This paper describes the first use of aerial observations by a drone as an additional means for choosing sampling points during field studies of cyanobacterial harmful algal blooms (CyanoHABs) in selected Bulgarian waterbodies and the use of HPLC analysis of marker pigments for the fast determination of phytoplankton composition and biomass. The selection of waterbodies was based on the authors’ personal expertise and data collected over a 25-year period. In all sites chosen by drone, there were high levels of cyanobacteria and cyanotoxins were present: microcystins (MC-LR, MC-RR, MC-YR in Durankulak Lake and MC-LR and MC-RR in the Sinyata Reka Reservoir), cylindrospermopsin (in the Vaya Lake and in the Mandra Reservoir) and saxitoxins (in Durankulak Lake). The finding of cylindrospermopsin is the first in Bulgaria, the detection of saxitoxins is the first for Durankulak Lake and the microcystins records are the first for Sinyata Reka Reservoir. Considering the high total number of wetlands in Bulgaria, many of which are lowland, small and shallow and therefore vulnerable to CyanoHABs, we recommend further use of drones and HPLC in monitoring, which should speed up detection and reduce sampling efforts while enabling valuable information to be gathered.

With the increasing demand for natural and safe products in cosmetics, algae with their diverse and valuable bioactive compounds are gaining vital importance. Until now, cosmetics have focused mainly on the use of freshwater and marine algae. However, algae are not restricted to aquatic habitats. They are found in essentially every type of aeroterrestrial and extreme environment on the Earth. There, they have to cope with harsh ecological conditions and have developed special strategies to thrive in these inimical habitats. Although not thoroughly studied, their adaptations include protective biochemical compounds which can find their application or are already used in the field of cosmetics. With proper cultivation techniques, algae from these habitats can provide novel sources of high-value functional products for the cosmetics industry, which have the advantage of being obtained in eco-friendly and cost-effective processes. However, it has to be considered that a few aeroterrestrial and extremophilic algae can be toxin producers, and in order to ensure conformity to the safe quality standards, all new ingredients must be properly tested. The aim of the present review is to unveil the hidden and underestimated potential of the enigmatic algae of aeroterrestrial and extreme habitats for the rapidly developing modern cosmetic industries.

The scientific and public awareness of hazardous photosynthetic prokaryotes (cyanobacteria/cyanoprokaryotes) and especially the contamination of drinking-water reservoirs with cyanotoxins is world-wide increasing. Recently much more attention has been paid to the events and results of mass proliferation of these toxic organisms even in South-East European countries in spite of the fact that, as a rule, they are not controlled by national legislation.The present paper presents a summary of results of such studies carried out in summer-autumn periods of the last 15 years (2000-2015) in Bulgarian water bodies differing by location, morphometry and trophic status, incl. drinking water reservoirs, recreational lakes and sites of nature conservation importance. A multivariate analysis allowed to outline the distribution patterns and environmental drivers of the planktonic cyanoprokaryote assemblages in relation with the available data on the water bodies, highlighting species composition and abundance of the main taxa, including potentially toxic species. Samples analysis by HPLC-DAD and/or LC/MS, ELISA and in vitro cytotoxicity tests allowed detection of microcystins, nodularins and saxitoxins. Toxin concentration ranged between 0.1 and 26.5 µg L-1 in water samples and between 10.9 and 1070 µg g-1 (d.w.) in concentrated (net) samples. Despite the fact that microcystins were not found in all studied water bodies and that the recorded levels were still lower in comparison with some other European countries, the fact that cyanotoxins were detected in 16 water bodies (incl. 3 drinking water reservoirs) could serve as an alert for the need of recognition of cyanotoxins as a new health risk factor in the country. Therefore, permanent monitoring with identification of toxins in water bodies at risk and activities for limitation and control of toxic blooms are urgently needed, in combination with increase of the attention to the effects of cyanotoxins on both human health and health of aquatic ecosystems in Bulgaria.

