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A New Plant Expression System for Producing Pharmaceutical Proteins

Abd-Aziz N, Tan BC, Rejab NA, Othman RY, Khalid N

Mol Biotechnol, 2020 Apr;62(4):240-251.
PMID: 32108286 DOI: 10.1007/s12033-020-00242-2

In the past decade, interest in the production of recombinant pharmaceutical proteins in plants has tremendously progressed because plants do not harbor mammalian viruses, are economically competitive, easily scalable, and capable of carrying out complex post-translational modifications required for recombinant pharmaceutical proteins. Mucuna bracteata is an essential perennial cover crop species widely planted as an underground cover in oil palm and rubber plantations. As a legume, they have high biomass, thrive in its habitat, and can fix nitrogen. Thus, M. bracteata is a cost-efficient crop that shows ideal characteristics as a platform for mass production of recombinant protein. In this study, we established a new platform for the transient production of a recombinant protein in M. bracteata via vacuum-assisted agro-infiltration. Five-week-old M. bracteata plants were vacuum infiltrated with Agrobacterium tumefaciens harboring a plasmid that encodes for an anti-toxoplasma immunoglobulin (IgG) under different parameters, including trifoliate leaf positional effects, days to harvest post-infiltration, and the Agrobacterium strain used. Our results showed that vacuum infiltration of M. bracteata plant with A. tumefaciens strain GV3101 produced the highest concentration of heterologous protein in its bottom trifoliate leaf at 2 days post-infiltration. The purified anti-toxoplasma IgG was then analyzed using Western blot and ELISA. It was demonstrated that, while structural heterogeneity existed in the purified anti-toxoplasma IgG from M. bracteata, its transient expression level was two-fold higher than the model platform, Nicotiana benthamiana. This study has laid the foundation towards establishing M. bracteata as a potential platform for the production of recombinant pharmaceutical protein.

Matched MeSH terms: Toxoplasma/immunology
Molecular investigation on the occurrence of Toxoplasma gondii oocysts in cat feces using TOX-element and ITS-1 region targets

Chemoh W, Sawangjaroen N, Nissapatorn V, Sermwittayawong N

Vet J, 2016 Sep;215:118-22.
PMID: 27325616 DOI: 10.1016/j.tvjl.2016.05.018

One of the most important routes of transmission for Toxoplasma gondii infection is the ingestion of foods contaminated with cat feces containing sporulated oocysts. The diagnosis of T. gondii infection by fecal microscopy is complicated, as other similar coccidian oocysts are often present in the same fecal specimen. This study aimed to identify T. gondii oocysts in cat feces using a novel PCR technique. Feline fecal specimens (n = 254) were screened for coccidian oocysts by light microscopy using the Sheather's flotation method. PCR analysis performed on the same specimens targeted a 529 bp repeat element and internal transcribed spacer-1 (ITS-1) regions were used to confirm the presence of Toxoplasma oocysts. By light microscopy, 49/254 (19.3%) of specimens contained coccidian oocysts. PCR analysis demonstrated 2/254 (0.8%) and 17/254 (6.7%) positive results using Tox and ITS-1 primers, respectively. However, coccidian oocysts were not identified on microscopic examination of specimens that were PCR-positive by Tox primers. Coccidian oocysts were identified on microscopic examination of 6/17 (35.3%) of the PCR-positive fecal specimens using ITS-1 primers. The BLAST results of 16 ITS-1 sequences were identified as T. gondii (n = 12; 4.7%) and Hammondia hammondi (n = 4; 1.6%). There was slight agreement between the 529 bp and ITS-1 PCR results (κ = 0.148). This is the first report of the detection of Toxoplasma oocysts using PCR analysis on feline fecal specimens from Southern Thailand. The ITS-1 region has potential as an alternative marker to identify T. gondii oocysts in feline fecal specimens.

