Exploring the use of fatty acid ethyl esters as a potential natural solution for the treatment of fish parasitic diseases.

Alternatives to conventional chemical treatments for parasitic diseases in fish are needed. Microalgal-sourced fatty acid ethyl esters (FAEEs) have shown an antiparasitic effect against Gyrodactylus turnbulli infection in guppies. Here, we tested a range of commercial FAEEs of various carbon chain lengths and unsaturation levels against two fish parasites. Guppies and barramundi infected with G. turnbulli and Trichodina sp., respectively, were used. The most effective FAEE, after excluding those toxic to fish, was ethyl laurate (12:0). For both parasites, the LD50 was 18.75 µM within 250 min of incubation. Ethyl eicosapentaenoate (20:5n3) was the next most effective FAEE against G. turnbulli, and dihomo-γ-linolenic acid ethyl ester (20:3n6) and ethyl α-linolenate (18:3n3) were the next most effective against Trichodina sp. In addition, FAEEs prepared from the microalga Phaeodactylum tricornutum residue, after fucoxanthin extraction, were examined against Trichodina sp. infection in barramundi for the first time. LD85 and LD100 was achieved at 2.5 and 5 µL mL-1 of the FAEE preparation, respectively. In vivo, immersion of infected barramundi in 1.25 µL mL-1 of this preparation for 24 h reduced infection prevalence from 100% to 53% and was non-toxic to fish.

A new species of Urocleidoides Mizelle & Price, 1964 from the gills of Schizodon nasutus Kner, 1858 (Characiformes, Anostomidae) in southeastern Brazil.

Based on an integrative approach, this study describes a new species of Urocleidoides infesting Schizodon nasutus in the Paranapanema River basin, Brazil. The new species can be distinguished from its congeners by specific morphological features, including the shape of the male copulatory organ and accessory piece, the ventral bar shape, and the shape and size of the hooks. Molecular analyses, particularly of the 28S rDNA gene, suggest a close relationship between the new species and Urocleidoides paradoxus. The phylogenetic and taxonomic arrangement of Urocleidoides is discussed, as the analyses of the 28S rDNA and COI mtDNA resolved the genus as non-monophyletic, with Diaphorocleidus, Rhinoxenus, and Cacatuocotyle nested within it. Additional morphological and molecular data of other congeneric species are required to investigate the phylogenetic position and classification of Urocleidoides. This study underscores the significance of using integrative approaches in understanding host-parasite associations and phylogenetic relationships, contributing to the description of the freshwater fish parasite biodiversity in South America, particularly in the Paranapanema river basin.

Morphological and genetic identification of the gill monogenean parasite (Diclidophora merlangi) that infects Twobar Seabream Fish (Acanthopagrus bifasciatus) in the Arabian Gulf, Saudi Arabia.

Ectoparasites, particularly monogeneans, negatively affect fish health and growth. This study identified monogenean parasites in the twobar seabream, Acanthopagrus bifasciatus (Sparidae), inhabited the Arabian Gulf (Saudi Arabia). Following that, forty A. bifasciatus fish samples were visually examined for monogeneans. Parasite species were collected from the gills and then analyzed morphometrically, morphologically, and molecularly using the partial regions of the large subunit of ribosomal RNA (28S rRNA) and mitochondrial cytochrome C oxidase subunit I (COI) genes. Fish species were also identified using a DNA barcoding approach based on the COI gene. The monogenean species of Diclidophora merlangi (Diclidophoridae) were found in 45% of the fish species studied. The generic features of the Diclidophora genus distinguish this species. This species discriminated itself from congeners by having a muscular bulb with 17 grooved and recurved hooks, 218±10 (184-267) post-ovarian testes, and four pairs of pedunculated clamps of relative sizes. Partial 28S rRNA sequencing from monogeneans revealed that they grouped with members of the genus Diclidophora, forming a monophyletic group that supported the morphological descriptions. Molecular identification revealed that D. merlangi has a unique barcode made up of a COI sequence. The host identity was established as A. bifasciatus based on the COI gene sequences. Furthermore, a molecular phylogenetic study was performed to determine the phylogenetic affinity of parasite species and fish hosts. This study on Diclidophora species is considered the first record of this genus in the examined area.

