Journal of Oceanography and Marine Research
Open Access

Ecology of the Epiphytic Potentially Harmful Dinoflagellate Ostreopsis cf. ovata (Fukuyo) from Coastal Waters off Alexandria Egypt

Shimaa Hosny^* and Wagdy Labib ^*Correspondence: Shimaa Hosny, Taxonomy & Biodiversity of Aquatic Biota Lab, Marine Environment Division, National Institute of Oceanography and Fisheries, Alexandria, Egypt, Email:

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Introduction

Macroalgae provide a suitable substrate for microalgae found as epiphytes, which often provide support for higher trophic levels and might have important implications for food web dynamics [1,2]. Other ecological part played by epiphytes in ecosystem as a protection layer for seagrasses from excessive UV radiance was discussed [3]. However, as epiphyte load increases, it can negatively affect seagrasses in different ways [4].

Species of the dinoflagellate genus Ostreopsis Schmidt are unicellular algae preferentially epiphytic/epibenthic and grow in shallow waters of tropical and subtropical areas on abiotic and biotic substrates [5,6]. But cells may detach from the substrate and are found planktonic in the water column [6,7]. However, in the last few decades an increasing number of reports have been documented for a worldwide geographical expansion, and intensive bloom outbreaks of O. ovata and O. siamensis in warm and cold temperate areas [8,7]. Ostreopsis sp. has been expanding geographically in recent decades, especially in temperate regions, with 67% of studies focused on these regions [9]. There are currently eleven described species of Ostreopsis, four of which are known to be toxic; O. cf. ovata, O. siamensis, O. mascarenensis, and O. lenticularis to which scientists, the aquaculture industry, and government are keenly attentive [10,11]. The toxicity is associated with the presence of palytoxin and its derivatives [12].

Ostreopsis cf. ovata (Fukuyo) populations typically proliferate in rocky coasts, forming rusty-brown coloured mucilaginous film [6]. Widespread occurrence of the toxic benthic dinoflagellates genus Ostreopsis Schmidt has increased during the last decades [5]. It is recently found in the Mediterranean Sea where the first known bloom recorded in 1998 along the Tuscany coasts [13].

The presence of O. ovata in coastal waters may pose a threat to coastal food web and fishery [14]. In the Mediterranean Sea, neurotoxic effects due to toxin accumulation in food web have not yet been reported and Ostreopsis species are implicated thus far only in respiratory affections and skin or eyes irritations, causing health problems as dyspnea, fever, conjunctivitis and dermatitis for people near the shore exposed to marine aerosols during bloom events [15]. High levels of ovatoxins were recorded in natural samples; episodes of death of both benthic invertebrates (limpets, sea urchins and mussels) and macroalgae were commonly observed during O. ovata blooms.

A review of the existing literatures over the last three decades on harmful red tide blooms in Alexandria coastal waters shows a great attention towards planktonic microalgal bloom forming species while, there is a paucity of information in the literature on the ecology of attached potentially harmful epiphytic dinoflagellates in relation to biotic and abiotic regulating factors [7,16]. Ismael and Halim reported epiphytic Ostreopsis cf. ovata east of Alexandria [17]. Such studies are of increasing importance in the years ahead and that is what this study sets out to address. This in situ study investigates the interactions of habitat and ecosystem characteristics in the facilitation/regulation of the abundance of the epiphytic dinoflagellate O. ovata living attached to diverse macroalgal species in Alexandria waters, considering multiple environmental stressors assumed to regulate or influence its growth on biotic substrates. The study might provide additional information for better understanding the ecology of the Ostreopsis population dynamics in order to depict some considerations on its occurrence trend in this area and provide a basis for effective bloom management and mitigation. We quantified the abundances of O. ovata on different macroalgal communities dominated by specific macroalgal species, and measure multiple environment and weather factors to determine their effects on O. ovata abundances and endeavored to identify a correlation between O. ovata abundance and ambient abiotic factors.

