Experimental studies on the migration of Anisakis sp. larvae (Nematoda: ascaridida) into the flesh of herring, Clupea harengus L.

In two experiments, a batch of 38 herring from Shetland was gutted immediately on capture (0 h) and further batches were gutted 14 and 37 h after capture and storage on ice. The numbers of Anisakis larvae in the body-cavity and in the flesh (detected by pepsin-HCl digest) were then counted. The proportion of fish with worms in the flesh was higher at 14 and 37 h than at 0 h. The percentage of the total worm burden in the flesh increased between cash each time interval, which suggests that a large-scale migration of larvae into the flesh occurs in ungutted fish. There was a positive association between the numbers of larvae in the viscera and the numbers in the flesh. Most larvae occurred in the hypaxial muscles. The results are discussed in relation to the observations of earlier workers.     

Larval anisakids and other helminths in the hake, Merluccius gayi (Guichenot) from Chile

One hundred and twenty hake Merluccius gayi caught off central Chile were examined for helminth parasites. Sixty hake were kept in ice from the catch of a trawler were examined upon arrival at port, while the other 60 hake were dissected on board immediately after collection. Ninety one per cent of hake were infected with one or more species of the following helminths: trypanorhynch plerocerci of Grillotia dollfusi Carvajal, 1971; adult worms of the genus Clestobothrium; larval nematodes of the genera Anisakis and Phocanema; 86% of the total were infected with Anisakis sp. and 42.5% with Phocanema sp. Infection with larval nematodes increased with length of the fish and there was a greater incidence of infection among the females. There was a greater frequency of occurrence of the parasites in the viscera than in the musculature.     

Commercial blast-freezing of third-stage Anisakis simplex larvae encapsulated in salmon and rockfish

Sixty-four fish were blast-frozen to -35 C for 15 hr to determine the effects of commercial blast-freezing on the viability of third-stage larvae of Anisakis simplex encapsulated in the muscle and viscera of sockeye salmon (Oncorhynchus nerka) and canary rockfish (Sebastes pinniger). Parallel tests were conducted on larval nematodes in 16 whole (round) salmon, 16 dressed salmon (heads and viscera removed), and 32 whole (round) rockfish. After blast-freezing, 4 in-the-round salmon, 4 dressed salmon, and 8 in-the-round rockfish were examined at 1, 24, 48, and 72 hr. A total of 3,539 dead and 6 live larvae were collected from the fish tissues after standard enzymatic digestion. Salmon were infected with 1,245 of these larvae, and rockfish with 2,300. The 6 live worms, 2 from salmon and 4 from rockfish rounds, were recovered from muscle 1 hr after freezing; they were slightly motile and showed severe internal damage. No viable worms were found at or after 24 hr. The commercial blast-freezing process effectively killed larval nematodes in whole or dressed fish. Market-ready samples of previously blast-frozen silver salmon (O. kisutch) and chum salmon (O. keta) fillets and chum salmon steaks yielded no live worms, thereby confirming the efficacy of this process.     

Anisakis simplex (s.s.) larvae in wild Alaska salmon: no indication of post-mortem migration from viscera into flesh

The prevalence, mean intensity and distribution of Anisakis nematode third-stage larvae (L3) in the muscle and viscera of wild-caught chum salmon Oncorhynchus keta, pink salmon O. gorbuscha and sockeye salmon O. nerka were compared immediately after catch. Salmon were collected during the fishing season in July 2007 in Bristol Bay and Prince William Sound close to Cordova, Alaska (USA). All fish were infected, and more than 90% of the nematode larvae were found in the edible muscle meat. The isolated anisakid L3 were genetically identified as A. simplex (s.s.). The distribution of nematodes in the muscle meat of fresh-caught salmon was examined in 49 O. keta, 50 O. nerka and 12 O. gorbuscha from Cordova. Most of the larvae were detected in the muscle parts around the body cavity, but nematodes were also found in the tail meat and epaxial muscle (loins). The mean intensity of Anisakis larvae in the edible part was 21 individuals for O. gorbuscha, 62 individuals for O. keta and 63 individuals for O. nerka. No difference in the intensity of Anisakis larvae in the hypaxial muscle was found between fresh-caught and immediately gutted salmon and individuals stored ungutted for 24 h either on ice or in refrigerated sea water.     