An insight into the functional genomics and species classification of Eudiplozoon nipponicum (Monogenea, Diplozoidae), a haematophagous parasite of the common carp Cyprinus carpio.

BACKGROUND: Monogenea (Platyhelminthes, Neodermata) are the most species-rich class within the Neodermata superclass of primarily fish parasites. Despite their economic and ecological importance, monogenean research tends to focus on their morphological, phylogenetic, and population characteristics, while comprehensive omics analyses aimed at describing functionally important molecules are few and far between. We present a molecular characterisation of monogenean representative Eudiplozoon nipponicum, an obligate haematophagous parasite infecting the gills of the common carp. We report its nuclear and mitochondrial genomes, present a functional annotation of protein molecules relevant to the molecular and biochemical aspect of physiological processes involved in interactions with the fish hosts, and re-examinate the taxonomic position of Eudiplozoon species within the Diplozoidae family. RESULTS: We have generated 50.81 Gbp of raw sequencing data (Illumina and Oxford Nanopore reads), bioinformatically processed, and de novo assembled them into a genome draft 0.94 Gbp long, consisting of 21,044 contigs (N50 = 87 kbp). The final assembly represents 57% of the estimated total genome size (~ 1.64 Gbp), whereby repetitive and low-complexity regions account for ~ 64% of the assembled length. In total, 36,626 predicted genes encode 33,031 proteins and homology-based annotation of protein-coding genes (PCGs) and proteins characterises 14,785 (44.76%) molecules. We have detected significant representation of functional proteins and known molecular functions. The numbers of peptidases and inhibitors (579 proteins), characterised GO terms (16,016 unique assigned GO terms), and identified KEGG Orthology (4,315 proteins) acting in 378 KEGG pathways demonstrate the variety of mechanisms by which the parasite interacts with hosts on a macromolecular level (immunomodulation, feeding, and development). Comparison between the newly assembled E. nipponicum mitochondrial genome (length of 17,038 bp) and other diplozoid monogeneans confirms the existence of two distinct Eudiplozoon species infecting different fish hosts: Cyprinus carpio and Carassius spp. CONCLUSIONS: Although the amount of sequencing data and characterised molecules of monogenean parasites has recently increased, a better insight into their molecular biology is needed. The E. nipponicum nuclear genome presented here, currently the largest described genome of any monogenean parasite, represents a milestone in the study of monogeneans and their molecules but further omics research is needed to understand these parasites' biological nature.

Morphological and molecular characterization of Encotyllabe vallei Monticelli, 1907 (Monopisthocotylea, Monogenea) from the gilthead seabream Sparus aurata Linnaeus (Teleostei, Sparidae) from the southwestern Mediterranean and notes on host specificity of the genus Encotyllabe Diesing, 1850.

Incomplete original descriptions, the unavailability or poor conditions of specimens and the lack of detailed redescriptions have caused the validity of several species of the genus Encotyllabe Diesing, 1850 to be questioned. To date, seven of the recognized species were described upon one or two specimens, hindering study of intraspecific variations. This was made worse by considering few morphoanatomical differences sufficient to erect new species. Among Encotyllabe spp. occurring in Mediterranean waters, E. vallei was first described from the gilt-head bream Sparus aurata (Sparidae) off Italy. Although beautifully illustrated for a paper from that century, morphometric data for E. vallei from the type-host S. aurata remain unavailable. Previous records of E. vallei provided either morphometrical or molecular data, and its validity was questioned. We provide a redescription of E. vallei based on newly collected specimens from the S. aurata from the southwestern Mediterranean (off Algeria) using integrative taxonomy. Analysis of cox1 sequences of E. vallei from S. aurata, compared to sequences from other sparid hosts, mainly Pagellus bogaraveo, revealed a divergence not exceeding 2%, suggesting a stenoxenic specificity for this monogenean. Given that P. bogaraveo is the type-host for Encotyllabe pagelli, we were tempted to suggest a synonymy between E. vallei and E. pagelli. We refrained from doing so because E. pagelli was first described from the Atlantic coast off Brest, France. Morphological data for Encotyllabe from P. bogaraveo are warranted assessing the host specificity of E. vallei and whether there might be a species complex within individual sparid fish species.

