Publications by authors named "María Prado-Alvárez"

11 Publications

  • Page 1 of 1

First detection of OsHV-1 in the cephalopod Octopus vulgaris. Is the octopus a dead-end for OsHV-1?

J Invertebr Pathol 2021 Feb 14:107553. Epub 2021 Feb 14.

Marine Molecular Pathobiology Group, Marine Research Institute, Spanish National Research Council, Eduardo Cabello 6, 36208 Vigo, Spain. Electronic address:

The ostreid herpes virus (OsHV-1), associated with massive mortalities in the bivalve Crassostrea gigas, was detected for the first time in the cephalopod Octopus vulgaris. Wild adult animals from a natural breeding area in Spain showed an overall prevalence of detection of 87.5% between 2010 and 2015 suggesting an environmental source of viral material uptake. Overall positive PCR detections were significantly higher in adult animals (p = 0.031) compared to newly hatched paralarvae (62%). Prevalence in embryos reached 65%. Sequencing of positive amplicons revealed a match with the variant OsHV-1 µVar showing the genomic features that distinguish this variant in the ORF4. Gill tissues from adult animals were also processed for in situ hybridization and revealed positive labelling. Experimental exposure trials in octopus paralarvae were carried out by cohabitation with virus injected oysters and by immersion in viral suspension observing a significant decrease in paralarval survival in both experiments. An increase in the number of OsHV-1 positive animals was detected in dead paralarvae after cohabitation with virus injected oysters. No signs of viral replication were observed based on lack of viral gene expression or visualization of viral structures by transmission electron microscopy. The octopus response against OsHV-1 was evaluated by gene expression of previously reported transcripts involved in immune response in C. gigas suggesting that immune defences in octopus are also activated after exposure to OsHV-1.
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http://dx.doi.org/10.1016/j.jip.2021.107553DOI Listing
February 2021

A Naïve Population of European Oyster Ostrea edulis with Reduced Susceptibility to the Pathogen Bonamia ostreae: Are S-Strategy Life Traits Providing Protection?

Integr Comp Biol 2020 08;60(2):249-260

Aquaculture and Fisheries Development Centre, School of Biological, Earth and Environmental Sciences, University College Cork, Cork, Ireland.

European populations of the native flat oyster, Ostrea edulis, have been heavily depleted by two protozoan parasites, Marteila refringens and Bonamia ostreae, with mortalities of up to 90% reported in naïve populations. However, in studies carried out over a 10-year period, researching the parasite-host relationship of B. ostreae and O. edulis in several age cohorts within a naïve O. edulis population from Loch Ryan (LR), Scotland, 1364 specimens were challenged and only 64 (5%), across multiple testing protocols, screened positive for B. ostreae. This article presents a case for the development of S-strategy life traits in the LR population that coincide with enhanced immune function and survival. Oysters are considered typical r-strategists (small in size with fast development and high fecundity) while S-strategists, as outlined in Grime's (1977) competitor-stress tolerant-ruderal (C-S-R) triangle theory, are characterized by slow growth and investment in the durability of individuals. This study hypothesizes that slower growth and reduced reproductive output in LR oysters has resulted in the investment of an enhanced immune function and reduced susceptibility to B. ostreae that is, r-strategists with S-strategy life traits equates to protection from significant pathogens. The findings presented here within provide a strong case study for local adaptation of energy allocation and provides empirical support for the C-S-R triangle theory in a marine organism.
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http://dx.doi.org/10.1093/icb/icaa071DOI Listing
August 2020

Is pallial mucus involved in Ostrea edulis defenses against the parasite Bonamia ostreae?

J Invertebr Pathol 2020 01 2;169:107259. Epub 2019 Dec 2.

Institut Français de Recherche pour ĺExploitation de la Mer (IFREMER), Laboratoire de Génétique et Pathologie (LGP), Avenue Mus de Loup, 17390 La Tremblade, France.

