Research Progress on the Association between Schizophrenia and Toxoplasma gondii Infection

Toxoplasma gondii(T.gondii or Tg),is an obligatory intracellular parasite with humans as its intermediate hosts.In recent years,significant correlations between T.gondii infection and schizophrenia have been reported,including the possible mediating mechanisms.Currently,mechanisms and hypotheses focus on central neurotransmitters,immunity,neuroinflammation,and epigenetics;however,the exact underlying mechanisms remain unclear.In this article,we review the studies related to T.gondii infection and schizophrenia,particularly the latest research progress.Research on dopamine(DA)and other neurotransmitters,the blood-brain barrier,inflammatory factors,disease heterogeneity,and other confounders is also discussed.In addition,we also summarized the results of some new epidemiological investigations.

弓形虫(Toxoplasma gondii)的PCR检测方法在笼养猕猴群中的应用

目的建立快速、灵敏、特异及检测结果易判断的PCR方法,并应用于大规模猕猴种群的弓形虫常规检测中。同时比较巢式PCR和单一PCR的一致性。方法根据弓形虫保守基因p30(SAG1)设计了内、外两对进行巢式PCR扩增以及B1基因设计一对引物进行单一PCR扩增,将DNA样本进行10倍倍比稀释,以检测巢式PCR反应的灵敏度;并对医学生物学研究所自繁猕猴共150只进行了弓形虫检测。结果巢式PCR检测法检测限度可达10-3ng/uL,而且方法特异。两种PCR法检测结果基本一致,其中巢式PCR检测阳性率(10%)稍高于单一PCR检测阳性率(8.67%)。结论巢式PCR和一次PCR方法都可应用于猕猴弓形虫的常规检测中,并提示巢式PCR比单一PCR更敏感、检出率更高。

Biological role of surface Toxoplasma gondii antigen in development of vaccine

AIM： To analyze the biological role of the surface antigen of Toxoplasma gondii（Tgondii） in development of vaccine. METHODS： The surface antigen of Tgondii （SAG1） was expressed in vitro. The immune response of the host to the antigen was investigated by detection of specific antibody reaction to SAG1 and production of cytokines. Mice were immunized with recombinant SAG1 and challenged with lethal strain of Tgondii RH. The monoclonal antibody to r-SAG1 was prepared and used to study the effects of SAG1 on Tgondii tachyzoites under electromicroscope. RESULTS： The mice immunized with recombinant SAG1 delayed death for 60 h compared to the control group. The recombinant SAG1 induced specific high titer of IgG and IgM antibodies as well as IFN-y, IL-2 and IL-4 cytokines in mice. In contrast, IL-12, IL-6 and TNF-α were undetectable. When T gondii tachyzoites were treated with the monoclonal antibody to r-SAG1, the parasites were gathered together, destroyed, deformed, swollen, and holes and gaps formed on the surface. CONCLUSION： SAG1 may be an excellent vaccine candidate against T gondii. The immune protection induced by SAG1 against Tgondii may be regulated by both hormone- and cell-mediated immune response.

Investigation on the co-infections of Toxoplasma gondii with PRRSV, CSFV or PCV-2 in swine in part of China

