Modulatory actions of Echinococcus granulosus antigen B on macrophage inflammatory activation.

Cestodes use own lipid-binding proteins to capture and transport hydrophobic ligands, including lipids that they cannot synthesise as fatty acids and cholesterol. In E. granulosus s.l., one of these lipoproteins is antigen B (EgAgB), codified by a multigenic and polymorphic family that gives rise to five gene products (EgAgB8/1-5 subunits) assembled as a 230 kDa macromolecule. EgAgB has a diagnostic value for cystic echinococcosis, but its putative role in the immunobiology of this infection is still poorly understood. Accumulating research suggests that EgAgB has immunomodulatory properties, but previous studies employed denatured antigen preparations that might exert different effects than the native form, thereby limiting data interpretation. This work analysed the modulatory actions on macrophages of native EgAgB (nEgAgB) and the recombinant form of EgAg8/1, which is the most abundant subunit in the larva and was expressed in insect S2 cells (rEgAgB8/1). Both EgAgB preparations were purified to homogeneity by immunoaffinity chromatography using a novel nanobody anti-EgAgB8/1. nEgAgB and rEgAgB8/1 exhibited differences in size and lipid composition. The rEgAgB8/1 generates mildly larger lipoproteins with a less diverse lipid composition than nEgAgB. Assays using human and murine macrophages showed that both nEgAgB and rEgAgB8/1 interfered with in vitro LPS-driven macrophage activation, decreasing cytokine (IL-1ß, IL-6, IL-12p40, IFN-ß) secretion and ·NO generation. Furthermore, nEgAgB and rEgAgB8/1 modulated in vivo LPS-induced cytokine production (IL-6, IL-10) and activation of large (measured as MHC-II level) and small (measured as CD86 and CD40 levels) macrophages in the peritoneum, although rEgAgB8/1 effects were less robust. Overall, this work reinforced the notion that EgAgB is an immunomodulatory component of E. granulosus s.l. Although nEgAgB lipid's effects cannot be ruled out, our data suggest that the EgAgB8/1 subunit contributes to EgAgB´s ability to regulate the inflammatory activation of macrophages.

Larvae trypanorhynch (Cestoda) infecting the dusky flounder, Syacium papillosum (Paralichthyidae: Pleuronectiformes) in the continental shelf of the Yucatan Peninsula, Mexico.

Trypanorhynch cestode larvae were found parasitizing specimens of dusky flounder Syacium papillosum (Linnaeus, 1758) in the Southeastern Gulf of Mexico (s-GoM). Plerocercoids were recovered from a total of 194 flatfish, embedded in the intestine and stomach wall. Trypanorhynch were identified using morphology and a molecular phylogeny using newly sequenced partial 28S (region D1-D3) ribosomal DNA in combination with data derived from other species of trypanorhynchs available from GenBank. Larvae representing three genera of trypanorhynch cestodes, Nybelinia Poche, 1926; Kotorella Euzet & Radujkovic, 1989 and Oncomegas Dollfus, 1929 were found in dusky flounder specimens in the s-GoM. These plerocercoids represent six species: Nybelinia sp. 1, Nybelinia sp. 2, Nybelinia sp. 3, Nybelinia sp. 4, Kotorella pronosoma (Stossich, 1901) and Oncomegas wageneri (Linton, 1890) Dollfus, 1929. All cestode specimens in this study represent new locality records for their respective species.

Thermal tolerance and survival are modulated by a natural gradient of infection in differentially acclimated hosts.