Infection status of the sea eel (Astroconger myriaster) purchased from the Noryangjin fish market with anisakid larvae]

Although the sea eel (Astroconger myriaster) is suspected as one of the most important fish host for human anisakiasis in Korea, no report has been made on the infection status of the sea eel with anisakid larvae. In the present study, 26 sea eels (Astroconger myriaster) were purchased from the Noryangjin fish market in Seoul, and anisakid larvae were collected from their viscera, muscle, head and skin. The collected larvae were classified by their morphological types. A total of 1,351 anisakid larvae were collected from 15 of 26 fish examined. Among them, 1,269 were recovered from the viscera, 66 from the muscle, and 16 from the head and skin. Morphologically, most of the anisakids were classified into 6 known larval types, Anisakis type I (564 larvae) of Berland (1961), Contracaecum type A (409) and type D (5) of Koyama et al. (1969), Contracaecum type C' (83) and type D' (117) of Chai et al. (1986), and Contracaecum type V (1) of Yamaguti (1935). Remaining 172 specimens were new in the available literature, hence, designated as Contracaecum type A' (new type). The present results revealed that the sea eels caught in the Korean waters are heavily infected with anisakid larvae, not only in their viscera but also in the muscle, and Anisakis type I was the most common among the 7 larval types.     

Interaction between halofenozide and the entomopathogenic nematode Heterorhabditis marelatus for control of Japanese beetle (Coleoptera: Scarabaeidae) larvae

Japanese beetle, Popillia japonica Newman is a major pest of turf and ornamentals. Laboratory bioassays were conducted to evaluate the potential interactions between a biological control agent, Heterorhabditis marelatus (Nematoda: Heterorhabditidae) IN strain and the insecticide halofenozide against both overwintered and nonoverwintered 3rd instars of Japanese beetle. Treatments consisted of all combinations of 2 rates of halofenozid with H. marelatus nematodes Imidacloprid was used as a standard. Percentage larval mortality was evaluated at 7, 14, and 21 d after treatment. No deleterious effects were observed. The nematode treatments generally produced significantly greater larval mortality relative to both chemical treatments. Twenty-one days after treatment, both rates of nematodes resulted in 100% mortality, whereas insecticide treatments did not surpass 60% mortality. No synergism was detected in any of the combination treatments. There were no significant differences in nematode reproduction in larvae exposed to halofenozide and nematodes versus larvae exposed to only nematodes.     

Recovery of parasitic nematodes from fish by digestion or elution.

Two methods, digestion and elution, were used to recover parasitic nematodes from 470 flatfish belonging to species in the family Pleuronectidae. Samples of similar fish were collected from market lots; half of each sample was subjected to digestion, and half was subjected to elution (sedimentation). The edible (flesh) and the inedible (viscera) portions of each fish were analyzed separately. The total number of nematodes recovered by digestion was 1,110, which was not significantly greater than the 922 nematodes recovered by elution. However, digestion recovered 1,062 nematodes of the anisakine genera Anisakis and Phocanema, which are potentially pathogenic for human consumers of raw of semiraw fish. This number is significantly greater than the 608 pathogenic nematodes recovered by elution. Digestion also recovered 242 more nematodes from the edible flesh than did elution. Conversely, more nonpathogenic nematodes were recovered by elution. Approximately half the fish (240) had been collected in Boston markets, and the other half (230) had been collected in San Francisco markets. Fish from San Francisco each contained an average of eight nematodes, and those from Boston contained an average of less than one nematode per fish.     

'Cebiche'--a potential source of human anisakiasis in Mexico?

Five fish species used for preparation of a popular dish (cebiche) made with raw fish flesh in Mexico were obtained from five localities of the coast of Yucatan. Lutjanus synagris, Gerres cinereus, Sphyraena barracuda, Epinephelus morio and Haemulon plumieriwere examined for the presence of larvae of anisakid nematodes, causative agents of human anisakiasis. The nematode Pseudoterranova sp. was found in E. morio and S. barracuda with a total prevalence of 83% and 6.5 +/- 6.2 worms per fish for E. morio, and a prevalence of 33% and 10.2 +/- 30.0 worms per fish for S. barracuda. Contracaecumsp. was found to infect G. cinereus with a prevalence of 57% and 7.6 +/- 11.4 worms per fish. The relatively high prevalence of Pseudoterranova sp. indicates that this parasite is a potential causal agent of anisakiasis on the coast of Yucatan. Although all larvae were found only in the mesentery of the fish host, their importance as a potential source of human infection cannot be excluded as larval migration to the muscles in dead fish is possible.     

