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As a key component of cell-cultured fish, fish skin gelatin (FSG)-based cell scaffold provides support structures for cell growth, proliferation, and differentiation. However, there are potential allergenicity risks contained in FSG-based scaffolds. In this study, 3D edible scaffolds were prepared by phase separation method and showed a contact angle of less than 90°, which indicated that the scaffolds were favorable for cell adhesion. Besides, the swelling ratio was greater than 200%, implying a great potential to support cell growth. The sequence homology analysis indicated that FSG was prone to cross-reaction with collagen analogues. Additionally, a food allergic model was constructed and represented that mice gavaged with cod FSG exhibited higher levels of specific antibodies, mast cell degranulation, vascular permeability, and intestinal barrier impairment than those gavaged with pangasius and tilapias FSG. Its higher allergenicity might be attributed to a higher number of digestion-resistant linear epitopes. Moreover, the higher hydrolysis degree linked to the exposure of linear epitopes to promote the combination with IgE, which was also responsible for maintaining the higher allergenicity of cod FSG. This study clarifies allergenic risks in cell-cultured fish and further study will focus on the allergenicity reduction of FSG-based cell scaffolds.

Background: IgE reactivity to fish allergens in atopic dogs, which are used as models for food allergy, has not been elucidated to date. We investigated IgE reactivity to crude extracts and purified allergens derived from the Pacific cod (Gadus macrocephalus) in atopic dogs to identify the allergenic proteins of cod., Results: The levels of specific IgE to crude cod extracts were measured in the sera of 179 atopic dogs, including 27 dogs with cod allergy, using enzyme-linked immunosorbent assay (ELISA). Specific IgE to crude cod extracts were present in 36 (20%) of the 179 atopic dogs and in 12 (44%) of the 27 dogs with cod allergy. The allergens in crude cod extracts were analyzed by ELISA, immunoblotting, and liquid chromatography-tandem mass spectrometry. In allergen component analysis, IgE reactivity to tropomyosin and enolase was observed in the sera of dogs with cod allergy. IgE reactivity to parvalbumin, collagen, and tropomyosin was evaluated using the sera of atopic dogs that tested positive for specific IgE to crude cod extracts. Among the 36 dogs with IgE reactivity to crude cod extracts, 9 (25%), 14 (39%), and 18 (50%) dogs tested positive for specific IgE to parvalbumin, collagen, and tropomyosin, respectively., Conclusions: The IgE reactivity to cod allergens observed in dogs was similar to that in humans, and this finding further supports the use of atopic dogs with fish allergy as a model for fish allergy in humans.

The aim of this work was to evaluate some biological properties of hake head oil (HHO) as well its lipid composition. The fatty acid profiles showed a dominance of unsaturated fatty acids overtaking 55% of the total fatty acids. Omega-3 polyunsaturated fatty acid profiles exhibited a dominance of EPA (eicosapentaenoic acid) (3.96%) and DHA (docosahexaenoic acid) (25.39%). The antioxidant activity was determined through two different assays: DPPH scavenging activity and β-carotene bleaching by linoleic acid assay. Eighteen mice were excised on their back and divided into 3 groups, treated with sterile saline, commercial healing cream and HHO, respectively. The wound closure rate, the hydroxyproline contents and the histopathology evolution in skin tissue were elaborated. Also, the anti-inflammatory activity was studied by carrageenan-induced mouse paw edema. Mice were divided into 3 groups treated respectively with sterile saline, anti inflammatory drug reference and HHO. The anti-inflammatory evaluation of HHO in mice exhibited an important inhibition of carrageenan-induced hind paws edema, as confirmed by the histological analysis, the superoxide dismutase (SOD), catalase (CAT) and malondialdehyde (MDA) level. HHO displayed a significant wound healing effect probably due to the anti-inflammatory and antioxidant activities of its EPA and DHA contents. The overall results proved that HHO might be favorable drugs who exert a great therapeutic potential wound healing and anti-inflammatory effects in animal model., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest., (Copyright © 2020 Tissue Viability Society. Published by Elsevier Ltd. All rights reserved.)

