Viviane Boulo - Academia.edu (original) (raw)

Papers by Viviane Boulo

HAL (Le Centre pour la Communication Scientifique Directe), Jun 2, 2022

Abstract.-Hybridoma technology is reviewed, and the characteristics of monoclonal antibodies are ... more Abstract.-Hybridoma technology is reviewed, and the characteristics of monoclonal antibodies are compared with those of polyclonal antibodies. The contribution of monoclonal antibodies to molluscan pathology is developed with special emphasis on their use as diagnostic tools. The results of studies with monoclonal antibodies prepared against the protozoan oyster pathogen Bonamia ostreae are briefly described. The development of hybridoma technology as elaborated by Kohler and Milstein (1975) has made an impact in many fields ofbiological research such as immunology, biochemistry, and pathology (Yelton and Scharff 1981; Krakauer 1985; Seiler et al. 1985). In pathology, mouse monoclonal anti bodies have been used in diagnosis (Van Der Auwera 1987) and therapy (Blythman et al. 1981; Frankel 1985). In this article, we briefly review hybridoma technology. We compare the properties of monoclonal and polyclonal antibodies, and we consider the prospects for use of monoclonal anti bodies in ...

Aquaculture, 2017

Fasting and refeeding occur naturally in predators but this is largely ignored when dealing with ... more Fasting and refeeding occur naturally in predators but this is largely ignored when dealing with farmed fish. Therefore, the effects of 3-week fasting and re-alimentation (2.5% of the individual body mass) were investigated using two genetically selected populations (F2 generation) of 250 g juvenile sea bass (Dicentrarchus labrax L.). Blood osmolarity, gill and intestinal morphology and expression of the sodium pump (Na + , K +-ATPase, NKA) were studied on two phenotypes showing different degrees of body mass loss during food deprivation: one group losing body mass rapidly during fasting (F +) and the other one limiting body mass loss during the same period (F-). Blood osmotic pressure significantly decreases due to re-alimentation in both groups, but this is compensated in the F + group. In this group, gill ionocytes are smaller and less numerous, but a significantly higher NKA gene expression is noted in the gills in comparison to the F-individuals 48 and 72 h after re-alimentation, and also in the posterior intestine 72 h after re-alimentation. This most probably occurs to compensate for a higher salt intake during nutrient absorption in comparison to the F-group. Furthermore, refed F-fish absorb more lipids along the proximal anterior intestine, and take longer to digest than the F + group, and show enterocyte vacuolization in the posterior intestine. Therefore, the two selected populations have different postprandial digestive strategies: the F-fish optimize feed efficiency first at the cost of optimal hydromineral adjustment, while the F + group invests in osmoregulatory performance at the expense of digestive physiology. Please note that this is an author-produced PDF of an article accepted for publication following peer review. The definitive publisher-authenticated version is available on the publisher Web site. Statement of relevance Our paper is highly relevant to the general field of commercial aquaculture. There is an increasing number of research articles dealing with fasting and refeeding in commercial fish and how to improve fish nutrition based on these physiological data and genetic selection. Highlights ► This study focused on differences observed in the gills and intestine of two selected populations with contrasting tolerance to fasting. ► The two phenotypes show different degrees of body mass loss during food deprivation: one group losing body mass rapidly during fasting (F +) and the other one limits body mass loss during the same period (F-). ► During feeding, these two phenotypes have different physiological optimization / prioritization and energetic benefits. ► We evidence that the fast feeders (F +) are also the bad fasters.

Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2016

In this comparative study, osmoregulatory mechanisms were analyzed in two closely related species... more In this comparative study, osmoregulatory mechanisms were analyzed in two closely related species of palaemonid shrimp from Brazil, Macrobrachium pantanalense and Macrobrachium amazonicum. A previous investigation showed that all postembryonic stages of M. pantanalense from inland waters of the Pantanal are able to hyper-osmoregulate in fresh water, while this species was not able to hypoosmoregulate at high salinities. In M. amazonicum originating from the Amazon estuary, in contrast, all stages are able to hypo-osmoregulate, but only first-stage larvae, late juveniles and adults are able to hyper-osmoregulate in fresh water. The underlying molecular mechanisms of these physiological differences have not been known. We therefore investigated the expression patterns of three ion transporters (NKA α-subunit, VHA B-subunit and NHE3) following differential salinity acclimation in different ontogenetic stages (stage-V larvae, juveniles) of both species. Larval NKAα expression was at both salinities significantly higher in M. pantanalense than in M. amazonicum, whereas no difference was noted in juveniles. VHA was also more expressed in larvae of M. pantanalense than in those of M. amazonicum. When NHE3 expression is compared between the larvae of the two species, further salinity-related differences were observed, with generally higher expression in the inland species. Overall, a high expression of ion pumps in M. pantanalense suggests an evolutionary key role of these transporters in freshwater invasion.

Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 2016

The role of the main ion transporting enzyme Na+/K+-ATPase in osmoregulation processes was invest... more The role of the main ion transporting enzyme Na+/K+-ATPase in osmoregulation processes was investigated in Litopenaeus stylirostris. The development and localization of the osmoregulation sites were studied during ontogenesis by immunodetection of Na + K +-ATPase using monoclonal antibodies and transmission electron microscopy (TEM). Osmoregulation sites were identified as the pleurae and branchiostegites in the zoeae and mysis stages. In the subsequent post-metamorphic stages the osmoregulatory function was mainly located in the epipodites and branchiostegites and osmotic regulation was later detected in the gills. The presence of ionocytes and microvilli in these tissues confirmed their role in ionic processes. The complete open reading frame of the mRNA coding for the α-subunit of Na+K+-ATPase was characterized in L. stylirostris. The resulting 3092-bp cDNA (LsNKA) encodes a putative 1011-amino-acid protein with a predicted molecular mass of 112.3 kDa. The inferred amino acid sequence revealed that the putative protein possesses the main structural characteristics of the Na+K+-ATPase α-subunits. Quantitative RT-PCR analyses indicated that LsNKA transcripts did not significantly vary between the different developmental stages. The number of transcripts was about 2.5-fold higher in the epipodites and gills than in any other tissues tested in juveniles. A reverse genetic approach was finally implemented to study the role of LsNKA in vivo. Knock down of LsNKA expression by gene-specific dsRNA injection led to an increase of shrimp mortality following an abrupt salinity change compared to control animals. These data strongly suggest that LsNKA plays an important role in osmoregulation when the shrimp are challenged by changing salinities.

The flat oyster Ostrea edulis was originally reared in several regions of the European waters. Th... more The flat oyster Ostrea edulis was originally reared in several regions of the European waters. The tidal saline ponds on the Atlantic coasts of France were highly productive before being hit by two consecutive epizooties. Parasitic diseases were described on this species, both of them leading 10 economic losses, as heavy mortalities have usually been recorded before the oysters reach the commercial size. Repeated experiments showed that the two diseases, Marteilia refringens and Bonamia ostreae are still active in this environment. Recently, some evidences were given that haemocytoblasts of Crassostrea gigas had the ability to lyze the Bonamia cells. Therefore, in an experiment of contiguous culture of the two species of oysters, the growth, mortality and prevalence of the two diseases were recorded of two populations of the fiat oyster. The prevalence of Marteilia was higher then the one of Bonamia (49.8 % against 7.4 %). The oysters originated from the area exhibited a better grow...

Frontiers in Physiology, 2011

The successful establishment of a species in a given habitat depends on the ability of each of it... more The successful establishment of a species in a given habitat depends on the ability of each of its developing stages to adapt to the environment. In order to understand this process we have studied the adaptation of a euryhaline fish, the sea-bass Dicentrarchus labrax, to various salinities during its ontogeny. The expression and localization of Aquaporin 1a (AQP1a) mRNA and protein were determined in different osmoregulatory tissues. In larvae, the sites of AQP1a expression are variable and they shift according to age, implying functional changes. In juveniles after metamorphosis (D32-D48 post-hatch, 15-25 mm) and in pre-adults, an increase in AQP1a transcript abundance was noted in the digestive tract, and the AQP1a location was observed in the intestine. In juveniles (D87-D100 post-hatch, 38-48 mm), the transcript levels of AQP1a in the digestive tract and in the kidney were higher in sea water (SW) than at lower salinity. These observations, in agreement with existing models, suggest that in SW-acclimated fish, the imbibed water is absorbed via AQP1a through the digestive tract, particularly the intestine and the rectum. In addition, AQP1a may play a role in water reabsorption in the kidney. These mechanisms compensate dehydration in SW, and they contribute to the adaptation of juveniles to salinity changes during sea-lagoon migrations. These results contribute to the interpretation of the adaptation of populations to habitats where salinity varies.

