As markers of exposure anti-malaria antibody responses can help characterise heterogeneity in malaria transmission. In the present study antibody responses to Plasmodium falciparum AMA-1, MSP-119 and CSP were measured with the aim to describe transmission patterns in meso-endemic settings in Lake Victoria. Two cross-sectional surveys were conducted in Lake Victoria in January and August 2012. The study area comprised of three settings: mainland (Ungoye), large island (Mfangano) and small islands (Takawiri, Kibuogi, Ngodhe). Individuals provided a finger-blood sample to assess malaria infection by microscopy and PCR. Antibody response to P. falciparum was determined in 4,112 individuals by ELISA using eluted dried blood from filter paper. The overall seroprevalence was 64.0% for AMA-1, 39.5% for MSP-119, and 12.9% for CSP. Between settings, seroprevalences for merozoite antigens were similar between Ungoye and Mfangano, but higher when compared to the small islands. For AMA-1, the seroconversion rates (SCRs) ranged from 0.121 (Ngodhe) to 0.202 (Ungoye), and were strongly correlated to parasite prevalence. We observed heterogeneity in serological indices across study sites in Lake Victoria. These data suggest that AMA-1 and MSP-119 sero-epidemiological analysis may provide further evidence in assessing variation in malaria exposure and evaluating malaria control efforts in high endemic area.
Toxoplasma gondii primary infection in pregnancy is associated with poor obstetric outcomes. This study aimed to determine the seroprevalence of Toxoplasma infection in pregnant migrant and refugee women from Myanmar attending antenatal care in Thailand. A random selection of 199 residual blood samples from first antenatal screen in 2014-2015 was tested for Toxoplasma IgG and IgM antibodies. Seroprevalence of Toxoplasma infection was 31.7% (95% confidence interval = 25.6-38.4). Avidity testing in the three positive IgM cases indicated all were past infections. Multiparity (≥ 3 children) was significantly associated with higher Toxoplasma seropositivity rates. Seroprevalence of T. gondii infection in this pregnant population is similar to the only other report from Myanmar, where multiparity was also identified as a significant association. Toxoplasma infection is important in pregnant women. Nevertheless, in this marginalized population, this infection may be given less priority, due to resource constraints in providing the most basic components of safe motherhood programs.
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.
Most studies on human immunity to malaria have focused on the roles of immunoglobulin G (IgG), whereas the roles of IgM remain undefined. Analyzing multiple human cohorts to assess the dynamics of malaria-specific IgM during experimentally induced and naturally acquired malaria, we identified IgM activity against blood-stage parasites. We found that merozoite-specific IgM appears rapidly in Plasmodium falciparum infection and is prominent during malaria in children and adults with lifetime exposure, together with IgG. Unexpectedly, IgM persisted for extended periods of time; we found no difference in decay of merozoite-specific IgM over time compared to that of IgG. IgM blocked merozoite invasion of red blood cells in a complement-dependent manner. IgM was also associated with significantly reduced risk of clinical malaria in a longitudinal cohort of children. These findings suggest that merozoite-specific IgM is an important functional and long-lived antibody response targeting blood-stage malaria parasites that contributes to malaria immunity.
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.
Plasmodium vivax is the most widespread and difficult to treat cause of human malaria. The development of vaccines against the blood stages of P. vivax remains a key objective for the control and elimination of vivax malaria. Erythrocyte binding-like (EBL) protein family members such as Duffy binding protein (PvDBP) are of critical importance to erythrocyte invasion and have been the major target for vivax malaria vaccine development. In this study, we focus on another member of EBL protein family, P. vivax erythrocyte binding protein (PvEBP). PvEBP was first identified in Cambodian (C127) field isolates and has subsequently been showed its preferences for binding reticulocytes which is directly inhibited by antibodies. We analysed PvEBP sequence from 316 vivax clinical isolates from eight countries including China (n = 4), Ethiopia (n = 24), Malaysia (n = 53), Myanmar (n = 10), Papua New Guinea (n = 16), Republic of Korea (n = 10), Thailand (n = 174), and Vietnam (n = 25). PvEBP gene exhibited four different phenotypic clusters based on the insertion/deletion (indels) variation. PvEBP-RII (179-479 aa.) showed highest polymorphism similar to other EBL family proteins in various Plasmodium species. Whereas even though PvEBP-RIII-V (480-690 aa.) was the most conserved domain, that showed strong neutral selection pressure for gene purifying with significant population expansion. Antigenicity of both of PvEBP-RII (16.1%) and PvEBP-RIII-V (21.5%) domains were comparatively lower than other P. vivax antigen which expected antigens associated with merozoite invasion. Total IgG recognition level of PvEBP-RII was stronger than PvEBP-RIII-V domain, whereas total IgG inducing level was stronger in PvEBP-RIII-V domain. These results suggest that PvEBP-RII is mainly recognized by natural IgG for innate protection, whereas PvEBP-RIII-V stimulates IgG production activity by B-cell for acquired immunity. Overall, the low antigenicity of both regions in patients with vivax malaria likely reflects genetic polymorphism for strong positive selection in PvEBP-RII and purifying selection in PvEBP-RIII-V domain. These observations pose challenging questions to the selection of EBP and point out the importance of immune pressure and polymorphism required for inclusion of PvEBP as a vaccine candidate.
