Anisakis Simplex Larvae Research Articles

Acetylcholinesterase secreted by Anisakis simplex larvae (Nematoda: Anisakidae) parasitizing herring, Clupea harengus: an inverse relationship of enzyme activity in the host–parasite system

Acetylcholinesterase (AChE) is a key enzyme involved in nerve impulse transmission in both vertebrates and invertebrates. In addition to neuromuscular AChE, many parasitic nematodes synthesize AChE in secretory glands and release the enzyme into their external environment. In this study, we evaluate the activities of both somatic and secreted AChE from larvae (L3) of the parasitic nematode Anisakis simplex, and compare these to the AChE activity in its host, herring, Clupea harengus. A. simplex larvae were obtained from a herring sampled in three areas of the southern Baltic. Enzyme kinetics were determined for excretory/secretory (E/S) products and somatic extracts of larvae as well as for herring muscle tissue. The results reveal that mean AChE activity is approximately fourfold higher in E/S products and eightfold higher in somatic extracts of post-secretory A. simplex larvae than in host muscle tissue. The level of AChE activity in nematodes is inversely related to the enzyme activity in their hosts, i.e. reduced AChE activity in herring was accompanied by increased enzyme activity in its parasites. The physiological function of AChE secreted by parasitic nematodes has been widely discussed in the literature, and numerous roles for this form of enzyme have been suggested. The results of our investigation indicate that AChE secretion by A. simplex larvae may constitute an adaptive mechanism that promotes survival under adverse environmental conditions. Larvae probably increase secretion of AChE in response to a direct and/or indirect effect of neurotoxic compounds. This is the first report of such a phenomenon in A. simplex.

Oral inoculation of live or dead third-stage larvae of Anisakis simplex in rats suggests that only live larvae induce production of antibody specific to A. simplex

Live Anisakis simplex third-stage larvae (L3) penetrate gastrointestinal mucosa after they are ingested in raw or undercooked seafood, thereafter causing gastrointestinal manifestations and allergic manifestations such as urticaria and anaphylaxis. These allergic reactions are mediated by specific IgE to L3 allergens, especially excretory-secretory (ES) allergens. Recent evidences suggest that only live larvae can cause allergic reactions, although cases attributable to ingestion of cooked, frozen seafood have been reported. Therefore the risk of Anisakis-associated hypersensitivity by ingestion of properly cooked and frozen fish remains controversial. No prior report describes the kinetics of antibody production in experimental animals after oral inoculation with dead L3. This study used ELISA to assess antibody production in rats inoculated orally with dead L3. Positive absorbance value in IgG, IgM, and IgE specific to ES antigen from L3 were found in rats inoculated with live L3 but not with dead L3 (frozen, heated, cut, or homogenized). At one week post re-inoculation with live or frozen L3 to the initially sensitized rats, the absorbance value of the specific IgM and IgE to ES antigen elevated quickly and highly in rats that had been re-inoculated with live L3, but they decreased slightly or did not change in rats inoculated with frozen L3. These results suggest that only ingestion of live L3 can produce the specific antibody and induce initial and secondary sensitizations to L3.

Activity of tea tree (Melaleuca alternifolia) essential oil against L3 larvae of Anisakis simplex.

Nematicidal activity of Melaleuca alternifolia essential oil, commonly known as tea tree oil (TTO), was assayed in vitro against L3 larvae of Anisakis simplex. The results showed a mortality of 100% for concentrations between 7 and 10 μL/mL after 48 h of incubation, obtaining an LD50 value of 4.53 μL/mL after 24 hours and 4.27 μL/mL after 48 hours. Concentration-dependent inhibition of acetylcholinesterase was observed for tea tree essential oil showing inhibition values of 100% at 100 μL/mL. This fact suggests that TTO may act as an AChE inhibitor. Terpinen-4-ol was discarded as main larvicide compound as it did not show larvicidal or anticholinesterase activity. The data obtained suggest that the essential oil of Melaleuca alternifolia may have a great therapeutic potential for the treatment of human anisakiasis.

