As filter feeders, bivalves and ascidians can accumulate contaminants present in the environment and pass them on to higher food chain levels as vectors. The consumption of bivalves contaminated with ...the potent neurotoxin domoic acid (DA) can cause amnesic shellfish poisoning in humans. The aim of this study was to determine seasonal differences in occurrence and accumulation of this phycotoxin in European oysters (
Linnaeus, 1758) (
= 46), Queen scallops (
Linnaeus, 1758) (
= 53), and edible ascidians of the
spp. (
= 107), originating from the same harvesting area in the Northern Adriatic Sea. The quantification was performed using ultra-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) preceded by derivatization with dansyl chloride. DA was found in very low concentrations throughout the year, with a maximum value of 810 μg/kg in Queen scallops. This study reveals differences in the occurrence and accumulation of DA between Queen scallops and the other two investigated species (oysters and ascidians) and the highest concentrations during the colder part of the year. Even though DA was detected in all of them, Queen scallops showed higher DA accumulation compared to the other two (
< 0.001), hence representing a sentinel species suitable for the monitoring of DA level in seafood.
The aim of this study was to determine the occurrence of the most representative cyclic imines (CIs) gymnodimine (GYM), pinnatoxin G (PnTX-G), and 13-desmethyl SPX C (SPX1) in Mediterranean mussels ...(Mytilus galloprovincialis Lamarck, 1819) (n = 416), European oysters (Ostrea edulis Linnaeus, 1758) (n = 104), Queen scallops (Aequipecten opercularis Linnaeus, 1758) (n = 52) and edible ascidians of the Microcosmus spp. (n = 104) originating from nine harvesting and breeding areas in the northern part of the Adriatic Sea using ultra-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). All CI concentrations were far below the guidance level of 400 μg SPXs/kg proposed by the EU Reference Laboratory for Marine Toxins. In contrast to Queen scallops and ascidians, in Mediterranean mussels and European oysters CIs were found throughout the year. Our data reveal the differences between species predisposed for CIs accumulation, as well as seasonal and locational variations in CIs occurrence.
Uzgoj, izlov i promet školjkašima reguliran je brojnim zakonima kojima je cilj stavljanje na tržište zdrave hrane. Zakonodavstvo ne predviđa kontrolu Vibrio spp., što može rezultirati stavljanjem na ...tržište i konzumacijom školjkaša kontaminiranih bakterijama ovog roda te posljedično prouzročiti pojavu bolesti u ljudi. U radu je prikazano značenje najvažnijih potencijelno patogenih halofilnih vibrija poput V. cholerae, V. parahaemolyticus i V. vulnificus u infekciji ljudi. Tijekom zadnjih dvadesetak godina u zemljama mediteranske regije provedena su istraživanja, koja su redom potvrdila prisutnost vibrija u morskoj vodi, školjkašima, sedimentu i u otpadnim vodama. U Hrvatskoj je u rijetkim istraživanjima školjkaša dokazana duž Jadranske obale prisutnost Vibrio spp., uz dominaciju vrste V. parahaemolyticus. Sve izraženija pojava globalnog zatopljenja podudara se i s neočekivanim pojavama infekcija s Vibrio spp. na sjeveru Europe, ponajprije na području Baltičkog mora. Četiri su važne značajke pripadnika roda Vibrio koje ih čine pogodnima u procjeni klimatskih promjena. To su: osjetljivost na temperaturu (jako dobro rastu pri temperaturama iznad 15 °C), brzo umnažanje, pojava u područjima gdje se nikada nisu javljali (kao što su sjeverna Europa, Aljaska, Čile, sjevero - istočno područje SAD) i mogućnost ulaska u organizam putem hrane i izlaganjem kontaminiranoj vodi. S gledišta sigurnosti hrane i školjkaša kao hrane, globalno zatopljenje i klimatske promjene imaju značajan utjecaj na dinamiku i rasprostranjenost potencijalno patogenih halofilnih vibrija na područja u kojima nisu povijesno dokazani kao uzročnici oboljenja ljudi. Posljednjih su desetljeća razvijane različite metode s ciljem pouzdanije identifikacije Vibrio vrsta. Radilo se na poboljšanju klasičnih mikrobioloških metoda, a većina je istraživanja usmjerena prema razvoju i primjeni molekularnih tehnika. Istraživanjima kojima su uspoređivani rezultati kulturelnih mikrobioloških metoda i lančane reakcije polimerazom (PCR), dokazana je značajno veća osjetljivost i pouzdanost molekularnih metoda u dokazu uzročnika.
