Microglia are resident immune cells in the central nervous system (CNS) and play a crucial role in immune surveillance and tissue repair in the brain. Microglia are involved in processes such as neurogenesis, antigen presentation, synaptic pruning, and phagocytosis. They can be activated by pathogenic infections and various other pathological factors, thereby exerting effects of mediating neuroinflammation or neuroprotection. Meanwhile, when the homeostasis of the neuronal microenvironment is disrupted, microglia can become hyperactivated, leading to multi-dimensional damage and exacerbation of CNS lesions. This article systematically reviews the activation pathways and phenotypic states of microglia, their involvement in CNS injury during pathogen infections, and their roles in the infection-related CNS diseases. It aims to provide insights into the mechanisms underlying the involvement of microglia in infectious brain diseases and to identify potential therapeutic targets for CNS damage.

Toxoplasma gondii (T. gondii) has the ability to disseminate widely in the host, including crossing the blood brain barrier (BBB) to establish persistent brain cysts. However, the molecular mechanism by which T. gondii hijacks host immune cell migration to facilitate its own dissemination remain incompletely understood. Here we show that T. gondii infection upregulates S100A6 via a calcium and P65-dependent pathway; This elevated S100A6 expression promoted its interaction with filamin A, leading to promoted cytoskeletal reorganization, enhanced cell migration and T. gondii replication. Furthermore, S100A6 knockout in C57BL/6 J mice decreases BBB permeability, inhibits tachyzoite traversal into the brain, and results in fewer and smaller brain cysts. These findings identify S100A6 as a critical regulator of T. gondii induced cell migration and dissemination, and suggest that targeting S100A6 or its downstream signaling can offer therapeutic strategies to limit parasite spread and prevent chronic neuro-toxoplasmosis.

​Toxoplasma gondii, an opportunistic protozoan, causes severe toxoplasmosis in immunocompromised individuals and adverse pregnancy outcomes. Its genome is critical for studying intracellular pathogens and drug targets, yet existing first/second-generation assemblies remain incomplete due to repetitive sequences. Using third-generation nanopore sequencing, we refined the T. gondii genome, reconfirming the fusion of Chromosomes VIIb and VIII across GT1, PRU, and VEG strains, and identifying multiple previously unassigned novel sequences. We corrected misreported copy numbers of diagnostic markers: the B1 gene ranged from 9–19 copies, and 529 bp repeats from 86-212 copies, lower than prior estimates. No correlation was found between virulence-related gene clusters and strain virulence. Notably, the VEG strain’s apicoplast genome was 5-6 times larger than GT1/PRU, with type III strains showing greater divergence than type I/II strains. This high-quality T. gondii genome resource advances parasitology research.

ABSTRACT Enterococcus faecalis (E. faecalis) is an opportunistic pathogen capable of causing various life-threatening infections, including urinary tract infections, bloodstream infections, infective endocarditis, and meningitis. As a major etiological agent of healthcare-associated infections (HAIs), its global prevalence continues to rise, a trend closely linked to the increasing problem of multidrug resistance driven by overuse of antibiotics. Therefore, rapid and accurate detection is essential for timely treatment and improved prognosis. In this study, the pheS gene of E. faecalis was rapidly amplified using recombinase polymerase amplification (RPA), and detection was achieved via a CRISPR/Cas12a system. The Cas12a-crRNA complex specifically recognized the amplification product and triggered nonspecific cleavage of a single-stranded DNA (ssDNA) reporter, generating a fluorescent signal that could be quantified in a real-time PCR system or visualized directly under ultraviolet (UV) light. After optimization of key parameters—including RPA primers, reaction conditions, crRNA sequence, and the crRNA/Cas12a combination—the assay achieved a limit of detection (LOD) of 10−2 ng/μL within a short turnaround time, and showed no cross-reactivity with other common pathogen detection results from clinical isolates and spiked samples were fully consistent with those obtained through PCR/qPCR, confirming high reliability. In summary, the RPA-CRISPR/Cas12a detection method established in this study is sensitive, specific, and reliable. Its simplicity, minimal equipment requirements, and cost-effectiveness make it a promising tool for rapid clinical detection of E. faecalis.IMPORTANCEEnterococcus faecalis is a major opportunistic pathogen responsible for severe healthcare-associated infections, with rising prevalence linked to antibiotic resistance. Rapid and accurate detection is critical for timely treatment and infection control. Conventional methods are often time-consuming or require complex laboratory infrastructure, limiting their use at the point of care. This study developed a rapid detection assay by integrating recombinase polymerase amplification with the CRISPR/Cas12a system, targeting the pheS gene of E. faecalis. The method is sensitive and specific, providing visual results under UV light within a short turnaround time. It offers a simple, cost-effective, and requires minimal equipment, suitable for clinical and resource-limited settings, potentially improving diagnostic efficiency and supporting antimicrobial stewardship.

