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.     

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.     

A study of the migration of larval Anisakis simplex (Nematoda: Ascaridida) in the Chilean hake, Merluccius gayi (Guichenot)

Hake were collected during four trips to investigate a possible migration from viscera to somatic musculature of larval Anisakis simplex. On each trip 150 hake were examined and separated in five groups of 30 hake. One group consisted of fish gutted immediately after capture; two other groups were gutted 15 h after capture, one of which had been left at environmental temperature, and the other put in ice; the remaining two groups (exposed to the same treatment) were gutted 30 h after capture. All fish flesh was digested (HCl-pepsin digest); all nematodes found in the digest and in the viscera were counted. There was no increase of larval nematodes in flesh at 15 and 30 h. Temperature did not affect the migration of larvae. A regression test did not show significant differences between groups. Finally, there was no correlation between number of larvae in viscera and in musculature.     

Comparison of fatty acid profiles of spawning and non-spawning Pacific herring, Clupea harengus pallasi

Crude lipid and fatty acid composition from liver, intestine, roe, milt and flesh of spawning and non-spawning Pacific herring (Clupea harengus pallasi) were examined to determine the relative effects of spawning on the nutritional value of herring. Depletion of lipid due to spawning condition was significant (P < 0.01) in all organ tissues and flesh of spawning herring. The lipid content ranged from an average of 1.9 to 3.4% (wet weight basis) in different organ tissues of spawning herring, to 10.5 to 16% in non-spawning fish. The fatty acid profile exhibited many differences in the relative distribution of individual fatty acids among organ tissues and between the two fish groups. Oleic acid (C18:1n-9), a major monounsaturated fatty acid (MUFA) found in all tissue lipids, decreased significantly (P < 0.01) in spawning fish. The two monoenes, C20:1n-9 and C22:1n-11, occurred at high concentrations in the flesh but at only minor proportion in the digestive organs and gonads. Spawning herring also had significantly (P < 0.01) higher polyunsaturated fatty acids (PUFA) content in the organ tissues, particularly in the milt and ovary, with docosahexaenoic acid (C22:6n-3, DHA) having the greatest proportion. Among the n-6 fatty acids, only C18:2n-6 and C20:4n-6 occurred at notable amounts and were present in higher proportions in spawning fish. We concluded that although relatively higher n-3 fatty acid content was found in the organ lipids of spawning herring, they are not an energy-dense prey food source due to the fact that both flesh and gonads contain a very low amount of lipid.     

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.     

Biological features of the common fish species in Olyutorsky-Navarin region and the adjacent waters of the Bering Sea: 2. Families Macrouridae,Clupeidae, and Osmeridae

Biological features of the four common fish species, giant grenadier Albatrossia pectoralis (Macrouridae), Pacific herring Clupea pallasii (Clupeidae), Pacific rainbow smelt Osmerus mordax dentex, and Pacific capelin Mallotus villosus catervarius (Osmeridae), were studied under the 20-year dataset (1995?2015). These species inhabit the northwestern Bering Sea in the summer–autumn period and form the schoolings in the Olyutorsky-Navarin region. The size–age parameters of the fish caught by different sampling gear, as well as the peculiarities of the body length and body weight dynamics, spawning periods, spawning range, and conditions, were analyzed. The largest specimens of giant grenadier, Pacific herring, and Pacific rainbow smelt were observed in the catches performed by the bottom setline and the gill nets; the smallest fish were found in the trawl catches. The body length and body weight of Pacific herring were larger in the pelagic trawls compared to the bottom trawls; an opposite pattern was observed for the Pacific capelin. The abundant year-class in the species with short life cycle (capelin and herring) is well tracked on the longterm plots of the fish body size; this is accompanied by the decrease of their biological parameters. Herring stock covers large growing grounds; smaller body size was observed for the herring grazing in the coastal waters; young specimens dominate here.     

