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Allelopathic effects in dinoflagellates A case study of the benthic genus Ostreopsis and two non-
toxic species from the genera Prorocentrum and Coolia *G. Portela M.1, Riobó P.2, Franco J.M.2, Rodríguez F.1
1 Instituto Español de Oceanografia (IEO), Centro Oceanográfico de Vigo (UA Microalgas Nocivas CSIC-IEO), Subida a Radio Faro 50-52, Cabo Estay, Canido, 36390 Vigo, Spain. *email: [email protected] 2 Instituto de Investigaciones Marinas, CSIC (UA Microalgas Nocivas CSIC-IEO), Eduardo Cabello, 6, 36208 Vigo, Spain.
4. CONCLUSION
•“Cells experiment” showed that non-toxic strains grew normally in contrast with Ostreopsis spp., suggesting a competitive advantage through nutrients, space competition and/or some allelopathic effect.
•VGO 1001 possessed higher OVTX amounts, C. monotis only seems to be affected by VGO 1000 supernatant by means of reducing movement capacity. It would be advisable to study if any morphological or physiological differences are also observed when exposed to supernatant.
•Different toxins profile found in Ostreopsis could be explained by 2genetic differences between these strains.
•Further studies will be necessary to clarify if there is an allelopathic interaction that could be related with toxins amount or profile in Ostreopsis.
0
0,05
0,1
0,15
0,2
0,25
0,3
0,35
0,4
0,45
0,5
r
Coolia monotis growth rate from day 0 to day 6
Control
Supern_VGO1000_500
Supern_VGO1000_1000
Supern_VGO1001_500
Supern_VGO1001_1000
vs. VGO1000_500
vs. VGO1000_1000
vs. VGO1001_500
vs. VGO1001_1000
1. AIM
The epibenthic dinoflagellate Ostreopsis is a potentially toxic bloom-forming organism. Ostreopsis spp. have the ability to produce palytoxin-like compounds which have demonstrated to be harmful for both human health and 4benthic invertebrates (sea urchins). This genus often co-occurs with other dinoflagellates (i.e. Coolia, Prorocentrum) but little is known about the potential interactions between these dinoflagellates in the benthic environment. The aim of the present study was to explore the potential inhibitory effects on growth and/or cell behavior by toxic strains of Ostreopsis cf. ovata (VGO 1001) and Ostreopsis sp. (Lanzarote type VGO 1000), on two non-toxin producer dinoflagellates (Coolia monotis and Prorocentrum elegans) isolated from the same geographical area (Canary Islands, NE Atlantic).
2. METHODS
•Growth and culture conditions: 25ºC, 277.41 µE/m2s-1, 12:12 L:D cycle, 150 mL glass flasks, L1 medium. 1000 cells/mL (non-toxic strains), 1000 and 500 cells/mL (both Ostreopsis). •Culture samples: fixed with Lugol and counted using Sedgewick rafter chamber. •Growth rates: 3instantaneous rate of increase: r. •Toxins extraction: glass microfiber filters MFV5 (47mm), extracted in 2 mL of MeOH, homogenized and vortexed. •Control cultures experiment: separated species. •Mixed cells cultures experiment: different Ostreopsis concentrations + non-toxic species. •Supernatant cultures experiment: clarified Ostreopsis medium added to the non-toxic species. •Toxin analyses by liquid chromatography – High resolution mass spectrometry1. Toxins identification is function of the characteristic fragment.
3. RESULTS
3.1 ”Cells experiment” •Ostreopsis sp. (VGO 1000) evidenced reduced growth rate in mixed cultures relative to the controls (Fig. 1B). This effect was not apparent in O. cf. ovata (VGO 1001) (Fig. 1D). However, both Ostreopsis strains declined fast their numbers and reached a senescent phase whereas controls remained in stationary phase. •Growth of C. monotis and P. elegans did not seem to be affected when co-cultured with Ostreopsis cells and P. elegans growth rates were slightly higher compared with the control.
3.2 “Supernatant experiment” •Supernatant apparently affects C. monotis population from day 0 to 4 by delayin its growth (Fig. 2C). Later on, Coolia population recovered and grew similarly to control cultures (Fig. 2B). •Reduced mobility was observed in C. monotis with VGO 1000 supernatant, as evidenced in video records obtained with a camera. Such effect was not evidenced with VGO 1001 treatment. •No effects were observed in P. elegans even though a small population increase was observed compared with the control.
3.3 Toxins profile
•OVTX-a+b: major component of O. cf. ovata (VGO 1001) in control initial cultures (88.7%) (Fig. 3A) and those at the end of the experiment (90.6-96.7%). •OVTX-c: higher at O. cf. ovata (VGO 1001) in the “cells experiment” with Coolia (96.8%) and Prorocentrum(91.1 %) (Figs. 3C, 3D). •OVTX-a: major component of Ostreopsis sp. (Lanzarote type VGO 1000) in control initial cultures (Fig. 3B), while trace putative PLTX (0.53 and 0.19 pg/cell) was found in mixed cultures with P. elegans.
REFERENCES 1 Ciminiello, P., Dell’Aversano, C., Dello Iacovo, E., Fattorusso, E., Forino, M., Grauso, L. et al. 2010. Complex palytoxin-like profile of Ostreopsis ovata. Identification of four new ovatoxins by high-resolution liquid chromatography/mass spectrometry. Rapid Commun Mass Spectrum 24: 2735-2744. DOI: 10.1002/rcm.4696. 2 Ciminiello, P., et al., Investigation of toxin profile of Mediterranean and Atlantic strains of Ostreopsis cf. siamensis (Dinophyceae) by liquid chromatography–high resolution mass spectrometry. Harmful Algae (2013), http://dx.doi.org/ 10.1016/j.hal.2012.12.002 3 Gotelli, N. J. 1995. A Primer of Ecology. Sinauer, Sunderland, Massachusetts, 206 pp. 4 Tuya, F., Martin, J. A., Reuss, G.M. & Luque, A., 2001. Feeding preferences of sea urchin Diadema antillarum in Gran Canaria Island (Central-East Atlantic Ocean). JMBA, 81, 1-5.
ACKNOWLEDGEMENTS
We thank to Patricio Díaz Gómez for R plotting help. This experiment was carried out at Instituto Español de Oceanografía (IEO) in Vigo.
Fig. 1. A) O. sp. (Lanzarote type VGO 1000) culture B) growth curves and
C) O. cf. ovata (VGO 1001) culture D) growth curves in control,
supernatant and cells experiments.
Fig. 2. A) Coolia monotis culture B) growth curves and C) growth rates
in control (black), supernatant (red) and cells (green) experiments.
B)
Fig. 3. Total ion chromatogram (TIC)
of A) VGO 1001 and B) VGO 1001 in
control cultures (exponential phase)
and VGO 1001 in cells experiment
mixed with C) P. elegans and D) C.
monotis in cells experiments at
senescent phase. E) P. elegans
culture.
PLTX 327.1919
OVTX-a 327.1919
OVTX-a+b 371.2181
OVTX-c 371.2179
D)
B)
C)
A) B)
C)
D)
A)
A)
C)
E)
