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The Optically Stimulated Luminescence (OSL) dating started in 2004 in the Geological Institute of Hungary, then continued in the successor Geological and Geophysical Institute of Hungary, and the Mining and Geological Survey of Hungary. Our OSL ages have contributed to the reconstructions of palaeohydrography and palaeomorphology as well as the timing of neotectonic movements and archaeological sites among others. Principally, quartz is used for dating because it is an abundant and resistant mineral of the sediments and it serves as a natural retrospective dosimeter that is trapping and storing charges in the point defects of its crystal lattice. During the dating of Upper Pleistocene and Holocene sediments, it has been recognised that the luminescence properties of quartz and its suitability for OSL dating show local differences. For example, in some places of Hungary, quartz has much dimmer luminescence than in other areas of the country, or it is saturated due to lower radioactive radiation, which makes OSL dating challenging. The aim of this study was to find the causes of these local differences. Quartz was separated from some plutonic, volcanic, metamorphic and older sedimentary rocks and sediments of Hungary, which can be source of some Upper Pleistocene and Holocene sediments in Hungary. Beside OSL measurements, for the detection of the chemical impurities that can cause defects in the crystal lattice, Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS), Micro-Fourier Transform Infrared Spectrometry (Micro-FTIR) and Infrared Spectrometry were applied. Thermal Analysis and X-Ray Diffraction were applied for the characterization of the quartz separates. The concentration of the radioactive elements in the studied formations was also determined based on laboratory High-Resolution Gamma-Ray Spectrometry. The results of the OSL measurements indicated that sediments, sedimentary rocks and some volcanic tuffs give brighter or more intensive luminescence than the quartz of the studied metamorphic and plutonic rocks. The quartz grains of only a few formations are appropriate for OSL dating if they get into the Pleistocene or Holocene sediments. They are solely sediments and sedimentary rocks, certain Upper-Miocene, Upper-Miocene–Pliocene and Upper-Oligocene sands of Kalla Gravel, Zagyva and Torokbalint Sandstone formations. The results of the LA-ICP-MS measurements indicated Li, Na, Al, P, K, Ti and Ba impurities in the quartz grains, while Micro-FTIR analysis detected coupled substitution of Al3++ H+ at the position of Si4+ incorporated as AlOH structural hydroxyl, and molecular water as well. These impurities partly are present in mineral phases, as clay minerals or smectite, and mica, according to the results of XRD, Thermal Analysis and Infrared Spectrometry. The causes of the local differences in luminescence properties of quartz grains and their suitability for OSL dating are probably their different source rocks, and diverse thermal and sedimentary history. The quartz grains that are originated directly from metamorphic and plutonic rocks by erosion show unfavourable OSL properties, probably because, during their long cooling period, they relax most of the point defects in their crystal lattice that were formed during their formation on high temperature. The OSL properties of the quartz grains of the sediments and sedimentary rocks are more favourable due to the repeated cycles of radioactive irradiation and the zeroing of the OSL in sunshine which increase the sensitivity of quartz.

Hungarian wine is not in such a bad position as it is considered. We have outstanding specialists and the country’s natural conditions are also proper. In Hungary, there are wine production and wine consumption has a major traditions. The role of marketing in our wine industry is growing more and more, and this trend is expected to continue in the future. The development of tourism in wine-growing regions and advertising would considerably promote the popularity of Hungarian wine.

Érmellék is an area of the Partium. It is situated between Szilágy and Bihar counties and between River Kraszna and Berettyó. Its name is from Ér, the small brook, which crosses the area. Érmellék is famous for its wine production, because its climate is favourable for this kind of trade. Only a few information sheets of the small Partium-belonging micro-region, Érmellék are published nowadays. Érmellék has extremely picturesque architectural wine cellar rows, of which ”szalacsi” wine cellar row (wine cellar row system) with its unique structure consists of about a thousand cellars.

