Shellfish and fish are sources of potent allergens in sensitised individuals. In view of the limited information available from published studies, especially on mollusc allergens, more detailed characterisation of the major allergens from different shellfish and fish is necessary to accurately predict cross-allergenicity. The need for a detailed characterisation of the immune response and the allergens in an indigenous mollusc species, followed a recent increase in patients presenting in the Cape Town area, with TY.pe I allergic reactions after ingestion of abalone (Haliotis midae). The first objective of this study was to determine the frequency and spectrum of reported hypersensitivity to abalone and other related seafood species in the Western Cape of South Africa. 105 volunteer subjects with suspected seafood allergy were recruited by means of a detailed seafood allergy questionnaire, advertised in the local press. The analysis of the questionnaire demonstrated clearly the importance of abalone, which was the third most frequently reported species (35%), after rock lobster and shrimp, of the 26 seafood species implicated in allergic reactions. Allergy to seafood was confirmed by the presence of specific IgE using in-house and commercial radioimmunoassays (RASTs). A novel Abalone-RAST identified specific IgE to abalone in 17/38 subjects who reported adverse reaction following the ingestion of abalone. The novel Abalone-RAST correlated positively not only with the phylogenetic closely related Snail-RAST (p<0.01) but also with squid and several indigenous crustacean species. Among subjects with multiple seafood allergies, the immune responses to molluscs species were found to be diverse. The presence of species specific allergens in the mollusc group was supported by the very low frequency of concurrent sensitivity to all mollusc species (13%) compared to concurrent sensitivity between mollusc and among the crustacea and fish group (42% and 56% respectively). The persistence of specific IgE, following a period of more than three years of seafood avoidance, was demonstrated in the mollusc species studied. The second objective was to investigate the specific immune responses to local seafood in more detail using RAST-inhibition experiments, skin prick tests (SPTs) and Western blots to demonstrate the presence of species-specific allergens. Sensitivity for the detection of abalone allergy was improved using an additional in-house SPT in six RAST negative subjects (23/38, 61 %). RAST-inhibition experiments with abalone extract demonstrated the highest degree of cross-reactivity with the Snail-RAST. In addition, a strong inhibition was achieved using commercial crustacean RASTs, confirming the presence of cross-reacting allergens in species of the same and other seafood groups. However, the low inhibitions achieved with the indigenous black mussel and squid extracts indicated that their allergen compositions differ from the species utilised in the commercial RASTs. Unique species-specific protein bands could be detected by SDS-gel electrophoresis, which clearly distinguished related mollusc species. These have not been demonstrated previously. Western blot analysis of different mollusc species identified several prominent allergens. An unexpected finding was the appearance of novel specific IgE binding reactivity after cooking the abalone. Several IgE binding proteins with similar molecular weights could be detected in immuno blots of indigenous crustacean and fish species. The next objective was to characterise the hypersensitive reactions to the local abalone species (H midae) in detail and identify the allergens found in this mollusc species. The questionnaire on abalone sensitive subjects revealed that asthma-like symptoms and the delayed onset of symptoms were frequent in sensitised individuals (42% and 34% respectively). This has previously only been reported for snail and in one case report on abalone. Surprisingly, the five abalone sensitive subjects who were studied in more detail had concurrent sensitivity to HDM and, analysed by RAST-inhibition, demonstrated two distinct types of responses. One set of subjects demonstrated a strong inhibition by HDM, indicating clearly for the first time that cross-reacting allergens or epitopes must exist between the food allergens from abalone and the air borne allergens found in HDM. SDS-gel electrophoresis demonstrated common but in addition species-specific protein bands even between very closely related abalone species from South Africa, Australia and Japan. Western blot analysis revealed two major allergens with molecular weights of about 38 and 45 kDa. Their remarkable thermal stability was demonstrated by various in-vitro and in-vivo assays. These two allergens were also present in extracts of other indigenous mollusc species, and surprisingly in some local crustacean and fish species. The individual immune responses to mollusc species were very heterogeneous for each analysed individual. The 38 kDa allergen is believed to belong to the protein family of tropomyosins, as was supported by specific IgE binding to recombinant tropomyosin of shrimp. This novel 45 kDa allergen of the South African abalone (H midae), was registered with the WHO International Union of Immunological Societies (IUIS) as Hal m 1. It is only the second allergen recognised for a mollusc species after Tod p 1 from squid. A further aim of these studies was to generate monoclonal antibodies (MoAbs) to the mollusc allergens using the hybridoma technology. The MoAbs were used, due to their consistent specific binding, to identify cross-reacting allergens among species of different seafood groups. In addition, I attempted to develop an immunologic test to distinguish between abalone species from different parts of the world. This test is of importance for the South African police to identify for forensic purposes unequivocally, fresh or processed abalone tissue exported illegally from South Africa to the Far East. Three ELISA- and Western blot assay positive clones were analysed. They demonstrated highly individual binding profiles when binding to fresh and cooked mollusc species was analysed. MoAb clones 2.11 and 2.12 (generated to the same protein of abalone) lacked complete binding to four and two of the ten abalone species respectively, allowing for very distinct species identification. Furthermore, antibody binding to cross-reacting proteins in crustacea and fish was also detected. Western blot results demonstrated clearly that these two antibodies bind to different epitopes on the same protein, making them very useful as tool for allergen and species identification. Finally, an amino acid analysis of the 3 purified antibody binding fractions of abalone was conducted. These proteins were rich in glutamine and asparagine, like tropomyosin, but differed significantly from tropomyosin with respect to serine content. The immunological findings using different patient specific sera and the monoclonal antibodies generated, provide important new information and insights into the concordant and multiple positive sensitivity to molluscs, crustacea and fish, and new information about the complexity and stability of immune responses to seafood and mollusc allergens observed in allergic subjects.

