AbstractToxoplasma gondii is an obligate intracellular protozoan parasite, which can infect almost all warm-blooded animals, including humans, leading to toxoplasmosis. Currently, the effective treatment for human toxoplasmosis is the combination of sulphadiazine and pyrimethamine. However, both drugs have serious side-effects and toxicity in the host. Therefore, there is an urgent need for the discovery of new anti-T. gondii drugs with high potency and less or no side-effects. Our findings suggest that lumefantrine exerts activity against T. gondii by inhibiting its proliferation in Vero cells in vitro without being toxic to Vero cells (P ≤ 0.01). Lumefantrine prolonged mice infected with T. gondii from death for 3 days at the concentration of 50 μg L−1 than negative control (phosphate-buffered saline treated only), and reduced the parasite burden in mouse tissues in vivo (P ≤ 0.01; P ≤ 0.05). In addition, a significant increase in interferon gamma (IFN-γ) production was observed in high-dose lumefantrine-treated mice (P ≤ 0.01), whereas interleukin 10 (IL-10) and IL-4 levels increased in low-dose lumefantrine-treated mice (P ≤ 0.01). The results demonstrated that lumefantrine may be a promising agent to treat toxoplasmosis, and more experiments on the protective mechanism of lumefantrine should be undertaken in further studies.

Background: Toxoplasma gondii is an obligate intracellular protozoan parasite, which can infect almost all warm-blooded animals, including humans, leading to toxoplasmosis. Currently, the effective treatment for human toxoplasmosis is the combination of sulfadiazine and pyrimethamine. However, both drugs have serious side effects and toxicity in the host. Therefore, there is an urgent need for the discovery of new anti- Toxoplasma drugs with high potency and less or no side-effects. Methods: The cytotoxicity of sulfadiazine and lumefantrine to Vero cells was evaluated by the methyl thiazolyl tetrazolium (MTT) assay. And MTT assay was also used to detect the inhibitory effects of lumefantrine on parasites invasion and proliferation. Flow cytometry was conducted to further verify parasites proliferation. qPCR was performed to evaluate the parasite load in the mice after lumefantrine treatment. In order to determine whether lumefantrine treatment enhances Th1 or Th2 cytokine response, IFN-γ, IL-4, and IL-10 levels in the serum of mice were determined. Results: Our findings suggest that lumefantrine exerts activity against T. gondii by inhibiting its replication and invasion of Vero cells in vitro without being toxic to the cells. Furthermore, lumefantrine protected mice with acute toxoplasmosis from death to a certain extent and reduced the parasite burden in mouse tissues in vivo . In addition, a significant increase in IFN-γ production was observed in high dose lumefantrine-treated mice while IL-10 and IL-4 levels increased in low dose lumefantrine-treated mice. Conclusions: The results of this study demonstrated that lumefantrine may be a promising agent to treat toxoplamosis, and more experiments on the protective mechanism of lumefantrine should be undertaken in further studies. Key words: Toxoplasma gondii , Lumefantrine, anti- Toxoplasma gondii , Invasion, Proliferation

Many obligate intracellular apicomplexan parasites have adapted a distinct invasion mechanism involving a close interaction between the parasite ligands and the sialic acid (SA) receptor. We found that sialic acid binding protein-1 (SABP1), localized on the outer membrane of the zoonotic parasite Toxoplasma gondii, readily binds to sialic acid on the host cell surface. The binding was sensitive to neuraminidase treatment. Cells preincubated with recombinant SABP1 protein resisted parasite invasion in vitro. The parasite lost its invasion capacity and animal infectivity after the SABP1 gene was deleted, whereas complementation of the SABP1 gene restored the virulence of the knockout strain. These data establish the critical role of SABP1 in the invasion process of T. gondii. The previously uncharacterized protein, SABP1, facilitated T. gondii attachment and invasion via sialic acid receptors.

