Papel de las actividades superóxido dismutasa y catalasa en la ...

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RESULTS AND DISCUSSION reason, once aqueous and ethanolic extracts were assayed in vitro, oral administration of whole alga cells was performed. In fish, as in other aquatic organisms, administration of whole microorganisms has focused mainly on bacterial species, such as probiotics (Verschuere et al., 2000; Nikoskelainen et al., 2001; Irianto and Austin, 2003; Salinas et al., 2005; Balcázar et al., 2006; Díaz-Rosales et al., 2006; Salinas et al., 2006), but studies about whole algae are still very scarce. On the other hand, P. cruentum acumulates large amounts of lipids, specially arachidonic acid and noticeable amounts of eicosapentaenoic acid (Kinsella et al., 1990; Koven et al., 2001); carbohydrates (Fujiki et al., 1992; Santarém et al., 1997; Bagni et al., 2000; Morris et al., 2000; Esteban et al., 2001; Jeney and Jeney, 2002; Cook et al., 2003; Couso et al., 2003; Castro et al., 2004; Bagni et al., 2005; Kumar et al., 2005); carotenes (Tachinaba et al., 1997; Amar et al., 2004); vitamins (Hardie et al., 1990, 1991; Ortuño et al., 1999; Jeney and Jeney, 2002). Due to the fact that this alga contains different immunostimulant substances, its use could generate a more general immune response as has been proposed for other microorganisms such as yeasts (Ortuño et al., 2002; Rodríguez et al., 2003). Thus, three groups of sole specimens received daily for four weeks one of the different diets assayed on a daily basis: commercial diet supplemented with lyophilized alga, diet consisting on non-supplemented commercial diet (control group) or diet composed of commercial diet containing immunostimulant, Sanostim. Besides, to evaluate the possible synergic effect of immunostimulant with a vaccine, two weeks after the beginning of the feeding trial, a group of fish per treatment were intraperitoneally inoculated with a bacterin of P. damselae subsp. piscicida, a formalinkilled aqueous vaccine. Results obtained show that after four weeks of algal administration, the superoxide anion production increases in immunized fish. This increase is statistically significant, not only compared to fish fed with normal diet, but also compared to fish fed with commercial immunostimulant. The combined action of immunostimulant and vaccine has been described by numerous authors, who concluded that combination of vaccination and immunostimulant administration increases vaccine potency (Jeney and Anderson, 1993; RØsrstad et al., 1993; Aakre et al., 1994; Sakai et al., 1995; Baulny et al., 1996; Sakai, 1999). 102
RESULTS AND DISCUSSION Once immunostimulant effect of algal cells was demonstrated, the potential effect of the extracellular polysaccharidic fraction of P. cruentum on the respiratory burst activity of sole phagocytes was evaluated (article 2.2., Article section). Results obtained indicate that in vitro none of the assayed concentrations of the extracellular polysaccharide from P. cruentum stimulate the respiratory burst of sole phagocytes, after 30 min contact with fish cells. These results are not in agreement with the data obtained by Castro et al. (2004, 2006), who suggest that stimulation of respiratory burst activity in turbot (Psetta maxima) phagocytes incubated with algal extracts, is due to algal polysaccharides. On the other hand, according to Castro et al. (2004) the modulatory ability of the respiratory burst activity of fish phagocytes varies greatly among algal species. Therefore, the non stimulation of respiratory burst in vitro by P. cruentum polysaccharides could be due to the absence of immunostimulant activity in this fraction, presence in low concentration or short incubation time with phagocytes. Whereas, the positive control, a commercial β-glucan, induced an increase in the respiratory burst, activity when applied at 10 mg ml -1 . Additionally, fish were intraperitoneally inoculated with 500 µg of extracellular polysaccharidic fraction. Later, fish were immunized with a bacterin, composed by P. damselae subsp. piscicida formol-inactivated cells. Sampling time was carried out at 24 h and seven days post-vaccination. Results obtained indicate that the concentration and time assayed, 1 and 7 days after polysaccharidic fraction inoculation, do not produce an increase in the respiratory burst activity of sole phagocytes, not even immunized fish. Moreover, 24 h post-inoculation, the respiratory burst decreases in phagocytes from fish inoculated with polysaccharidic fraction or with the bacterin. This decrease may be due to an immunosupression by stress after handling (Thompson et al., 1993; Pulsford et al., 1995) as it is not observed after 7 days of the inoculation. To sum up, the polysaccharidic fraction of P. cruentum, in assayed conditions, does not enhance the respiratory burst activity in sole phagocytes. These results suggest that stimulation observed after oral administration of alga cells would be due to other compounds with immunostimulant properties. However, we cannot rule out the possibility that the polysaccharide stimulate another immunological parameter. Thus, a 103
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FACULTAD DE CIENCIAS DEPARTAMENTO D
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FACULTAD DE CIENCIAS DEPARTAMENTO D
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Los ensayos que constituyen esta Te
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- Díaz-Rosales, P, Arijo, S, Chabr
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Sabe esperar, aguarda que la marea
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ÍNDICE / INDEX Papel de las activi
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ÍNDICE / INDEX 2. Photobacterium d
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RESUMEN Photobacterium damselae sub
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INTRODUCCIÓN 1. LA ACUICULTURA. EL
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INTRODUCCIÓN saxatilis) (Hawke et
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INTRODUCCIÓN 2.2. SINTOMATOLOGÍA
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INTRODUCCIÓN las epidemias surgida
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INTRODUCCIÓN et al., 1997). En P.
