MOLECULAR CHARACTERIZATION OF ANISAKID NEMATODES HYSTEROTHYLACIUM SPECIES FROM JAPANESE THREADFIN BREAM NEMIPTERUS JAPONICUS (BLOCH, 1791) (PERCIFORMES, NEMIPERIDAE) FROM IRAQI MARINE WATER FISH

The present study provides a new insight into valuable information on the diverse structure of the Anisakid population and discusses the limited species richness in the Nemipterus japonicus (Bloch,1791) (Perciformes, Nemiperidae). The fishing area consists of various locations in the Arabian Gulf (29°58 0 33 00 N48°28 0 20 E). A total of 315 marine fish were examined, (n=287) were infected. Larval stages (n= 763) encysted within the mesenteries peritoneum and viscera of fish organs were isolated, with a prevalence of 91.11% of infection and, the intensity was 2.65. Molecular analysis was carried out on thirty individuals who have examined the morphology and showed some appearance differences, by amplifying internal transcribed spacers ITS and ITS-1 of nuclear rDNA (rDNA) by PCR using the primer sets NC5/NC2 and SS1/NC13R of thirteen DNA products. Evolutionary analyses were conducted in MEGA X. based on the identity percentage in the GenBank database showed that they belong to anisakid nematodes, in particular, they belong to eleven distinct taxa within the Hysterothylacium Ward & Magath, 1917 (Rhabditida, Raphidascarididae) and one identified species H. amoyense (Hsü, 1933) Deardorff & Overstreet, 1980. The current study records eleven species that belong to a genus of Hysterothylacium; some of the alignment of sequences polymorphisms reveals new different individuals of larvae species that may be adopted as new species if their adult stage is detected, and N. japonicus fish considered as a new host record. The current study provides some insights on the systematic taxonomy of these parasites, in addition, it supports similar studies that have been published elsewhere.

Novel Omics Studies on Anisakid Nematodes

Parasitic nematodes infecting humans and animals are widely distributed in marine and terrestrial environments, causing considerable morbidity and mortality globally [...]

Metagenomics Analysis Reveals an Extraordinary Inner Bacterial Diversity in Anisakids (Nematoda: Anisakidae) L3 Larvae

L3 larvae of anisakid nematodes are an important problem for the fisheries industry and pose a potential risk for human health by acting as infectious agents causing allergies and as potential vectors of pathogens and microrganisms. In spite of the close bacteria–nematode relationship very little is known of the anisakids microbiota. Fresh fish could be contaminated by bacteria vectored in the cuticle or in the intestine of anisakids when the L3 larvae migrate through the muscles. As a consequence, the bacterial inoculum will be spread, with potential effects on the quality of the fish, and possible clinical effects cannot be discarded. A total of 2,689,113 16S rRNA gene sequences from a total of 113 L3 individuals obtained from fish captured along the FAO 27 fishing area were studied. Bacteria were taxonomically characterized through 1803 representative operational taxonomic units (OTUs) sequences. Fourteen phyla, 31 classes, 52 orders, 129 families and 187 genera were unambiguously identified. We have found as part of microbiome an average of 123 OTUs per L3 individual. Diversity indices (Shannon and Simpson) indicate an extraordinary diversity of bacteria at an OTU level. There are clusters of anisakids individuals (samples) defined by the associated bacteria which, however, are not significantly related to fish hosts or anisakid taxa. This suggests that association or relationship among bacteria in anisakids, exists without the influence of fishes or nematodes. The lack of relationships with hosts of anisakids taxa has to be expressed by the association among bacterial OTUs or other taxonomical levels which range from OTUs to the phylum level. There are significant biological structural associations of microbiota in anisakid nematodes which manifest in clusters of bacteria ranging from phylum to genus level, which could also be an indicator of fish contamination or the geographic zone of fish capture. Actinobacteria, Aquificae, Firmicutes, and Proteobacteria are the phyla whose abundance value discriminate for defining such structures.