Host cell invasion by Toxoplasma gondii is crucial for the survival and proliferation of parasite. The process of T. gondii tachyzoite invasion requires interaction between parasite proteins and receptors on the surface of host cells. Sialic acid is one of the important receptors for host cell invasion by T. gondii. However, the parasite-derived proteins interacting with sialic acid have not been well characterized. In this study, a novel protein named putative TCP-1 chaperonin (TGME49_318410) in T. gondii (TgTCP-1) was targeted and characterized. TgTCP-1 protein colocalized with MIC3 protein, which could be secreted from T. gondii tachyzoites, and this protein showed a specific binding activity to sialic acid, and DC and Vero cells in vitro. The binding of TgTCP-1 protein to DC and Vero cells were inhibited by either pre-incubation with free sialic acid or neuraminidase treatment of the cells. Moreover, a significant reduction of T. gondii invasion in Vero cells was observed after pre-incubation of the cells with recombinant TgTCP-1 protein. These results illustrated that TgTCP-1 is an important molecule involved in sialic acid-dependent host cell invasion by T. gondii.

原生动物寄生虫感染是当今全球面临的最紧迫的健康问题之一,且某些种类寄生虫可造成巨大的经济损失.耐药性寄生虫的不断产生,迫切需要揭示其分子致病机理,希望找到新的潜在药物靶点.通过干扰赖氨酸乙酰化破坏基因表达对原生动物寄生虫(疟原虫、弓形虫、锥虫等)的存活具有重要影响,赖氨酸乙酰化的bromodomain蛋白已经成为基因表达的关键调控因子,甚至有希望成为新一类药物靶标.虫体bromodomain蛋白质在寄生虫的生存活力、侵袭和生长阶段转换中发挥重要作用,并且bromodomain抑制剂具有作为新型抗寄生虫药的功效.论文对代表性寄生虫的bromodomain蛋白等相关方面的研究进展做一综述.

Toxoplasma gondii is a unicellular protozoan parasite of the phylum Apicomplexa. The parasite repeatedly goes through a cycle of invasion, division and induction of host cell rupture, which is an obligatory process for proliferation inside warm-blooded animals. It is known that the biology of the parasite is controlled by a variety of mechanisms ranging from genomic to epigenetic to transcriptional regulation. In this study, we investigated the global protein posttranslational lysine crotonylation and 2-hydroxyisobutyrylation of two T. gondii strains, RH and ME49, which represent distinct phenotypes for proliferation and pathogenicity in the host. Proteins with differential expression and modification patterns associated with parasite phenotypes were identified. Many proteins in T. gondii were crotonylated and 2-hydroxyisobutyrylated, and they were localized in diverse subcellular compartments involved in a wide variety of cellular functions such as motility, host invasion, metabolism and epigenetic gene regulation. These findings suggest that lysine crotonylation and 2-hydroxyisobutyrylation are ubiquitous throughout the T. gondii proteome, regulating critical functions of the modified proteins. These data provide a basis for identifying important proteins associated with parasite development and pathogenicity.

弓形虫是一种专性细胞内寄生原虫,宿主类型众多,寄生于人、畜、野生动物及禽类等几乎所有的有核细胞中,引发人兽共患寄生虫病,在世界范围内流行,对人类和畜牧业都具有潜在威胁.猫科动物是弓形虫唯一的终末宿主;弓形虫的中间宿主极其广泛,包括各种哺乳类动物、禽类和人等,其中感染最为严重的动物是猪.猪弓形虫病在不同地区均有流行发生,感染率也较高,平均感染率达22.3％.而猪肉是人类主要的肉食消费品之一,人在摄入未煮熟的含有弓形虫包囊的猪肉或内脏时可获得弓形虫感染,危害人类健康.故本试验对辽宁省不同地区的猪弓形虫病进行流行病学调查,并分析其感染因素.目前,检测弓形虫的金标准是改良凝集实验(MAT),其特点是操作简单、检测快速、结果准确,避免了酶联免疫吸附试验(ELISA)、胶体金试验、间接血凝试验等造成的假阳性和假阴性.从辽宁省不同地区采集猪的血清样品,采用MAT方法对血清样品进行猪弓形虫血清抗体检测并对其感染因素进行分析,结果显示:各个地区弓形虫感染率有所不同,抚顺感染率最高(27.9％),本溪感染率最低(6.28％);不同发育阶段的猪均有感染,繁殖母猪感染率(25.14％)>育肥猪感染率(12.23％)>仔猪感染率(6.04％),且繁殖母猪经产次数越多其感染率越高,经产5胎的繁殖母猪的感染率约是经产1胎繁殖母猪的13倍;大白猪较长白猪更易感弓形虫.另外,弓形虫可以通过母猪的胎盘传播方式(即垂直传播)传染给仔猪,使其获得先天性感染;仔猪也可以通过水平传播方式获得感染,即通过接触环境中的卵囊感染弓形虫.研究结果可为预防和控制猪弓形虫的感染提供一定的数据参考.