Background: Fish are one of eight major allergens defined in the US Food Allergen Labeling and Consumer Protection Act, and cod and pollock are two of the major fish allergens. This paper describes development and validation of a method to detect cod and pollock in complex food matrices using real-time polymerase chain reaction (PCR). Mitochondrial cytochrome oxidase I (COI) sequences obtained through DNA barcoding were used to design a single set of primers and probe which detected three species in the genus Gadus: Atlantic cod, Pacific cod, and walleye pollock., Results: Cod spiked into three different food matrices (cooking oil, clam chowder, and hushpuppy mix) yielded high linearity, dynamic range spanning six orders of magnitude, and lower limits of detection at 1-10 ppm (ppm; mg kg -1 ). Frying had an adverse effect on the lower limit of detection, but not on linearity., Conclusions: This work shows that COI DNA barcoding sequences can be used to effectively design real-time PCR assays for detection of food allergens in complex matrices. While full-length DNA barcodes distinguish individual species, the PCR assay designed here detected three different species. This is likely because real-time PCR assays are tolerant to basepair mismatches and do not utilize the full length of the DNA barcode. Published 2018. This article is a U.S. Government work and is in the public domain in the USA., (Published 2018. This article is a U.S. Government work and is in the public domain in the USA.)

Interleukin 1b (IL-1b) is a member of the cytokine family that serves as major mediators for early pro-inflammatory responses. During artificial breeding of Pacific cod (Gadus macrocephalus), outbreaks of NNV in the early development stages could lead to high mortality. Recent research reported that IL-1b participated in the host-virus interaction against NNV infection, however, no IL-1b gene has been identified from Pacific cod so far. In this study, we described the identification and characterization of a IL-1b from Pacific cod transcriptome at both transcript and protein level. Sequence alignment and phylogenetic analysis were conducted to determine and confirm their evolutionary relationship. Expression analysis revealed that IL-1b was expressed in all tested tissues and the western blot analysis confirmed the same expression profiles using purified recombinant IL-1b protein. In addition, significant up-regulation was detected in IL-1b transcripts during early development stages at 30dph and 50dph. After challenged with poly I:C, IL-1b was significantly induced after 24 h. Taken together, these results strongly suggested that IL-1b might play an essential role in innate defense mechanism of Pacific cod., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: Anaphylaxis and immediate-type fish roe allergies have been reported worldwide, and, in Japan, fish roe is the sixth most common food allergen. No oral food challenges (OFCs) have used pollock roe (PR), which is reported to have high cross-reactivity with salmon roe (SR). Therefore, we administered an OFC using cooked PR to evaluate PR- and SR-specific immunoglobulin E (IgE) levels and allergic reactions in patients with PR sensitivity., Methods: This retrospective study evaluating patient characteristics and responses to OFCs was conducted with 10-20 g of cooked PR, between April 2006 and November 2016., Results: We assessed 51 patients (median age: 6.8 years). All had PR sensitization, 6 (12%) with a history of immediate reactions to PR, and 18 (35%) of immediate reactions to SR. Median PR-specific and SR-specific IgE values were 3.4 kUA/L and 9.9 kUA/L, respectively. Seven patients (14%) had a positive OFC. There was no anaphylaxis. Induced symptoms were mild and included localized urticaria, throat pruritus, intermittent cough, and mild abdominal pain. We treated one patient with mild abdominal pain with oral antihistamines. There were no significant differences in history of immediate reaction to PR and PR-specific IgE titers between OFC-positive and OFC-negative patients, although significant differences were found for PR-specific IgE titers adjusted for SR-specific IgE (p = 0.025) and PR-specific IgE/SR-specific IgE ratio (p = 0.009)., Conclusions: Increased PR-specific IgE/SR-specific IgE ratio or PR-specific IgE levels adjusted for SR-specific IgE levels were risk factors for OFC positivity., (Copyright © 2017 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

