Enolase 2 (ENO2) is a crucial an important glycolytic enzyme involved in glycolysis and is also recognised as a multifunctional protein. However, its interacting proteins in intestinal porcine epithelial cells (IPEC-J2 cells) remain unclear. This study investigates the interacting proteins of Cystoisospora suis (Biester et Murray, 1934) ENO2 (CsENO2) in IPEC-J2 cells. According to our previous research, CsENO2 is highly expressed in sporulated oocysts. Co-immunoprecipitation/liquid chromatography-mass spectrometry (Co-IP/LC-MS) was conducted using the pcDNA3.1-Flag vector carrying the enolase 2 gene, which was transfected into IPEC-J2 cells to co-immunoprecipitate with cellular proteins. The Co-IP/LC-MS-based protein sequencing identified three cellular proteins: NME2, RPS3, and Vimentin. Subsequently, cDNAs encoding the complete coding sequences of these proteins were cloned into the pGEX-4T-1 expression vector. Immunofluorescence assays revealed the localisation of CsENO2 in sporulated oocysts and sporozoites. Most CsENO2 protein was found in the cytoplasm of sporulated oocysts. In sporozoites, CsENO2 protein was primarily distributed around and outside the nucleus. This study aims to identify CsENO2-interacting proteins in IPEC-J2 cells and determine the localisation of CsENO2 within sporulated oocysts and sporozoites.
The species diversity of reptile parasites in the New World, including South America, remains poorly understood, and this also applies to tapeworms (Cestoda: Onchoproteocephalidea). Only 17 species of the non-monophyletic genus Ophiotaenia La Rue, 1911 have been recorded in reptiles in the Neotropical Region. In this paper, three new species are described from dipsadine snakes of the genus Erythrolamprus Boie (Colubridae) in Brazil and Paraguay, and a new genus, Penetrocestus, is proposed to accommodate these tapeworms that penetrate the intestinal wall of their hosts. The new genus differs from other proteocephalids parasitising snakes by the presence of a large apical glandular organ, which is larger than the suckers, and a large vaginal sphincter lined with a thick layer of chromophilic cells. Penetrocestus gen. n. is also characterised by a long and wide region ('metascolex') posterior to the scolex and a well-developed inner longitudinal musculature, which is weakly developed in most proteocephalids parasitising Neotropical snakes. Ophiotaenia joanae (de Chambrier et Paulino, 1997) from Xenodon neuwiedii Günther (Colubridae) in Brazil is transferred to Penetrocestus as its type species and as a new combination. Penetrocestus gorgonae sp. n. from Erythrolamprus aesculapii (Linnaeus) in Brazil, Penetrocestus jaegeri sp. n. from Erythrolamprus jaegeri (Günther) in Paraguay, and Penetrocestus tristani sp. n. from Erythrolamprus poecilogyrus (Wied-Neuwied) in Paraguay differ from Penetrocestus joanae (de Chambrier et Paulino, 1997) comb. n. and from each other by the position of the metascolex, width of the scolex, morphology of the terminal (distal) portion of the vagina, and the number of testes, as summarised in the key to their identification. The phylogenetic relationships of the new species are unknown, as they were collected more than 35 years ago and no tissues were fixed in ethanol for genetic characterisation, but the new data support the assumption of strict host specificity of reptilian tapeworms in the Neotropical Region.
The monogenean ectoparasite Gyrodactylus salaris Malmberg, 1957 was detected in 2001 on juveniles of landlocked Salmo salar Linnaeus in three rivers draining to Lake Kuitozero in the White Sea basin. The ancestry of the colony was studied by internal transcribed spacers of the nuclear ribosomal operon (ITS), mitochondrial COI gene and by 12 microsatellite markers. The maternal ancestor was a frozen heterozygotic and heteroplasmic diploid clone RBT marked by a ITS-1068R, spread by rainbow trout and observed in Denmark, Norway, Finland and Russia. The clone combined genomes from two divergent matrilines, probably from grayling (Thymallus Linck) in the Southern Baltic basin (COI alien, 84 % bootstrap support) and in Balkans (COI rbt, 99 %). The genetic distance of the 1545 bp sequences was dMCL = 0.025. The population in Kuitozero was a progeny of RBT females repeatedly inseminated by conspecific males. It was sexual but propagated as parthenogenetic clones. The derived population carried only COI rbt.. The microsatellite alleles of the RBT clone segregated with the paternal alleles following Mendelian model, with an excess of heterozygotes: He = 0.608, Ho = 0.883. Among 244 genotyped worms, 27 multilocus clones were observed, as segregated among the parr (FST = 0.824). Parental genotypes were not detected. The largest clone (104 worms) was heterozygotic in all 12 loci and was identified on fifteen fish, in two rivers and four years. The balancing selection (excess of heterozygotes) equalised the allelic frequencies and maximised the genetic variance available for purifying and adaptive natural selection on a new host. The two sexual episodes associated with three host switch steps offers a model to understand the evolution of the diverse host spectrum of host-specific Gyrodactylus von Nordmann, 1832.
The discovery of multiple metabolic enzymes encoded by genes of apparent plant or cyanobacterial origin in trypanosomatids led to the influential hypothesis that the common ancestor of Euglenozoa harboured a plastid that was subsequently lost in kinetoplastids. Here, we critically re-evaluate this hypothesis using an expanded, phylogenetically balanced dataset comprising 299 eukaryotic and 102 bacterial species. We reassess the evolutionary histories of 16 genes encoding proteins previously interpreted as evidence for an ancestral euglenozoan plastid using state-of-the-art maximum-likelihood and Bayesian phylogenetic approaches, supplemented by topology tests. Our analyses reveal that none of the examined genes provides compelling support for a plastid-bearing ancestor of Euglenozoa. Instead, these enzymes display heterogeneous evolutionary origins consistent with multiple independent horizontal gene transfer events between euglenozoans and diverse bacterial (distinct from cyanobacteria) and eukaryotic donors. Only the gene encoding a vacuolar H⁺-pyrophosphatase, an electrogenic proton pump, shows limited affinity to chloroplast-bearing lineages, and this signal alone is insufficient to infer plastid ancestry. Taken together, our results strongly suggest a horizontal gene transfer from various non-plastid bearing lineages over the hypothesis of plastid presence in the euglenozoan common ancestor with subsequent loss in kinetoplastids, diplonemids, and non-photosynthetic euglenids.