The apicoplast, a relic plastid organelle derived from secondary endosymbiosis, is crucial for many medically relevant Apicomplexa. While it no longer performs photosynthesis, the organelle retains several essential metabolic pathways. In this study, we examine the four primary metabolic pathways in the Toxoplasma gondii apicoplast, along with an accessory pathway, and identify conditions that can bypass these. Contrary to the prevailing view that the apicoplast is indispensable for T. gondii, we demonstrate that bypassing all pathways renders the apicoplast non-essential. We further show that T. gondii lacking an apicoplast (T. gondii-Apico) can be maintained indefinitely in culture, establishing a unique model to study the functions of this organelle. Through comprehensive metabolomic, transcriptomic, and proteomic analyses of T. gondii-Apico we uncover significant adaptation mechanisms following loss of the organelle and identify numerous putative apicoplast proteins revealed by their decreased abundance in T. gondii-Apico. Moreover, T. gondii-Apico parasites exhibit reduced sensitivity to apicoplast targeting compounds, providing a valuable tool for discovering new drugs acting on the organelle. The capability to culture T. gondii without its plastid offers new avenues for exploring apicoplast biology and developing novel therapeutic strategies against apicomplexan parasites.

The phylum Apicomplexa comprises eukaryotic parasites that cause fatal diseases affecting millions of people and animals worldwide. Their mitochondrial genomes have been significantly reduced, leaving only three protein-coding genes and highly fragmented mitoribosomal rRNAs, raising challenging questions about mitoribosome composition, assembly and structure. Our study reveals how Toxoplasma gondii assembles over 40 mt-rRNA fragments using exclusively nuclear-encoded mitoribosomal proteins and three lineage-specific families of RNA-binding proteins. Among these are four proteins from the Apetala2/Ethylene Response Factor (AP2/ERF) family, originally known as transcription factors in plants and Apicomplexa, now repurposed as essential mitoribosome components. Cryo-EM analysis of the mitoribosome structure demonstrates how these AP2 proteins function as RNA binders to maintain mitoribosome integrity. The mitoribosome is also decorated with members of lineage-specific RNA-binding proteins belonging to RAP (RNA-binding domain abundant in Apicomplexa) proteins and HPR (heptatricopeptide repeat) families, highlighting the unique adaptations of these parasites. Solving the molecular puzzle of apicomplexan mitoribosome could inform the development of therapeutic strategies targeting organellar translation.