Explosive networking: The role of adaptive host radiations and ecological opportunity in a species-rich host-parasite assembly.

Many species-rich ecological communities emerge from adaptive radiation events. Yet the effects of adaptive radiation on community assembly remain poorly understood. Here, we explore the well-documented radiations of African cichlid fishes and their interactions with the flatworm gill parasites Cichlidogyrus spp., including 10,529 reported infections and 477 different host-parasite combinations collected through a survey of peer-reviewed literature. We assess how evolutionary, ecological, and morphological parameters determine host-parasite meta-communities affected by adaptive radiation events through network metrics, host repertoire measures, and network link prediction. The hosts' evolutionary history mostly determined host repertoires of the parasites. Ecological and evolutionary parameters predicted host-parasite interactions. Generally, ecological opportunity and fitting have shaped cichlid-Cichlidogyrus meta-communities suggesting an invasive potential for hosts used in aquaculture. Meta-communities affected by adaptive radiations are increasingly specialised with higher environmental stability. These trends should be verified across other systems to infer generalities in the evolution of species-rich host-parasite networks.

Impact of high temperature, CO2 and parasitic infection on inflammation, immunodepression and programmed cell death in Colossoma macropomum at the transcriptional level.

The production of tambaqui Colossoma macropomum has recently reached a milestone, being considered the main native species produced in South American continental waters. Despite the importance of this fish, its immunity is poorly understood, and global warming could pose severe risks to its health as increasing water temperature leads to an increase in the incidence of parasitic diseases. In an experimental context based on the high-emission scenario of the 5th Intergovernmental Panel on Climate Change (IPCC) report, we evaluated the synergistic effect of exposure to the extreme climate change scenario (RCP8.5) during two exposure periods (7 and 30 days) and two levels of parasitism by monogeneans (low and high). The goal was to understand how the tambaqui immune system will react to this challenge. To achieve this goal, we analyzed the expression of nine immunity-related genes (jak3, stat3, il-10, socs1, casp1, il-1ß, tp53, bcl2, and hif-1α) in the spleen. Our main findings showed downregulation in the jak3/stat3 pathway, genes related to the control of inflammation and apoptosis, in addition to upregulation of proinflammatory genes and those related to pyroptosis during the first 7 days of exposure to the extreme climate scenario, also indicating a stage of immunodepression in these animals. After 30 days of exposure, all genes tended to return to similar levels in the current scenario, possibly due to the decrease in parasite load caused by chronic exposure to the extreme scenario. Our data strongly suggest that the increase in parasitism intensity caused by the extreme climate change scenario is responsible for disturbances in the host's immune system. However, more studies are needed to clarify this poorly understood cascade of events.

Effects of different regimes of low temperature on egg hatching of Dactylogyrus vastator (Monogenea: Dactylogyridae).

The development of dactylogyrids is dependent on water temperature, and their eggs fail to hatch below 5 °C. In the field, however, mean abundance of Dactylogyrus species increases and reaches a high level in winter, which suggests that infective oncomiracidia hatch from eggs in winter. Therefore, the effect of low water temperature on in vitro egg hatching of D. vastator was determined in laboratory. D. vastator hatching success was 65.3%, 62.7%, 42.6% and 22.3% when eggs were firstly incubated for 0, 7, 14 and 21 days at 5 °C and then consecutively maintained for 15 days at 20 °C. When eggs were directly incubated at 5 °C, eggs failed to hatch within one month. However, hatching success was 69.8% and 66.7%, respectively, when maintained at 5 °C after 12 and 24 h incubation at 20 °C. The results suggested that egg incubation for more than 1 week at 5 °C had significant impacts on hatching success of D. vastator subsequently maintained at 20 °C. But low temperature (5 °C) had no effect on hatching success when eggs were firstly exposed to room temperature (20 °C) for one day.

