Search

Ciguatera Fish Poisoning (CFP) is primarily caused by consumption of tropical and sub-tropical fish contaminated by Ciguatoxins (CTXs). These lipid-soluble, polyether neurotoxins are produced by dinoflagellates in the genera Gambierdiscus and Fukuyoa. While there is no regulatory level in Europe for CTXs, the European Food Safety Authority (EFSA) adopted the United States guidance level of 0.01 μg P-CTX1B eq.kg−1 of fish. This limit is extremely low and requires significant improvement in the detection of CTXs. In this study, we compared analytical protocols based on liquid chromatography coupled to tandem low or high resolution mass spectrometry (LC-LRMS or HRMS) to find the best conditions for sensitivity and/or selectivity. Different approaches such as LC conditions, ion choice and acquisition modes, were evaluated to detect the Pacific-ciguatoxins (P-CTXs) on a triple quadrupole (API4000 Qtrap, Sciex) or a quadrupole time of flight (QTOF 6550, Agilent Technologies) spectrometer. Moreover, matrix effects were calculated using matrix-matched calibration solutions of P-CTX1B and P-CTX3C prepared in purified fish extract. Subsequently, the method performance was assessed on naturally contaminated samples of seafood and phytoplankton. With LRMS, the ammoniated adduct ion used as a precursor ion showed an advantage for selectivity through confirmatory transitions, without affecting signal-to-noise ratios, and hence limits of detection (LODs). As also reported by some studies in the literature, methanol-based mobile phase gave better selectivity and sensitivity for the detection of P-CTXs. While the LOD for P-CTX1B and P-CTX3C met the EFSA recommendation level when using LRMS, the findings suggested careful evaluation of instrumental parameters for determination of CTXs. LODs were significantly higher for HRMS, which currently results in the need for a significantly higher sample intake. Nevertheless, HRMS allowed for the identification of artefacts and may allow for improved confirmation of the identity of P-CTXs analogues. Consequently, LRMS and HRMS are considered complementary to ensure adequate quantitation and identification of P-CTXs.

Ciguatera fish poisoning (CFP) is a form of food poisoning caused by consumption of varieties of toxic ciguatera fish species from tropical and subtropical waters. In the present review, significant features and perspective of LC/MS methods for CTXs as confirmatory methods are reviewed on the discussion for one of the best LC/MS/MS method reported by Yogi in 2011. More practical approach on the basis of the previous method is proposed with a single LC/MS by using selected ion monitoring (SIM). High resolution mass spectrometry (HRMS) coupled with LC is also proven as a reliable approach for identification of CTX analogues even in the situation that analytical standards are not available.

Species in the epi-benthic dinoflagellate genus Gambierdiscus produce ciguatoxins (CTXs) and maitotoxins (MTXs), which are among the most potent marine toxins known. Consumption of fish contaminated with sufficient quantities of CTXs causes Ciguatera Fish Poisoning (CFP), the largest cause of non-bacterial food poisoning worldwide. Maitotoxins, which can be found in the digestive system of fish, could also contribute to CFP if such tissues are consumed. Recently, an increasing number of Gambierdiscus species have been identified; yet, little is known about the variation in toxicity among Gambierdiscus strains or species. This study is the first assessment of relative CTX- and MTX-toxicity of Gambierdiscus species from areas as widespread as the North-Eastern Atlantic Ocean, Pacific Ocean and the Mediterranean Sea. A total of 13 strains were screened: (i) seven Pacific strains of G. australes , G. balechii , G. caribaeus , G. carpenteri , G. pacificus , G. scabrosus and one strain of an undetermined species ( Gambierdiscus sp. Viet Nam), (ii) five strains from the North-Eastern Atlantic Ocean (two G. australes , a single G. excentricus and two G. silvae strains), and (iii) one G. carolinianus strain from the Mediterranean Sea. Cell pellets of Gambierdiscus were extracted with methanol and the crude extracts partitioned into a CTX-containing dichloromethane fraction and a MTX-containing aqueous methanol fraction. CTX-toxicity was estimated using the neuro-2a cytoxicity assay, and MTX-toxicity via a human erythrocyte lysis assay. Different species were grouped into different ratios of CTX- and MTX-toxicity, however, the ratio was not related to the geographical origin of species (Atlantic, Mediterranean, Pacific). All strains showed MTX-toxicity, ranging from 1.5 to 86 pg MTX equivalents (eq) cell −1 . All but one of the strains showed relatively low CTX-toxicity ranging from 0.6 to 50 fg CTX3C eq cell −1 . The exception was the highly toxic G. excentricus strain from the Canary Islands, which produced 1426 fg CTX3C eq cell −1 . As was true for CTX, the highest MTX-toxicity was also found in G. excentricus . Thus, the present study confirmed that at least one species from the Atlantic Ocean demonstrates similar toxicity as the most toxic strains from the Pacific, even if the metabolites in fish have so far been shown to be more toxic in the Pacific Ocean.

