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PubMed2002

Growth of whaleworm (Anisakis simplex, Nematodes, Ascaridoidea, Anisakidae) third-stage larvae in paratenic fish hosts.

The growth of Anisakis simplex L3 larvae in the three paratenic hosts saithe ( Pollachius virens), cod (Gadus morhua) and redfish (Sebastes marinus) was studied based on material collected off the island of Vega on the west coast of Norway over a period of 1 year. In all three fish species, a positive correlation between the length of larvae and the age of the fish was shown. The number of large larvae, here defined as L3>28 mm, increases with increasing age of the host. These findings clearly indicate an accumulation of large L3 larvae and larval growth in the fish host. The larvae were additionally grouped according to the tissue--muscle, liver or other viscera--they were located in. When the average lengths of the three groups of larvae were compared, the gadoids saithe and cod had somewhat higher values for L3 larvae found in the liver. Redfish, on the other hand, which store fat to a relatively higher degree in the viscera, had the highest average length of larvae in this tissue. The lowest average values in all three species were found in the musculature. These trends suggest that the growth of A. simplex L3 larvae is positively correlated with the fat content of the tissue in which it is encapsulated. Of the three host species included in this study, saithe seemed to be the best suited for A. simplex L3, as the growth rate and average length of the parasite was greatest in this species. The growth and average length of L3 was least pronounced in redfish while cod held an intermediate position.

Age Distribution↗
PubMed2004

Growth, fecundity and sex ratio of adult whaleworm (Anisakis simplex; Nematoda, Ascaridoidea, Anisakidae) in three whale species from the North-East Atlantic.

The growth rate, fecundity, and sex ratio of Anisakis simplex in minke whales (Balaenoptera acutorostrata), porpoises (Phocoena phocoena) and long-finned pilot whales (Globicephala melas) was studied on the basis of material from the North Atlantic. A total of 8,135 mature A. simplex were collected from 24 minke whales, 11 porpoises and eight pilot whales. For both males and females, the prevalence was 100% for all three host species, with a mean intensity of 1,727, 262 and 139, respectively. The mean body length of adult female A. simplex was 126 mm in minke whales, 71 mm in the porpoises and 73 mm in pilot whales; and for males the averages were, respectively, 106 mm, 57 mm and 68 mm. Eggs from the uteri of 32 females of length 87-176 mm collected in minke whale stomachs were counted in a Fuchs-Rosenthal chamber. Total egg production was measured in 14 females cultivated at sea. The female growth period was estimated to be 30-60 days, and apparently all eggs were shed during the last week of life. A female of size 150 mm produces approximately 1.5 million eggs. In the cultivation experiment, about 85% of the total egg production was shed during the first 3 days after spawning started.

Animals↗
PubMed2004

Several allergens from Anisakis simplex are highly resistant to heat and pepsin treatments.

Ingestion of raw or undercooked fish can lead to infection with Anisakis simplex. Sensitized patients show specific IgE to proteins from this parasite. The aim of this study was to assess the frequency of specific IgE recognition directed to heat and/or pepsin-resistant allergens from A. simplex among sensitized patients. Twenty-seven patients with positive specific IgE and immunoblotting with a crude parasite extract were included in the study. Specific IgE detection against allergens resistant to boiling for 30 min and/or a pepsin digestion of an A. simplex extract was performed by immunoblotting. A total of 81% of the patients showed specific IgE to pepsin-resistant allergens and 67% had specific IgE to heat-resistant allergens. Thirty percent of patients recognized allergens after both treatments, one being the allergen detected by 75% of the patients of this group. Heat- and/or pepsin-resistant allergens from A. simplex could explain reactions and symptoms after the ingestion of well-cooked or canned fish.

Allergens↗
PubMed2004

The life cycle of Anisakis simplex in the Norwegian Deep (northern North Sea).

