Le Grapevine fanleaf virus (GFLV) est le principal agent de la maladie du court-noué de la vigne. Sa protéine de capside (CP) permet la formation des virions indispensables à la protection du génome viral, au mouvement de cellule à cellule au sein de tubules formés par la protéine de mouvement (MP) du virus, et à la transmission du GFLV par son nématode vecteur Xiphinema index. Principaux résultats : 1. un motif exposé à la surface de la CP dont la nature est critique pour transmission du GFLV par X. index a été identifié et pourrait constituer un déterminant de la spécificité de transmission. 2. Des tubules fluorescents ont été produits de façon constitutive in planta. Ils permettent de complémenter en trans un GFLV dépourvu de MP. 3. L’expression transitoire de la CP conduit à la production de pseudo-particules. Celles-ci sont modifiables à façon et font de la capside du GFLV une plateforme biotechnologique unique. De plus, c’est un puissant outil pour étudier la biologie du virus. / Grapevine fanleaf virus (GFLV) is the main agent of grapevine fanleaf degeneration disease. Its coat protein (CP) self-assembles in virions necessary for viral genome protection, for cell-to-cell movement using tubules formed by the movement protein (MP) of the virus, and for the transmission of GFLV by its nematode vector Xiphinema index.Main results: 1. An outer surface-exposed CP motif has been identified as critical for GFLV transmission by X. index and could be a determinant of transmission specificity. 2. Fluorescent tubules have been produced by constitutive expression in planta. They allow the complementation in trans of a GFLV deleted of its MP coding sequence. 3. Transient expression of the GFLV CP leads to the production of virus-like particles. They can be easily modified and show that GFLV capsid is a unique biotechnology platform. In addition, they are a powerful tool to study the biology of the virus.

Natural mortality is a poorly known aspect of fisheries biology, despite its importance in stock assessments and population analysis. Of the many potential sources of mortality and morbidity in fishes, the effect of internal parasites is perhaps the least studied. Intestinal parasites may inhibit nutrient uptake as well as stimulate an inflammatory response in fish. Intestinal parasites of several tropical pelagic elasmobranchs, including silky and night sharks (Carcharhinus falciformis and C. signatus), the pelagic stingray (Pteroplatytrygon violacea), and mesopelagic fishes including sailfin lancetfish (Alepisaurus ferox), oilfish (Ruvettus pretiosus), snake mackerel (Gempylus serpens), escolar (Lepidocybium flavobrunneum), and Atlantic pomfret (Brama brama) are described from the South Atlantic Bight in the western North Atlantic. Parasites recovered include cestodes, trematodes, acanthocephalans and nematodes. Total gastrointestinal parasite loads were compared against the size (both length and weight) of the host, showing no correlation in pelagic elasmobranchs and a small correlation in mesopelagic teleosts. Sex and parasite loads were also compared for elasmobranchs showing a weak correlation. Capture seasons were also compared, and other results of this research showed that the parasites in this study are not host-species specific. The observed parasite classes and total helminth loads are the first described for these five mesopelagic fishes and will serve as a baseline for further studies. Future research is suggested to ascertain if commercially valuable, co-occurring, co-existing pelagic fishes may also be at risk for similar intestinal parasite fauna and burdens.