Materials and Methods

Description of the study sites

The study site is located at the western head of Abu Qir Bay, east of Alexandria City. It is a relatively sheltered rocky shore, characterized by chains of natural exposed rocks extends for about 100 m seaward from the coast. Its massive rocky outcrops provide a suitable substratum for a rich algal flora [18]. This site was selected due to the presence of abundant and highly productive macroalgae community [7]. A transect stretched for about 25 m long was positioned perpendicular to the coast, among which two sites at 0.5 m and 1m depth were chosen. Sampling was carried out in May, July, and October of 2017 and February of 2018 (Figure 1).

Figure 1. Location of the studied site.

Sampling methods

A total of 26 samples of macroalgae including 14 macroalgal species of the different groups Rhodophyta (7 species), Phaeophyta (2 species), and Chlorophyta (5 species) were collected seasonally at a rocky substrate. Depending on their growth cycle and the environmental disturbance, some macroalgae species were missing, particularly during rough sea periods in winter, and therefore could not be collected. Thus, the collected macroalgae here represents the whole algal species present during the different months. The handpicked macroalgae of ca. 20- 100 g wet weight collected carefully was placed into individual plastic bags with surrounding seawater, and transported on ice cooler back to the laboratory. In laboratory, each macroalgae was put in a plastic container with 250 mL of fresh filtered seawater, vigorously shaken to dislodge epiphytic cells, and the material passed through several meshed filter (500, 100 and 20 μm mesh) to separate macroalgae and water containing microalgae [19]. The material retained by the filter was suspended in sterile filtered seawater (25 mL) and fixed with the addition of Lugol's acid solution at 1%. For qualitative and quantitative analyses, 2 mL of preserved sample from determined wet weight of each macroalgal sample was counted triplicate using counting chamber. Cell abundances of epiphytic cells expressed here as cells g^-1 wet weight of macroalgae, following the methodology described by Okolodkov [20]. Identification and nomenclature of microorganisms followed appropriate keys while macroalgae following standard taxonomic keys as described by Aleem et al. [21-24].

Measurement of environmental factors

Water temperature (°C), salinity, dissolved oxygen (mg l-1 and saturation %) and pH values determined using the (hydro lab. HANA, Model HI 9828). Samples intended for nutrient analysis were kept in the dark at 4°C. Concentrations of nitrite, nitrate, ammonia, phosphate and silicate were measured according to the methodology of Strickland and Parsons [25].

Statistical analyses

The Pearson's correlation coefficient “r” and multiregression analyses (STATISTICA 7.0, Stat Soft, Inc.) were applied on the whole data set in order to highlight eventual relationships between Ostreopsis abundance and environmental parameters, as well as between the parameters themselves across the two sites.

Results

Environmental characteristics of sites

The sampled sites were natural rocky areas and were highly exposed to hydrodynamic conditions. Each measured environmental parameter showed approximately the same range of values at sampled sites. The physical and chemical parameters of water from locations collecting samples are given in Table 1.

Table 1: Temporal distribution of physical and chemical parameters.

The total temperature range encountered varied within 10.74°C between the lowest in February and the highest in July, reflecting a clear seasonality. Salinity range varied within 1.77, with no particular seasonal variation. The hydrogen ion concentration values lie in the alkaline side and with a very narrow range of fluctuation (0.54). Dissolved oxygen in May and July increased by 3.5-3.9 times greater compared with the minimum in October. Concerning nutrients, nitrite followed typical summer minimum, and it was highest in May. Nitrate concentrations were always high, and expressed 63.21-78.91% of the total nitrogen. Ammonia concentrations were often <3 μM, and reached minimum in October. Silicate values were the lowest in May, and were at a maximum of 5.64 μM in July. Phosphate trend followed that of silicate.

Macroalgae species

The encountered macroalgae species in the present study (14 species) are given in Table 2 Clear seasonality is evident, and Ulva compressa represents a perennial form.

Table 2: Monthly occurrence of the host macroalgae.

