The study of antigenic epitopes from Toxoplasma gondii has not only enhanced our understanding of the structure and function of antigens, the reactions between antigens and antibodies, and many other aspects of immunology, but it also plays a significant role in the development of new diagnostic reagents and vaccines. In the present study, T. gondii GRA6 epitopes were identified using bioinformatics tools and a synthetic peptide technique. The potential B cell epitopes of GRA6 predicted by bioinformatics tools concentrated upon 3 regions of GRA6, 1-20 aa, 44-103 aa, and 172-221 aa. Ten shorter peptides from the 3 regions were synthesized and assessed by ELISA using pig sera from different time points after infection. Three of the 10 peptides (amino acids 44-63, 172-191, and 192-211) tested were recognized by all sera and determined to be immunodominant B-cell epitopes of GRA6. The results indicated that we precisely and accurately located the T. gondii GRA6 epitopes using pig sera collected at different time points after infection. The identified epitopes may be very useful for further studies of epitope-based vaccines and diagnostic reagents.
Allergy and Immunology ; Antibodies ; Computational Biology ; Enzyme-Linked Immunosorbent Assay ; Epitopes ; Epitopes, B-Lymphocyte* ; Indicators and Reagents ; Peptides ; Toxoplasma* ; Vaccines

The sodium hydrogen exchanger 1 (NHE1), which functions in maintaining the ratio of Na+ and H+ ions, is widely distributed in cell plasma membranes. It plays a prominent role in pH balancing, cell proliferation, differentiation, adhesion, and migration. However, its exact subcellular location and biological functions in Toxoplasma gondii are largely unclear. In this study, we cloned the C-terminal sequence of T. gondii NHE1 (TgNHE1) incorporating the C-terminal peptide of NHE1 (C-NHE1) into the pGEX4T-1 expression plasmid. The peptide sequence was predicted to have good antigenicity based on the information obtained from an immune epitope database. After induction of heterologous gene expression with isopropyl-b-D-thiogalactoside, the recombinant C-NHE1 protein successfully expressed in a soluble form was purified by glutathione sepharose beads as an immunogen for production of a rabbit polyclonal antiserum. The specificity of this antiserum was confirmed by western blotting and immunofluorescence. The antiserum could reduce T. gondii invasion into host cells, indicated by the decreased TgNHE1 expression in T. gondii parasites that were pre-incubated with antiserum in the process of cell entry. Furthermore, the antiserum reduced the virulence of T. gondii parasites to host cells in vitro, possibly by blocking the release of Ca2+. In this regard, this antiserum has potential to be a valuable tool for further studies of TgNHE1.
Animals ; Cell Line ; Immune Sera/genetics/immunology/*metabolism ; Male ; Mice ; Protozoan Proteins/genetics/*metabolism ; Rabbits ; Recombinant Proteins/immunology ; Sheep ; Sodium-Hydrogen Antiporter/genetics/immunology/*metabolism ; Toxoplasma/genetics/immunology/*metabolism ; Toxoplasmosis/parasitology/prevention & control

The breeding of domestic rabbits (Oryctolagus cuniculus) for human consumption has a long tradition in China. Infections that can affect the production of meat or even be transmitted from animals to humans are important to monitor, especially for public health reasons as well as for their impact on animal health. Thus, a total of 1,132 domestic rabbit sera from 4 regions in China were collected for serological screening for Encephalitozoon cuniculi and for Toxoplasma gondii by ELISA and modified agglutination test (MAT), respectively. Antibodies to E. cuniculi were detected in 248/1,132 (21.9%) sera tested while antibodies against T. gondii revealed a seroprevalence of 51/1,132 (4.5%). We believe that the present results are of epidemiological implications and public health importance due to the acknowledged susceptibility of humans to E. cuniculi and T. gondii infections. Therefore, routine screening tests of domestic rabbits are proposed considering the zoonotic potential of these parasites.
Animals ; Animals, Domestic/blood/microbiology/parasitology ; Antibodies, Fungal/*blood ; Antibodies, Protozoan/*blood ; China/epidemiology ; Encephalitozoon cuniculi/*immunology/isolation & purification ; Encephalitozoonosis/blood/microbiology/*veterinary ; Female ; Male ; Rabbits/blood/microbiology/parasitology ; Seroepidemiologic Studies ; Toxoplasma/*immunology/isolation & purification ; Toxoplasmosis, Animal/*blood/parasitology

