5. Vaccine development: basic considerations. Asexual blood stage vaccines: from merozoites to peptides

Asexual blood stage proliferation is responsible for the morbidity and mortality associated with malaria infection in man. These developmental stages are therefore obvious targets for the development of malaria vaccines. Several asexual blood stage components have been identified as potential candidates for the development of vaccines and some of them have been shown, following immunization, to induce at least partial protection in a variety of Plasmodium-host combinations. Studies on defined parasite components and on synthetic peptides derived from them have revealed new insights at the molecular level into parasite mechanisms involved in propagation and survival in the infected host, and into the interaction between parasite components and the host immune system. Practical application of these findings is likely to provide the basis for the design of more appropriate antigens for the development of vaccine

Daño, producción de semillas y persistencia del hongo endófito Epichloë occultans en plantas de Lolium multiflorum bajo un ataque de herbívoros lepidópteros y contaminación por ozono

Plants are expected to face novel challenges as consequence of human-driven global change. Outbreaks of pests and higher incidence of contaminants are increasing. Plants can improve tolerance to stress factors through associations with symbiotic microorganisms. Certain grasses establish persistent and asymptomatic symbioses with Epichloë fungal endophytes, which are known to confer protection against Herbivores and improve plant tolerance to abiotic stress factors. Nonetheless, accumulating evidence suggest the symbiosis outcome is context dependent. We evaluated the capacity of the endophyte fungus E. occultans in protecting the annual grass Lolium multiflorum against a spontaneous larva attack of the generalist herbivore Agrotis ipsilon under episodic exposure of plants to ozone. Symbiotic and non-symbiotic plants were individually grown outdoors and exposed to ozone at different stages resulting in four treatments: control (plant never exposed to ozone), plant exposed to ozone at vegetative stage, plant exposed to ozone at reproductive stage, and plant exposed to ozone at both stages. After the last exposure, there was an outbreak of A. ipsilon larvae. We evaluated herbivore damage, seed production per plant, and endophyte transmission to the seeds. Frequency of attacked plants was irrespective of both the endophyte and ozone exposure. However, the damage level per plant was only reduced by the endophyte. Seed production was slightly lower in endophyte-symbiotic plants and not affected by ozone. Interestingly, herbivore damaged and undamaged endophyte-symbiotic plants contributed equally to seed production. However, in plants exposed to ozone once at vegetative or reproductive stage, endophyte-free undamaged plants had higher seed production than endophyte-free damaged plants. Ozone treatments did not affect the transmission efficiency of endophytes to the seeds. Mean endophyte transmission efficiency per plant was 95%. Medium doses of ozone seem to have undetectable effects on grass-endophyte symbiosis, not affecting the defensive mutualism nor the persistence of the symbiont across generations.Las plantas enfrentan nuevos desafíos debido al cambio global impulsado por las actividades humanas. Mientras los brotes de plagas y la incidencia de contaminantes están en aumento, las plantas pueden mejorar su tolerancia al estrés asociándose con microorganismos. Ciertas gramíneas establecen simbiosis persistentes y asintomáticas con endófitos fúngicos Epichloë, y así obtienen protección contra herbívoros y tolerancia al estrés abiótico. No obstante, los resultados de la simbiosis dependen del contexto ecológico. Evaluamos la capacidad del hongo endófito E. occultans para proteger el pasto anual Lolium multiflorum contra un ataque de larvas del herbívoro generalista Agrotis ipsilon bajo exposición episódica de las plantas al ozono. Cultivamos plantas simbióticas y no-simbióticas en macetas y las expusimos a ozono en diferentes etapas, resultando en cuatro tratamientos: control (nunca expuesta al ozono), planta expuesta al ozono en etapa vegetativa, planta expuesta al ozono en etapa reproductiva y planta expuesta al ozono en ambas etapas. Luego, hubo un brote de larvas de A. ipsilon. Evaluamos el daño causado por las larvas, la producción de semillas por planta y la transmisión del endófito a las semillas. La frecuencia de plantas atacadas fue independiente de la presencia del endófito y del ozono. Sin embargo, el nivel de daño por planta sólo fue afectado y reducido por el endófito. La producción de semillas fue ligeramente menor en las plantas con endófito y no afectada por el ozono. Curiosamente, las plantas simbióticas dañadas y no-dañadas presentaron igual producción de semillas. No obstante, las plantas no-simbióticas no-dañadas produjeron más semillas que las dañadas cuando fueron expuestas una vez a ozono en etapa vegetativa o reproductiva. El ozono no afectó la eficiencia de transmisión de los endófitos a las semillas, la cual fue, en promedio, 95%. El ozono, en dosis medias, parece tener efectos indetectables sobre la simbiosis planta-endófito.Fil: Gundel, Pedro Emilio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentina. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Recursos Naturales y Ambiente; ArgentinaFil: Bigaxzoli, Fernando. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Métodos Cuantitativos y Sistemas de Información; ArgentinaFil: Freitas, Priscila P.. Ministry For Primary Industries; Nueva ZelandaFil: Landesmann, Jennifer Brenda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto de Investigaciones en Biodiversidad y Medioambiente. Universidad Nacional del Comahue. Centro Regional Universidad Bariloche. Instituto de Investigaciones en Biodiversidad y Medioambiente; Argentina. Universidad Nacional del Comahue. Centro Regional Universitario Bariloche. Laboratorio de Ecotono; ArgentinaFil: Martínez Ghersa, María Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentina. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Recursos Naturales y Ambiente; ArgentinaFil: Ghersa, Claudio Marco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentina. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Recursos Naturales y Ambiente; Argentin