Bonamia ostreae is an intrahemocytic parasite that has been responsible for severe mortalities in the flat oyster Ostrea edulis since the 1970́s. The Pacific oyster Crassostrea gigas is considered to be resistant to the disease and appears to have mechanisms to avoid infection. Most studies carried out on the invertebrate immune system focus on the role of hemolymph, although mucus, which covers the body surface of molluscs, could also act as a barrier against pathogens. In this study, the in vitro effect of mucus from the oyster species Ostrea edulis and C. gigas on B. ostreae was investigated using flow cytometry. Results showed an increase in esterase activities and mortality rate of parasites exposed to mucus from both oyster species. In order to better understand the potential role of mucus in the defense of the oyster against parasites such as B. ostreae, liquid chromatography and tandem mass spectrometry were used to describe and compare mucus protein composition from both species. In all oyster species, pallial mucus contains a high level of proteins; however, O. edulis mucus produced a variety of proteins that could be involved in the immune response against the parasite, including Cu/Zn extracellular superoxide dismutase, thioxiredoxin, peroxiredon VI, heat shock protein 90 as well as several hydrolases. Conversely, a different set of antioxidant proteins, hydrolases and stress related proteins were identified in mucus from C. gigas. Our results suggest an innate immunity adaptation of oysters to develop a specific response against their respective pathogens. The mucosal protein composition also provides new insights for further investigations into the immune response in oysters.
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http://dx.doi.org/10.1016/j.jip.2019.107259DOI Listing
January 2020

Occurrence of OsHV-1 in Cultured in Ireland during an Exceptionally Warm Summer. Selection of Less Susceptible Oysters.

Front Physiol 2016 8;7:492. Epub 2016 Nov 8.

Aquaculture and Fisheries Development Centre, School of Biological, Earth and Environmental Science and Environmental Research Institute, University College Cork Cork, Ireland.

The occurrence of OsHV-1, a herpes virus causing mass mortality in the Pacific oyster was investigated with the aim to select individuals with different susceptibility to the infection. Naïve spat transferred to infected areas and juveniles currently being grown at those sites were analyzed using molecular and histology approaches. The survey period distinguishes itself by very warm temperatures reaching up to 3.5°C above the average. The virus was not detected in the virus free area although a spread of the disease could be expected due to high temperatures. Overall mortality, prevalence of infection and viral load was higher in spat confirming the higher susceptibility in early life stages. OsHV-1 and oyster mortality were detected in naïve spat after 15 days of cohabitation with infected animals. Although, infection was associated with mortality in spat, the high seawater temperatures could also be the direct cause of mortality at the warmest site. One stock of juveniles suffered an event of abnormal mortality that was significantly associated with OsHV-1 infection. Those animals were infected with a previously undescribed microvariant whereas the other stocks were infected with OsHV-1 μVar. Cell lesions due to the infection were observed by histology and true infections were corroborated by hybridization. Survivors from the natural outbreak were exposed to OsHV-1 μVar by intramuscular injection and were compared to naïve animals. The survival rate in previously exposed animals was significantly higher than in naïve oysters. Results derived from this study allowed the selection of animals that might possess interesting characteristics for future analysis on OsHV-1 resistance.
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http://dx.doi.org/10.3389/fphys.2016.00492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5099240PMC
November 2016

Whole-genome amplification: a useful approach to characterize new genes in unculturable protozoan parasites such as Bonamia exitiosa.

Parasitology 2015 Oct 18;142(12):1523-34. Epub 2015 Aug 18.

Laboratoire de Génétique et Pathologie des Mollusques Marins,IFREMER,Avenue de Mus de Loup,17390 La Tremblade,France.

Bonamia exitiosa is an intracellular parasite (Haplosporidia) that has been associated with mass mortalities in oyster populations in the Southern hemisphere. This parasite was recently detected in the Northern hemisphere including Europe. Some representatives of the Bonamia genus have not been well categorized yet due to the lack of genomic information. In the present work, we have applied Whole-Genome Amplification (WGA) technique in order to characterize the actin gene in the unculturable protozoan B. exitiosa. This is the first protein coding gene described in this species. Molecular analysis revealed that B. exitiosa actin is more similar to Bonamia ostreae actin gene-1. Actin phylogeny placed the Bonamia sp. infected oysters in the same clade where the herein described B. exitiosa actin resolved, offering novel information about the classification of the genus. Our results showed that WGA methodology is a promising and valuable technique to be applied to unculturable protozoans whose genomic material is limited.
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http://dx.doi.org/10.1017/S0031182015000967DOI Listing
October 2015

Effect of okadaic acid on carpet shell clam (Ruditapes decussatus) haemocytes by in vitro exposure and harmful algal bloom simulation assays.

Cell Biol Toxicol 2013 Jun 9;29(3):189-97. Epub 2013 May 9.

Department of Cell and Molecular Biology, Faculty of Sciences, University of A Coruña, Campus A Zapateira s/n, 15071, A Coruña, Spain.