Three hundred and one sera of HIV/AIDS patients were tested for anti-Toxoplasma IgG antibody by ELISA technique. The seroprevalence of toxoplasmosis was 41.2% (95% CI: 35.5-46.9) in HIV/AIDS patients. The seroprevalence was significantly higher in the Malay (57.9%) than the Chinese (38.7%), followed by the Indian patients (29.6%) (p<0.05). No possible risk factor, such as contact with cats, consumption of uncooked meat, and history of blood transfusions was found to have any significant association with the presence of anti-Toxoplasma antibody in the study sample (p>0.05). Multivariate analysis was employed to find any association between Toxoplasma seroprevalence and a single subject having single or multiple risk factors. It was found that the association was not statistically significant (p>0.05). Among the HIV/AIDS study samples, 124 (41.2%) samples were found to have positive anti-Toxoplasma antibody, the association between the presence of anti-Toxoplasma antibody and CD4 cell count was determined but no statistically significant association was found (p>0.05). During the study period, only one case of active CNS toxoplasmosis was registered and the diagnostic criteria included: clinical presentations, CT scan finding, serological evidence of anti-Toxoplasma IgG antibody, and respose to anti-Toxoplasma therapy.
Cattle in Peninsular Malaysia were examined for evidence of infection with Babesia ovata, B. bigemina and B. bovis by an enzyme-linked immunosorbent assay for detection of antibody to the three Babesia species. All of the test samples when assayed with B. ovata antigen, resulted in low value indicating low probability of cattle infected with B. bigemina, 74.4% were positive for B. bovis and 72.6% were positive for both Babesia species. In addition, a serological survey with regard to age difference was carried out on a milk production farm. High reactivity antibody to B. bigemina and B. bovis was detected in calves less than 1 month of the age. The reactivity decreased in calves 1-3 months of the age. Then, the reactivity increased for both Babesia species in 6 months old calves. These results suggested that cattle infected with B. bigemina and B. bovis were widespread throughout Peninsular Malaysia and that both parasites might exist as an enzootical parasite.
Indirect fluorescent antibody (IFA) tests and enzyme-linked immunosorbent assays (ELISA) were used to measure antibodies to Plasmodium falciparum in an indigenous population in an area of Malaysia with high malaria prevalence. The results of three surveys were analyzed to examine the relation of these serologic measures with age, parasite rate, and spleen size. For children 0-4 years old, increasing spleen size was associated with an increasing likelihood of malaria parasitemia, while for 5-9 year olds the two variables were unrelated. Parasite rate declined with age and ELISA titre increased with age in all surveys; IFA titre was consistently high and did not vary with age. Neither antibody measure was significantly correlated with either the presence or the actual density of parasitemia. These antibody measures are most useful as adjuncts to the more traditional techniques of malaria assessment.
The IgG avidity assay is an important tool in the management of suspected toxoplasmosis in pregnant women. This study aimed to produce new Toxoplasma gondii recombinant proteins and to assess their usefulness in an IgG avidity assay. Toxoplasma positive and negative serum samples were used, the former were categorized into low (LGA) and high (HGA) IgG avidity samples. Immunoblots were performed on 30 T. gondii cDNA clones to determine the reactivity and IgG avidity to the expressed proteins. Two of the clones were found to have diagnostic potential and were analyzed further; AG12b encoded T. gondii apical complex lysine methyltransferase (AKMT) protein and AG18 encoded T. gondii forkhead-associated (FHA) domain-containing protein. The His-tagged recombinant proteins, rAG12b and rAG18, were expressed and tested with LGA and HGA samples using an IgG avidity western blot and ELISA. With the IgG avidity western blot, rAG12b identified 86.4% of LGA and 90.9% of HGA samples, whereas rAG18 identified 81.8% of both LGA and HGA samples. With the IgG avidity ELISA, rAG12b identified 86.4% of both LGA and HGA samples, whereas rAG18 identified 77.3% of LGA and 86.4% of HGA serum samples. This study showed that the recombinant antigens were able to differentiate low avidity and high avidity serum samples, suggesting that they are potential candidates for use in the Toxoplasma IgG avidity assay.