Toxoplasma gondii, a parasitic protozoan affecting approximately one-third of global population, causes opportunistic toxoplasmosis. It penetrates barriers to immune-privileged sites, causing encephalitis, retinochoroiditis, and fetal damage. The infection may be linked to neurodegenerative and psychiatric disorders. The T. gondii–host interaction mechanism remains central to understanding its pathogenesis. The changes in small molecule metabolites after infection, which affects the central nervous system (CNS) normal function, have been poorly characterized. The metabolic alterations in brain tissues of sv129 mice infected by T. gondii at 9 days post-infection (DPI) were analyzed through untargeted metabolomic detection. Cholesterol metabolic reprogramming was assessed through analysis of related gene’s transcription with quantitative reverse transcription polymerase chain reaction (qRT-PCR). The primary target cells responsible for cholesterol metabolic dysregulation were identified through detection of the secreted cytokines with enzyme-linked immunosorbent assay (ELISA). The T. gondii replication in host cells treated with 25-HC was evaluated using immunofluorescence assay (IFA). Transcriptomic analysis was performed to identify the differentially expressed genes (DEGs) in the host cells infected by T. gondii and/or treated with 25-HC, and the host cell M1 polarization was confirmed by qRT-PCR. Brain metabolomic profiling identified 19 differentially expressed metabolites (including 25-HC), primarily involved in amino acid metabolism and cholesterol metabolism pathways (biosynthesis of primary bile acids and steroids). Toxoplasma gondii infection triggered host cholesterol metabolic reprogramming and promoted 25-HC secretion from glial cells, which indirectly inhibited T. gondii’s proliferation in host cells. Transcriptomic analysis revealed that 25-HC upregulated the expression of chemokines, C-type lectin receptors, and inflammation-related genes. Notably, 25-HC was verified to confer host resistance against T. gondii infection by promoting microglial M1 polarization. Our study demonstrated that T. gondii infection activates the CH25H-25-HC axis to induce microglial M1 polarization and cytokine secretion, thereby establishing an anti-Toxoplasma defense. These findings highlight the central role of cholesterol metabolism in T. gondii pathogenesis and provide innovative strategies for the diagnosis, prevention, and treatment of toxoplasmosis.

Acute-on-chronic liver failure (ACLF) is a leading cause of global liver-related mortality. Bacterial infection, especially in patients with decompensated cirrhosis, commonly triggers ACLF and is difficult to treat with antibiotics. Therefore, finding alternative strategies for preventing and managing bacterial infection is an urgent priority. Here, we observed that patients with bacterial infection and decompensated cirrhosis, as well as ACLF mice, exhibited lower fecal panose levels than uninfected controls. Megamonas funiformis, with 4α-glucanosyltransferase (4αGT) as a key enzyme for panose production, was identified as a potential panose producer. Animal experiments demonstrated that panose efficiently reduced liver injury and extended survival in ACLF mice by mitigating bacterial infection. Further results revealed that panose enhanced resistance to bacterial infection by inhibiting oxidative stress-induced gut barrier disruption, thereby limiting bacterial dissemination. Mechanistically, panose interacted with the solute carrier family 7 member 11 (SLC7A11, also known as xCT) protein to boost antioxidant glutathione levels in intestinal epithelial cells. These findings highlight panose's potential in preventing bacterial infection, offering a valuable insight into mitigating ACLF progression.

IntroductionZika virus (ZIKV) is a mosquito-borne arbovirus. Maternal infection may cause severe complications such as neonatal microcephaly and neurological defects. To date, there is no clinically approved vaccine or specific drug against ZIKV infection. The host calcium-binding protein, S100A6, is a member of S100 protein family, regulates various cellular processes, and has been recognized as a host-dependent factor for Flavivirus infection.MethodsS100A6 expression in host cells after ZIKV infection was detected by western blotting (WB). The effects of host S100A6 on ZIKV replication as indicated by the RNA and protein levels of nonstructural protein 3 (NS3) were detected by qRT-PCR, plaque assay, immunofluorescence assay (IFA), and WB respectively. The interaction and co-localization of S100A6 with NS3 were examined through co-immunoprecipitation (Co-IP) and IFA. Proteasome inhibitor and lysosomal acidification inhibitor were used to explore the degradation pathway of NS3 mediated by S100A6.ResultsZIKV infection induced a dose- and time-dependent increase in host S100A6 expression. Overexpression of S100A6 in HeLa cells did not affect ZIKV binding or entry into host cells but significantly inhibited viral replication. Conversely, S100A6 knockdown led to a significant increase in ZIKV replication. Moreover, S100A6 was found binding to ZIKV-NS3, leading to NS3 degradation without affecting genome copies. The use of lysosomal acidification inhibitor NH4Cl significantly reversed S100A6-mediated downregulation of NS3 protein levels, suggesting that S100A6 degrades NS3 via the lysosomal pathway.ConclusionZIKV infection upregulated host S100A6, which acted as an anti-infection factor by specifically targeting ZIKV-NS3 for degradation, thereby inhibiting viral replication. These findings provide insights into a potential mechanism of host resistance to ZIKV infection and enhance our understanding of the ZIKV-host interaction.