Antioxidant, anti-adipocyte differentiation, antitumor activity and anthelminticactivities against Anisakis simplex and Hymenolepis nana ofyakuchinone A from Alpinia oxyphylla

BackgroundAlpinia oxyphylla is a common remedy in traditional Chinese medicine. Yakuchinone A is a major constituent of A. oxyphylla and exhibits anti-inflammatory, antitumor, antibacterial, and gastric protective activities.MethodsAntioxidant and antitumor characteristics of yakuchinone A in skin cancer cells as well as novel mechanisms for the inhibition of adipocyte differentiation, cestocidal activities against Hymenolepis nana adults, and nematocidal activities against Anisakis simplex larvae are investigated.ResultsYakuchinone A presents the ability of the removal of DPPH·and ABTS+ free radicals and inhibition of lipid peroxidation. Yakuchinone A suppresses intracellular lipid accumulation during adipocyte differentiation in 3 T3-L1 cells and the expressions of leptin and peroxisome proliferator-activated receptor γ (PPARγ). Yakuchinone A induces apoptosis and inhibits cell proliferation in skin cancer cells. The inhibition of cell growth by yakuchinone A is more significant for non-melanoma skin cancer (NMSC) cells than for melanoma (A375 and B16) and noncancerous (HaCaT and BNLCL2) cells. Treatment BCC cells with yakuchinone A shows down-regulation of Bcl-2, up-regulation of Bax, and an increase in cleavage poly (ADP-ribose) polymerase (PARP). This suggests that yakuchinone A induces BCC cells apoptosis through the Bcl-2-mediated signaling pathway. The anthelmintic activities of yakuchinone A for A. simplex are better than for H. nana.ConclusionsIn this work, yakuchinone A exhibits antioxidative properties, anti-adipocyte differentiation, antitumor activity, and anthelmintic activities against A. simplex and H. nana.

Expression of immune relevant genes in rainbow trout following exposure to live Anisakis simplex larvae

Basic immune response mechanisms in vertebrates against helminths are still poorly understood. Fish-nematode models may prove valuable for elucidation of this question. In this study we orally challenged rainbow trout (Oncorhynchus mykiss) with larvae of Anisakis simplex (Nematoda: Anisakidae) and subsequently investigated the expression of 18 immune relevant genes in spleen and liver 1, 4 and 8days post infection (d.p.i.). Gene expression data were analysed with regard to the infection status of the challenged rainbow trout at the time of necropsy; “worms rejected” (÷worms), “worms present” (+worms) and a combined group consisting of samples pooled from both previous groups (÷/+worms). No significant regulation of cytokine genes was recorded but fish which had rejected worms up-regulated the CD4 gene (6.1-fold change, 8d.p.i.) in liver. The gene encoding CD8 was significantly down-regulated 24h post challenge in livers in fish still carrying worms (2.7-fold change) but not in the worm-free group. The immunoglobulin gene IgM was significantly down-regulated (2.9-fold change, 8d.p.i.) in liver samples from the +worms group. Complement factor C3 and precerebellin genes were significantly up-regulated twofold in liver samples from infected fish 4d.p.i. Significant up-regulation of the acute-phase protein SAA was observed in all three groups and in both tissues. To our knowledge, this is the first study to describe the expression of immune genes in a fish host challenged with live nematode larvae.

Is Anisakis simplex s.l. a biological marker for stock identification of Strangomera bentincki from Chile?

The absence of the larval Anisakis simplex s. l. in samples from central Chile but heavy infections in fish from southern localities suggest the existence of two stocks of Strangomera bentincki, one closely associated with the central coast of Chile (landing port San Antonio and Talcahuano, c. 33° 30' S-36° 40' S) and the other associated with southern Chile (landing port Puerto Montt, c. 41° 30' S). Results confirm the usefulness of metazoan parasites, like A. simplex s. l. as biological markers.

