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Toxocariasis is a neglected parasitic disease caused predominantly by larvae of Toxocara canis. While this zoonotic disease is of major importance in humans and canids, it can also affect a range of other mammalian hosts. It is known that mucins secreted by larvae play key roles in immune recognition and evasion, but very little is understood about the molecular interactions between host cells and T. canis. Here, using an integrative approach (affinity pull-down, mass spectrometry, co-immunoprecipitation and bioinformatics), we identified 219 proteins expressed by a murine macrophage cell line (RAW264.7) that interact with prokaryotically-expressed recombinant protein (rTc-MUC-1) representing the mucin Tc-MUC-1 present in the surface coat of infective larvae of T. canis. Protein-protein interactions between rTc-MUC-1 and an actin binding protein CFL1 as well as the fatty acid binding protein FABP5 of RAW264.7 macrophages were also demonstrated in a human embryonic kidney cell line (HEK 293T). By combing predicted structural information on the protein-protein interaction and functional knowledge of the related protein association networks, we inferred roles for Tc-MUC-1 protein in the regulation of actin cytoskeletal remodelling, and the migration and phagosome formation of macrophage cells. These molecular interactions now require verification in vivo. The experimental approach taken here should be readily applicable to comparative studies of other ascaridoid nematodes (e.g. T. cati, Anisakis simplex, Ascaris suum and Baylisascaris procyonis) whose larvae undergo tissue migration in accidental hosts, including humans. Author summary: Toxocariasis is a neglected parasitic disease of humans caused mainly by larvae of Toxocara canis. Given that T. canis is zoonotic and can infect a range of mammals, there has been substantial interest in host-parasite relationships, with studies showing that T. canis larvae secrete abundant mucins that effect/modulate immune responses and disease pathogenesis. To improve the understanding of immunomolecular interactions, we investigated the role(s) of the protein component of a mucin (Tc-MUC-1) secreted by infective larvae using a well-defined murine macrophage line (RAW264.7). The non-glycosylated recombinant protein of Tc-MUC-1 (designated rTc-MUC-1) was shown to interact with at least 219 proteins of RAW264.7 cells, particularly with the actin binding protein (CFL1) and a fatty acid binding protein (FABP5), which are involved in cell migration and phagocytosis, respectively. Based on these findings, we propose that Tc-MUC-1 regulates cytoskeletal organisation and signal transduction in host macrophages. It would be interesting to establish, using the integrative experimental approach employed here, whether the role(s) of Tc-MUC-1 protein homologues of related ascaridoids are conserved. [ABSTRACT FROM AUTHOR]

Simple Summary: Toxoplasmosis is one of the most common parasitic zoonoses in the world. It not only threatens human health and safety but also causes considerable losses to the global animal husbandry industry. Swine, as an intermediate host of Toxoplasma gondii, has important economic and public health significance, and the global prevalence of T. gondii in swine varies between 10% and 60%. Therefore, it is important to establish simple, effective, sensitive and specific diagnostic methods for the detection and prevention of T. gondii infection in swine. In the present study, we developed a colloidal gold immunochromatographic strip based on rTgROP14, recombinant protein A and monoclonal antibody TgROP14-5D5 for serological detection of T. gondii in swine populations. This new ICT achieved good specificity and sensitivity and has potential for routine serological detection of T. gondii in swine. Toxoplasma gondii, a worldwide distributed apicomplexan protozoan, can infect almost all warm-blooded animals and may cause toxoplasmosis. In order to provide a point-of-care detection method for T. gondii infection, an immunochromatographic test (ICT) was established. The proposed test uses recombinant T. gondii rhoptry protein 14 (ROP14) conjugated with 20 nm gold particles, recombinant protein A as the detection line and monoclonal antibody TgROP14-5D5 as the control line. The specificity, sensitivity, positive predictive value, negative predictive value and stability of this new ICT were evaluated. rTgROP14 was specifically recognized by positive serum of T. gondii but not negative serum. mAb TgROP14-5D5 showed higher specific recognition of T. gondii antigens and was therefore selected for subsequent colloidal gold strip construction. The new ICT based on TgROP14 exhibited good diagnostic performance with high specificity (86.9%) and sensitivity (90.9%) using IHA as a "reference standard". Among 436 field porcine sera, ICT and IHA detected 134 (30.7%) and 99 (22.7%) positive samples, respectively. The relative agreement was 87.8%. These data indicate that this new ICT based on TgROP14 is a suitable candidate for routine testing of T. gondii in the field. [ABSTRACT FROM AUTHOR]