Fish eggs and larvae in the diet of herring Clupea harengus membras Linnaeus, 1758 and the sprat Sprattus sprattus balticus (Schneider, 1904) (Clupeidae) in the Southeastern Baltic Sea

The qualitative and quantitative composition of fish eggs and larvae in the stomach contents of Baltic herring and sprat was examined. The occurrence of fish eggs and larvae in the diet of herring was markedly higher, compared to sprat. Eggs of four and larvae of nine fish species were found in the herring stomachs. The prey species composition of the herring and sprat diets was related to the ichthyoplankton composition and to the degree of overlapping of the prey and predator distributions. Sprat eggs were the predominant prey in the herring diet in May and sprat larvae prevailed in August. The daily ration estimates indicate that the sprat larvae consumption by herring in August exceeded the sprat egg consumption in May by many times. Cod eggs in the herring stomachs were only found at the optimal oxygen content and salinity for the cod eggs in the near-bottom water layer.     

Larval and juvenile Pacific herring Clupea pallasii are not susceptible to infectious hematopoietic necrosis under laboratory conditions

Infectious hematopoietic necrosis (IHN) leads to periodic epidemics among certain wild and farmed fish species of the Northeast (NE) Pacific. The source of the IHN virus (IHNV) that initiates these outbreaks remains unknown; however, a leading hypothesis involves viral persistence in marine host species such as Pacific herring Clupea pallasii. Under laboratory conditions we exposed specific pathogen-free (SPF) larval and juvenile Pacific herring to 10(3) to 10(4) plaque-forming units (pfu) of IHNV ml(-1) by waterborne immersion. Cumulative mortalities among exposed groups were not significantly different from those of negative control groups. After waterborne exposure, IHNV was transiently recovered from the tissues of larvae but absent in tissues of juveniles. Additionally, no evidence of viral shedding was detected in the tank water containing exposed juveniles. After intraperitoneal (IP) injection of IHNV in juvenile herring with 10(3) pfu, IHNV was recovered from the tissues of sub-sampled individuals for only the first 5 d post-exposure. The lack of susceptibility to overt disease and transient levels of IHNV in the tissues of exposed fish indicate that Pacific herring do not likely serve a major epizootiological role in perpetuation of IHNV among free-ranging sockeye salmon Oncorhynchus nerka and farmed Atlantic salmon Salmo salar in the NE Pacific.     

Discrimination of Baltic herring populations with respect to Anisakis simplex larvae infection

Samples of herring Clupea harengus in the Baltic were collected during the spawning season in western, middle and eastern waters off the Polish coast. Two samples were taken per area, several morphometric characters were measured and some meristic characters (vertebrae and gill rakers) were counted. Infection with Anisakis simplex larvae, total length, age and maturity stage were recorded. Univariate and multivariate statistical tests were used to compare the samples, discriminant analysis was applied to separate herring populations and the fraction of herring populations in each area was estimated. The results obtained indicated that infected fish represented a mixed group of western and central Baltic herring. In the first samples collected from each area, the share of the western population decreased from 40% in the western to 18% in the middle and 13% in the eastern coasts. The share of western herring was higher in the second samples collected in the same area and decreased from 91% in the western coast to 14 and 32% in the middle and eastern ones, respectively.     

Morphological and molecular characterization of Anisakis larvae (Nematoda: Anisakidae) in Beryx splendens from Japanese waters