A major challenge in the international fish trade remains to reliably discriminate hundreds of fish species from possible substitutes which are perhaps not recommended for consumption. Recurring instances of ciguatera fish poisoning in the EU were associated with the consumption of mislabeled imported tropical fishes, mostly the red snapper Lutjanus malabaricus. Because the underlying fish species substitutions were only brought to the public attention due to resulting severe disease symptoms of this foodborne illness, we suggested a significant dark figure of undetected species substitutions with associated health risks. For the first time, the species identity of 975 individual fish imported into the EU at Frankfurt Airport from various provenances was correlated with their potential risk to cause ciguatera intoxication over a three-year period. Based on DNA sequence analysis and supporting peer-review of specimen photographs an overall mislabeling rate of 31% was determined which varied massively between 0% and 100% across fish families. Even though the sampling was not strictly random, this suggests massive fish species substitutions already at early stages of the value chain in the EU and over long time periods. From a health risk perspective, the occurrence of ciguatera-prone species in the cohort of DNA-identified substituted fish was dramatically higher compared to the correctly labeled fish (46% vs. 17%). Since our sampling largely reflects standardized official import procedures on fish labeling, these numbers suggest that the import of mislabeled tropical fishes, in particular snappers (Lutjanidae) and groupers (Serranidae), poses an underestimated health risk for seafood consumers in Europe.

Graphical abstract, Highlights • P-CTX-1B in 84 Spanish Mackerel from NSW and QLD were quantified using LC–MS/MS. • Liver & flesh from 6 fish and liver from 4 other fish were positive for P-CTX-1B. • Liver had significantly higher concentration (∼6 times) of P-CTX-1B than flesh. • No relationship was observed between length/weight of fish & detection of P-CTX-1B., Ciguatera Fish Poisoning (CFP) is a tropical disease caused by the consumption of fish contaminated with ciguatoxins (CTXs). Currently, the only feasible prevention methods for CFP are to avoid the consumption of fish of certain species from some regions, avoid larger fish of certain species, or avoid all fish caught from specific regions. Here, we quantified levels of P-CTX-1B in Spanish Mackerel (Scomberomorus commerson), which is the main fish species that causes CFP in New South Wales and Queensland, Australia, using LC–MS detection against a toxin standard. We found detectable P-CTX-1B in both flesh and liver tissues in fish from New South Wales (n = 71, 1.4% prevalence rate, with a confidence interval of 1%–4%, and 7% prevalence, 1%–12%, in flesh and liver, respectively). In the small sample of fish from Queensland, there was a 46% prevalence (19–73%, n = 13). Toxin levels found were 0.13 μg kg−1 to