Obesity is a current epidemic in the United States. Greater than one third of adults and nearly one fifth of children are classified as obese (Ogden, 2014). Increased awareness of this problem has triggered interest into the potentially causal agents in our environment. These agents, termed obesogens, are a type of teratogen hypothesized to work during embryonic development by programming offspring to store excess fat. A type of teratogen, obesogens are also typically reprotoxic, affecting germline development and embryonic viability. These noxious teratogens span a variety of different chemical classes including estrogen mimics, metals and metalloids, biocides, and even voluntary exposures like cigarette smoke. The goal of this work is to use the soil nematode, Caenorhabditis elegans to model the reprotoxic and obesogenic events that take place as a result of chronic, low-dose exposure to these agents. C. elegans is especially well suited to this end. The organism has a straightforward biology, a short lifespan on the order of days, and relevant lipid metabolic gene homology to humans. We characterize here the resultant effects on egg-laying, hatching, and embryo viability after chronic parental exposure to obesogens. Through this, we are able to best recapitulate the low-dose exposure for our obesogen screen. Here, chronic, sub-lethal parental exposure to the obesogens screened yielded viable offspring that exhibited increased percentages of body area lipid stained and induced differences in stained lipid localization. Reduced pharyngeal pumping rates in two of our offspring groups highlights the potential for lasting changes on behavioral phenotype as well. Lastly, we employed phylogenetic guided analysis to functionally assess putative C. elegans nuclear hormone receptors (NHRs) to human Peroxisome Proliferator Activated Receptors (PPARs). These human PPARs have implications in lipid metabolism and obesity. We report here novel findings that NHR-85 and SEX-1 may act to repress lipid storage in the nematode, thereby serving as potential homologs of human PPARα. The results presented in this study highlight the lasting effects of chronic parental exposure to noxious environmental teratogens on the function of the reproductive system. Furthermore, the nematode can be used as an in vivo model to screen sublethal doses of these teratogens for their potential to act as obesogens. Bioinformatics guided analysis of C. elegans NHR homologs to human NHRs can continue to yield novel insight on this fat storage and its regulation in the nematode.

Abstract Collagen prolyl 4-hydroxylases (C-P4Hs) and related enzymes catalyze the hydroxylation of certain proline residues in animal collagens and plant hydroxyproline-rich proteins, respectively. Animal C-P4Hs and their isoenzymes have been characterized to date from humans, rodents, insects and nematodes. Most of the animal C-P4Hs are α2β2 tetramers in which protein disulphide isomerase (PDI) serves as the β subunit, but the nematode C-P4Hs characterized so far have unique molecular compositions. Two P4Hs have been cloned from the plant Arabidopsis thaliana and one from the Paramecium bursaria Chlorella virus-1, these being monomeric enzymes. This thesis reports on the identification of a large P4H family in the green alga Chlamydomonas reinhardtii and the cloning and characterization of one member, Cr-P4H-1. This is a soluble monomer that hydroxylates in vitro several peptides representing sequences found in C. reinhardtii cell wall proteins. Lack of its activity led to a defective cell wall structure, indicating that Cr-P4H-1 is essential for proper cell wall assembly and that the other P4Hs cannot compensate for the lack of its activity. Two C. elegans genes, Y43F8B.4 and C14E2.4, predicted to code for C-P4H α subunit-like polypeptides were analyzed. Three transcripts were generated from Y43F8B.4, one of them coding for a functional C-P4H α subunit named PHY-4.1. C14E2.4 turned out not to be a C-P4H α subunit gene, as a frame-shift led to the omission of codons for two catalytically critical residues. PHY-4.1 formed active tetramers and dimers with PDI-2 and had unique substrate requirements in that it hydroxylated certain other proline-rich sequences besides collagen-like peptides. Inactivation of the Y43F8B.4 gene led to no obvious morphological abnormalities. Spatial expression of the phy-4.1 transcript and PHY-4.1 polypeptide was localized to the pharynx and the excretory duct. Taken together, these data indicate that PHY-4.1 is not involved in the hydroxylation of cuticular collagens but is likely to have other substrates in vivo. Cloning and characterization of the PHY-1 and PHY-2 subunits from the closely related nematode Caenorhabditis briggsae revealed distinct differences in assembly properties between the C. elegans and C. briggsae PHY-2 subunits in spite of their high amino acid sequence identity. Genetic disruption of C. briggsae phy-1 resulted in a less severe phenotype than that observed in C. elegans, evidently on account of its more efficient assembly of the C. briggsae PHY-2 to an active C-P4H explaining the milder phenotype. Rescue of C. elegans and C. briggsae phy-1 mutants was achieved by injection of a wild-type phy-1 gene from either species.