The objective of the present investigation was to estimate the prevalence of Toxoplasma gondii infection and co-infection with porcine reproductive and respiratory syndrome virus(PRRSV), classical swine fever virus(CSFV) and porcine circovirus type 2(PCV-2) in pigs in China. A total of 372 tissues or serum samples collected from pigs distributed in 9 provinces/municipalities of China during the period from February 2011 to November 2012 were assayed for T. gondii antigens and antibodies using enzyme linked immunosorbent assay(ELISA) technique, while the PCR was designed for the detection of the PRRSV, CSFV and PCV-2, respectively. The total positive rate of T. gondii, PRSSV, CSFV and PCV-2 was 9.14%(34/372), 50.00%(186/372), 37.10%(138/372) and 3.23%(12/372), respectively. Among the 34 T. gondii positive samples, 26 samples were simultaneously infected with T. gondii and viruses, while the remaining eight samples were infected with T. gondii alone. In addition, the co-infection rate of T. gondii with PRSSV, T. gondii with PRSSV and CSFV, T. gondii with PRSSV and PCV-2, T. gondii with CSFV and PCV-2, T. gondii with PRSSV, CSFV and PCV-2 was 1.61%(6/372), 4.03%(15/372), 0.27%(1/372), 0.27%(1/372) and 0.81%(3/372), respectively. The results of the present survey revealed that PRRSV and CSFV were the common pathogens co-existing with porcine toxoplasmosis in China, and both of them could increase the chances of T. gondii infection in pig. This is the first report of T. gondii co-infections with viruses in pigs. It is very important to understand the interactions of parasite and virus, and can be used as reference data for the control and prevention of co-infections of T. gondii and viruses in pigs.

Isolation and Molecular Characterization of Toxoplasma gondii from Chickens in China

One strain of Toxoplasma gondii was successfully isolated from chickens in China by bioassay in mice. Antibodies and circulating antigens of T. gondii were assayed by the ELISA kits in 100 free range chickens from a rural area surrounding Funing, China. Fifty-three chickens were antibody-positive and 21 chickens were antigen positive. Hearts, brains, spleens, lungs, livers, and kidneys of 21 antibody or antigen-positive chickens were bioassayed in mice. One strain of T. gondii was isolated from 1 of 21 （4.76%） chickens. The isolated T. gondii killed all of the inoculated mice. Genotyping of this isolate using polymorphisms at the loci 5′-SAG2, 3′-SAG2, SAG3, cB21-4, L358, BTUB, and GRA6 revealed that it was Type I. These indicated that it was virulent for mice. This is the first report of isolation of T. gondii from chickens in China.

Toxoplasma gondii infection among chronic hepatitis C patients:A casecontrol study

Objective:To determine the detection rate of anti-Toxoplasma gondii(T.gondii) IgG and IgM in chronic HCV patients attending the Department of Tropical Medicine Mansoura University hospital in Egypt.Methods:This study included 120 adult chronic HCV patients.81 decompensate cirrhosis(late-stage)and 39 chronic HCV non cirrhotic patients(early-stage) and40 healthy blood donors as controls.Serum samples uere examined for anti-Toxoplasma IgM and anti-Toxoplasma IgG antibodies by ELISA.Real-time RT-polymerase chain reaction assay was done for quantitation of hepatitis C virus.Results:Anti-T.gondii IgG antibodies were detected in 75(92.6%) of 81 late-stage cirrhotic patients.30(76.9%) of the 39 chronic HCV non cirrhotic patients(early-Stage) and in 6(15ft) of 40 controls with statistically significant difference(P<0.001).Anti-T.gondii IgM antibodies were found in 11(13.6%) in late stage patients,5(12.8%)in early stage and in 3(7.5%) of controls with no statistical significant difference(P=0.610).There was no correlation between stage of fibrosis and IgM or IgG antibodies positivity in our studied groups(P=0.526).High IgG levels significantly correlated with high viral load(P=0.026).Conclusions:Our findings suggest that the serious opportunistic T.gondii infection represent a potential significant risk for chronic HCV patients.So.toxoplasmosis should be considered in their investigations and follow-up.