Light and scanning electron microscopy of the effects of Macrogyrodactylus congolensis (Prudhoe, 1957) on the skin of the African sharptooth catfish Clarias gariepinus (Burchell, 1822).

Clarias gariepinus (Burchell, 1822) is one of the two most actively cultured freshwater fish in Africa and therefore, economically important. Specimens of this species were purchased from a fish farm near Hartbeespoort Dam (North West, South Africa) and introduced into the tanks of the research aquarium in the Department of Zoology at the University of Johannesburg. However, the skin of these fish was infected with Macrogyrodactylus congolensis (Prudhoe, 1957), which proliferated profusely in the favourable conditions of the aquarium, posing a potential threat to its host. The current study was aimed at examining the pathology caused by M. congolensis on the skin of C. gariepinus. Species identification of the parasite was confirmed using light microscopy (LM), scanning electron microscopy (SEM) and DNA barcoding of the internal transcribed spacer region. Examination of the pathology was studied using LM of haematoxylin and eosin-stained sections (epoxy embedded) and SEM of parasites attached to the hosts' skin. Infected skin exhibited excessive mucus production, corroborated by an increased number of mucus cells alongside proliferated and abnormally enlarged club cells, resulting in varying thickness of the epidermal layer. At the site of attachment, the basement membrane detached from the dermis. Hamulus points and marginal hooks of the parasite pierce through the hosts' skin resulting in tearing. Epidermal cells and melanin granules were observed in the intestinal lumen of the parasite. Melanin granules were absorbed by the parasite's intestinal epithelium confirming that the parasite feeds on host skin tissue.

Molecular (cox1), geographical, and host record investigation of monogeneans Mazocraes australis (Mazocraeidae), Polylabris sillaginae, and P. australiensis (Microcotylidae).

This study determines the occurrence and molecular characterisation of Monogenea from three commercially important Australian fish: Australian sardine Sardinops sagax (Jenyns), Australian anchovy Engraulis australis (White), and eastern school whiting Sillago flindersi McKay. Earlier studies have provided only morphological species identification, whereas this study combines both morphological and molecular methods. A total of 247 fish across 3 species, sourced from the New South Wales and Victorian coasts, were examined for Monogenea. A total of 187 monogenean parasites were recovered from the gills. The overall prevalence, mean intensity, and mean abundance were 34%, 2.23, and 0.78, respectively. The parasites were initially classified morphologically as three species across two families. Family Mazocraeidae was represented by Mazocraes australis Timi et al. J Parasitol 85:28-32, 1999, and family Microcotylidae by Polylabris sillaginae (Woolcock, Parasitology 28:79-91, 1936) Dillon, Hargis, and Harrises, 1983 and P. australiensis Hayward, 1996. Molecular identification of parasites was conducted through sequencing of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. The fish hosts in the present study were also barcoded (mitochondrial cox1 gene) to confirm specific identities. There was no comparable cox1 sequence available in GenBank for the parasites found in the present study. However, the phylogenetic tree clustered the monogenean species identified in this study according to their familial groups of Mazocraeidae and Microcotylidae. The presence of M. australis on E. australis and S. sagax was confirmed in this study. Polylabris australiensis was only found on S. sagax but Si. flindersi was found to be a host for both Polylabris species. This study is the first to explore the mitochondrial cox1 genes of these three-monogenean species. These findings will serve as a foundation for future monogenean research in Australian waters and elsewhere.

Under the radar: co-introduced monogeneans (Polyopisthocotylea: Gastrocotylinea) of the invasive fish Scomberomorus commerson in the Mediterranean Sea.