刚地弓形虫(Toxoplasma gondii)作为顶复门中一种寄生原虫,可以感染有核细胞.研究表明,钙离子与弓形虫侵染、运动等生理活动相关.内质网中含有大量的钙离子,许多钙离子结合蛋白质在弓形虫的生理活动中起重要作用.钙离子结合蛋白质包含多种EF手型(EF-hand)结构域.目前记载在TOXO DB中含有EF-hand的蛋白质有55种;在Plasmo DB含有EF-hand的蛋白质有69种.本文对钙离子结合蛋白质的各种作用进行阐述.

Invasion of erythrocytes by merozoites is required in the life cycle of malarial parasites. Proteins derived from the invasive merozoites are essential ligands for erythrocyte recognition and penetration. In this study, we report a novel protein that possesses a Trx domain-like structure of the thioredoxin family and is expressed on the surface of merozoites of the malaria parasite Plasmodium falciparum. This protein, namely, PfTrx-mero protein, displayed a mutated sequence character at the Trx domain, but with a specific binding activity to human erythrocytes. Specific antibodies to the protein inhibited merozoite invasion into human erythrocytes. Immunization with a homologous protein of Plasmodium berghei strain ANKA also showed significant protection against lethal infection in mice. These results suggested that the novel PfTrx-like-mero protein expressed on the surface of merozoites is an important ligand participating in erythrocyte invasion and a potential vaccine candidate.

目的 对刚地弓形虫钙依赖性蛋白激酶2B(TgCDPK2B)进行生物信息学分析,预测其免疫原性及其功能.方法 利用生物信息学在线预测软件ProtParam、SOSUI、ProtScale、TMHMM、SignalP-4.1、Targetp1.1Server、SOPMA、Motif Scan、SWISS-MODEL、SYFPEITHI等分析预测TgCDPK2B蛋白的理化性质、可溶性、亲疏水性、跨膜结构域、信号肽、亚细胞定位、二级结构、蛋白翻译后修饰位点,以及三级结构、可溶性、柔韧性、表面可及性、B细胞和T细胞抗原表位等. 结果 CDPK2B蛋白含有604个氨基酸,分子式为C2965H4633N823O890S23,相对分子质量(Mr)为66.78676×103,理论等电点为6.40,不稳定指数为31.70,脂溶性指数为77.43,平均疏水性系数为-0.440 562,属于亲水性蛋白.TgCDPK2B蛋白无信号肽和跨膜区,在该蛋白的二级结构中,α-螺旋、β-折叠、β-转角、无规则卷曲所占比例分别为38.58％、15.4％、11.42％和34.60％;TgCDPK2B蛋白含有11个亲水性和11个柔韧性参数＞2.0的区域,28个表面可及性＞1.9的区域,8个暴露表面得分＞2.3的区域.TgCDPK2B蛋白共有4种翻译后修饰位点,33个B细胞抗原表位,13个CTL细胞抗原表位,34个Th细胞抗原表位. 结论 生物信息学预测TgCDPK2B为可溶性蛋白,含有多个B、T细胞抗原表位,可作为弓形虫疫苗抗原候选.