Journal of Experimental Biology, 2006

SUMMARYThis study combines a cellular and molecular analysis of the Na+/K+/2Cl- cotransporter (NK... more SUMMARYThis study combines a cellular and molecular analysis of the Na+/K+/2Cl- cotransporter (NKCC) to determine the osmoregulatory role of this protein in different tissues during the ontogeny of the sea bass. We have characterized the complete sequence of the NKCC1 isoform isolated from the sea bass gills and have identified, by immunofluorescence, NKCC1, and other isoforms, within the epithelium of the major osmoregulatory organs. Different (absorptive and secretory) functions have been attributed to this protein according to the tissue and salinity. The effects of short- (1-4 days), medium- (7-21 days) and long (6 months)-term freshwater (FW) adaptations were investigated, in comparison with seawater(SW)-maintained sea bass. In adult sea bass after long-term adaptation to FW and SW, the gills had the highest expression of NKCC mRNA compared with the median/posterior kidney and to the posterior intestine. Expression of NKCC mRNA in the kidney was 95% (SW) and 63% (FW) lower, and...

Research on bonamiasis, an epizootic disease of the edible oyster Ostrea edu/is caused by the pro... more Research on bonamiasis, an epizootic disease of the edible oyster Ostrea edu/is caused by the protozoan Bonamia ostreae, is discussed in relation to oyster fanning, research technology, epizootiology, and management. Morphological and infectious characteristics of the parasite are described. Recent progress in isolation and purification of the parasite have permitted investigations into host defense mechanisms, parasite infectivity, and the development of a mol1usc-pathogen mode!.

Microbes and Infection, 2006

The strategies used by bacterial pathogens to circumvent host defense mechanisms remain largely u... more The strategies used by bacterial pathogens to circumvent host defense mechanisms remain largely undefined in bivalve molluscs. In this study, we investigated experimentally the interactions between the Pacific oyster (Crassostrea gigas) immune system and Vibrio aestuarianus strain 01/32, a pathogenic bacterium originally isolated from moribund oysters. First, an antibiotic-resistant V. aestuarianus strain was used to demonstrate that only a limited number of bacterial cells was detected in the host circulatory system, suggesting that the bacteria may localize in some organs. Second, we examined the host defense responses to V. aestuarianus at the cellular and molecular levels, using flow-cytometry and real-time PCR techniques. We showed that hemocyte phagocytosis and adhesive capabilities were affected during the course of infection. Our results also uncovered a previouslyundescribed mechanism used by a Vibrio in the initial stages of host interaction: deregulation of the hemocyte oxidative metabolism by enhancing the production of reactive oxygen species and downregulating superoxide dismutase (Cg-EcSOD) gene expression. This deregulation may provide an opportunity to the pathogen by impairing hemocyte functions and survival. These findings provide new insights into the cellular and molecular bases of the host-pathogen interactions in C. gigas oyster.

Marine Biotechnology, 2009

Summer mortality of the Pacific oyster Crassostrea gigas is the result of a complex interaction b... more Summer mortality of the Pacific oyster Crassostrea gigas is the result of a complex interaction between oysters, their environment, and pathogens. Heredity appears to be a major factor determining the sensitivity of oysters to summer mortality, allowing resistant (R) and susceptible (S) lines to be produced. We conducted genome-wide expression profiling of R and S gonads during the 3-month period preceding a summer mortality event, using a cDNA microarray that we designed. ANOVA analysis revealed that 34 genes were differentially expressed between R and S lines on four dates preceding the mortality event. Annotation of some of these genes highlights reproduction and its allocation and antioxidant defenses as the main pathways that operate differentially between R and S lines. This transcriptional analysis provides new indications to define markers for quantitative trait loci searches and functional studies and evaluate the potential role of each gene in the resistance to summer mortality.

Journal of Invertebrate Pathology, 1992

Abstract Based on parasite purification and hemocyte primary culture, an in vitro system was esta... more Abstract Based on parasite purification and hemocyte primary culture, an in vitro system was established for studying early interactions between Bonamia ostreae and the hemocytes of sensitive Ostrea edulis and resistant Crassostrea gigas. Infections were observed ...

Marine biotechnology (New York, N.Y.)

The eastern oyster, Crassostrea virginica, and the Pacific oyster, C. gigas, are species of globa... more The eastern oyster, Crassostrea virginica, and the Pacific oyster, C. gigas, are species of global economic significance as well as important components of estuarine ecosystems and models for genetic and environmental studies. To enhance the molecular tools available for oyster research, an international group of collaborators has constructed a 27,496-feature cDNA microarray containing 4460 sequences derived from C. virginica, 2320 from C. gigas, and 16 non-oyster DNAs serving as positive and negative controls. The performance of the array was assessed by gene expression profiling using gill and digestive gland RNA derived from both C. gigas and C. virginica, and digestive gland RNA from C. ariakensis. The utility of the microarray for detection of homologous genes by cross-hybridization between species was also assessed and the correlation between hybridization intensity and sequence homology for selected genes determined. The oyster cDNA microarray is publicly available to the res...