Cell membrane-derived nanovesicles (CMNVs) are nanoscale lipid bilayer structures obtained from cellular membranes that serve as biomimetic drug delivery platforms, offering immune evasion, targeting, and surface functionalization capabilities. While most CMNVs originate from mammalian cells, Toxoplasma gondii (T. gondii), a genetically tractable protozoan with a structurally distinct membrane, offers a high-yield and underexplored source for producing T. gondii-derived CMNVs (TgCMNVs). These vesicles are obtained from the parasite's plasma membrane and inner membrane complex and retain unique features including abundant GPI-anchored SRS proteins, phosphatidylthreonine-rich lipids, and an editable genome, enabling versatile engineering via genetic and chemical strategies. We review methods for TgCMNV fabrication, purification, and functionalization, and evaluate their potential in immunomodulation, attenuation of tissue injury, cancer immunotherapy, and self-adjuvanting vaccine design. By combining intrinsic immune engagement with programmable surface architecture, TgCMNVs could serve as a complementary and adaptable platform alongside established CMNV systems. Finally, we discuss key translational considerations, including scalable production, immunogenicity control, regulatory compliance, and stability testing, which will be essential for assessing the feasibility of TgCMNVs in clinical applications.

Toxoplasma gondii (T. gondii) is an opportunistic pathogen affecting about 1/3 of world population. While often asymptomatic in immunocompetent individuals, it can lead to severe toxoplasmosis in immunocompromised patients. Recent research has unveiled a potential link between T. gondii infection and neuropsychiatric diseases. We implemented both a cohort study and a case control study to further identify this association. In the cohort study, we analyzed data from the UK Biobank database, which included 8,814 individuals tested for T. gondii SAG1 antibodies and free of neuropsychiatric disorders at baseline. Among them, 22.52% (n=1,985) tested positive for SAG1 antibody. Over an average follow-up period of 12.26 years, Cox proportional hazards models and logistic regression analysis revealed a significant association between the SAG1 seropositivity at baseline and the incidence of schizophrenia (HR: 5.89; 95% CI: 1.69-20.53). In our case-control study, 239 patients diagnosed with schizophrenia and 455 healthy individuals were involved. Using the modified agglutination test (MAT) to detect T. gondii antibodies, logistic regression analysis showed a higher prevalence of T. gondii infection among schizophrenia patients (10.04%) compared to healthy controls (3.74%). T. gondii infection emerged as a significant risk factor for schizophrenia (OR: 3.33; 95% CI: 1.68-6.61). However, our investigations did not reveal a robust association between T. gondii infection and other neuropsychiatric conditions, including Alzheimer’s disease, dementia, anxiety, depression, neurodegenerative disorders, and peripheral neurological disorders such as neurological and plexus disorders.

Background: Accumulating evidence suggests that latent infection with Toxoplasma gondii (T. gondii) is associated with a variety of neuropsychiatric and behavioral conditions. This research aims to explore the potential correlation between T. gondii antibody positivity and neuropsychiatric disorders through a comprehensive prospective cohort study. Methods: The cohort study utilized the UK Biobank database to recruit 8814 individuals with no prior diagnosis of neuropsychiatric disorders. Cox proportional hazards models were employed to investigate the associations between T. gondii P22 antibody seropositivity (P22+) + ) and the development of various types of neuropsychiatric disorders. Results: Of the population, 14.65 % tested positive for T. gondii P22 antibody. The presence of T. gondii P22 antibody showed a slight inverse association with epilepsy (HR: 0.28; 95 % CI: 0.10-0.77), while it was positively associated with an increased risk of developing anxiety disorders (HR: 1.38; 95 % CI: 1.04-1.83). Limitations: The study sample consisted mostly of white British individuals aged 40 to 69 years old. Although we adjusted for potential confounders, there may be other unmeasured and residual confounding factors that could have influenced our reported associations. Conclusions: The findings suggested an increased risk of anxiety and potential evidence of epilepsy associated with T. gondii P22+. + . However, our analysis did not reveal an increased risk of several other neuropsychiatric conditions including Alzheimer's disease, dementia, substance abuse disorders, depression, and neurodegenerative disorders, associated with P22 antibody seropositivity.