Multiple functions of caspases include normal cell turnover, proper development and function of the immune system, hormone-dependent atrophy, embryonic development and chemical-induced cell injury. During artificial propagation of Pacific cod, Gadus macrocephalus, high mortality occurred during early development stages. Here, we performed various analyses on the cDNA and protein sequences of six different G. macrocephalus caspases namely GmCasp3, 6, 7, 8, 9 and 10, and tried to investigate the contributions of caspase family to the development and Nervous Necrosis Virus (NNV) resistance. Sequence analysis of GmCaspase proteins showed that each caspase shared conserved domains like "HG", "QACXG (X for R, G or Q)" and "GSWF" except GmCasp10. Sequence alignment and phylogenetic tree showed that GmCasp8 and GmCasp10 were quite different from those of other fishes. 3-D models indicated that structure of GmCasp3 is very conservative, but GmCasp6, 7, 8, 9 and 10 are less conservative. Tissue distribution analysis showed that six Gmcaspases mRNA transcripts were detected in tissues of intestine, gill, thymus, head-kidney and spleen with different abundance, but Gmcasp7 were not detected in the brain. GmCasp3 transcript was kept at very low level in the early development stages, while the expression levels of GmCasp6, 7, 8, 10 were different at various development stages. GmCasp8 level seemed to be much higher than other caspases in the heads of 65dph and 75dph juveniles. To understand the role of caspases during NNV outbreak, modulation in expression of each Gmcaspases were investigated. The results showed that GmCasp3 transcript level increased significantly when NNV broke out, while GmCasp7, 8, 9 and 10 in cod heads decreased obviously at 69dph and 77dph. The results suggest that caspases in Pacific cod should be diverse in their structure and function, and their unique features and response to NNV outbreak add more evidences for the specificity of immune system in Pacific cod., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: The relationship between anaphylaxis and cardiovascular events has been reported in the past. While skin and respiratory symptoms are usually the most common and the first to appear, cardiovascular complications play a key role and represent the leading cause of death in anaphylaxis., Methods: We report 3 episodes of atrial fibrillation triggered by anaphylaxis. Allergy and cardiology studies were performed. In both patients, the etiological agent was identified: Anisakis simplex hypersensitivity and food allergy., Results: The heart is the source and target of chemical mediators released during an allergic reaction. In the heart, there are plenty of mast cells, and they are predominantly located around the coronary adventitia and in close contact with small vessels in the muscle wall. The release of mediators can influence ventricular function, heart rate, and coronary artery tone. Anaphylaxis can trigger any kind of arrhythmia. In these cases, the very interesting point of discussion was: which should be first, treating anaphylaxis or cardiac events? The other controversial point was the use of epinephrine, the first line of treatment for anaphylaxis. Recommendations about epinephrine in cardiac patients during an anaphylactic event are still a major dilemma., Conclusions: We emphasize the importance of the priority of establishing protocols between cardiologist and allergist in treatment of cardiac complications during anaphylaxis, and we warn about the correct diagnosis of arrhythmias in anaphylaxis in order to treat them as soon as possible, to prevent other consequences and complications., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Interferon regulatory factor 7 (IRF7) plays an important role in regulating the response of type I interferon (IFN) to viral infection. To understand the mechanisms underlying immune reactions in the Pacific cod, Gadus macrocephalus, the gene encoding G. macrocephalus IRF7 was cloned and characterized. The cDNA of G. macrocephalus IRF7 was also cloned and sequenced. A cDNA sequence of 2032 bp was assembled using polymerase chain reaction (PCR) products. It contains an open reading frame of 1323 bp in length, which encoded a 440-amino acid polypeptide that comprised a DNA-binding domain (DBD), an IRF association domain (IAD), and a serine-rich domain (SRD). In the DBD, the tryptophan cluster consisted of only four tryptophans, which is a unique characteristic in fish IRF7. The mRNA of IRF7 was detected in various tissues, including in the spleen, thymus, kidney, intestine, and gills, using relative quantification PCR (R-qPCR). Dynamic expression of IRF7 was observed in larvae throughout post-hatching (ph) development, with the highest level detected at day of ph (dph) 25. Response to immune stimulation was examined by challenging larvae with polyriboinosinic polyribocytidylic acid (pIC) to mimic viral infection and elicit an immune reaction. R-qPCR revealed that the expression of IRF7 significantly increased in pIC-treated groups relative to that in the control groups, in a time-dependent manner, with peak responses at 48 and 72 h after pIC-treatment. These results show that IRF7 is expressed in various tissues of adult fish and larvae and is sensitive to viral infection, suggesting that it plays a role in antiviral immune defense in G. macrocephalus., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Mortality (>90%) is a big concern in larval rearing facilities of Pacific cod, Gadus macrocephalus, limiting its culture presently still in the experimental stages. Understanding the immune system development of G. macrocephalus is crucial to optimize the aquaculture of this species, to improve the use of economic resources and to avoid abuse of antibiotics. For the transcriptome analysis, using an Illumina sequencing platform, 61,775,698 raw reads were acquired. After a de novo assembly, 77,561 unigenes were obtained. We have classified functionally these transcripts by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). 27 genes mainly related to hematopoietic or lymphoid organ development and somatic diversification of immune receptors have been reported for the first time in Pacific cod, and 14 Ig heavy chain (μ chain) locuses were assembled using Trinity. Based on our previous achievement, we have chosen Rag1 and Igμ as immune system development biomarkers. Full length cDNA of Rag1 and Igμ as biomarkers were obtained respectively using RACE PCR. Concerning Rag1, the deduced amino acid of Rag1 and protein immunodetection revealed a Rag1 isoform of 69 kDa, significantly different from other fish orthologs, such as Oncorhynchus mykiss (121 kDa). Phylogenetic analysis reveals a unique immune system for the Gadus genre, not exclusive for Atlantic cod, among vertebrates. Meanwhile, full length cDNA of Igμ included an ORF of 1710 bp and the deduced amino acid was composed of a leader peptide, a variable domain, CH1, CH2, Hinge, CH3, CH4 and C-terminus, which was in accordance with most teleost. Absolute quantification PCR revealed that significant expression of Rag1 appeared earlier than Igμ, 61 and 95 dph compared to 95 dph, respectively. Here we report the first transcriptomic analysis of G. macrocephalus as the starting point for genetic research on immune system development towards improving the Pacific cod aquaculture., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Genes encoding the major histocompatibility complex (MHC) have been thought to play a vital role in the adaptive immune system in all vertebrates. The discovery that Atlantic cod (Gadus morhua) has lost important components of the MHC II pathway, accompanied by an unusually high number of MHC I genes, shed new light on the evolution and plasticity of the immune system of teleosts as well as in higher vertebrates. The overall aim of this study was to further investigate the highly expanded repertoire of MHC I genes using a cDNA approach to obtain sequence information of both the binding domains and the sorting signaling potential in the cytoplasmic tail. Here we report a novel combination of two endosomal sorting motifs, one tyrosine-based associated with exogenous peptide presentation by cross-presenting MHCI molecules, and one dileucine-based associated with normal MHC II functionality. The two signal motifs were identified in the cytoplasmic tail in a subset of the genes. This indicates that these genes have evolved MHC II-like functionality, allowing a more versatile use of MHC I through cross-presentation. Such an alternative immune strategy may have arisen through adaptive radiation and acquisition of new gene function as a response to changes in the habitat of its ancestral lineage.