Epiphytic species

Generally, the phytoplankton epiphytic species other than O. cf. ovata found during the study comprised mainly diatom species of the genera Achnanthes (Agardh), Bacillaria (Gemlin), Climacosphenia (Ehrenberg), Cocconies (Ehrenberg), Cyclotella (Chang), Licmophora (Agardh), Melosira (Pritchard), Navicula (Cleve & Grunow), Odentella (Agardh), Pleurosigma (Smith), Pseudonitzschia (Cleve), and Synedra (Nitzsch), and few rather dinoflagellates mostly belong to the genera Prorocentrum (Ehrenberg), Protoperidinum (Balech), Pyrophacus (Stein), and Scrippsiella (Loeblich III). The abundance of epiphytic cells is given in Table 3.

Table 3: Abundance of the Ostreopsis cf. ovata (average cells g^-1 wet weight x 10³) on the different macroalgae groups.

Temporal trend of Ostreopsis

The surface distribution of O. ovata reflects its nature an epiphytic form that attached on the thalli of the whole tested macroalgal groups with different frequencies, and during the different seasons. Based on average values (Table 3), and neglecting multiple factors as the seasonal variation of both macroalgal and microalgal species diversity and community structure, the magnitude of the abundance is arranged as Chlorophyta>Rhodophyta> Phaeophyta. The results highlighted the highest abundance observed between spring and autumn culminated the peak on chlorophycean species in July, while, it was sharply declined in winter when the abundance was at minimum in February.

The diatom Navicula sp. during February was the major contributor to total epiphytic diatom density (35% of the total diatom abundance). Meanwhile, no epiphytic O. ovata were found on Phaeophyta sp. in October. This species contributes significantly to the total epiphytic counts at the two sampling sites with higher abundances at the relatively far station comparing with each other’s, following the same seasonal trend.

Abundance on macroalgae species

The field work involved the manipulation of naturally occurring macroalgal community omplexities in shallow rocky shore. The results show noticeable differences in the abundance of O. ovata depending on macroalgal host species, without statistically highlighting a clear substrate preference. The Chlorophytes U. linza, U. compressa and U. fasciata were particularly able to support very high abundances of Ostreopsis during May and July, and it was relatively less on U. lactuca in October. The maximum abundance on U. linza during May attained almost 1900 times the values estimated during February on the same species. The results declared Chlorophytes might be the optimal macroalgae for epiphytic Ostreopsis. As for the Rhodophyta, the maximum abundance was found on C. officianalis in July and October. The occurrence of the Phaeophyta species was restricted to May and July, and hence, abundance on the species was very low, among them, P. pavonia sustained the highest abundances.

In May (Table 4), the highest abundance of Ostreopsis appeared attached mainly to the Chlorophyta species in order of magnitude U. linza>U. fasciata>U. compressa, representing 42.53-99.44% of the total epiphytes. Much lower densities found on Rhodophyta species, but, mainly were restricted to the thalli of C. officianalis, contributing >94% of the total. A few thousand cells were gathered on the surface of L. papillosa, higher than on P. capillacea. The lowest abundance appeared on the Phaeophyta C. spinosa.

Table 4: Abundance (cells g^-1 wet weight x10³ (of Ostreopsis cf. ovata cells on macroalgae species during May.

The macroalgal community changed in July compared with May as C. pelluceda replaced U. linza, Rhodophyta species were well diversified (4 spp.), and species belong to Phaeophyta were not recorded. The diversified algal community might contribute to the abundance of O. ovata observed all over the year (Table 5).

Table 5: Abundance (cells g ^-1 wet weight x10³ (of Ostreopsis cf. ovata cells on macroalgae species during July.

The Chlorophytes were still the hosts having the highest abundances of Ostreopsis in the order of magnitude U. compressa> U. fasciata> C. pelluceda. Among the Rhodophytes, J. rubens seems the preferable host, where Ostreopsis contributed 77.86-91.04% of the total epiphytes. Again, the lowest abundance detected on P. capillacea.

