Seroprevalence and incidence of Toxoplasma gondii infection in the Legnano area of Italy.

The decreasing prevalence of anti-Toxoplasma antibodies in Europe has re-opened the question of the appropriateness of serological screening during pregnancy. A study of 3426 pregnant women, resident in the Legnano area of Italy, revealed that the IgG seroprevalence according to ELISA was 21.5%, and that of IgM according to ELISA and enzyme-linked fluorescent assay was 1.2% and 0.9%, respectively. The incidence of infection, estimated on the basis of IgG avidity, was 0.9%. These results confirm a decrease in the prevalence of IgG, but indicate a high incidence of infection, thus suggesting that screening for anti-Toxoplasma antibodies during pregnancy should be maintained.     

孕期弓形虫感染对胎盘的入侵和病理损害的研究

目的探讨孕期弓形虫感染对胎盘的入侵和病理损害情况。方法孕8dBALB/c小鼠经腹腔接种弓形虫速殖子,在孕10、12、14、16和18d剖腹取胎鼠,分离胎盘并精确称重。采用HE染色观察胎盘的病理损害情况,RNA原位杂交检测胎盘组织弓形虫速殖子的动态入侵。结果在孕10、12d,感染组胎盘重量与对照组比较无明显差别。而在孕14、16、18d,感染组的胎盘重量明显低于对照组（P〈0.01）。随着感染时间的延长,胎盘组织切片可见有大量淋巴细胞、单核细胞、嗜酸性粒细胞浸润,绒毛血管减少,显示有炎症表现;滋养层细胞破坏明显,可见凋亡小体形成。感染组小鼠在孕10d未检测到虫体,感染孕12d偶见虫体。孕14、16、18d虫体数目明显增多（P〈0.01）,虫体主要位于滋养层细胞内、细胞间质和血管中,并有假包囊形成。结论孕期弓形虫感染可影响胎盘生长,胎盘出现炎症反应和凋亡等病理损害。虫体可侵入胎盘组织细胞间质和血管中,并侵入滋养层细胞内增殖,跨越胎盘屏障传播给胎儿。     

社区居民弓形虫健康教育效果评价

目的评价对社区居民开展弓形虫病健康教育和重点人群监测干预的效果。方法以社区卫生服务机构为服务载体，对社区居民开展有计划的多种形式的健康教育；对社区居民中育龄妇女、孕妇及宠物饲养者建立健康档案，定期监测；参照“KABP”调查标准，设计调查问卷调查，用SPSS10．0软件分析数据。结果健康教育前社区居民对弓形虫病防治知识、态度信念、行为平均得分为46．5、61．8、81．3；健康教育实施半年后平均得分分别为69．6、77．8、96．6，差异有统计学意义（P〈0．001）。健康教育实施前社区居民特殊人群中育龄妇女、孕妇、宠物饲养者对弓形虫病防治知识知晓率、态度与信念、行为形成率有明显差异（P〈0．01）。结论在社区居民中开展弓形虫病防治的健康教育是有效的。     

96-Well plates providing high optical resolution for high-throughput, immunofluorescence-based screening of monoclonal antibodies against Toxoplasma gondii

We have developed a method for high resolution, high magnification immunofluorescence-based screening in a multi-well format, using a recently introduced 96-well plate specifically developed for fluorescence microscopy. We report here on the use of these plates to screen hybridoma supernatants for reactivity with specific subcellular compartments of the protozoan parasite Toxoplasma gondii. This has proven to be a powerful screening strategy, particularly when combined with high-throughput immunoblotting, and has enabled us to generate nine different monoclonal antibodies (MAbs) against either the periphery or structures within the apical end of T. gondii. The availability of a disposable, inexpensive, 96-well plate with optical properties suitable for high magnification imaging could lead to applications in a variety of fluorescence-based screening protocols.     