Matched MeSH terms: Toxoplasma/isolation & purification*
Fulltext An evaluation of a recombinant multiepitope based antigen for detection of Toxoplasma gondii specific antibodies

Hajissa K, Zakaria R, Suppian R, Mohamed Z

BMC Infect Dis, 2017 12 29;17(1):807.
PMID: 29284420 DOI: 10.1186/s12879-017-2920-9

BACKGROUND: The inefficiency of the current tachyzoite antigen-based serological assays for the serodiagnosis of Toxoplasma gondii infection mandates the need for acquirement of reliable and standard diagnostic reagents. Recently, epitope-based antigens have emerged as an alternative diagnostic marker for the achievement of highly sensitive and specific capture antigens. In this study, the diagnostic utility of a recombinant multiepitope antigen (USM.TOXO1) for the serodiagnosis of human toxoplasmosis was evaluated.
METHODS: An indirect enzyme-linked immunosorbent assay (ELISA) was developed to evaluate the usefulness of USM.TOXO1 antigen for the detection of IgG antibodies against Toxoplasma gondii in human sera. Whereas the reactivity of the developed antigen against IgM antibody was evaluated by western blot and Dot enzyme immunoassay (dot-EIA) analysis.
RESULTS: The diagnostic performance of the new antigens in IgG ELISA was achieved at the maximum values of 85.43% and 81.25% for diagnostic sensitivity and specificity respectively. The USM.TOXO1 was also proven to be reactive with anti- T. gondii IgM antibody.
CONCLUSIONS: This finding makes the USM.TOXO1 antigen an attractive candidate for improving the toxoplasmosis serodiagnosis and demonstrates that multiepitope antigens could be a potential and promising diagnostic marker for the development of high sensitive and accurate assays.

Matched MeSH terms: Toxoplasma/immunology*
Fulltext Induction of Protective Immunity against Toxoplasmosis in BALB/c Mice Vaccinated with Toxoplasma gondii Rhoptry-1

Sonaimuthu P, Ching XT, Fong MY, Kalyanasundaram R, Lau YL

Front Microbiol, 2016;7:808.
PMID: 27303390 DOI: 10.3389/fmicb.2016.00808

Toxoplasma gondii is the causative agent for toxoplasmosis. The rhoptry protein 1 (ROP1) is secreted by rhoptry, an apical secretory organelle of the parasite. ROP1 plays an important role in host cell invasion. In this study, the efficacy of ROP1 as a vaccine candidate against toxoplasmosis was evaluated through intramuscular or subcutaneous injection of BALB/c mice followed by immunological characterization (humoral- and cellular-mediated) and lethal challenge against virulent T. gondii RH strain in BALB/c mice. Briefly, a recombinant DNA plasmid (pVAX1-GFP-ROP1) was expressed in CHO cells while expression of recombinant ROP1 protein (rROP1) was carried out in Escherichia coli expression system. Immunization study involved injection of the recombinant pVAX1-ROP1 and purified rROP1 into different group of mice. Empty vector and PBS served as two different types of negative controls. Results obtained demonstrated that ROP1 is an immunogenic antigen that induced humoral immune response whereby detection of a protein band with expected size of 43 kDa was observed against vaccinated mice sera through western blot analysis. ROP1 antigen was shown to elicit cellular-mediated immunity as well whereby stimulated splenocytes with total lysate antigen (TLA) and rROP1 from pVAX1-ROP1 and rROP1-immunized mice, respectively, readily proliferated and secreted large amount of IFN-γ (712 ± 28.1 pg/ml and 1457 ± 31.19 pg/ml, respectively) and relatively low IL-4 level (94 ± 14.5 pg/ml and 186 ± 14.17 pg/ml, respectively). These phenomena suggested that Th1-favored immunity was being induced. Vaccination with ROP1 antigen was able to provide partial protection in the vaccinated mice against lethal challenge with virulent RH strain of tachyzoites. These findings proposed that the ROP1 antigen is a potential candidate for the development of vaccine against toxoplasmosis.

Matched MeSH terms: Toxoplasma
Fulltext Knowledge and practice on Toxoplasma infection in pregnant women from Malaysia, Philippines, and Thailand

Andiappan H, Nissapatorn V, Sawangjaroen N, Khaing SL, Salibay CC, Cheung MM, et al.