Critical factors required by the nematode Steinernema feltiae for the control of the leafminers Liriomyza huidobrensis, Liriomyza bryoniae and Chromatomyia syngenesiae

The critical factors required by entomopathogenic nematodes for the control of dipteran leafminers were investigated and compared. The foliar application of Steinernema feltiae and Heterorhabditis sp. (strain UK 211) caused a significant reduction in the numbers of dipteran leafminer larvae. There was no significant difference in efficacy between these nematode species against Liriomyza huidobrensis at 20°C and 85–90% r.h. Application of S. feltiae to all three larval instars of L. huidobrensis significantly reduced larval survival, with application to the second instar being the most effective at 20 ± 2°C and 80 ± 10% r.h. Humidities of >90% significantly enhanced the efficacy of S. feltiae. When S. feltiae was applied to second instar larvae, it was equally effective throughout the temperature range 10–30°C. At 20°C and >90% r.h., sufficient nematodes were able to enter the leaf tissues within the first 12 h after application to reduce larval survival to c. 15%. A comparison between L. huidobrensis, L. bryoniue and Chromatomyia syngenesiue indicated that the efficacy of S. feltiae was affected by the same critical factors for all three species.     

Caging experiment to examine mortality during metamorphosis of coral reef fish larvae

All previous attempts to estimate early postsettlement mortality of coral reef fishes using either caging experiments or disappearance of new recruits have examined fish that had already settled, and therefore did not include the metamorphosis process. Crest nets capture unharmed transparent larvae during their migration from the open ocean to lagoon reefs before metamorphosis. We released these presettlement larvae at night into cages surrounding patch reefs and measured larval survivorship after two nights. This caging experiment involved cages enclosing the natural resident fish fauna, including predators, and others cleared of fish before releasing the larvae. The analyses of variance showed that (1) there was no difference in survivorship between the seven trials, (2) there was a significant difference between cleared and uncleared cages, and (3) there were significant differences between larval species tested. For the seven species that had a significant difference in survivorship between cleared and uncleared cages, average mortality of the larvae was 14% (range 0-26%) in cleared cages and 67% (range 29-76%) in cages with predators. The difference in mortality between species was related to the size of the larvae, as larger species exhibited reduced mortality compared to smaller species. Mortality was related to the abundance of resident fish that could act as predators or competitors. Predation can have a significant impact on the survival of metamorphosing fish larvae on coral reefs.     

Pasture populations of cattle nematode larvae in Guadeloupe (French West Indies)

The development of numbers of third-stage (L3) larvae of gastrointestinal nematodes after pat (faeces) deposition by heifers is described, using a mathematical model, for seven experiments carried out in Guadeloupe (French West Indies). A dramatic rise in L3 population size occurred on herbage near pats, 17-25 days after pat deposition. There was no clear relationship between climatic data and L3 population dynamics. However, 5 successive days of rainfall induced the resumption of larval migration in two experiments. Changing the unit of expression of L3 population size (100 L3 m-2 vs 1000 L3 per kg DM) did not alter estimates of the model parameters. The evolution of L3 population of Haemonchus and Cooperia were similar.     

A simple technique for recovering larval ascaridoid nematodes from the flesh of marine fish

This paper describes and evaluates the efficiency of a simple technique for recovering larval ascaridoid nematodes (Anisakis simplex and Pseudoterranova decipiens) from the flesh of marine fish. The technique involves mechanical disintegration of the flesh in a domestic food processor, followed by visual inspection of diluted portions of the resulting homogenate under short-wave ultraviolet light. The nematodes, which remain intact, fluorescence brightly and are easily detected, particularly if the musculature has been frozen and thawed previously. The technique recovers a much higher proportion of the total number of nematodes than candling and slicing, is more rapid than pepsin-HCl digestion, and would therefore be suitable for large-scale surveys of ascaridoid nematodes in the flesh of marine fish.     

The suitability of the free-living nematode Panagrellus redivivus as live food for first-feeding fish larvae

The free‐living nematode Panagrellus redivivus has been recommended as a suitable food source for first‐feeding fish. A new technology for mass production of P. redivivus enables fish hatchery operators to rely on an inexpensive, standardized and permanently available live food for first‐feeding fish larvae. The proximate composition, and the fatty acid and amino acid profiles of nematodes mass produced on oat‐based and purified ingredient media were determined. The quality of nematodes was significantly influenced by the culture medium used. The lipid content and fatty acid composition of nematodes could be modified by using lipid‐enriched media. Mass‐produced nematodes were tested on first‐feeding common carp (Cyprinus carpio L.) and whitefish (Coregonus lavaretus) larvae. Carp larvae, grown on nematodes cultured on oat medium enriched with sunflower oil, showed a higher survival rate (87.1%) than the control group fed frozen zooplankton (82.9%) at the end of the 1‐week feeding experiment. Differences in larval mass between the treatments disappeared after subsequent feeding of a dry diet for 2 weeks. Whitefish larvae can be reared exclusively on a dry diet; here, the initial feeding of nematodes had no effect on final biomass and survival of larvae.     