Introduction of protocols for mass production of Toxoplasma gondii tachyzoites of the genotype II PRU strain

Background:Few investigations of genotype II of Toxoplasma gondii,the most preva-lent form of the Toxoplasma parasite in humans,have been carried out on due to the rapid conversion of tachyzoites to bradyzoites in its life cycle.The current study aimed to create animal and in vitro models for production of the tachyzoites of the Prugniaud(PRU)genotype II strain.Methods:To develop an immunocompromised model and obtain tachyzoites of the PRU strain,BALB/c mice were orally treated with dexamethasone(10 mg/kg),cyclo-phosphamide(36 mg/kg),and cyclosporine(18 mg/kg)from 5 days prior to inocula-tion.Then,10-15 tissue cysts of PRU strain were inoculated intraperitoneally into the mice.The tachyzoites obtained from mice were then cultivated in a HeLa cell culture.The resulting yield of tachyzoites was cryopreserved in 92%fetal calf serum,8%dimethyl sulfoxide.The infectivity of these tachyzoites was evaluated using in vivo and in vitro examinations.Results:Numerous tachyzoites were observed in the peritoneal fluid of the immuno-suppressed mice within 10-15 days after inoculation,and many tachyzoites were har-vested from the HeLa cell culture.Trypan Blue staining showed 80%viability of the tachyzoites recovered from cryopreservation and this was confirmed by HeLa cell culture.In addition,mice infected intraperitoneally with the recovered tachyzoites presented with cysts in the brain after 2 months.Conclusion:We have developed an animal model for mass production of T.gondii tachyzoites of the PRU strain.This method can provide fresh viable tachyzoites of Toxoplasma gondii for use as and when required in future investigations.

The level of chemerin and adipocyte fatty acid binding protein in Toxoplasma gondii seropositive obese individuals

Objective: To know the difference between chemerin and adipocyte fatty acid-binding protein(AFABP) levels in obese individuals with positive Toxoplasma gondii(T. gondii)immunoglobulin G(IgG) compared with negative T. gondii IgG.Methods: This study is a cross-sectional study by using consecutive sampling methods conducted from January to April 2013. The subjects were 57 obese individuals who were divided into obese group of positive and negative T. gondii IgG. The level of chemerin,AFABP and T. gondii IgG was done by ELISA. The data were analyzed by independent t test.Results: The results showed that the level of chemerin of positive T. gondii IgG group was significantly higher than the negative T. gondii IgG group [(70.0 ± 16.5) vs.(64.4 ± 16.1) pg/mL; P = 0.003], but there was not significant AFABP difference between seropositive and negative IgG groups [(83.6 ± 41.9) vs.(74.2 ± 36.7) pg/mL; P = 0.598].Conclusions: It can be concluded that the level of chemerin of seropositive T. gondii IgG was higher than that in the negative T. gondii IgG group.

CD2 deficiency partially prevents small bowel inflammation and improves parasite control in murine Toxoplasma gondii infection

To investigate whether bowel inflammation and/or parasite control is altered in the absence of the T cell adhesion molecule CD2.

Toxoplasma gondii infection can induce retinal DNA damage: an experimental study

AIM:To detect whether Toxoplasma gondii(T.gondii)infection of mice can induce retinal DNA damage.METHODS:A total of 20 laboratory-bred male Swiss albino mice were used and divided into four groups:control group(non-infected animals);T.gondii infected group;immunosuppressed infected group;and infected group treated with sulfadiazine and pyrimethamine.Mice eyes were collected 6wk post infection and retinas were obtained.Each retina was immediately processed for comet assay and the frequency of tailed nuclei(DNA damage)was calculated.In addition,retinal DNA damage was revealed by various comet assay parameters that were provided by the image analysis software including tail length,percentage of DNA in the tail,percentage of tailed cells and tail moment.RESULTS:The obtained results showed that T.gondii infection induced a statistically significant increase in the frequency of tailed nuclei,tail length,percentage of DNA in the tail,and tail moment in mice retinal cells compared to the control group(which showed some degree of DNA damage).In immunosuppressed infected group,retinal DNA damage was severing and there was significant increase in various comet assay parameters compared to both control and infected groups.After treatment with sulfadiazine and pyrimethamine,retinal DNA damage decreased and all comet assay parameters showed a statistical significant decrease compared to infected groups.CONCLUSION:T.gondii infection can induce DNA damage in mice retinal cells.