Front Microbiol, 2014;5:291.
PMID: 24966855 DOI: 10.3389/fmicb.2014.00291

Toxoplasma gondii, is one of the infectious agents of congenital TORCH infections, causes severe clinical outcomes in fetus and newborns. Nevertheless this life-threatening parasitic disease is preventable by simple preventive measures related to lifestyle during pregnancy. We aim to study on the knowledge about toxoplasmosis and practices that prevents this infection among the pregnant women. Total of 2598 pregnant women from Malaysia, Philippines, and Thailand were randomly surveyed to determine the knowledge and their practices on Toxoplasma infection. The questionnaire covered respondents' general information and knowledge on plausible risks factors, symptoms, timing of infection, prevention knowledge, and preventive behavior regarding Toxoplasma infection. Majority of these pregnant women were in their age group of 20-29 years (50.9%), completed secondary level of education (51.7%), in their second trimester of pregnancies (38.1%), non-parous (36.6%), and had no history of abortion (90.4%). Based on this survey, only 11% of these pregnant women had read, heard, or seen information regarding toxoplasmosis and 3.5% of them were aware of being tested for the infection. A small percentage of these pregnant women knew that T. gondii were shed in the feces of infected cats (19.4%) and sometimes found in the raw or undercooked meat (11.0%). There was 16.1% of responding women knew that toxoplasmosis is caused by an infection. Demographic profiles such as age group, level of education, pregnancy term, and number of children of the pregnant women showed significant association with their responses toward prevention knowledge and preventive behavior related questions (P < 0.05). Thus, it is suggested that health education on toxoplasmosis and primary behavioral practices should be consistently offered to reproductive age women in general and pregnant women in particular. This information could help to reduce vertical transmission of Toxoplasma infection during pregnancy.

Matched MeSH terms: Toxoplasma
Fulltext Evaluation of Immunoprotection Conferred by the Subunit Vaccines of GRA2 and GRA5 against Acute Toxoplasmosis in BALB/c Mice

Ching XT, Fong MY, Lau YL

Front Microbiol, 2016;7:609.
PMID: 27199938 DOI: 10.3389/fmicb.2016.00609

Toxoplasmosis is a foodborne disease caused by Toxoplasma gondii, an obligate intracellular parasite. Severe symptoms occur in the immunocompromised patients and pregnant women leading to fatality and abortions respectively. Vaccination development is essential to control the disease. The T. gondii dense granule antigen 2 and 5 (GRA2 and GRA5) have been targeted in this study because these proteins are essential to the development of parasitophorous vacuole (PV), a specialized compartment formed within the infected host cell. PV is resistance to host cell endosomes and lysosomes thereby protecting the invaded parasite. Recombinant dense granular proteins, GRA2 (rGRA2) and GRA5 (rGRA5) were cloned, expressed, and purified in Escherichia coli, BL21 (DE3) pLysS. The potential of these purified antigens as subunit vaccine candidates against toxoplasmosis were evaluated through subcutaneous injection of BALB/c mice followed by immunological characterization (humoral- and cellular-mediated) and lethal challenge against virulent T. gondii RH strain in BALB/c mice. Results obtained demonstrated that rGRA2 and rGRA5 elicited humoral and cellular-mediated immunity in the mice. High level of IgG antibody was produced with the isotype IgG2a/IgG1 ratio of ≈0.87 (p < 0.001). Significant increase (p < 0.05) in the level of four cytokines (IFN-γ, IL-2, IL-4, and IL-10) was obtained. The antibody and cytokine results suggest that a mix mode of Th1/Th2-immunity was elicited with predominant Th1-immune response inducing partial protection against T. gondii acute infection in BALB/c mice. Our findings indicated that both GRA2 and GRA5 are potential candidates for vaccine development against T. gondii acute infection.

Matched MeSH terms: Toxoplasma
Fulltext Seroepidemiology of Toxoplasmosis among People Having Close Contact with Animals

Brandon-Mong GJ, Che Mat Seri NA, Sharma RS, Andiappan H, Tan TC, Lim YA, et al.