© 2016 Elsevier B.V. Ciguatera fish poisoning (CFP) has been reported for many years in Rarotonga, Cook Islands, and has had the world's highest reported incidence of this illness for the last 20 years. Following intensive sampling to understand the distribution of the causative organisms of CFP, an undescribed Gambierdiscus species was isolated from the Rarotongan lagoon. Gambierdiscus cheloniae sp. nov. has the common Gambierdiscus Kofoidian plate formula (except for a variability in the number of precingular plates in aberrant cells): Po, 3′, 6″ (7″), 6C?, 6 or 7S, 5′″, 1p and 2″″. The 2′ plate is hatchet shaped and the dorsal end of 1p is pointed and the relatively narrow 1p plate. Morphologically G. cheloniae is similar to the genetically closely related species G. pacificus, G. toxicus and G. belizeanus, although smaller (depth and length) than G. toxicus. The apical pore plate varies from those of G. belizeanus and G. pacificus, which are shorter and narrower, and from G. toxicus, which is larger. G. cheloniae also differs from G. pacificus in the shape of the 2′ plate. The description of this new species is supported by phylogenetic analyses using three different gene regions. G. cheloniae produced the putative maitotoxin-3 analogue, MTX-3, but neither maitotoxin or monitored ciguatoxin. Extracts of G. cheloniae were shown to be highly toxic to mice by intraperitoneal (i.p.) injection, although they were less toxic by gavage. It is possible that this species produces toxins other than putative MTX-3.

International audience; Ciguatera Fish Poisoning (CFP) is a non-bacterial seafood poisoning well characterized in the remote archipelagos of French Polynesia, yet poorly documented in the Society archipelago, most notably on Moorea, the second most populated island in French Polynesia, which counts a high proportion of fishermen fishing on a regular basis. To address this knowledge gap, a holistic study of the ciguatera issue was conducted on Moorea. First, ciguatera risk was analysed in terms of incidence rate, fish species most commonly involved and risk stratification in Moorea lagoon based on 2007–2013 epidemiological data. A mean incidence rate of 8 cases per 10,000 inhabitants for the study period and an average under-reporting rate of 54% were found. Taking into account hospitalization and medication fees, and loss of productive days, the health-related costs due to CFP were estimated to be USD $1613 and $749 for each reported and unreported case, respectively, with an overall cost of USD $241,847 for the study period. Comparison of the present status of CFP on Moorea with a risk map established in the late 1970’s showed that the spatial distribution of the risk has stayed relatively stable in time, with the north shore of the island remaining the most prone to ciguatera. Evaluation of the current knowledge on CFP among different populations groups, i.e. fishermen, residents and visitors, was also conducted through direct and indirect interviews. About half of the fishermen interviewed were actually able to identify risky fishing areas. While, overall, the CFP risk perception in the fishing community of Moorea seemed accurate, although not scientifically complete, it was sufficient for the safe practice of their fishing activities. This may be due in part to adaptive responses adopted by 36% of the fishermen interviewed, such as the avoidance of either high-risk fishing sites or toxic species. At the residents and visitors’ level, the study points out a striking lack of awareness of the CFP issue among visitors, as compared to local residents. Indeed, less than 25% of Moorea visitors vs. an average of 98% in residents were aware of CFP or of its presence on the island. Interestingly, evaluation of the fish consumption preferences showed that 70% of visitors do not consume lagoon fish during their stay, not for fear of CFP, but mainly due to the lack of availability of these species in recreational facilities or because they have nutritional preference for pelagic fish. This lack of awareness, along with the report by several CFP patients of the consumption of fish species yet banned for sale, stress the need for improved communication efforts on this critical issue among both residents and visitors on Moorea. The implementation of a public outreach strategy is proposed, based on both existing information networks and low-cost communication actions through information displays at various strategic locations, e.g. Tahiti-Faa’a international airport, the ferry boat station, recreational facilities, as well as the major trading points on Moorea Island.

Ciguatera fish poisoning (ciguatera) is a common clinical syndrome in areas where there is dependence on tropical reef fish for food. A subset of patients develops recurrent and, in some instances, chronic symptoms, which may result in substantial disability. To identify possible biomarkers for recurrent/chronic disease, and to explore correlations with immune gene expression, peripheral blood leukocyte gene expression in 10 ciguatera patients (7 recurrent and 3 acute) from the U.S. Virgin Islands, and 5 unexposed Florida controls were evaluated. Significant differences in gene expression were noted when comparing ciguatera patients and controls; however, it was not possible to differentiate between patients with acute and recurrent disease, possibly due to the small sample sizes involved.