This study investigates basic chemical composition and fatty acid profile of Mediterranean mussels (Mytillus galloprovincialis) cultivated in aquaculture facilities in six locations on the North ...Adriatic Istrian peninsula over an annual farming cycle. Mussels were sampled on a monthly basis. Mussels sampled during summer had a lower water content (p = 0.022), higher protein content (p = 0.006) and higher lipid content (p < 0.001), with the highest nutritional quality seen in samples from Limski Bay. Additionally, geographical location significantly influenced the fatty acid profile, in which dominated saturated fatty acids (SFA) followed by monounsaturated (MUFA) and polyunsaturated fatty acids (PUFA). The shares of SFA, MUFA and PUFA were highest in Savudrija Bay, Budava and Limski Bay, respectively. As compared to SFA and MUFA, the amount of PUFA varied in an opposite fashion, being higher during warmer months (p < 0.001). The major PUFA was eicosapentaenoic acid, whose share was significantly higher as compared to docosahexaenoic acid during the entire annual farming cycle at all sampling locations (p < 0.001). Based on the basic chemical composition and fatty acid profile, an optimal location and timing for harvesting Mediterranean mussels is Limski Bay during summer.
Fikotoksini predstavljaju skupinu prirodnih kemijskih kontaminanata u koje spadaju azaspiracidi (AZA), okadaična kiselina (OA) i njeni derivati, pektenotoksini (PTX), jesotoksini (YTX), ciklički ...imini (CI), domoična kiselina (DA), saksitoksini (STX), brevetoxini (BTX), ciguatoksini (CTX) i palitoksini (PlTX). Rezultati dosadašnjih istraživanja ukazuju na učestalu pojavnost fikotoksina u morskim organizmima, a poznato je da ove tvari mogu imati štetan učinak na okoliš i zdravlje ljudi, budući da su pored akutne toksičnosti pojedini ujedno i tumor promotori, odnosno da već i u malim koncentracijama pokazuju mutageno i toksično djelovanje u organizmu. Stoga su s ciljem zaštite zdravlja potrošača zakonodavstvom određene najveće dopuštene količine (NDK) fikotoksina u školjkaša, bodljikaša, plaštenjaka i morskih puževa (EZ 853/2004; EU 786/2013), iako neki od njih, poput cikličkih imina, još nisu uključeni u programe monitoringa, odnosno za njih legislativom nije određena NDK. U daljnjim istraživanjima, uz primjenu potvrdnih visokospecifičnih analitičkih metoda u ispitivanjima pojavnosti fikotoksina, potrebno je ispitati njihove učinke na osjetljiviji dio populacije te zbog mogućeg sinergičkog učinka u organizmu ustvrditi i supojavnost, kao i provesti procjenu izloženosti potrošača zbog prisustnosti ovih toksičnih tvari u različitim morskim organizmima namijenjenih konzumaciji. U ovom radu dan je pregled literaturnih spoznaja o svojstvima najznačajnijih predstavnika fikotoksina, njihovim toksičnim učincima u organizmu i pojavnosti u različitim vrstama morskih organizama.
Primjenom mikrobioloških i molekularnih tehnika, pretraženo je 328 uzoraka školjkaša dagnji (Mytilus galloprovincialis), uzgojenih na području istarskoga akvatorija. Rezultati ukazuju na smanjenu ...osjetljivost mikrobiološke tehnike (2.44 %) u odnosu na molekularnu dijagnostiku kojom je dokazana prisutnost bakterije V. parahaemolyticus u 21,34 % uzoraka. Izgledi za dokazivanje prisutnosti ove bakterije su 7,19 puta veći kada je površinska temperatura mora veća od 15 °C, dok su geni za kodiranje termostabilnog direktnog hemolizina (TDH) i hemolizina ovisnog o TDH (TRH) u uzorcima dokazani pri temperaturama mora iznad 22,5 °C, odnosno 24,7 °C. Udio TDH i TRH pozitivnih uzoraka iznosio je 10 % od ukupnih 70 V. parahaemolyticus-pozitivnih uzoraka. S obzirom na nedostatak zakonske obveze praćenja prisutnosti ove bakterije moguće je oboljevanje ljudi nakon konzumacije sirovih ili termički nedostatno obrađenih školjkaša.