Abstract Background Toxoplasma gondii infection affects a significant portion of the global population, leading to severe toxoplasmosis and, in immunocompromised patients, even death. During T. gondii infection, disruption of gut microbiota further exacerbates the damage to intestinal and brain barriers. Therefore, identifying imbalanced probiotics during infection and restoring their equilibrium can regulate the balance of gut microbiota metabolites, thereby alleviating tissue damage. Methods Vimentin gene knockout (vim−/−) mice were employed as an immunocompromised model to evaluate the influence of host immune responses on gut microbiota balance during T. gondii infection. Behavioral experiments were performed to assess changes in cognitive levels and depressive tendencies between chronically infected vim−/− and wild-type (WT) mice. Fecal samples were subjected to 16S ribosomal RNA (rRNA) sequencing, and serum metabolites were analyzed to identify potential gut probiotics and their metabolites for the treatment of T. gondii infection. Results Compared to the immunocompetent WT sv129 mice, the immunocompromised mice exhibited lower levels of neuronal apoptosis and fewer neurobehavioral abnormalities during chronic infection. 16S rRNA sequencing revealed a significant decrease in the abundance of probiotics, including several species of Lactobacillus, in WT mice. Restoring this balance through the administration of Lactobacillus murinus and Lactobacillus gasseri significantly suppressed the T. gondii burden in the intestine, liver, and brain. Moreover, transplantation of these two Lactobacillus spp. significantly improved intestinal barrier damage and alleviated inflammation and neuronal apoptosis in the central nervous system. Metabolite detection studies revealed that the levels of various Lactobacillus-related metabolites, including indole-3-lactic acid (ILA) in serum, decreased significantly after T. gondii infection. We confirmed that L. gasseri secreted much more ILA than L. murinus. Notably, ILA can activate the aromatic hydrocarbon receptor signaling pathway in intestinal epithelial cells, promoting the activation of CD8+ T cells and the secretion of interferon-gamma. Conclusion Our study revealed that host immune responses against T. gondii infection severely disrupted the balance of gut microbiota, resulting in intestinal and brain damage. Lactobacillus spp. play a crucial role in immune regulation, and the metabolite ILA is a promising therapeutic compound for efficient and safe treatment of T. gondii infection. Graphical Abstract

Abstract Background Toxoplasma gondii is an opportunistic pathogenic protozoan that infects all warm-blooded animals, including humans, and causes zoonotic toxoplasmosis. The bradyzoite antigen 1 (BAG1), known as heat-shock protein (HSP)30, is a specific antigen expressed during the early stage of T. gondii tachyzoite–bradyzoite conversion. Methods A bag1 gene knockout strain based on the T. gondii type II ME49 was constructed and designated as ME49Δbag1. The invasion, proliferation, and cyst formation efficiency in the cell model and survival in the mouse model were compared between the ME49 and ME49Δbag1 strains after infection. Quantitative polymerase chain reaction (qPCR) was used to detect the transcriptional level of important genes, and western-blot was used to detect protein levels. Results ME49Δbag1 displayed significantly inhibited cyst formation, although it was not completely blocked. During early differentiation induced by alkaline and starvation conditions in vitro, the proliferation of ME49Δbag1 was significantly accelerated relative to the ME49 strain. Meanwhile, the transcription of the HSP family and bradyzoite formation deficient 1 (bfd1) were significantly enhanced. The observed upregulation suggests a compensatory mechanism to counterbalance the impaired stress responses of T. gondii following bag1 knockout. On the other hand, the elevated transcription levels of several HSP family members, including HSP20, HSP21, HSP40, HSP60, HSP70, and HSP90, along with BFD1, implied the involvement of alternative regulatory factors in bradyzoite differentiation aside from BAG1. Conclusions The data suggested that when bag1 was absent, the stress response of T. gondii was partially compensated by increased levels of other HSPs, resulting in the formation of fewer cysts. This highlighted a complex regulatory network beyond BAG1 influencing the parasite’s transformation into bradyzoites, emphasizing the vital compensatory function of HSPs in the T. gondii life cycle adaptation. Graphical Abstract