2 p. / Alfalfa varieties differ in fall dormancy, defined as growth during the fall. Nondormant alfalfa varieties are usually planted in mild winter areas for their ability to grow in the fall. However, fall growth of nondormant alfalfa may be undesirable in areas subject to repeated frosts or freezes. Nondormant, very nondormant, and extremely nondormant alfalfa varieties (fall dormancy class 8, 9, and 10) are adapted to elevations below 4000 feet in Arizona. Other dormancy classes not included in this publication are moderately nondormant varieties (fall dormancy class 7) which may be grown from 3000 to 5000 feet, and semi-dormant and dormant varieties (fall dormancy 6 and below) which are adapted to colder winter areas above 4000 feet.

Les entérobactéries du genre Xenorhabdus sont pathogènes de larves d'insectes et symbiotiques de nématodes du genre Steinernema. En lutte biologique, les couples Steinernema-Xenorhabdus sont utilisés contre un large spectre d'insectes ravageurs de culture. Les deux partenaires du couple modèle Steinernema carpocapsae-Xenorhabdus nematophila peuvent être expérimentalement dissociés tout en restant pathogènes pour les insectes. En revanche, certaines souches de Xenorhabdus sont non-virulentes lorsqu'elles sont injectées directement dans une larve d'insecte. L'objectif de cette thèse est de caractériser deux souches non-virulentes de Xenorhabdus, X. poinarii G6 (Xp G6) et X. bovienii CS03 (Xb CS03). Les souches appartenant à l'espèce non-virulente X. poinarii possèdent des génomes de petite taille. Nous avons mis en évidence un phénomène de réduction génomique due à la délétion de larges régions génomiques chez la souche Xp G6. Cette évolution pourrait avoir eu lieu suite à un transfert des fonctions bactériennes de virulence à son nématode hôte et/ou à sa spécialisation envers certains coléoptères. Au sein de l'espèce X. bovienii, Xb CS03 est non-virulente par injection dans les lépidoptères Spodoptera littoralis et Galleria mellonella. Par rapport à d'autres couples némato-bactériens Steinernema sp.-X. bovienii, le couple formé par Xb CS03 et son nématode symbiotique S. weiseri 583 présente également une virulence atténuée sur ces lépidoptères. Le génome de Xb CS03 est de très grande taille et contient un grand nombre de gènes dégradés (pseudogènes). Une comparaison génomique entre Xb CS03 et une souche virulente appartenant à la même espèce, X. bovienii SS-2004 (Xb SS-2004), montre que Xb CS03 est plus riche que Xb SS-2004 en gènes codant des chaînes d'assemblage enzymatiques NRPS/PKS (non-ribosomal peptide synthase/polyketide synthethase) produisant des métabolites antimicrobiens potentiels. A l'inverse, Xb SS-2004 contient davantage de gènes codant des facteurs de virulence de type hémolysine, adhésine ou systèmes de sécrétion. Ceci suggère deux scénarios évolutifs différents, privilégiant une forte virulence pour Xb SS-2004 et l'élimination des compétiteurs au sein du cadavre de l'insecte pour Xb CS03. Enfin, une recherche de facteurs de virulence potentiels a été effectuée par une approche de génomique comparative entre les souches non-virulentes Xp G6 et Xb CS03, d'une part et trois souches de Xenorhabdus virulentes, d'autre part. L'analyse fonctionnelle de gènes candidats a été entamée. En conclusion, la caractérisation de nouveaux modèles bactériens dans le genre Xenorhabdus ouvre le champ à l'identification de nouvelles stratégies de virulence et de nouveaux facteurs de virulence chez les bactéries entomopathogènes. / Xenorhabdus are enterobacteria pathogenic of insect larvae and symbiotic of nematodes from the Steinernema genus. The Steinernema-Xenorhabdus associations are used against a wide range of insect pests. The two partners of the model Steinernema carpocapsae-Xenorhabdus nematophila association can be experimentally dissociated. Each partner is pathogenic for insect larvae. Contrarily, some other Xenorhabdus strains are non-virulent when injected directly into insect larvae. In this thesis, we characterized two non-virulent Xenorhabdus strains, X. poinarii G6 (Xp G6) and X. bovienii CS03 (Xb CS03). Strains from the X. poinarii species had small-sized genomes. We showed that the Xp G6 strain had undergone a genome reduction due to the deletion of large genomic regions. Transfer of virulence functions from the bacteria to the nematode and/or the specialization of the association towards coleopteran insects are likely the cause of this evolution. Within the X. bovienii species, Xb CS03 was non-virulent strain when injected into the Spodoptera littoralis and Galleria mellonella lepidopteran insects. When compared to other Steinernema-X. bovienii pairs, the association between Xb CS03 and its symbiotic nematode S. weiseri 583 had also a lower virulence on those insects. Xb CS03 had a large-sized genome and harbored numerous degraded genes (pseudogenes). Genome comparison between Xb CS03 and a virulent strain from the same species, X. bovienii SS-2004 (Xb SS-2004), showed that Xb CS03 contained more loci encoding NRPS/PKS enzymes (non-ribosomal peptide synthase/polyketide synthethase), producing potential antimicrobial metabolites, than Xb SS-2004. On the other hand, Xb SS-2004 contained more genes encoding virulence factors such as hemolysins, adhesins or secretion systems. This suggests that the two strains followed different evolutionary scenarios, favoring strong virulence in Xb SS-2204 and elimination of competitors for Xb CS03.Finally, we searched for potential virulence factors by comparing the genomes of the non-virulent strains Xp G6 and Xb CS03 with three virulent strains. Functional analyses of the candidates are in progress. In conclusion, characterizing new bacterial models in the Xenorhabdus genus paves the way for the identification of new virulence strategies and new virulence genes in entomopathogenic bacteria.