Front Immunol, 2015;6:143.
PMID: 25972863 DOI: 10.3389/fimmu.2015.00143

A cross-sectional study was conducted to determine the seroepidemiology of Toxoplasma infection and its risk association among people having close contact with animals. A total of 312 blood samples were collected from veterinary personnel (veterinarian, technicians, and students) and pet owners from veterinary clinics and hospitals in the area of Klang Valley, Malaysia. About 4 cc of blood samples drawn from agreed participants were processed for measurement of anti-Toxoplasma IgG and IgM antibodies as well as avidity test of Toxoplasma IgG by ELISA I, II, and III kits. Meanwhile, the demographic profiles and possible risk factors of these participants were also recorded in the standardized data collection sheets. Overall seroprevalence of toxoplasmosis was observed in 62 (19.9%) participants being 7 (18.4%) in veterinarians, 15 (33.3%) in veterinary technicians, 29 (14.9%) in veterinary students, and 11 (31.4%) in pet owners. Of 19.9% Toxoplasma seropositive samples, 18.3% was positive for IgG antibody, 1.0% for IgM antibody, and 0.6% for both IgG and IgM antibodies. Of three different IgG avidity ELISA kits, ELISA III showed high avidity in all five seropositive samples (IgM and IgG/IgM antibodies) indicating chronic Toxoplasma infection which is consistent with no evidence of clinical toxoplasmosis diagnosed during the time of this study. Univariate analysis showed that age group, gender, study population, gardening, task performance, and working duration were significantly associated with Toxoplasma seropositivity. Further analysis by multivariate analysis using logistic regression showed that age group of ≥30 years old (OR = 0.34, 95% CI = 0.18-0.63, p = 0.001) and working or study duration of >10 years having close contact with animals (OR = 5.07, 95% CI = 1.80-14.24, p = 0.002) were identified as significant risks for Toxoplasma infection. Based on the results obtained, a comprehensive Toxoplasma screening and health surveillance program on toxoplasmosis should be implemented among people having close contact with animals in general and confirmed Toxoplasma seronegative individuals in particular to prevent seroconversion.

Matched MeSH terms: Toxoplasma
Fulltext Bilateral Jensen disease in a healthy young lady

Ghani Siti-Ilyana, Koh, Yi-Ni, Embong Zunaina

Journal of Biomedical and Clinical Sciences, 2018;3(1):19-24.
MyJurnal

Toxoplasmic optic neuropathy is rare and usually occurs monoocularly. This case report demonstrates a rare presentation of bilateral
juxtapapillary retinochoroiditis (Jensen disease) due to toxoplasma infection in
a young healthy patient. A 20-year-old lady presented with bilateral painless
blurring of central vision for 5 days duration. It was preceded by fever, upper
respiratory tract symptoms and headache. There was no history of contact or
being scratched by a cat. Visual acuity was counting fingers for the right eye
and 6/45 for the left eye. There was presence of relative afferent pupillary
defect in the right eye. Optic nerve functions were impaired bilaterally which
was severe in the right eye. Both eyes showed the presence of mild anterior
segment inflammation and vitritis. Fundus examination revealed juxtapapillary
retinochoroiditis bilaterally with swollen optic disc. Optical coherence
tomography (OCT) showed presence of intra-retinal and sub-retinal fluid at
macular area bilaterally. Serology for anti-toxoplasma Immunoglobulin G (IgG)
was positive with titre of 1450 IU/ml. Computed tomography scan (CT scan) of
brain and orbit was normal. A diagnosis of bilateral juxtapapillary
retinochoroiditis or Jensen disease was made. Oral azithromycin 500 mg daily
and guttae prednisolone 4 hourly for 6 weeks was commenced. Oral
prednisolone 50 mg daily (1 mg/kg/day) was added after completion of 1 week
of antibiotic and was tapered down within 5 weeks. There was improvement of
vision as early as 3 weeks post initiation of the treatment. Upon 6 weeks
completing the treatment, her vision has improved to 6/7.5 on both eyes with
resolution of optic disc swelling and sub-retinal fluid. Early recognition and
initiation of treatment in toxoplasma infection associated with juxtapapillary
retinochoroiditis usually result in good visual prognosis.

Matched MeSH terms: Toxoplasma
Fulltext Epidemiological and clinical features of Talaromycosis (Penicilliosis) Marneffei among human immunodeficiency virus-infected patients in Malaysia

Syaziah, I., Azura, S.S., Tzar, M.N.

Medicine & Health, 2018;13(2):103-113.
MyJurnal

The AIDS epidemic in Southeast Asia has led to a marked rise in the incidence of talaromycosis (penicilliosis) marneffei. The aim of this cross-sectional study was to determine the clinico-epidemiological features and outcome predictors of talaromycosis in Malaysia. We identified Talaromycosis marneffei cases from cultures of sterile specimens from 191 human immunodeficiency virus (HIV)- infected patients. Talaromycosis increased from 20-25 (2010-2011) to 45-50 cases per year (2012-2014). Talaromycosis was the HIV-presenting illness in 117 (61.3%) patients. The incidence of talaromycosis as HIV-presenting illness showed an increasing trend from 10.7 (2010) to 26.4 (2014) cases per 1000 new HIV patients. The patients were between 19 and 74 of age (mean 37.2+9.4 years) and the male to female ratio was 7.7:1. Malay (73, 38.2%) and Chinese (70, 36.3%) were the most prevalent ethnic groups. Common clinical manifestations included loss of weight (85.9%), fever (84.8%) and cough (67%), while skin lesions were only present in 42.9% cases. Common concurrent infections were oral candidiasis (79.6%), tuberculosis (36.1%) and hepatitis C infection (20.9%). Most patients (93.7%) were anaemic with mean haemoglobin level of 9.9+2.3 g/dL, 39% had impaired liver function, and 18.8% were neutropaenic. Median CD4 cell count was 16 cells/L. Most patients (70.4%) received intravenous amphotericin B followed by itraconazole. At 8-month follow up, 148 (81.8%) patients were alive while 33 (18.2%) had died. Intravenous drug abuse, concurrent toxoplasma encephalitis and concurrent Pneumocystis jiroveci pneumonia independently predicted death outcome in both univariate and multivariate logistic regression analyses.