Research Progress in NTPase of Toxoplasma gondii and Other Protozoa

Nucleoside triphosphate hydrolase （NTPase） is a multifunctional enzymatic family widely existing in vivo. They can hydrolyze NTP to NMP or dNTP to dNMP to produce energy. In this article, the structure of Toxoplasma gondii NTPase is analyzed. The research progress in NT- Pase of Toxoplasrna gondii, Trypanosoma cruzi, Sarcocystis neurona and Neospora caninum was briefly reviewed.

刚地弓形虫(Toxoplasma gondii)不同地理株对钴^(60)丙线敏感性的比较研究

对中国NT株弓形虫和美国的ME—49株及TS—2株弓形虫的包囊进行了抗丙线耐受性的比较，试验采用上述三株弓形虫包囊为材料，分设0．4、0．5、0．6、0．7、1．OKGy丙线剂量组和不经照射处理的对照组。通过感染小鼠和幼猫的生物学检测，求得使包囊丧失感染性的丙线剂量，得出不同株包囊对丙线的敏感性。结果表明：丙线控制中国NT株的最小有效剂量为0．55KGy，控制ME—49株和TS—2株的感染性的最小有效剂量为0．60KGy。两者接近，提示这三个地理株的弓形虫包囊对钴60丙线的敏感性虽略有差异，但其差异并不明显。

Detection of Antibodies against Toxoplasma gondii by ELISA with Recombinant Microneme Protein 3

[Objective] To develop a new method for serodiagnosis of swine toxoplasmosis. [Method] With the purified recombinant microneme protein 3 （rMIC3） as coating antigens, an indirect ELISA was developed for detection of antibodies against Toxoplasma gondii. [ Result] The optimal working concentration of rMIC3 was 3. 40 ug/ml, and the optimal degree of dilution of sera was 1：160. Cross-reaction was not observed between the Toxoplasma gondii-positive sera and the positive sera against classical swine fever virus or some other pathogens. The developed ELISA had 92.56% coincidence rate with latex agglutination test. [ Conclusion] The developed ELISA is sensitive, rapid, specific and reproducible, and thus it can be applied in serodiagnosis and seroprevalence investigation of swine toxoplasmosis.

Pituitary prolactin adenoma with Toxoplasma gondii infection

Objective: To report two recent cases of pituitary adenoma associated with Toxoplasma gondii (T.Gondii) infection.Methods: Histological changes were observed in H & E and PAS staining sections microscopically.Immunohistochemistry was performed to classify the pituitary tumors and to confirm the diagnosis of T.gondii.Results: The cases were 43- and 19-year-old females, in which the latter one was a recurring case, and radiology examination showed that tumors existed in sellar region.Microscopically, the tumors consisted of small homogenous polygonal or round cells with abundant eosinophilic granular cytoplasm.Immunohistochemistry revealed they were prolactin-producing adenomas.Interestingly, we found toxoplasma infection in the tumor tissues, being confirmed by T.gondii sepicific antibody immunohistochemistry.Conclusion: The association of pituitary adenoma with toxoplasma raises the possibility that T.gondii may be involved in the development of certain cases of pituitary adenoma.

Low Sero-Prevalence of Toxoplasma gondii IgG and IgM Antibodies in HIV Sero-Positive Patients Attending National Hospital, Abuja, Nigeria