Crude soluble antigen (CSA) produced from Entamoeba histolytica trophozoite is conventionally used for serodiagnosis of invasive amoebiasis. However, high background seropositivities by CSA-assay in endemic areas complicate the interpretation of positive result in clinical settings. Instead, incorporating a second assay which indicates active or recent infection into the routine amoebic serology could possibly complement the limitations of CSA-assay. Hence, the present study aimed to evaluate the diagnostic efficacies of indirect ELISAs using CSA and excretory-secretory antigen (ESA) for serodiagnosis of amoebic liver abscess (ALA). Reference standard for diagnosis of ALA at Hospital Universiti Sains Malaysia is based on clinical presentation, radiological imaging and positive indirect haemagglutination assay (titer ≥256). Five groups of human serum samples collected from the hospital included Group I - ALA diagnosed by the reference standard and pus aspirate analysis using real-time PCR (n=10), Group II - ALA diagnosed by the reference standard only (n=41), Group III - healthy control (n=45), Group IV - other diseases control (n=51) and Group V - other infectious diseases control (n=31). For serodiagnosis of ALA serum samples (Group I and II), CSA-ELISA showed sensitivities of 100% for both groups, while ESA-ELISA showed sensitivities of 100% and 88%, respectively. For serodiagnosis of non-ALA serum samples (Group III, IV and V), CSA-ELISA showed specificities of 91%, 75% and 100%, respectively; while ESA-ELISA showed specificities of 96%, 98% and 100%, respectively. Indirect ELISAs using CSA and ESA have shown distinct strength for serodiagnosis of ALA, in terms of sensitivity and specificity, respectively. In conclusion, parallel analysis by both assays improved the overall efficacies of amoebic serology as compared to either single assay.
It was reported that a 47kDa antigenic polypeptide of Plasmodium falciparum had been strongly presented by the sera from 1) imported Japanese malaria patients with severe symptoms and 2) symptomatic and parasitemic inhabitants in endemic areas in the Sudan, Malaysia and the Philippines. In the present study, we observed the reactivity of the sera from falciparum malaria patients who had been hospitalized in the Bangkok Hospital for Tropical Diseases, Faculty of Tropical Medicine, Mahidol University, and compared the antibody response against the 47kDa antigenic polypeptide according to the severity of the patients. It was observed that antibodies to this molecule were more commonly shared in sera from severer patients, although the IFAT titers against the whole P. falciparum parasite antigen were lower in the group, which suggested that this antibody against the 47kDa molecule was playing a specific role at a severe stage of the infection. Determination of the immunological features of the antigenic molecules of parasites by this type of sero-epidemiological study will provide a new assay system for evaluation of immune status of individuals in different severity and suggest a way of vaccine development.
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.
Toxoplasma gondii infects all warm-blooded animals, including humans. Early diagnosis and determining the infective stage are critical for effectively treating immunosuppressed individuals and pregnant women with toxoplasmosis. Among the rhoptry proteins of the parasite, Rhoptry protein 8 (ROP8), is known to be expressed during the early stages of T. gondii infection and is involved in parasitophorous vacuole formation. In this study, we have investigated the diagnostic efficacy of recombinant ROP8 (rROP8).
Toxoplasmosis, being one of the TORCH's infections in pregnant women, is caused by Toxoplasma gondii, an obligate intracellular protozoan parasite. This parasitic infection in pregnancy congenitally causes severe outcomes to their fetus and newborn. This study aimed to determine the seroprevalence and stages of Toxoplasma infection in pregnant women and its associated risks exposures.
Toxoplasmosis is an infection caused by the parasite Toxoplasma gondii. Chronically-infected individuals with a compromised immune system are at risk for reactivation of the disease. In-vivo induced antigen technology (IVIAT) is a promising method for the identification of antigens expressed in-vivo. The aim of the present study was to apply IVIAT to identify antigens which are expressed in-vivo during T. gondii infection using sera from individuals with chronic toxoplasmosis. Forty serum samples were pooled, pre-adsorped against three different preparations of antigens, from each in-vitro grown T. gondii and Escherichia coli XLBlue MRF', and then used to screen a T. gondii cDNA expression library. Sequencing of DNA inserts from positive clones showed eight open reading frames with high homology to T. gondii genes. Expression analysis using quantitative real-time PCR showed that SAG1-related sequence 3 (SRS3) and two hypothetical genes were up-regulated in-vivo relative to their expression levels in-vitro. These three proteins also showed high sensitivity and specificity when tested with individual serum samples. Five other proteins namely M16 domain peptidase, microneme protein, elongation factor 1-alpha, pre-mRNA-splicing factor and small nuclear ribonucleoprotein F had lower RNA expression in-vivo as compared to in-vitro. SRS3 and the two hypothetical proteins warrant further investigation into their roles in the pathogenesis of toxoplasmosis.
Plasmodium knowlesi is now known as the fifth Plasmodium species that can cause human malaria. The Plasmodium merozoite surface protein (MSP) has been reported to be potential target for vaccination and diagnosis of malaria. MSP-1(33) has been shown to be immunogenic and its T cell epitopes could mediate cellular immune protection. However, limited studies have focused on P. knowlesi MSP-133. In this study, an approximately 28-kDa recombinant P. knowlesi MSP-1(33) (pkMSP-1(33)) was expressed by using an Escherichia coli system. The purified pkMSP-1(33) reacted with serum samples of patients infected with P. knowlesi (31 of 31, 100%) and non-P. knowlesi malaria (27 of 28, 96.43%) by Western blotting. The pkMSP-1(33) also reacted with P. knowlesi (25 of 31, 80.65%) and non-P. knowlesi malaria sera (20 of 28, 71.43%) in an enzyme-linked immunosorbent assay (ELISA). Most of the non-malarial infection (49 of 52 in by Western blotting and 46 of 52 in the ELISA) and healthy donor serum samples (65 of 65 by Western blotting and ELISA) did not react with recombinant pkMSP-1(33).