliably due to the higher variances of hybrid taxa and the tendency of backcrosses to resemble their recurrent parents. Visual assessments of seedling morphology may provide a filter allowing screening of the large numbers needed to monitor gene flow, but will need to be combined with other hybrid detection methods to ensure hybrids are detected.

Area Optimized H.264 Intra Prediction Architecture for 1080p HD Resolution

High performance video standards use prediction techniques to achieve high picture quality at low bit rates. The type of prediction decides the bit rates and the image quality. Intra Prediction achieves high video quality with significant reduction in bit rate. This paper present an area optimized architecture for Intra prediction, for H.264 decoding at HDTV resolution with a target of achieving 60 fps. The architecture was validated on Virtex-5 FPGA based platform. The architecture achieves a frame rate of 64 fps. The architecture is based on multi-level memory hierarchy to reduce latency and ensure optimum resources utilization. It removes redundancy by reusing same functional blocks across different modes. The proposed architecture uses only 13% of the total LUTs available on the Xilinx FPGA XC5VLX50T.

Gene enrichment using antibodies to DNA/RNA hybrids : mapping the ribosomal DNA of slime mold and rat

A novel method for gene enrichment has been developed and applied to mapping the rRNA genes of two eucaryotic organisms. The method makes use of antibodies to DNA/RNA hybrids prepared by injecting rabbits with the synthetic hybrid poly(rA)•poly(dT). Antibodies which cross-react with non-hybrid nucleic acids were removed from the purified IgG fraction by adsorption on columns of DNA-Sepharose, oligo(dT)-cellulose, and poly(rA)-Sepharose. Subsequent purification of the specific DNA/RNA hybrid antibody was carried out on a column of oligo(dT)-cellulose to which poly(rA) was hybridized. Attachment of these antibodies to CNBr-activated Sepharose produced an affinity resin which specifically binds DNA/RNA hybrids.

In order to map the rDNA of the slime mold Dictyostelium discoideum, R-loops were formed using unsheared nuclear DNA and the 178 and 268 rRNAs of this organism. This mixture was passed through a column containing the affinity resin, and bound molecules containing R- loops were eluted by high salt. This purified rDN A was observed directly in the electron microscope. Evidence was obtained that there is a physical end to Dictyostelium rDN A molecules approximately 10 kilobase pairs (kbp) from the region which codes for the 268 rRNA. This finding is consistent with reports of other investigators that the rRNA genes exist as inverse repeats on extra-chromosomal molecules of DNA unattached to the remainder of the nuclear DNA in this organism.

The same general procedure was used to map the rRNA genes of the rat. Molecules of DNA which contained R-loops formed with the 188 and 288 rRNAs were enriched approximately 150- fold from total genomal rat DNA by two cycles of purification on the affinity column. Electron microscopic measurements of these molecules enabled the construction of an R-loop map of rat rDNA. Eleven of the observed molecules contained three or four R-loops or else two R-loops separated by a long spacer. These observations indicated that the rat rRNA genes are arranged as tandem repeats. The mean length of the repeating units was 37.2 kbp with a standard deviation of 1.3 kbp. These eleven molecules may represent repeating units of exactly the same length within the errors of the measurements, although a certain degree of length heterogeneity cannot be ruled out. If significantly shorter or longer repeating units exist, they are probably much less common than the 37.2 kbp unit.

The last section of the thesis describes the production of antibodies to non-histone chromosomal proteins which have been exposed to the ionic detergent sodium dodecyl sulfate (SDS). The presence of low concentrations of SDS did not seem to affect either production of antibodies or their general specificity. Also, a technique is described for the in situ immunofluorescent detection of protein antigens in polyacrylamide gels.

Comparative growth and viability of hybrids between two populations of Chinese shrimp (Fennropenaeus chinensis)