Toxoplasma gondii is an intracellular opportunistic parasite that exists in a latent form within the human central nervous system (CNS), even in immune-competent hosts. During acute infection, T. gondii traverses the blood–brain barrier (BBB). In the subsequent chronic infection phase, the infiltration of immune cells into the brain, driven by T. gondii infection and the formation of parasitic cysts, leads to persistent activation and proliferation of astrocytes and microglia. This process results in neuronal damages that are fatal in some cases. Through inducing systemic immune responses, T. gondii infection can dramatically alter the behavior of rodents and increase the risk of various neuropsychiatric disorders in humans. In this review, we explore some recent research progress on the major events involved in BBB disruption, glial cell activation and neuronal damage following T. gondii infection in hosts. It further discusses potential pathological mechanisms and the feasible treatment approaches for the neurodegenerative and neuropsychiatric disorders caused by T. gondii infection to extend our understanding for pathogenesis and preventive control of toxoplasmosis in humans.

ABSTRACT Protein disulfide isomerase, containing thioredoxin (Trx) domains, serves as a vital enzyme responsible for oxidative protein folding (the formation, reduction, and isomerization of disulfide bonds in newly synthesized proteins) in the endoplasmic reticulum (ER). However, the role of ER-localized PDI proteins in parasite growth and their interaction with secretory proteins remain poorly understood. In this study, we identified two ER-localized PDI proteins, TgPDI8 and TgPDI6, in Toxoplasma gondii. Conditional knockdown of TgPDI8 resulted in a significant reduction in intracellular proliferation and invasion abilities, leading to a complete block in plaque formation on human foreskin fibroblast monolayers, whereas parasites lacking TgPDI6 did not exhibit any apparent fitness defects. The complementation of TgPDI8 with mutant variants highlighted the critical role of the CXXC active site cysteines within its Trx domains for its enzymatic activity. By utilizing TurboID-based proximity labeling, we uncovered a close association between PDI proteins and canonical secretory proteins. Furthermore, parasites lacking TgPDI8 showed a significant reduction in the expression of secretory proteins, especially those from micronemes and dense granules. In summary, our study elucidates the roles of TgPDI8 and sets the stage for future drug discovery studies.IMPORTANCEApicomplexans, a phylum of intracellular parasites, encompass various zoonotic pathogens, including Plasmodium, Cryptosporidium, Toxoplasma, and Babesia, causing a significant economic burden on human populations. These parasites exhibit hypersensitivity to disruptions in endoplasmic reticulum (ER) redox homeostasis, necessitating the presence of ER-localized thioredoxin (Trx) superfamily proteins, particularly protein disulfide isomerase (PDI), for proper oxidative folding. However, the functional characteristics of ER-localized PDI proteins in Toxoplasma gondii remain largely unexplored. In this study, we identified two ER-localized proteins, namely, TgPDI8 and TgPDI6, and demonstrated the indispensable role of TgPDI8 in parasite survival. Through a comprehensive multi-omics analysis, we elucidated the crucial role of TgPDI8 in the processing of secretory proteins in T. gondii. Additionally, we introduced a novel ER-anchored TurboID method to label and identify canonical secretory proteins in T. gondii. This research opens up new avenues for understanding oxidative folding and the secretory pathway in apicomplexan parasites, laying the groundwork for future advancements in antiparasitic drug development.