Prevalence of larval Anisakis simplex in pen-reared and wild-caught salmon (Salmonidae) from Puget Sound, Washington

The abundance of parasites of public health significance in pen-reared salmon and wild-caught salmon was compared. Two hundred eighty-seven salmon from Puget Sound, Washington, were examined for third-stage larvae of Anisakis simplex. Of these fish, 237 Atlantic salmon (Salmo salar), coho salmon (Oncorhynchus kisutch) and chinook salmon (O. tshawytscha) were reared in commercial salmon pens and 50 sockeye salmon (O. nerka) were caught during their spawning migration. All wild-caught salmon were found to be infected with larval A. simplex; conversely, all pen-reared fishes lacked such infections. Edible musculature of wild salmon were infected with 581 (87%) nematode larvae. Of other salmon parasites known to infect humans, one Diphyllobothrium sp. plerocercoid was collected from each of three of the 50 wild-caught salmon. The study showed that farmed salmon may increase the margin of safety for consumers of raw seafood.     

Survival and behavioral characteristics of amphidromous goby larvae of Sicyopterus japonicus (Tanaka, 1909) during their downstream migration

To understand the ecology and environmental tolerances of newly hatched larvae of the amphidromous fish Sicyopterus japonicus during their downstream migration, the salinity tolerance of eggs, 0-15 day old larvae, and adults, and the temperature tolerance, specific gravity and phototaxis of hatched larvae were examined. Tolerances of adults were measured as survival after a 24 h challenge in freshwater (FW), brackish water (1/3 SW) and seawater (SW). The survival rate of adult S. japonicus was 100% in FW and 1/3 SW, while none survived in SW. Hatching success of eggs (30 eggs each) was significantly higher in FW (mean: 73%) and 1/3 SW (73%) than in SW (19%). Tolerance of newly hatched larvae to salinity and temperature was investigated in different combinations of salinities (FW, 1/3 SW and SW) and temperatures (18, 23 and 28 °C). Larval survival was significantly different in each salinity and temperature. Survival rate was significantly higher in 1/3 SW than in FW and higher in SW than in FW at 23 °C and 28 °C. At the latter part of the experiment, there was no survival in FW and at 28 °C. Survival was higher in lower temperatures, but larval development did not occur in FW. Specific gravity of newly hatched larvae was 1.036 at 28 °C and 1.034 at 23 °C. When exposed to a light source on one side of an aquarium, larval distribution was not affected. Our results indicated larval S. japonicus are more adapted to brackish water and seawater than freshwater, while the adults and eggs are more adapted to freshwater and brackish water than seawater. This is consistent with their amphidromous life history with growth and spawning occurring in freshwater and the larval stage utilizing marine habitats.     

The abundance of Anisakis simplex L3 in the body-cavity and flesh of marine teleosts

Smith J. W. 1984. The abundance of Anisakis simplex L3 in the body-cavity and flesh of marine teleosts. International Journal for Parasitology14: 491–495. In experiments conducted at sea, whole (ungutted) fish of three species were stored on ice at 3–5 °C for periods from 0 to 72 h. Some Anisakis simplex L3 migrated from the body-cavity into the flesh in mackerel Scomber scombrus, but not in either blue whiting Micromesistius poutassou or whiting Merlangius merlangus. Earlier work has shown that larvae migrate into the flesh of herring Clupea harengus post mortem but not of walleye pollock Theragra chalcogramma. It seems, therefore, that larvae migrate post mortem into the flesh of ‘fatty’ species (e.g. herring, mackerel) but not of ‘non-fatty’ species (e.g. blue whiting, whiting, walleye pollock). No significant larval excapsulation occurred in isolated mackerel or whiting viscera. In vivo, most encapsulated L3 occur in the body-cavity of euphausiid-feeding fish (herring, mackerel, blue whiting, walleye pollock) but are more widely distributed throughout the tissues of piscivorous fish (whiting, cod). These observations are discussed in relation to the apparent importance of euphausiids as intermediate hosts of A. simplex.     

