Experimental challenge of Anisakis simplex sensu stricto and Anisakis pegreffii (Nematoda: Anisakidae) in rainbow trout and olive flounder

The third-stage larvae of Anisakis simplex sensu lato (s.l.) are found in many marine fishes. To ensure food safety, it is important to determine whether these larvae are present in the body muscle of commercial fish species. However, there is little information regarding the tissue specificity of Anisakis and two of its sibling species, A. simplex sensu stricto (s.s.) and Anisakis pegreffii, that are common in marine fish in Japanese waters. We orally challenged rainbow trout (Oncorhynchus mykiss (Walbaum)), and olive flounder (Paralichthys olivaceus (Temminck and Schlegel)) with L3 larvae of these two sibling species and monitored infection for 5weeks. In rainbow trout, A. simplex s.s., but not A. pegreffii larvae, migrated into the body muscle. A small number of freely moving A. pegreffii larvae were recovered within the body cavity. In olive flounder, A. simplex s.s. larvae were found in both the body cavity and body muscle. A. pegreffii larvae were found only in the body cavity and primarily encapsulated in lumps. Our results indicate that there are differences in the sites of infection and host specificity between the two sibling species of A. simplex s.l.     

Molecular characterization of larval anisakid nematodes from marine fishes of Madeira by a PCR-based approach, with evidence for a new species

One-hundred and fifteen anisakid larvae from 3 different fish hosts, Aphanopus carbo, Scomber japonicus, and Trachurus picturatus, caught in Madeiran waters, were identified by PCR-RFLP. Three distinct species were identified in A. carbo, namely Anisakis simplex sensu srricto, Anisakis pegreffii, and Anisakis ziphidarum; 5 in S. japonicus, i.e., A. simplex s.s., A. pegreffii, Anisakis physeteris, Anisakis typica, and A. ziphidarum; and 3 in T. picturatus, i.e., A. simplex s.s., A. pegreffii, and A. typica. Anisakis simplex s.s. was the most frequent species in both A. carbo and S. japonicus (54% and 23.5%, respectively). Anisakis pegreffii and A. physeteris occurred with a frequency of 20.6% in S. japonicus, whereas in T. picturatus the most frequent species was A. typica (41.9%), followed by A. simplex s.s. (32.3%). Furthermore, A. carbo and S. japonicus were infected by an apparently undescribed taxon, provisionally named Anisakis sp. A. Based on estimations of the genetic distance, this new taxon seems to be more similar to A. ziphidarum (0.0335) than to other species of the genus.     

Molecular identification of Anisakis species from Pleuronectiformes off the Portuguese coast

Anisakid nematodes belonging to the Anisakis simplex complex are highly prevalent in several fish species off the coast of Portugal and are an important zoonotic problem in the Iberian Peninsula. Two reproductively isolated sibling species of the Anisakis simplex complex were identified from Pleuronectiformes inhabiting the Portuguese coast using restriction fragment length polymorphism (RFLP). Recombinant genotypes corresponding to presumptive Anisakis simplex sensu stricto and Anisakis pegreffii hybrids were also detected by this technique, as well as the species Anisakis typica. Although 25 species of Pleuronectiformes were investigated, Anisakis spp. larvae were only found in seven: Arnoglossus imperialis, Arnoglossus laterna, Lepidorhombus boscii, Citharus linguatula, Platichthys flesus, Dicologlossa cuneata and Solea senegalensis. The occurrence of hybrids in relatively sedentary fishes such as the Pleuronectiformes suggests that the Portuguese coast may constitute an area of hybridization and, therefore, is of particular interest for the study of the process of hybridization and speciation for these anisakids.     

Distribution of Anisakis species larvae from fishes of the Japanese waters

Human anisakiasis is caused by the consumption of raw, marinated or undercooked fish and squid infected with nematodes of the genus Anisakis Dujardin, 1845. In view of food safety, this study was carried out to examine the distribution of Anisakis species in marine fishes within Japanese waters. Seven fish species from six localities were collected and examined for Anisakis infection. Morphological and molecular (ITS region and mtDNA cox2 gene) characterization revealed the presence of two, among the three sibling species of Anisakis simplex, viz. A. simplex sensu stricto (s.s.) and A. pegreffii. Distribution data were collated with the results from the previous researches to better understand Anisakis distribution in Japanese waters. Distributions of Anisakis species were found to be locality-specific rather than host-specific, particularly between the two major species, A. simplex s.s. and A. pegreffii. Anisakis simplex s.s. is mainly found in fishes from northern Japan to Pacific sides, whereas A. pegreffii is in fishes from the Sea of Japan to East China Sea sides.     