The third-stage (L3) larvae of Anisakis, which are the etiological agents of human anisakiasis, have been categorized morphologically into Anisakis Type I larvae and Anisakis Type II larvae. Genetic analysis has allowed easy identification of these larvae: Anisakis Type I larvae include the species Anisakis simplex sensu stricto, Anisakis pegreffii, Anisakis simplex C, Anisakis typica, Anisakis ziphidarum, and Anisakis nascettii, whereas Anisakis Type II larvae include the species Anisakis physeteris, Anisakis brevispiculata, and Anisakis paggiae. Since human consumption of raw fish and squid is common in Japan, we investigated Anisakis L3 larvae in 44 specimens of Beryx splendens from Japanese waters. A total of 730 Anisakis L3 larvae collected from B. splendens were divided morphologically into 4 types: Type I, Type II, and 2 other types that were similar to Anisakis Type III and Type IV described by Shiraki (1974). Anisakis Type II, Type III, and Type IV larvae all had a short ventriculus, but their tails were morphologically different. In addition, data from genetic analysis indicated that Anisakis Type II, Type III, and Type IV larvae could be identified as A. physeteris, A. brevispiculata, and A. paggiae, respectively. Therefore, A. physeteris, A. brevispiculata, and A. paggiae can be readily differentiated not only by genetic analysis but also by morphological characteristics of L3 larvae.     

Spatial and Temporal Variability in Juvenile Pacific Herring, Clupea pallasi, Growth in Prince William Sound, Alaska

We examined the spatial and temporal variability of juvenile Pacific herring, Clupea pallasi, growth within Prince William Sound, Alaska. Pacific herring, ranging from post-larval to mature fish, were collected from four spatially segregated bays between October 1995 and March 1998. Linear growth equations from each bay were similar. However, growth rates and wet weight-at-length, reflecting condition, of juvenile Pacific herring cohorts varied seasonally and annually. The short term spatial variability in juvenile Pacific herring growth suggested that each bay was a unique nursery area. The physical and biological conditions within each bay appeared to dictate Pacific herring growth rate.     

Molecular characterization of Anisakis larvae from fish caught off Sardinia

Anisakis spp. larvae are parasitic, and potentially zoonotic, nematodes transmitted by marine fish and cephalopods, which are the main intermediate hosts of the third larval stage. The accidental consumption of infected raw or poorly cooked fish may cause gastroenteric diseases and allergies in humans. The aim of the present study was to use polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to define the occurrence, species variability, and host preferences of Anisakis spp. larvae in fish caught off the coast of Sardinia. Necropsy was used on 285 samples; 552 Anisakis spp. L3 larvae were isolated from 87 fish that tested positive for this nematode. Anisakis pegreffii was most frequently encountered (90.6%), with a primary preference for Scomber scombrus, Zeus faber, and Trachurus mediterraneus. In contrast, the prevalence of Anisakis physeteris was only 1.3%. A hybrid genotype of Anisakis simplex sensu stricto and Anisakis pegreffii was also observed, which confirms the results of previous studies carried out in the western Mediterranean. Interestingly, no Anisakis simplex s.s. larvae were recovered. These results indicate that the diversity of Anisakis species is low in Sardinia waters, probably because of its geographic position.     

The incidence of Anisakis sp. larvae (Nematoda: Ascaridata) in the commercially exploited stocks of herring (Clupea harengus L., 1758,) (Pisces: Clupeidae) in British and adjacent waters.

10 000 herrings from the commercial fisheries around the coasts of Great Britain and Ireland were examined for the nematode larva Anisakis sp. between October 1968 and October 1970. Heavy infestations were found in the North Sea stocks, moderate infestations west of Scotland and minimal infestations elsewhere; but nowhere was the parasite completely absent. Younger fish were often found to be more heavily infested than older fish and it is therefore suggested that the accumulation of larvae in the body cavity may be affected by annual fluctuations either in the population of an infested first intermediate host or in the extent to which the herrings are feeding on this host. A small but constant proportion of the parasites penetrate the musculature of the herring regardless of other factors such as the age or length of the host, or the number of other larvae present.     