Wild ectotherms are exposed to multiple stressors, including parasites, that can affect their responses to environmental change. Simultaneously, unprecedented warm temperatures are being recorded worldwide, increasing both the average and maximum temperatures experienced in nature. Understanding how ectotherms, such as fishes, will react to the combined stress of parasites and higher average temperatures can help predict the impact of extreme events such as heat waves on populations. The critical thermal method (CTM), which assesses upper (CTmax) and lower (CTmin) thermal tolerance, is often used in acclimated ectotherms to help predict their tolerance to various temperature scenarios. Despite the widespread use of the CTM across taxa, few studies have characterized the response of naturally infected fish to extreme temperature events or how acute thermal stress affects subsequent survival. We acclimated naturally infected pumpkinseed sunfish (Lepomis gibbosus) to four ecologically relevant temperatures (10, 15, 20 and 25°C) and one future warming scenario (30°C) for 3 weeks before measuring CTmax and CTmin. We also assessed individual survival the week following CTmax. Parasites were counted and identified following trials to relate infection intensity to thermal tolerance and survival. Interestingly, trematode parasites causing black spot disease were negatively related to CTmax, suggesting that heavily infected fish are less tolerant to acute warming. Moreover, fish infected with yellow grub parasites showed decreased survival in the days following CTmax implying that the infection load has negative survival consequences on sunfish during extreme warming events. Our findings indicate that, when combined, parasite infection and high prolonged average temperatures can affect fish thermal tolerance and survival, emphasizing the need to better understand the concomitant effects of stressors on health outcomes in wild populations. This is especially true given that some parasite species are expected to thrive in warming waters making host fish species especially at risk.

New species of Echinobothrium van Beneden, 1849 (Cestoda: Diphyllidea) from Indo-Pacific maskrays (Neotrygon Castelnau) off Australia and Borneo.

Four new diphyllidean species of the genus Echinobothrium van Beneden, 1849 are described from Indo-Pacific maskrays (Neotrygon Castelnau, Dasyatidae). Echinobothrium giraffaeous sp. n. from Neotrygon leylandi (Last) off northern Australia, Echinobothrium ivanovae sp. n. from Neotrygon orientalis Last, White et Serét off Borneo, and Echinobothrium bethae sp. n. from Neotrygon varidens (Garman) off Borneo are distinguished from all but one of the 33 valid species of the genus by the possession of the outermost A hooks with an extended base into which the bases of the three outermost B hooks are inserted. Echinobothrium rhynchobati (Khalil et Abdul-Salam, 1989) is the only known species with this unique feature, but its rostellum has a system of interlocking knobs and sockets that articulate bases of the A and B type hooks with one another, which is not present in any of the newly described species. Echinobothrium tyleri sp. n. from Neotrygon australiae Last, White et Serét off northern Australia is distinguished from all known species of Echinobothrium by its unique rostellar hook formula {2-3 18/17 2-3}. With the present addition of four new species, the central Indo-Pacific realm becomes the major hotspot for Echinobothrium, from which 13 species have been reported.

First Report of Intestinal Cestode Infection in a Berylline Hummingbird (Saucerottia beryllina) in Mexico.

BACKGROUND: Up to now, five cestode species have been reported infecting five hummingbird species. To date, there have been no reports of cestode infections in hummingbirds in Mexico. METHODS: A Berylline hummingbird (Saucerottia beryllina) was found dead in a backyard at Toluca City, Mexico, and a window collision was assumed as the cause of death. The bird was preserved in 10% neutral buffered formalin for routine histological examination. RESULTS: At the histological study, liver parenchymal disruption was observed. This lesion could be the result of the assume collision. No lesions were observed in other tissues examined. Conspicuous cestode structures were observed in the lumen of the small intestine. Structure of cestodes, as revealed from histological sections, suggests their position in the genus Anonchotaenia Cohn, 1900 (family Paruterinidae). CONCLUSION: This is the first report of intestinal cestodosis in a Berylline hummingbird (S. beryllina) in Mexico.

Integrative taxonomy of metazoan parasites of the bluntnose sixgill shark Hexanchus griseus (Bonnaterre, 1788) in the Mediterranean Sea, with the resurrection of Grillotia acanthoscolex Rees, 1944 (Cestoda: Trypanorhyncha).

