The protozoan parasite Toxoplasma gondii produces a family of microneme proteins that are thought to play diverse roles in aiding the parasite's intracellular existence. Among these, TgMIC2 has a putative function in parasite adhesion to the host cell to initiate the invasion process. The invasion process may be localized and inhibited by monoclonal antibodies against the protein(s) involved. Here we report on the construction of a phage-displayed single-chain variable fragment (scFv) library from mice immunized with whole T. gondii parasites. The library was subsequently panned against recombinant TgMIC2 (rpTgMIC2) and 2 different groups of antibody clones were obtained, based on fingerprinting and sequencing data. The expressed recombinant scFv antibody was able to recognize rpTgMIC2 in a Western blot detection experiment. These results show that the phage display technology allows quick and effective production of monoclonal antibodies against parasite antigens. By panning the scFv-displayed library, we should be able to obtain a plethora of multi-functional scFv antibodies towards T. gondii proteins.
Management of inflammation after surgery for recalcitrant anterior uveitis is challenging. Herein, we report successful treatment using intracameral injection of recombinant tissue plasminogen activator (rtPA) in two patients with recalcitrant anterior uveitis, due to infective uveitis and Vogt-Koyanagi-Harada disease, respectively. A 40-year-old woman presented with bilateral redness and vision reduction that had persisted 2 weeks. She also had bilateral anterior uveitis, vasculitis, retinitis, and optic disc swelling. Serology was positive for Bartonella henselae and Toxoplasma gondii. She was treated using long-term systemic corticosteroids and appropriate antibiotics. Our second case; a healthy 30-year-old man with bilateral eye redness and reduced vision without pain, and associated with headache and tinnitus for 1 weeks. He showed bilateral granulomatous inflammation with vitritis, choroiditis, retinitis, and hyperemic optic disc. The patient was diagnosed with Vogt-Koyanagi-Harada disease and treated with systemic corticosteroids. Both patients developed secondary cataracts and glaucoma that necessitated surgical intervention. Persistent chronic inflammation led to the formation of a thick fibrin membrane anterior to the intraocular lens (IOL) after phacoemulsification surgery with IOL implantation. This membrane was removed surgically, and intracameral injection of rtPA (25 μg) was carried out. The persistent inflammation had resolved and visual acuity had significantly improved within 1 week of intracameral rtPA injection. There were no reported ocular or systemic side effects. Intracameral rtPA is beneficial in patients with recalcitrant anterior uveitis who have undergone intraocular surgery. In most cases, surgical intervention improves the patients' vision. Intracameral rtPA should be considered in cases of persistent inflammation of varying etiology.
Toxoplasma gondii (T. gondii) is a zoonotic infection that may be transmitted to human beings either by consumption of raw or uncooked meat or by ingesting oocysts. Toxoplasma organisms can cross blood placenta barrier and may result in congenital toxoplasmosis. About 80% of immunocompetent individuals do not show any clinical manifestations and are silent carriers of this disease. Pregnant women especially in highly prevalent areas are recommended to be screened for this disease in order to prevent the potential vertical transmission. To our knowledge no such study has been conducted in this region of Saudi Arabia. This study attempted to carry out two objectives: first, to find out the seroprevalence of T. gondii infection in pregnant women attending prenatal care services in our hospital; second, to find out risk factors associated with T. gondii seroprevalence in our patients. It was carried out in Teaching Hospital in Al-Kharj over a period of one year. All 306 pregnant women attending antenatal clinic were involved in the study. A pretested selfexplanatory questionnaire was filled out by the patients and their sera were collected to be tested for IgG and/or IgM against T. gondii. The results were then statistically analyzed using SPSS software and p-value was calculated using Pearson Chi Square test. Out of the 306 blood samples tested, 99 (32.4%) were seropositive for specific anti T. gondii IgG antibodies and 3(1%) were seropositive for IgM. This show that seroprevalence of T. gondii antibodies was high among pregnant women and the prevalence showed a significant association with age. The study recommends conducting educational programs to raise awareness among women about risk factors and precautions to be taken.
Toxoplasmosis is a parasitic zoonotic disease caused by Toxoplasma (T.) gondii. Limited data are available on the occurrence of T. gondii in women especially pregnant women in Pakistan. The present study aimed to determine the occurrence and risk factors associated with T. gondii in pregnant and non-pregnant women in Punjab Province, Pakistan. A cross-sectional study was conducted and 593 samples were collected from pregnant (n = 293) and non-pregnant (n = 300) women of District Headquarter Hospitals of Chiniot, Faisalabad, Jhang and Okara, Pakistan. Data related to demographic parameters and risk factors were collected using a pretested questionnaire on blood sampling day. Serum samples were screened for antibodies (IgG) against T. gondii using ELISA. A univariant and binomial logistic regression was applied to estimate the association between seropositive and explanatory variables considering the 95% confidence interval. P value ⩽0.05 was considered statistically significant for all analysis. Out of 593, 44 (7.42%) women were seropositive for T. gondii IgG antibodies. Occupation, age, sampling location, socioeconomic status, contact with cat, pregnancy status and trimester of pregnancy were significantly associated with seropositivity for T. gondii antibodies. Location and trimester of pregnancy were identified as potential risk factors for T. gondii seropositivity based on binomial logistic regression. Toxoplasma gondii is prevalent in pregnant and non-pregnant women. Therefore, now a necessitated awareness is required to instruct the individuals about these infectious diseases (toxoplasmosis) and their control strategies to maintain the health of human population. Moreover, health awareness among public can help the minimization of T. gondii infection during pregnancy and subsequent risk of congenital toxoplasmosis.
AbstractToxoplasma gondii infects a broad range of warm-blooded hosts, including humans. Important clinical manifestations include encephalitis in immunocompromised patients as well as miscarriage and fetal damage during early pregnancy. Toxoplasma gondii dense granule antigen 2 and 5 (GRA2 and GRA5) are essential for parasitophorous vacuole development of the parasite. To evaluate the potential of GRA2 and GRA5 as recombinant DNA vaccine candidates, these antigens were cloned into eukaryotic expression vector (pcDNA 3.1C) and evaluated in vaccination experiments. Recombinant DNA vaccines constructed with genes encoding GRAs were validated in Chinese hamster ovary cells before evaluation using lethal challenge of the virulent T. gondii RH strain in BALB/c mice. The DNA vaccines of pcGRA2 and pcGRA5 elicited cellular-mediated immune response with significantly higher levels of interferon-gamma, interleukin-2 (IL-2), IL-4, and IL-10 (P < 0.05) compared with controls. A mixed T-helper cell 1 (Th1)/Th2 response was associated with slightly prolonged survival. These findings provide evidence that DNA vaccination with GRA2 and GRA5 is associated with Th1-like cell-mediated immune responses. It will be worthwhile to construct recombinant multiantigen combining full-length GRA2 or/and GRA5 with various antigenic proteins such as the surface antigens and rhoptry antigens to improve vaccination efficacy.