The Levant Basin is in many ways the world's most invaded marine ecosystem owing to the existence of the man-made Suez Canal. The invasion of free-living organisms through this pathway is increasingly documented and monitored in the past two decades, and their ecological impact recognized. Nonetheless, while tremendous scientific effort is invested in documenting introduced fishes, co-introduction events of these fishes and their parasites have drawn relatively little interest. In our research, we examined the presence of gill parasites (Monogenea) on the invasive narrow barred Spanish mackerel Scomberomorus commerson which has been known in the Mediterranean Sea for 80 years. The gills of S. commerson supported numerous, relatively large monogeneans (Monogenea: Gastrocotylinea), reaching prevalence levels of 100% with a mean intensity of ~ 80 worms per host. Using an integrated molecular and morphological approach, four gastrocotylinean species were identified: Gotocotyla acanthura, Cathucotyle cathuaui, Pricea multae, and Pseudothoracocotyla ovalis. Two species, C. cathuaui and P. ovalis, are reported here for the first time from the Mediterranean. Sequences of the 28S rRNA gene of G. acanthura from native hosts, Pomatomus saltatrix and Trachinotus ovatus, differed from individuals collected from S. commerson by 1.8%. We therefore suggest that the taxonomic status and distribution of G. acanthura should be revisited, and we recommend an integrated approach as essential to accurately detect co-introductions.

Occurrence and molecular identification of Monogenea from blue-spotted flathead Platycephalus caeruleopunctatus (Scorpaeniformes: Platycephalidae) in Australian waters.

This study describes the occurrence and molecular identification of Monogenea from blue-spotted flathead Platycephalus caeruleopunctatus (McCulloch) (Scorpaeniformes: Platycephalidae) from waters off the NSW coast, Australia. Platycephalus spp. are favoured by consumers for delicate, white, mild flavoured flesh and therefore are commercially important species within Australia. Platycephalus spp. are also extensively targeted by Australian recreational fishers. There has been no previous study that has exclusively focused on Pl. caeruleopunctatus in Australia or globally. Although a single study by Dillon (1985), of monogeneans infecting Platycephalus spp. from Australian waters, identified Microcotyle bassensis Murray, 1931 in Pl. caeruleopunctatus. The present study combines both morphological and molecular methods to identify both host and parasites. A total of 116 fish, sourced from the waters off the coast of New South Wales, Australia, were examined. A total of 1498 Monogenea were recovered from the gills. The overall prevalence, mean intensity, and mean abundance were 72%, 18.05, and 12.91, respectively. Monogenea were initially classified morphologically as two different species M. bassensis (family: Microcotylidae) and Platycephalotrema bassense (Hughes, 1928) Kritsky & Nitta, 2019 (family: Ancyrocephalidae). Molecular identification of Monogenea was conducted through sequencing of their mitochondrial cytochrome c oxidase subunit 1 (cox1) and nuclear 28S genes. The specific identification of host Pl. caeruleopunctatus was confirmed through sequencing the cox1 gene. There was no comparable sequence for cox1 and 28S genes available in GenBank for the monogenean species found in the present study. Only a single sequence (obtained from the nuclear ITS2-rDNA) was deposited in GenBank for M. bassensis. However, the phylogenetic analyses of mitochondrial and nuclear sequences revealed that the identified Monogenea clustered according to their familial groups. Platycephalotrema bassense was identified for the first time in Pl. caeruleopunctatus in the present study. This study has provided the first evidence for the exploration of both cox1 and 28S sequences of all Monogenea. The findings of this study serve as a foundation for future monogenean research on other Platycephalus spp. from Australian waters.

Morphological and genetic variability of the cryptic Gyrodactylus sphinx and Gyrodactylus gerasevi n. sp. (Platyhelminthes: Monogenea) from the Mediterranean Sea and Black Sea: two new members of the cross-ocean distributed Gyrodactylus orecchiae species group.