Appropriate diagnoses of parasites of apex marine predators are crucial to understand their biodiversity, host specificity, biogeography, and life cycles. Such diagnoses are also informative of ecological and biological characteristics of both host and environment in which the hosts and their parasites live. We here (i) investigate the parasite fauna of a bluntnose sixgill shark Hexanchus griseus (Bonnaterre, 1788) obtained from the Gulf of Naples (Tyrrhenian Sea), (ii) characterize molecularly all its metazoan parasites, and (iii) resurrect and report the main morphological features and phylogenetic position of Grillotia acanthoscolex, a cestode species previously synonymized with Grillotia adenoplusia. A rich parasite fauna represented by eight different taxa was found, including two monogeneans (Protocotyle grisea and Protocotyle taschenbergi), one digenean (Otodistomum veliporum), four cestodes (Crossobothrium dohrnii, Clistobothrium sp., G. acanthoscolex, and G. adenoplusia), and one copepod (Protodactylina pamelae). Sequencing of these samples accounts for an important molecular baseline to widen the knowledge on the parasitic fauna of bluntnose sixgill sharks worldwide and to reconstruct their correct food chains. The bluntnose sixgill shark was found to be a definitive host for all endoparasites found here, confirming that it occupies an apex trophic level in the Mediterranean Sea. The taxa composition of the trophic parasite fauna confirms that the bluntnose sixgill shark mostly feeds on teleost fish species. However, the occurrence of two phillobothrid cestodes (C. dohrnii and Clistobothrium sp.) suggests that it also feeds on squids. Finally, we emphasize the importance of using integrative taxonomic approaches in the study of parasites from definitive and intermediate hosts to elucidate biology and ecology of taxa generally understudied in the Mediterranean Sea.

Corrigendum: Trophic diversification and parasitic invasion as ecological niche modulators for gut microbiota of whitefish.

[This corrects the article DOI: 10.3389/fmicb.2023.1090899.].

Manual for the study of tapeworms (Cestoda) parasitic in ray-finned fish, amphibians and reptiles.

Based on long-term and often frustrating experiences with the poor quality of tapeworms (Cestoda) collected throughout the world for taxonomic and phylogenetic studies, and considering the increasing obstacles to obtaining new material, a simple, easy-to-use and illustrated methodological guide (manual) is provided. It focusses mainly on key steps in examining hosts, collecting cestodes from poikilothermous vertebrates except elasmobranchs, i.e., from ray-finned fish (Actinopterygii), amphibians and 'reptiles' (a paraphyletic group comprising all sauropsids except birds), and fixing them for subsequent morphological and molecular study. It is proposed that the following methodological points should be followed: (i) ideally only freshly euthanised hosts (not previously frozen) should be used for parasitological examination; (ii) hosts examined should be documented by photographs; host tissue should also be preserved for future genotyping if necessary; (iii) tapeworms should be detached carefully to keep the scolex intact and properly cleaned before fixation; (iv) a small piece of cestode tissue should be always preserved in molecular grade ethanol for DNA sequencing; (v) tapeworms should be fixed as quickly as possible after collecting them and while they are still alive, always using hot (heated) fixatives; this prevents unnatural contraction or deformation and ensures uniform fixation; (vi) each sample (vial) should be properly labelled (a unique code should be given to every cestode sample); (vii) vouchers of sequenced specimens (hologenophores or paragenophores) should always be preserved for identification, and deposited in internationally recognised collections. It is hoped that this guide helps researchers and students to properly process valuable material of cestodes to make it suitable for reliable identification including genotyping and comparative anatomy, which is a prerequisite for any subsequent ecological, biogeographical, phylogenetic life cycle or molecular study.

Metapopulation Structure of Two Species of Pikeworm (Triaenophorus, Cestoda) Parasitizing the Postglacial Fish Community in an Oligotrophic Lake.