No description available.

No description available.

The polar lobe of Ilyanassa is asymmetrically partitioned into the D lineage of cells. Two of these cells, 3D and 4d, induce proper axial cell fate patterning in the embryo. Based on known embryological data in Ilyanassa, I hypothesized that Notch signaling would be required for this induction. I found that Notch signaling is required for cell fates induced by 4d and is temporally required well after 4d induction. Based on these results, I hypothesize that Notch signaling is involved in a reciprocal induction between the micromeres and the macromeres (endoderm) resulting in the maintenance of micromere fate induction and endoderm specification.Loss of the polar lobe results in the loss of cell fate induction by 3D/4d. Therefore, I hypothesized that proteins are asymmetrically bequeathed to the inducing D lineage cells by the polar lobe. To test this hypothesis, I compared global protein differences between two cell stage intact embryos, lobeless embryos, and isolated polar lobes by 2-Dimensional Electrophoresis analysis. I found several (12) quantitative differences between these samples including four spots enriched in the polar lobe isolates. I identified voltage-dependent anion-selective channel (VDAC) as one of the candidate proteins enriched in polar lobe isolates. I propose that VDAC is asymmetrically distributed by the polar lobe to the D cell and that it may function in D cell induction and mesendoderm fate specification.Lastly, I identify an acetylated tubulin antigen as a marker for cilia. I describe the pattern of cilia differentiation in the developing larvae that results in the formation of two ciliary bands, the prototroch and the metatroch, required for locomotion and feeding. These ciliary bands are conserved among annelid and mollusc larvae. Interestingly, the metatroch is derived from third quartet derivatives in the annelid Polygordius and from second quartet derivatives in the mollusc Crepidula. I provide evidence that the metatroch is derived from the first quartet derivatives in the mollusc Ilyanassa. Thus while the larval metatroch is conserved, its clonal origin is not. Based on these results, I provide support for the hypothesis that the metatroch is not homologous between annelids and molluscs or even among molluscs.

Despite having similar brains, related species can display divergent behaviors. Investigating the neural basis of such behavioral divergence can elucidate the neural mechanisms that allow behavioral change and identify neural mechanisms that influence the evolution of behavior. Fewer than three percent of Nudipleura (Mollusca, Opisthobranchia, Gastropoda) species have been documented to swim. However, Tritonia diomedea and Pleurobranchaea californica express analogous, independently evolved swim behaviors consisting of rhythmic, alternating dorsal and ventral flexions. The Tritonia and Pleurobranchaea swims are produced by central pattern generator (CPG) circuits containing homologous neurons named DSI and C2. Homologues of DSI have been identified throughout the Nudipleura, including in species that do not express a dorsal-ventral swim. It is unclear what neural mechanisms allow Tritonia and Pleurobranchaea to produce a rhythmic swim behavior using homologous neurons that are not rhythmic in the majority of Nudipleura species. Here, C2 homologues were also identified in species that do not express a dorsal-ventral swim. We found that certain electrophysiological properties of the DSI and C2 homologues were similar regardless of swim behavior. However, some synaptic connections differed in the non-dorsal-ventral swimming Hermissenda crassicornis compared to Tritonia and Pleurobranchaea. This suggests that particular CPG synaptic connections may play a role in dorsal-ventral swim expression. DSI modulates the strength of C2 synapses in Tritonia, and this serotonergic modulation appears to be necessary for Tritonia to swim. DSI modulation of C2 synapses was also found to be present in Pleurobranchaea. Moreover, serotonergic modulation was necessary for swimming in Pleurobranchaea. The extent of this neuromodulation also correlated with the swimming ability in individual Pleurobranchaea; as the modulatory effect increased, so too did the number of swim cycles produced. Conversely, DSI did not modulate the amplitude of C2 synapses in Hermissenda. This indicates that species differences in neuromodulation may account for the ability to produce a dorsal-ventral swim. The results indicate that differences in synaptic connections and neuromodulatory dynamics allow the expression of rhythmic swim behavior from homologous non-rhythmic components. Additionally, the results suggest that constraints on the nervous system may influence the neural mechanisms and behaviors that can evolve from homologous neural components.