International audience; Ciguatera Fish Poisoning (CFP) is a foodborne disease classically related to the consumption of tropical coral reef fishes contaminated with ciguatoxins (CTXs), neurotoxins produced by dinoflagellates of the Gambierdiscus genus. Severe atypical ciguatera-like incidents involving giant,clams, a marine resource highly consumed in the South Pacific, are also frequently reported in many Pacific Islands Countries and Territories. The present study was designed to assess the ability of giant clams to accumulate CTXs in their tissues and highlight the potential health risks associated with their consumption. Since giant clams are likely to be exposed to both free-swimming Gambierdiscus cells and dissolved CTXs in natural environment, ex situ contamination experiments were conducted as follows: giant clams were exposed to live or lyzed cells of TB92, a highly toxic strain of G. polynesiensis containing 5.83 +/- 0.85 pg P-CTX-3C equiv. cell(-1) vs. HIT0, a weakly toxic strain of G. toxicus containing only (2.05 +/- 1.16) x 10(-3) pg P-CTX-3C equiv. cell-1, administered over a 48 h period at a concentration of 150 cells mL(-1). The presence of CTXs in giant clams tissues was further assessed using the mouse neuroblastoma cell-based assay (CBA-N2a). Results showed that giant clams exposed to either lyzed or live cells of TB92 were able to bioaccumulate CTXs at concentrations well above the safety limit recommended for human consumption, i.e. 3.28 +/- 137 and 2.92 +/- 1.03 ng P-CTX-3C equiv. g(-1) flesh (wet weight), respectively, which represented approximately 3% of the total toxin load administered to the animals. In contrast, giant clams exposed to live or lyzed cells of HIT0 were found to be free of toxins, suggesting that in the nature, the risk of contamination of these bivalves is established only in the presence of highly toxic blooms of Gambierdiscus. Liquid chromatography-mass spectrometry (LC-MS/MS) analyses confirmed CBA-N2a results and also revealed that P-CTX-3B was the major CTX congener retained in the tissues of giant clams fed with TB92 cells. To the best of our knowledge, this study is the first to provide evidence of the bioaccumulation of Gambierdiscus CTX.s in giant clams and confirms that these bivalve molluscs can actually constitute another pathway in ciguatera poisonings. While most monitoring programs currently focus on fish toxicity, these findings stress the importance of a concomitant surveillance of these marine invertebrates in applicable locations for an accurate assessment of ciguatera risk.

The isolation and culturing of toxic Gambierdiscus species is desirable for obtaining reliable sources of toxins responsible for Ciguatera Fish Poisoning (CFP) in order to provide reference materials and purified standards for chemical analyses and bioassays. Gambierdiscus polynesiensis CAWD212, G. pacificus CAWD213 and G. australes CAWD149 and 216 have been isolated from samples of sea water from the Cook Islands. G. polynesiensis produced ciguatoxins (CTX; 18.2 pg per cell) and G. australes CAWD149 produced maitotoxin (MTX; 8.3 pg per cell) and in both extracts putative MTX-3 was detected. G. pacificus also produced putative MTX-3. The toxicities of extracts of G. polynesiensis and G. pacificus were similar by i.p. injection (LD50s of 1.0 mg/kg and 0.8 mg/kg respectively), but when administered by gavage, the MTX-3 analogue producing G. pacificus (LD50 251 mg/kg) was 30-times less toxic than the CTX-producing G. polynesiensis (LD50 7.9 mg/kg). Consumption of fish contaminated by CFP has caused illness and even death in the Cook Islands and throughout the Pacific region and is now considered a globally neglected disease. Large-scale cultures of these organisms are now available, and isolation of the toxins they produce is in progress.