Several researchers have investigated the association of numerous opportunistic pathogens with HIV, little is documented on its association with T. gondii in our environment. We investigated the prevalence of T. gondii immunoglobulins G and M （IgG and IgM） in HIV positive individuals in relation to their cluster of differentiation 4 （CD4） cells count. IgG, IgM and CD4 were assayed using enzyme immunoassay （EIA） and flowcytometry respectively. 341 HIV positive individuals were studied in the present research, 30 （8.7%） of them had T. gondii IgG and IgM, 297 subjects had CD4 cells count a range of 200-400 cells/μL, 27 （9.7%） and 2 （0.6%） of which had T. gondii IgG and IgM respectively. Of the 44 HIV positive subjects with CD4 〉 400 cells/μL, one （2.2%） was positive for T. gondii IgG. In the control group, all the 177 had CD4 〉 400 cells/μL of which, one （0.5%） had T. gondii IgG. The prevalence of T. gondii infection was significantly higher in HIV positive individuals than in controls （P 〈 0.05）. Male subjects in the age bracket 18-30 years had significantly higher prevalence when compared to other groups （P 〈 0.05）. Although the present findings revealed a low prevalence of T. gondii antibodies in HIV infection, this suggests that a differential toxoplasmosis diagnosis is also necessary in cases of encephalitis in HIV infection.

The First Report on Sero-Prevalence of Toxoplasma gondii in Working Horses and Donkeys in the Sudan

A serological survey of Toxoplasma gondii was conducted using LAT （Latex Agglutination Test） on 205 sera collected from working horses and donkeys in Khartoum State, Sudan. The overall sero-prevalence of the investigated equines was 32.7%. Antibodies to T. gondii were found in 38% of 100 horses and 27.6% of 105 donkeys. The titers were 1：2 （4 heads）, 1：4 （11 heads）, 1：8 （13 heads）, 1：16 （17 heads）, 1：32 （5 heads）, 1：64 （10 donkeys） and 1：128 （7 donkeys）. Neither the area nor animal species showed significant differences between the investigated groups. However, age was reported to show significant （P 〈 0.05） effect on equine toxoplasmosis in the Sudan. This is the first report on equine toxoplasmosis in the Sudan.

Protein Phosphatase 2C of Toxoplasma Gondii Interacts with Human SSRP1 and Negatively Regulates Cell Apoptosis

Objective The protozoan Toxoplasma gondii expresses large amounts of a 37 kDa Type 2C serine-threonine phosphatase,the so-called TgPP2 C which has been suggested to contribute to parasite growth regulation.Ectopic expression in mammalian cells also indicated that the enzyme could regulate growth and survival.In this study,we aimed to investigate the interaction of TgPP2 C with human SSRP1（structure-specific recognition protein 1） and the effects of TgPP2 C on cell viability.Methods The yeast two hybrid system,His-tag pull-down and co-immunoprecipitation assays were used to confirm the interaction of TgPP2 C with SSRP1 and determine the binding domain on SSRP1.The evaluation of cell apoptosis was performed using cleaved caspase-3 antibody and Annexin-V/PI kit combined with flow cytometry.Results We identified human SSRP1 as an interacting partner of TgPP2 C.The C-terminal region of SSRP1 including the amino acids 471 to 538 was specifically mapped as the region responsible for interaction with TgPP2 C.The overexpression of TgPP2 C down-regulated cell apoptosis and negatively regulated apoptosis induced by DRB,casein kinase II（CKII） inhibitor,through enhanced interaction with SSRP1.Conclusion TgPP2 C may be a parasitic factor capable of promoting cell survival through interaction with the host protein SSRP1,thereby creating a favorable environment for parasite growth.

Analysis of Proteotranscriptomics Landscape Reveals Differentially Regulated Pathways in Toxoplasma gondii Infected Mouse Liver