The content of lysozyme as an indicator of nonspecific immunity in kidney, liver, spleen, and blood serum in 8 fish species of the order Gadiformes and the order Pleuronectiformes was analyzed. The Gadiformes are characterized by a very low level of enzyme or the absence of its activity in individual organs and serum, regardless of the fish habitat location. The Pleuronectiformes demonstrated diversity in content and distribution of lysozyme in the body of various fish species. Two plaice species with the unique distribution inhabiting the Sea of Okhotsk were found. This feature is due, first, to the highest lysozyme content in serum of the previously studied fish; second, to the 2-3-fold excess of the enzyme content in serum as compared with the studied immunocompetent organs. The obtained results confirm the opinion about the existence of various related groups among fishes which essentially differ in the lysozyme role in nonspecific immunity.

A multibiomarker approach was developed on juvenile Atlantic tomcod (Microgadus tomcod) to evaluate the pertinence of this approach for low-cost screening assessment of the environmental quality of various coastal sites within estuaries. Several biometric indices and biomarkers (ethoxyresorufin-O-deethylase (EROD) activity, metallothionein concentration, and immune responses) were investigated on immature and maturing tomcods (≤ 31 months) collected in four environmentally contrasted sites in the St. Lawrence Estuary (SLE). Simultaneous examination of various age classes provides the opportunity to detect short-term responses in sensitive young-of-the-year fish (e.g., EROD induction) and longer-time effects associated with chronic exposure and bioaccumulation (e.g., metallothionein induction). Principal component analysis was helpful to discriminate between responses possibly related to contaminant exposure (EROD, metallothionein) and responses that could be affected by upstream-downstream gradient (immune response, biometric indices). Measurement of a battery of biomarkers in young tomcods at several sites along the shore of the SLE is a low-cost screening investigation useful to identify hot spots requiring further investigation with chemical analysis and additional reference sites.