Toxoplasmosis is considered as an important risk factor for bad obstetric history (BOH) and one of the major causes of congenitally acquired infections. The present study aimed to estimate the seropositivity of T. gondii infection and associated risk factors among the attendees of high risk pregnancy and low risk antenatal care clinic of Minia Maternity and Pediatric University Hospital, Minia, Egypt. The study was carried out from April 2013 to April 2014 through 2 phases, the first phase was case-control study, and the second phase was follow-up with intervention. A total of 120 high risk pregnant and 120 normal pregnant females were submitted to clinical examinations, serological screening for anti-Toxoplasma IgM and IgG antibodies by ELISA, and an interview questionnaire. Seropositive cases were subjected to spiramycin course treatment. The results showed that the seroprevalence of toxoplasmosis in high-risk pregnancy group was 50.8%, which was significantly different from that of normal pregnancy group (P<0.05). Analysis of seropositive women in relation to BOH showed that abortion was the commonest form of the pregnancy wastage (56.5%). The high prevalence of T. gondii seropositive cases was observed in the age group of 21-30 years. Post-delivery adverse outcome was observed in 80.3% of high-risk pregnancy group compared to 20% of normal pregnancy group. There was a statistically significant relationship between seropositivity and living in rural area, low socioeconomic level, and undercooked meat consumption (P<0.05). Serological screening for anti-Toxoplasma antibodies should be routine tests especially among high-risk pregnant women.
Adolescent ; Adult ; Antibodies, Protozoan/*blood ; Case-Control Studies ; Egypt ; Enzyme-Linked Immunosorbent Assay ; Feeding Behavior ; Female ; Follow-Up Studies ; Hospitals, University ; Humans ; Immunoglobulin G/blood ; Immunoglobulin M/blood ; Pregnancy ; Pregnancy Complications, Infectious/*epidemiology ; Risk Factors ; Rural Population ; Seroepidemiologic Studies ; Socioeconomic Factors ; Surveys and Questionnaires ; Toxoplasma/*immunology ; Toxoplasmosis/*epidemiology ; Young Adult

Toxoplasmosis, caused by Toxoplasma gondii, is a parasitic zoonosis with worldwide distribution. The present study investigated the prevalence of T. gondii in dogs in Zhanjiang city, southern China, using both serological and molecular detection. A total of 364 serum samples and 432 liver tissue samples were collected from the slaughter house between December 2012 and January 2013 and were examined for T. gondii IgG antibody by ELISA and T. gondii DNA by semi-nested PCR based on B1 gene, respectively. The overall seroprevalence of T. gondii IgG antibody was 51.9%, and T. gondii DNA was detected in 37 of 432 (8.6%) liver tissue samples. These positive DNA samples were analyzed by PCR-RFLP at 3'- and 5'-SAG2. Only 8 samples gave the PCR-RFLP data, and they were all classified as type I, which may suggest that the T. gondii isolates from dogs in Zhanjiang city may represent type I or type I variant. This study revealed the high prevalence of T. gondii infection in dogs in Zhanjiang city, southern China. Integrated measures should be taken to prevent and control toxoplasmosis in dogs in this area for public health concern.
Animals ; Antibodies, Protozoan/blood ; China/epidemiology ; Dog Diseases/epidemiology/*parasitology ; Dogs ; Female ; Genotype ; Liver/parasitology ; Male ; Toxoplasma/classification/genetics/immunology/*isolation & purification ; Toxoplasmosis, Animal/blood/epidemiology/*parasitology

