Occurrence of larval anisakids in horse mackerel (Trachurus trachurus) caught in Portuguese waters.

The third-stage larvae (L3) of the Anisakidae family are parasitic nematodes with zoonotic impact and are frequently encountered in the organs and musculature of various fish intended for human consumption. Since Anisakis simplex (s.s.) and A. pegreffii are the major aetiological agents of human disease, this study aims to combine the morphological and molecular data on the recovered anisakid larvae to contribute to a simplified morphological distinction of those species and conducted a survey of anisakid larvae infection in horse mackerel (Trachurus trachurus). Here, 116 horse mackerel caught in Portuguese waters were analysed for the presence of L3 of anisakids, and 3148 larvae were collected, of which only 30% were retrieved during visual inspection. As such, visual inspection does not appear to be very effective in anisakid detection. A prevalence of 84.5% of infected fish was found, and the mean intensity and mean abundance were 32.1 and 27.1 parasites per fish, respectively. The morphological and molecular analyses of 196 L3 randomly chosen from the total sample of parasites demonstrated the presence of L3 of mostly Anisakis spp., with only one L3 of Hysterothylacium aduncum. Relative frequencies of 62.9% for A. pegreffii and 37.1% for A. simplex (s.s.) were obtained. The morphometry differences between these two sibling species were evaluated, and the results demonstrated significant differences between the length of the ventriculus and the length of the oesophagus. Precisely, A. simplex (s.s.) has a longer oesophagus and ventriculus than A. pegreffii. As such, these differences may be used to distinguish the two species through morphological analysis.

Zoonotic nematode parasites infecting selected edible fish in New South Wales, Australia.

Despite increases in the annual consumption of seafood in Australia, studies on the occurrence and prevalence of zoonotic parasites in fish and the risk they may pose to human health are limited. The present study was aimed at determining the occurrence of zoonotic nematodes in commonly consumed fish in New South Wales, Australia's most populous state. Three species of fish, including the Australian pilchard, Australian anchovy, and eastern school whiting, were purchased from a fish market and examined for the presence of nematode parasites. All Australian pilchards examined in this study were infected (100%; n = 19), followed by the eastern school whiting (70%; n = 20) and Australian anchovy (56%; n = 70). Nematodes were in the larval stage and, therefore, classified by morphotype, followed by specific identification through sequencing of their internal transcribed spacer (ITS) regions. Seven different larval types with zoonotic potential, belonging to the families Anisakidae (Contracaecum type II and Terranova type II) and Raphidascarididae (Hysterothylacium types IV [genotypes A and B], VIII, XIV and a novel Hysterothylacium larval type, herein assigned as type XVIII), were found. The new larval type was identified as Hysterothylacium thalassini, based on ITS sequence data. The presence of the infective stage of a range of zoonotic parasites in fish commonly consumed in New South Wales is important, particularly as, in some dishes, these fish are used whole, raw or undercooked. This study provides the basis for future research on other aspects of these parasites, in regards to public health.

A new species of Raphidascaris (Nematoda: Raphidascarididae) infecting the fish Gymnogeophagus balzanii (Cichlidae) from the Pantanal wetlands, Brazil and a taxonomic update of the subgenera of Raphidascaris based on molecular phylogeny and morphology.

Raphidascaris (Sprentascaris) andersoni n. sp. (Nematoda: Raphidascarididae) collected in the intestine of the humphead cichlid Gymnogeophagus balzanii (Perugia) from the Pantanal wetlands, State of Mato Grosso do Sul (Brazil) is described and genetically characterized. The new species differs from its congeners mainly by having a conspicuous papilla-like formation slightly anterior to the cloacal aperture. Furthermore, males of R. (S.) lanfrediae and R. (S.) mahnerti have caudal alae, and R. (S.) hypostomi and R. (S.) pimelodi lack lateral alae, whereas in the new species caudal alae are absent and lateral alae present. The remaining congeners, namely, R. (S.) marano and R. (S.) saltaensis differ from Raphidascaris (Sprentascaris) andersoni n. sp. mainly because males have three pairs of postcloacal papillae (vs five pairs). In the phylogenetic reconstructions, using three nuclear genetic markers (18S, ITS1-5.8S-ITS2 and 28S rDNA) and one mitochondrial (cox1 mtDNA), the new species was separated from other representatives of Raphidascarididae, and the absence of monophyly in Hysterothylacium and Raphidascaroides was confirmed. Moreover, the subgenera Sprentascaris and Ichthyascaris appeared to be monophyletic. Therefore, even though Raphidascaris (Raphidascaris) was apparently not monophyletic, the subgenera of Raphidascaris should be re-erected as valid genera. The updated diagnoses of Ichthyascaris, Raphidascaris and Sprentascaris are given. The present study represents the first parasitological survey in G. balzanii.