M.Sc. (Nematology) / Please refer to full text to view abstract

Experiments were performed to assess the suitability of Rhabditis pellio, a nematode found in earthworms, as a challenge organism for use in development of a biomarker assay to determine the potential of chemicals to suppress the immunocompetence of the non-specific immune system. To accomplish this goal, information on the life cycle of R. pellio was determined; including effects of incubation time and temperature on growth rates; along with information on the immune response elicited in the earthworm, Lumbricus terrestris. Immune parameters measured were coelomocyte migration toward and attachment to R. pellio larvae and adults. Preliminary background information showed that R. pellio has potential as a challenge organism for development of a biomarker assay.

Orientador: Silvia Renata Siciliano Wilcken / Resumo: Meloidogyne incognita é considerado um dos nematoides de maior importância econômica em termos mundiais. Para seu adequado manejo, uma das estratégias mais eficientes é o uso de cultivares resistentes. Portanto, estratégias para otimizar os programas de melhoramento genético são necessárias. A análise de expressão gênica tem ampliado a compreensão dos mecanismos de interação planta-nematoide e acelerado o desenvolvimento de genótipos resistentes. A cultivar de aveia branca IPR Afrodite, resistente a M. incognita, e cultivares suscetíveis foram avaliadas em relação à sua tolerância a este nematoide, além de ter sido estudado o comportamentode genes candidatos que regulam sua resistência, desafiados com densidades populacionais iniciais crescentes de M. incognita, durante o seu estabelecimento e manutenção do parasitismo. Para a avaliação da tolerância, plantas cultivadas em vasos de 3.600 cm3 foram inoculadas com 0,0625 (187,5); 0,125 (375); 0,25 (750); 0,5 (1500); 1 (3000); 2 (6000); 4 (12000); 8 (24000); 16 (48000) e 32 (96000) exemplares por cm3 de solo e avaliadas aos 67 dias após a inoculação (DAI), através da mensuração do desenvolvimento das plantas, além da multiplicação do nematoide. Os resultados mostraram que, em densidades menores, IPR Afrodite apresentou fator de reprodução próximo a 1,0, sugerindo menor expressão da resistência. Em função desses resultados, experimento semelhante foi conduzido para análise da expressão gênica dessa cultivar ao nematoide, com aval... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Meloidogyne incognita is considered one of the main economic important nematodes worldwide. For its management, the use of resistant cultivars is amongst the more efficient strategy. Therefore, strategies to optimize the genetic breeding programs are necessary. The gene expression analysis has amplified the comprehension about the mechanisms involved in the plant-nematode interaction and accelerated the development of resistant genotypes. The oat cultivar IPR Afrodite, resistant to M. incognita, and cultivars suscetibles was evaluated in relation to its tolerance to this nematode and, besides, the behavior of candidate genes regulating the resistance reaction was studied, challenged with crescent initial population densities of M. incognita, during its establishment and parasitism maintenance. To tolerance evaluation, plants cropped in 3,600 cm3 -pots were inoculated with 0,0625 (187,5); 0,125 (375); 0,25 (750); 0,5 (1500); 1 (3000); 2 (6000); 4 (12000); 8 (24000); 16 (48000) e 32 (96000) exemplars.cm-3 of soil and evaluated at 67 days after inoculation (DAI), through the mensuration of the plant development and nematode multiplication. Results showed that, in lower densities, IPR Afrodite showed reproduction factor values closed to 1.0, suggesting a lower resistance expression. In face of these results, a similar experiment was conducted to gene expression analysis of this cultivar to nematode, with evaluations at 0 (immediately before inoculation), 2 and 9 DAI. From RT-qPCR... (Complete abstract click electronic access below) / Doutor