The paper presents results from the first application of polyphasic approach in studies of field samples from Bulgaria. This approach, which combined the conventional light microscopy (LM) and molecular-genetic methods (based on PCR amplified fragments of microcystin synthetase gene mcyE), revealed that almost all microcystin-producers in the studied eutrophic waterbodies belong to the genus Microcystis. During the molecular identification of toxin-producing strains by use of HEPF × HEPR pair of primers, we obtained 57 sequences, 56 of which formed 28 strains of Microcystis, spread in six clusters of the phylogenetic tree. By LM, seven Microcystis morphospecies were identified (M. aeruginosa, M. botrys, M. flos-aquae, M. natans, M. novacekii, M. smithii, and M. wesenbergii). They showed significant morphological variability and contributed from <1% to 98% to the total biomass. All data support the earlier opinions that taxonomic revision of Microcystis is needed, proved the presence of toxigenic strains in M. aeruginosa and M. wesenbergii, and suppose their existence in M. natans. Our results demonstrated also that genetic sequencing, and the use of HEPF × HEPR pair in particular, can efficiently serve in water quality monitoring for identifying the potential risk from microcystins, even in cases of low amounts of Microcystis in the water.

Carotenoids have received particular attention both for their importance in algal systematics and hydrobiology and for their health benefits for humans, along with other applications in various industries. Here, we provide new data on the carotenoid content of five Vischeria/ Eustigmatos strains isolated from aeroterrestrial habitats in Bulgaria and kept in the Algal Collection of Sofia University (ACUS). The obtained pigment pattern with nine carotenoids and generalization of literature data allowed us to update the knowledge on the chemotaxonomic characteristics of Eustigmatophyceae with a total of 47 pigments reported for the group, out of which 37 are carotenoids. The important photosynthetic pigment lutein, considered also a high-value product with extensive applications in feed, food, nutraceutical and pharmaceutical industries, was proved as a novel carotenoid for the group. The results also confirmed the presence of luteoxanthin, which is rarely reported in algae. All strains had a high content of the commercially valuable health-promoting xanthophyll astaxanthin. There were also different amounts of beta-carotene, violaxanthin, vaucheriaxanthin, canthaxanthin, zeaxanthin and antheraxanthin. Based on the differences in the pigment composition of the members of the ecological groups of aquatic and aeroterrestrial species, we propose the existence of two main pigment types in the class Eustigmatophyceae, with the aquatic type further divided into freshwater and marine pigment subtypes.

The rising interest in harmful cyanoprokaryote blooms promotes an increase of phycological and ecological research on potentially toxic species and their hazardous substances. The present study aimed to identify the main microcystin (MC) producers and their contribution to the phytoplankton of shallow waterbodies in Bulgaria, applying different methods. The sampling was performed in August 2019 in nine lakes and reservoirs, two of which (reservoirs Kriva Reka and Izvornik 2) were studied for the first time. The high contribution of cyanoprokaryotes to the total species composition and phytoplankton abundance was proved by light microscopic (LM) observations and HPLC analysis of marker pigments. The LM identification of potential MC-producers was supported by PCR amplification of mcyE and mcyB genes. The MCs amounts, detected by HPLC-DAD, varied by sites with a range from undetectable concentrations to 0.46 µg L−1 with only one recorded variant, namely MC-LR. It was found only in the reservoirs Mandra and Durankulak, while toxigenic MC-strains were obtained by PCR from five more waterbodies. Both LM and PCR demonstrated that the MC-producers were Microcystis aeruginosa and M. wesenbergii, despite their occurrence in low amounts (<0.5–5% of the total biomass) when filamentous cyanoprokaryotes dominated.

The review summarizes the available knowledge on toxins and their producers from rather disparate algal assemblages of aeroterrestrial, airborne and other versatile extreme environments (hot springs, deserts, ice, snow, caves, etc.) and on phycotoxins as contaminants of emergent concern in soil and plants. There is a growing body of evidence that algal toxins and their producers occur in all general types of extreme habitats, and cyanobacteria/cyanoprokaryotes dominate in most of them. Altogether, 55 toxigenic algal genera (47 cyanoprokaryotes) were enlisted, and our analysis showed that besides the “standard” toxins, routinely known from different waterbodies (microcystins, nodularins, anatoxins, saxitoxins, cylindrospermopsins, BMAA, etc.), they can produce some specific toxic compounds. Whether the toxic biomolecules are related with the harsh conditions on which algae have to thrive and what is their functional role may be answered by future studies. Therefore, we outline the gaps in knowledge and provide ideas for further research, considering, from one side, the health risk from phycotoxins on the background of the global warming and eutrophication and, from the other side, the current surge of interest which phycotoxins provoke due to their potential as novel compounds in medicine, pharmacy, cosmetics, bioremediation, agriculture and all aspects of biotechnological implications in human life.

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