Okadaic acid (OA), produced by dinoflagellates during harmful algal blooms (HAB), belongs to the Diarrheic Shellfish Poisoning toxins that cause gastrointestinal symptoms in humans after consumption. In the present work, Ruditapes decussatus haemocytes were selected to evaluate the effect of OA on cell viability, enzymatic status and immune capacity through the measure by flow cytometry of apoptosis-cell death, non-specific esterase activity and phagocytosis. In order to compare different exposure conditions, two experiments were developed: in vitro exposure to OA and HAB simulation by feeding clams with the OA producer, Prorocentrum lima. Apoptosis was not OA dose-dependent and cell death increased in both assays. Phagocytosis of latex beads and esterase activity decreased in haemocytes incubated with OA. In contrast, esterases increased during the feeding with P. lima. Our results showed that OA and the simulated HAB caused damages on haemocyte functions and viability.
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http://dx.doi.org/10.1007/s10565-013-9246-1DOI Listing
June 2013

Heat shock protein 90 of Bonamia ostreae: characterization and possible correlation with infection of the flat oyster, Ostrea edulis.

J Eukaryot Microbiol 2013 May-Jun;60(3):257-66. Epub 2013 Mar 12.

IFREMER, Laboratoire de Génétique et Pathologie, Avenue de Mus de Loup, 17390, La Tremblade, France.

In this study, we described the cytosolic HSP90 of Bonamia ostreae, an intracellular parasite of Ostrea edulis hemocytes. The complete open reading frame was assembled by Rapid Amplification cDNA Ends reactions on cDNA of B. ostreae-infected hemocytes. HSP90 amplification was corroborated in infected oysters and B. ostreae purified cells. The functionality of the HSP90, studied by inhibitory assays with radicicol, suggests that this protein may play a role in hemocyte invasion. Our results inform the molecular basis that governs B. ostreae-O. edulis interactions.
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http://dx.doi.org/10.1111/jeu.12031DOI Listing
October 2013

Effects of okadaic acid on haemocytes from Mytilus galloprovincialis: a comparison between field and laboratory studies.

Mar Environ Res 2012 Oct 7;81:90-3. Epub 2012 Sep 7.

Department of Cell and Molecular Biology, Campus A Zapateira s/n, University of A Coruña, A Coruña, Spain.

Individuals of Mytilus galloprovincialis, contaminated with Diarrheic Shellfish Poisoning (DSP) toxins, were studied with the aim to correlate the okadaic acid (OA) body burden and the percentage of damaged haemocytes by quantifying annexin V positive cells by flow cytometry. Results showed less percentage of damaged haemocytes in high OA contaminated samples. These data were compared with results of in vitro assays of mussel haemocytes exposed to increased concentrations of OA. Similarly, haemocytes exposed to the most concentrated OA solution were less damaged.
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http://dx.doi.org/10.1016/j.marenvres.2012.08.011DOI Listing
October 2012

Evaluation of genotoxicity in gills and hemolymph of clam Ruditapes decussatus fed with the toxic dinoflagellate Prorocentrum lima.

J Toxicol Environ Health A 2011 ;74(15-16):971-9

Department of Cell and Molecular Biology, Faculty of Sciences, University of A Coruña, Spain.

Diarrheic shellfish poisoning (DSP) is a gastrointestinal (GIT) disease that appears a few hours after ingesting okadaic acid (OA)-contaminated mollusks; okadaic acid is present in dinoflagellates of the genera Dinophysis and Prorocentrum. Toxic manifestations occur all year round at a higher or lesser intensity, and as a consequence, extractive production factories need to be closed during these periods which affects the economy of aquaculture industries. Although the concentration of harmful algae is usually found at high levels in clam digestive gland, bivalve mortality was not increased. In this study, the genotoxic effects produced by OA in clam Ruditapes decussatus were determined using the comet assay. In vitro (exposing hemocytes to different concentrations of OA) and in vivo (feeding clams with toxic dinoflagellate Prorocentrum lima) experiments were conducted in order to determine the genotoxic effects of OA on bivalve cells. Hemocytes and gill cells were analyzed by in vivo and in vitro approaches. While the in vitro study showed a rapid effect of OA on hemocytes, data obtained in the in vivo experiment reflected contradictory results dependent upon the concentration of OA and cell type evaluated. An increase in DNA damage was observed at the lower concentration and only in gill tissue. The results obtained may contribute to a better understanding of the mechanisms underlying genotoxic effects induced by OA on bivalves.
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http://dx.doi.org/10.1080/15287394.2011.582025DOI Listing
September 2011

Expression of Mytilus immune genes in response to experimental challenges varied according to the site of collection.