Mutualism effectiveness and vertical transmission of symbiotic fungal endophytes in response to host genetic background

838-849Certain species of the Pooideae subfamily develop stress tolerance and herbivory resistance through symbiosis with vertically transmitted, asexual fungi. This symbiosis is specific, and genetic factors modulate the compatibility between partners. Although gene flow is clearly a fitness trait in allogamous grasses, because it injects hybrid vigor and raw material for evolution, it could reduce compatibility and thus mutualism effectiveness. To explore the importance of host genetic background in modulating the performance of symbiosis, Lolium multiflorum plants, infected and noninfected with Neotyphodium occultans, were crossed with genetically distant plants of isolines (susceptible and resistant to diclofop-methyl herbicide) bred from two cultivars and exposed to stress. The endophyte improved seedling survival in genotypes susceptible to herbicide, while it had a negative effect on one of the genetically resistant crosses. Mutualism provided resistance to herbivory independently of the host genotype, but this effect vanished under stress. While no endophyte effect was observed on host reproductive success, it was increased by interpopulation plant crosses. Neither gene flow nor herbicide had an important impact on endophyte transmission. Host fitness improvements attributable to gene flow do not appear to result in direct conflict with mutualism while this seems to be an important mechanism for the ecological and contemporary evolution of the symbiotum

Occurrence of Alkaloids in Grass Seeds Symbiotic With Vertically-Transmitted \u3cem\u3eEpichloë\u3c/em\u3e Fungal Endophytes and Its Relationship With Antioxidants

Host organisms can acquire new functional traits through symbiosis. Seed-transmitted Epichloë fungal endophytes are known to protect host plants against herbivores and increase tolerance to abiotic stresses by alkaloids and antioxidants, respectively (currencies of mutualism). Whereas, alkaloids are fungal products with demonstrated effects at plant vegetative stage, few studies have focused on alkaloids in seeds. We assessed the occurrence of fungal alkaloids and determined their concentrations in seeds of two host grasses, Festuca rubra and Lolium multiflorum. Then, we sought for a relationship with the antioxidants tocochromanols and glutathione, which are involved in the control of oxidative stress. Different alkaloids were detected depending on the species and plant genotype. Most notably, loline alkaloids were not detected in F. rubra seeds, whereas ergovaline and peramine were absent in L. multiflorum. In F. rubra, ergovaline concentration was dependent on the maternal line in interaction with the production year, diminishing in seeds after 1 year of storage. The exposure of L. multiflorum plants to ozone had no effect on the seed concentration of lolines. There was a significant positive relationship between the concentrations of ergovaline and tocochromanols in both species, and between ergovaline concentration and EGSSG/2GSH (glutathione half-cell reduction potential) in RAB maternal line of F. rubra. These results suggest that alkaloid and antioxidants have a close association in seeds of host grasses, and that the alkaloid bioactivity could be related with the antioxidant capacity to control stress. This has important implications for the ecology of partner species, thus supporting its consideration for further research

Occurrence of Alkaloids in Grass Seeds Symbiotic With Vertically-Transmitted Epichloë Fungal Endophytes and Its Relationship With Antioxidants

Host organisms can acquire new functional traits through symbiosis. Seed-transmitted Epichloë fungal endophytes are known to protect host plants against herbivores and increase tolerance to abiotic stresses by alkaloids and antioxidants, respectively (currencies of mutualism). Whereas, alkaloids are fungal products with demonstrated effects at plant vegetative stage, few studies have focused on alkaloids in seeds. We assessed the occurrence of fungal alkaloids and determined their concentrations in seeds of two host grasses, Festuca rubra and Lolium multiflorum. Then, we sought for a relationship with the antioxidants tocochromanols and glutathione, which are involved in the control of oxidative stress. Different alkaloids were detected depending on the species and plant genotype. Most notably, loline alkaloids were not detected in F. rubra seeds, whereas ergovaline and peramine were absent in L. multiflorum. In F. rubra, ergovaline concentration was dependent on the maternal line in interaction with the production year, diminishing in seeds after 1 year of storage. The exposure of L. multiflorum plants to ozone had no effect on the seed concentration of lolines. There was a significant positive relationship between the concentrations of ergovaline and tocochromanols in both species, and between ergovaline concentration and EGSSG/2GSH (glutathione half-cell reduction potential) in RAB maternal line of F. rubra. These results suggest that alkaloid and antioxidants have a close association in seeds of host grasses, and that the alkaloid bioactivity could be related with the antioxidant capacity to control stress. This has important implications for the ecology of partner species, thus supporting its consideration for further research