Predation efficiency of indigenous larvivorous fish species on Culex pipiens L. larvae (Diptera: Culicidae) in drainage ditches in Argentina.

Two neotropical freshwater fish species, Cnesterodom decemmaculatus (Poeciliidae) and Jenynsia multidentata (Anablepidae), were collected from human-made ditches, a common habitat of the house mosquito Culex pipiens in La Plata, Argentina. Cnesterodom decemmaculatus was recorded in 62 of the 100 examined ditches, whereas J. multidentata was collected from only 21 ditches sympatrically with C. decemmaculatus. Culex pipiens was the only mosquito species collected, and its larvae and pupae were found in 38 of the 100 ditches. Fish and mosquito larvae and pupae were collected together in only two ditches and were significantly negatively correlated. Siphons of larval Culex and remnants of chironomid larvae, copepods, aquatic mites, and fish were present in the gut contents of two C. decemmaculatus from mosquito-positive ditches, while diatoms and filamentous algae were recorded in every fish dissected. Adult C. decemmaculatus and J. multidentata needed approximately 6.2 h to completely digest one Cx. pipiens 4th instar larva under laboratory conditions. When fish were confined with a density of 60 or fewer Cx. pipiens 4th instar larvae, C. decemmaculatus and J. multidentata adults consumed 100% of them in one day but only 35% and 42%, respectively, when confined with 150 larvae. Eradication of Cx. pipiens from a ditch, where densities had averaged 250 immatures per dip, was achieved 17 days after the introduction of 1,700 C. decemmaculatus.     

Developmental dynamics of myogenesis in the shipworm <Emphasis Type="Italic">Lyrodus pedicellatus</Emphasis> (Mollusca: Bivalvia)

Background

The shipworm Lyrodus pedicellatus is a wood-boring bivalve with an unusual vermiform body. Although its larvae are brooded, they retain the general appearance of a typical bivalve veliger-type larva. Here, we describe myogenesis of L. pedicellatus revealed by filamentous actin labelling and discuss the data in a comparative framework in order to test for homologous structures that might be part of the bivalve (larval) muscular ground pattern.

Results

Five major muscle systems were identified: a velum retractor, foot retractor, larval retractor, a distinct mantle musculature and an adductor system. For a short period of larval life, an additional ventral larval retractor is present. Early in development, a velum muscle ring and an oral velum musculature emerge. In late stages the lateral and dorsal mantle musculature, paired finger-shaped muscles, an accessory adductor and a pedal plexus are formed. Similar to other bivalve larvae, L. pedicellatus exhibits three velum retractor muscles, but in contrast to other species, one of them disappears in early stages of L. pedicellatus. The remaining two velum retractors are considerably remodelled during late larval development and are most likely incorporated into the elaborate mantle musculature of the adult.

Conclusions

To our knowledge, this is the first account of any larval retractor system that might contribute to the adult bodyplan of a (conchiferan) mollusk. A comparative analysis shows that a pedal plexus, adductors, a larval velum ring, velum retractors and a ventral larval retractor are commonly found among bivalve larvae, and thus most likely belong to the ground pattern of the bivalve larval musculature.

     

Infectivity and reproductive potential of the Oswego strain of Heterorhabditis bacteriophora associated with life stages of the clover root curculio,Sitona hispidulus

The infectivity and reproductive potential of the entomopathogenic nematode Heterorhabditis bacteriophora (Oswego strain), at different concentrations, was studied. Seventy to 80.0% mortality to late instar larvae of the clover root curculio, Sitona hispidulus, and 40.0-76.0% mortality to pupae, was observed at concentrations of 15-100 infective juveniles. There were no significant differences in mortality among nematode concentrations. LC(50) levels of 4.0 and 21.4 nematodes were determined for clover root curculio larvae and pupae, respectively. Nematodes did not cause significant mortality to adult or first instar clover root curculio. H. bacteriophora was able to complete its development and reproduce in 74.0-95.0% of clover root curculio late instar larvae and pupae. Reproductive potential in curculio larvae and pupae ranged from 0 to 7040 infective juveniles per host. Larvae exposed to 100 nematodes had a reproductive potential significantly higher than in those larvae exposed to 15 and 50 nematodes. Reproductive potential in pupae decreased with an increased nematode dose, indicating potential crowding effects. Host larval and pupal mass were positively correlated with nematode progeny production.