Molecular identification of the etiological agent of the human anisakiasis in Japan

Anisakis simplex complex presently comprises three sibling species, A. simplex sensu stricto, A. pegreffii and A. simplex C. A. simplex is a common parasite in fishes and cephalopods and capable of causing anisakiasis in humans. Therefore, identification of sibling species of A. simplex was important for human health. In this study, one hundred Anisakis type I larvae isolated from eighty five patients with anisakiasis in Hokkaido and Kyushu in Japan were analyzed by adapting the new molecular method that can identify the sibling species of A. simplex complex. Based on the restriction fragment length polymorphism (RFLP) pattern of ITS regions including 5.8 subunit rRNA gene, we identified two sibling species, A. simplex s. str. and A. pegreffii. However, the infection rate of A. simplex s. str. was significantly higher than that of A. pegreffii. Eighty four (98.8%) out of the eighty five patients were infected with A. simplex s. str. On the contrary, one patients (1.2%) in Kyushu infected with A. pegreffii. This study provided basic information about human infection with A. simplex complex. Furthermore, we suggested that A. simplex s. str. is the most important etiological agent in Japan.     

Molecular characterization of Anisakis pegreffii larvae in Pacific cod in Japan

It is now recognized that the morphospecies Anisakis simplex is not a single species but a complex composed of three sibling species, A. simplex sensu stricto, A. pegreffii and A. simplex C. In Japan, A. simplex-like larvae have been isolated from a variety of fish and humans, but the larvae collected have been identified as A. simplex by only light microscopy. Therefore, the epidemiology of the A. simplex complex, composed of three sibling species, is still unclear in Japan. In the present study, 26 A. simplex-like larval isolates were obtained from two Pacific cod landed in Hokkaido, Japan, and examined genetically by PCR-RFLP and direct sequencing of the ITS region of rDNA. Among the 26 isolates, 24 were identified as A. simplex sensu stricto, the other two as A. pegreffii. The present study is the first to confirm the distribution of A. pegreffii in Japan, and to detect A. pegreffii larvae in Pacific cod.     

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.     

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.     

Evidence for a new species of <Emphasis Type="Italic">Anisakis</Emphasis> Dujardin, 1845: morphological description and genetic relationships between congeners (Nematoda: Anisakidae)

In the present study, a new biological species of Anisakis Dujardin, 1845, was detected in Kogia breviceps and K. sima from West Atlantic waters (coast of Florida) on the basis of 19 (nuclear) structural genes studied by multilocus allozyme electrophoresis. Fixed allele differences at 11 enzyme loci were found between specimens of both adults and larvae of the new species and the other Anisakis spp. tested. Reproductive isolation from A. brevispiculata Dollfus, 1968 was demonstrated by the lack of hybrid or recombinant genotypes in mixed infections in K. breviceps. Genetic distance of the new species from its closest relative, A. brevispiculata, was D(Nei)=0.79. The new species is morphologically different from the other species which have been genetically characterised and from the other Anisakis retained by Davey (1971) as valid or as species inquirendae: the name of Anisakis paggiae n. sp. is proposed for the new taxon. Anisakis Type II larvae (sensu Berland, 1961) from the European hake Merluccius merluccius in the northeastern Atlantic Ocean (Galician coast) and from the scabbard fish Aphanopus carbo in Central Atlantic waters (off Madeira), were identified as A. paggiae n. sp. Its genetic relationships with respect to the seven species previously characterised (A. simplex (Rudolphi, 1809) sensu stricto), A. pegreffii Campana-Rouget & Biocca, 1955, A. simplex, (A. typica (Diesing, 1860), A. ziphidarum Paggi et al., 1998, A. physeteris Baylis, 1923 and A. brevispiculata) were also inferred. Overall, a low genetic identity was detected at allozyme level between the eight Anisakis species. Interspecific genetic identity ranged from I(Nei)=0.68, between the sibling species of the A. simplex complex, to I(Nei)=0.00 (no alleles shared at the considered loci) when A. physeteris, A. brevispiculata and the new species were compared with the other species of the genus. Concordant topologies were obtained using both UPGMA and NJ tree analyses for the considered species. In both analyses, A. paggiae n. sp. clustered with A. brevispiculata. They also indicated two main clades, the first including A. physeteris, A. brevispiculata and A. paggiae n. sp., the second containing all of the remaining species (i.e. A. simplex (s.s.), A. pegreffii, A. simplex, A. typica and A. ziphidarum). A deep separation between these two main Anisakis clades, also supported by high bootstrap values at the major nodes, was apparent. This is also supported by differences in adult and larval morphology, as well as with respect to their main definitive hosts. A morphological key for distinguishing adult A. paggiae n. sp., A. physeteris and A. brevispiculata is presented. Allozyme markers for the identification of any life-history stage of the Anisakis spp. so far studied, as well as ecological data on their definitive host preferences and geographical distribution, are updated.     