Copepoda (Calanus finmarchicus n = 1,722, Paraeuchaeta norvegica n = 1,955), Hyperiidae (n = 3,019), Euphausiacea (Meganyctiphanes norvegica n = 4,780), and the fishes Maurolicus muelleri (n = 500) and Pollachius virens (n = 33) were collected in the Norwegian Deep (northern North Sea) during summer 2001 to examine the importance of pelagic invertebrates and vertebrates as hosts of Anisakis simplex and their roles in the transfer of this nematode to its final hosts (Cetaceans). Third stage larvae (L3) of A. simplex were found in P. norvegica, M. muelleri and P. virens. The prevalence of A. simplex in dissected P. norvegica was 0.26%, with an intensity of 1. Prevalences in M. muelleri and P. virens were 49.6% and 100.0%, with mean intensities of 1.1-2.6 (total fish length >or=6.0-7.2) and 193.6, respectively. All specimens of C. finmarchicus and M. norvegica examined were free of anisakid nematode species and no other parasites were detected. P. norvegica, which harboured the third stage larvae, is the obligatory first intermediate host of A. simplex in the investigated area. Though there was no apparent development of larvae in M. muelleri, this fish can be considered as the obligatory second intermediate host of A. simplex in the Norwegian Deep. However, it is unlikely that the larva from P. norvegica can be successfully transmitted into the cetacean or pinniped final hosts, where they reach the adult stage. An additional growth phase and a second intermediate host is the next phase in the life cycle. Larger predators such as P. virens serve as paratenic hosts, accumulating the already infective stage from M. muelleri. The oceanic life cycle of A. simplex in the Norwegian Deep is very different in terms of hosts and proposed life cycle patterns of A. simplex from other regions, involving only a few intermediate hosts. In contrast to earlier suggestions, euphausiids have no importance at all for the successful transmission of A. simplex in the Norwegian Deep. This demonstrates that this nematode is able to select definite host species depending on the locality, apparently having a very low level of host specificity. This could explain the wide range of different hosts that have been recorded for this species, and can be seen as the reason for the success of this parasite in reaching its marine mammal final hosts in an oceanic environment.

Animals↗
PubMed2005

CO(2)-fixing enzymes during moulting from third larval to fourth larval stage of Anisakis simplex and Hysterothylacium aduncum (Nematoda: Anisakidae).

The fixing of CO(2) is an important metabolic process for many organisms. In the anisakid nematodes, CO(2) has been shown to be necessary for their development, at least in vitro. The presence of CO(2) stimulates the moulting (M3) of the larvae from the third (L3) to the fourth (L4) stage and prolongs the survival, at least, in vitro. We determined the activity of CO(2)-fixing enzymes, common to many organisms, in two anisakids: Anisakis simplex, a parasite of cetaceans, and Hysterothylacium aduncum, a parasite of fish. Although no activity was detected for pyruvate carboxylase or carboxylating-malic enzyme, we detected phosphoenolpyruvate carboxykinase (PEPCK) and phosphoenolpyruvate carboxylase (PEPC) activity. In A. simplex, PEPCK was clearly higher than that of PEPC throughout the moulting process studied. In H. aduncum, although the activity of both enzymes was of similar magnitude, they showed different behaviour; PEPCK activity decreased after the moulting to L4, PEPC activity increased so that the ratio PEPCK/PEPC activity decreased from 1.90 before moulting to 0.59 after.

Animals↗
PubMed2005

Isolation of a heat-resistant allergen from the fish parasite Anisakis simplex.

The thermal stability of allergenic peptides from the fish parasite Anisakis simplex has not been fully elucidated. This is of special relevance for physicians who should clearly indicate if sensitized patients should avoid ingestion of raw fish only or whether well-cooked fish should also be avoided, if allergenic peptides derived from the parasite remain immunologically detectable. An allergen was purified after heating a crude parasite extract for 30 min. The allergen was further purified by an ethanol fractionation procedure followed by a reversed-phase HPLC. The N-terminal amino acid sequence was obtained. This allergen was detected by 27% of sensitized subjects. The N-terminal amino acid sequence of the 9 kDa allergen showed no similarities to other known proteins. A minor low molecular weight allergen from A. simplex is highly resistant to heating and it could therefore have significant clinical relevance.

Allergens↗
PubMed2005

New locality records for third-stage larvae of Anisakis simplex (sensu lato) (Nematoda: Ascaridoidea) in euphausiids Euphausia pacifica and Thysanoessa raschii from Prince William Sound, Alaska.