Gyrodactylus sphinx Dmitrieva & Gerasev, 2000 is the only species of Gyrodactylus originally described from Aidablennius sphynx (Valenciennes) in the Black Sea. In the present study, monogeneans similar to G. sphinx are reported from the same host and from two other species of Blenniidae from the Black Sea, as well as from the Mediterranean Sea. This study aims to verify the taxonomic status of the specimens found in different hosts and localities, other than the type ones of G. sphinx. Twenty-two measurements of the haptoral structures of 169 gyrodactylids were used for the morphological study. Morphometric variability between different samples was analysed using analysis of variance, multivariate analysis of variance (MANOVA) and principal component analysis (PCA). Molecular studies were carried out using the nuclear internal transcribed spacer 2 and 5.8S ribosomal DNA regions. Network, Bayesian phylogenetic and species delimitation analyses were performed to infer the number of taxonomic units and the phylogeographic relationships occurring within and among them. MANOVA revealed a significant dependence of the morphometry of hamuli and marginal hooks on host species and regions, but a clear differentiation between samples was not confirmed by PCA. Moreover, the ranges of all dimensions overlapped between samples. However, molecular analyses suggested the occurrence of at least two taxonomic entities. The most common entity was present in individuals of the Black and Mediterranean seas, and is described here as Gyrodactylus gerasevi n. sp., whereas a second entity recognized as a G. sphinx was found only in individuals from two localities off Crimea. The monophyletic cluster grouping of these two species was placed within a large clade that also included a separate sister cluster with seven other species of the Gyrodactylus orecchiae cross-ocean species group.

First morphological and phylogenetic data on Ligophorus kaohsianghsieni (Platyhelminthes: Monogenea) from the Black Sea and the Sea of Japan and molecular evidence of deep divergence of sympatric Ligophorus species parasitizing Planiliza haematocheilus.

Ligophorus kaohsianghsieni (Gusev, ) Gusev, was collected from the so-iuy mullet Planiliza haematocheilus (Temminck & Schlegel, 1845) from the Black Sea and the Sea of Japan. DNA sequences data for L. kaohsianghsieni, as well as its morphological characters from the Sea of Japan were obtained for the first time. Significant morphometric and genetic diversity between specimens of L. kaohsianghsieni from the Black-Azov Sea region and the Sea of Japan were not found. For the first time, the molecular phylogeny of L. kaohsianghsieni based on three fragments of the nuclear DNA ribosomal cluster (18S, internal transcribed spacer 1 and 28S) was reconstructed. Molecular analysis of Ligophorus species from the Atlantic and Pacific Oceans revealed a significant phylogenetic distance between L. kaohsianghsieni and two others species (Ligophorus pilengas and Ligophorus llewellyni) from the same host (P. haematocheilus) and region. This result indicates the lack of correspondence between the phylogenetic and geographical closeness of the hosts and the relation of their parasites from the genus Ligophorus.

Checklist of The Monogenea (Platyhelminthes) Parasitic in Tunisian Aquatic Vertebrates.

153 species of monogeneans have been recorded in Tunisian aquatic vertebrates (89 hosts). A list of these species with hosts is presented. A comparison of the Monogenea diversity off the coast of Tunisia with other regions of the Mediterranean Sea and the world is provided. The number of parasites depends on the number of hosts examined and their diversity in the region. This list shows that Monopisthocotylea is the richest group. In addition, new records have been reported during a survey of the diversity of monogeneans fish in the southern coast of Tunisia (Gulf of Gabes) including: Benedenia monticellii (Parona and Perugia, 1895) Johnston, 1929, Lamellodiscus bidens Euzet, 1984, Lamellodiscus confusus Amine, Euzet & Kechemir-Issad, 2007, Lamellodiscus ergensi Euzet & Oliver, 1966, Lamellodiscus hilii Euzet, 1984, Lamellodiscus impervius Euzet, 1984, Lamellodiscus knoepffleri Oliver, 1969, Lamellodiscus theroni Amine, Euzet & Kechemir-Issad, 2007, Ligophorus acuminatus Euzet & Suriano, 1977, Ligophorus angustus Euzet & Suriano, 1977, Ligophorus heteronchus Euzet & Suriano, 1977, Ligophorus macrocolpos Euzet & Suriano, 1977, Ligophorus minimus Euzet & Suriano, 1977, Capsala maccallumi Price, 1939 and Pseudanthocotyloides heterocotyle (Van Beneden, 1871) Euzet & Prost, 1969.