Genetic identification of Anisakis larvae in European hake from Atlantic and Mediterranean waters for stock recognition

The occurrence of seven species of the larval parasitic nematode Anisakis , which can be used as a biological tag for hake Merluccius merluccius stocks throughout their geographical range, is reported. Hake were collected from 14 localities in the Mediterranean Sea and the Atlantic Ocean. Anisakis larvae ( n  = 1950), which were recovered, were identified to species by means of genetic markers (allozymes). Within Anisakis type I, the larvae of A. pegreffii , A. simplex s.s ., A. typica and A. ziphidarum were detected, while within Anisakis type II, A. physeteris , A. brevispiculata and Anisakis sp. were identified. There were significant differences in the relative proportions of the various Anisakis species identified in hake samples from the Mediterranean Sea and Atlantic Ocean, suggesting the existence of different stocks of M. merluccius in European waters.     

Larval anisakids parasitizing the blue whiting, Micromesistius poutassou, from Motril Bay in the Mediterranean region of southern Spain

A total of 301 blue whiting, Micromesistius poutassou Risso, 1826, ranging in length from 17 to 28 cm, from Motril Bay (Mediterranean coast, south Spain) were examined for anisakid nematodes, as these fish are common items in the Spanish Mediterranean diet. Three anisakid species were morphologically identified with a total prevalence of 10.63%. Anisakis simplex s.l. Rudolphi, 1809 had a prevalence value of 6.65%, compared with 2.66% for A. physeteris Baylis, 1923 and 2.33% for Hysterothylacium aduncum Rudolphi, 1802. Variations in prevalence values with season and host size are discussed. Allozyme markers (leucine aminopeptidase-1) were used to identify anisakid nematodes assigned to the A. simplex complex and all examined larvae were found to correspond genetically to A. pegreffii Nascetti et al., 1986.     

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.     

Genetic markers in ribosomal DNA for the identification of members of the genus Anisakis (Nematoda: ascaridoidea) defined by polymerase-chain-reaction-based restriction fragment length polymorphism

Polymerase-chain-reaction-based restriction fragment length polymorphism analysis was performed to establish genetic markers in rDNA, for the identification of the three sibling species of the Anisakis simplex complex and morphologically differentiated Anisakis species, i.e. Anisakis physeteris, Anisakis schupakovi, Anisakis typica and Anisakis ziphidarum. Different restriction patterns were found between A. simplex sensu stricto and Anisakis pegreffii with two of the restriction endonucleases used (HinfI and TaqI), between A. simplex sensu stricto and A. simplex C with one endonuclease (HhaI), and between A. simplex C and Aniskis pegreffii with three endonucleases (HhaI, HinfI and TaqI), while no variation in patterns was detected among individuals within each species. The species A. physeteris, A. schupakovi, A. typica and A. ziphidarum were found to be different from each other and different from the three sibling species of the A. simplex complex by distinct fragments using 10-12 of the endonucleases tested. The polymorphisms obtained by restriction fragment length polymorphisms have provided a new set of genetic markers for the accurate identification of sibling species and morphospecies.     

Molecular systematics, phylogeny and ecology of anisakid nematodes of the genus Anisakis Dujardin, 1845: an update

Advances in the taxonomy and ecological aspects concerning geographical distribution and hosts of the so far genetically recognised nine taxa of the nematodes belonging to genus Anisakis (i.e. A. pegreffii, A. simplex s.s., A. simplex C, A. typica, A. ziphidarum, Anisakis sp., A. physeteris, A. brevispiculata and A. paggiae) are here summarized. Genetic differentiation and phylogenetic relationships inferred from allozyme (20 enzyme-loci) and mitochondrial (sequences of cox-2 gene) markers, are revised and compared. The two genetic analyses are congruent in depicting their phylogenetic relationships. Two main clusters are showed to exist in the obtained trees, one encompassing the species A. pegreffii, A. simplex s.s., A. simplex C, A. typica, A. ziphidarum and Anisakis sp.; while, the second including A. physeteris, A. brevispiculata and A. paggiae. The existence of two clades is also supported by their morphological differentiation in adult and larval morphology. Comparison of phylogenetic relationships among Anisakis spp. with those currently available for their cetacean definitive hosts suggests parallelism between host and parasite phylogenetic tree topologies. Preliminary data for reconstruction of a possible co-evolutionary scenario between cetacean hosts and their Anisakis endoparasites suggests that cospeciation and host-switching events may have accompanied the evolution of this group of parasites. Finally, genetic/molecular markers for the identification of the so far genetically recognized taxa of Anisakis at any life-stage and both sexes were given also in relation to human anisakiosis is discussed.     