目的 筛选并分析刚地弓形虫慢性感染小鼠脑转录组差异表达基因(DEG),分析与抑郁相关的犬尿氨酸(KYN)通路DEG的相对转录水平,为探究弓形虫慢性感染导致小鼠抑郁样症状的机制提供理论依据.方法 18只SV129雄性小鼠随机平均分为感染组和对照组.感染组小鼠腹腔注射ME49株速殖子120个(200 μl),对照组注射等体积的磷酸缓冲液,感染后3个月收集感染组和对照组小鼠脑组织,提取小鼠脑组织总RNA进行转录组测序,筛选DEG,对DEG进行聚类分析、基因本体(GO)功能注释分析、京都基因与基因组百科全书(KEGG)功能注释和富集分析.选取与抑郁症相关的KYN通路的8个DEG,分别为 γ 干扰素(IFN-γ)、吲哚胺2,3-双加氧酶1(IDO1)、IDO2、犬尿氨酸酶(KYNU)、犬尿氨酸-3-单氧化酶(KMO)、3-羟基邻氨基苯甲酸3,4-二加氧酶(3-HAO)、波形蛋白(Vim)和脑源性神经营养因子(BDNF),以甘油醛-3-磷酸脱氢酶基因为内参,实时荧光定量PCR(qRT-PCR)检测各个基因的相对转录水平.结果 感染组和对照组小鼠脑转录组的DEG共2 295个,其中上调2016个,下调279个.GO分析结果显示,生物过程中富集最显著的是定位,共257个DEG;细胞组分中富集最显著的是蛋白复合物,共425个DEG;分子功能中富集最显著的是分子转导活性,共177个DEG.生物过程、细胞组分和分子功能富集DEG数量最多的分别是细胞过程、细胞组分和结合,分别有1 039、1 240、1 088个DEG.KEGG分析结果显示,功能注释分析上调居前3位的代谢通路分别为免疫系统、信号转导、病毒性传染病,下调居前3位的分别为信号转导、信号分子和相互作用、免疫系统;功能富集分析结果显示77条通路富集显著.与抑郁症相关的信号通路有肿瘤坏死因子、神经活性配体-受体相互作用、NF-kappa B、JAK-STAT、坏死性凋亡、细胞凋亡、趋化因子、KYN通路等.qRT-PCR结果显示,以对照组小鼠相对转录水平为100％,感染组小鼠IFN-γ、IDO1、IDO2、KYNU、KMO、3-HAO 和 Vim 等 7 个基因的相对转录水平分别为 3 023.08％、355.52％、190.17％、496.55％、339.92％、212.74％、507.34％,较对照组的转录水平显著上调(t=3.782、3.749、3.226、2.908、2.533、5.656、2.948,均P＜0.05或0.01);BDNF的相对转录水平为63.32％,转录水平显著下调(t=2.398,P＜0.05).IFN-γ、IDO1、IDO2、KYNU、KMO、3-HAO、BDNF、Vim等 8 个基因 qRT-PCR获得的差异倍数分别为4.96、1.74、0.89、2.10、1.60、1.06、-0.94、2.18,转录组测序获得的差异倍数分别为7.30、0.55、0.80、3.83、2.75、3.53、-0.86、1.93.qRT-PCR与转录组测序获得的转录趋势一致.结论 筛选获得刚地弓形虫慢性感染小鼠脑转录组DEG,弓形虫慢性感染小鼠中枢神经系统免疫持续激活,与抑郁症相关的KYN通路的7个DEG的转录水平上调.

Mosquito-borne diseases, such as malaria, dengue, Zika, and West Nile, have caused substantial disease burdens to the people living in tropical and subtropical regions. Despite decades of efforts in control, the prevalence, geographic distribution, and severity of these diseases are still deteriorating and the options for vector control are limited. Insect-specific viruses (ISVs) can naturally infect and replicate in mosquitoes, and even be vertically transmitted from generation to generation, interfering with arbovirus' proliferation, and disturbing mosquito's physiology, which are harmless to vertebrates and have great potential for bio-control. Insect-specific RNA viruses (ISRVs) infection will inhibit the replication of mosquito-borne viruses, which may be due to the close phylogeny of ISRVs and the arboviruses. Mosquito baculoviruses (MBVs) and mosquito densoviruses (MDVs) belong to insect-specific DNA viruses. The former has limited pathogenicity to mosquitoes, while the latter can be horizontally and vertically transmitted in mosquitoes, resulting in habitat pollution, abnormal development or death of larvae and adults. In addition, ISVs can be used as fusion expression vectors to express insect virulence proteins in mosquitoes or directly inactivate important genes of mosquitoes. However, there are still many deficiencies in ISVs research studies. Clarification of the host limit mechanism for each ISVs and application of more targeted and efficient ISVs or toxins will be the future research directions.