经口感染弓形虫在小鼠组织内的动态分布

目的观察经口感染弓形虫速殖子在小鼠小肠组织、实质器官内的动态分布。方法BALB／c小鼠96只．随机分为8组，0d组用0．5ml PBS／只灌胃，其余组用RH株弓形虫速殖子1×10^4个灌胃感染，感染后2、4、6、8、10、12、14d处死小鼠，每次处死12只，取十二指肠、空肠、回肠、肝脏、脾、肾、肺、心和脑做组织印片，吉-瑞氏染色，镜检。结果感染后2d在十二指肠、空肠、回肠、肺和心，4d在脾，6d在肾和肝脏发现虫体，脑内未发现虫体。十二指肠、空肠、回肠组织内虫体数量呈上升趋势，第6天达高峰后稍有下降。肝内虫体数量呈上升趋势，第6天最多；脾内虫体数量在6d达峰值后保持较高水平；实验期间肾、肺和心内虫体数量保持较低水平。结论弓形虫经口感染小鼠后2d，小肠组织出现大量速殖子；同时肺和心内有少量速殖子；4d在脾、6d在肾和肝脏发现虫体；速殖子在上述组织内增殖，并形成假包囊，脑内未发现虫体。     

2005年湛江市、韶关市、汕头市人兽弓形虫感染状况调查

目的了解广东省湛江市、韶关市、汕头市的人兽弓形虫感染情况。方法收集3个地区人、猪、鼠血清，采用酶联免疫吸附试验（ELISA）间接法检测弓形虫IgG抗体。结果湛江市检测猪血清279份，阳性13份，阳性率为4．66％，检测鼠血清93份，阳性7份，阳性率7．53％；韶关市检测人血清42份，阳性0份，检测鼠血清95份，阳性3份，阳性率3．16％；汕头市检测人血清50份，阳性6份，阳性率为12．00％，检测鼠血清100份，阳性3份，阳性率为4．00％。结论汕头市人群存在弓形虫感染，湛江市、韶关市、汕头市兽类存在弓形虫感染，3个地区的鼠类弓形虫感染率无统计学意义，家鼠、野鼠的弓形虫感染率也无统计学意义。     

弓形虫依钙蛋白激酶Calpain—like基因的克隆及表达

目的克隆弓形虫RH株Calpain—like基因片段，构建原核表达载体，诱导表达Calpain-like基因重组蛋白。方法收集、纯化弓形虫RH株速殖子，提取总RNA，在设计合成的引物中引入SalI和EcoRI酶切位点。应用FIT—PCR扩增弓形虫RH株Calpain—like基因片段，插入pGEM—T载体，提取重组质粒，双酶切获得目的基因，亚克隆到原核表达质粒pET32a，重组子经双酶切、PCR和DNA序列分析鉴定，转化大肠杆菌BL21并以IPTG诱导表达。结果从弓形虫RH株速殖子eDNA中扩增出316bp的Calpain-like基因片段；含pET32a／Calpain—like的重组质粒在宿主菌经诱导后，获得与预期分子量相符的表达产物。结论成功地克隆和表达弓形虫RH株Calpain—like基因，弓形虫Calpain—like基因的克隆表达为进一步筛选弓形虫疫苗候选分子和治疗药物的靶位提供了研究基础。     

A seroprevalence study of poliovirus antibody in the population of northern Greece

This study assessed immunity to poliomyelitis in a representative sample of 1064 persons living in northern Greece. Antibody prevalences in the individuals tested were 91.1% (95% confidence interval (CI): 89.4-92.8), 92.1% (95% CI: 90.5-93.7) and 83.1% (95% CI: 80.8-85.4) for poliovirus types 1, 2 and 3, respectively. For poliovirus type 3, a gap in immunity was found in individuals aged 10-29 years. Re-vaccination of adolescents living in northern Greece is suggested to ensure herd immunity and to minimise the risk of importation of wild poliovirus from endemic countries.     

弓形虫速殖子入侵对宿主细胞窖蛋白-1的表达变化

胞膜窖Caveolae,又称细胞膜穴样凹陷,是细胞表面特异性的内陷微区,Caveolin-1是其的标记蛋白.Caveolae结构和caveolin蛋白在多种细胞生理功能方面如细胞信号转导、胆固醇转运、内吞和肿瘤抑制等发挥着重要作用.在动脉硬化、肌营养不良和Alzheimer'S综合征等多种疾病中均发现存在caveola...     