International audience; Numerous assessment tools based on biological units (i.e. bioassays) have been developed to complement physicochemical or immunochemical methods. Bioassays provide complex information on the overall toxic potencies or biological activities of cyanobacterial samples including isolated cyanotoxins as well as natural blooms. The present chapter provides rationale beyond the use of bioassays and shows key examples for the assessment of toxic cyanobacteria covering both the toxicology (or more specifically toxinology for natural toxins) and ecotoxicology arenas for all levels of biological organization (from in vitro to in vivo, from plants to animals). At the present time, bioassays for potential use in routine monitoring need further research and validation, and there is no clear agreement among scientists on the list of recommended bioassays. Nevertheless, bioassays play an important role in the primary screening of cyanobacteria for toxic compounds and for their detection under specific situations such as limited access to advanced instrumental analytical equipment or indications of toxic metabolites other than (or in addition to) the known cyanotoxins.

A bor egy olyan nemes ital, amelyet egyes nemzetek büszkén fogyasztanak és készítenek. A szőlőművelésnek és borkészítésnek hagyományai vannak. A nemzetközi borpiacon a hagyományos bortermelő országok mellett megjelentek a „hódító” bornemzetek. A magyar borászat nincs rossz helyzetben. Kiváló szakembereink vannak és a természeti adottságaink is megfelelőek. A borfogyasztás szokásainak vizsgálatából kiderült, hogy itthon az egyik legkedveltebb alkohol a bor, amelyet a magyarok kiváltképpen ünnepekkor fogyasztanak szívesen. Míg a hazai borfogyasztás évről évre csökken, a fiatalok körében mégis divat lett érteni a borokhoz. A magyar borkultúra szintje viszonylag alacsony, bár fejlődési pályán van. Szükséges tehát a borkultúrát javító szerveződések támogatása, kitalálása.

Aims: In this study, we report a new, simple methodology for the monitoring of Cryptosporidium oocysts and Giardia cysts in drinking water samples, ranging from 10- to 1000-l, which combines a new ARAD microfibre filtration of the (oo)cysts from drinking water and loop-mediated isothermal amplification (LAMP) of a human pathogenic Cryptosporidium parvum, Cryptosporidium hominis, Cryptosporidium meleagridis and Giardia duodenalis Assemblage A and B specific DNA sequence. Methods and Results: During the evaluation of the new concentration and detection technique, spiked reagent and matrix water samples plus blank samples were filtered and tested. In total, 27 samples have been investigated. The results clearly demonstrate that the methodology of using a new ARAD filter, which passed through 1000 l of drinking water with high turbidity (2 NTU), and followed by the LAMP assay was able to detect at least one (oo)cyst in 10 l of drinking water based on a 1000-l sample, taken over a 24-h period. Conclusions: The described protozoa detection methodology is sensitive, rapid and cost-effective. Significance and Impact of the Study: This effective procedure will be useful for small waterworks to achieve continuous monitoring and is also of value for screening catchments to identify those that require further treatment and more detailed microscopic counts.

Safe drinking water is a top priority in preventing disease outbreaks and is of general concern to everyone. This study examines the occurrence of Cryptosporidium and Giardia in Hungarian drinking water supplies for the first time. A total of 76 raw and drinking water samples were examined using the U.S. EPA Method 1623. From these 15 of 34 (48.4%) raw water samples tested positive for Giardia and 7 (26.6%) for Cryptosporidium. Twelve of 45 (26.7%) drinking water samples were positive for Giardia and 6 (13.3%) for Cryptosporidium. Overall, Giardia cysts and/or Cryptosporidium oocysts were detected in 48% of the raw water samples and 35% of the drinking water samples. The highest levels in drinking water were found to be 3 oocysts/100 litres of Cryptosporidium and 63.6 cysts/100 litres for Giardia, enough to cause giardiasis. The highest levels in raw water were 1,030 cysts/100 litres for Giardia and 50 oocysts/100 litres for Cryptosporidium and higher oocyst densities were associated with source water receiving effluents from sewage treatment plants or originating from a forest environment. In addition to this monitoring, riverbank filtrated water and raw water from the River Danube in Budapest were monitored in order to ascertain protozoan removal efficiency of riverbank filtration (RBF). A total of 157 samples, including 87 samples from the River Danube and 70 samples post RBF, were examined. Cryptosporidium and Giardia were detected regularly in the river water but never in riverbank filtered water suggesting the effectiveness of RBF as a purification method. The occurrence of Cryptosporidium oocysts and Giardia cysts in the investigated water supplies may require the water utilities and water authorities in Hungary to apply additional monitoring and treatment and/or watershed controls.