Fenneropenaeus chinensis is confined to the Yellow Sea and Bohai Sea in China and the West Coast of the Korean Peninsula. Intra- and intercross populations were produced between Rushany (YP) and Korean (KN) populations. Seven traits were recorded. The heterosis of hybrids was computed and comparison between treatments was performed by ANOVA. At the fourth month after post-larvae, six indexes of growth trait and viability showed a range of heterosis, ranging from 0.514% to 14.950%. At the fifth month after post-larvae, six indexes of growth trait and viability ranged from -9.000% to 19.090%. The negative heterosis was observed in CL, HST and viability. The heterosis of KN female xYP male tended generally to increase as the age of the Chinese shrimp increased while the heterosis of YP female xKN male tended to decrease. The results indicated that the viability of reciprocal hybrids were not significantly different (P > 0.05) from their parents during the experiment. The result of ANOVA indicated that the F1 hybrids were significantly different (P < 0.05) in WST and TW at the fourth month. The multiple comparisons of LSD test indicated that the hybrids of YP female xKN male were significantly different (P < 0.05) from their parents in TW. The hybrids of YP female xKN male were significantly different (P < 0.05) from the other three combinations in WST. At the fifth months, the F1 hybrids had significant difference (P < 0.05) in body weight while other traits showed no significant differences (P > 0.05) from their parents. The multiple comparisons of LSD test indicated that the hybrids of KN female xYP male were significantly different (P < 0.05) from the KN parents in TW. The results indicate that in experimental conditions, the F-1 hybrids created from two populations of Chinese shrimp showed a certain level of heterosis for growth performance and viability. The crossing scheme may improve growth performance and viability in Chinese shrimp, but the improvement may be limited because effective crossbreeding requires the maintenance of pure, preferably inbred, lines and possibly involves specialized sire and dam lines. Therefore, the exploitation of heterosis through single crossbreeding in Chinese shrimp is of limited utility in practical commercial shrimp aquaculture in spite of the potential of significant heterosis. The crossbreeding of different populations can be applied in the establishment of base populations.

Intra-habitat heterogeneity of microbial food web structure under the regime of eutrophication and sediment resuspension in the large subtropical shallow Lake Taihu, China

Planktonic microbial community structure and classical food web were investigated in the large shallow eutrophic Lake Taihu (2338 km(2), mean depth 1.9 m) located in subtropical Southeast China. The water column of the lake was sampled biweekly at two sites located 22 km apart over a period of twelve month. Site 1 is under the regime of heavy eutrophication while Site 2 is governed by wind-driven sediment resuspension. Within-lake comparison indicates that phosphorus enrichment resulted in increased abundance of microbial components. However, the coupling between total phosphorus and abundance of microbial components was different between the two sites. Much stronger coupling was observed at Site 1 than at Site 2. The weak coupling at Site 2 was mainly caused by strong sediment resuspension, which limited growth of phytoplankton and, consequently, growth of bacterioplankton and other microbial components. High percentages of attached bacteria, which were strongly correlated with the biomass of phytoplankton, especially Microcystis spp., were found at Site 1 during summer and early autumn, but no such correlation was observed at Site 2. This potentially leads to differences in carbon flow through microbial food web at different locations. Overall, significant heterogeneity of microbial food web structure between the two sites was observed. Site-specific differences in nutrient enrichment (i.e. nitrogen and phosphorus) and sediment resuspension were identified as driving forces of the observed intra-habitat differences in food web structure.

The intra-Traveller debate on ‘Traveller ethnicity’ in the Republic of Ireland. A critical discourse analysis

This dissertation assesses from an under-explored angle the enduring contention over Travellers’ ethnic recognition in the Republic of Ireland, particularly over the last decade. The novelty of this study concerns not only its specific focus on and engagement with the debate on ‘Traveller ethnicity’ among Traveller activists. It also pertains to the examination of Travellers’ arguments for and against ethnicity in light of critical theorisations as well as insights from identity politics. Furthermore, the adoption of a Critical Discourse Analytical framework offers new perspectives to this controversy and its potential implications. Finally, this thesis’ relevance extends beyond the contention on ‘Traveller ethnicity’ in itself. It also draws attention to the complex dynamics of colonisation and appropriation between the global and the local. Particularly, it points to the interplay between international human rights discourses and the local ones, formulated by NGOs struggling for equality. In this way it sheds light on more general issues such as the dialectical potential of human rights discourses: the benefits and pitfalls of framing recognition claims in the legalistic terms of human rights. In this study it is argued that the contention on ‘Traveller ethnicity’ defies a simplistic polarisation between Irish Travellers and the Irish State since it has been simultaneously played out within the Travelling community. Specifically, this study explores how ‘Traveller ethnicity’ has been introduced, embraced, promoted and contested within Traveller politics to the point of becoming a hotly debated and divisive issue among Traveller activists and at the heart of the community itself. Putting Traveller activists centre-stage, their discourses for and against ‘Traveller ethnicity’ are examined and assessed against one another and their potential implications for Traveller politics, policies and identities are pointed out. Contending discourses are historically contextualised as the product of specific structural, material and discursive configurations of power and socio-economic relations within Irish society. Discourses for and against ‘Traveller ethnicity’ are assessed as being significant beyond the representational level. They are regarded as contributing to dialectically constitute Travellers’ ways of being, representing and acting. Furthermore these discourses are considered as sites and means of power struggles, whose stakes are not only words, but relate to issues of power and leadership within the Travelling community; adjudications over material resources; the adoption of certain policy approaches over others; and, finally, the consolidation of certain subject positions over others for Travellers to draw upon and relate to mainstream society. This study highlights an ongoing ideological struggle for the naturalisation of ‘Traveller ethnicity’ as a self-evident ‘fact’, which involves no active choice by Travellers themselves. Overall, ‘Traveller ethnicity’ appears to constitute an enduring source of dilemmas for the Travelling community. These revolve around the contradictory potential of ethnicity claims-making —both its perils and advantages— and its status as a potent political strategic resource that can both challenge and reinforce existing power relations, policies and identities.