Absence of anisakis nematodes in smoked farmed Atlantic salmon (Salmo salar) products on sale in European countries

The increase of global demand of aquaculture products as compensation for the lowering of fishery sustainability, has shown a parallel awareness by the consumers on the importance of the safety and quality of fish products. Among these, salmon industry has reached a leading position demonstrating the negligible risk of presence of zoonotic helminths such as anisakid nematodes in farmed salmon. Despite the massive production of data in literature on parasitological surveys carried out on fresh salmon, no data are published on processed farmed salmon such as smoked products. In 2016, 270 fillets of smoked farmed Atlantic salmon (Salmo salar) and 13 smoked fillets from wild sockeye salmon (Oncorhynchus nerka) have been analyzed by visual inspection and UV-press method searching for the presence of anisakid nematodes. No parasites were detected in fillets from farmed Atlantic salmon, while 10 out of 13 fillets from wild salmon were positive for Anisakis simplex s.s. larvae. This first survey on the possible presence of anisakid nematodes in processed smoked salmon confirms that this risk in farmed Atlantic salmon products has to be considered negligible.

Assessment on the Occurrence of Anisakid and other Endoparasitic Nematodes Infecting Commercially-Important Fishes at Tayabas Bay

Anisakid nematodes are parasites commonly present in the marine environment. Parasites belonging to the family Anisakidae or the genus Anisakis can cause two different clinical manifestations: gastrointestinal disorders and allergic reactions known as anisakiasis. In this study, we examined 7,126 marine fishes belonging to four different commercially-important fish species; Rastrelliger kanagurta, Sardinella lemuru, Atule mate, and Selar crumenophthalmus for the presence of anisakid and other endoparasitic nematode infection. The fishes caught from Tayabas Bay were bought from three different landing sites from March 2017 to February 2018. The gonads, liver, and stomach of each fish species were incubated for 12-18 hours for rapid isolation and endoparasite evaluation. After the isolation of parasites, anisakid nematodes were fixed in vials with 70% ethanol for morphological analysis under the microscope. Six anisakid groups of genera, including Hysterothylacium, Terranova, Anisakis, Contracaecum, Raphidascaris, and Camallanus, and a non-anisakid group Echinorhynchus were identified. The results showed that the prevalence of anisakid infection in all species was 24.18 %, with a mean intensity of infection of 1.91. Rastrelliger kanagurta (Dalahican), Atule mate, and Selar crumenophthalmus were the most infected with 50.90%, 38.98%, and 30.52% prevalence rate, respectively, followed by Rastrelliger kanagurta (San Francisco) (24.18%) and Sardinella lemuru (7.46%). The collected data suggest that commercially-important fish caught in the Tayabas Bay waters are susceptible to parasitization by larvae of the genus Camallanus followed by Hysterothylacium and Terranova in their visceral organs. The prevalence of anisakid infection was almost similar between female (45.3 %) and male (47.21 %) fishes with a mean intensity of 1.95 & 1.96, respectively. Also, larger fishes were heavily infected with anisakid larvae than small fishes. Thus, the intensity and prevalence of the fish parasite can be used as a biological tag for benchmarking and stock assessment purposes.

Parasitic Anisakid Nematode Isolated from Stranded Fraser’s Dolphin (Lagenodelphis hosei Fraser, 1956) from Central Philippine Waters

Cetaceans, including dolphins, serve as definitive hosts of zoonotic anisakid nematodes, which are important etiological agents for human anisakiasis and allergy-associated health risks. With limited knowledge of these zoonotic parasites from the marine environment in the Philippine waters, the stranding of a Fraser’s dolphin (Lagenodelphis hosei Fraser, 1956) off the central Philippines made it possible to identify the worm species isolated from its gut. Parasitological examinations were carried out using morphological and molecular tools. Morphologically, the SEM and LM data revealed that the specimens belong to the genus Anisakis of the Type 1 group. Molecularly, PCR-RFLP results of the ITS region generated only a single fragment pattern on all worm samples corresponding to the reported molecular keys for A. typica. Further sequence and phylogenetic analyses of both ITS rDNA and mtDNA COX2 genes confirmed the anisakid nematodes’ identity as A. typica. The molecular data obtained in this study support previous findings on the possible existence of local variants of A. typica in this region.