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INTRODUCCIÓN 3.1. ESTALLIDO RESPIR
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INTRODUCCIÓN radicales generados e
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INTRODUCCIÓN Tabla 2 Ejemplos de m
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INTRODUCCIÓN de parasitismo en el
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INTRODUCCIÓN razón de la búsqued
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INTRODUCCIÓN araquidónico, están
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INTRODUCCIÓN simplificar la admini
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INTRODUCCIÓN sino también cuando
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INTRODUCCIÓN La necesidad de mejor
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OBJETIVOS El trabajo planteado en e
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MATERIA L Y MÉTODOS La metodologí
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RESULTADO S Y DISCUSIÓN El primer
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RESULTADO S Y DISCUSIÓN 1996; Lync
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RESULTADO S Y DISCUSIÓN Una vez de
Page 73 and 74: RESULTADO S Y DISCUSIÓN inmunoesti
Page 75 and 76: RESULTADO S Y DISCUSIÓN cruentum s
Page 77 and 78: RESULTADO S Y DISCUSIÓN permiten c
Page 79: RESULTADO S Y DISCUSIÓN harían fa
Page 83 and 84: CONCLUSIONES Tras los estudios real
Page 85: FACULTAD DE CIENCIAS DEPARTAMENTO D
Page 89: ABSTRACT Photobacterium damselae su
Page 93 and 94: INTRODUCTION 1. AQUACULTURE. THE CU
Page 95 and 96: INTRODUCTION With regard to Solea s
Page 97 and 98: INTRODUCTION 3.1. RESPIRATORY BURST
Page 99 and 100: INTRODUCTION Some catalases have al
Page 101 and 102: INTRODUCTION pathogens, study of th
Page 103 and 104: INTRODUCTION increase their bacteri
Page 105 and 106: INTRODUCTION 4.3.1. Porphyridium cr
Page 107 and 108: INTRODUCTION Different mechanisms h
Page 109: A I M S
Page 113: M A T E R I A L S A N D M E T H O D
Page 117: R E S U L T S A N D D I S C U S S I
Page 120 and 121: RESULTS AND DISCUSSION resist 100 m
Page 122 and 123: RESULTS AND DISCUSSION present work
Page 126 and 127: RESULTS AND DISCUSSION great number
Page 128 and 129: RESULTS AND DISCUSSION dominant spe
Page 131 and 132: CONCLUSIONS Studies carried out on
Page 133: R R E F E R E N C I A S E F E R E N
Page 136 and 137: REFERENCIAS / REFEREN CES Aoki, T,
Page 138 and 139: REFERENCIAS / REFEREN CES Bell, MV,
Page 140 and 141: REFERENCIAS / REFEREN CES Dalmo, RA
Page 142 and 143: REFERENCIAS / REFEREN CES Hamaguchi
Page 144 and 145: REFERENCIAS / REFEREN CES Kono, Y &
Page 146 and 147: REFERENCIAS / REFEREN CES Magariño
Page 148 and 149: REFERENCIAS / REFEREN CES Ortuño,
Page 150 and 151: REFERENCIAS / REFEREN CES Salinas,
Page 152 and 153: REFERENCIAS / REFEREN CES Thompson,
Page 155: S E C C I Ó N D E A R T Í C U L O
Page 159: A RTÍCULO 1.1. A RTICLE 1.1.
Page 162 and 163: Journal of Fish Diseases 2003, 26,
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Journal of Fish Diseases 2006, 29,
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A RTÍCULO 2.1. A RTICLE 2.1.
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Abstract The stimulatory effect of
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alga Porphyridium cruentum. For thi
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Feeding assays were carried out wit
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(10 8 bacterias ml -1 ) and used to
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are in agreement with the data repo
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unvaccinated fish. However, the ser
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immune response of gilthead seabrea
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[27] Duncan, PL and Klesius, PH. Ef
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(Macrogard) and alginic acid (Ergos
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[66] Salinas, I, Díaz-Rosales, P,
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Figure 1a 3 2.5 Stimulation index 2
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Figure 1c Stimulation index 3 2.5 2
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Figure 2b Stimulation index 3 2.5 2
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Effect of the extracellular polysac
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1. Introduction Solea senegalensis
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On the other hand, β-1,3-glucan fr
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eaction was tested by adding supero
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Intraperitoneal inoculation of the
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defense mechanisms and protective i
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phagocytes from sole (Solea senegal
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Figure 1a Stimulation index 3 2.5 2
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Figure 2 Stimulation index 1.4 1.2
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Effect of dietary administration of
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1. Introduction Senegalese sole cul
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2.2. Fish and experimental design S
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2.5. Challenge with Photobacterium
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the densitometric curves of the pro
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most efficient probiotics for aquac
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acterial diversity only the dominan
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[9] Verschuere, L, Rombaut, G, Sorg
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[28] Burr, G, Gathin, D and Ricke,
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WB, editors. Techniques in Fish Imm
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and host-specific communities of ac
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Table 1 Primer Sequence (5’-3’)
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Figure 1b Stimulation index 2.5 2 1
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Table 2 Primer Control Pdp11 Pdp13
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Figure 3 Pearson correlation [0.0%-
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que perdieras parte de tu tiempo en
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