DEVELOPMENT OF A SENSITIVE AND SPECIFIC TIME-RESOLVED FLUOROMETRIC IMMUNOASSAY FOR THE BOVINE ACUTE-PHASE PROTEIN HAPTOGLOBIN (HP)

Haptoglobin (Hp) is recognised as a major acute phase protein in the bovidae and its presence in serum is used as an indicator of inflammation. A mouse monoclonal antibody (1D9) specific for bovine Hp was labelled with a lanthanide (Eu) chelate and used to develop a competitive immunoassay. This competitive immunoassay allowed direct measurement of Hp in serum and was validated for intra- and interassay coefficients of variation (below 8%). Cross-reactivity with other serum proteins was measured (less than 0.1%) and limits of detection for Hp in serum were established for adult male (0.344 mu g/ml) and adult female cattle (1.589 mu g/ml). The immunoassay was compared with an established haptoglobin-haemoglobin binding assay.

Somatic hybrids of higher plants obtained from amiprophosmethyl-resistant mutant Nicotiniana plumbaginifolia

Interspecific and intertribal somatic hybrids were obtained to study the composition and function of microtubules in hybrid plants. The amiprophosmethyl-resistant mutant Nicotiana plumbaginifolia L. was used as donor; canamycin-resistant mutants N. sylvestris L. and Atropa belladonna served as recipients. Cytogenetic analysis confirmed the hybrid nature of the clones selected. Immunoflourescent analysis showed that constitutions of mitotic spindles in regenerating protoplast, isolated from the hybrid NpAb-107 and the mutant N. plunbaginifolia, show no change after a 2-hour treatment with 5 mu M of amiprophosmethyl; in A. belladonna, the division spindle is completely destroyed under these conditions. Tubulin was isolated from the hybrid NpAb-107 and separated by two-dimensional electrophoresis. The results showed that NpAb-107 has the beta-tubulin isoform specific for N. plumbaginifolia in addition to all isoforms of A. belladonna.

Identification of systemic immune response markers through metabolomic profiling of plasma from calves given an intra-nasally delivered respiratory vaccine

Vaccination procedures within the cattle industry are important disease control tools to minimize economic and welfare burdens associated with respiratory pathogens. However, new vaccine, antigen and carrier technologies are required to combat emerging viral strains and enhance the efficacy of respiratory vaccines, particularly at the point of pathogen entry. New technologies, specifically metabolomic profiling, could be applied to identify metabolite immune-correlates representative of immune protection following vaccination aiding in the design and screening of vaccine candidates. This study for the first time demonstrates the ability of untargeted UPLC-MS metabolomic profiling to identify metabolite immune correlates characteristic of immune responses following mucosal vaccination in calves. Male Holstein Friesian calves were vaccinated with Pfizer Rispoval® PI3 + RSV intranasal vaccine and metabolomic profiling of post-vaccination plasma revealed 12 metabolites whose peak intensities differed significantly from controls. Plasma levels of glycocholic acid, N-[(3α,5β,12α)-3,12-Dihydroxy-7,24-dioxocholan-24-yl]glycine, uric acid and biliverdin were found to be significantly elevated in vaccinated animals following secondary vaccine administration, whereas hippuric acid significantly decreased. In contrast, significant upregulation of taurodeoxycholic acid and propionylcarnitine levels were confined to primary vaccine administration. Assessment of such metabolite markers may provide greater information on the immune pathways stimulated from vaccine formulations and benchmarking early metabolomic responses to highly immunogenic vaccine formulations could provide a means for rapidly assessing new vaccine formulations. Furthermore, the identification of metabolic systemic immune response markers which relate to specific cell signaling pathways of the immune system could allow for targeted vaccine design to stimulate key pathways which can be assessed at the metabolic level.