Although Toxoplasma gondii infection in primary school children has been investigated in many countries, limited surveys have been available in primary school children in China. In the present study, we report the seroprevalence of T. gondii infection in primary school children in Shandong province, China. Sera from 6,000 primary school children were evaluated for T. gondii antibodies with ELISA. The overall seroprevalence of T. gondii infection was 16.0% (961/6,000), of which 14.5% (870/6,000) were positive for anti-T. gondii IgG antibodies, 3.4% (206/6,000) positive for IgM, and 1.9% (115/6,000) were positive for both IgG and IgM. The results of the present investigation indicated a high seroprevalence of T. gondii infection in primary school children in Shandong province, China. Therefore, effective measures should be taken to prevent and control T. gondii infection in primary school children in this province. To the best of our knowledge, this is the first report of T. gondii seroprevalence in primary school children in Shandong province, China.
Antibodies, Protozoan/blood ; Child ; China/epidemiology ; Female ; Humans ; Male ; Seroepidemiologic Studies ; Students ; Toxoplasma/genetics/immunology/*isolation & purification ; Toxoplasmosis/blood/*epidemiology/*parasitology

The aim of this cross sectional case control study was to examine the serofrequency and serointensity of Toxoplasma gondii (Tg) IgG, IgM, and DNA among patients with schizophrenia. A total of 101 patients with schizophrenia and 55 healthy controls from Sungai Buloh Hospital, Selangor, Malaysia and University Malaya Medical Center (UMMC) were included in this study. The diagnosis of schizophrenia was made based on the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV). The presence of Tg infection was examined using both indirect (ELISA) and direct (quantitative real-time PCR) detection methods by measuring Tg IgG and IgM and DNA, respectively. The serofrequency of Tg IgG antibodies (51.5%, 52/101) and DNA (32.67%, 33/101) among patients with schizophrenia was significantly higher than IgG (18.2%, 10/55) and DNA (3.64%, 2/55) of the controls (IgG, P=0.000, OD=4.8, CI=2.2-10.5; DNA, P=0.000, OD=12.9, CI=2.17-10.51). However, the Tg IgM antibody between patients with schizophrenia and controls was not significant (P>0.005). There was no significant difference (P>0.005) in both serointensity of Tg IgG and DNA between patients with schizophrenia and controls. These findings have further demonstrated the strong association between the active Tg infection and schizophrenia.
Adolescent ; Adult ; Aged ; Antibodies, Protozoan/*blood ; Case-Control Studies ; Cross-Sectional Studies ; DNA, Protozoan/*blood ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Immunoglobulin G/blood ; Immunoglobulin M/blood ; Malaysia ; Male ; Middle Aged ; Real-Time Polymerase Chain Reaction ; Schizophrenia/*complications ; Seroepidemiologic Studies ; Toxoplasma/classification/genetics/immunology/*isolation & purification ; Toxoplasmosis/*epidemiology/immunology/*parasitology ; Young Adult