Effects of excretory/secretory products from Anisakis simplex (Nematoda) on immune gene expression in rainbow trout (Oncorhynchus mykiss)

Excretory/secretory (ES) products are molecules produced by parasitic nematodes, including larval Anisakis simplex, a parasite occurring in numerous marine fish hosts. The effects of these substances on host physiology have not been fully described. The present work elucidates the influence of ES substances on the fish immune system by measuring immune gene expression in spleen and liver of rainbow trout (Oncorhynchus mykiss) injected intraperitoneally with ES products isolated from A. simplex third stage larvae. The overall gene expression profile of exposed fish showed a generalized down-regulation of the immune genes tested, suggesting a role of ES proteins in immunomodulation. We also tested the enzymatic activity of the ES proteins and found that lipase, esterase/lipase, valine and cysteine arylamidases, naphthol-AS-BI-phosphohydrolase and α-galactosidase activities were present in the ES solution. This type of hydrolytic enzyme activity may play a role in nematode penetration of host tissue. In addition, based on the notion that A. simplex ES products may have an immune-depressive effect (by minimizing immune gene expression) it could also be suggested that worm enzymes directly target host immune molecules which would add to a decreased host immune response and increased worm survival.

RELATIONSHIP BETWEEN RED VENT SYNDROME AND ANISAKID LARVAE BURDEN IN WILD ATLANTIC SALMON (SALMO SALAR)

The pathogenesis of the recently recognized "red vent syndrome" in wild Atlantic salmon (Salmo salar) is not fully understood. Pathologic observations indicate that this syndrome is associated with the presence of nonencapsulated larvae of the nematode Anisakis simplex in the body wall, the lower intestinal wall, and the visceral cavity surrounding the vent region. We evaluated the relationship between the occurrence of red vent syndrome and intensity of infection with Anisakis sp. larvae in naturally infected fish. Salmon caught by sport anglers were opportunistically evaluated to detect red vent syndrome. We included 106 salmon with red vent syndrome and 98 without red vent syndrome in this study. Intensity of infection was established by counting the total number of perivisceral larvae and by determining the number of larvae per gram in 10 g of pepsin-digested perianal tissue. The severity of inflammatory changes was also evaluated in standard histologic sections of the perianal area using a semiquantitative scale. Salmon with red vent syndrome had significantly higher intensity of inflammation than salmon without red vent syndrome (P=0.008). The odds of having red vent syndrome increased with the number of perianal larvae per gram of perianal tissue (P=0.002; odds ratio [OR]=1.12; 95% confidence interval: [1.05; 1.22]) but not with the number of perivisceral larvae, fish length, or gender. Although these results support the association between this syndrome and intensity of infection by A. simplex, the relationship is not strong (OR near 1), suggesting that the clinical expression of red vent syndrome at an individual level, and the emergence of this disease on a global scale, must be determined by other factors, such as timing of infection.

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.

The role of muscle-invading anisakid larvae on bacterial contamination of the flesh of post-harvest blue whiting (Micromesistius poutassou)

