According to farm records, cultured Haliotis midae (50-70 g.abalone⁻¹) were growing 10% slower in winter when compared to summer. This reduction in growth rate also coincided with enlarged gonads. Initial trials showed that there were differences in mean monthly growth rates ranging from 1.97 – 5.14 g abalone⁻¹ month⁻¹, and gonad bulk index (GBI) also varied between months (GBI range: 26.88 ± 12.87 to 51.03 ± 34.47). The investment of energy into gonad tissue growth did not compromise whole body growth as the abalone continued to gain weight throughout the reproductive periods, probably due to gonadal growth. Growth of this size class of abalone was not influenced by water temperature or day length, suggesting favourable on-farm culture conditions (regression analyses, p > 0.05). There is no need to implement a seasonal dietary regime. Cultured H. midae were fed artificial diets with different protein sources, including only soya, only fishmeal, a combination of soya and fishmeal, and these were compared to kelp-fed abalone. Kelp-fed abalone grew slower than those fed artificial feeds (p>0.05). Gonad growth was the greatest when soya meal was included in the diet (average GBI: 74.91 ± 23.31), while the average gonad size of abalone fed the fishmealbased diet had gonads which were 38% smaller, and kelp-fed abalone had gonads which were 75% smaller than those of the abalone fed on diets containing soya meal. The increased gonad mass in abalone fed on diets including soya meal could be attributed to phytoestrogenic activity, as a result of the presence of isoflavones found in the soya plant; this remains to be tested. The use of soya in brood stock diet development is advised. The influence of dietary protein to energy ratio (1.41 – 2.46 g MJ⁻¹) on growth and gonad size was tested. Protein and energy levels within the ranges tested (22 and 33% protein; 13.5 and 15.6 MJ kg⁻¹) did not interact to influence growth rates of cultured H. midae. GBI increased from 50.67 ± 4.16 to 83.93 ± 9.35 units as a function of dietary protein to energy ratio (y = 42.02 x⁰·⁸¹; r² = 0.19; regression analysis: F₁¸₃₈ = 8.9; p = 0.005). In addition, protein level influenced gonad size, with gonad growth being greater in abalone fed the high protein diet (factorial ANOVA: F₁¸₃₂ = 7.1, p = 0.012). Canning yields were reduced by 7% when the protein content was increased, while increasing the quantity of dietary energy improved canning yields by ~ 6% (one-way ANOVA: F₁¸₂₈ = 14.4, p= 0.001). The present study provided evidence that although growth rates are varying seasonally, reproductive investment is not hindering weight gain. Gonad growth can be influenced if desired by farms, depending on the level of soya inclusion, as well as the protein to energy ratio in the diet. Monthly variation in growth and gonad size, as well as the influence of diet on gonad growth were highlighted, and the implications for farm application and further research were discussed.