Five euphausiid species (n=41,037) were collected in October 1997 from Prince William Sound, Alaska, and examined for parasites. New locality records are claimed for third-stage larvae (L3) of whaleworm Anisakis simplex (sensu lato) in two (0.019%) of 10,437 Thysanoessa raschii, and in one (0.013%) of 7,443 Euphausia pacifica. The L3 were about 19.5, 21.3 and 30.5 mm long. L3 were absent from 8,026 T. inermis, 10,302 T. longipes and 4,829 T. spinifera collected at the same time, and from 6,648 euphausiids representing the five species collected in July 1998. No individual harboured more than one parasite-a whaleworm L3, the ellobiopsid Thalassomyces fagei, a trypanorhynchid (tetrarhynchid) cestode (probably Nybelinia sp.), or a possible rhizocephalan. Controversy regarding the number of moults that occur in the egg of marine ascaridoids is discussed briefly.

Alaska↗
PubMed2006

Freezing infective-stage larvae from Anisakis simplex and their produce at -20 degrees C for 24 h does not prevent the occurrence of autonomic imbalance in rat ileum.

UNLABELLED: In the present report, we analyze the efficacy of the sanitary regulations to kill the Anisakis simplex larvae (As L3) (heat or freeze) to avoid the gastrointestinal alterations that it provokes. We studied the effects on intestinal contractility (muscular tone, amplitude, and frequency of the twitches and cholinergic and alpha-adrenergic stimulus) of As L3, their crude extracts (CE) and excretory-secretory products (ESP), untreated or heated (60 degrees C for 15 min) or frozen (-20 degrees C for 24 h) using rat ileum and an isometric system. Carbachol and noradrenaline have been used as cholinergic and alpha-adrenergic stimulus, respectively. We determined that viable As L3, their untreated CE and ESP, as well as all their frozen counterparts, altered the intestinal contractile activity and its autonomic control. In contrast, heated As L3, CE, and ESP were incapable of provoking any change in rat ileum motility, suggesting an inhibitory effect by the heating procedure. IN CONCLUSION: 1) The gastrointestinal alterations provoked by As are not necessarily associated with a prior infection; and 2) the storage of the seafood at -20 degrees C during 24 h does not prevent the intestinal autonomic imbalance provoked by As, whereas, the prior heating at 60 degrees C for 15 min may completely prevent such process.

Animals↗
PubMed2000

Distribution of Anisakis simplex in fish caught in the Ligurian Sea.

A total of 822 individuals of 5 fish species were sampled in the western coastal area of the Mediterranean Sea and examined for the presence of Anisakis simplex larvae in the body cavity, muscle, and viscera. A. simplex mean intensity (mI) and prevalence (Pr) values were very high in the common scad (Trachurus trachurus; Pr 80-100%, mI 19.3-36.8) and the blue scad (T. picturatus; Pr 80-100%, mI 18.2-70.7), moderate in the blue whiting (Micromesistius poutassou), and low in the red mullet (Mullus barbatus) and the striped mullet (M. surmuletus). The larval burden was strongly related to host length in the common and blue scad.

Animals↗
PubMed2001

Experimental infections of copepods and sticklebacks Gasterosteus aculeatus with small ensheathed and large third-stage larvae of Anisakis simplex (Nematoda, Ascaridoidea, Anisakidae).

Free-swimming ensheathed larvae of Anisakis simplex were shown experimentally to be ingested by the copepods Oitona similis and Acartia tonsa and by the nauplii of barnacles Balanus sp. The larvae did not grow in the copepod hemocoel. Experimental infections of various malacostracans were unsuccessful. Sticklebacks Gasterosteus aculeatus are naturally infected with larvae of A. simplex in coastal brackish water. Such sticklebacks may have acquired the infection by eating either a crustacean host or third-stage larvae (L3) from fish. Experimental infections of cod Gadus morhua with L3 from viscera of herring Clupea harengus showed that about one-third of the ingested larvae passed through the cod alimentary tract and were extruded whole but dead. Experiments showed that larvae from herring viscera survived and remained infective after at least 6 weeks in brackish water and seawater and that L3 from herring viscera were ingested by sticklebacks (and flounder Platichthys flesus), where they reencapsulated on the viscera; the L3 were alive 2 years later.