The occurrence of lysozyme and immunoglobulin (Ig) in semen of different teleost species (brown trout-Salmo trutta, perch-Perca fluviatilis, burbot-Lota lota) was studied. In all investigated species lysozyme activities (1.13-1.45 U ml(-1)) and Ig concentrations (T-Ig: 1.11-1.61 microg ml(-1), IgG [measured only in brown trout]: 1.49 microg ml(-1)) were detected in seminal plasma. Ig was also found in spermatozoa (T-Ig: 0.234-0.357 microg/g protein, IgG: 0.198 microg ml(-1)) while spermatozoal lysozyme activities were low and fluctuating (0.093-0.164 U/g protein). In Salmo trutta lysozyme activities and immunoglobulin levels were compared between semen samples with high and low sperm motility as motility is an indicator for sperm fertility. Lysozyme activities were higher in seminal plasma of samples with high motility than in those with low motility while seminal plasma and spermatozoal immunoglobulin concentrations (T-Ig, IgG) were increased in samples with low motility in comparison to samples with high motility. Seminal plasma and spermatozoal IgG concentrations and seminal plasma lysozyme activities showed significant correlations with the sperm motility rate and swimming velocity. Moreover, lysozyme improved the viability of spermatozoa in in vitro experiments. Possible physiological meanings of these results are discussed., (Copyright 2010 Elsevier Inc. All rights reserved.)

In this study, susceptibility and potential carrier status of burbot, Lota lota, were assessed for five important fish pathogens. Burbot demonstrated susceptibility and elevated mortality following challenge with infectious haematopoietic necrosis virus (IHNV) by immersion and to Aeromonas salmonicida by intraperitoneal (i.p.) injection. IHNV persisted in fish for at least 28 days, whereas A. salmonicida was not re-isolated beyond 17 days post-challenge. In contrast, burbot appeared refractory to Flavobacterium psychrophilum following intramuscular (i.m.) injection and to infectious pancreatic necrosis virus (IPNV) by immersion. However, i.p injection of IPNV resulted in re-isolation of virus from fish for the duration of the 28 day challenge. Renibacterium salmoninarum appeared to induce an asymptomatic carrier state in burbot following i.p. injection, but overt manifestation of disease was not apparent. Viable bacteria persisted in fish for at least 41 days, and bacterial DNA isolated by diagnostic polymerase chain reaction was detected from burbot kidney tissue 90 days after initial exposure. This study is the first to investigate susceptibility of burbot to selected fish pathogens, and this information will aid in efforts to culture and manage this species.

This paper reports the cloning and sequencing of interleukin (IL)-22 in two gadoid fish, cod (Gadus morhua) and haddock (Melanogrammus aeglefinus). The complete transcript of this gene was 1002 and 1154 bp respectively, of which 492 bp was the open reading frame (ORF) in both genes. High amino acid identity (88.3%) was found between these genes but was less than 50% aa identity to other known genes. The gene organisation of haddock IL-22 consisted of five exons and four introns, as with other IL-10 family members. Expression studies showed that IL-22 is constitutively expressed in gill, with low level expression also observed in gut, gonad and head kidney. In a vaccination experiment, haddock were injected intraperitoneally with formalin-killed Vibrio anguillarum or with PBS, and 2 months later challenged by immersion in 10(7)cfu/ml V. anguillarum for 30 min. Head kidney and gill samples were collected prior to challenge and 24, 48 and 72 h post-challenge (hpc) for Real-time PCR analysis of IL-22 expression. No significant changes in IL-22 expression were observed in head kidney tissue but vaccinated fish showed a significantly increased expression of IL-22 24 hpc in gill and no mortalities were seen in these fish. In contrast, control fish, which started to succumb to the disease from 72 hpc, showed no significant increase in gill expression after challenge.