Cytogenetic and histological studies of the brook trout (Salvelinus fontinalis) x arctic charr (Salvelinus alpinus) hybrids

In aquaculture, application of fish hybrids has increased. This technique permits improvement of the fish production by providing specimens showing better growth rate when compared to the parental species. Indeed, sterile individuals are highly demanded because quite frequently parental fish mature before they reach the market size, which impairs their growth and decrease their economic value. Throughout the last years, the commercial and scientific interest in salmonids has increased rapidly, among them, the brook trout (Salvelinus fontinalis), Arctic charr (Salvelinus alpinus) are species that can be crossed to produce hybrids that might by cultured in the fish farms. In the present thesis, we have assessed chromosome numbers and evaluate gonadal sex in the brook trout X Arctic charr hybrid progenies. In our populations, the karyotype of the brook trout comprises 84 chromosomes: 16 bi-armed chromosomes (meta-submetacentric) and 68 one-armed chromosomes (telo-acrocentrics) and the chromosome arm number, NF= 100. Arctic charr karyotype shows variation related to the chromosome number (2n= 81-82) and stable chromosome arm number (NF= 100). 2n= 81 chromosomes consisted of 19 bi-armed and 62 one-armed chromosomes, while 2n= 82 karyotype was organized into 18 meta-submetacentric and 64 acrocentrics. The cytogenetic and histological analysis of the brook trout X Arctic charr hybrids (sparctics) was carried out to asses chromosome and chromosome arm number and gonadal sex of the studied specimens. Diploid chromosome number in the hybrids varied from 81 to 84 and individuals with 83 and 84 chromosomes were predominant. Most of the fish had chromosome arm number equal to 100. Robertsonian fusion in the Arctic charr and chromosome behaviour in the hybrid fish cells might lead to the observed variation in chromosome numbers in the hybrids. Among studied fish, 12 were males, 3 were females and 9 had intersex gonads. No correlation between chromosome number and disturbances in the gonadal development was found. This might suggest that intersex gonads might have been developed as a consequence of disturbances in the genetic sex determination process. Genetic sex determination acts properly in the parental species but in the hybrids this may not be as efficient.

The identification and characterization of inter- and intra-species genetic diversity derived from retrotransposons in humans

Retrotransposons, which used to be considered as “junk DNA”, have begun to reveal their immense value to genome evolution and human biology due to recent studies. They consist of at least ~45% of the human genome and are more or less the same in other mammalian genomes. Retrotransposon elements (REs) are known to affect the human genome through many different mechanisms, such as generating insertion mutations, genomic instability, and alteration in gene expression. Previous studies have suggested several RE subfamilies, such as Alu, L1, SVA and LTR, are currently active in the human genome, and they are an important source of genetic diversity between human and other primates, as well as among humans. Although several groups had used Retrotransposon Insertion Polymorphisms (RIPs) as markers in studying primate evolutionary history, no study specifically focused on identifying Human-Specific Retrotransposon Element (HS-RE) and their roles in human genome evolution. In this study, by computationally comparing the human genome to 4 primate genomes, we identified a total of 18,860 HS-REs, among which are 11,664 Alus, 4,887 L1s, 1,526 SVAs and 783 LTRs (222 full length entries), representing the largest and most comprehensive list of HS-REs generated to date. Together, these HS-REs contributed a total of 14.2Mb sequence increase from the inserted REs and Target Site Duplications (TSDs), 71.6Kb increase from transductions, and 268.2 Kb sequence deletion of from insertion-mediated deletion, leading to a net increase of ~14 Mb sequences to the human genome. Furthermore, we observed for the first time that Y chromosome might be a hot target for new retrotransposon insertions in general and particularly for LTRs. The data also allowed for the first time the survey of frequency of TE insertions inside other TEs in comparison with TE insertion into none-TE regions. In summary, our data suggest that retrotransposon elements have played a significant role in the evolution of Homo sapiens.