The wine is still in Hungary in Hungarian history, culture, and one of the key elements of the economy are considered . Last, but not to the point of producing and selling the wine regions of livelihoods. Thousands of years in wine-producing traditions preserved in the modern winemakers, wine competitions demonstrate their skills. The Tokaj wine region of the World's major and significant wine regions in Hungary.

At the Oregon State University Workshop we utilized two ‘culture/maintenance-free’ microbiotests, the Thamnotoxkit F™ with the anostracan crustacean Thamnocephalus platyurus and the Daphtoxkit F™ magna with the cladoceran crustacean Daphnia magna, to determine the toxicity of water samples in a blind format. The Thamnotoxkit was applied to 7 samples and the Daphtoxkit to 12 samples. The chemical composition of the water samples to which the Toxkit microbiotests had been applied was disclosed a posteriori by the organizers and appeared to contain chlordimeform, colchicine, phosdrin, mercuric chloride, sodium arsenite, metham sodium, sodium cyanide, trimethylol propane phosphate, p-chlorophenol and a natural sediment sample containing mercury. Three of the water samples were blanks that had not been spiked with chemicals. No false positives were obtained with the two Toxkits and all the toxic waters were earmarked as such by the microbiotests, except trimethylol propane phosphate, which was not found to induce acute effects in Daphnia magna in the non-diluted water sample containing 100 mg l−1 of this compound. Copyright © 2004 John Wiley & Sons, Ltd.

Various types of toxicity classification systems have been elaborated by scientists in different countries, with the aim of attributing a hazard score to polluted environments or toxic wastewaters or of ranking them in accordance with increasing levels of toxicity. All these systems are based on batteries of standard acute toxicity tests (several of them including chronic assays as well) and are therefore dependent on the culturing and maintenance of live stocks of test organisms. Most systems require performance of the bioassays on dilution series of the original samples, for subsequent calculation of L(E)C50 or threshold toxicity values. Given the complexity and costs of these toxicity measurements, they can only be applied in well-equipped and highly specialized laboratories, and none of the classification methods so far has found general acceptance at the international level. The development of microbiotests that are independent of continuous culturing of live organisms has stimulated international collaboration. Coordinated at Ghent University, Belgium, collaboration by research groups from 10 countries in central and eastern Europe resulted in an alternative toxicity classification system that was easier to apply and substantially more cost effective than any of the earlier methods. This new system was developed and applied in the framework of a cooperation agreement between the Flemish community in Belgium and central and eastern Europe. The toxicity classification system is based on a battery of (culture-independent) microbiotests and is particularly suited for routine monitoring. It indeed only requires testing on undiluted samples of natural waters or wastewaters discharged into the aquatic environment, except for wastewaters that demonstrate more than 50% effect. The scoring system ranks the waters or wastewaters in 5 classes of increasing hazard/toxicity, with calculation of a weight factor for the concerned hazard/toxicity class. The new classification system was applied during 2000 by the participating laboratories on samples of river water, groundwaters, drinking waters, mine waters, sediment pore waters, industrial effluents, soil leachates, and waste dump leachates and was found to be easy to apply and reliable.

Cylindrospermopsis raciborskii is a planktonic freshwater cyanobacterium that has become increasingly prevalent in tropical and temperate water bodies world-wide. This species is of concern from a water-quality perspective because of its known ability to produce toxins that can affect the health of humans and other animals. This study investigates genetic vari- ation between strains of C. raciborskii isolated from freshwater rivers and reservoirs in Australia, Brazil, Germany, Hungary, Portugal and the USA. Strains were first character- ized by analysis of their 16S rRNA gene nucleotide sequences and were found to have a sequence divergence of 99.1%. A phylogenetic tree, constructed using the 16S rRNA gene sequences showed that strains grouped into Australian, European and North/South American phylotypes. To investigate further the observed separation of strains into geographically distinct groups, we applied a cyanobacterium-specific short tandem repeat sequence tech- nique, HIP1. An electrophoretic comparison of the HIP1 polymerase chain reaction products showed clear distinctions between the C. raciborskii strains. A phylogenetic tree, based on the repeat element banding patterns, also revealed three distinct groups of C. raciborskii strains. The first group consisted of strains from the USA and Brazil; the second comprised European strains from Germany, Hungary and Portugal; and the third were strains from Australia. In general, between-country variation was greater than within-country variation, indicating that this fingerprinting technique can successfully distinguish C. raciborskii strains taken from different global locations. The relationship between toxicity and the observed HIP1 polymerase chain reaction fingerprint profiles was less clear, although it is interesting to note that of the strains analysed in this study, only Australian strains are known to produce cylindrospermopsin and only Brazilian strains have been reported to produce paralytic shellfish poisoning toxins.