Behaviors are a product of underlying neural circuits, yet there is a paucity of mechanistic information about how nervous systems contribute to the repeated evolution of similar behaviors. Theoretical studies have predicted that the same behavioral output can be generated by neural circuits with different properties. Here, we test the theory in biological circuits by comparing the central pattern generator (CPG) circuits underlying swimming behaviors in nudibranchs (Mollusca, Gastropoda, Euthyneura, Nudipleura). In comparative studies of neural circuits, neurotransmitter content can serve as landmarks or molecular markers for neuron types. Here, we created a comprehensive map of GABA-immunoreactive neurons in six Nudipleura species. None of the known swim CPG neurons were GABA-ir, but they were located next to identifiable GABA-ir neurons/clusters. Despite strong conservation of the GABA-ergic system, there were differences, particularly in the buccal ganglia, which may represent adaptive changes. We applied our knowledge of neurotransmitter distribution along with morphological traits to identify the neuron type Si1 in Flabellina, a species that swims via whole body left-right (LR) flexions and in Tritonia, a dorsal-ventral (DV) swimming species. Si1 is a CPG member of the LR species Melibe, whereas its homologue in the LR species Dendronotus is not. In Flabellina, Si1 was part of the LR CPG and despite having similar synaptic connections as Flabellina and Melibe, Si1 in Tritonia was not part of its DV swim CPG. Side by side circuit comparison of Flabellina, Melibe and Dendronotus revealed different combinations of circuit architecture and modulation resulting in different circuit configurations for LR swimming. This includes differences in the role and activity pattern of Si1, sensitivity to curare and the effect of homologues of C2, a DV CPG neuron, on the LR motor pattern. These results collectively reveal three different circuit variations for generating the same behavior. It suggests that the neural substrate from which behaviors arise is phylogenetically constrained. While this neural substrate can be configured in multiple different ways to generate the same outcome, the possibilities are finite and, as seen here, similar structural and functional neural motifs are used in the evolution of these circuits.

No description available.

Water management that alters riverine ecosystem processes has strongly influenced deltas and the people who depend on them, but a full accounting of the trade-offs is still emerging. Using palaeoecological data, we document a surprising biogeochemical consequence of water management in the Colorado River basin. Complete allocation and consumptive use of the river's flow has altered the downstream estuarine ecosystem, including the abundance and composition of the mollusc community, an important component in estuarine carbon cycling. In particular, population declines in the endemic Colorado delta clam, Mulinia coloradoensis, from 50-125 individuals m(-2) in the pre-dam era to three individualsm-2 today, have likely resulted in a reduction, on the order of 5900-15 000 tCyr(-1) (4.1-10.6 mol Cm-2 yr(-1)), in the net carbon emissions associated with molluscs. Although this reduction is large within the estuarine system, it is small in comparison with annual global carbon emissions. Nonetheless, this finding highlights the need for further research into the effects of dams, diversions and reservoirs on the biogeochemistry of deltas and estuaries worldwide, underscoring a present need for integrated water and carbon planning.

Parmi la diversité des types microstructuraux que l’on peut trouver dans les coquilles des Mollusques, les architectures entrecroisées (lamellaire-croisé, lamellaire-croisé complexe, folié-croisé) sont de loin les plus abondantes. Mais elles restent, de par leurs organisations tridimensionnelles complexes, peu documentées : la majorité des études de biominéralisation, et les différents modèles qui en découlent, sont en effet essentiellement basés sur des architectures plus simples (telles que les couches nacrées ou prismatiques). Or il convient, si l'on souhaite aboutir à une meilleure compréhension des mécanismes fondamentaux qui dirigent la minéralisation de ces biocarbonates, de vérifier dans quelle mesure les modèles développés à partir de coquilles ”simples” sont aussi applicables à des organisations microstructurales plus complexes. Cette étude se focalise donc sur les couches entrecroisées des coquilles de quelques Mollusques, dans le but de mettre en évidence les différents niveaux du contrôle biologique que l'organisme exerce sur leur formation et leur croissance. À cette fin, des techniques de caractérisation in-situ des assemblages organiques sont privilégiées, en relation étroite avec l’analyse des organisations microstructurales et des mécanismes de biocristallisation à fine échelle. Quelques aspects de la diagénèse de ces microstructures seront aussi abordés, à travers l’étude de coquilles fossiles de Patella sp (~100 ka) et Velates perversus (~50 Ma). / Among the variety of microstructural types that can be found within Molluscs shells, intercrossed structures (crossed-lamellar, complex crossed-lamellar and cross-foliated) are by far the most commonly found. They are, however, still poorly documented - mainly due to their complex 3D organization. The majority of biomineralization studies, and the resulting models, are indeed essentially based on simple architectures (such as nacreous or prismatic layers). In order to achieve a better understanding of the fundamental mechanisms that drive the mineralization of such biocarbonates, it is therefore mandatory to check to which extent models developed from « simple » shells stay consistent when applied to more complex microstructural organizations. The present study focuses on intercrossed layers of several Mollusk shells, in order to highlight the various levels of biological control exerted by the organism on their formation and growth. In-situ techniques are used to characterize biochemical compositions, in close correlation with microstructural patterns, as well as fine-scale biocrystallization processes. Some peculiar features of the diagenesis of these microstructures are illustrated, through the study of fossil shells from Patella sp (~100 ky) and Velates perversus (~50 My).

No description available.