Sector-Specific on-the-Job Training: Evidence from U.S. Data.

Using data from the National Longitudinal Survey of Youth (NLSY), we re-examine the effect of formal on-the-job training on mobility patterns of young American workers. By employing parametric duration models, we evaluate the economic impact of training on productive time with an employer. Confirming previous studies, we find a positive and statistically significant impact of formal on-the-job training on tenure with the employer providing the training. However, the expected net duration of the time spent in the training program is generally not significantly increased. We proceed to document and analyze intra-sectoral and cross-sectoral mobility patterns in order to infer whether training provides firm-specific, industry-specific, or general human capital. The econometric analysis rejects a sequential model of job separation in favor of a competing risks specification. We find significant evidence for the industry-specificity of training. The probability of sectoral mobility upon job separation decreases with training received in the current industry, whether with the last employer or previous employers, and employment attachment increases with on-the-job training. These results are robust to a number of variations on the base model.

Modulation de l’expression du gène CFTR par le produit du gène FIC1 responsable de la cholestase familiale intra-hépatique progressive de type 1 : Identification des mécanismes moléculaires impliqués

Réalisé en cotutelle avec l'Université de Cergy-Pontoise

Étude du polymorphisme intra- et inter-spécifique du gène β-tubuline chez des espèces de champignons mycorhiziens à arbuscules en vue de développer des marqueurs moléculaires

Les champignons mycorhiziens à arbuscules (CMA), classés dans le phylum Glomeromycota, ne peuvent pas être facilement identifiés par la morphologie de leurs spores et leurs mycélia à l'intérieur ou à l'extérieur des racines de leurs hôtes. Ce problème fondamental d'identification rend l'étude de leur diversité, en particulier dans leur habitat naturel (sol et racine) extrêmement difficile. Les gènes ribosomaux ont été largement utilisés pour développer des amorces spécifiques et en inférer des arbres phylogénétiques. Cependant, ces gènes sont très polymorphes et existent en plusieurs copies dans le génome des CMA, ce qui complique l’interprétation des résultats. Dans notre étude, nous avons étudié le polymorphisme intra- et inter-spécifique du gène β-tubuline, présent en faible nombre de copies dans le génome des CMA, afin d’obtenir de nouvelles séquences nucléotidiques pour développer des marqueurs moléculaires. Les gènes β-tubuline amplifiés à partir de l'ADN génomique de cinq espèces du genre Glomus ont été clonés et séquencés. L’analyse des séquences indique un polymorphisme intraspécifique chez trois espèces de CMA. Deux séquences paralogues très variables ont été nouvellement identifiées chez les G. aggregatum, G. fasciculatum et G. cerebriforme. Aucun polymorphisme n’a été détecté chez les G. clarum et G. etunicatum. Toutes les séquences montrent la présence de deux introns hautement variables. La majorité des substitutions ont été localisées dans les exons et sont synonymes à 90%. La conservation des acides aminés suggère un niveau élevé de sélection négative sur le gène β-tubuline et nous permet de confirmer que les CMA représentent un ancien groupe fongique (400 million d’années). L’analyse phylogénétique, réalisée avec vingt et une séquences nucléotidiques du gène β-tubuline, a révélé que les séquences des Glomaceae forment un groupe monophylétique bien supporté, avec les Acaulosporaceae et Gigasporaceae comme groupe frère. Les séquences paralogues nouvellement identifiées chez les G. aggregatum et G. fasciculatum n'ont pas été monophylétiques au sein de chaque espèce. Les oligonucléotides ont été choisis sur la base des régions variables et conservées du gène β-tubuline. Le test PCR des amorces β-Tub.cerb.F/ β-Tub.cerb.R a révélé des bandes spécifiques de 401 pb pour les séquences paralogues du G. cerebriforme. Deux paires d’amorces ont été développées afin d’identifier les séquences du groupe nommé Tub.1. Les tests PCR nous ont permis d’identifier certaines séquences du groupe Tub.1. Une paire d’amorce β-Tub.2.F/ β-Tub.2.R nous a permis d’identifier certaines séquences paralogues du groupe nommé Tub.2. L’analyse d’autres gènes combinée à celle du gène β-tubuline permettra le développement de marqueurs moléculaires plus spécifiques pour l’identification de CMA.