Toxoplasma gondii is an obligatory intracellular parasite that causes a common infection in many warm-blooded animals. During infection, the host’s immune system plays an important role in confining the dissemination of the parasites in the hosts. T cell immunoglobulin- and mucin domain–containing molecule 3 (Tim-3) has been characterized as an important regulator in cell-mediated immune responses in various infections. Here, we compared Tim-3 expression on splenic and circulatory T, B cells and a few cytokines in the sera of mice infected with the more virulent type I (RH) vs. the low virulent type II (ME49) strain. Tim-3 expression on the splenic and circulatory T cells of mice infected with T. gondii (RH strain) was higher than that in mice infected with T. gondii (ME49 strain). T. gondii infection reduced the proportion of splenic helper T cells (Th) and cytotoxic T cells (Tc) and increased Tim-3 expression. Further, serum levels of interleukin (IL)-2, interferon γ, tumor necrosis factor (TNF)-α, IL-12p70, IL-22, IL-17A, and IL-5 increased significantly after infection. Mice infected with T. gondii (ME49 strain) showed higher levels of TNF-α, IL-17A, IL-12p70, and IL-22 than that infected by the RH strain. Our study revealed that T. gondii strains may have their inherent ability in triggering different host immune responses, which may explain the clinical variation in diseases severity after infection.

Toxoplasma gondii is an obligate intracellular zoonotic parasite, infecting warm-blood animals including humans. Previous serological surveys of T. gondii infection have focused on people of different occupations and special groups, such as slaughterhouse workers, AIDS patients and pregnant women. To investigate the potential impact of T. gondii infection on the health of young students, the prevalence of T. gondii infection and associated risk factors among the newly enrolled undergraduates and postgraduate students were investigated. A total of 3,569 newly enrolled students (age range: 15- to 37-years-old, median 26 years) from various regions of China were recruited in this study. The serum samples were tested for the presence of T. gondii specific IgG by the modified agglutination test (MAT). Questionnaires were used to collect information on risk factors for T. gondii infection. Sixty-five (1.82%) out of 3,569 participants were seropositive for IgG antibodies to T. gondii by MAT (titer≥1:20). Four variables were found to be positively associated with T. gondii infection, including primary geographical location, living in rural areas, gardening or agriculture, and drinking unboiled water by the univariate logistic regression, and only gardening or agriculture was the independent risk factor for T. gondii positivity by using multivariate logistic regression in this study, which may provide information to guide future research and control policies.

Serum from 2 063 pigs from a slaughterhouse in Liaoning Province were collected and detected by modified agglutination test (MAT).The results showed that the samples with titer≥25 were 233 and the positive rate was 11.3 ％.A total of 67 pigs with titer≥100 were chosen for Toxoplasma gondii isolation,in which 23 T.gondii strains were isolated from the heart tissues with an isolation rate of 34.3％.In this study,T.gondii strains were isolated successfully,showing the infectious status of pig toxoplasmosis in Liaoning Province,which poses a huge potential threat to public health.

BACKGROUND:Genetic information for Toxoplasma gondii isolates from pigs in eastern, south, and southwestern regions of China has been reported previously. However, there are no data from pigs in the northeastern area of the country. To better understand the epidemiology of T. gondii, we determined the seroprevalence and genotypes of T. gondii from pigs slaughtered for human consumption in Liaoning province, northeastern China.FINDINGS:Out of 2063 pigs examined, 233 (11.26%) were seropositive for T. gondii by the modified agglutination test (MAT), and viable parasites were isolated by bioassay in mice from 23 (9.87%) of the 233 seropositive pigs. Fifteen out of 23 isolates were genotyped using 10 PCR-restriction fragment length polymorphism (RFLP) genetic markers including SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico. One isolate was identified as ToxoDB genotype #3 (type II-variant), and one was genotype #1 or #3. The other 13 isolates were ToxoDB #9 (type Chinese 1).CONCLUSIONS:To our knowledge, this is the first report of T. gondii isolation and genotyping from pigs in northeastern China. This study indicates that pigs are a potential source for transmission of T. gondii to humans, therefore poses a potential public health concern. The genotyping results revealed the presence of genotype Chinese 1 in northeastern China, enriching the scope of T. gondii genotypes distribution in eastern Asia.