Multivariate morphometric analysis of Pacific herring, <Emphasis Type="Italic">Clupea pallasii</Emphasis> (Clupeiformes: Clupeidae) in Sakhalin waters: Investigation of intraspecific differentiation

In the Pacific herring, Clupea pallasii, eight samples of spawning, mature fish (collected in 1978 and 1981) and six samples of immature fish individuals (collected in 1979, 1980 and 1984) were analyzed. Mature fish were analyzed by 24 conventional morphometric characters or morphometric indices. By means of discriminant and factor analyses (principal component mode) the existence of differentiating groupings of vector scores was found that have obvious biological meaning as local Pacific herring stocks. In concordance with former morphobiological and genetic data, current multivariate observations supported the existence in the surveyed area of Sakhalin waters of at least three stocks. One of them is the Sakhalin-Hokkaido local stock of Pacific herring, which in the past provided the bulk of herring catches in the region and showed a distinct and sharper morphometric differentiation as compared with other herring assemblages.     

Anisakis pegreffii Larvae in Sea Eels (Astroconger myriaster) from the South Sea,Republic of Korea

Anisakis simplex sensu stricto (s.s.), Anisakis pegreffii, Anisakis berlandi (=A. simplex sp. C), and Anisakis typica are the 4 major species of Anisakis type I larvae. In the Republic of Korea (Korea), A. pegreffii, A. berlandi, and A. typica larvae in fish hosts has seldom been documented. In this study, molecular analysis was performed on Anisakis larvae from the sea eels (Astroconger myriaster), the major source of human anisakiasis in Korea, collected from Tongyeong City, a southern coastal area of Korea. All 20 sea eels examined were infected with Anisakis type I larvae (160 larvae; 8 per fish). Their species were analyzed using PCR-RFLP patterns and nucleotide sequences of internal transcribed spacers (ITS1, 5.8 subunit gene, and ITS2) and mitochondrial cytochrome c oxidase 2 (cox2). Most (86.8%; 112/129) of the Anisakis type I larvae were A. pegreffii, and 7.8% (10/129) were A. typica. The remaining 5.4% (7/129) was not identified. Thus, A. pegreffii is the major species of anisakid larvae in sea eels of the southern coast of Korea.     

Distribution of Anisakis larvae, identified by genetic markers, and their use for stock characterization of demersal and pelagic fish from European waters: an update

In the present paper, recent results obtained on the use of different distributions observed in larval species of Anisakis, genetically identified by means of allozyme markers, for stock characterization of demersal (Merluccius merluccius), small (Trachurus trachurus) and large pelagic (Xiphias gladius) finfish species in European waters, are reviewed and discussed. Several species of Anisakis were identified in the three fish hosts: A. simplex (s.s.), A. physeteris, A. typica, A. ziphidarum, A. pegreffii, A. brevispiculata and A. paggiae. Canonical discriminant analysis performed on all the samples of the three fish species collected in areas comprising their geographical range, according to the different species of Anisakis identified, showed distinct fish populations in European waters. In all the three fish hosts, the pattern of distribution of Anisakis larvae allowed discrimination of Mediterranean stocks from Atlantic stocks. In the case of swordfish, the possible existence of a southern Atlantic stock separated from a northern one is also suggested. Congruence and discordance with the population genetic data inferred from allozyme markers on the same samples of the three fish species are also discussed.     

Electrophoretic identification of Anisakis sp. larvae (Ascaridida: Anisakidae) from Clupea harengus L. in Baltic Sea

On the basis of electrophoretic studies carried out on 15 gene-enzyme systems, 80 Anisakis sp. larvae from the herring Clupea harengus, fished in South Baltic Sea and Gdanisk Bay, were identified as Anisakis simplex B. This is the first record of Anisakis simplex B in the Baltic Sea. The spawning migration of the herrings from the North Sea to the Baltic Sea and the distribution of A. simplex B are briefly discussed.     