The aim of this cross sectional case control study was to examine the serofrequency and serointensity of Toxoplasma gondii (Tg) IgG, IgM, and DNA among patients with schizophrenia. A total of 101 patients with schizophrenia and 55 healthy controls from Sungai Buloh Hospital, Selangor, Malaysia and University Malaya Medical Center (UMMC) were included in this study. The diagnosis of schizophrenia was made based on the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV). The presence of Tg infection was examined using both indirect (ELISA) and direct (quantitative real-time PCR) detection methods by measuring Tg IgG and IgM and DNA, respectively. The serofrequency of Tg IgG antibodies (51.5%, 52/101) and DNA (32.67%, 33/101) among patients with schizophrenia was significantly higher than IgG (18.2%, 10/55) and DNA (3.64%, 2/55) of the controls (IgG, P=0.000, OD=4.8, CI=2.2-10.5; DNA, P=0.000, OD=12.9, CI=2.17-10.51). However, the Tg IgM antibody between patients with schizophrenia and controls was not significant (P>0.005). There was no significant difference (P>0.005) in both serointensity of Tg IgG and DNA between patients with schizophrenia and controls. These findings have further demonstrated the strong association between the active Tg infection and schizophrenia.
Adolescent ; Adult ; Aged ; Antibodies, Protozoan/*blood ; Case-Control Studies ; Cross-Sectional Studies ; DNA, Protozoan/*blood ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Immunoglobulin G/blood ; Immunoglobulin M/blood ; Malaysia ; Male ; Middle Aged ; Real-Time Polymerase Chain Reaction ; Schizophrenia/*complications ; Seroepidemiologic Studies ; Toxoplasma/classification/genetics/immunology/*isolation & purification ; Toxoplasmosis/*epidemiology/immunology/*parasitology ; Young Adult

OBJECTIVETo prepare and characterize rabbit polyclonal antibodies against Toxoplasma gondii vacuolar proton pyrophosphatase type I (TgVP1).

METHODS AND RESULTSTwo synthesized peptides TgVP1-1 and TgVP1-2 as the haptens were conjugated with KLH to immunize rabbits. Indirect ELISA showed that the titers of rabbit anti-TgVP1-1 polyclonal antibody and rabbit anti-TgVP1-2 polyclonal antibody reached 1:128 000. Western blotting results revealed that both purified polyclonal antibodies could specifically bind to a purified 85 kD T. gondii protein predicted as TgVP1. The protein detected by these two polyclonal antibodies was distributed in the cytoplasm of T. gondii tachyzoite, and this distribution pattern was consistent with that of acidocalcisome.

CONCLUSIONThe peptide-based method of antibody generation is efficient and the obtained TgVP1 polyclonal antibodies possess a high specificity to facilitate further study of T. gondii acidocalcisome and the diagnosis of toxoplasmosis.

Animals ; Antibodies ; immunology ; Blotting, Western ; Enzyme-Linked Immunosorbent Assay ; Protozoan Proteins ; immunology ; Pyrophosphatases ; immunology ; Rabbits ; Toxoplasma ; enzymology

OBJECTIVETo study the changes and significance of lymphocyte sunsets and serum interferon-γ (IFN-γ) levels in children with toxoplasma infection.

METHODSThirty-four children who were newly diagnosed with toxoplasma infection (TOX-IgM+ group) between January 2011 and April 2014, 12 children who had ever been diagnosed with toxoplasma infection (TOX-IgG+ group), and 54 healthy children (control group) were enrolled. The percentages of CD4+, CD8+, CD19+ and NK cells in peripheral blood lymphocytes were detected by flow cytometry. Serum levels of IFN-γ were measured using ELISA.

RESULTSThe percentages of CD4+ cells and the CD4+/CD8+ ratio in the TOX-IgM+ group were significantly lower than in the TOX-IgG+ and control groups, while the percentages of CD8+ and NK cells and serum IFN-γ levels were significantly higher than in the other two groups (P<0.05). The TOX-IgG+ group had higher serum IFN-γ levels than the control group (P<0.05). There was a positive correlation between the percentage of CD8+ cells and serum IFN-γ levels in the TOX-IgM+ group (r=0.756; P<0.05).

CONCLUSIONSCD4+, CD8+ and NK cells may play important roles in the resistance against toxoplasma infection by promoting the secretion of cytokines.

Antibodies, Protozoan ; blood ; CD4-CD8 Ratio ; Child, Preschool ; Female ; Humans ; Immunoglobulin G ; blood ; Immunoglobulin M ; blood ; Infant ; Interferon-gamma ; blood ; Lymphocyte Subsets ; immunology ; Male ; Toxoplasma ; immunology