弓形虫PRU株慢性感染小鼠脑部包囊的时空分布特征

目的观察实验小鼠脑内包囊的时空分布特点,为探讨弓形虫感染引起的情志和行为改变提供病理基础。方法弓形虫PRU株经口感染小鼠,经HE染色和免疫组化检测,在显微镜下计算前额、海马、丘脑、小脑和杏仁核部位的包囊个数,然后随机选取上述部位的5个视野拍照,计算包囊的平均密度,并用方差分析比较不同脑组织包囊密度有无统计学差异。结果弓形虫感染30和90d时,HE染色和免疫组化后显微镜观察发现,小鼠不同位置弓形虫包囊密度差异有统计学意义,其中,丘脑的包囊密度最大,其次是前额皮质、海马、杏仁核,小脑的包囊密度最小。丘脑的包囊密度显著高于其他4个脑区（P〈0.01）,小脑的包囊密度显著低于其它4个部位（P〈0.01）,而前额皮质、海马和杏仁核所含的包囊密度差异无统计学意义（P〉0.05）;弓形虫感染1个月时,杏仁核包囊密度明显低于感染3个月（F=18.314,P〈0.001）,但是小脑的包囊密度高于感染3个月（F=18.314,P〈0.001）。结论弓形虫包囊在慢性感染小鼠不同脑组织内的分布具有时空特异性,这可能是其临床表现的病理学基础。     

Different manifestations of Toxoplasma gondii infection in F344 and LEW rats

There is evidence that not only the immune status, but also the genetic predisposition of certain hosts influence the clinical outcome of Toxoplasma gondii infection. By far the majority of our knowledge on genetic and immunological mechanisms involved in control of T. gondii infection has been obtained by studying mouse models, which in terms of clinical outcome of infection differ considerably from humans. Rats which show a rather similar course of infection in comparison to humans have not so far been investigated for effects of genetic differences on course of the infection. In this study we show that, like mice, different strains of rats exhibit a remarkable variation in the number of brain cysts arising from chronic infection. LEW rats seem to be highly resistant to cyst formation, in contrast to F344 rats that are susceptible. In addition, F344 rats express high numbers of γδ T cells during the acute phase of infection, whereas LEW rats express elevated but comparably low numbers of γδ T cells. The RT1 (rat MHC) haplotypes of both strains are identical in the RT1A and RT1B/D regions, which encode the restriction elements for conventional peptide antigens. Consequently, rat strain-specific differences may be useful to define MHC-independent mechanisms of resistance against T. gondii, which may also act in humans. Received: 24 July 1998     

Three years of multi-laboratory external quality control for the molecular detection of Toxoplasma gondii in amniotic fluid in France

Between 2002 and 2004, panels of amniotic fluid containing varying concentrations of Toxoplasma gondii were sent to up to 23 laboratories in France for molecular (PCR-based) detection as part of a national quality assurance initiative in the molecular prenatal diagnosis of toxoplasmosis. Participants were free to enroll and no fees were required. The general level of sensitivity was high, and the rate of false-positive reactions was relatively low. Considerable diversity among PCR methods and primers was revealed. This external quality assurance scheme provided the opportunity to improve laboratory practice and performance, and to increase communication among laboratories involved in making this diagnosis.     

Diagnosis of central nervous system Toxoplasma gondii from the cerebrospinal fluid in a patient with acquired immunodeficiency syndrome

The cytologic diagnosis of central nervous system (CNS) Toxoplasma gondii was established from the cerebrospinal fluid (CSF) in a patient with acquired immunodeficiency syndrome (AIDS) who presented with obstructive hydrocephalus and progressive obtundation. Free extracellular tachyzoites, phagocytized tachyzoites, and cyst forms were identified with Wright staining of cytocentrifuge preparations. Diagnosis was confirmed with indirect immunofluorescent antibody studies. Initial CSF examination was remarkable for low cellularity (24 wbc/mm3) with 42% neutrophils and 58% mononuclear cells on differential examination. Subsequent differential counts of CSF showed persistence of a mixed inflammatory pattern until day 10 at which time a mononuclear pleocytosis was observed. Cerebrospinal chemistries were notable for a low glucose (20 mg/dl), which persisted for 10 days. The cytocentrifuge Wright stain preparation was found to be superior to routine millipore filter Papanicolaou stain preparation for the identification of free tachyzoites and is recommended in the setting of immunodeficient patients with suspected CNS toxoplasmosis.     