ABSTRACT Toxoplasma gondii is an important intracellular protozoan, which needs to exploit host nutrition for successful parasitism. We previously reported that Casitas B-lineage lymphoma-b (Cbl-b) was screened as a host dependency factor of T. gondii by using lentiviral CRISPR-Cas9-single guide RNA (sgRNA) libraries. Furthermore, the detailed mechanism of Cbl-b being required by T. gondii infection was explored in this study. The proliferation of T. gondii was found to be significantly inhibited in Cbl-b knockdown cell lines, and the Cbl-b expression level increased with prolonged T. gondii infection, while the MyD88 level was significantly decreased in the T. gondii infection group. Toll-like receptor (TLR)/MyD88 is a conserved innate cellular immune signaling pathway against pathogens infection. Cbl-b was found to interact with MyD88 and mediate MyD88 ubiquitination in the fluorescence resonance energy transfer and co-immunoprecipitation experiments. A Cbl-b knockout (KO) C57BL/6J lineage was then constructed and, together with the wild-type (WT) mice, was infected with the same amount of T. gondii tachyzoites of ME49 strain. At 13 days post infection, all the mice in the WT group died, while 80% of the mice in the Cbl-b KO group still survived, even to the end of the experiment. The parasitic burden in the liver, lung, and brain of the Cbl-b KO mice was significantly lower than that of the WT mice (P < 0.05). In Cbl-b KO infected mice, the percentage of B cells was higher, whereas that of macrophages was lower, and the interferon-γ and interleukin-6 levels in the serum were higher than that in the WT infected mice; the difference was significant (P < 0.05). Furthermore, when host cells were infected by T. gondii, host’s Cbl-b was found to interact with MyD88 to ubiquitinate MyD88 for ribosome-dependent degradation. Therefore, the host’s innate immunity against T. gondii through the TLR/MyD88 pathway was negatively regulated by Cbl-b. IMPORTANCE This is the first report that a human E3 ubiquitin ligase, Casitas B-lineage lymphoma proto-oncogene B (Cbl-b), functions as a host dependency factor for the intracellular protozoan Toxoplasma gondii and the mechanism for how T. gondii infection inhibits the TLR/MyD88 innate immunity pathway through MyD88 degradation mediated by Cbl-b. This finding is an impactful contribution for understanding the host cell immunity against T. gondii infection.

Toxoplasma gondii is the causative agent of toxoplasmosis, a zoonotic disease that poses a threat to human health and a considerable loss to livestock farming. At present, clinical therapeutic drugs mainly target T. gondii tachyzoites and fail to eradicate bradyzoites. Developing a safe and effective vaccine against toxoplasmosis is urgent and important. Breast cancer has become a major public health problem and the therapeutic method needs to be further explored. Many similarities exist between the immune responses caused by T. gondii infection and the immunotherapy for cancers. T. gondii dense granule organelles secrete immunogenic dense granule proteins (GRAs). GRA5 is localized to the parasitophorous vacuole membrane in the tachyzoite stage and the cyst wall in the bradyzoite stage. We found that T. gondii ME49 gra5 knockout strain (ME49Δgra5) was avirulent and failed to form cysts but stimulated antibodies, inflammatory cytokines, and leukocytes infiltration in mice. We next investigated the protective efficacy of ME49Δgra5 vaccination against T. gondii infection and tumor development. All the immunized mice survived the challenge infection of either wild-type RH, ME49, VEG tachyzoites, or ME49 cysts. Moreover, ME49Δgra5 tachyzoite inoculation in situ attenuated the growth of murine breast tumor (4T1) in mice and prevented 4T1's lung metastasis. ME49Δgra5 inoculation upregulated the levels of Th1 cytokines and tumor-infiltrating T cells in the tumor microenvironment and triggered anti-tumor responses by increasing the number of natural killer, B, and T cells, macrophages, and dendritic cells in the spleen. Collectively, these results suggested that ME49Δgra5 was a potent live attenuated vaccine against T. gondii infection and breast cancer.