New and known zoonotic nematode larvae within selected fish species from Queensland waters in Australia.

Zoonotic fish-borne nematodes are a significant and growing global health concern. However, in Australia their occurrence and geographical distribution are not well known. Moreton Bay in Queensland, Australia, is a popular tourist destination for domestic and international visitors where seafood is a popular food item. The aim of this study was to determine the occurrence of zoonotic nematodes within four commercially significant teleost fish species from Moreton Bay; Surf Bream, Sea Mullet, Yellow-Fin Tripod fish and Gold-Lined Sea Bream. Overall, 34.03% of fish examined (n = 144) were found to be a host to at least one parasitic nematode. The seasonal dynamics and prevalence of nematode infection was tested by statistical means. Results indicated a significant difference in seasonal infection for Surf Bream (p < 0.005) only. Several potentially zoonotic larval morphotypes were found, including Contracaecum types I and II, and Terranova type II (Family Anisakidae), and Hysterothylacium types VI and XIV and a new Hysterothylacium larval type herein assigned as type XVII (Family Raphidascarididae). ITS sequence data showed that some Contracaecum type II and Hysterothylacium types VI found in the present study are genetically distinct from those previously reported. The new genotypes were assigned to Contracaecum type II Genotype B and Hysterothylacium type VI, Genotypes B and C. This study provides essential information for future research on specific identification and differentiation of infective stages of zoonotic nematoda in edible fish.

Nematode parasites of commercially important fish from the southeast coast of Brazil: Morphological and genetic insight.

Studies of nematofauna of teleost fish from the Brazilian coast are relatively scarce and limited to identification based on morphology. The objective of the present study was to determine the diversity and prevalence of nematode parasites in teleost fish from the southeast Atlantic coast of Rio de Janeiro, through morphological, molecular, and ecological approaches. Parasites were collected from sixty specimens each of Genypterus brasiliensis, Micropogonias furnieri, and Mullus argentinae obtained in winters and summers of 2012­2014. Morphological and genetic characterization was conducted using light microscopy and the molecular targets 18S rDNA, ITS1, and mtDNA cox2. Nematodes identified in M. furnieri were Cucculanus genypteri (n = 1575, P = 98.3%) and Hysterothylacium deardorffoverstreetorum (s.l.) (n = 2, P = 3.3%); in G. brasiliensis were Dichelyne (Cucullanellus) sciaenidicola (n = 99, P = 33.3%), Cucculanus pulcherrimus (n = 45, P = 18.3%), Hysterothylacium deardorffoverstreetorum (s.l.) (n = 3, P = 5%), and Anisakis typica (n = 1, P = 1.7%); and, in M. argentinae, were H. deardorffoverstreetorum (s.l.) (n = 146, P = 48.3%), and Procamallanus (Spirocamallanus) halitrophus (n = 4, P = 6.7%). DNA sequence data of C. genypteri, C. pulcherrimus, D. (C.) sciaenidicola, and P. (S.) halitrophus were reported for the first time. New host records are M. argentinae for P. (S.) halitrophus, M. furnieri for A. typica, while H. deardorffoverstreetorum (s.l.) was found in all three fish species. Intestine showed significantly higher intensity than other sites, and no significant seasonal variation in parasitological indices was observed. Hysterothylacium specimens (n = 6) were found in fish muscle, potentially a public health concern.

Primeiro relato de Hysterothylacium deardorffoverstreetorum (Raphidascarididae) em bijupirá de criação, Rachycentron canadum (Linnaeus 1766), no Brasil / First report of Hysterothylacium deardorffoverstreetorum (Raphidascarididae) in cobia (Rachycentron canadum (Linnaeus 1766) farming in Brazil

O objetivo do presente estudo foi registrar a ocorrência de larva de nematoide da família Raphidascarididae, Hysterothylacium deardorffoverstreetorum, em bijupirá Rachycentron canadum (Linnaeus, 1766), criado e alimentado com ração e "trashfish" em fazendas marinhas localizadas no Rio de Janeiro, Brasil, alertando para possíveis riscos zoonóticos ao se usar esse tipo de alimentação para criação de peixes. Foram necropsiados 15 animais. Os parasitas encontrados foram coletados, fixados e, posteriormente, conservados em álcool a 70°GL, clarificados e identificados. Em todos os animais necropsiados, apenas um espécime estava parasitado com uma larva de Hysterothylacium deardorffoverstreetorum na serosa do fígado do peixe e conclui-se que a presença desse parasita em bijupirá de criação alimentado por "trashfish pode estar associada à alimentação, indicando, assim, um potencial risco zoonótico.(AU)