The Artemia salina bioassay was successfully applied to the analysis of the hepatotoxic cyanobacterial alkaloid and protein synthesis inhibitor, cylindrospermopsin. A dose-dependent response in mortality was observed for purified cylindrospermopsin and LC(50) values decreased with time from 8.1 to 0.71 microg/ml(-1), between 24 and 72 h, respectively. Cylindrospermopsin was slightly less potent than micro cystin-LR, with similar LC(50) values on a gravimetric basis, but was more toxic to A.salina than the protein synthesis inhibitors, cycloheximide, chloramphenicol and tetracycline. Cylindrospermopsin-containing strains of the cyanobacterium Cylindrospermopsis raciborskii were found to be toxic to A.salina and the LC(50) concentration for these strains over time was greater than the LC(50) for purified cylindrospermopsin, with the exception of C. raciborskii strain CR1.

(2000). Phytoplankton composition, biodiversity and a pilot survey of toxic cyanoprokaryotes in a large cascading reservoir system (Tiete basin, Brazil) SIL Proceedings, 1922-2010: Vol. 27, No. 5, pp. 2734-2742.

Lake Velencei is a shallow lake in Hungary. The first cyanobacterial bloom of Microcystis aeruginosa appeared in the lake in 1991. This cyanobacterial species was also found earlier but blooms were not recorded. The toxins of M. aeruginosa were studied in Lake Velencei in 1992. The present study deals with the toxicity of a M. aeruginosa biomass of different origin detected by intraperitoneal (ip.) mouse assay, Thamnotoxkit F microbiotest, and toxin analysis by high-performance liquid chromatography (HPLC). The biomass was collected by plankton net and the isolated strains were cultured from sediments and from bloom material. Minimal lethal doses (MLD) by ip. mouse assay ranged from 20 to 50 mg/kg -1 . Comparing the results from 1992 and 1997-1998, no change in the toxicity of M. aeruginosa was detected. According to the HPLC analysis the combination of microcystins varied in a narrow range with microcystin-LR found in each sample. The cyanobacterial blooms caused by M. aeruginosa in Lake Velencei considerably decreased, though it remained in the sediment with the same toxicity. The Thamnotoxkit F microbiotest was an alternative method for detecting the cyanotoxins.

The Thamnotoxkit F was evaluated for detecting cyanobacterial toxins. This is a 24 hour bioassay using, as test organisms larvae of the freshwater anostracan crustacean Thamnocephalus platyurus hatched from cysts. Freeze-dried Microcystis aeruginosa samples from freshwaters throughout Hungary were tested. Good correlation was obtained between the results of intraperitoneal mouse bioassays and Thamnotoxkit F assays. The LC 50 's for purified microcystins- LR, -RR, and -YR ranged from 0.1 to 2.27 μg per mL, respectively in the Thamnotoxkit assay. The LC 50 of laboratory cultures of Cylindrospermopsis raciborskii isolated from samples from Lake Balaton, was 0.574 mg lyophilized material per mL, in a four-laboratory trial. Although not developed originally for detecting cyanobacterial toxins, the Thamnotoxkit F has good potential as an initial screening procedure for toxicity assessment of cyanobacterial scums and blooms in freshwaters.

The transmission route of giardiasis not yet understood and why some infected individuals remain asymptomatic while others become quite ill. The drinking water quality is supposedly responsible for the prevalence of asymptomatic Giardia duodenalis infections in different areas, therefore asymptomatic giardiasis has been investigated in three water supply areas of Hungary: three hundred stool samples from inhabitants of Budapest, Füzér and Mátrafüred were examined by immunological and molecular methods for the presence of G. duodenalis infections. Individuals were asked to fill out a validated questionnaire at the time of stool collection and the interview covered demographic data, family life, education and travel history.In Budapest and in Mátrafüred in one stool sample G. duodenalis Assemblage A, whereas in Füzér once G. duodenalis Assemblage A, once Assemblage B and twice mixed infection were detected. We found higher prevalence rate of 4% of G. duodenalis infections of asymptomatic people in the village Füzér, where the removal of the Giardia cysts of the drinking water treatment plant was not effective. This study throws a light the need to look into the possibility of other risks of Giardia infections such as water transmission routes. To our knowledge, this is the first study evaluating the prevalence of G. duodenalis infections in asymptomatic persons in Hungary.