Animals↗
PubMed2001

Cellular immune responses in mice immunized with Anisakis simplex larval antigens.

Cellular immune responses to Anisakis simplex L3 antigens were investigated in BALB/c mice injected subcutaneously with a homologous crude extract (CE). Popliteal lymph nodes (PLN) were found to be increased in size and weight after A. simplex CE footpad injection. The effects of A. simplex CE in vitro proliferation were assayed with non-fractionated PLN cells or nylon-wool purified T cells derived from pooled lymph node cells of mice subcutaneously injected with CE. Spleen cells from immunized animals (antigen alone, or larva alone, or antigen plus larva) were studied by flow cytometry. The immunization induced a high proportion of CD4 + and TCRalphabeta + T cells. The number of B cells (CD45 + and TCRalphabeta-) in pre-immunized and infected mice was lower than that observed in animals subjected to infection only. The number of CD4+ T cells increased in the infected and in the pre-immunized and infected mice. In the latter, a decrease of CD8a + T cells was noted. The greatest increase in CD8a+ and TCRalphabeta- T cells was found in mice that had been subjected to infection only. Histological analysis showed that the most prominent lesions were gastric and intestinal in animals infected orally with one larva.

Animals↗
PubMed1996

Antigenic cross-reactivity in mice between third-stage larvae of Anisakis simplex and other nematodes.

We used ELISA and immunoblotting to investigate antigenic cross-reactivity in mice between third-stage larvae of Anisakis simplex and five other nematodes: the ascaridoids Ascaris suum, Toxocara canis and Hysterothylacium aduncum, and the nonascaridoids Trichinella spiralis and Trichuris muris. Two sera were raised against each species (including A. simplex, but excluding A. suum), by infection or by immunization with somatic antigens. Serum against A. suum was raised by immunization only. The reactivities of each serum with A. simplex somatic antigens (SA), excretion-secretion antigens (ES), pseudocoelomic fluid antigens (PF) and cuticular antigens (CA) were investigated. The results of ELISA indicated high antigenic cross-reactivity between A. simplex and the remaining ascaridoid nematodes, confirming that there is extensive antigenic similarity within this group of nematode parasites. Immunoblotting again confirmed the high degree of cross-reactivity between the SA of A. simplex and SAs of the other ascaridoids, although several A. simplex SA components in the 11-18 kDA range were only recognized by sera from mice infected with A. simplex. In addition, two A. simplex PF components of 22 and 27 kDA, were recognized only by sera from mice infected with, or immunized with the SA of, A. simplex. Finally, the anti-phosphorylcholine monoclonal antibody BH8 recognized only a small number of A. simplex antigens, indicating that phosphorylcholine epitopes are not significant contributors to the observed cross-reactivity with the other nematodes.

Animals↗
PubMed1997

Monoclonal antibodies against diagnostic Anisakis simplex antigens.

Five monoclonal antibodies (UA2, UA3, UA5, UA6, and UA8) specific for Anisakis simplex are described. All are IgG1/kappa monoclonal antibodies, except for UA2, which is an antibody IgM/kappa. The molecular weights of the major components recognized in immunoblotting are 48 and 67 kDa (UA2); 139 kDa (UA3 and UA5; same epitope); 35, 38, and 139 kDa (UA6); and 205 kDa (UA8). UA2 was the only monoclonal antibody to recognize both components of an excretion-secretion antigen preparation and antigens in the excretory cell and esophageal glands of third-stage A. simplex larvae; antigens in the excretory cell were also recognized by UA3 and UA6. Cross-reactivity studies using a hyperimmune polyclonal rabbit serum reacting with various ascaridoid nematodes indicated that the antigens captured by our monoclonal antibodies were specific for A. simplex. Finally, comparative studies of our monoclonal antibodies and An2 (the only monoclonal antibody currently available for serodiagnosis of human anisakiasis), based on the calculation of multiples of normal activity for human anisakiasis sera, indicated that our monoclonal antibodies (and particularly UA3) recognized antigens that are good candidates for serodiagnostic purposes.

Animals↗
PubMed1998

Distribution of whaleworm (Anisakis simplex, Nematoda, Ascaridoidea) L3 larvae in three species of marine fish; saithe (Pollachius virens (L.)), cod (Gadus morhua L.) and redfish (Sebastes marinus (L.)) from Norwegian waters.