Many commercially utilised marine fish species are regularly infected with the larvae of parasitic nematodes of the genus Anisakis. Most of the larvae are found in the intestinal lumen of the fish, or on the organs of the visceral cavity. Some may, however, migrate from the intestine into the flesh of the fish, sometimes deeply into the fillets. The intestine of fish has a rich and diverse microbiota, including members of the specific spoilage bacteria (SSB) of seafood. Thus, the muscle invading nematodes will inevitably have a high load of bacteria in their intestinal tract and on their surfaces. While many studies are concerned with the quality reducing or human pathogenic properties these larvae represent, the present study aims to investigate the role of muscle-invading larvae of the species Anisakis simplex with respect to bacterial contamination of the flesh of post-harvest pelagic fish. Tissue samples of Anisakis-positive and Anisakis-negative body muscle, and samples of the fish intestine were collected from freshly caught blue whiting (Micromesistius poutassou) and processed for subsequent histological and microbiological examination. In Gram stained sections, the parasite intestine was densely populated with Gram-positive bacteria. The culture based quantitative analyses showed seven times higher numbers of colony forming units (CFU)/g for plate counts and ten times higher for SSB/g, when comparing the Anisakis-negative samples with the Anisakis-positive samples. This indicates that the parasite larvae act as vector for bacteria which contaminate the fish flesh. Molecular qualitative analyses were performed by 16S rRNA gene examinations, PCR-DGGE and sequencing. The general microbiota was determined, and some bacteria such as Shewanella sp., Photobacterium phosphoreum, and Vibrio sp., all known contributors to the bacterial spoilage in fish, were identified. These findings, supported by examination of histological sections of Anisakis-positive samples, along with the increased bacterial load in the flesh indicate that certain shelf life reducing SSB are introduced into the fish flesh by A. simplex larvae.

Expression and characterization of α-methylacyl CoA racemase from Anisakis simplex larvae.

Larval excretory-secretory products of Anisakis simplex are known to cause allergic reactions in humans. A cDNA library of A. simplex 3rd-stage larvae (L3) was immunoscreened with polyclonal rabbit serum raised against A. simplex L3 excretory-secretory products to identify an antigen that elicits the immune response. One cDNA clone, designated as α-methylacyl CoA racemase (Amacr) contained a 1,412 bp cDNA transcript with a single open reading frame that encoded 418 amino acids. A. simplex Amacr showed a high degree of homology compared to Amacr orthologs from other species. Amacr mRNA was highly and constitutively expressed regardless of temperature (10-40℃) and time (24-48 hr). Immunohistochemical analysis revealed that Amacr was expressed mainly in the ventriculus of A. simplex larvae. The Amacr protein produced in large quantities from the ventriculus is probably responsible for many functions in the development and growth of A. simplex larvae.

A technique for the intra-gastric administration of live larvae of Anisakis simplex in mice

To understand the mechanisms of infection and to attempt to simulate human infection by the Anisakidae family, many in vivo experimental approaches have been developed. The aim was to develop and present a technique for the induction of an oral infection through the use of an intra-gastric gavage of live Anisakis simplex in mice. A commercial pediatric gastric tube (No. 4) was cut longitudinally to produce a 3-cm slit at the distal end where the larva was placed to then be administered to the stomach of the mouse. There were no abnormal clinical complications before, during or after the procedure. In conclusion oral infection through the direct delivery of larvae in the stomach is simple and effective.

The effect of anti-Anisakis simplexantibody levels on C3 and C4 complement components in human sera

Previously, an in vitro effect was observed on the complement system not only of the excretory-secretory products but also of somatic antigens from L3 Anisakis simplex larvae. In the present work the effect of anti-A. simplex specific antibodies on C3 and C4 levels in human sera was investigated. Up to 309 samples of sera were tested to determine levels of C3 and C4 and anti-A. simplex antibodies, including immunoglobulins IgG, IgM, IgA and IgE. Significant differences were observed between levels of C3 and C4 and all immunoglobulins except for IgE. In the case of immunoglobulins, the probability that an anti-A. simplex positive subject has a C3 deficiency was 3.8 times higher than a subject without specific antibodies. In conclusion, an association between elevated levels of anti-A. simplex antibodies and C3 and C4 deficiency was demonstrated.

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.