In the present study, we estimated the levels of infestation of the main fish species that are hosts for two Triaenophorus species: T. crassus and T. nodulosus. The prevalence of T. crassus and T. nodulosus infestations in the intestine of their definitive host-pike Esox lucius was similar (71.0% and 77.4%, respectively). At the same time, the prevalence of T. crassus infestation in muscle tissue was significantly different between the second intermediate hosts, Coregonus lavaretus pidschian (31.4%) and Cor. l. pravdinellus (91.2%), due to considerable differences in their diets. For T. nodulosus, we found significant variations in the levels of prevalence among the second intermediate hosts-100% for Lota lota, 81.8% for Cottus sibiricus 31.9% for Thymallus arcticus, and 24.5% for Perca fluviatilis-that we also explained using different diets. Moreover, analysis of the symmetry of parasite infestations did not reveal any asymmetry between the number of cysts in the left and right body surfaces of the "planktivorous" form/species of whitefish, whereas in the ''benthivorous", an asymmetry of parasite infestations was found.

microRNA silencing in a whole worm cestode model provides insight into miR-71 function.

Parasites belonging to the class Cestoda include zoonotic species such as Echinococcus spp. and Taenia spp. that cause morbidity and mortality in endemic areas, mainly affecting pastoral and rural communities in low income countries but also upper middle income countries. Cestodes show remarkable developmental plasticity, implying tight regulation of gene expression throughout their complex life cycles. Despite the recent availability of genomic data for cestodes, little progress was made on postgenomic functional studies. MicroRNAs (miRNAs) are key components of gene regulatory systems that guide diverse developmental processes in multicellular organisms. miR-71 is a highly expressed miRNA in cestodes, which is absent in vertebrates and targets essential parasite genes, representing a potential key player in understanding the role of miRNAs in cestodes biology. Here we used transfection with antisense oligonucleotides to perform whole worm miRNA knockdown in tetrathyridia of Mesocestoides vogae (syn. Mesocestoides corti), a laboratory model of cestodes. We believe this is the first report of miRNA knockdown at the organism level in these parasites. Our results showed that M. vogae miR-71 is involved in the control of strobilation in vitro and in the establishment of murine infection. In addition, we identified miR-71 targets in M. vogae, several of them being de-repressed upon miR-71 knockdown. This study provides new knowledge on gene expression regulation in cestodes and suggests that miRNAs could be evaluated as new selective therapeutic targets for treating Neglected Tropical Diseases prioritised by the World Health Organization.

ADULT TAPEWORM (PLATYHELMINTHES: CESTODA) PARASITES OF NORTH AMERICAN HERPETOFAUNA: CHECKLIST OF SPECIES AND IDENTIFICATION KEY TO FAMILIES AND GENERA.

An updated checklist of adult tapeworms (Platyhelminthes: Cestoda) that parasitize wild North American amphibians and reptiles is presented: A total of 58 species grouped in 15 genera, 5 families, and 3 orders, are registered; these infect a total of 90 species of reptiles and 88 species of amphibians in the region. An illustrated identification key for the families and genera listed is proposed.

Identification and phylogenetic analysis of Taenia spp. parasites found in wildlife in the Emilia-Romagna region, northern Italy (2017-2022).

The recent expansion of the habitat of several wildlife species, comprising anthropized areas, is a relevant risk factor for many zoonotic diseases and should be considered in national and regional sanitary monitoring systems. We evaluated adult intestinal Taenia spp. parasites isolated from wild carnivores and cystic larval forms isolated from wild mammals analysed at the Istituto Zooprofilattico Sperimentale della Lombardia ed Emilia-Romagna (IZSLER) as part of the regional wildlife sanitary surveillance plan. Then, we assessed parasite species through molecular analysis (multiplex PCR followed by ribosomal 12S subunit gene sequencing) in order to update the epidemiological situation on Taeniids infection in the Emilia-Romagna wildlife, reporting the prevalence of each isolated species. The most commonly isolated species was Taenia serialis, which we detected in both wolves and foxes as definitive hosts and in roe deer as intermediate host. More attention on the distribution of Taeniids in wildlife should be paid, considering their potential zoonotic role: several Taenia spp. (Taenia solium, Taenia multiceps, Taenia serialis, Taenia brauni, Taenia glomerulatus) are known for causing coenurosis in humans, with possible severe or fatal outcomes.