Bacterial indicator organisms (e.g., coliforms, E. coli) and some chemical parameters (e.g., turbidity, ammonia) are basic monitoring tools used to measure both changes in drinking-water quality and the presence of hard-to-detect pathogenic bacteria, viruses and protozoa. Microscopically detectable free-living organisms, such as some groups of bacteria, fungi, nematodes, rotifers and protozoa, are usable as additional indicators of fecal or environmental contamination of drinking water as well as any changes in the drinking-water quality. Our aim in this paper is to summarize the results of microscopic examination of 913 drinking-water samples from different water sources in Hungary in 2004 and 2005 and to demonstrate how these results can be used to maintain safe and good-tasting drinking-water quality. A total of 277 drinking-water samples failed Hungarian microscopic water quality standards as a result of helminths (58%), protozoa (41%), iron bacteria (16%), sulfur bacteria (13%), fungi (11%), algae (5%) and multiple biological contaminants (34%). Based on these results, pipe washing or water storage tank cleaning was deemed necessary. In addition, a number of disinfection or filtration failures were found. Two detailed case studies show the usefulness of monitoring microscopic parameters to avoid disease outbreaks. To our knowledge this is the first paper discussing drinking-water microscopy based on Hungarian experience and practice, which could be useful and informative for other countries.

The aim of this study is to compare the responses of two different ecotoxicological methods for the determination of the toxic hazard of river or creek sediments in Hungary. Since water quality is intrinsically linked to sediment quality, the ecotoxicological control of sediments is also very important in the water quality policy. The 2000/60 EC Water Framework Directive aims at achieving a good qualitative and quantitative status of all water bodies by 2015. Fifteen sediments and four sludges were collected at different sites in Hungary, which are contaminated by industrial sewage. The first assay is the acute toxicity test with the crustacean Daphnia magna Straus (OECD 202) using larvae less than 1 day old, and an exposure time of 2 days. The second assay is the Ostracodtoxkit F with freshly hatched larvae of the crustacean Heterocypris incongruens and an exposure time of 6 days. The Ostracodtoxkit F test is a "direct sediment contact" bioassay, in which the test organisms are in continuous contact with the sediments. The Daphnia tests were applied on water extracts of the sediment without any contact of the test organisms with the contaminated sediment. The same 1:4 water/sediment ratio has been applied to both tests. The results showed higher toxic effects of the sediments to H. incongruens than to D. magna confirming the need to complement "water only" tests with "solid phase" assays for a meaningful evaluation of the toxic hazard of aquatic environments. The sensitivity of H. incongruens is similar to that of the other test species which are currently used for solid-phase assays. The growth inhibition of H. incongruens is a very sensitive endpoint for sediment toxicity testing. The sediment toxicity tested by ostracods showed strong correlation with concentration of total chromium, lead, and cadmium together.

In this study, we report a new, simple methodology for the monitoring of Cryptosporidium oocysts and Giardia cysts in drinking water samples, ranging from 10- to 1000-l, which combines a new ARAD microfibre filtration of the (oo)cysts from drinking water and loop-mediated isothermal amplification (LAMP) of a human pathogenic Cryptosporidium parvum, Cryptosporidium hominis, Cryptosporidium meleagridis and Giardia duodenalis Assemblage A and B specific DNA sequence.During the evaluation of the new concentration and detection technique, spiked reagent and matrix water samples plus blank samples were filtered and tested. In total, 27 samples have been investigated. The results clearly demonstrate that the methodology of using a new ARAD filter, which passed through 1000 l of drinking water with high turbidity (2 NTU), and followed by the LAMP assay was able to detect at least one (oo)cyst in 10 l of drinking water based on a 1000-l sample, taken over a 24-h period.The described protozoa detection methodology is sensitive, rapid and cost-effective.This effective procedure will be useful for small waterworks to achieve continuous monitoring and is also of value for screening catchments to identify those that require further treatment and more detailed microscopic counts.

