Toxoplasma gondii is a parasite that generates latent cysts in the brain; reactivation of these cysts may lead to fatal toxoplasmic encephalitis, for which treatment remains unsuccessful. We assessed spiramycin pharmacokinetics coadministered with metronidazole, the eradication of brain cysts and the in vitro reactivation. Male BALB/c mice were fed 1,000 tachyzoites orally to develop chronic toxoplasmosis. Four weeks later, infected mice underwent different treatments: (i) infected untreated mice (n = 9), which received vehicle only; (ii) a spiramycin-only group (n = 9), 400 mg/kg daily for 7 days; (iii) a metronidazole-only group (n = 9), 500 mg/kg daily for 7 days; and (iv) a combination group (n = 9), which received both spiramycin (400 mg/kg) and metronidazole (500 mg/kg) daily for 7 days. An uninfected control group (n = 10) was administered vehicle only. After treatment, the brain cysts were counted, brain homogenates were cultured in confluent Vero cells, and cysts and tachyzoites were counted after 1 week. Separately, pharmacokinetic profiles (plasma and brain) were assessed after a single dose of spiramycin (400 mg/kg), metronidazole (500 mg/kg), or both. Metronidazole treatment increased the brain spiramycin area under the concentration-time curve from 0 h to ∞ (AUC(0-∞)) by 67% without affecting its plasma disposition. Metronidazole plasma and brain AUC(0-∞) values were reduced 9 and 62%, respectively, after spiramycin coadministration. Enhanced spiramycin brain exposure after coadministration reduced brain cysts 15-fold (79 ± 23 for the combination treatment versus 1,198 ± 153 for the untreated control group [P < 0.05]) and 10-fold versus the spiramycin-only group (768 ± 125). Metronidazole alone showed no effect (1,028 ± 149). Tachyzoites were absent in the brain. Spiramycin reduced in vitro reactivation. Metronidazole increased spiramycin brain penetration, causing a significant reduction of T. gondii brain cysts, with potential clinical translatability for chronic toxoplasmosis treatment.
Molecular methods are used increasingly for the detection of Toxoplasma gondii infection. This study developed a rapid, sensitive, and specific conventional triplex PCR for the detection of the B1 gene and ITS1 region of T. gondii using newly designed primers and an internal control based on the Vibrio cholerae HemM gene. The annealing temperature and concentrations of the primers, MgCl(2), and dNTPs were optimized. Two sets of primers (set 1 and 2) were tested, which contained different segments of the T. gondii B1 gene, 529 repeat region and ITS1 region. A series of sensitivity tests were performed using parasite DNA, whole parasites, and spiked human body fluids. Specificity tests were performed using DNA from common protozoa and bacteria. The newly developed assay based on set 2 primers was found to be specific and sensitive. The test was capable of detecting as little as 10 pg T. gondii DNA, 10(4) tachyzoites in spiked body fluids, and T. gondii DNA in the organ tissues of experimentally infected mice. The assay developed in this study will be useful for the laboratory detection of T. gondii infection.
A total of 1,885 blood and stool samples of four main protozoan parasitic infections were retrospectively reviewed from January, 2000 to April, 2004. Eleven of the 1,350 stool samples were shown positive for Cryptosporidium and Giardia infections; one of the 5 cases was clinically diagnosed as gastrointestinal cryptosporidiosis, while 6 cases were giardiasis. In patients with giardiasis, children were among the high-risk groups, making up 66.7% of these patients. The common presenting signs and symptoms were: diarrhea (83.3%), loss of appetite (83.3%), lethargy (83.3%), fever (66.7%), nausea/vomiting (50.0%), abdominal pain (16.7%), dehydration (16.7%) and rigor and chills (16.7%). Metronidazole was the drug of choice and was given to all symptomatic patients (83.3%). For the blood samples, 28 of the 92 peripheral smears for Plasmodium spp infection were diagnosed as malaria. The age range was from 4 to 57, with a median of 32.5 years. The sex ratio (M:F) was 3.6:1, while the age group of 30-44 years was the most commonly affected in both sexes. The majority of patients were foreigners (60.7%) and non-professional (39%). Plasmodium vivax (71%) infection was the most common pathogen found in these patients, along with a history of traveling to an endemic area of malaria (31%). The predominant presenting signs and symptoms were: fever (27%), rigor and chills (24%), nausea/vomiting (15%) and headache (8%). Chloroquine and primaquine was the most common anti-malarial regimen used (78.6%) in these patients. The seroprevalence of toxoplasmosis in different groups was 258/443 (58%): seropositive for IgG 143 (32.3%); IgM 67 (15%); and IgG + IgM 48 (10.8%). The age range was from 1 to 85, with a mean of 34 (+/- SD 16.6) years. The predominant age group was 21 to 40 years (126; 28.4%). The sex ratio (M:F) was 1.2:1. Subjects were predominantly male (142; 32%) and the Malay (117; 26.4%). Of these, 32 cases were clinically diagnosed with ocular toxoplasmosis. The range of age was from 10 to 56 years with a mean of 30.5 (+/- SD 12.05) years. The sex ratio (M:F) was 1:1.7. The majority were in the age group of 21 to 40 years, female (20; 62.5%), and Malay (17; 53%). They were also single (16; 50%), unemployed (12; 37%), and resided outside Kuala Lumpur (21; 65.6%). The more common clinical presentations were blurring of vision (25; 78%), floaters (10; 31%) and pain in the eye (7; 22%). We found that funduscopic examination (100%) and seropositivity for anti-Toxoplasma antibodies (93.7%) were the main reasons for investigation. Choroidoretinitis was the most common clinical diagnosis (69%), while clindamycin was the most frequently used antimicrobial in all cases. Among HIV-infected patients, 10 cases were diagnosed as AIDS-related toxoplasmic encephalitis (TE) (9 were active and 1 had relapse TE). In addition, 1 case was confirmed as congenital toxoplasmosis.
The neglected tropical diseases, echinococcosis, schistosomiasis and toxoplasmosis are all globally widespread zoonotic diseases with potentially harmful consequences. There is very limited data available on the prevalence of these infections, except for schistosmiasis, in underdeveloped countries. This study aimed to determine the seroprevalence of Echinococcus multilocularis, Schistosoma mansoni, and Toxoplasma gondii antibodies in populations from the Monduli and Babati districts in Tanzania.
Toxoplasma gondii is an important pathogen in veterinary and human medicine. In this study, a new multiplex TaqMan real-time PCR for detection of T. gondii DNA was developed. This assay consisted of new sets of primers and probes which targeted B1 gene and ITS-1 region of T. gondii, with Vibrio cholera gene as internal control. The B1 gene primers were designed to detect T. gondii RH strain, while the ITS-1 region primers detected most T. gondii strains. Specificity test using common protozoal and bacterial DNA revealed that the assay was very specific to T. gondii. Standard curves constructed using human body fluids spiked with T. gondii (RH and ME49 strains) showed that the sensitivity of the assay was one parasite, with R² value of 0.975 to 0.999 and efficiency of 97% to 99% for all types of samples. The assay performed on DNA extracted from tissues of mice infected with T. gondii showed that liver contained the highest parasite load for both strains of T. gondii. The multiplex real-time PCR developed in this study would be potentially useful for detection of T. gondii in human and animal samples.
We describe the results of serology for parasitic infection of 250 foreign workers who were seen at the University of Malaya Medical Centre, UMMC during 7-months period. The 250 foreign workers participated included 114 from Indonesia, 142 from Bangladesh, two from Myanmar and two from Pakistan. Blood samples were taken from these workers and eight tests (amoebiasis, echinococcosis, filariasis, leishmaniasis, malaria, schistosomiasis, toxoplasmosis, and trypanosomiasis) were performed on serum. Among the 250 sera tested, 92 (36.8%) were found to be positive for at least one parasitic infection. There was one case where the serum was found positive for 5 tests. The most common antibody detected in those positive sera was antibody for toxoplasma (80.%), followed by filaria (32.8%) and amoeba (30%). Other tests showed low percentage of infection with schistosomiasias, 10%; echinococcosis, 6% and malaria, 3.6%. None of the foreign workers were found positive for leishmaniasis or trypanosomiasias.
Infection by Toxoplasma gondii is prevalent worldwide. The parasite can infect a broad spectrum of vertebrate hosts, but infection of fetuses and immunocompromised patients is of particular concern. Easy-to-perform, robust, and highly sensitive and specific methods to detect Toxoplasma infection are important for the treatment and management of patients. Rapid diagnostic methods that do not sacrifice the accuracy of the assay and give reproducible results in a short time are highly desirable. In this context, rapid diagnostic tests (RDTs), especially with point-of-care (POC) features, are promising diagnostic methods in clinical microbiology laboratories, especially in areas with minimal laboratory facilities. More advanced methods using microfluidics and sensor technology will be the future trend. In this review, we discuss serological and molecular-based rapid diagnostic tests for detecting Toxoplasma infection in humans as well as animals.