Abalone, Haliotis midae, farmers in South Africa that feed formulated diets reported a periodic drop in abalone growth during periods of increased gonad development. A large drop in abalone biomass was noticed after presumed spawning events. This study was aimed to determine the effect of diet and sex-sorting on gonad development in abalone. Experiments were conducted on a commercial abalone farm from July 2012 to the end of June 2013. Isonitrogenous and isoenergetic diets were formulated with two protein sources. A fishmeal and soybean meal (S-diet) diet and a fishmeal only (F-diet) diet were fed to abalone (50 - 70 g abalone⁻¹) over 12 months. Weight and length gain, gonad bulk index (GBI), visceral index (%) and meat mass index (%) were determined monthly and seasonally. A histological study on the female gonads was conducted. This study also included an experiment to test the effect of sex-sorting (70 - 80 g abalone⁻¹) on growth and body composition with treatments including males (M), females (F) and equal numbers of males and females (MF). Weight gain and length gain were faster in S-diet-fed abalone (RM-ANOVA, F ₍₁, ₁₆₎ = 7.77, p = 0.01; F ₍₁, ₆₉₎ = 49.9, p < 0.001, respectively). Gonad development was significantly affected by the inclusion of soybean meal with S-diet-fed abalone showing higher GBI-values than F-diet-fed abalone (RM-ANOVA, F ₍₁, ₃₃)= 16.22, p = 0.0003). Male abalone had higher GBI-values than females (RM-ANOVA, F ₍₁, ₃₃₎ = 39.87, p < 0.0001). There was no significant difference in average feed conversion ratio (FCR) between diets over time (RM-ANOVA, F ₍₁, ₂₁₎ = 0.008, p = 0.97). However, average FCR-values were significantly highest between November 2012 and March 2013, the presumed spawning season. The visceral mass (gut and gonad) as a proportion of whole mass (visceral index, %) was significantly higher in abalone fed the S-diet (RM-ANOVA; F ₍₁, ₆₉₎ = 68.06, p < 0.0001). There was no difference in meat mass index (%) between diets for both male and female abalone (RM-ANOVA; F ₍₇, ₂₄₈₎ = 0.80, p = 0.60; F ₍₇, ₂₄₁₎ = 1.7, p = 0.11,respectively). Meat mass index significantly decreased from September 2012 to February 2013 coinciding with the period of high GBI-values. The distribution of oocyte maturity stages differed between diets. The majority of oocytes within S-diet-fed abalone were fully mature stage 8 oocytes compared to a majority of stage 7 oocytes in F-diet-fed abalone. Histology corroborated peaks in GBI-values for abalone fed both diets. There was no significant difference in growth, GBI, visceral index (%) and meat mass index (%) between abalone sorted into monosex and mixed-sex populations. Thus, the presence of the opposite sex did not have an effect on growth and gonad mass in H. midae. The phytoestrogens daidzin, glycitin, genistin, daidzein, glycitein and genistein were present in soybean meal and only traceable amounts were found in the F-diet. This study provided evidence that soybean meal present in formulated feed affected growth and gonad development in H.midae. The difference in the distribution of the maturity stages of oocytes was affected by diet. Sex-sorting abalone into monosex and mixed-sex populations had no influence on weight and length gain and gonad development.

Dans les écosystèmes marins côtiers, l‘activité anthropique et les paramètres naturels induisent chez lesorganismes aquatiques des situations de «multistress». Parmi ces paramètres, deux d‘entre eux ont été étudiés:La contamination métallique et l’infection bactérienne. Dans ce contexte la compréhension des réponsesmoléculaires et physiologiques de l’ormeau rouge Haliotis rufescens a été étudiée, en particulier, face l’impactmétallique (cuivre), et au pathogène (Vibrio parahaemolyticus) en interaction avec des bactéries probiotiquespendant le stade juvénile de ce mollusque en conditions contrôlées. Dans un première temps, une évaluation surl’effet d’un consortium probiotique formé de trois souches bactériennes: Vibrio sp C21, Agarivorans albus F1et Vibrio sp F15 a été testée et les paramètres de survie et de croissance des juvéniles de l’ormeau ont été suivis.Les résultats montrent que les probiotiques augmentent les performances de ces deux paramètres de trait de viecomparés aux individus témoins sans probiotiques. Dans un second temps, une approche génomique parbanques soustractives a été réalisée afin d’identifier les transcrits régulés par l’impact métallique. Cetteméthode a permis d’identifier 108 gènes potentiellement impliqués dans la réponse cellulaire et physiologiquedes organismes face au stress. La cinétique de l’expression génique a été suivie à l’aide de 14 transcrits et leurvariation temporelle pendant la phase critique révèle des régulations parfois complexes et différentielles