Molecular identification of Atlantic goliath grouper Epinephelus itajara (Lichtenstein, 1822) (Perciformes: Epinephelidae) and related commercial species applying multiplex PCR

The Atlantic goliath grouper, Epinephelus itajara , is a critically endangered species, threatened by illegal fishing and the destruction of its habitats. A number of other closely related grouper species found in the western Atlantic are also fished intensively. While some countries apply rigorous legislation, illegal harvesting followed by the falsification of fish products, which impedes the correct identification of the species, is a common practice, allowing the catch to be marketed as a different grouper species. In this case, molecular techniques represent an important tool for the monitoring and regulation of fishery practices, and are essential for the forensic identification of a number of different species. In the present study, species-specific primers were developed for the Cytochrome Oxidase subunit I gene, which were applied in a multiplex PCR for the simultaneous identification of nine different species of Epinephelidae: Epinephelus itajara , E. quinquefasciatus , E. morio , Hyporthodus flavolimbatus , H. niveatus , Mycteroperca acutirostris , M. bonaci , M. marginata , and M. microlepis . Multiplex PCR is a rapid, reliable and cost-effective procedure for the identification of commercially-valuable endangered fish species, and may represent a valuable tool for the regulation and sustainable management of fishery resources.(AU)

O mero, Epinephelus itajara , encontra-se criticamente ameaçado, resultado da pesca ilegal e destruição dos habitas. Filogeneticamente relacionadas a este táxon encontram-se garoupas que atualmente são intensamente pescadas no Atlântico Oeste. Apesar de leis mais restritivas aplicadas em alguns países, a captura ilegal com a descaracterização morfológica é uma prática comum que impossibilita a identificação correta da espécie permitindo que seja comercializada como garoupas, badejos ou chernes. A este respeito, técnicas moleculares representam ferramentas importantes para o monitoramento e fiscalização da pesca, provando ser essencial, na identificação forense de diversas espécies. Primers espécie-específicos foram desenvolvidos com base no gene Citocromo Oxidase subunidade I que aplicados em PCR-Multiplex possibilitam a identificação simultânea de nove espécies Epinephelidae: Epinephelus itajara , E. quinquefasciatus , E. morio , Hyporthodus flavolimbatus , H. niveatus , Mycteroperca acutirostris , M. bonaci , M. marginata e M. microlepis . A identificação via PCR multiplex de espécies de peixes ameaçadas e comercialmente importantes é um método rápido, prático, seguro e de baixo custo, que poderá ser útil o controle do uso e manejo sustentável de recursos pesqueiros.(AU)

IgE Sensitization Patterns to Commonly Consumed Foods Determined by Skin Prick Test in Korean Adults

Offending food allergens can vary with regional preferences in food consumption. In this study, we analysed sensitization rates to commonly consumed foods in Korean adults suspected of having food allergy. One hundred and thirty four subjects underwent a skin prick test (SPT) with 55 food allergens, of which 13 were made by our laboratory and the rest were commercially purchased. Of the 134 patients, 73 (54.5%) were sensitized to one or more food allergens. Sensitization to chrysalis was detected most frequently, at a rate of 25.4%. Sensitization rates to other food allergens were as follows: maize grain (13.4%), shrimp (11.9%), almond (11.1%), wheat flour (8.2%), lobster (8.2%), buckwheat (8.2%), mackerel (5.2%), pollack (5.2%), halibut (4.5%), peanut (4.5%), anchovy (4.4%), squid (3.7%), saury (3.0%), common eel (3.0%), yellow corvina (3.0%), hairtail (2.2%), octopus (2.2%), and others. In addition to well-known food allergens, sensitivity to mackerel, chrysalis, pollack, and halibut, which are popular foods in Korea, was observed at high rates in Korean adults. We suggest that the SPT panel for food allergy in Korea should include these allergens.

Identification of the major allergens of Indian scad (Decapterus russelli).

The purpose of this study was to characterize major allergens of Indian scad (Decapterus russelli) which is among the most commonly consumed fish in Malaysia. Raw and cooked extracts of the fish were prepared. Protein profiles and IgE binding patterns were produced by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting using sera from subjects with fish allergy. The major allergens of the fish were then identified by two-dimensional electrophoresis (2-DE), followed by mass spectrometry of the peptide digests. The SDS-PAGE of the raw extract revealed 27 protein fractions over a wide molecular weight range, while the cooked extract demonstrated only six protein fractions. The 1-DE immunoblotting detected 14 IgE-binding proteins, with a molecular weight range from 90 to < 6.5 kDa. Three protein fractions with molecular weights of approximately 51, 46 and 12 kDa were identified as the major allergens of this fish. The approximately 12 kDa band was a heat-resistant protein while the approximately 51 and 46 kDa proteins were sensitive to heat. The 2-DE gel profile of the raw extract demonstrated > 100 distinct protein spots and immunoblotting detected at least 10 different major IgE reactive spots with molecular masses as expected and isoelectric point (pI) values ranging from 4.0 to 7.0. A comparison of the major allergenic spot sequences of the 12 kDa proteins with known protein sequences in databases revealed extensive similarity with fish parvalbumin. In conclusion, this study demonstrated that a parvalbumin which is similar to Gad c 1 is the major allergen of Indian scad. Interestingly, we also detected heat-sensitive proteins as major allergenic components in our fish allergy patients.

Role of pepsin in modifying the allergenicity of bhetki (Lates calcarifer) and mackerel (Rastrelliger kanagurta) fish.

The effect of pepsin digestion on the allergenicity of raw and thermally processed (boiled and fried) fish muscle extracts of two widely consumed fishes bhetki (Lates calcarifer) and mackerel (Rastrelliger kanagurta) was studied. Sere were collected from 110 patients who were hypersensitive to fish, as evidenced by their clinical history, symptoms and positive skin-prick test results. The various extracts after digestion with pepsin at different times of incubation were tested for specific IgE-binding activity by ELISA and immunoblotting with patients' sera. All the extracts of both the fishes retained their allergenicity as evidenced by ELISA and immunoblotting. In bhetki, maximum allergenicity was found in the pepsin-digested fried extract, whereas similar treatment decreased the allergenicity in fried mackerel. Results showed that raw as well as thermally processed allergens of both the fishes maintained strong allergenicity, even after digestion with pepsin for different time periods. The study revealed that the fish proteins played an important role in manifestation of allergy, due to their stable structure, which was retained even after pepsin and heat treatment.