Thesis (M. Sc.(Agronomy)) -- University of Limpopo, 2017. / Cleome gynandra is increasingly becoming an important strategy for achieving food and nutrition security among rural households in many developing countries. Root-knot (Meloidogyne species) nematodes, with limited nematode management strategies, limit the successful production of this vegetable crop. Nemafric-BL and Nemarioc-AL phytonematicides are separately being developed in South Africa for sustainable crop production systems. However, the two products have not been simultaneously tested for managing the notorious Meloidogyne species and absorption of phosphorus, with a combination of Vesicular arbuscular mycorrhiza (VAM). The objective of this study therefore was to determine the interactive effects of VAM and each of the two phytonematicides on nutrient content, growth of C. gynandra. A 2 × 2 × 2 factorial experiment, with the first, second and third factors being VAM (V), nematode (N) and Nemafric-BL phytonematicide (P). The eight treatments included (1) untreated control (V0N0P0), (2) nematodes alone (V0N1P0), (3) VAM alone (V1N0P0) (4) Nemarioc-AL phytonematicide alone (V0N0P1), (5) V1N1P0, (6) V0N1P1, (7) V1N0P1 and (8) V1N1P1, were laid out in a randomised complete block design, with ten replications. The same layout experiment was done for the Nemarioc-AL phytonematicide trial which had a similar layout. Seedlings were irrigated with 250 ml chloride-free tapwater every other day for 56 days. Multifeed and NPK (2:3:2(22) fertilisers were applied at transplanting. The second order interaction (V1N1P1), was highly significant (P ≤ 0.01) for plant height contributing 54% in TTV (Total Treatment Variation) of the variable. Among the main factors (N, P and V), only nematode had highly significant effects on stem diameter. All interactions of VAM, nematode and Nemarioc-AL phytonematicide and main factors each had no significant effect on Cleome. The second order (V1N1P1) and the first order interaction (V1N1P1) did not have significant effects on the three nutrient elements except for the first order interaction (V1N0P1) which was significant on foliar Zn contributing 42% in TTV of the variable. Also nematode had highly significant effect on foliar K and significant effect on foliar Zn contributing 49 and 31% in TTV of the respective variables. Using the two-way table, VAM and Nemafric-BL phytonematicide each increased foliar Zn by 27% and 29%, respectively. The second and first order interactions of VAM, N and Nemarioc-AL phytonematicide and the main factors did not have significant effect on foliar K, Fe and Zn. The second order interaction of VAM, nematode and Nemafric-BL phytonematicide had significant effects on gall rating, contributing 2% in TTV of the variable. VAM, nematode and Nemarioc-AL phytonematicide showed that the second and first order interaction except for V1N0P1 interaction on gall rating, were not significant for nematode variables. The V1N0P1 interaction contributed 20% in TTV of gall rating. Using a two-way table, VAM and phytonematicide each increased root galls by 7% and 74%, respectively. Combined, VAM and phytonematicide reduced root galls by 64%. The innovative products interacted together and that Nemafric-BL and Nemarioc-AL phytonematicides and VAM alone could be used in managing nematodes. / National Research Foundation, Agricultural Research Council-Universities Collaboration Centre