Helminths in Invasive Raccoons (Procyon lotor) from Southwest Germany.

As hosts of numerous zoonotic pathogens, the role of raccoons needs to be considered in the One Health context. Raccoons progressively expand their range as invasive alien species in Europe. This study aimed to investigate the intestinal helminth fauna of raccoons in Baden-Wuerttemberg, Germany, as no such screening had ever been conducted there. In total, we obtained 102 animals from hunters in 2019 and 2020. Intestinal helminths were retrieved using the SSCT (segmented sedimentation and counting technique) and identified morphologically and by PCR-based Sanger sequencing. Fecal samples were assessed using the ELISA PetChekTM IP assay (IDEXX, Germany) and flotation technique. The artificial digestion method was employed for analyzing muscle tissue. We detected species of four nematode genera (Baylisascaris procyonis, Toxocara canis, Capillaria spp., and Trichuris spp.), three cestode genera (Atriotaenia cf. incisa/procyonis, Taenia martis, and Mesocestoides spp.), and three trematode genera (Isthmiophora hortensis/melis, Plagiorchis muris, and Brachylaima spp.). Echinococcus spp. and Trichinella spp. were not found. The invasive behavior and synanthropic habits of raccoons may increase the infection risk with these helminths in wildlife, domestic and zoo animals, and humans by serving as a connecting link. Therefore, it is crucial to initiate additional studies assessing these risks.

First insights into the activity of major digestive enzymes in the intestine of the European catfish Silurus glanis and protective anti-enzymatic potential of its gut parasite Silurotaenia siluri.

The European catfish Silurus glanis is attracting growing interest as an object of fisheries and aquaculture, which is reinforced by the expansion of its natural range under climate change. Shaping the effective exploitation strategy for this valuable species requires detailed knowledge of its biology, including feeding and digestion processes, especially near the natural limits of the species range. Meanwhile, the digestion physiology of the European catfish remains poorly explored, including the activity of major digestive enzymes and the possible effect of intestinal parasites on this activity. In this regard, the activity of proteinases and α-amylase in the intestinal mucosa of the catfish was studied. Adult catfish were collected in the Rybinsk reservoir (Upper Volga) located close to the northern limit of the species range. It was shown that all subclasses of intestinal digestive proteinases, including serine proteinases, metalloproteases and cysteine (thiol) proteinases, function in the gut mucosa of the catfish. The mucosal levels of total proteolytic activity depended on fish size, in contrast to those of trypsin, chymotrypsin and α-amylase. The level of chymotrypsin activity was significantly higher than that of trypsin activity. It was also found that the incubation medium and extract of the cestodes Silurotaenia siluri parasitizing the catfish gut had a significant inhibitory effect on the activity of serine proteases (trypsin and chymotrypsin) operating in the intestines of the host fish.

Targeted and non-targeted proteomics to characterize the parasite proteins of Echinococcus multilocularis metacestodes.