chezles organismes. Ensuite, les réponses immunitaires des hémocytes d’ormeau face l'exposition au cuivre ont étérecherchées. Les résultats montrent que l’exposition au métal provoque des disfonctionnement importants desactivités hémocytaires. Enfin, l’interaction entre les bactéries pathogènes et probiotiques ont étés étudiées etleurs effets sur le mollusque ont été également recherchés. La présence des bactéries pathogènes régulel’expression génique des transcrits impliqués dans des fonctions physiologiques clés des organismes (lesdéfenses immunitaires et l’état énergétique). De plus, les résultats de cette interaction nous renseignent sur lerôle capital accompli par les bactéries probiotiques sur les performances en termes d’amélioration de la survieet de la croissance des jeunes ormeaux en condition de stress. / In coastal ecosystems, anthropogenic activity and natural factors induce “multi-stress” situations. Inthis study, we focused on two such stress-inducing factors: metal contamination and bacterial infection.Specifically, under controlled conditions, we examined the molecular and physiological responses of redabalone Haliotis rufescens toward the impact of a metal (copper) and a pathogen (Vibrio parahaemolyticus)with respect to this mollusk’s interaction with probiotic bacteria during the juvenile stage. First, we assessed theeffect of a consortium of three probiotic bacterial strains—Vibrio sp. C21, Agarivorans albus F1, and Vibrio sp.F15—on two abalone lifecycle-related parameters, namely, survival and growth. The results showed that,compared to the control treatment (no probiotics), the probiotics improved the performance of these twolifecycle parameters. In a second step, the molecular mechanisms underlying the juvenile abalone response tocopper stress were studied under experimental conditions, using a suppression subtractive hybridizationmethod. This method identified 108 partial sequences of genes involved in the cellular and physiologicalresponses to stress. The kinetics of gene expression were followed using 14 of these transcripts, and theirtemporal variations during the critical phase showed complex and differential regulation exerted by copper inthese organisms. We then investigated the immune response of abalone hemocytes to copper exposure. Theresults showed that exposure to the metal causes a significant dysfunction of hemocyte activities. Finally, theinteraction between probiotics and pathogenic bacteria (transcriptomic and cellular aspects) was studied and theeffects of these bacteria on mollusk performance were also investigated. The presence of pathogenic bacteriawas found to regulate gene expression of transcripts involved in key physiological functions, including thosethat regulate immune responses and energy metabolism. Examination of these interactions thus indicates therole of probiotic bacteria in performance in terms of the improved survival and growth of juvenile abaloneunder stress conditions.

Depuis une quinzaine d’années, des mortalités estivales d’ormeaux européens, Haliotis tuberculata, surviennent sur le littoral breton et normand, et en structures aquacoles. Ces mortalités sont attribuées à l’espèce bactérienne Vibrio harveyi, et se produisent chez des ormeaux sexuellement matures lorsque la température de l’eau dépasse 17°C.Ce travail de thèse visait en une approche multifactorielle de l’étude de cette interaction hôte-parasite, afin de spécifier les conditions intrinsèques aux ormeaux dans le déclenchement de cette vibriose, le cycle infectieux de V. harveyi chez l’ormeau européen et le rôle de la température dans l’accomplissement de ce cycle infectieux, et enfin la réponse physiologique de l’ormeau lors d’une exposition à V. harveyi.Les principaux résultats montrent un différentiel d’expression génomique entre des ormeaux résistants et des ormeaux sensibles au cours d’une exposition à V. harveyi, attestant ainsi l’importance du statut physiologique de l’hôte dans la survie à la vibriose chez l’ormeau européen. Ce constat est supplémenté de la mise en évidence de sensibilité à cette maladie chez des ormeaux sexuellement immatures, habituellement résistants, acclimatés à 19°C et exposés à des conditions contraignantes de type manipulation. Par ailleurs, l’étude de la voie d’entrée et de la dynamique d’infection de V. harveyi chez l’ormeau européen a révélé un tropisme particulier de ce vibrion pathogène vers les tissus branchiaux dès les premières heures de contact, et son invasion dans le système circulatoire dès 24h de contact. L’étude de la réponse hémocytaire des ormeaux et du métabolisme branchial, à l’échelle moléculaire et cellulaire, lors des premières heures de contact, démontre 1/ la genèse d’un stress oxydatif au niveau des branchies d’ormeaux sensibles