Allergens and allergoids can be characterized by means of physicochemical methods, resulting in a description of the protein on different structural levels. Several techniques are available and their suitability depends on the composition of the particular sample. Current European legislation on allergen products demands characterization of final products in particular focusing on identity, degree of modification (for allergoids) and stability of the composition. Structural parameters of allergens may be used to investigate the stability of an allergen product. The challenge is to identify and optimize techniques that allow determination of protein structure in the context of a formulated pharmaceutical product. As the majority of the products currently marketed are formulated with aluminium hydroxide or aluminium phosphate as a depot, most of the methods and techniques used for protein characterization in solution are not applicable. An additional hurdle is that allergen products are based on natural extracts, comprising a mixture of proteins, both allergens and non-allergens, sometimes in the presence of other uncharacterized components from the raw material. This paper describes which methods are suitable for the different stages of allergen product manufacturing, and how these relate to the current regulatory requirements. Some of the techniques are demonstrated using a model allergen, cod parvalbumin, and a chemically modified form thereof. We conclude that a variety of methods is available for characterization of proteins in solution, and that a limited number of techniques appear to be suitable for modified allergens (allergoids). Adaptation of existing methods, e.g. mass spectroscopy and infrared spectroscopy may be helpful to obtain protein parameters of allergens in a formulated allergen product. By choosing a combination of techniques, including those additional to physicochemical approaches, relevant parameters of allergens in formulated allergen products can be assessed in order to achieve a well-characterized pharmaceutical product.

Allergic reaction following fish consumption can trigger life-threatening reactions in predisposed individuals. Parvalbumins from different species have been identified as the major fish allergens. There are two distinct phylogenetic lineages of parvalbumins, alpha and beta. Most allergic reactions are caused by beta-parvalbumins. We cloned and expressed cDNAs encoding cod (Gadus morhua) and carp (Cyprinus carpio) beta-parvalbumins and purified natural cod beta-parvalbumin. CD spectra of the purified proteins showed that their overall secondary structure contents were very similar. No differences in thermal stability were monitored in the calcium-bound or calcium-depleted form of natural cod parvalbumin. IgE reactivity was assessed using 26 sera of fish allergic patients from Spain, The Netherlands, and Greece in immunoblot and ELISA experiments. Twenty-five of the 26 patients with IgE reactivity to native and recombinant cod parvalbumin also reacted to the recombinant carp parvalbumin. IgE inhibition assays were performed using cod and carp extracts and purified recombinant parvalbumin of cod and carp. High crossreactivity among cod and carp parvalbumins was observed in immunoblots as well as in fluid phase assays. Natural and recombinant parvalbumins gave comparable results when performing various in vitro diagnostic assays.

Vibrio anguillarum is one of the primary pathogens responsible for high levels of fish mortality in the aquaculture industry, and among gadoids O2a and b are the most common pathogenic serotypes. In this paper a variety of studies were performed to assess the optimal route by which to challenge haddock against this pathogen, and an optimal regime to vaccinate haddock. The most efficient method to challenge haddock with V. anguillarum in this study was immersion in a bath containing 10(7)cfu/ml, where 60% mortality was seen. Subsequent experiments showed that juvenile haddock could be protected against bacterial challenge with V. anguillarum, with a significant reduction in mortalities observed amongst the vaccination treatments when compared to the unvaccinated controls. However, as seen previously in cod studies, vaccination did not induce a specific antibody response.

The full sequence of the heavy chain of Immunoglobulin M (IgM) and a partial fragment of the recombination activating gene-1 (RAG-1) gene were sequenced in haddock, Melanogrammus aeglefinus, L. The complete transcript of haddock IgM consisted of 1865 bp and translated into a 572-aa peptide. The RAG-1 fragment was 1776 bp and was identified as the core region of RAG-1. These two immune genes were used in expression studies as markers of early development in haddock larvae. A DIG labelled oligoprobe of the RAG-1 gene was used in whole-mount in situ hybridisation (WISH). A hybridisation signal for RAG-1 was first detected in larvae at 25 days post-hatching (dph) in two bilateral symmetric regions of the head identified as the thymus. Further expression studies were carried out by RT-PCR analysis of RAG-1 and IgM on larval samples obtained during early development, i.e. from fertilisation to weaning. Haddock RAG-1 expression was detected after 21 days post-fertilisation (dpf) whilst IgM transcripts were not detected until 40 dpf, equivalent to day 29 post-hatching. These results suggest that the immune system in haddock starts to develop in larvae of 6-7 mm in length (25-29 dph).