Metagenomics Analysis Reveals An Extraordinary Bacterial Diversity in Anisakids (Nematoda: Anisakidae) L3 Larvae

Background: L3 larvae of anisakid nematodes are an important problem for the fisheries industry and pose a potential risk for human health by acting as agents causing allergies and as potential vectors of pathogens microrganisms. In spite of the close bacteria-nematode relationship very little is known of the Anisakids microbiote. Fresh fish could be contaminated by bacteria vectored in the cuticle or in the intestine of Anisakids when the L3 larvae migrate through the muscles. As a consequence the bacterial inoculum will be spread, affecting the quality of the fish, and possible clinical effects cannot be discardedResults: A total of 2,689,113 16S rRNA gene sequences from a total of 113 L3 individuals obtained from fish captured along the FAO 27 area were studied. Bacteria were taxonomically characterized through 1803 representative OTUsequences. Fourteen Phyla, 31 Classes, 52 Orders, 129 Families and 187 genera were unambiguously identified. We have found as part of microbiome an average of 123 OTUs per L3 individual. Diversity Indices (Shannon and Simpson indices) indicate an extraordinary diversity of bacteria at a OTU level. There are clusters of Anisakids individuals (samples) defined by the associated bacteria which however are not significantly related with fish hosts or anisakid taxa. This suggests that association or relationship among bacteria in Anisakids, exists without the influence of fishes or nematodes. The lack of relationships with hosts of Anisakids taxa has to be necessarily expresssed by the association among bacterial OTUs or other taxonomical levels which are ranging from OTUs to phylum level. Conclusions: There are significant biological structural associations of microbiota in anisakid nematodes which manifest in clusters of bacteria ranging from plyllum to genus level which could also be anindicator of degree of fish contamination or from the geographic zone of fish capture. Actinobacteria, Aquificae, Firmicutes, and Proteobacteria are the phyla whose abundance value discriminate for difining such structures.

Advances in Omic Studies Drive Discoveries in the Biology of Anisakid Nematodes

Advancements in technologies employed in high-throughput next-generation sequencing (NGS) methods are supporting the spread of studies that, combined with advances in computational biology and bioinformatics, have greatly accelerated discoveries within basic and biomedical research for many parasitic diseases. Here, we review the most updated “omic” studies performed on anisakid nematodes, a family of marine parasites that are causative agents of the fish-borne zoonosis known as anisakiasis or anisakidosis. Few deposited data on Anisakis genomes are so far available, and this still hinders the deep and highly accurate characterization of biological aspects of interest, even as several transcriptomic and proteomic studies are becoming available. These have been aimed at discovering and characterizing molecules specific to peculiar developmental parasitic stages or tissues, as well as transcripts with pathogenic potential as toxins and allergens, with a broad relevance for a better understanding of host–pathogen relationships and for the development of reliable diagnostic tools.

Nematode Parasites in Baltic Sea Mammals, Grey Seal (Halichoerus grypus (Fabricius, 1791)) and Harbour Porpoise (Phocoena phocoena (L.)), from the German Coast

Purpose Endoparasitic nematodes of six harbour porpoises Phocoena phocoena and four grey seals Halichoerus grypus, stranded at the eastern coast of the Baltic Sea in Germany in winter 2019, were analysed in order to identify nematode parasites and to compare with recent studies from the same area. Methods Endoparasitic nematodes were identified by using both morphological and molecular characters. The successfully obtained sequences of the rDNA marker regions ITS-1, 5.8S, ITS-2 from 29 anisakid and the rDNA marker region ITS-2 of 11 pseudalid nematodes were amplified. Results Analyses revealed the presence of three parasite species, the anisakid nematode Contracaecum osculatum from grey seals and the pseudalid nematodes Pseudalius inflexus and Stenurus minor from the harbour porpoises. Other anisakid nematodes regularly occurring in the Baltic Sea, e.g. Anisakis simplex or Pseudoterranova decipiens, were not found. Conclusions The prevalence of 100% and a severe parasite load in grey seals demonstrated a very high C. osculatum infection of Baltic Sea fish as their regular prey. Prevalence of 33% for parasites in harbour porpoises and minor infection rates, combined with a distinct lack of anisakid nematodes, are typical for the current situation of the porpoise parasite fauna in the Baltic Sea. Invasive parasite species as possible indicators for climate change could not be detected.