A new species of Anisakis Dujardin, 1845 (Nematoda, Anisakidae) from beaked whales (Ziphiidae): allozyme and morphological evidence

Larvae and adults of Anisakis, recovered from the beaked whales Mesoplodon layardii and Ziphius cavirostris from the Mediterranean Sea and South African waters, were analysed morphologically and by molecular markers (allozymes). A new Anisakis species was identified, showing fixed allele differences at a number of loci from the other Anisakis spp. tested (A. simplex complex, A. physeteris). The lack of hybrid or recombinant genotypes in mixed infections with A. pegreffii, A. simplex C and A. physeteris, as well as the high values of genetic distance (average D_Nei = 1.65 from the members of the A. simplex complex, and D_Nei = 3.09 from A. physeteris) showed that the new species is reproductively isolated. This new Anisakis species is morphologically different from the other Anisakis retained by Davey (1971) as either good species or species inquirendae. The name Anisakis ziphidarum n. sp. is proposed for the new species.     

Genetic relationships among Anisakis species (Nematoda: Anisakidae) inferred from mitochondrial cox2 sequences, and comparison with allozyme data

The genetic relationships among 9 taxa of Anisakis Dujardin, 1845 (A. simplex (sensu stricto), A. pegreffii, A. simplex C., A. typica, A. ziphidarum, A. physeteris, A. brevispiculata, A. paggiae, and Anisakis sp.) were inferred from sequence analysis (629 bp) of the mitochondrial cox2 gene. Genetic divergence among the considered taxa, estimated by p-distance, ranged from p = 0.055, between sibling species of the A. simplex complex, to p = 0.12, between morphologically differentiated species, i.e., A. ziphidarum and A. typica. The highest level was detected when comparing A. physeteris, A. brevispiculata, and A. paggiae versus A. simplex complex (on average p = 0.13) or versus A. typica (on average p = 0.14). Sequence data from the newly identified Anisakis sp. poorly aligned with other Anisakis species but was most similar to A. ziphidarum (p = 0.08). Phylogenetic analyses based upon Parsimony and Bayesian Inference, as well as phenetic analysis based upon Neighbor-Joining p-distance values, generated similar tree topologies, each well supported at major nodes. All analyses delineated two main claides, the first encompassing A. physeteris, A. brevispiculata, and A. paggiae as a sister group to all the remaining species, and the second comprising the species of the A. simplex complex (A. simplex (s.s.), A. pegreffii and A. simplex C), A. typica, A. ziphidarum, and Anisakis sp. In general, mtDNA-based tree topologies showed high congruence with those generated from nuclear data sets (19 enzyme-loci) and with morphological data delineating adult and larval stages of the Anisakis spp.; however, precise positioning of A. typica and A. ziphidarum remain poorly resolved, though they consistently clustered in the same clade as Anisakis sp. and the A. simplex complex. Comparison of anisakid data with those currently available for their cetacean-definitive hosts suggests parallelism between host and parasite phylogenetic tree topologies.     

Allozyme and PCR-RFLP markers in anisakid nematodes,aethiological agents of human anisakidosis

In the framework of the researches granted by MURST COFIN97, studies on anisakid nematodes, aethiological agents of human anisakidosis, were carried out. The project was aimed to implement the knowledge on the systematics, genetics, ecology and epizootiology of species of the genera Anisakis and Pseudoterranova by applying genetic markers obtained from multilocus allozyme electrophoresis and from PCR-based techniques. The results obtained by allozyme studies allowed to extend the geographic distribution and to detect new definitive and intermediate/paratenic hosts of two sibling species of the A. simplex complex, i.e. A. simplex s.s. and A. pegreffii and to characterise the species A. simplex C, a new sibling species within the A. simplex complex as well as a new species belonging to the genus Anisakis, A. ziphidarum. Combined allozyme and morphological analyses provided markers for the identification of the four sibling species of the Pseudoterranova decipiens complex and their nomenclatural designations. New markers based on PCR-RFLP analysis were used for the identification of sibling species of the Anisakis simplex complex and of another four species of the genus. Genetic markers based on three diagnostic restriction enzymes allowed the identification as A. pegreffii of a larva obtained by endoscopy in a case of human anisakidosis in Southern Italy.     