Ciguatera, an ichtyosarcotoxism linked to the consumption of usually healthy coral fish is a common poisoning in the Pacific, Caribbean and Indian Ocean where it is endemic. However, increased tourism and commercial transportation of tropical fish for consumption make it an unexceptional intoxication in countries away from its endemic area. Environmental stresses such as climate changes also contribute to the expansion of its geographical area. The non-specific clinical symptomatology is characterized by the occurrence of gastrointestinal, cardiovascular, nervous and general signs few hours after eating a ciguatoxic fish. The diagnosis is clinical and relatively easy in endemic areas but much less for physicians who are rarely confronted with, which is a source of prolonged diagnostic delays and a significant increase in spending. Treatment of ciguatera is symptomatic but new treatments, still experimental, give a real hope for the future.

The present study used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to investigate retrospectively ciguatoxin (CTX)-positive samples as determined by mouse bioassay (MBA) in the past 10 years in Hong Kong. The results showed that Pacific CTXs (P-CTX-1, -2 and -3) were the most commonly observed toxins found in the samples, indicating Pacific Ocean areas as the most important origin of ciguatera fish poisoning. Clinical diagnosis from ciguatera patients also revealed the predominance of neurological illnesses in most cases, supporting intoxication of Pacific origin. This study demonstrated the ability of laboratory analysis to identify and quantify Pacific CTXs in suspected fish samples, so as to support the clinical diagnosis of ciguatera. Comparative analysis (Student's t-test and Spearman's rank correlation analysis) on the two CTX detection methods showed approximate linearity for overall P-CTXs (P-CTX-1, -2 and -3)/P-CTX-1 alone as derived by LC-MS/MS and total toxicity levels (P-CTX-1 equivalent) as determined by MBA. The LC-MS/MS method coupled with the rapid extraction method could allow the detection of trace amount of CTXs at levels below the clinically relevant limit, 0.1 ppb P-CTX-1 in fish flesh. For practical application, the adoption of a two-tiered approach for testing, chemical analysis by LC-MS/MS for toxic fish screening, coupled with biological assay by MBA for final toxicity confirmation, was proposed for first-line screening of CTX in potentially contaminated fish samples in the market, with an aim to minimizing the use of laboratory mice and at the same time providing reasonably effective means for routine analysis.

A cooked meal remnant and uncooked portion of a Caribbean barracuda suspected in ciguatera fish poisoning were examined for the presence of ciguatoxins (CTX). Samples were analysed using a tiered method of CTX analysis consisting of in vitro cell (N2a) assay to assess composite toxicity and liquid chromatography-tandem mass spectrometry (LC–MS/MS) for structural confirmation. Meal remnant and uncooked fish extracts were cytotoxic by N2a cell assay and Caribbean ciguatoxin congener C-CTX-1 was structurally confirmed. Sample extracts were fractionated by LC and fractions analysed by the cell assay. The cytotoxicity profiles of cooked meal remnant and uncooked fish were similar. Cytotoxicity-guided LC–MS/MS analyses identified several CTX congeners contributing to the composite toxicity of the samples. C-CTX-1 was a major contributor, supporting its utility as a biomarker of Caribbean ciguatoxic fish.

To review the clinical features and laboratory investigations of ciguatera patients in Hong Kong between 2004 and 2007 in order to show the timely sampling of implicated fish from ciguatera victims and application of validated mouse bioassay for confirming suspected clinical cases of ciguatera.Diagnosis of the ciguatera victims was based on history of coral fish consumption and clinical presentations stated in official guidelines for clinical diagnosis of ciguatera fish poisoning in Hong Kong. Food remnants of coral fish samples were collected swiftly from ciguatera victims between 2004 and 2007 for ciguatoxins (CTXs) analysis.Major clinical symptoms in ciguatera patients included gastrointestinal and neurological effects including limb numbness and diarrhoea, which developed at 0.5 to 15 hours after consumption of fish. In most cases, neurological symptoms were more common than gastrointestinal symptoms. A broad range of attack rate (10%-100%) was observed in each ciguatera outbreak. Validated mouse bioassay on ether extracts of the food remnant samples confirmed that all were CTXs-positive (0.5 - 4.3 MU/20 mg ether extract) and directly linked to the corresponding ciguatera cases.Consistency between clinical and laboratory analysis for ciguatera poisoning illustrates the application of laboratory mouse bioassay in a timely fashion for confirming ciguatera poisoning cases and implementing effective public health measures. With further improvement in laboratory techniques, features of ciguatera fish poisoning cases can be better defined. Further studies are needed to determine the risk of each class of CTXs (Pacific-, Indian- and Caribbean-CTXs) in Hong Kong.