… International pour l' …, 1991

Diseases of Aquatic Organisms, 1991

Monoclonal antibodies against Bonamia ostreae (Protozoa: Ascetospora), an intrahaemocytic parasit... more Monoclonal antibodies against Bonamia ostreae (Protozoa: Ascetospora), an intrahaemocytic parasite of flat oyster Ostrea edulis (Mollusca: Bivalvia) Herve Rogierl, Dominique Hervio2, Viviane ~o u l o~, Christine claviesl, Eliane Hervaudl, Evelyne ache re^, Eric ~i a l h e~, Henri ~r i z e l~, Bernard pau3, Francis ~a o l u c c i l ' Sanofi Recherche, Rue du Professeur,

Diseases of Aquatic Organisms, 1992

Currently recognltlon of molluscan dlseases relles heavlly upon llght mlcroscoplc examlnatlon of ... more Currently recognltlon of molluscan dlseases relles heavlly upon llght mlcroscoplc examlnatlon of stained histological sections of molluscan tissues Such histopathological methods are timeconsuming and relatively inefficient In the study of ongolng epizootics Alternatively, more effluent investigative tools are needed to accelerate d~s e a s e recognltlon and quantif~catlon Such methods are needed to prevent control and eradicate molluscan dlseases Thls paper reports the development of a direct sandwich enzyme-linked lmmunosorbent assay (ELISA) for detecting the ascetosporan intrahemocytic parasite B o n a m~a ostreae of the flat oyster Ostrea eduhs Bonamiasls 1s a commercially important epizootic dlsease of oysters A Bonam~a-speciflc monoclonal antlbody IS employed in the assay The assay is performed In antibody-coated microplates employng standardized reagent solutions and 50 ul oyster hemolymph samples This procedure was developed into a commercial product, available as a kit Based upon cllnical studles, there was a 90 O/o agreement In test results when ELISAs were compared to the reference standard, hlstopathologic light microscopic examlnatlon for the parasites

Diseases of Aquatic Organisms, 1994

Digestive glands of mussels Mytilus edulis from Brittany, France, infected with Marteilia sp. (As... more Digestive glands of mussels Mytilus edulis from Brittany, France, infected with Marteilia sp. (Ascetospora) were used to purify the parasite. A modification of a previously used purification protocol increased punfication efficiency, permitting sporangial primordia and sporangia of Marteilia sp. to be obtained. Mouse (Balb/c) monoclonal antibodies were generated against this parasite. From the fusion, 26 rnonoclonal antibodies against Marteilia sp. were obtained. Antibodies from 6 clones reacted only with Marteilia sp. cells and not with normal host tissues. Four of these antibodies (1/1-3, 311-1, 4/1-1 and 6/2-3) reacted with the sporangia wall and two with the spore cytoplasm (9/1-1 and 12/51), Antibodies cross-reacted with A4arteilia refringens from Mytilus galloprovincialis obtained in the Ria de Vigo, Spain.

Developmental & Comparative Immunology, 2007

The work presented here reports the expression of immune-related genes during ontogenesis in the ... more The work presented here reports the expression of immune-related genes during ontogenesis in the oyster Crassostrea gigas. Expression patterns of 18 selected genes showed that RNAs detected in oocytes and 2-4 cell embryos are of maternal origin and that gene transcription starts early after fertilization. The expression patterns of 4 genes (Cg-timp, Cg-tal, Cg-EcSOD and Drac3) suggested that hemocytes appear in the gastrula-trochophore stages. The localization of Cg-tal expression suggested that hematopoietic cells were derived from vessels and/or artery endothelia cells. Moreover, a bacterial challenge affected the level of expression of genes. Indeed, a change in expression levels was observed for Cg-LBP/BPI, Cg-timp, Drac3 and Cg-MyD88 genes in larval stages upon exposure to non-pathogenic bacteria. In early juveniles, a modulation was also observed for Cg-LBP/BPI, Cg-timp, Cg-MyD88 and for Cg-tal, according to the concentration of bacteria. Altogether, the results showed that studying the appearance of immunocompetent cells through their ability to express immune-related genes is a tool to gain insight the ontogenesis of the oyster immune system.