Multiplex PCR for the identification of Anisakis simplex sensu stricto, Anisakis pegreffii and the other anisakid nematodes

A multiplex PCR method was established for the rapid identification of Anisakis simplex sensu stricto, A. pegreffii, A. physeteris, Pseudoterranova decipiens, Contracaecum osculatum and Hysterothylacium aduncum. The sequence alignment of the internal transcribed spacer 1 region (ITS-1) between A. simplex s. str. and A. pegreffii showed a high degree of similarity, but only two C-T transitions were observed. To differentiate A. simplex s. str. from A. pegreffii, an intentional mismatch primer with an artificial mismatched base at the second base from the primer 3' end was constructed. This intentional mismatch primer, which produced a PCR band only from A. pegreffii DNA, was able to differentiate the two morphologically indistinguishable sibling species of A. simplex. Specific forward primers for other anisakid species were also designed based on the nucleotide sequences of the ITS region. The multiplex PCR using these primers yielded distinct PCR products for each of the anisakid nematodes. The multiplex PCR established in this study would be a useful tool for identifying anisakid nematodes rapidly and accurately.     

Anisakis simplex larvae: infection status in marine fish and cephalopods purchased from the Cooperative Fish Market in Busan, Korea

The infection status of marine fish and cephalopods with Anisakis simplex third stage larva (L3) was studied over a period of 1 year. A total of 2,537 specimens, which consisted of 40 species of fish and 3 species of cephalopods, were purchased from the Cooperative Fish Market in Busan, Korea, from August 2006 to July 2007. They were examined for A. simplex L3 from the whole body cavity, viscera, and muscles. A. simplex L3 were confirmed by light microscopy. The overall infection rate reached 34.3%, and average 17.1 larvae were parasitized per infected fish. Fish that recorded the highest infection rate was Lophiomus setigerus (100%), followed by Liparis tessellates (90%), Pleurogrammus azonus (90%), and Scomber japonicus (88.7%). The intensity of infection was the highest in Gadus macrocephalus (117.7 larvae per fish), followed by S. japonicus (103.9 larvae) and L. setigerus (54.2 larvae). Although abundance of A. simplex L3 was not seasonal in most of the fish species, 10 of the 16 selected species showed the highest abundance in February and April. A positive correlation between the intensity of L3 infection and the fish length was obvious in S. japonicus and G. macrocephalus. It was likely that A. simplex L3 are more frequently infected during the spring season in some species of fish. Our study revealed that eating raw or undercooked fish or cephalopods could still be a source of human infection with A. simplex L3 in Korea.     

Larval Anisakid Infections in Marine Fish from Three Sea Areas of the Republic of Korea

The present study was performed to determine the infection status of anisakid larvae in marine fish collected from 3 sea areas of the Republic of Korea. Total 86 marine fish (8 species) collected from the East Sea (Goseong-gun, Gangwon-do), 171 fish (10 species) from the South Sea (Sacheon-si, Gyeongsangnam-do), and 92 fish (7 species) from the Yellow Sea (Incheon Metropolitan City) were examined by both naked eyes and artificial digestion method. Among the total of 349 fish examined, 213 (61.0%) were infected with 8 species of anisakid larvae, i.e., Anisakis simplex, 6 types of Contracaecum spp., and Raphidascaris sp., and the mean larval density was 13.8 per infected fish. Anisakid larvae were detected in 45 fish (52.3%) from the East Sea, 131 fish (76.6%) from the South Sea, and 37 fish (40.2%) from the Yellow Sea. The average numbers of larvae detected were 4.0, 16.6, and 15.9, respectively. Anisakis simplex larvae were detected in 149 fish (42.7%), and the mean larval density was 9.0 per infected fish. They were found in 26 fish (30.2%) collected from the East Sea, 96 fish (56.1%) from the South Sea, and 27 fish (29.3%) from the Yellow Sea. The average numbers of larvae detected were 2.9, 10.3, and 10.5, respectively. Conclusively, the present study suggests that the infection rate and density of anisakid larvae are more or less higher in the fish from the South Sea than those from the East Sea or the Yellow Sea.