Background

Many phytoplankton species cause blooms and red tides in the aquatic ecosystems. Marine coastal areas are important ecosystems and are source for fishing, aquaculture and tourism. However, they are subject of pollution, eutrophication and harmful algal blooms which cause a widespread problem (Smayda, 1997).

Through the eutrophication period (1970-1995) of the Black Sea ecosystem in the water in front of the Bulgarian coast were observed numerous cases of intense and prolonged phytoplankton blooms leading to reduced biodiversity and destruction of bottom biocenoses (Kamburska et al., 2003; Moncheva and Doncheva, 1998; Moncheva et al., 1995; Nesterova, 1987; Petrova - Karadjova, 1984; Shiganova et al., 2008; Velikova et al., 1999; Zaitsev and Mamaev, 1997;). Dozens of phytoplankton species were recorded into a state of blooms.

Reduction of phytoplankton development in the Bulgarian marine aquatories has started after 1995 (Bodeanu et al., 1998; Velikova et al., 2001). Specifically in Varna Bay, cape Galata and cape Emine reduction of phytoplankton quantitative development has been registered after 1998 (so called post-eutrophication period) (Gerdzhikov and Petrova, 2008).

Reduction in the number of blooming phytoplankton species, e.g., in Varna Bay for the period of the 90’s a total of 37 "blooming" species were observed (Velikova and Petrova, 1999), and in the period 2005-2014 were registered only 10 species, growing at much lower quantitative values (Klisarova et al., 2015).

Akashiwo sanguinea is a cosmopolitan dinoflagellate that has been observed to form major blooms in coastal ecosystems around the world. A. sanguinea plays a major role in the ecology of many marine environments, including coastal ecosystems with variable salinities, where its euryhaline character makes it competitive (Badylak et al., 2014).

Akashiwo sanguinea (K.Hirasaka) G.Hansen & Ø.Moestrup in N.Daugbjerg, G.Hansen, J.Larsen, & Ø.Moestrup 2000 (Daugbjerg et al., 2000) Synonyms: Gymnodinium sanguineum K.Hirasaka 1922, Gymnodinium splendens Lebour 1925, Gymnodinium nelsonii Martin, 1929. The genus name akashiwo is Japanese for red tide.

A. sanguinea is exclusively planktonic and has a worldwide distribution in temperate and tropical waters. It is nearly always found in coastal and estuarine locations. It is one of the largest dinoflagellates with about 40-80 µm length and 30-50 µm width. According to Bockstahler and Coats (1993), A. sanguinea is mixotrophic, being primarily photosynthetic but also feeding on ciliate protozooplankton if necessary.  It is also susceptible to parasitic dinoflagellates such as Amoebophrya (Coats and Park, 2002). In general, this species is not considered to be toxic. Reports of toxicity in A. sanguinea are mostly anecdotal and based on its abundance in natural populations in which mortality has occurred. However, toxicity of mice was reported by Tindall et al., (1984). Jessup et al., (2009) reported extensive seabird mortality caused by surfactant-like protein exudates derived from A. sanguinea, which coated their feathers and neutralized natural water repellency and insulation (Hargraves P. E, 2011, Akashiwo sanguinea. Protists of the Indian River Lagoon. Smithsonian Institution, http://www.sms.si.edu/IRLSPEC/Akashi_sangui.htm). For instance, Gymnodinium splendens and Gonyaulax polyedra release toxins that kill or exclude zooplankton and fish. Without herbivores, such blooms may be quasi-permanent (Longhurst, 2007).

Gymnodinium splendens (A.sanguinea) is a common Black Sea phytoplankton species but was observed in bloom concentrations for the first time in the 90's (1995, 1999) (Moncheva et al, 2001; Velikova et al., 1999). Near the coast of Florida blooms of A.sanguinea are detected at salinities from 7 to 23 ‰ (Phlips et al., 2012), in Monterey Bay the species blooms even at 33.53 ‰. In its mass development A.sanguinea can cause sea bird kills due to surfactant-like proteins that coat feathers and neutralize water repellency and insulation (Lewitus et al., 2012).