We investigated the prevalence of Giardia and Cryptosporidium species and analysed the genotypes in 36 samples collected from different water sources and various geographic areas in Hungary. Samples were collected from drinking water and sewage treatment plants and from the recreation area of Lake Balaton. The (oo)cysts were purified according to the US EPA 1623 method and they were detected by immunofluorescence test (IFT). Genomic DNA was extracted from all samples and then the GDH target gene for Giardia and the SSUrDNA for both Giardia and for Cryptosporidium species were amplified by PCR. 24 out of 36 samples (67%) were Giardia positive and 15 (42%) were Cryptosporidium positive by IFT. PCR confirmed that 13 out of 36 samples (36%) were Giardia positive and 10 (28%) contained Cryptosporidium. Twelve Giardia and two Cryptosporidium PCR products were successfully sequenced. In seven samples G. lamblia Assemblage A and in one sample Assemblage B and in four cases Assemblages A and B have been found. In one sample C. parvum and in the other separate sample C. meleagridis were detected. Sequence analysis revealed a new subtype of G. duodenalis complex, clustered close to the Assemblage A group. This study provides the first report on simultaneous detection and genotyping of G. duodenalis and Cryptosporidium species from water supplies in Hungary.

Greater and greater demand is needed to detect and identify the new emerging biological agents in drinking water supply. The so called indicator organisms like bacteria (E. coli, coliform bacteria etc.) often do not indicate the real danger for human health because of their higher sensitivity to chlorination for e.g. The other pathogen organisms like Cryptosporidia, Giardia, Cyanobacteria and their toxins, viruses, Legionella, fungi have higher resistance in other words lower sensitivity to chlorine. So some water borne disease can emerge beside the indicator parameters are satisfying. A case study in Hungary could prove this. Calicivirus caused outbreak in two villages spreading by the network of potable water. There are some indicators in legislation used in Hungary with similar sensitivity like pathogens to point out the possibility of occurrence of pathogens. These organisms are not pathogens but their appearance indicate the neglected condition of the network and the procedure of treatment plant. On the other hand the organisms can be used for detecting hazards like chemicals, genotoxic agents etc. Two technical workshops were organised on Toxicity for Biodefense in Oregon State University, USA and University of Pecs, Hungary. Both of the workshops were organised on the idea that each participant/group brought were given identical samples, and results were compared. Both toxicity and

Microbiological agents can appear in the drinking water network by accidental events like breaking the pipe and the drinking water can be contaminated with sewage or contaminated soil, or neglected procedure and handling of water treatment plant and pipe system. On the other hand bioterrorism should be considered when different biological agents like viruses, bacteria, rickettsia, fungi, toxins, chimeras and prions can be used as biological weapons. If producing biological weapons is supported by governments like in the period of cold war this term of production can be regulated and controlled by international contracts, agreements. But nowadays the main threat is bioterrorism when agreements are ineffective. The primary goal of terrorism is to erode the security of a nation, to disrupt the continuity of society, and to destroy the nation’s social capital – its morale, cohesion, and values. In case only the international collaboration can decrease the

Cyanobacteria occur widely in lakes, reservoirs, ponds, and slow flowing rivers. Many species are known to produce toxins (cyanotoxins), a number of which are of concern for health. Cyanotoxins vary in chemical structure and may be found intracellular or released into water. There is not only a wide variation in the toxicity of known cyanotoxins but a substantial number of toxins have to date not been identified chemically. Chemical analysis of cyanotoxins is nowadays not used for routine monitoring because it is time consuming, it requires specialized equipment and expertise, and is hence expensive. There is hence an urgent need for rapid tests in surface waters to detect cyanobacterial toxins because of the need for safe drinking water and safe natural bathing waters, which may be burdened by cyanobacterial blooms or scums. Previous investigations have already shown that larvae of the anostracan crustacean Thamnocephalus platyurus are quite sensitive to neurotoxic and hepatotoxic cyanotoxins. The present paper reports on the sensitivity comparison of the (1 h) Rapidtoxkit (based on a sublethal endpoint) and the (24 h) Thamnotoxkit microbiotest (based on mortality). Both assays make use of larvae of T. platyurus. The Rapidtoxkit is a new microbiotest that determines the decrease of ingestion of colored particles by the crustacean larvae, which are stressed by a short exposure to toxicants. Fifteen cyanobacterial samples composed of laboratory strains and natural bloom samples were tested by both microbiotests. All samples were also analyzed concurrently by HPLC for microcystins and cylindrospermopsin. The correlation coefficient between the two microbiotests (r = 0.82) showed the very good correspondence between the sublethal and the lethal effects. No known toxins could be detected in some samples, although the latter were found highly toxic to the test organisms in both bioassays. These results point to the presence of unknown toxin(s) produced by some cyanobacteria such as e.g., the Cylindrospermopsis raciborskii strain isolated from Lake Balaton in Hungary. This comparative study clearly showed that the 1 h Rapidtoxkit is an attractive rapid alternative to the Thamnotoxkit microbiotest.

