In vitro effects of the polyphenols resveratrol, mangiferin and (–)-epigallocatechin-3-gallate on the scuticociliate fish pathogen Philasterides dicentrarchi

Abstract

This study investigated the in vitro effects of the polyphenols resveratrol, mangiferin and (-)-epigallocatechin-3-gallate (EGCG) on the histiophagous ciliate Philasterides dicentrarchi, which causes fatal scuticociliatosis in farmed turbot Scophthalmus maximus L. Of the 3 polyphenols, resveratrol showed strongest antiprotozoal activity, reducing ciliate density after 1 wk culture by, on average, 91% at 50 µM, and 96% at 500 µM. EGCG reduced ciliate density by, on average, 93% at 500 µM, with no significant effect at 50 µM. Mangiferin reduced ciliate density by, on average, 56% at 500 µM, again with no significant effect at 50 µM. In view of these findings, we discuss the potential utility of chemotherapy with polyphenols as a strategy for the control of scuticociliatosis in farmed turbot. KEY WORDS: Polyphenols · Philasterides dicentrarchi · Resveratrol · Mangiferin · Epigallocatechin Resale or republication not permitted without written consent of the publisherDis Aquat Org 59: [171][172][173][174] 2004 toalexin (phytoestrogen) synthesized by Vitis vinifera in response to injury or fungal attack, and is present at high concentrations in grape skin and red wine. MATERIALS AND METHODSThe scuticociliate Philasterides dicentrarchi was obtained from ascitic fluid from the body cavity of naturally infected turbot Scophthalmus maximus, and maintained in complete L-15 medium (Leibovitz, Sigma) under the culture conditions previously described , which maintain the capacity of the parasite to infect turbot (Iglesias et al. 2002). The polyphenols trans-resveratrol (RESV), mangiferin (MA) and (-)-epigallocatechin-3-gallate (EGCG) (Fig. 1) were purchased from Sigma (Sigma Química) and maintained as 100 mM stock solutions in dimethyl sulfoxide (DMSO) at -20°C in the dark until use. To investigate antiprotozoal effects, the polyphenols (RESV, MA, or EGCG) in 1 ml of complete L-15 medium were added to the wells of 24-well sterile culture plates (Corning) containing 5 × 10 3 ciliates per well, to a final concentration of 50 or 500 µM, with subsequent incubation for 1 wk at 16°C. After this incubation period, the number of ciliates present was quantified in a 25 µl aliquot from each well: the ciliates were first inactivated with glutaraldehyde (final concentration 0.25%) for 15 min at room temperature, then counted in a hemocytometer (Iglesias et al. 2002). All experiments were done in quintuplicate. To rule out possible effects of the solvent DMSO, quintuplicate wells with L-15 medium containing the highest DMSO concentration used (0.5%) were also included. The results are expressed as means ± standard error (SEM), and statistical significances were assessed by 1-way ANOVA followed by TukeyKramer tests for multiple comparisons. RESULTS AND DISCUSSIONThe effects of RESV, MA, and EGCG (50 or 500 µM) on the Philasterides dicentrarchi trophozoite count in culture after 1 wk incubation are summarized in Fig. 2. As can be seen, the solvent DMSO had no significant effect; in previous studies we have found that DMSO at up...

Hydrogenosome Metabolism Is the Key Target for Antiparasitic Activity of Resveratrol against Trichomonas vaginalis

“…The effects of RESV and MDZ on the in vitro growth of T. vaginalis were determined as previously described, with minor modifications (18). For all experiments, stock solutions of RESV and MDZ were prepared in dimethyl sulfoxide (DMSO), to a concentration of 100 mM, and stored away from light at Ϫ80°C.…”

“…RESV, which is found naturally in grapes and red wine (14), acts as a precursor for stilbene compounds of higher fungotoxicity that accumulate in grapevine as a result of infection or stress (15). RESV has also been shown to be active against bacteria, fungi, protozoa, and viruses (16)(17)(18)(19)(20)(21)(22). In addition to its antimicrobial activity, RESV has also attracted attention on the basis of its health benefits to humans, which include anti-inflammatory effects; reduced risks of cardiovascular disease, cancer, obesity, diabetes, and neurodegenerative diseases; and also extension of life span by mimicking the caloric restriction effect (23).…”

Curcumin synergizes with resveratrol to stimulate the MAPK signaling pathway in human articular chondrocytes in vitro

“…Several lines of evidence indicate that Resveratrol is also capable of immunomodulatory, antioxidative, anti-apoptotic and anti-inflammatory functions [14,20,29,44]. Curcumin (diferuloylmethane), a dietary non-toxic pigment in curry, acts as a potent inhibitor of nuclear transcription factor jB (NF-jB)-activation in several cell types [10,47,53].…”

In vitro treatments for the theront stage of the ­ciliate protozoan Cryptocaryon irritans

“…Some studies have demonstrated the low toxicity of certain polyphenols in mice and humans (García et al 2003). It has therefore been hypothesized that these polyphenols would not be toxic to fish when used under in vivo conditions (Leiro et al 2004). Suzuki et al (2006) studied the efficacy and safety of GTE in salmonids under hatchery conditions, but the concentrations used were much lower than the efficient concentration tested in the present study against C. irritants.…”

“…EGCG comprises 14% of GTE, and it was shown that an application of pure EGCG was able to reduce I. necator infections (Suzuki et al 2006). Leiro et al (2004) showed that in vitro exposure to 500 µM of EGCG over 1 wk reduced the population density of the ciliate Philasterides dicentrarchi Dragesco et al, 1995 by 93%. In the present study, EGCG managed to kill all theronts when applied at 50, 250 or 500 mg l -1 over a period of 3 h, whereas its efficacy was considerably lower when application times were shorter and doses were lower (Table 1).…”