Fish Shellfish Immunol 2010 Apr 1;28(4):640-8. Epub 2010 Jan 1.

Ecosystèmes Lagunaires, UMR 5119, Université de Montpellier 2-CNRS, cc093, place E. Bataillon, F-34095 Montpellier cedex 05, France.

Mussels live in diverse coastal environments experience various physical, chemical and biological conditions, which they counteract with functional adjustments and heritable adaptive changes. In order to investigate possible differences in immune system capabilities, we analyzed by qPCR the expression levels of 4 immune genes (defensin, mytilin B, myticin B, lysozyme) and HSP70 in the Mediterranean mussel, Mytilus galloprovincialis collected in 3 European farming areas {Atlantic Ocean-Ría de Vigo-Spain (RV), French Mediterranean Gulf of Lion-Palavas-Prévost lagoon (PP) and Northern Adriatic Sea-Venice-Italy (VI)} in response to one injection of one of the 3 bacterial species (Vibrio splendidus LGP32, Vibrio anguillarum, Micrococcus lysodeikticus), and to heat shock or cold stress. We confirmed that the 5 genes are constitutively expressed in hemocytes, defensin being the less expressed, myticin B the highest. As suspected, the same gene resulted differently expressed according to mussel group, with the biggest difference being for HSP70 and lysozyme and lowest expression of all the 5 genes in mussels from RV. In addition, gene expression levels varied according to the challenge. Most frequent effect of bacterial injections was down-regulation, especially for mytilin B and myticin B. Heat shock enhanced transcript levels, particularly in mussels from RV, whereas cold stress had no effect. In situ hybridization of labelled probes on mussel hemocytes indicated that bacterial injections did not change the mRNA patterns of defensin and myticin B whereas mytilin B mRNA almost disappeared. In conclusion, these results demonstrated that constitutive level, nature and intensity of immune gene expression regulations strongly depended from mussel group, and support the concept of gene-environment interactions.
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http://dx.doi.org/10.1016/j.fsi.2009.12.022DOI Listing
April 2010

Immune responses of mussel hemocyte subpopulations are differentially regulated by enzymes of the PI 3-K, PKC, and ERK kinase families.

Dev Comp Immunol 2008 6;32(6):637-53. Epub 2007 Nov 6.

Department of Immunology, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Mexico City DF-04510, Mexico.

Various hemocyte cell types have been described in invertebrates, but for most species a functional characterization of different hemocyte cell types is still lacking. In order to characterize some immunological properties of mussel (Mytilus galloprovincialis) hemocytes, cells were separated by flow cytometry and their capacity for phagocytosis, production of reactive oxygen species (ROS), and production of nitric oxide (NO), was examined. Phosphatidylinositol 3-kinase (PI 3-K), protein kinase C (PKC), and extracellular signal-regulated kinase (ERK) inhibitors were also used to biochemically characterize these cell responses. Four morphologically distinct subpopulations, designated R1-R4, were detected. R1, R2, and R3 cells presented different levels of phagocytosis towards zymosan, latex beads, and two bacteria species. Similarly, R1 to R3, but not R4, cells produced ROS, while all subpopulations produced NO, in response to zymosan. Internalization of all phagocytic targets was blocked by PI 3-K inhibition. In addition, internalization of latex particles, but not of bacteria, was partially blocked by PKC or ERK inhibition. Interestingly, phagocytosis of zymosan was impaired by PKC, or ERK inhibitors, only in R2 cells. Zymosan-induced ROS production was blocked by PI 3-K inhibition, but not by PKC, or ERK inhibition. In addition, zymosan-stimulated NO production was affected by PI 3-K inhibition in R1 and R2, but not in R3 or R4 cells. NO production in all cell types was unaffected by PKC inhibition, but ERK inhibition blocked it in R2 cells. These data reveal the existence of profound functional and biochemical differences in mussel hemocytes and indicate that M. galloprovincialis hemocytes are specialized cells fulfilling specific tasks in the context of host defense.
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http://dx.doi.org/10.1016/j.dci.2007.10.004DOI Listing
August 2008