Microsporidian EnP1 alters host cell H2B monoubiquitination and prevents ferroptosis facilitating microsporidia survival.

Microsporidia are intracellular eukaryotic pathogens that pose a substantial threat to immunocompromised hosts. The way these pathogens manipulate host cells during infection remains poorly understood. Using a proximity biotinylation strategy we established that microsporidian EnP1 is a nucleus-targeted effector that modifies the host cell environment. EnP1's translocation to the host nucleus is meditated by nuclear localization signals (NLSs). In the nucleus, EnP1 interacts with host histone H2B. This interaction disrupts H2B monoubiquitination (H2Bub), subsequently impacting p53 expression. Crucially, this inhibition of p53 weakens its control over the downstream target gene SLC7A11, enhancing the host cell's resilience against ferroptosis during microsporidian infection. This favorable condition promotes the proliferation of microsporidia within the host cell. These findings shed light on the molecular mechanisms by which microsporidia modify their host cells to facilitate their survival.

Overcoming research challenges: In vitro cultivation of Ameson portunus (Phylum Microsporidia).

Ameson portunus is an intracellular pathogen that infects marine crabs Portunus trituberculatus and Scylla paramamosain, causing significant economic losses. However, research into this important parasite has been limited due to the absence of an in vitro culture system. To address this challenge, we developed an in vitro cultivation model of A. portunus using RK13 cell line in this study. The fluorescent labeling assay indicated a high infection rate (∼60 %) on the first day post-infection and quantitative PCR (qPCR) detection demonstrated successful infection as early as six hours post-inoculation. Fluorescence in situ hybridization (FISH) and qPCR were used for the detection of A. portunus infected cells. The FISH probe we designed allowed detection of A. portunus in infected cells and qPCR assay provided accurate quantification of A. portunus in the samples. Transmission electron microscopy (TEM) images revealed that A. portunus could complete its entire life cycle and produce mature spores in RK13 cells. Additionally, we have identified novel life cycle characteristics during the development of A. portunus in RK 13 cells using TEM. These findings contribute to our understanding of new life cycle pathways of A. portunus. The establishment of an in vitro culture model for A. portunus is critical as it provides a valuable tool for understanding the molecular and immunological events that occur during infection. Furthermore, it will facilitate the development of effective treatment strategies for this intracellular pathogen.

Expression profiles of host miRNAs and circRNAs and ceRNA network during Toxoplasma gondii lytic cycle.

Toxoplasma gondii is an opportunistic protozoan parasite that is highly prevalent in the human population and can lead to adverse health consequences in immunocompromised patients and pregnant women. Noncoding RNAs, such as microRNAs (miRNAs) and circular RNAs (circRNAs), play important regulatory roles in the pathogenesis of many infections. However, the differentially expressed (DE) miRNAs and circRNAs implicated in the host cell response during the lytic cycle of T. gondii are unknown. In this study, we profiled the expression of miRNAs and circRNAs in human foreskin fibroblasts (HFFs) at different time points after T. gondii infection using RNA sequencing (RNA-seq). We identified a total of 7, 7, 27, 45, 70, 148, 203, and 217 DEmiRNAs and 276, 355, 782, 1863, 1738, 6336, 1229, and 1680 DEcircRNAs at 1.5, 3, 6, 9, 12, 24, 36, and 48 h post infection (hpi), respectively. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses revealed that the DE transcripts were enriched in immune response, apoptosis, signal transduction, and metabolism-related pathways. These findings provide new insight into the involvement of miRNAs and circRNAs in the host response to T. gondii infection.

Evaluation of protective immunity induced by a DNA vaccine encoding SAG2 and SRS2 against Toxoplasma gondii infection in mice.

Toxoplasma gondii is an important protozoan pathogen, which can cause severe diseases in the newborns and immunocompromised individuals. Developing an effective vaccine against Toxoplasma infection is a critically important global health priority. Immunofluorescence staining analysis revealed that TgSAG2 and TgSRS2 are membrane associated and displayed on the surface of the parasite. Immunizations with pBud-SAG2, pBud-SRS2 and pBud-SAG2-SRS2 DNA vaccines significantly increased the production of specific IgG antibodies. Immunization with pBud-SAG2-SRS2 elicited cellular immune response with higher concentrations of IFN-γ and IL-4 compared to the control group. Antigen-specific lymphocyte proliferations in the pBud-SRS2 and pBud-SAG2-SRS2 groups were significantly higher compared to that in the control group. Furthermore, 30 % of mice immunized with pBud-SAG2-SRS2 survived after the challenge infection with virulent T. gondii RH tachyzoites. This study revealed that immunization with pBud-SAG2-SRS2 induced potent immune responses, and has the potential as a promising vaccine candidate for the control of T. gondii infection.

Anti-Toxoplasma gondii effect of tylosin in vitro and in vivo.

BACKGROUND: Toxoplasma gondii is an important protozoan pathogen with medical and veterinary importance worldwide. Drugs currently used for treatment of toxoplasmosis are less effective and sometimes cause serious side effects. There is an urgent need for the development of more effective drugs with relatively low toxicity. METHODS: The effect of tylosin on the viability of host cells was measured using CCK8 assays. To assess the inhibition of tylosin on T. gondii proliferation, a real-time PCR targeting the B1 gene was developed for T. gondii detection and quantification. Total RNA was extracted from parasites treated with tylosin and then subjected to transcriptome analysis by RNA sequencing (RNA-seq). Finally, murine infection models of toxoplasmosis were used to evaluate the protective efficacy of tylosin against T. gondii virulent RH strain or avirulent ME49 strain. RESULTS: We found that tylosin displayed low host toxicity, and its 50% inhibitory concentration was 175.3 µM. Tylsoin also inhibited intracellular T. gondii tachyzoite proliferation, with a 50% effective concentration of 9.759 µM. Transcriptome analysis showed that tylosin remarkably perturbed the gene expression of T. gondii, and genes involved in "ribosome biogenesis (GO:0042254)" and "ribosome (GO:0005840)" were significantly dys-regulated. In a murine model, tylosin treatment alone (100 mg/kg, i.p.) or in combination with sulfadiazine sodium (200 mg/kg, i.g.) significantly prolonged the survival time and raised the survival rate of animals infected with T. gondii virulent RH or avirulent ME49 strain. Meanwhile, treatment with tylosin significantly decreased the parasite burdens in multiple organs and decreased the spleen index of mice with acute toxoplasmosis. CONCLUSIONS: Our findings suggest that tylosin exhibited potency against T. gondii both in vitro and in vivo, which offers promise for treatment of human toxoplasmosis.