Matched MeSH terms: Toxoplasma
Fulltext Serological and molecular detection of toxoplasmosis among blood donors in tertiary hospital of Malaysia

Aisha Khodijah Kholib Jati, Suharni Mohamad, Zeehaida Mohamed, Wan Haslindawani Wan Mahmood, Wan Muhamad Amir W Ahmad, Wan Suriana Wan Ab Rahman1

Malaysian Journal of Medicine and Health Sciences, 2020;16(3):73-78.
MyJurnal

Introduction: This preliminary cross-sectional study aimed to investigate the prevalence of toxoplasmosis among blood donors in Kelantan, Malaysia. Methods: A total of 56 blood donors were screened by an enzyme-linked immu- nosorbent assay (ELISA) for anti-T. gondii Immunoglobulin G (IgG) and Immunoglobulin M (IgM) antibodies. Positive
T. gondii IgG and IgM were further tested for IgG avidity ELISA. All extracted deoxyribonucleic acids (DNAs) from whole blood samples were analyzed for the presence of the Toxoplasma B1 gene and the ITS1 region by polymerase chain reaction (PCR). The socio-demographic data of donors was assessed using a data collection form. Results: Out of 56 blood donors, 24 (42.86%) donors were IgG+/IgM-, and 2 (3.57%) donors were IgG+/IgM+ with one of them having a high avidity index indicating as past infection for more than 20 weeks and the other with a low avidity index indicating as recent infection within 20 weeks. None of the samples tested positive for the presence of the Toxoplasma B1 gene and the ITS1 region. A univariate analysis showed that only employment status was significantly associated with Toxoplasma seropositivity. Conclusion: The seroprevalence of toxoplasmosis among blood donors in Kelantan, Malaysia, was 46.43%. Nevertheless, direct detection by PCR showed that this parasite was absent in the blood. These results highlight that the blood donors in this study had previously been exposed to T. gondii infection. The parasite may still remain in certain tissues but does not freely circulate in the blood.

Matched MeSH terms: Toxoplasma
Seroprevalence, associated risk factors and hematological impacts of toxoplasmosis in small ruminants of Multan, Punjab-Pakistan

Ullah MZ, Awais MM, Akhtar M, Anwar MI, Navid MT, Khan I, et al.

Trop Biomed, 2018 Dec 01;35(4):1028-1040.
PMID: 33601850

Toxoplasmosis is a protozoal infection of zoonotic potential with worldwide geographical distribution which affects nearly all warm-blooded animals including mammals and birds. Keeping in view, this study was conducted to determine the seroprevalence of toxoplasmosis along with associated risk factors and its haematological impacts in small ruminants of district Multan, Pakistan. In this study, a total of 250 sera samples collected from sheep (n=125) and goats (n=125) from three tehsils of Multan were examined using commercially available Latex agglutination test kit for the presence of anti-T. gondii antibodies. The haematological profiles of Toxoplasma seropositive and seronegative animals were determined by using automated haematology analyser. Overall seroprevalence of toxoplasmosis in small ruminants was 42.80% with a higher prevalence rate (44.80%) in sheep as compared to goats (40.80%). Sex, existence of co-morbid conditions, feeding pattern and presence of pet cats and dogs were identified as significant (P<0.05) risk factors associated with the presence of antibodies against toxoplasmosis. The breed was found to be a significant (P=0.026) risk factor for the seroprevalence of toxoplasmosis in goats but not in sheep. Haematological analysis revealed significantly altered leukocytic counts (P<0.05) in seropositive sheep and goats as compared to seronegative ones. Our findings showed that small ruminants of the Multan District in Pakistan are toxoplasma seropositive and may pose a serious threat of public health concern in the region.