Purpose: To clarify the positivity of specific IgE to salmon and cod roe in outpatients., Methods: Specific IgE were assayed using CAP RAST system in 91 pediatric outpatients. They were 47 males and 44 females, including 22 allergic, 29 infectious, 10 neurological, 8 gastrointestinal, 7 urological, 6 hematologic, 3 metabolic disease and 6 other disorders. For control, 30 sera from healthy normal adult volunteers were assayed. Additionally, sera from 653 allergic patients were also collected in our laboratory. Specific IgEs against salmon and cod meat were also assayed simultaneously., Results: In 91 pediatric patients, two children were salmon roe positive and one child was cod roe positive. Three children scored class 1, borderline positive in salmon roe, and one child scored class 1 in cod roe. Other children were negative in all allergens. No positive sera were found in normal adult volunteers. Among 653 specimens in our laboratory, the positivity of specific IgE to salmon and cod roe were 25%, and 9%, respectively. Infants younger than two years old had higher ratio than older children. There was a significant correlation (r = 0.676) between the titers of IgE to salmon and cod roe. On the other hand, the titers of IgE to their meats correlated less than those to their roes. Our results support previous reports that fish roe from different species have common antigen apart from those of fish meat., Conclusion: Positive ratio of salmon and cod roe specific IgE were 2.2 and 1.1% in pediatric outpatients.

Introduction: Food protein-induced enterocolitis syndrome (FPIES) occurs in infants. Typical symptoms include profuse vomiting and/or diarrhea several hours after ingestion of a given food. The disorder is a non-IgE mediated food hypersensitivity. The most frequently involved foods are milk and soy, but some cases of FPIES induced by solid foods have been described. We report 14 patients with FPIES due to fish protein., Material and Methods: History and physical examination, skin prick test (SPT) with fish allergens and Anisakis simplex, prick-by-prick test with implicated fish and determination of specific IgE antibodies against fish were performed. In eight children atopy patch test (APT) were also performed. In nine patients an open oral food challenge with the implicated fish was carried out., Results: There were six boys and eight girls, aged from 9 to 12 months at diagnosis, with between two and six reactions to the offending fish proteins before the diagnosis was established. Four patients had a previous history of atopy. Presenting symptoms included diarrhea in two patients, profuse vomiting in six patients, and recurrent vomiting and subsequent diarrhea in three patients. In addition to these symptoms, associated septic appearance, apathy and lethargy were present in the remaining three patients. Onset of symptoms occurred a few minutes after fish ingestion in two patients and from 60 minutes to 6 hours in the 12 remaining patients. SPT to fish were negative in all patients. Serum food-specific IgE antibodies were negative in all patients except one. APT was positive in three patients. Open oral challenge (OC) was performed in nine infants and was positive in all. The patients were followed-up for between 1 and 7 years after diagnosis, and follow-up OC tests were performed after fish had been eliminated from the patients' diet for 3-4 years. Four patients became clinically tolerant to the causal food. Three patients currently tolerate only one type of fish (swordfish)., Conclusions: We report 14 patients with FPIES caused by fish protein. The symptoms suggest a form of cell-mediated, non-IgE mediated food hypersensitivity. The gold standard for diagnosis is OC, although caution should be exercised in infants with several reactions or a recent diagnosis. After a period of elimination of the causal food from the diet, tolerance can develop.

Increased fish consumption has led to frequent reporting of fish allergy and adverse reactions. Alaska pollack (Theragra chalcogramma) is a globally important commercial fish species, belonging to the Gadidae family. This family of fish also includes cod whose parvalbumin, Allergen M (Gad c 1), has been thoroughly studied and considered as a reference to sensitization in fish allergy. In the present study, parvalbumin from Alaska pollack, designated The c 1, was purified by use of anion exchange chromatography. To demonstrate the homogeneity of the purified protein, reverse phase high performance liquid chromatography was performed and showed two distinct fractions which had similar IgG and IgE binding capacities. Accordingly, cDNA cloning revealed two isotypic parvalbumin transcripts in pollack muscle. Recombinant parvalbumins of pollack exhibited low IgG and IgE binding capacities, in contrast to the native counterparts, which were almost as potent as cod Gad c 1. The allergenicity of The c 1 was assayed by ELISA inhibition, and compared to cod, the concentration required for obtaining 50% ELISA inhibition (C 50%) was only 18% higher for The c 1.