The frequency distribution of Anisakis simplex L3 larvae between host tissues was investigated in three host species: saithe, cod and redfish. Fish were sampled from Norwegian coastal waters and examined for the presence of A. simplex in muscle and viscera. In all three of the host species, A. simplex larvae were most frequently detected in the viscera; the percentages of total infection for saithe, cod and redfish were 99.6%, 97.8% and 88.0%, respectively. In general, the distribution patterns of A. simplex L3 between muscle and viscera were not significantly affected by host size. The observations that distributions vary between species and are not affected by host size do not support an earlier hypothesis which states that A. simplex L3 distributions are determined by an optimal pre-encapsulation migratory distance within host tissues. In contrast, it is suggested that A. simplex L3 distributions are governed by the conditions encountered within host tissues, and are possibly related to the availability of nutrients.

Animals↗
PubMed1999

Analysis of the antigenicity in mice of biotinyl enzymes from Anisakis simplex and other nematodes.

We analyzed the antigenicity of biotinyl enzymes contained in somatic extracts from Anisakis simplex and other parasite nematodes and show in this report that these molecules are an important source of cross-reactivity problems among these nematodes. Cross-reactivity was most pronounced among members of the superfamily Ascaridoidea. These results suggest that the presence of biotinyl enzymes in whole somatic extracts of A. simplex and other parasites may make serodiagnostic assays based on this kind of antigenic preparation unreliable.

Animals↗
PubMed1996

In vitro study on the effect of larval excretory/secretory products and crude extracts from Anisakis simplex on blood coagulation.

The anticoagulant action of Anisakis simplex larvae on human blood in vitro was examined. Anticoagulant activity was assessed by routine screening tests that evaluate the overall competency of the coagulant mechanism. A slight prolongation of the prothrombin time (PT) was observed with the larval crude extracts. Prolongation of the PT was seen at a concentration of excretory/secretory (ES) products greater than 62.5 micrograms/ml. No prolongation of the activated partial thromboplastin time (PTT) was observed using crude extracts. There was a prolongation of the PTT with ES products at concentrations greater than 62.5 micrograms/ml. ES products of the larvae were able to prolong coagulation times indicating that they contain an inhibitory or anticoagulant property. Preparation of crude extracts of A. simplex showed only minimal anticoagulant activity. The results obtained by measurements of the PT and the PTT suggest a probable alteration of one of the coagulation proteins namely factors Xa, IIa or Va. These findings suggest that the anticoagulant activity demonstrated in the ES products may play an important role during invasion of the gastric or intestinal mucosa by larvae and could have biological significance in infected patients.

Animals↗
PubMed2006

Sensitization to Anisakis simplex s.l. in a healthy population.

The main objective of this study was to determine the degree of sensitization to Anisakis simplex s.l. in healthy population. Using the determination of specific IgE, we studied the seroprevalence against A. simplex s.l. in blood donors selected at random in the region of Antequera (Southern Spain). We detected 22.1%. The immunoblotting technique revealed a band of approximately 60 kDa in the serum of individuals who also exhibited high values of specific IgE against A. simplex s.l. This band would be useful for diagnosis.

Adolescent↗
PubMed2005

Efficacy of albendazole against Anisakis simplex larvae in vitro.

BACKGROUND: The growing popularity in western countries of eating uncooked seafood has resulted in an increased incidence of anisakidosis. AIM: To study the in vitro activity of different concentrations of albendazole against Anisakis simplex larvae under different media pH. METHODS: A. simplex larvae were obtained from fresh hakes acquired from the fish market of Madrid. They were divided into groups and placed in culture dishes (15 larvae each) containing RPMI-1640, in the presence or absence of different concentrations of albendazole (300, 400 and 500 microg/mL). RESULTS: Albendazole dose-dependently reduced the survival of the larvae, its maximum activity being at 500 microg/mL when it killed almost all larvae at 48 h. Acidic medium pH significantly reduced the efficacy of albendazole. CONCLUSION: Albendazole is effective in killing A. simplex larvae at different pH in vitro, suggesting that this molecule could be useful in treating clinical manifestations of human anisakidosis.

Albendazole↗