弓形虫可遗传及可诱导的RNAi载体系统的构建

目的构建弓形虫可遗传及可诱导的RNAi载体系统，为弓形虫基因功能研究提供工具。方法通过PCR和酶切连接，首先构建弓形虫主要表面抗原1（SAG1）基因启动子驱动的绿色荧光蛋白基因载体pBSK—SAG1／5UTR-eGFP-SAG1／3UTR（pBSK-SAG1／GFP），然后构建以弓形虫热休克蛋白HSPT0基因启动子驱动的反向重复序列RNAi载体pBSK-HSP70／5UTR—IntronC—HSP70／3UTR，将载体pBSK—SAG1／GFP中的SAG1／5UTR—eGFP—SAG1／3UTR片段克隆到载体pBSK—HSP70／5UTR-IntronC—HSP70／3UTR中形成载体pBSK-GFP-Hairpin，再将该载体中的GFP-Hairpin片段克隆到载体pHANA-0．5中形成弓形虫可遗传及可诱导的RNAi载体系统pHANA—hairpin。通过PCR分别扩增SAG1和缓殖子蛋白1（BAG1）基因的正向和反向序列，通过酶切连接，将正向和反向序列克隆到载体pHANA—hairpin中，分别构建靶向SAG1和BAG1基因的RNAi载体pHANA．hairpin／SAG1和pHANA—hairpin／BAGl。结果酶切鉴定和测序结果表明成功构建载体pHANA—hairpin、pHANA-hairpin／SAGl和pHANA-hairpin／BAG1。结论弓形虫可遗传及可诱导的RNAi载体系统成功构建，为下一步基因功能研究奠定基础。     

Interferon-gamma suppresses the growth of Toxoplasma gondii in human fibroblasts through starvation for tryptophan

The effect of human recombinant interferon-γ (IFN-γ) on Toxoplasma gondii in cultured human fibroblasts is predominantly parasitostatic. This effect is dependent upon the induction in the host cell of a potent indoleamine 2,3-dioxygenase that converts tryptophan to N-formylkynurenine. This product is, in turn, degraded to kynurenine by a formamidase. Within 24 h of treatment with IFN-γ most of the tryptophan originally present in the medium is converted to these products together with some minor metabolites. When added to the medium of infected cultures at concentrations equimolar to the tryptophan content, neither N-formylkynurenine nor kynurenine suppresses the growth of T. gondii, although at higher concentrations they are effective. The medium of uninfected cultures treated with IFN-γ for 24 h has no effect on the growth of T. gondii, when transferred to fresh cultures provided that the residual IFN-γ is first removed by ultrafiltration or neutralized with a specific monoclonal antibody. Thus minor metabolites produced from tryptophan in response to IFN-γ and excreted into the medium are not parasitostatic. When cultures treated with IFN-γ for 24 h are incubated with medium that contains [³H]tryptophan, the radioactive amino acid is converted to N-formylkynurenine and kynurenine as rapidly as it enters the cell. This degradation not only results in a very low intracellular concentration of tryptophan but also produces intracellular concentrations of tryptophan metabolites that are significantly higher than the tryptophan concentration in control cells. However, it is unlikely that either metabolite reaches intracellular concentrations that are sufficient to suppress the growth of the parasite. The parasitostatic effect of IFN-γ is most likely to result from the starvation of T. gondii for tryptophan.     

Toxoplasma prevalence among pregnant women in Norway: a cross‐sectional study

Infection by Toxoplasma gondii may lead to complications in the foetus if the mother suffers from primary infection during pregnancy . Previously infected women have produced toxoplasma‐specific IgG antibodies. The most recent study on prevalence of toxoplasma IgG in the Norwegian pregnant population was conducted 20 years ago. The present study is part of a research programme initiated by the Norwegian Institute of Public Health. We aimed to update the knowledge regarding the prevalence of toxoplasma IgG among pregnant women in Norway. In this cross‐sectional study, sera from 1922 pregnant women in Buskerud (992) and Sør‐Trøndelag counties (930) in Norway were collected consecutively. The presence of toxoplasma IgG was identified by values ≥8 IU/mL using an ELISA test. The overall prevalence of toxoplasma IgG seropositivity was 9.3% (95% CI 8.1–10.7); Sør‐Trøndelag 10.4% (95% CI 8.6–12.6) and Buskerud 8.3% (95% CI 6.7–10.2). There was no difference between the counties (p = 0.13), and the result did not differ from prevalences found in 1974 (12.1%) and 1994 (10.7%). We found a higher prevalence among women ≥40 years (OR 2.65, 95% CI 1.30–5.42). The prevalence of toxoplasma IgG among pregnant women in Norway is low and has been stable during the last decades.     