Eudiplozoon nipponicum (Monogenea, Diplozoidae) and its adaptation to haematophagy as revealed by transcriptome and secretome profiling.

BACKGROUND: Ectoparasites from the family Diplozoidae (Platyhelminthes, Monogenea) belong to obligate haematophagous helminths of cyprinid fish. Current knowledge of these worms is for the most part limited to their morphological, phylogenetic, and population features. Information concerning the biochemical and molecular nature of physiological processes involved in host-parasite interaction, such as evasion of the immune system and its regulation, digestion of macromolecules, suppression of blood coagulation and inflammation, and effect on host tissue and physiology, is lacking. In this study, we report for the first time a comprehensive transcriptomic/secretome description of expressed genes and proteins secreted by the adult stage of Eudiplozoon nipponicum (Goto, 1891) Khotenovsky, 1985, an obligate sanguivorous monogenean which parasitises the gills of the common carp (Cyprinus carpio). RESULTS: RNA-seq raw reads (324,941 Roche 454 and 149,697,864 Illumina) were generated, de novo assembled, and filtered into 37,062 protein-coding transcripts. For 19,644 (53.0%) of them, we determined their sequential homologues. In silico functional analysis of E. nipponicum RNA-seq data revealed numerous transcripts, pathways, and GO terms responsible for immunomodulation (inhibitors of proteolytic enzymes, CD59-like proteins, fatty acid binding proteins), feeding (proteolytic enzymes cathepsins B, D, L1, and L3), and development (fructose 1,6-bisphosphatase, ferritin, and annexin). LC-MS/MS spectrometry analysis identified 721 proteins secreted by E. nipponicum with predominantly immunomodulatory and anti-inflammatory functions (peptidyl-prolyl cis-trans isomerase, homolog to SmKK7, tetraspanin) and ability to digest host macromolecules (cathepsins B, D, L1). CONCLUSIONS: In this study, we integrated two high-throughput sequencing techniques, mass spectrometry analysis, and comprehensive bioinformatics approach in order to arrive at the first comprehensive description of monogenean transcriptome and secretome. Exploration of E. nipponicum transcriptome-related nucleotide sequences and translated and secreted proteins offer a better understanding of molecular biology and biochemistry of these, often neglected, organisms. It enabled us to report the essential physiological pathways and protein molecules involved in their interactions with the fish hosts.

Morphological and molecular characterization of Ameloblastella pirarara sp. n. (Monogenoidea: Dactylogyridae) parasitizing the large Amazonian catfish Phractocephalus hemioliopterus.

In this study, integrative taxonomy is applied to describe a new dactylogyrid species, Ameloblastella pirarara sp. n. from the gills of Phractocephalus hemioliopterus, a commercially and ecologically important Amazonian catfish. Ameloblastella pirarara sp. n. can be distinguished from its congeners mainly by the morphology of the male copulatory organ (MCO), accessory piece, and anchors. The new species most resembles Ameloblastella unapi, from the Peruvian Amazon, but differs from it by the number of MCO rings, morphology of the vaginal canal and sclerotized structures of the haptor. Phylogenetic analyses based on sequences of the partial 28S rDNA (D1-D2 domains) gene placed the new species in a well-supported subclade of Ameloblastella spp. parasites of Neotropical siluriform fish, as a sister taxon to Ameloblastella unapioides. Thus, the new species described herein expands our knowledge of the diversity of monogenoid parasites from Amazonian freshwater fish.

Molecular communication between the monogenea and fish immune system.