Reactive oxygen species‐mediated activation of eosinophils induced by Anisakis simplex

Anisakiasis is parasitic disease caused by accidental ingestion of third stage larvae of Anisakis simplex carried by marine fishes or cephalopods. After ingestion, larvae can penetrate the stomach or intestinal wall and cause eosinophilic infiltration, inflammation, and allergic reactions. Although eosinophils are the principal effector cells of nematodes infection and allergic inflammation, the effect of A. simplex larvae on activation of human eosinophils remains largely unknown. Crude extract of A. simplex larvae caused phosphorylation of p38 MAPK and superoxide anion production, and increased surface expression of CD11b and CD69 on eosinophils. Pretreatment of eosinophils with p38 MAPK inhibitor SB202190 or NADPH oxidase inhibitor diphenyleneiodonium chloride reduced superoxide anion production and up‐regulation of CD11b and CD69 on eosinophils induced by A. simplex. However, pretreatment of eosinophils with diphenyleneiodonium chloride did not affect phosphorylation of p38 MAPK. A. simplex induced degranulation of human eosinophils and pretreatment of eosinophils with DPI blocked it. These results suggest that p38 MAPK‐mediated ROS production is required for the Anisakis‐induced activation of eosinophils.

Ani s 10, a new Anisakis simplex allergen: Cloning and heterologous expression

Anisakiasis is a human disease caused by accidental ingestion of larval nematodes belonging to the Anisakidae family. Anisakiasis is often associated with a strong allergic response. Diagnosis of A. simplex allergy is currently carried out by test based on the IgE reactivity to a complete extract of L3 Anisakis larvae although the specificity of these diagnostic tests is poor. Improving the specificity of the diagnostic test is possible using purified recombinant allergens. A new Anisakis allergen, named Ani s 10, was detected by immunoscreening an expression cDNA library constructed from L3 Anisakis simplex larvae. The new allergen was overproduced in Escherichia coli; it is a protein of 212 amino acids and it was localized as a 22 kDa protein band in an ethanol fractionated extract from the parasite. Ani s 10 has no homology with any other described protein, and its sequence is composed by seven almost identical repetitions of 29 amino acids each. A total of 30 of 77 Anisakis allergic patients (39%) were positive both to rAni s 10 and natural Ani s 10 by immunoblotting. The new allergen could be useful in a component-resolved diagnosis system for Anisakis allergy.

A 24 kDa Excretory-Secretory Protein ofAnisakis simplexLarvae Could Elicit Allergic Airway Inflammation in Mice

We have reported that a 24 kDa protein (22U homologous; As22U) of Anisakis simplex larvae could elicit several Th2-related chemokine gene expressions in the intestinal epithelial cell line which means that As22U may play a role as an allergen. In order to determine the contribution of As22U to allergic reactions, we treated mice with 6 times intra-nasal application of recombinant As22U (rAs22U). In the group challenged with rAs22U and ovalbumin (OVA), the number of eosinophils in the bronchial alveolar lavage fluid (BALF) was significantly increased, as compared to the group receiving only OVA. In addition, mice treated with rAs22U and OVA showed significantly increased airway hyperresponsiveness. Thus, severe inflammation around the airway and immune cell recruitment was observed in mice treated with rAs22U plus OVA. The levels of IL-4, IL-5, and IL-13 cytokines in the BALF increased significantly after treatment with rAs22U and OVA. Similarly, the levels of anti-OVA specific IgE and IgG1 increased in mice treated with rAs22U and OVA, compared to those treated only with OVA. The Gro-α (CXCL1) gene expression in mouse lung epithelial cells increased instantly after treatment with rAs22U, and allergy-specific chemokines eotaxin (CCL11) and thymus-and-activation-regulated-chemokine (CCL17) gene expressions significantly increased at 6 hr after treatment. In conclusion, rAs22U may induce airway allergic inflammation, as the result of enhanced Th2 and Th17 responses.