Chlorophyta in October comprised U. compressa and U. lactuca, but, with almost similar Ostreopsis abundances. Among the four recorded Rhodophytes, the highest abundance, in May was found on C. officianalis, followed by C. elongata as the second important host, while, the lowest density observed on C. mediterranean (Table 6).

Table 6: Abundance (cells g^-1 wet weight x10³ (of Ostreopsis cf ovata cells on macroalgae species during October.

Despite the well diversified macroalgal community in February (8 species), the epiphyte density dropped to minimum compared with other months. Ostreopsis abundance did not exceed 12, 80 and 35 cell g^-1 wet weight on the Chlorophyta, Rhodophyta and Phaeophyta species. The highest Ostreopsis abundance was restricted to the Rhodophytes C. elongata and P. capillacea. Moreover, its contribution to the total epiphytes was low, almost nil on P. capillacea and P. pavonia. The thalli of Phaeophyta P. pavonia and C. spinosa sustained the lowest abundance.

Interactions of the abundance of Ostreopsis cf. ovata with measured physical and chemical environmental parameters

The results of the statistical analyses (Tables 7-9) revealed water temperature the major environmental contributory affecting the occurrence O. ovata on examined macroalgae. The nutrients, NO₃ and PO₄ contribute significantly to the abundance of Ostreopsis on Chlorophyta spp., while, NO₂, SiO₄ beside PO₄ link significantly with its abundance on Rhodophyta. Salinity variations exhibit a weak, insignificant influence on the abundance attached to Chlorophyta spp., and it is almost of a negligible effect for Rhodophyta spp. A linear insignificant negative correlation was found with the pH variations. The highly significant correlation between salinity and nutrient concentrations indicate discharged fresh water the main source of nutrients supply.

Table 7: Abundance (cell g^-1 wet weight x10³(of Ostreopsis cf ovata cells on macroalgae species during February.

Table 8: Pearson’s correlation values for Ostreopsis cf. ovata on Chlorophyta species.

Table 9: Pearson’s correlation values for Ostreopsis cf. ovata on Rhodophyta species.

The multiple regression equation for O. ovata (Cell g^-1 wet weight) on Chlorophyta is expressed as:

= -147.49 - 4.87* °C - 28.92* NO₃ + 20.22*PO₄….. R₂ =0.88, n=27

explain the significant importance of these parameters in combination, on Rhodophyta as:

= -20.44+2.09* °C - 6.47*NO₂ - 2.10* SiO₄ + 2.2 *PO₄… R₂ =0.36, n=39

explain about 36% of the O. ovata variations.

The relation between Ostreopsis abundance on the green algae U. compressa and measured environmental factors was also done. This macroalgal species was chosen being a perennial form. The results Table 10 support others previously mentioned factors for all Chlorophyta spp., but, salinity negatively correlated with its abundance and PO₄ concentration seem more effective.

Table 10: Pearson’s correlation values for Ostreopsis cf. ovata on Ulva compressa.

The Pearson linear correlation between the epiphytic O. ovata concentrations was significant (p<0.01, N = 55, rs = 0.71), with an explained percentage of variation of R₂ = 0.50.

Discussion

The present study represents the first specific contribution on the epiphytic O. ovata in the Egyptian Mediterranean waters. Previously, this species was recorded planktonic form the investigated area, with a maximum abundance over 1980 cell l^-1 [7]. The study declared O. cf. ovata is not an obligate epiphytic species; since it is able to colonize the macroalgae thalli of 14 species of different groups include 5 Chlorophyta, 7 Rhodophyta and 2 Phaeophyta. A reasonable body of ecological studies has been conducted on the potential role of environmental on the dynamic occurrence of Ostreopsis. However, in many cases, contrasting results are found for the same taxon from different areas and the role of ecological habitats in the facilitation of attachment has rarely been considered [7,11]. Most of the research focused on the effects of seawater temperature, nutrients, and salinity on Ostreopsis sp. populations.