à la vibriose et 2/ une altération du fonctionnement des hémocytes, ce qui présume de l’une des stratégies majeures de virulence de V. harveyi. / For fifteen years, summer mortalities have been observed in wild and farmed populations of European abalone, Haliotis tuberculata, along the north French coast. These mortalities are attributed to the bacterial species Vibrio harveyi and occur in sexually mature animals, when the seawater temperature exceeds 17°C.A multifactorial approach to the study of this host-parasite interaction was done during this thesis, in order to specify the intrinsic abalone conditions in vibriosis mortalities, the infectious cycle of V. harveyi in European abalone and the role of temperature in the fulfillment of this infectious cycle, and finally the physiological response of abalone, at cellular and molecular level, when exposed to V. harveyi.The main results showed a differential gene expression between resistant and susceptible abalone during exposure to V. harveyi, indicating the importance of the physiological status of the host, in survival to vibriosis. This hypothesis is supplemented by the susceptibility of sexually immature abalone at 19°C to this disease, usually resistant, and exposed to manipulation stressor. Moreover, the study of the portal of entry and the dynamics of infection by V. harveyi in European abalone revealed a particular tropism of this vibrio pathogen for gill tissues, in the earlier hours of contact, and its invasion into the circulatory system from the first 24 hours of contact. The study of the response of abalone hemocyte and gill metabolism, at the cellular and molecular level, in the earlier hours of contact, shows 1/ a genesis of oxidative stress in gills of susceptible abalone, and 2/ an alteration of hemocyte functions, which may constitute one of the major strategies of virulence in V. harveyi.

Thesis (Msc Food Sc (Food Science))--University of Stellenbosch, 2009. / An 84-day study was conducted to find a suitable diet and feeding level for the culture of South African abalone (Haliotis midae) in Namibia. Two experimental diets, namely, a seaweed diet (SWD) Laminaria pallida (macrophytic algae) and a formulated diet (FD) (macro-algae), for use in abalone (Haliotis midae) feed development, were evaluated. The animals used in this study were juveniles (24.33 ± 3.14 mm shell length; 2.72 ± 0.83 g live weight, mean ± SE) and sub-adults (58.07 ± 10.33 mm shell length and 41.96 ± 20.61 g live weight, mean ± SE). The nutrient profile of the SWD and FD displayed no differences in the protein and carbohydrate levels. Crude protein levels ranged from 4.91 to 17.68% (dry matter (DM) basis). The lipid levels in the FD (0.25%) were almost 0.56% lower than that in the SWD (0.76%). The feed conversion ratio (FCR) and protein efficiency ratio (PER) for the sub-adult abalone ranged from 2.80 to 10.90 and 0.10 to 0.40, respectively. The juvenile abalone fed on the FD yielded significantly lower (P < 0.05) FCRs (0.8) and higher PERs (1.20) than their counterparts fed on the SWD. A similar trend was observed for the sub-adult abalone although the differences were not significant (P > 0.05). The relative growth rate (RGR) of juvenile fed on the FD was 25% lower compared to those fed on the SWD, while that of the sub-adult abalone fed on the FD was 29% lower compared to the abalone fed on the SWD. From the daily growth rate (DGR) in terms of daily body weight (DGRBW) calculated after the 84-day period, repeated-measures ANOVA (RANOVA) indicated no interaction between time period and diet. Although slightly lower, the DGRBW for the juvenile abalone fed on the SWD diet (0.033 g/day) did not differ significantly from the DGRBW of abalone fed on the FD (0.079 g/day). In contrast, sub-adult abalone fed on the SWD exhibited significantly higher DGRBW compared to those fed on the FD. Although the abalone fed on the FD was slightly higher in nutritional content, there was no significantly difference (P > 0.05) in the nutritional profile of the abalone soft body tissue fed on either the SWD or FD. There was no significant (P > 0.05) difference in preference when comparing the aroma of the abalone meat samples fed on either the SWD or FD. However, there was a significant difference (P < 0.05) in the consumers’ preference in terms of flavour for the abalone sample fed on the FD. The trained taste panel results indicated that there was no difference in the aroma and flavour of the abalone fed on the different diets (P > 0.05). This study showed that cultured juvenile H. midae, readily accepted a FD, producing high consumption and survival rates. The FD still warrants further refinement and testing for it to become a more effective mariculture feed with commercial potential.