Matched MeSH terms: Toxoplasma
Seroprevalence and molecular detection of Toxoplasma gondii in young healthy blood donors in Northern Iran

Kalantari N, Sheikhansari MR, Ghaffari S, Alipour J, Gorgani-Firouzjaee T, Tamadoni A, et al.

Trop Biomed, 2018 Dec 01;35(4):1017-1027.
PMID: 33601849

T. gondii is a life-threatening infection in immunocompromised patients which may be transmitted through blood transfusion. The present study aimed to evaluate the seroprevalence and molecular detection of T. gondii infection and the associated risk factors among young healthy blood donors in the central part of Mazandaran province, northern Iran. Blood samples were taken from 500 participants and the serum was separated. All serum samples were tested for the presence of anti-T. gondii antibodies (IgG) and then all positive samples were evaluated for IgM antibodies using commercial ELISA kits. All IgM positive samples and 66 randomly selected IgG positive samples were further tested by PCR of the REP-529 gene. Anti-Toxoplasma antibodies (IgG) avidity test was performed for 142 IgG positive samples which were randomly selected. In the current study, anti-T. gondii antibodies (IgG) and (IgM) were found in 316 (63.2%) and 3 (0.95 %) participants, respectively. Seropositivity rate of Toxoplasma was higher among blood donors living in rural areas (P=0.000) and those with a history of soil and animal contact (P<0.05). PCR of the REP-529 gene showed T. gondii DNA in 21 out of 66 samples. The REP-529 gene was not detected in IgM positive samples. Low avidity antibodies (IgG) was found in 23.2% of the IgG positive samples. In conclusions, this study found that the prevalence of toxoplasmosis among young healthy blood donors in north of Iran was high. To reduce the risk of parasite transmission, leukofilteration method are recommended for donated blood used for immunosuppressed patients.

Matched MeSH terms: Toxoplasma
Toxoplasma gondii infection in native village chickens (Gallus domesticus) in Selangor and Melaka, Malaysia

Sabri AR, Hassan L, Sharma RSK, Noordin MM

Trop Biomed, 2019 Sep 01;36(3):604-609.
PMID: 33597482

Toxoplasmosis is a worldwide zoonosis caused by the protozoa Toxoplasma gondii which affects human and animals. Village chickens (Gallus domesticus) most commonly known as Ayam Kampung or free-range chickens, have been suggested to play a role in the epidemiology of toxoplasmosis. This study determines the presence of T. gondii in the village chicken populations in two states of Malaysia. A total of 50 serum samples from the chickens from Selangor (n=20) and Melaka (n=30) were collected and analysed using commercial serological kits. T. gondii antigen was detected in 20% (Selangor 30%; Melaka 13%) samples using ELISA test and anti-T. gondii antibody was detected in all positive ELISA samples using the indirect haemagglutination test (IHAT). Histopathological examination revealed tissue changes such as inflammation and degeneration in brain and liver of seropositive chickens. This is the first report of T. gondii infection in the village chickens in Malaysia.

Matched MeSH terms: Toxoplasma
Fulltext Food and water-borne protozoal diseases (FWBPD)

Baha Latif

Medical Health Reviews, 2009;2009(2):27-46.
MyJurnal

This review places emphasis on the food and water borne protozoal diseases like, cryptosporidiosis, giardiasis, cyclosporiasis, and toxoplasmosis which are endemic in many parts of the world. The article addresses cryptosporidiosis, giardiasis, cyclosporiasis, and toxoplasmosis. Cryptosporidium and Giardia are widespread in the environment and major outbreaks have occurred as a result of contaminated drinking water. Cyclospora has significance in the preparation and consumption of fresh food that have not received heat treatment. Toxoplasma gondii, transmission occur through contaminated water, unpasteurised milk and also the consumption of raw meat, which has been regarded as a major route of transmission to humans .These parasitic protozoa do not multiply in foods, but they may survive in or on moist foods for months especially in cool and damp environment. Their ecology makes control of these parasites difficult. Transmission, clinical manifestations, outbreaks, public health surveillance and control measures that are used for those diseases will be reviewed in this article.