The statistical analyses illustrate how environmental factors in the sampling sites might influence epiphytic Ostreopsis abundance, particularly water temperature and the dissolved nitrate and phosphate concentrations; temperature was the major factor. However, the impact of temperature alone during the summer period, which was represented here by July, seems less evident may be due to strong interactions with other unmeasured physical parameters as light intensity and photoperiod wave activity and some biotic factors that could also be involved such as the availability of some seasonal macroalgal substrates and the competition with other microphytes [20,26]. Also allelopathy has been often suggested among the biotic factors crucial to Ostreopsis [27]. A concise summary of the effects of surface seawater temperature, salinity and nutrient concentrations on Ostreopsis sp. populations in temperate regions can be found in Accoroni and Totti’s review [11].

The maximum abundance of Ostreopsis in July took place at the highest water temperature (28.37°C), and a relative decrease of abundances observed in May and October at temperature between 22.34°C and 25.1°C, reaching its minimum in February associated with the lowest temperature all over the year (17.63°C). A bloom event reported with temperature not exceeding 22.5°C while, the results of Cohu et al. highlighted substantial seasonal variations in the abundance of Ostreopsis within an optimum growth temperature ranging from 23°C to 27.5°C. Moreover, temperatures from 22 to 30°C seem to favor Ostreopsis sp. blooms in temperate zones areas the Mediterranean Sea and the New Zealand coasts but the link between temperature and Ostreopsis growth is still not so clear and the relationship with the seawater temperature is not the same in all the geographic areas [28-30]. Also, Ostreopsis strains sampled from the tropics have exhibited growth optima in the range 25-30°C [26]. Until now, O. ovata and O. siamensis have been detected at the minimal temperatures of 13°C- 14.5°C in the Mediterranean Sea, yet were also recorded when water temperature dropped below 8°C [31,32]. Such results reveal a high tolerance to extreme climate changes [33]. Therefore, Yamaguchi et al. based on field and experimental studies demonstrated a wide tolerance ability of O. ovata to a wide temperature ranges, offering some reasons to explain its recent geographical expansion and occurrence in temperate areas [34]. These conflicting results may likely reflect geographical differences [8]. In the Mediterranean Sea, the seasonal trend of Ostreopsis sp. clearly follows temperature seasonality [31]. The observed marked heterogeneity in the abundance of O. ovata among the examined macroalgae of different groups, beside other substrates could partially explain its spatial variability [35].

Other ecological parameters of weak contribution to the variation of O. ovata were salinity and pH, in particular on Chlorophyta. The maximum abundance of this species occurred at 35.43 in July, while, at the same salinity level (35.8) in February, the abundance was at minimum. The results thus explain the insignificant correlation of salinity with abundance of Ostreopsis, and its negligible effect on Rhodophyta sp. The intermediate salinity levels, a sign of incoming discharged waters might have positive impact on the epiphytic Ostreopsis to proliferate. According to Delgado et al. freshwater source should have a negative impact on O. ovata growth via a decrease of salinity, as observed on O. lenticularis and others dinoflagellates [36]. Morton et al. reported that O. siamensis and O. heptagona displayed maximal growth at a salinity value of 33 [26]. In contrast, O. ovata isolated from Hawaii were found to be negatively correlated with salinity in agreement with Carlson and Tindall [37,38].

The role of depth was assessed at two target sites where samples were collected at 0.5 m and 1 m depth. Despite the very narrow distance between the two sites, the abundance of Ostreopsis was always higher at the relatively far site; more strong hydrodynamic action at St. II may be involved affecting abundance [6]. The abundance at St. II compared with St. I contributed 71.43-82.91% for Chlorophyta, 58.78-83.19% for Rhodophyta and 61.44-99.03% for Phaeophyta. While several authors found a significant negative relation of Ostreopsis with depth, other denied such conclusion [39,40]. However, one study contradicts these conclusions indicating that Ostreopsis sp. is better adapted to ‘slightly shaken’ environments [41]. The oxygen saturation decrease in summer was related to the increase in seawater temperature. It is might also be linked to the higher consumption of oxygen related to organic matter decomposition during O. ovata main occurrence; the negative correlation between these two factors might explain such observation.