The aim of this study was to record the occurrence of nematode larvae of the Raphidascarididae family, Hysterothylacium deardorffoverstreetorum in cobia Rachycentron canadum (Linnaeus, 1766) grown and fed with trashfish in marine farms located in Rio de Janeiro, Brazil, warning regarding possible zoonotic risk using this type of food for fish. Fifteen animals were necropsied. Parasites found were collected, fixed and later preserved in alcohol 70°GL, clarified and identified. In all animals necropsied, only one specimen was infested with a Hysterothylacium deardorffoverstreetorum larvae in the serosa of fish liver. It is concluded that the presence of the parasite in cobia culture fed with trashfish may be associated with a potential zoonotic risk.(AU)

Survey for the presence of ascaridoid larvae in the cinnamon flounder Pseudorhombus cinnamoneus (Temminck & Schlegel) (Pleuronectiformes: Paralichthyidae).

The cinnamon flounder Pseudorhombus cinnamoneus is a frequently consumed marine fish in China. However, the occurrence of ascaridoid larvae in P. cinnamoneus remains unclear. In the present study, a total of 85 P. cinnamoneus caught from the Yellow Sea (off Shidao, 36°52'57″N, 122°26'42″E) in 2011, which is located between mainland China and the Korean Peninsula, was investigated for ascaridoid larval infection. Four ascaridoid larval types, including Anisakis type I of Berland (1961), Hysterothylacium type of Smith (1983), Hysterothylacium type HL of Guo et al. (2014) and Raphidascaris type of Zhao et al. (2016), were detected in this important food fish. These larval types were identified as Anisakis pegreffii, Hysterothylacium aduncum, H. sinense and Raphidascaris lophii, respectively, using PCR-based restriction fragment length polymorphism (PCR-RFLP) analysis and sequencing of the ITS region of nuclear ribosomal DNA (rDNA). The third-stage larvae of H. sinense are reported from Chinese waters for the first time. The prevalence of H. sinense was 100% and represents the predominant species of the ascaridoid larvae found in P. cinnamoneus. The prevalences of A. pegreffii and H. aduncum were 44.7% and 81.2%, respectively. Phylogenetic analyses based on ITS sequences were performed to elucidate the genetic relationships of these ascaridoid nematodes. The present study increases the knowledge and distribution of ascaridoid larvae in this area of Yellow Sea. The high prevalence of ascaridoid larvae in P. cinnamoneus shows that an assessment needs to be undertaken to assess the risk these parasites may pose to public health.

Integrative taxonomy of Anisakidae and Raphidascarididae (Nematoda) in Paralichthys patagonicus and Xystreurys rasile (Pisces: Teleostei) from Brazil.

Thirty-six Paralichthys patagonicus and 30 Xystreurys rasile were collected in the state of Rio de Janeiro, Brazil to investigate the presence of anisakid and raphidascaridid nematodes. Anisakis typica, Terranova sp., Contracaecum sp., Hysterothylacium deardorffoverstreetorum, and Raphidascaris sp. were identified using integrative taxonomy of morphological and genetic data. Morphological and morphometric analysis was conducted using bright field microscopy with scanning electron microscopy for topographic characterization of the cuticular surface. Phylogenetic analysis, using ITS and cox2 molecular targets, clearly demonstrated the species identification of A. typica and H. deardorffoverstreetorum and the high diversity of H. deardorffoverstreetorum. This is the first report of A. typica, H. deardorffoverstreetorum, and Raphidascaris sp. parasitizing P. patagonicus and X. rasile.

Characterisation of Ascaridoid larvae from marine fish off New Caledonia, with description of new Hysterothylacium larval types XIII and XIV.

Here we report occurrence of six different morphotypes of ascaridoid type larvae from 28 species of fish collected from New Caledonian waters. The larvae were morphologically identified as Anisakis type I, Hysterothylacium type VI and new larval types XIII and XIV, Raphidascaris larval type and Terranova larval type II. Representatives of each morphotype were subjected to the amplification of the second internal transcribed spacers (ITS-2) of ribosomal DNA (rDNA) and those sequences were compared with ITS-2 sequences of other ascaridoid nematodes previously deposited in GenBank. ITS-2 sequences of Anisakis larval type I were identical to those of A. typica. ITS-2 sequences of Hysterothylacium larval type VI in the present study were identical to those previously found in Eastern Australian waters. No match was found for ITS-2 sequences of Hysterothylacium larval types XIII and XIV; therefore, the specific identities of these larval types remain unclear. ITS-2 sequences of Raphidascaris larval type were identical to those of R. trichiuri, which have previously been reported in Taiwanese waters. Terranova larval type II in the present study had identical ITS-2 sequences with Terranova larval types reported from Australian waters, however, the specific identity is unknown. This taxonomic work is essential if further research on these zoonotic parasites is to be effective. This includes investigations into such aspects as life cycle studies, impacts on human health and risk assessment for their transmission to humans.