The larval stage of the cestode Echinococcus multilocularis is the causative agent of alveolar echinococcosis. To investigate the biology of these stages and to test novel compounds, metacestode cultures represent a suitable in vitro model system. These metacestodes are vesicles surrounded by an envelope formed by the vesicle tissue (VT), which is formed by the laminated and germinal layer, and filled with vesicle fluid (VF). We analyzed the proteome of VF and VT by liquid chromatography tandem mass spectrometry (LC-MS/MS) and identified a total of 2,954 parasite proteins. The most abundant protein in VT was the expressed conserved protein encoded by EmuJ_000412500, followed by the antigen B subunit AgB8/3a encoded by EmuJ_000381500 and Endophilin B1 (protein p29). In VF, the pattern was different and dominated by AgB subunits. The most abundant protein was the AgB8/3a subunit followed by three other AgB subunits. In total, the AgB subunits detected in VF represented 62.1% of the parasite proteins. In culture media (CM), 63 E. multilocularis proteins were detected, of which AgB subunits made up 93.7% of the detected parasite proteins. All AgB subunits detected in VF (encoded by EmuJ_000381100-700, corresponding to AgB8/2, AgB8/1, AgB8/4, AgB8/3a, AgB8/3b, and AgB8/3c) were also found in CM, except the subunit encoded by EmuJ_000381800 (AgB8/5) that was very rare in VF and not detected in CM. The relative abundance of the AgB subunits in VF and CM followed the same pattern. In VT, only the subunits EmuJ_000381500 (AgB8/3a) and EmuJ_000381200 (AgB8/1) were detected among the 20 most abundant proteins. To see whether this pattern was specific to VF from in vitro cultured metacestodes, we analyzed the proteome of VF from metacestodes grown in a mouse model. Here, the AgB subunits encoded by EmuJ_000381100-700 constituted the most abundant proteins, namely, 81.9% of total protein, with the same order of abundance as in vitro. Immunofluorescence on metacestodes showed that AgB is co-localized to calcareous corpuscles of E. multilocularis. Using targeted proteomics with HA-tagged EmuJ_000381200 (AgB8/1) and EmuJ_000381100 (AgB8/2), we could show that uptake of AgB subunits from CM into VF occurs within hours.

Trophic diversification and parasitic invasion as ecological niche modulators for gut microbiota of whitefish.

Introduction: The impact of parasites on gut microbiota of the host is well documented, but the role of the relationship between the parasite and the host in the formation of the microbiota is poorly understood. This study has focused on the influence that trophic behavior and resulting parasitism has on the structure of the microbiome. Methods: Using 16S amplicon sequencing and newly developed methodological approaches, we characterize the gut microbiota of the sympatric pair of whitefish Coregonus lavaretus complex and the associated microbiota of cestodes parasitizing their intestine. The essence of the proposed approaches is, firstly, to use the method of successive washes of the microbiota from the cestode's surfaces to analyze the degree of bacterial association to the tegument of the parasite. Secondly, to use a method combining the sampling of intestinal content and mucosa with the washout procedure from the mucosa to understand the real structure of the fish gut microbiota. Results and discussion: Our results demonstrate that additional microbial community in the intestine are formed by the parasitic helminths that caused the restructuring of the microbiota in infected fish compared to those uninfected. Using the desorption method in Ringer's solution, we have demonstrated that Proteocephalus sp. cestodes possess their own microbial community which is put together from "surface" bacteria, and bacteria which are weakly and strongly associated with the tegument, bacteria obtained after treatment of the tegument with detergent, and bacteria obtained after removal of the tegument from the cestodes.

Medical Parasitology Taxonomy Update, June 2020-June 2022.

The taxonomy of medically important parasites continues to evolve. This minireview provides an update of additions and updates in the field of human parasitology from June 2020 through June 2022. A list of previously reported nomenclatural changes that have not been broadly adapted by the medical community is also included.

Recent Advances and Potential Future Applications of MALDI-TOF Mass Spectrometry for Identification of Helminths.

Helminth infections caused by nematodes, trematodes, and cestodes are major neglected tropical diseases and of great medical and veterinary relevance. At present, diagnosis of helminthic diseases is mainly based on microscopic observation of different parasite stages, but microscopy is associated with limited diagnostic accuracy. Against this background, recent studies described matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry as a potential, innovative tool for helminth identification and differentiation. MALDI-TOF mass spectrometry is based on the analysis of spectra profiles generated from protein extracts of a given pathogen. It requires an available spectra database containing reference spectra, also called main spectra profiles (MSPs), which are generated from well characterized specimens. At present, however, there are no commercially available databases for helminth identification using this approach. In this narrative review, we summarize recent developments and published studies between January 2019 and September 2022 that report on the use of MALDI-TOF mass spectrometry for helminths. Current challenges and future research needs are identified and briefly discussed.