Monogeneans parasitise mainly the outer structures of fish, such as the gills, fins, and skin, that is, tissues covered with a mucous layer. While attached by sclerotised structures to host's surface, monogeneans feed on its blood or epidermal cells and mucus. Besides being a rich source of nutrients, these tissues also contain humoral immune factors and immune cells, which are ready to launch defence mechanisms against the tegument or gastrointestinal tract of these invaders. The exploitation of hosts' resources by the Monogenea must, therefore, be accompanied by suppressive and immunomodulatory mechanisms which protect the parasites against attacks by host immune system. Elimination of hosts' cytotoxic molecules and evasion of host immune response is often mediated by proteins secreted by the parasites. The aim of this review is to summarise existing knowledge on fish immune responses against monogeneans. Results gleaned from experimental infections illustrate the various interactions between parasites and the innate and adaptive immune system of the fish. The involvement of monogenean molecules (mainly inhibitors of peptidases) in molecular communication with host immune system is discussed.

Gastrodermis ultrastructure of the different life stages of the polyopisthocotylean monogenean gill parasite Discocotyle sagittata.

The polyopisthocotylean Discocotyle sagittata is a blood-feeding monogenean that infects the gill lamellae of rainbow trout, Oncorhynchus mykiss, and brown trout, Salmo trutta. The ultrastructure of their alimentary tract, at different stages of the life cycle, was previously unknown. Here, we show that the gastrodermis of the oncomiracidium, subadult, and adult D. sagittata follows the same structural organization as that of other blood-feeding polyopisthocotyleans, being composed of digestive cells alternating with a connecting syncytium. Digestive cells of the oncomiracidium are found in three developmental forms: undifferentiated, developing differentiated, and differentiated (presumably functioning) cells whereas those of adult and subadult are present in a single functioning state with variable size and content. The apical cytoplasm of adult digestive cells forms conical outgrowths, a feature which is absent in the oncomiracidium. The connecting syncytium of the oncomiracidium has no evidence of metabolic activity, while that of adult and subadult is metabolically active. The lamellae of the connecting syncytium of adults and subadults are more numerous and larger, and their terminal portions are expanded, compared with those of the oncomiracidium. Parallel, tubular, membranous structures are characteristic of the apical cytoplasm of the connecting syncytium of the oncomiracidium. Luminal lamella in the oncomiracidium, subadult, and adult form balloon-like structures enclosing some luminal contents, but those of the oncomiracidium are larger, bounded by nucleated cytoplasmic layer, and enclose more luminal contents. The possible functions of these structures and mechanism of digestion in both oncomiracidium and adult are discussed.

Ultrastructural observations on the oncomiracidium epidermis and adult tegument of Discocotyle sagittata, a monogenean gill parasite of salmonids.

During their different life stages, parasites undergo remarkable morphological, physiological, and behavioral "metamorphoses" to meet the needs of their changing habitats. This is even true for ectoparasites, such as the monogeneans, which typically have a free-swimming larval stage (oncomiracidium) that seeks out and attaches to the external surfaces of fish where they mature. Before any obvious changes occur, there are ultrastructural differences in the oncomiracidium's outer surface that prepare it for a parasitic existence. The present findings suggest a distinct variation in timing of the switch from oncomiracidia epidermis to the syncytial structure of the adult tegument and so, to date, there are three such categories within the Monogenea: (1) Nuclei of both ciliated cells and interciliary cytoplasm are shed from the surface layer and the epidermis becomes a syncytial layer during the later stages of embryogenesis; (2) nuclei of both ciliated cells and interciliary syncytium remain distinct and the switch occurs later after the oncomiracidia hatch (as in the present study); and (3) the nuclei remain distinct in the ciliated epidermis but those of the interciliary epidermis are lost during embryonic development. Here we describe how the epidermis of the oncomiracidium of Discocotyle sagittata is differentiated into two regions, a ciliated cell layer and an interciliary, syncytial cytoplasm, both of which are nucleated. The interciliary syncytium extends in-between and underneath the ciliated cells and sometimes covers part of their apical surfaces, possibly the start of their shedding process. The presence of membranous whorls and pyknotic nuclei over the surface are indicative of membrane turnover suggesting that the switch in epidermis morphology is already initiated at this stage. The body tegument and associated putative sensory receptors of subadult and adult D. sagittata are similar to those in other monogeneans.