Prevalence and estimated incidence of Toxoplasma infection among pregnant women in Poland: a decreasing trend in the younger population

This study investigated the prevalence of specific Toxoplasma gondii IgG in pregnancy, the incidence of congenital toxoplasmosis and the prevalence trend of T. gondii infection among pregnant Polish women between 1998 and 2003. The study population comprised 4916 women who were admitted to the Polish Mother's Memorial Hospital Research Institute in Łódź. Their sera were tested for specific IgG and IgM antibodies to T. gondii, and the incidence of T. gondii infection was calculated from the increase in prevalence rates of IgG antibodies in various age groups. Specific IgG antibody was found in 41.3% (95% CI 39.9–42.7) of pregnant women, and the prevalence of IgG increased with age. The linear trend was significant (p <0.001), with an annual seroconversion rate of 0.7% (95% CI 0.004–0.010). The risk of primary infection was estimated to be 0.5% for 9 months, i.e., an incidence of 5/1000 pregnancies. Assuming a 30% maternofetal transmission rate, 1.5/1000 neonates were infected in utero. Seroprevalence during the 6-year study period decreased from 45.4% in 1998 to 39.4% in 2003, with a yearly decline in prevalence of 1.0% (p 0.02). The most important contributory factor to this decline was the group of women aged 19–29 years, among whom seroprevalence decreased significantly (p 0.007). Specific IgM was found in 244 (4.9%) women.     

Quantitation of Antibodies to Toxoplasma gondii in Swine Sera by Enzyme-Linked Immunosorbent Assay

Abstract

An enzyme-linked immunosorbent assay (ELISA) was developed for quantitation of antibodies to Toxoplasma gondii in swine sera. Because a commercial anti-swine IgG conjugate was directed also against swine IgM, the conjugate was absorbed with the IgM fraction to eliminate the interference of naturally occurring IgM antibodies that appeared consistently in sera collected from slaughtered pigs at an abattoir. The ELISA values of 0.2 or more observed in most of the sera successfully decreased to less than 0.2 by the use of absorbed conjugate. An attempt to use a protein A conjugate has failed. Evaluation of this system by comparing it with the latex agglutination test provided a high significant correlation, indicating its usefulness for serodiagnosis of swine toxoplasmosis.     

Toxoplasma gondii infection in pregnancy: opportunities and pitfalls of serological diagnosis

Because of its life cycle, the recovery of Toxoplasma gondii from biological samples is often impracticable. Consequently, a serological diagnosis represents the first and the most widely used approach to defining the stage of infection. The detection of IgG, IgM, IgA, IgE and IgG avidity by different methods offers this opportunity. However, the results may be affected by difficulties in interpretation, as the same antibody pattern may have a different valency, contingent upon subjects and clinical settings, e.g., pregnant women vs. neonates, and treated vs. untreated patients. This review describes the various factors that should be taken into account when performing serological tests for T. gondii, as well as the pitfalls that may be encountered during the interpretative process.     

Influence of maternal infections with viral agents or Toxoplasma gondii during pregnancy on fetal IgE production

The importance of maternal infections with Toxoplasma gondii , cytomegalovirus (CMV), Parvovirus B19, respiratory syncytial virus (RSV), and influenza A and B on fetal IgE synthesis was studied in 153 pregnant women. No case of specific IgM activity or viral DNA in cord blood, indicating a congenital infection, was found. From gestational week 15 to delivery, maternal IgG-Ab seroconversion to Parvovirus B19, RSV, influenza A, or influenza B occurred in 47 women. At delivery, serologic signs of past infection with T. gondii were observed in 29 (19%) women, and the corresponding figure for CMV was 117 (77%). The number of women with positive IgG seroconversion during pregnancy or positive IgG-Ab activity toward the studied infectious agents at delivery did not differ significantly among infants with an increased (≥1.3 kU/1; n =51) or with an undetectable (<0.1 kU/l; n =102) cord-blood IgE level. These results show that genetic and other environmental factors probably have a greater influence on fetal IgE synthesis than do maternal infections during pregnancy.