Seroprevalence of Antibodies againstAnisakis simplexLarvae among Health-Examined Residents in Three Hospitals of Southern Parts of Korea

The present study was performed to estimate the seroprevalence of larval Anisakis simplex infection among the residents health-examined in 3 hospitals in southern parts of Korea. A total of 498 serum samples (1 serum per person) were collected in 3 hospitals in Busan Metropolitan city, Masan city, and Geoje city in Gyeongsangnam-do (Province) and were examined by IgE-ELISA and IgE-western blotting with larval A. simplex crude extract and excretory-secretory products (ESP). The prevalence of antibody positivity was 5.0% and 6.6% with ELISA against crude extracts and ESP, respectively. It was also revealed that infection occurred throughout all age groups and higher in females than in males. A specific protein band of 130 kDa was detected from 10 patients with western blot analysis against crude extract and ESP among those who showed positive results by ELISA. Our study showed for the first time the seroprevalence of anisakiasis in Korea. The allergen of 130 kDa can be a candidate for serologic diagnosis of anisakiasis.

A study of the infection and physicochemical characteristics of the marine fish infected by Anisakis L₃ caught in Zhoushan Fishery

The aim of this research was to investigate the third phase Anisakis simplex larvae (Anisakis L₃) infection in marine fish caught in Zhoushan Fishery and to find out its physicochemical and biological characteristics. A total of 444 fish belonging to 29 species were dissected to isolate anisakis larvae which were then morphologically identified. The survival tolerance of Anisakis L₃ were observed in various conditions, such as in different temperature and medium. A total of 218 fish from 21 species were infected by Anisakis simplex larvae, yielding an overall infection rate of 49.10% (218/444). Trichiurus haumela, pneumatophorus japonicus, miichthys miiuy, argyrosomus argentatus and anguilliformes had high infection rate and had an average infection intensity of 15.28 per fish. 3332 Anisakis larvae were detected in 218 fish, among which Anisakis L₃ and Pseudoterranova larvae accounted for 99.46% (3314/3332) and 0.54% (18/3332) respectively. Anisakis L₃ was highly resistant to common condiment. We found the liquor with high concentration of alcohol showed better insecticidal effect than that with low concentration of alcohol (t = 4.105, P < 0.05) and low concentration mebendazole composite was not only more effective than high concentration mebendazole composite (F = 45.198, P < 0.01) but also more effective than other drugs, such as albendazole and mebendazole. Anisakis L₃ could live up to 9 h and 12 h at -20°C, -10°C respectively, however they were very sensitive to high temperature. It has been shown that they could only survive for less than 11 s and 1 s at 50°C and 60°C respectively. The observed Anisakis L₃ infection rate in the marine fish found in Zhoushan Fishery was very high. Anisakis L₃ showed high resistance to low temperature but not to high temperature.

Seasonal Occurrence of Helminths in the Anadromous Fish Coilia nasus (Engraulidae): Parasite Indicators of Fish Migratory Movements

To understand the seasonal migration of the anadromous Coilia nasus , we attempted to identify the parasites infecting C. nasus and determine their seasonal occurrence. From June 2007 to July 2008, a survey of 775 C. nasus individuals from the estuary of the Yangtze River and the coast of the East China Sea revealed more than 7,300 parasites associated with the gills and alimentary tracts of C. nasus . The following 6 helminth taxa were identified, i.e., the monogeneans Heteromazocraes lingmueni and Helciferus tenuis, the digenean Elytrophallus coiliae, the acanthocephalan Acanthosentis cheni , and larvae of the nematodes Anisakis simplex and Contracaecum sp., all of which are marine or brackish-water parasites. The absence of freshwater helminths suggested that the parasites acquired in freshwater may have been accidentally, and easily, lost by the time the fish had reached the estuary and coast. Contrary to seasonal occurrence of the parasites' life cycles, the lowest mean abundance and prevalence of H. lingmueni and A. cheni occurred in August, which suggested the immigration of C. nasus from freshwater to the Yangtze estuary, with lower parasite burdens. The highest mean abundance and prevalence of the nematodes A. simplex and Contracaecum sp. in May and June, and the lowest in August, indicated the arrival of the fish from the coast and the Yangtze River, to the estuary, respectively. These findings suggested that a majority of the fish prepared for spawning migration in the estuary in spring and early summer and returned to the estuary after spawning in the lower and middle reaches of the Yangtze River in late summer.