Toxoplasmosis, caused by an intracellular protozoan parasite, Toxoplasma gondii, is widespread throughout the world. The disease is of major medical and veterinary importance, being a cause of congenital disease and abortion in humans and in domestic animals.[1] In addition, it has gained importance recently due to toxoplasma encephalitis in AIDS patients.[2] T. gondii was discovered 100 years ago. Its identification was rapidly followed by the recognition that it was a human pathogen. During the past 100 years, the spectrum of diseases caused by this ubiquitous pathogen has expanded to include both congenital and acute infections as well as the recognition of diseases caused by this pathogen in the immune-compromised host. Recent data on behavioural changes in animals due to chronic toxoplasmosis is leading to research on the effect of this pathogen on the behaviour of humans.[3] Experimental studies on T. gondii have resulted in it becoming a model organism for studies on host pathogen interactions. Integration of clinical and experimental data on T. gondii should continue to lead to important insights into improvements in diagnosis for clinical management and vaccine development for control of toxoplasmosis.
Replacement therapy for toxoplasmosis was not a clear-cut choice since most of anti-parasitic agents available are also associated with Steven Johnson Syndrome. Further more the therapy has to be effective to control infection, which was previously achieved by oral Fansidar in this patient. Oral Azithromycin was seen as a drug of choice for these reasons. Corticosteroids were maintained since it was relatively indicated in both toxoplasmosis and SJS. Both conditions can results in visual impairment. SJS can be a life threathening condition and its ocular complications include conjunctivitis, ectropion or entropion, symblepharon, vascularization of the cornea, chronic dry eyes, and ankylosymblepharon. Proper management in dealing with both diseases is mandatory in order to prevent mortality and minimize the ocular complications. It has been shown in this case that the challenging part in managing patient with both diseases is to balance out between prevention of fatal consequences and the need control to the infection and preserving vision. Decisions on medical treatment for both conditions will remain controversial till reliable prospective randomized control trials are done to address the issues
During pregnancy, Toxoplasma gondii can be transmitted from mother to foetus and trigger a primary infection that may be symptomatic. It is important to distinguish between recently acquired and past infections to ensure proper treatment to minimize irreversible foetal injury. We used PCR of the B1 gene to evaluate the accuracy of T. gondii IgG antibody avidity testing in discriminating recent from past infection. In a cross-sectional study, T. gondii IgG and IgM antibodies were detected by enzyme linked fluorescence assay (ELFA) in 2120 serum samples from pregnant women referred to Karaj medical laboratories, February 2013 through March 2015 with 40 samples found positive. IgM-positive samples were evaluated by IgG avidity testing and PCR to amplify the B1 gene. Avidity studies indicated 20 samples with high IgG avidity, 15 with low IgG avidity, and five showing borderline values. The B1 gene was amplified in the borderline samples, with nine of the 15 showing low avidity. The B1 gene was not amplified in the high avidity sera. Our findings suggest that IgG avidity alone may not be sufficient to discriminate recent from past T. gondii infection and should not be used as the sole confirmatory test in pregnant women with IgG and IgM T. gondii antibodies. IgG avidity testing in combination with PCR may be more reliable for distinguishing between high- and low-risk infection and decrease the frequency of unnecessary treatment of pregnant women.
Toxoplasma gondii, a zoonotic protozoan that has a worldwide distribution, is known to infect many warm-blooded vertebrates. The feline species including domestic cats are the definitive hosts for Toxoplama gondii and shed the infective oocyst. There is lack of information on the prevalence of Toxoplasma gondii in cats in Malaysia. The objective of this study was to determine both the seroprevalence of T. gondii and the prevalence of T. gondii DNA in cats' feces in Klang Valley, Malaysia. 198 blood and 201 fecal samples were collected from pet and stray cats from the local council, Dewan Bandaraya Kuala Lumpur (DBKL) and University Veterinary Hospital, Universiti Putra Malaysia respectively. The overall seroprevalence of Toxoplasma gondii in cats in the Klang Valley was found to be 5.5%. There was a high prevalence (10.5%) of T. gondii DNA detected in the cat fecal samples in both pet and stray cats suggestive of T. gondii oocyst shedding. Stray cats showed a higher seroprevalence and molecular prevalence of T. gondii than the pet cats. However, comparative analysis using Chi-square test showed no significant difference between both groups (P>0.05). Higher prevalence (10.5%) of cats shedding T. gondii DNA as compared to the seroprevalence (5.5%) was found in the cat population in the Klang Valley. The high prevalence of cats shedding T. gondii DNA is alarming as this may directly reflect the number of oocysts excreted into the environment posing a significant public health hazard.