The estimated duration of cell division was 1.29 h, and the estimated in situ growth rate was 0.45 d ^-1, cells of A. sanguinea actively divided in the deeper layer (3.5 to 5 m) from 18:00 (Katano et al., 2011).

Although the number of species in the Bulgarian marine aquatories rarely crosses the border of 100x106 cells.m^-3 due to its relatively large size (L = ~ 80μm), with its mass development the produced biomass reaches and exceeds bloom level of 10 000 mg.m^-3 according to Moncheva and Parr (2010).

Results

In tracing the available at IRR Varna data for the development of the species since 1993, a tendency for reduction of its quantitative development in the Bulgarian marine aquatories was established (Figure 1). Its biomass decreases with linear trend y = -387.6x + 7004.2 and R² = 0.1789, range= 53.64 ÷ 24018.82 mg.m^-3, mean= 2740.56 mg.m^-3, std.dev= 5685.58 and the number with y = -3.2589x + 71.116 and R² = 0.1393, range=2.54 ÷ 216.33 mln. cells.m^-3, mean=35.27 mln. cells.m^-3, std.dev= 54.17 (Figure 1).

The development of A. sanguinea in the brackish, hyper-eutrophic water areas of Varna and Beloslav Lakes is with lower values (3 times) (Figure 2). In the lakes, a decrease of development values over the years is also observed: the biomass decreases depending on regression y = -180.12x + 1696.6 and R² = 0.1107, range= 8.09÷ 4661.44 mg.m^-3, mean= 795.99 mg.m^-3, std.dev= 1482.89 and the number with y = -1.944x + 22.817 and R² = 0. 0525, range= 0.25÷ 74.23 mln. cells.m^-3, mean= 13.097 mln. cells.m^-3, std.dev= 23.24 (Figure 2).

The development of A.sanguinea takes place mainly in the surface water layer 0-10 m (Figure 3) and only in the periods from February to August, 1995 and from October, 2000 to June, 2001 exhibited high values in depth (till 25 m, thermocline and bottom) (Figure 4 and 5). In 2015 A. sanguinea was not detected in the samples.

Figure 1 Maximum annual values of number (mln. cells.m-3, right axis) and biomass (mg.m-3, left axis), of Akashivo sanguinea in the Bulgarian marine aquatories, 1993-2015

Figure 2 Maximum annual values of number (mln. cells.m-3, right axis) and biomass (mg.m-3, left axis), of Akashivo sanguinea in the Varna and Beloslav lakes, 1993-2015(data of IFR-Varna)

Figure 3 Distribution of the maximum numbers (mln. cells.m-3) and biomass (mg.m-3) of A.sanguinea by depth, 1995-2015

In the long-term set of samples from the water area along the entire Bulgarian coast, the highest bloom values of A. sanguinea were established in Varna Bay (24018.82 mg.m^-3),  (1995, August, station B-3, 0m), (Technical Reports of Institute of fish resources, 1995); 30 miles in front of cape Galata (12240 mg.m^-3), (2001, June, G-30, 5 m), (Velikova et al., 2004) and 10 miles in front of cape Emine (10644 mg.m^-3), (2001, June, E-10, 10 m) (Technical Reports of Institute of fish resources, 2001) (Figure 6). In all other cases the species is present in the samples, but develops below the bloom concentrations.

Figure 4 Distribution of the maximum number (mln. cells.m-3) of A. sanguinea in years, months and depths from 1995 to 2015

Figure 5  Distribution of the maximum biomass (mg.m-3) of A. sanguinea in years, months and depths from 1995 to 2015

In the investigated 20-year period, the maximum length of the observed A. sanguinea cells decreases (Figure 7) under a linear relationship: y = -2.2624x + 91.023 and R² = 0.6181.

Discussion

In the period 1995 - 2015 A.sanguinea was present in 740 numbers of analyzed samples. A. sanguinea is a relatively rare phytoplankton species in the Bulgarian marine aquatories with frequency of occurrence 18.54%.