At the Oregon State University Workshop we utilized two "culture/maintenance-free" microbiotests, the Thamnotoxkit F with the anostracan crustacean Thamnocephalus platyurus and the Daphtoxkit F magna with the cladoceran crustacean Daphnia magna, to determine the toxicity of water samples in a blind format. The Thamnotoxkit was applied to 7 samples and the Daphtoxkit to 12 samples. The chemical composition of the water samples to which the Toxkit microbiotests had been applied was disclosed a posteriori by the organizers and appeared to contain chlordimeform, colchicine, phosdrin, mercuric chloride, sodium arsenite, metham sodium, sodium cyanide, trimethylol propane phosphate, p-chlorophenol and a natural sediment sample containing mercury. Three of the water samples were blanks that had not been spiked with chemicals. No false positives were obtained with the two Toxkits and all the toxic waters were earmarked as such by the microbiotests, except trimethylol propane phosphate, which was not found to induce acute effects in Daphnia magna in the non-diluted water sample containing 100 mg l(-1) of this compound.

The hepatotoxin cylindrospermopsin (CYN) is a potent inhibitor of protein synthesis in mammalian cells. It is produced by freshwater cyanobacterial blooms in countries such as Australia, the United States, Israel, Thailand, and Brazil. An interlaboratory comparison was organized as a first step to evaluate the measurement of CYN in lyophilized cyanobacterial cells. Six laboratories from Europe, Israel, and Australia participated in the trial. All of the methods used for extraction of the toxin and the high-performance liquid chromatography (HPLC) analysis were satisfactory on the basis of statistical evaluation, according to ISO standards 5725-1 and -2. Further comparison of all the extraction methods by the organizer indicated that the most effective extraction procedure used 5% formic acid to prevent interference in chromatograms by contaminant compounds when analyzed using HPLC employing isocratic conditions of 5% (v/v) aqueous methanol plus 0.1% (v/v) trifluoroacetic acid as the mobile phase.

Cylindrospermopsis raciborskii is a planktonic freshwater cyanobacterium that has become increasingly prevalent in tropical and temperate water bodies world-wide. This species is of concern from a water-quality perspective because of its known ability to produce toxins that can affect the health of humans and other animals. This study investigates genetic variation between strains of C. raciborskii isolated from freshwater rivers and reservoirs in Australia, Brazil, Germany, Hungary, Portugal and the USA. Strains were first characterized by analysis of their 16S rRNA gene nucleotide sequences and were found to have a sequence divergence of 99.1%. A phylogenetic tree, constructed using the 16S rRNA gene sequences showed that strains grouped into Australian, European and North/South American phylotypes. To investigate further the observed separation of strains into geographically distinct groups, we applied a cyanobacterium-specific short tandem repeat sequence technique, HIP1. An electrophoretic comparison of the HIP1 polymerase chain reaction products showed clear distinctions between the C. raciborskii strains. A phylogenetic tree, based on the repeat element banding patterns, also revealed three distinct groups of C. raciborskii strains. The first group consisted of strains from the USA and Brazil; the second comprised European strains from Germany, Hungary and Portugal; and the third were strains from Australia. In general, between-country variation was greater than within-country variation, indicating that this fingerprinting technique can successfully distinguish C. raciborskii strains taken from different global locations. The relationship between toxicity and the observed HIP1 polymerase chain reaction fingerprint profiles was less clear, although it is interesting to note that of the strains analysed in this study, only Australian strains are known to produce cylindrospermopsin and only Brazilian strains have been reported to produce paralytic shellfish poisoning toxins.