Toxoplasma gondii is a public health risk in developing countries, especially those located in the tropics. Widespread infection may inflict a substantial burden on state resources, as patients can develop severe neurological defects and ocular diseases that result in lifelong loss of economic independence. We tested sera for IgG antibody from 493 eye patients in Malaysia. Overall age-adjusted seroprevalence was estimated to be 25% (95% CI: [21%, 29%]). We found approximately equal age-adjusted seroprevalence in Chinese (31%; 95% CI: [25%, 38%]) and Malays (29%; 95% CI: [21%, 36%]), followed by Indians (19%; 95% CI: [13%, 25%]). A logistic regression of the odds for T. gondii seroprevalence against age, gender, ethnicity and the occurrence of six types of ocular diseases showed that only age and ethnicity were significant predictors. The odds for T. gondii seroprevalence were 2.7 (95% CI for OR: [1.9, 4.0]) times higher for a patient twice as old as the other, with ethnicity held constant. In Malays, we estimated the odds for T. gondii seroprevalence to be 2.9 (95% CI for OR: [1.8, 4.5]) times higher compared to non-Malays, with age held constant. Previous studies of T. gondii seroprevalence in Malaysia did not explicitly adjust for age, rendering comparisons difficult. Our study highlights the need to adopt a more rigorous epidemiological approach in monitoring T. gondii seroprevalence in Malaysia.
Toxoplasma gondii is an obligate intracellular zoonotic parasite of the phylum Apicomplexa which infects a wide range of warm-blooded animals, including humans. In this study in-vivo induced antigens of this parasite was investigated using in-vivo induced antigen technology (IVIAT) and pooled sera from patients with serological evidence of acute infection.
Toxoplasmosis is one of the most common opportunistic parasitic diseases in patients living with HIV/AIDS. This study aimed to determine the seroprevalence of Toxoplasma infection in HIV-infected patients and to identify associated risk factors in Toxoplasma seropositive patients. This study was conducted at a regional public hospital in Hat Yai, southern Thailand during October 2009 to June 2010. Blood samples were collected from 300 HIV-infected patients. Each subject also answered a socio-demographic and risk factors associated with Toxoplasma infection. The prevalence of anti-Toxoplasma IgG antibodies in HIV-infected patients was 109 (36.3%), of which 83 (76.2%) had past infection and 26 (23.9%) had recently acquired Toxoplasma infection as indicated by their IgG avidity. Multivariate analysis using logistic regression showed that gender difference (adjusted OR = 1.69, 95% CI = 1.05-2.72) was the only factor associated with Toxoplasma infection. From the results obtained, these HIV-infected patients could be at high risk of developing clinical evidence of severe toxoplasmosis. Therefore, it is necessary to introduce primary behavioral practices to prevent Toxoplasma infection among HIV-infected patients.
Toxoplasmosis, a parasitic disease in human and animals, is caused by Toxoplasma gondii. Our previous study has led to the discovery of a novel RAP domain binding protein antigen (TgRA15), an apparent in-vivo induced antigen recognised by antibodies in acutely infected individuals. This study is aimed to evaluate the humoral response and cytokine release elicited by recombinant TgRA15 protein in C57BL/6 mice, demonstrating its potential as a candidate vaccine for Toxoplasma gondii infection. In this study, the recombinant TgRA15 protein was expressed in Escherichia coli, purified and refolded into soluble form. C57BL/6 mice were immunised intradermally with the antigen and CASAC (Combined Adjuvant for Synergistic Activation of Cellular immunity). Antigen-specific humoral and cell-mediated responses were evaluated using Western blot and ELISA. The total IgG, IgG1 and IgG2a antibodies specific to the antigen were significantly increased in treatment group compare to control group. A higher level of interferon gamma (IFN-γ) secretion was demonstrated in the mice group receiving booster doses of rTgRA15 protein, suggesting a potential Th1-mediated response. In conclusion, the rTgRA15 protein has the potential to generate specific antibody response and elicit cellular response, thus potentially serve as a vaccine candidate against T. gondii infection.