Highlights: [•] The potent protection in mice immunized with recombinant protein ROP18 + Re was analyzed. [•] The rROP18 triggered a stronger humoral and cellular response against T. gondii. [•] Re is a promising vaccine adjuvant against toxoplasmosis, deserves further evaluation. [Copyright &y& Elsevier]

• Haemonchus contortus protein Hc-DHS-28 has 3-hydroxyacyl CoA dehydrogenase activity. • Hc-dhs-28 is highly transcribed in the diapause worm and located in the peroxisome. • Hc-dhs-28 shares high similarity in gene function with Caenorhabditis elegans (Ce) -dhs-28. • Hc-dhs-28 might regulate the formation of diapause in H. contortus. • Immunisation with recombinant Hc-DHS-28 could protect the host after H. contortus infection. Haemonchus contortus could enter the diapause stage to avoid hostile conditions, however the inducing mechanism still remains poorly understood. A similar dauer strategy exists in Caenorhabditis elegans , and dauer phenomones, which are produced through a four step cycle of peroxisomal fatty acid β-oxidation, are essential in this stage. In this study, a novel gene, Hc-dhs-28 , was identified and characterised. Hc-DHS-28 was the homologue of Ce-DHS-28, a key enzyme in the oxidation cycle, and the protein contained a short chain dehydrogenase domain and a peroxisomal targeting signal 1. The expression pattern of Hc-DHS-28 detected by quantitative real-time PCR and indirect immunofluorescence assay revealed that this protein was mainly expressed in the intestine and subdermal regions of larvae at diapause and in free-living stages. Enzyme activity analysis confirmed its 3-hydroxyacyl CoA dehydrogenase activity with 121, 149, 162 and 166 as key functional sites; meanwhile co-localization in human embryonic kidney 293 cells indicated that Hc-DHS-28 was targeted to the peroxisome of cytoplasm under the guide of peroxisomal targeting signal 1, which was consistent with the functional domain prediction of Hc-dhs-28. Overexpression, rescue and RNA interference experiments were carried out to explore the function of Hc-dhs-28. Our results showed that Hc-dhs-28 was very similar to Ce-dhs-28 and partially rescued its function in C. elegans. RNAi with Hc-dhs-28 in C. elegans led to decreased transcription of genes in the peroxisomal fatty acid β-oxidation cycle, considerable fat accumulation and dauer formation defects. Furthermore, immunisation with recombinant Hc-DHS-28 protein in sheep was able to maintain the body weight of the host after infection and reduce the worm burden. In conclusion, Hc-DHS-28 is most likely involved in the peroxisome fatty acid β-oxidation as the third 3-hydroxyacyl CoA dehydrogenase to regulate the production of diapause-related pheromones, and then influence the formation of diapause in H. contortus. [ABSTRACT FROM AUTHOR]

Echinococcus granulosus is the causative pathogen of cystic echinococcosis, a serious disease endangering human and animal health. In this study, an immunochromatographic strip was developed based on the recombinant protein Heat shock protein 70 (HSP70) for the serological detection of E. granulosus . The protocol completes within 20 min requiring no specialized equipment or chemical reagents, while specificity tests confirmed no cross-reactivity with positive serum of Fasciola hepatica , Haemonchus contortus , Peste des petits ruminants virus (PPRV) and Foot-and-mouth disease virus (FMDV). The strips remained stable after storage at 4 °C for up to 8 months. Both immunochromatographic strip and ELISA tests were applied to detect E. granulosus antibody in a total of 728 serum samples obtained from slaughter houses in Zhejiang Province. Our data revealed positive rates of 2.61 and 1.65% by immunochromatographic strip and ELISA methods, respectively. The immunochromatographic strip test developed in this study provides a simple, specific and rapid method of E. granulosus antibody detection and infected sheep monitoring. [ABSTRACT FROM AUTHOR]