Summary
Marine biotoxins are thermostable, mostly non‐protein molecules with different chemical components. According to solubility, they are divided into water‐soluble (hydrophilic) and fat‐soluble ...(lipophilic). The regulations of the EU and the Republic of Croatia regarding the maximum limits (ML) in live shellfish cover several groups: azaspiracids (AZA), okadaic group (okadaic acid and dinophysistoxins), yessotoxins (YTX), domoic and epi‐domoic acid (DA) and paralytic toxins (PSP).
Lipophilic toxins (LT) of the okadaic group and azaspiracids cause poisoning manifested by symptoms of the digestive system, such as nausea, vomiting, spasm and diarrhea. Symptoms appear as early as 30 minutes to several hours after consuming contaminated shellfish and last approximately for three days. Yessotoxins are lipophilic marine biotoxins that have not been proven to cause human poisoning, however, when applied to experimental animals intraperitoneally, they show a high toxic effect (EFSA, 2009a). The same applies to pectenotoxins (PTX) for which acute reference doses (ARfD) have been determined (EFSA, 2009b), but are no longer covered by EU and Croatian regulations related to ML.
Hydrophilic marine biotoxins (HT) are the neurotoxins domoic (DA) and epi‐domoic acid, and a group of paralytic marine biotoxins that can cause serious poisoning in humans, sometimes with a fatal outcome. DA poisoning causes gastrointestinal symptoms, but also neurological symptoms such as disorientation, confusion, behavioral changes, headache, convulsions and coma (Todd, 1993; Perl et al., 1990). In the most severe cases, muscle paralysis and suffocation occur, and death occurs due to suffocation due to respiratory paralysis and collapse of the cardiovascular system, usually within twelve hours (Prakash et al., 1971).
Marine biotoxins accumulate in shellfish, and as they are thermostable, the harmful effect on human health is manifested acutely, after consumption, in the case when they are present in shellfish in concentrations where they can have a harmful effect on health. Cases of human poisoning are often not reported to the competent health institutions, because although unpleasant, they mostly pass without the need for medical intervention. For this reason, one gets the impression that cases of poisoning by marine biotoxins are extremely rare.
In the Republic of Croatia, continuous monitoring of marine biotoxins in shellfish has been carried out for years. The following types of shellfish are sampled: mussels (Mytilus galloprovincialis), clams (Venerupis decussata), oysters (Ostrea edulis), queen scallops (Aequipecten opercularis), smooth clam (Callista chione) and warty venus (Venus verrucosa).
This scientific opinion represents the results of the monitoring carried out between 2017 and 2022 for the group of amnesic marine biotoxins (ASP), for the group of azaspiracids (AZA), for the group of marine biotoxins of okadaic acid (OA), for the group of paralytic of marine biotoxins (PSP) and for the group of yessotoxins (YTX).
In order to determine the actual exposure of consumers to marine biotoxins from shellfish, the calculation of average values for the UB, MB, LB scenario and for the highest concentration was used, which were combined with the highest intake of shellfish, which was calculated according to food consumption data in the Republic of Croatia that were collected in 2011/12 years. In addition, the exposure was assessed in the case of the assumption that shellfish contain the maximum legally permitted amount (ML), as well as the case with the corrected portion size due to the possible influence of heat treatment on the concentration of marine biotoxins, and the portion size of 400 g used by EFSA when preparing of its scientific opinions, in which it established current acute reference doses (ARfD) for each of the marine biotoxins monitored.
An exposure assessment for AZA was not made because this marine biotoxin was not detected in any sample during the monitored time in an amount above the limit of detection/quantification (LOD/LOQ). Therefore, it can be concluded that the risk for consumers in this case is negligible.