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.
Toxoplasma gondii is a protozoan parasite that is capable of causing a zoonotic disease, known as toxoplasmosis. Vertical transmission of T. gondii from the mother to the fetus, during pregnancy may cause severe complications to the developing fetus. This current study aimed to determine the seroprevalence and investigate the associated risk factors of Toxoplasma infection in pregnant women (n=219) visiting the antenatal clinic at UMMC. While the elevated level of anti-Toxoplasma IgG and IgM antibodies indicates the presence of infection, it fails to differentiate between a past and a recent infection. Thus, the study also demonstrates the usefulness of IgG avidity in validating the timing of infection. The serum samples were tested for the presence of anti-Toxoplasma IgG and IgM antibodies by ELISA test, and the seropositive samples for both anti-Toxoplasma IgG and IgM antibodies were further evaluated by IgG avidity. The results showed that the overall prevalence of T. gondii seropositivity was 34.7%. Of these, 30.6% (67/219) were positive for anti-Toxoplasma IgG antibody only, 2.3% (5/219) were positive for anti-Toxoplasma IgM only, and the remaining 1.8% (4/219) was positive for both anti-Toxoplasma IgG and IgM antibodies. All of the pregnant women who were positive for both anti-Toxoplasma IgG and IgM antibody were found to have past infection when evaluated by IgG avidity. In this study, Malay ethnicity and the number of existing previous children were significantly associated with T. gondii seropositivity (p<0.05). Based on these findings, information and education on the transmission and prevention of congenital toxoplasmosis are very crucial as a public health effort towards a healthier society.
Schizophrenia is a pervasive neuropsychiatric disease of unknown cause. Previous studies have reported that toxoplasmosis may be a possible cause of schizophrenia. To ascertain possible relationship between Toxoplasma gondii and schizophrenia, a cross sectional study, employing an enzyme-linked immunosorbent assay (ELISA) was performed to study the seroprevalence of anti-T. gondii IgG antibody in schizophrenic patients. Furthermore, demographic data analysis from schizophrenic patients were analysed to associate toxoplasmosis with schizophrenia. A total of 288 serum samples from schizophrenic patients (n=144) and psychiatrically healthy volunteers (n=144) were recruited in this study. Interestingly, a significant result in the serointensity rate of anti-T. gondii IgG antibody (> 60 IU/mL) in schizophrenic patients (61.1%) was demonstrated as compared to psychiatrically healthy volunteers (40.8%) (X² = 4.236, p < 0.050). However, there was no significant difference between the seropositivity rate of anti-T. gondii IgG antibody between the two groups. Analysis from demographic data revealed that the seropositivity rate of anti-T. gondii IgG antibody in schizophrenic patients was significantly associated with age group of more than 40 years old (p=0.007) and between ethnic (p=0.046). Nevertheless, no significant association between seropositivity rate of anti-T. gondii IgG antibody with gender (p=0.897), duration of illness (p=0.344) and family history of schizophrenia (p=0.282) in these patients. Thus, this finding is essential as a preliminary data in Malaysia to establish the association between T. gondii and schizophrenia.
In this study we have cloned unreported gene fragments of Toxoplasma gondii GRA7 and SAG1 and expressed the corresponding recombinant proteins, followed by evaluation of their usefulness for the serological diagnosis of toxoplasmosis. Both recombinant proteins were expressed efficiently in insoluble form, purified by single step Ni-NTA affinity chromatography and their antigenicity to detect toxoplasma specific IgG antibodies were determined by immunoblotting. A total of 60 serum samples from three groups of individuals based on their anti-toxoplasma antibody profiles were tested, namely (I) IgM+, IgG+ (n=20), (II) IgM-, IgG+ (n=20) and (III) IgM-, IgG- (n=20). Both recombinant proteins exhibited high sensitivity (100%) with sera from Group I. rGRA7 and rSAG1 reacted 40% and 80% respectively with Group II sera. The specificity of the recombinant proteins based on reactivities with Group III sera were 100% and 80% with rGRA7 and rSAG1 respectively. Thus rGRA7 was found to be better at discriminating probable acute from chronic phases of toxoplasmosis, and it also showed higher specificity.