The results showed the measured nutrient concentrations values falling within the expected range for a shallow area subjected to a moderate/temporally anthropic impact [7]. The high NO₃ concentrations with the main Ostreopsis occurrence periods, and that of PO₄ in July as well certainly indicate arrival of nutrients supply through eutrophication processes. Since the study area is a summer resort that starts annually from May, the increased population density might represent a crucial anthropogenic reason to promote the occurrence of Ostreopsis in such urbanized area, similar to other data [42]. The statistical analyses declared highly significant correlation between salinity and nutrient concentration indicates same source of nutrient supply. The analyses showed the positive significant correlation between NO₃, PO₄ and abundance of O. ovata on Chlorophytes, and the same for NO₂ and PO₄ for Rhodophytes, and also with a significant positive correlation with SiO₄, in agreement with Parsons and Preskitt [37]. Such high nutrient concentrations provide an indication of eutrophication that might play a significant part to control the epiphytic Ostreopsis abundance. Subsequently, there are some conflicting results regarding the relationships of nutrients with Ostreopsis proliferation trends in field observations of tropical and temperate areas [39]. However, often, and particularly in coastal areas, eutrophication appears to be directly linked to the occurrence of harmful algal blooms and worldwide Ostreopsis sp. appears to proliferate both in eutrophicated and oligotrophic areas [43-45]. Meanwhile, anthropogenic nutrient enrichment might have an indirect impact on Ostreopsis sp. abundance by inducing a shift in macroalgae coverage frequency and/or composition, which lead to an additional difficulty in ecological studies [46]. Silicate seems to match with the abundance of O. ovata on Rhodophyta species; it is unexpected for dinoflagellates which do not specifically use silicate for their growth, in contrast to diatoms.

Another environmental factor seems to affect the abundance of O. ovata was the N:P ratios. The N:P ratios calculated based on the Redfield model [47]. Working on this principle, high N:P ratios during May and February may be taken as indicative of low P availability, whereas low values (<16) during July and October might indicate Nlimitation [48]. The N:P ratios matched in strong negatively correlation with the abundance of Ostreopsis on Chlorophyta sp. (r=0.38 and 0.25 at St. I and II, respectively, n=27, p at 0.5), and corresponded in high significance with its abundance on Rhodophytes (r=0.54 and 0.55). The indirect effect of N:P ratio through changed macroalgal composition was discussed [49,50].

The results declared O. ovata found on all the examined macroalgae, exhibits the different host preferences; Ulva species were the optimal macroalgae for the epiphytic O. ovata. However, factors as algae structure and surface area, regional occurrence and coverage percentage, and presence of allelopathic compounds may determine the host preference of the species [7]. According to the field observations, Ulva sp. predominated all the investigated period, with different frequency degrees. As far as macroalgae are concerned, Cohu et al. [42] were unsuccessful in establishing significant preferences, while, Accoroni et al. [11] noted Ostreopsis sp. to display algal host preference, and certain macoalgae species have the ability of support very high abundance of O. ovata offering evidences that algal host preferences is species-specific [8]. The present study documented the significant importance of Ulva sp. as the suitable host of O. ovata in Alexandria waters, in contrasting with others in the Mediterranean Sea that show O. ovata most often found on Phaeophyta and Rhodophyta sp. [31].

Conclusion

The potential toxic O. ovata represents another additional threat to economy and public health in Alexandria. To our knowledge, this is its first study as epiphytic form in SE Mediterranean Sea taking into account diverse ecological and biological factors. Temperature, PO₄ and NO₃ seem major contributors influencing its abundance. Ostreopsis ovata had the ability to attach to a diversified macroalgae species, with preference to Chlorophyta. Since O. cf. ovate cells are loosely attach to the substrata and can be easily removed and re-suspended in the water column, future study of wave action in Alexandria waters presents untapped field of research with great potential.

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