L’augmentation de température que subit la planète ces dernières décennies a de nombreuses conséquences dont la recrudescence de maladies infectieuses aussi bien chez l’homme que chez les animaux. Certaines populations de l’ormeau européen Haliotis tuberculata, vivant dans les zones les plus chaudes de Bretagne et de Normandie ont ainsi subi de très importantes mortalités depuis 1997, dues à la bactérie Vibrio harveyi. Cependant, certaines des populations les plus sévèrement touchées se sont aujourd’hui reconstruites et les mortalités semblent s’être arrêtées dans certaines de ces zones. La question se pose donc de l’apparition d’une résistance de l’ormeau face à cette maladie émergente. Pour répondre à cette question, les réponses à l’infection de plusieurs populations naturelles par cette bactérie ont été analysées. Une population présentant une forte résistance à la maladie a été identifiée.La voie d’entrée de la bactérie (ie. les branchies) a été identifiée comme jouant un rôle dans la résistance à l’infection. Par ailleurs, des infections successives ont permis de démontrer un effet d’amorçage immunitaire. Suite à une première exposition, une protection durant jusqu’à deux mois intervient contre l’effet d’inhibition de la phagocytose, provoquée normalement par une infection à V. harveyi. La différence d’expression de gènes des hémocytes d’ormeaux sensibles et résistants a été quantifiée par RNAseq pendant une infection expérimentale. Cette comparaison a montré une reconnaissance plus efficace du pathogène chez les résistants, par des récepteurs tels que les TLR ou les PGRP. La forte surexpression chez la population résistante, d’un gène impliqué dans la synthèse de mucine qui est l’un des composants principaux du mucus renforce l’hypothèse d’une forte implication des branchies dans la résistance. Enfin, une analyse in silico des séquences obtenues en RNAseq a permis d’apporter des preuves de l’existence d’un système de méthylation de l’ADN chez H. tuberculata ainsi qu’une possible implication de ce système dans l’adaptation de l’ormeau à son milieu. / Increasing global temperatures have numerous consequences for marine ecosystems, including the rise of infectious diseases. Certain populations of the European abalone Haliotis tucerculata have suffered from severe and recurrent mortality since 1997 due to infection caused by the bacterium Vibrio harveyi, particularly in areas with higher average summer temperatures. Given the spatial heterogeneity in mortalities, and the observation that the historically most severely impacted populations have recovered in recent years, the question of the emergence of resistance to the disease was addressed. The mortality rate in response to infection by V. harveyi was quantified experimentally in abalone originating from three natural populations, and one population exhibiting resistance to the disease was identified. In a subsequent experiment, the immune response of abalone was compared between infected individuals from a resistant and from a susceptible population. The portal of entry of the bacterium (ie. gills) was identified as playing a role in resistance. Furthermore, successive exposures of abalone to the bacterium demonstrated an immune priming effect, such that following a first exposure, phagocytosis was no longer inhibited by infection with V. harveyi, and that this improved protection against the disease lasted for at least two months. Differences in gene expression was quantified by RNAseq in the hemocytes of resistant and susceptible abalone following exposure to the pathogen. This comparison showed that resistant abalone had more effective recognition of the bacterium by receptors as the TLR or PGRP. The substantial over-expression of a gene involved in the synthesis of mucin, the main component of mucus, (UDP-GalNAC) in the resistant population, supports the interpretation of a strong involvement of gills in the resistance. Finally, an in-silico analysis of the sequences obtained from RNAseq indicate the existence of a DNA methylation system in H. tuberculata and suggested an involvement of epigenetic mechanisms in the adaptation of abalone to its environment.