The Effects of Seed Size on Hybrids Formed between Oilseed Rape (Brassica napus) and Wild Brown Mustard (B. juncea)

Background : Seed size has significant implications in ecology, because of its effects on plant fitness. The hybrid seeds that result from crosses between crops and their wild relatives are often small, and the consequences of this have been poorly investigated. Here we report on plant performance of hybrid and its parental transgenic oilseed rape (Brassica napus) and wild B. juncea, all grown from seeds sorted into three seed-size categories.[br/] Methodology/Principal Findings : Three seed-size categories were sorted by seed diameter for transgenic B. napus, wild B. juncea and their transgenic and non-transgenic hybrids. The seeds were sown in a field at various plant densities. Globally, small-seeded plants had delayed flowering, lower biomass, fewer flowers and seeds, and a lower thousand-seed weight. The seed-size effect varied among plant types but was not affected by plant density. There was no negative effect of seed size in hybrids, but it was correlated with reduced growth for both parents.[br/] Conclusions : Our results imply that the risk of further gene flow would probably not be mitigated by the small size of transgenic hybrid seeds. No fitness cost was detected to be associated with the Bt-transgene in this study

The ER-Bound RING Finger Protein 5 (RNF5/RMA1) Causes Degenerative Myopathy in Transgenic Mice and Is Deregulated in Inclusion Body Myositis

Growing evidence supports the importance of ubiquitin ligases in the pathogenesis of muscular disorders, although underlying mechanisms remain largely elusive. Here we show that the expression of RNF5 (aka RMA1), an ER-anchored RING finger E3 ligase implicated in muscle organization and in recognition and processing of malfolded proteins, is elevated and mislocalized to cytoplasmic aggregates in biopsies from patients suffering from sporadic-Inclusion Body Myositis (sIBM). Consistent with these findings, an animal model for hereditary IBM (hIBM), but not their control littermates, revealed deregulated expression of RNF5. Further studies for the role of RNF5 in the pathogenesis of s-IBM and more generally in muscle physiology were performed using RNF5 transgenic and KO animals. Transgenic mice carrying inducible expression of RNF5, under control of β-actin or muscle specific promoter, exhibit an early onset of muscle wasting, muscle degeneration and extensive fiber regeneration. Prolonged expression of RNF5 in the muscle also results in the formation of fibers containing congophilic material, blue-rimmed vacuoles and inclusion bodies. These phenotypes were associated with altered expression and activity of ER chaperones, characteristic of myodegenerative diseases such as s-IBM. Conversely, muscle regeneration and induction of ER stress markers were delayed in RNF5 KO mice subjected to cardiotoxin treatment. While supporting a role for RNF5 Tg mice as model for s-IBM, our study also establishes the importance of RNF5 in muscle physiology and its deregulation in ER stress associated muscular disorders

Distamycin A Inhibits HMGA1-Binding to the P-Selectin Promoter and Attenuates Lung and Liver Inflammation during Murine Endotoxemia

Background: The architectural transcription factor High Mobility Group-A1 (HMGA1) binds to the minor groove of AT-rich DNA and forms transcription factor complexes (“enhanceosomes”) that upregulate expression of select genes within the inflammatory cascade during critical illness syndromes such as acute lung injury (ALI). AT-rich regions of DNA surround transcription factor binding sites in genes critical for the inflammatory response. Minor groove binding drugs (MGBs), such as Distamycin A (Dist A), interfere with AT-rich region DNA binding in a sequence and conformation-specific manner, and HMGA1 is one of the few transcription factors whose binding is inhibited by MGBs. Objectives: To determine whether MGBs exert beneficial effects during endotoxemia through attenuating tissue inflammation via interfering with HMGA1-DNA binding and modulating expression of adhesion molecules. Methodology/Principal Findings: Administration of Dist A significantly decreased lung and liver inflammation during murine endotoxemia. In intravital microscopy studies, Dist A attenuated neutrophil-endothelial interactions in vivo following an inflammatory stimulus. Endotoxin induction of P-selectin expression in lung and liver tissue and promoter activity in endothelial cells was significantly reduced by Dist A, while E-selectin induction was not significantly affected. Moreover, Dist A disrupted formation of an inducible complex containing NF-κB that binds an AT-rich region of the P-selectin promoter. Transfection studies demonstrated a critical role for HMGA1 in facilitating cytokine and NF-κB induction of P-selectin promoter activity, and Dist A inhibited binding of HMGA1 to this AT-rich region of the P-selectin promoter in vivo. Conclusions/Significance: We describe a novel targeted approach in modulating lung and liver inflammation in vivo during murine endotoxemia through decreasing binding of HMGA1 to a distinct AT-rich region of the P-selectin promoter. These studies highlight the ability of MGBs to function as molecular tools for dissecting transcriptional mechanisms in vivo and suggest alternative treatment approaches for critical illness