Anisakidae and Raphidascarididae larvae parasitizing Selene setapinnis (Mitchill, 1815) in the State of Rio de Janeiro, Brazil / Larvas Anisakidae e Raphidascarididae parasitos de Selene setapinnis (Mitchill, 1815) no Estado do Rio de Janeiro, Brasil

Between February and August, 2012, thirty specimens of Atlantic moonfish, Selene setapinnis, were purchased in local markets in Niterói, State of Rio de Janeiro, Brazil, with the aim of analyzing the presence of anisakid nematodes, establishing their rates of parasitism and infection sites, due to importance in the sanitary inspection. A total of sixty nematode larvae, belonging to at least two species were found: nine larvae of Terranova sp., Anisakidae, with prevalence (P) of 13.3%, mean intensity (MI) of 2.25, mean abundance (MA) of 0.30 and range of infection intensity (RI) from 1 to 6; and 51 larvae of Hysterothylacium fortalezae, Raphidascarididae, with P = 26.7%, MI = 6.40, MA = 1.70, and RI = 1-17. The infection sites for Terranova sp. were the mesentery and liver serosa; and for H. fortalezae, the infection sites were the mesentery, abdominal cavity and liver serosa. New morphological data from scanning electron microscopy, on the external structures of H. fortalezae (mainly at the posterior end), are presented. This is the first report of H. fortalezae parasitizing S. setapinnis.

De Fevereiro a Agosto de 2012, trinta espécimes de peixe galo, Selene setapinnis, foram adquiridos de mercados locais em Niterói, Estado do Rio de Janeiro, Brasil, com o objetivo de analisá-los quanto à presença de nematoides anisaquídeos, estabelecendo seus índices parasitários e sítios de infecção, devido a sua importância na inspeção sanitária. Do total de sessenta larvas de nematoides, pelo menos duas espécies foram encontradas: nove larvas de Terranova sp., Anisakidae, com prevalência (P) de 13,3%, intensidade média (IM) de 2,25, abundância média (AM) de 0,30, e amplitude de variação da intensidade de infecção (AI) de 1 a 6; e 51 larvas de Hysterothylacium fortalezae, Raphidascarididae, com P = 26,7%, IM = 6,40, AM = 1,70, e AI = 1 a 17. Os sítios de infecção para Terranova sp. foram o mesentério e a serosa do fígado; e para H. fortalezae, os sítios de infecção foram o mesentério, cavidade abdominal e serosa do fígado. Novos dados morfológicos das estruturas externas, principalmente da extremidade posterior de H. fortalezae, são evidenciados por microscopia eletrônica de varredura. Este é o primeiro relato de H. fortalezae parasitando S. setapinnis.

Genetic and morphological characterisation of a new species of the genus Hysterothylacium (Nematoda) from Paralichthys isosceles Jordan, 1890 (Pisces: Teleostei) of the Neotropical Region, state of Rio de Janeiro, Brazil

Taking into account the difficulties of taxonomic identification of larval anisakid nematodes based on morphological characters, genetic analyses were performed, together with those usually applied, in order to identify anisakid larvae found in the flounder Paralichthys isosceles from the littoral of the state of Rio de Janeiro, Brazil. The analysis of 1,820 larvae revealed a new species, similar to Hysterothylacium MD, Hysterothylacium 2, Hysterothylacium KB and Hysterothylacium sp regarding the absence of the larval tooth, an excretory pore situated below the nerve ring level, and slender lateral alae. Moreover, the new species differs from Hysterothylacium fortalezae and Hysterothylacium reliquens with regard to the number and size of spines present on the tail end and from Hysterothylacium patagonicus by the absence of interlabia. The maximum parsimony and neighbour joining tree topologies based on the 18S ribosomal DNA gene, complete internal transcribed spacer region and cytochrome oxidase 2 (COII) gene demonstrated that the Brazilian larvae belong to Raphidascarididae and represent a unique genetic entity, confirmed as a new Hysterothylacium species. Furthermore, the new species presents COII genetic signatures and shares polymorphisms with Raphidascarididae members. This is the first description of a new anisakid species from Brazil through the integration of morphological and molecular taxonomy data.