Many studies have been published on the human health effects of cyanobacterial toxicity. As a public health concern, we have to mention that the development of contact dermatitis, asthmalike symptoms, and symptoms resembling hay fever during bathing in cyanobacterial blooms have been also described. Microcystis aeruginosa, Anabaena flos-aquae, Aphanizomenon flos-aquae, and Cylindrospermopsis raciborskii are the most common species found in Hungarian freshwaters. A sensitization test on albino guinea pigs, and intradermal reactivity, and occular irritation test on albino rabbits were carried out with freeze-dried algal suspension in physiological salt solution. The sensitivity of guinea pigs is similar to that of humans. Microcystis, Anabaena, Cylindrospermopsis, and Aphanizomenon bloom and strain samples were examined in sensitization and irritation tests and no correlation was found between the toxin content and the allergenic character. The most toxic one (Microcystis aeruginosa) was not the most allergenic sample, but the nontoxic Aphanizomenon was the most allergenic one. The axenic strains were not allergenic at all. The pure microcystin LR was only slightly allergenic even in high concentration (1.5 mg/ml). Water and lipid soluble fractions were obtained by water and chloroform extraction of lyophilized algal suspensions. The chloroform fraction was bound on C18 cartridges and eluted by methanol in nine fractions. Only one of the lipid soluble fractions was skin irritative whereas the strongest irritative effect was shown by the water soluble fraction.

In the present study 4 hazardous wastes originated from 4 different typical industrial activities (leachates of waste oil, waste stockpiles of aluminium factory, sewage of leather industry and of the chemical industry) were examined with several Toxkit microbiotests: the Algaltoxkit, the Protoxkit, the Daphtoxkit and the Thamnotoxkit. The data obtained with the microbiotests were compared to those obtained with the standard conventional acute toxicity tests. The goal of the study was to try out the reproducibility of the tests on the same samples, prepared in the same laboratory and provided to all the participants of the interlaboratory exercise. The first reproducibility evaluation of the tests was made with the reference toxicant potassium dichromate (K2Cr2O7). The outcome was that the lowest coefficients of variation were found at the Thamnotoxkit and highest with Algaltoxkit. The same findings were observed with the industrial waste samples. Different responses were obtained depending on the preparation and the character of the waste. For example, whereas a strong (and highest) toxic effect was found with the Thamnotoxkit, the algal cells in the Algaltoxkit test grew better than in the controls, when exposed to the same waste sample. The sensitivity of the Protoxkit ciliates was the lowest for all the wastes examined. In conclusion of this study, the experience gained and data obtained indicate that the Toxkit microbiotests are suited for routine monitoring after modification of some technical aspects.

An interlaboratory trial with 8 laboratories was organised for evaluating the potential of the Thamnotoxkit F™ microbiotest for toxicity detection of 5 toxin producing cyanobacteria (Anabaena flos-aquae, Microcystis aeruginosa, Cylindrospermopsis raciborskii, Aphanizomenon flos-aquae, Tychonema bourrellyi) and the green alga Selenastrum capricomutum (Raphidocelis subcapitata) as a control. The Thamnotoxkit F™ test is a 24h bioassay using, as test organisms, larvae of the freshwater anostracan crustacean Thamnocephalus. platyurus hatched from cysts. Dilution series were prepared from freeze-dried samples of microalgae which were submitted twice to freezing and melting before extraction of the water soluble cyanotoxins. The concentration series used for each microalgal extract ranged from 0.3 mg/ml to 5.0 mg/ml. All samples were tested twice at different time by each examiner, eventually yielding 16 independent results obtained from 3 parallels (48 data for each sample). The samples of C. raciborskii were only studied by 4 examiners and gave 8 results (24 data). The water extracts of An. flos-aquae and M. aeruginosa were the most toxic to T. platyurus inducing 100% mortality in all concentrations used, except the highest dilution in which the mortality was, however, still around 70-80%. The extracts of C. raciborskii also induced 100% mortality as from the 1.0 mg/ml concentration and Aph. flos-aquae killed all the test organisms at the 3.0 mg/ml dilution. Water extracts of T. bourrellyi only caused total mortality in the highest test concentration. The water extract of the control alga S. capricomutum only gave a significant toxicity effect (around 30%) on the crustaceans in the highest concentration.