After 1995, the only blooms of A. sanguinea were registered in 2001 (Velikova et al., 2004). On the contrary, during the Knorr cruise in spring, 2001 (Soydemir et al., 2002) was registered high abundance of dinoflagellates (98% of the total biomass) due to the proliferation of Gymnodinium splendens, co-dominated by the coccolothophoride E. huxley (42% of the total abundance) and an average biomass and abundance much higher than the reported values in the western Black Sea offshore area during the last decade (Moncheva S., State of knowledge of eutrophication and biological response in the Black Sea Ecosystem, Report-biology.doc, ftp://ocean.ims.metu.edu.tr/).

Since 2002 blooms of A. sanguinea have not been registered (Figure 1). This coincides with the general reduction of phytoplankton development in the Bulgarian Black Sea water (Bodeanu et al., 1998; Gerdzhikov and Petrova, 2008; Klisarova et al., 2015; Velikova et al., 2001). Another reason could be the lack of samples precisely at the time of high concentrations of the species, because of the standard scheme of monthly sampling.

Figure 6  Distribution of the maximum biomass (mg.m-3) of A. sanguinea along the Bulgarian coast, 1995-2015 (Ocean Data View v4.7.4, 2015 Reiner Schlitzer)

Figure 7 Dynamics of maximum linear dimensions (MLD) (µm) of the observed in the Bulgarian marine aquatories cells of A. sanguinea

According to Longhurst (2007), Gymnodinium splendens specially adapts to high nutrients and low turbulence. The ideal case to represent this quadrant are the red tides that occur in permanently stratified coastal waters where nutrient input is both continuous and strong. 

These characteristics of the development of A. sanguinea can explain the established by us 3 times lower concentrations of the species in nutrient-rich water of Varna and Beloslav lakes where due to no great depth and intense shipping traffic, a higher level of turbulence might be suggested compared with the water of the coastal areas (Figure 1 and 2).

In comparison, many developing in front of the Bulgarian coast blooming species were registered with fold higher concentrations in the catchment areas of lakes (Petrova and Gerdzhikov, 2006; 2007; 2011; 2012; 2013; Velikova and Petrova, 1999).

In contrast, blooms of A. sanguinea in the Bulgarian marine aquatories were registered only in Varna Bay and in water influenced by the Danube inflow (10-30 sea miles away from the coast). High numbers of the species were recorded also in Burgas Bay and one-mile area in front of Balchik, areas traditionally defined as "hot spots" in the coastal ecosystem (Figure 6).

The reducing of the size (Figure 7) of the representatives of A. sanguinea developing in the Bulgarian marine aquatories can probably be explained by the tendency of reduction of anthropogenic pressure. From this perspective, the average size of the species could be used as an index of measuring the degree of eutrophication of our sea water areas.

Materials and methods

The study was carried out within the period 1995 - 2015 in the Bulgarian Black Sea aquatories. The scientific expeditions were conducted on board the R/V Prof. Al. Valkanov. 3991 phytoplankton samples were collected from 127 stations at standard horizons (0, 10, 25, 50, 75 and 100 m) by bathometers type Niskin-5L or in shallow water (up to 15 m depth) at surface – bottom horizons (Figure 8). The samples were fixed onboard the ship in 2% formalin solution and concentrated by the sedimentary method (Morozova-Vodyanitskaya, 1954).

The qualitative and quantitative analyses of the samples were performed with a light microscope Nikon E400 and Olympus BX41 in counting cells Sedgewick Rafter - 1 ml and Palmer – Maloney - 0.05 ml, using standard methods (Moncheva and Parr, 2010). Software Phytomar 2.0 (IFR - Varna, 2008), Ocean Data View (v4.7.4, 2015 Reiner Schlitzer) and Excel 12 (Microsoft Office 2007) were used for calculations and graphs.

Figure 8 Map with sampling stations (Ocean Data View v4.7.4, 2015 Reiner Schlitzer)

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