Anisakis simplex: uncoupling of oxidative phosphorylation in the muscle mitochondria of infected fish

Adenosine triphosphatase activity stimulated by Mg2+ was greater in muscle mitochondria of fish infected with larval Anisakis simplex nematodes than in uninfected fish. When muscle mitochondria were isolated in a sucrose ethylene-glycol bis(beta-aminoethyl ether)N,N'-tetraacetic acid medium from fresh uninfected fish, they were loosely coupled, and their adenosine triphosphatase activity was comparable to that of mitochondria from rat tissue. Activity in infected fish was dose dependent, increasing with the number of worms per fish. Excretory secretory products or a cytoplasmic fraction of anisakines, when incubated with coupled rat mitochondria, also caused adenosine triphosphatase activity to increase. Storage of fish flesh caused an increase in adenosine triphosphatase activity, but such aging was not significant until 5 and 10 days after death in refrigerated and frozen samples, respectively. The Mg2+ stimulated adenosine triphosphatase activity of muscle mitochondria can be used to estimate the number of nematodes per market fish. The type of medium used to isolate the mitochondria is crucial in such studies; an ionic medium with Nagarse proteinase was optimal for fish muscle mitochondria.     

Population structure of three species of Anisakis nematodes recovered from Pacific sardines (Sardinops sagax) distributed throughout the California Current system

Members of the Anisakidae are known to infect over 200 pelagic fish species and have been frequently used as biological tags to identify fish populations. Despite information on the global distribution of Anisakis species, there is little information on the genetic diversity and population structure of this genus, which could be useful in assessing the stock structure of their fish hosts. From 2005 through 2008, 148 larval anisakids were recovered from Pacific sardine (Sardinops sagax) in the California Current upwelling zone and were genetically sequenced. Sardines were captured off Vancouver Island, British Columbia in the north to San Diego, California in the south. Three species, Anisakis pegreffii, Anisakis simplex 'C', and Anisakis simplex s.s., were identified with the use of sequences from the internal transcribed spacers (ITS1 and ITS2) and the 5.8s subunit of the nuclear ribosomal DNA. The degree of nematode population structure was assessed with the use of the cytochrome c oxidase 2 (cox2) mitochondrial DNA gene. All 3 Anisakis species were distributed throughout the study region from 32°N to 50°N latitude. There was no association between sardine length and either nematode infection intensity or Anisakis species recovered. Larval Anisakis species and mitochondrial haplotype distributions from both parsimony networks and analyses of molecular variance revealed a panmictic distribution of these parasites, which infect sardines throughout the California Current ecosystem. Panmictic distribution of the larval Anisakis spp. populations may be a result of the presumed migratory pathways of the intermediate host (the Pacific sardine), moving into the northern portion of the California Current in summer and returning to the southern portion to overwinter and spawn in spring. However, the wider geographic range of paratenic (large piscine predators), and final hosts (cetaceans) can also explain the observed distribution pattern. As a result, the recovery of 3 Anisakis species and a panmictic distribution of their haplotypes could not be used to confirm or deny the presence of population subdivision of Pacific sardines in the California Current system.     

The effects of salinity and temperature on the development and survival of fish parasites

In brackish water the variety of marine and freshwater parasite species is considerably reduced. The distribution in brackish water of most marine endoparasites is restricted by the salinity tolerance of their hosts, most of the parasite species are more tolerant than their hosts. The influence of salinity and temperature on nine species has been examined; first stage larvae of Contracaecum aduncum develop in 0-32‰ salinity; Cryptocotyle lingua proved to be infective at salinities down to 4‰. The greatest resistance was found in Anisakis larvae from herring Clupea harengus , which survived for more than half a year. Parasites in the fish intestines appear to be unaffected by changing water salinities, as the osmolarity in the intestines stays nearly constant. Marine ectoparasites ( Acanthochondria depressa, Lepeophtheirus pectoralis ) survive about three times longer than freshwater species ( Piscicola geometra, Argulus foliaceus ) when salinity is 16‰. High temperature increases the effects of adverse salinities on parasites. There is evidence that none of these ecto-parasitic species can develop within the range of 7-20‰ salinity.