Toxoplasma gondii (T. gondii) an intracellular protozoan parasite, infects mammals including human population world-wide. Upon primary infection, the parasite contributes to mild flu like symptoms in immune competent host, but life threatening complication is seen in immune compromised patients and in pregnant women. Understanding the host-parasite interaction is critical for understanding the pathogenesis and biology parasite reactivation in the host. In this study, we used proteotrasncriptomics analyses by integrating the transcriptomics and proteomics data of T. gondii infected mouse liver to uncover the effector molecules responsible for disease pathogenesis that can be used as candidate markers for diagnosis and drug target. With this aim, we systematically integrated transcriptomicand proteomic data, representing the parasite infected mouse liver. Out of 2758 differentially expressed genes (DEGs) and 301 differentially expressed proteins (DEPs), 159 overlapping genes were identified. Among them, 86 genes were upregulated and 72 were downregulated in their respective mRNA and protein levels in the infected condition. Gene Ontology (GO) analysis revealed that the upregulated genes were mostly associated with immune system processes whereas the downregulated genes were involved in oxidation-reduction process and metabolism of lipid, and fatty acids. Protein-protein interaction (PPI) network analysis uncovered an interaction-hub including, Psmb8, Psmb9 and Tap1 for upregulated proteins and Cyp1A2, Cyp4A10 and Cyp3A11 for down-regulated proteins. Further studies are needed to validating these effector molecules. These molecules are likely to play a vital role in disease pathogenesis, as well as can be used as potential diagnostic marker and drug target candidates.

Serologic Detection of <i>Toxoplasma gondii</i>in Cat Owners Residing at Dhaka Metropolitan Area of Bangladesh

Toxoplasma gondii is a zoonotic protozoan that can infect any warm-blooded mammal. T. gondii infects about one-third of the human population on the planet. Infection with the parasite in human causes toxoplasmosis that may pose a high risk in immunocompromised individuals under certain clinical conditions. Cats are the ultimate hosts of T. gondii where oocysts are formed through mating of male and female gametes. Infected cats can expel T. gondii oocysts in their feces, and thereby capable of pass on a disease to humans and other animals through consumption of foods, vegetables and water that are polluted with cat feces. The study was conducted to detect the presence of anti-T. gondii IgM and IgG antibodies in the blood of individuals with or without cat contact to determine if there is any relationship between cat contacts and T. gondii infection in humans. To address this, we enrolled subjects who contacted with the cat as target group and individuals with no cat contact as control group. Following register of different demographic data (including age, sex, education, foods habit, income status, etc.), whole blood from each enrolled subject of both the target group and control group was collected for serum preparation. T. gondii infected subjects were detected by Toxo Rapid test kit through identifying anti-T. gondii IgM and IgG antibodies in their serum. We found that only three out of twenty subjects who were in contact with cat showed positive IgG response while IgM antibody response was absent for all subjects. When compared with the data from control group, we did not find any significant association (p = 0.33) of cat contact with the transmission of T. gondii into human. However, with this small number of study subjects, we cannot conclusively say that there is no impact of cat contact on the transmission of T. gondii into human. Whether any association exists or not can be ascertained with a large number of subjects from different areas of Bangladesh in a future study in the population.

Genotyping of Toxoplasma gondii in Sheep and Cattle Meat Using PCR-RFLP Technique

distribution.The main source of infection for humans is livestock and meat-producer animals.The relationships between Toxoplasma genotype and biological characteristics of the parasite have already been identified.According to the pathogenicity of the parasite in laboratory animals,Toxoplasma is divided into three genotypes included type I,II and III.Understanding the genotype of the parasite,could help us to predict clinical features and severity of disease.The aim of this study was to identify genotypes of T.gondii in cattle and sheep meat and meat products in Ahvaz city southwest of Iran.One hundred and ninety samples of tongue,heart and muscles of sheep and cattle and meat products,including sausages and burgers,were collected from slaughterhouses and stores.To identify Toxoplasma gondii,DNA were extracted from samples and B1 gene were amplified by specific primers.To determine the genotype of T.gondii,PCR-RFLP was done on positive samples using by amplifying GRA6 gene and endonuclease Msel enzyme.Data analysis showed that the strain of the parasite in all positive samples belonged to genotype I.In this study the predominant Toxoplasma genotype was type I which can cause severe clinical symptoms in immunocompromised patients.Further research is needed to determine the genotype of the parasite in humans and other animals.