An exposure assessment for ASP in the assumed case of consumption of the proposed EFSA portion and in the case of a concentration equal to the NDK value, may lead to an intake of ASP that exceeds the ARfD value. ASP was quantified in some samples, but none had a concentration equal to or higher than ML. Therefore, the risk in this case can be considered negligible.
The exposure assessment for OA indicates the possibility of intake of OA greater than the ARfD in certain cases, the most certain of which is the case of the highest concentration and consumption of the largest portion in the Republic of Croatia for mussels, which means that there is a risk for consumers. The concentration for OA, in addition to being found above the LOD/LOQ, was also found to be above the ML in 2.22% of mussel samples, 0.62% of oyster samples and 0.15% of scallop samples.
The exposure assessment for PSP indicates the possibility of intake of PSP higher than the ARfD in certain cases, the most certain of which is the case of the highest concentration and consumption of the largest portion in the Republic of Croatia for mussels, which means that there is a risk for consumers. The concentration for PSP, in addition to being above the LOD/LOQ, was also above the ML in 0.51% of mussel samples and 0.31% of scallop samples.
The exposure assessment for YTX did not lead to YTX intake exceeding the ARfD value in any case. YTX was quantified in some samples, but none had a concentration equal to or greater than ML. Therefore, the risk in this case can be considered negligible.
When interpreting the results of this scientific opinion, it must be taken into account that the samples analyzed for marine biotoxins were collected directly from the shellfish breeding/harvesting areas for monitoring purposes. It's important to note that these shellfish weren't placed on the market, meaning consumers weren't exposed to marine biotoxins from these samples. This underscores the purpose and justification of the currently conducted extensive monitoring programme. Furthermore, recommendations for its continuation have been made, with certain improvements suggested.
Nakupljanje fikotoksina u plodovima mora može prouzročiti trovanja ljudi i znatne ekonomske gubitke u područjima koja se bave njihovim uzgojem. U cilju zaštite potrošača, zakonodavstvom su utvrđene ...najveće dopuštene količine (NDK) u dvoljušturnim školjkašima, kao i analitičke metode koje se primjenjuju u njihovom određivanju. Referentna metoda za određivanje lipofilnih fikotoksina ili lipofilnih toksina (LT) je tekućinska kromatografija u sprezi sa spektrometrijom masa (LC-MS/MS). Prilikom primjene ove metode, različite komponente matriksa mogu utjecati na ionizaciju analita i dovesti do pogrešne kvantifikacije. Cilj je ovog istraživanja bio ispitati
utjecaj matriksa dagnji, kamenica, kapica i mješčićnica na ionizaciju LT usporedbom nagiba kalibracijskih pravaca u matriksu i otapalu, a odnosi se na fikotoksine za koje su dostupne certificirane standardne otopine i za koje su Uredbom EU 853/2004 utvrđene NDK, uključujući pektenotoksin 2 (PTX2). Rezultati istraživanja ukazuju na to da su navedeni LT podložni utjecaju matriksa ukoliko se primjenjuje LC-MS/MS metoda za njihovo određivanje. Uočena je značajna supresija ionizacije većine analita u svim matriksima, osim u slučaju okadaične kiseline (OA) i dinofizistoksina 2 (DTX2) u školjkašima, kod kojih je uočeno značajno pojačanje ionizacije te dinofizistoksina 1 (DTX1) za kojeg nije uočen značajan utjecaj matriksa na ionizaciju u ekstraktu kamenica i kapica. Usporedbom nagiba kalibracijskog pravca u ekstraktu dagnji s onima u ekstraktu ostalih školjkaša nisu utvrđene značajne razlike, stoga se kalibracijski pravac u ekstraktu dagnji može primijeniti za kvantifikaciju LT i u ostalim vrstama obuhvaćenim ovim istraživanjem. Izuzetak su toksini OA skupine u mješčićnicama, za čiju je kvantifikaciju potrebno primijeniti kalibracijski pravac u ekstraktu navedene vrste. S obzirom na
opasnost koju fikotoksini predstavljaju za zdravlje ljudi, neophodno je neprestano razvijati pouzdane analitičke metode za njihovo određivanje, koje će doprinijeti kontroli kvalitete hrane i sigurnosti potrošača.
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