From 2001 to 2005, numerous cases of seafood poisonings were reported in a tribe from Lifou (Loyalty Islands Province, New Caledonia) of which 35 were thoroughly examined. Observations outlined by the epidemiological and clinical data (including severity and rapid onset of certain symptoms following consumption of either giant clams ( Tridacna spp.) or grazing and molluscivorous fish together with the apparent inefficacy of traditional remedies, were not in favour of a classical Ciguatera Fish Poisoning (CFP) outbreak. From 2005 onwards, an environmental offshore survey of the affected area was conducted. Screening of the damaged coral area revealed the presence of large populations of cyanobacteria identified as Hydrocoleum Kutzing, but the absence of Gambierdiscus spp., the well-known dinoflagellate causative agent of CFP. In vivo and in vitro toxicological studies of extracts obtained from cyanobacteria and giant clams, strongly suggested the co-occurrence of ciguatoxin-like, anatoxin-like and paralytic shellfish toxins in these samples. These new findings shed new light on the complexity of the CFP symptomatology and treatment and also on the diversity and origin of the CFP toxins. Furthermore they provide new evidence of the overall variability of seafood poisonings following the ingestion of different sea products living in a marine environment where significant harmful populations of microalgae and cyanobacteria coexist. This is the first report on the involvement of cyanobacteria in CFP-like outbreaks following the consumption of giant clams or fish specimens. Consequently, it is recommended that CFP risk assessment programs now include monitoring of cyanobacteria besides the obvious screening of CFP-promoting dinoflagellates.

Ciguatera fish poisoning is the most frequently reported seafood-toxin illness in the world, caused by the consumption of coral reef fishes contaminated with a group of natural toxins produced by minute phytoplankton (dinoflagellates). These toxins are potent and cause both acute and chronic neurologic disease in humans. Although ciguatera fish poisoning is associated with established neurotoxins in animal models, and with known peripheral nervous system effects in humans, this pilot study was the first to explore possible central nervous system effects associated with chronic ciguatera poisoning in humans. In a matched cohort design, 12 cases and 12 their age and gender matched friend controls underwent a battery of neuropsychological tests. The results indicated that although their scores on objective neuropsychological tests did not differ significantly from matched controls, persons with ciguatera reported a higher degree of toxicity-related symptomatology and endorsed significantly more depressive symptomatology than controls. Future research should explore larger numbers of persons with both acute and chronic ciguatera, with and without mannitol treatment.

W. R. Lange . Severity rating scales for ciguatera fish poisoning. Toxicon 31, 777–781, 1993.—Severity of ciguatera fish poisoning is often quite variable. Two symptom check list rating scales were developed for quantifying illness severity and for selectively monitoring response to therapy in patients with chronic toxicity. Content validity was ascertained, and internal consistency reliability was demonstrated by means of the Cronbach alpha correlation coefficient (alpha = 0.9475). It was concluded that these instruments were valid and reliable, and that they conveniently and accurately recorded illness severity and treatment efficacy. They should prove useful in clinical settings and epidemiologic investigations.

The structure of a new ciguatoxin congener, CTX3C (1). isolated from cultures of a dinoflagellate Gambierdiscus toxicus was elucidated mainly o

The known and theoretical pharmacology of poisons involved in toxic marine ingestions has assisted in the development of specific therapeutics for these afflictions. The clinical manifestations of several toxic marine ingestions have suggested toxins/cogeners that may be involved in the poisoning process, providing direction for the development of diagnostic laboratory tests, including those for cyclic ethers. Future investigations should involve utilization of acetaminophen and indomethacin for chronic ciguatera fish poisoning, and the evaluation of the role of polycyclic ethers in the temperature sensation reversal phenomenon of both ciguatera and neurotropic shellfish poisoning.