Matched MeSH terms: Toxoplasma
Fulltext Transcriptome Profiling of Toxoplasma gondii-Infected Human Cerebromicrovascular Endothelial Cell Response to Treatment with Monensin

Harun MSR, Taylor M, Zhu XQ, Elsheikha HM

Microorganisms, 2020 Jun 04;8(6).
PMID: 32512820 DOI: 10.3390/microorganisms8060842

Central to the progression of cerebral toxoplasmosis is the interaction of Toxoplasma gondii with the blood-brain barrier (BBB) endothelial cells. In the present work, we tested the hypothesis that inhibition of Wnt pathway signalling by the monovalent ionophore monensin reduces the growth of T. gondii infecting human brain microvascular endothelial cells (hBMECs) or microglial cells. The anti-parasitic effect of monensin (a Wnt signalling inhibitor) on the in vitro growth of T. gondii tachyzoites was investigated using two methods (Sulforhodamine B staining and microscopic parasite counting). The monensin inhibited T. gondii growth (50% inhibitory concentration [IC50] = 0.61 μM) with a selective index = 8.48 when tested against hBMECs (50% cytotoxic concentration [CC50] = 5.17 μM). However, IC50 of monensin was 4.13 μM with a SI = 13.82 when tested against microglia cells (CC50 = 57.08 μM), suggesting less sensitivity of microglia cells to monensin treatment. The effect of T. gondii on the integrity of the BBB was assessed by the transendothelial electrical resistance (TEER) assay using an in vitro human BBB model. The results showed that T. gondii infection significantly decreased hBMECs' TEER resistance, which was rescued when cells were treated with 0.1 µM monensin, probably due to the anti-parasitic activity of monensin. We also investigated the host-targeted effects of 0.1 µM monensin on global gene expression in hBMECs with or without T. gondii infection. Treatment of hBMECs with monensin did not significantly influence the expression of genes involved in the Wnt signalling pathway, suggesting that although inhibition of the Wnt signalling pathway did not play a significant role in T. gondii infection of hBMECs, monensin was still effective in limiting the growth of T. gondii. On the contrary, monensin treatment downregulated pathways related to steroids, cholesterol and protein biosynthesis and their transport between endoplasmic reticulum and Golgi apparatus, and deregulated pathways related to cell cycle and DNA synthesis and repair mechanisms. These results provide new insight into the host-modulatory effect of monensin during T. gondii infection, which merits further investigation.

Matched MeSH terms: Toxoplasma
Toxoplasma gondii infection among selected indigenous community in Sarawak, East Malaysia

Ngui R, Hassan NA, Chang LY, Teh SJC, Chua KH, Kee BP, et al.

Trop Biomed, 2020 Mar 01;37(1):155-164.
PMID: 33612726

Toxoplasma gondii is an obligate intracellular protozoan parasite that causes toxoplasmosis in humans. To date, little is known about T. gondii infection among the indigenous community, particularly in East Malaysia. This study was conducted to determine the status of T. gondii infection and to investigate associated risk factors among the indigenous community of Sarawak, East Malaysia. The sociodemographic data was obtained using a pretested questionnaire. A serological test was done to detect the presence of specific IgM and IgG antibodies against T. gondii in serum samples. A nested polymerase chain reaction (PCR) was used to determine acute infection among seropositive individuals. The overall seroprevalence of T. gondii infection was 50% (95% CI = 43.3 - 56.7). From this subset, 40.1%, 5.7%, and 4.2% were positive for anti-T. Gondii IgG antibodies, IgM, and both IgG and IgM, respectively. Four seropositive samples were amplified through PCR. None of the pregnant women tested positive for T. gondii infection based on the serological and PCR assays. A significant association was found between age, low monthly household income, unemployment, usage of untreated water and close contact with T. gondii seropositive cats. These results provide basic information on T. gondii infection and may be useful for policymakers to initiate prevention and control programs, especially amongst pregnant women and women of childbearing age in the indigenous community.

Matched MeSH terms: Toxoplasma
Seroprevalence of Toxoplasma gondii infection in people in southeast Hubei province, China

Shen ZZ, Li K, Li ZJ, Shang XL, Hu F, Zhou WJ, et al.

Trop Biomed, 2020 Jun 01;37(2):452-457.
PMID: 33612814

Toxoplasma gondii is a world-widely spread zoonotic parasite. However, scarce knowledge is known about the prevalence of T. gondii infection in people in Hubei province, China. This study herein was to perform epidemiological investigation of T. gondii infection in people in this region. A total 12527 blood samples were obtained during 2015-2018, and were assayed for T. gondii antibodies of IgG and IgM, respectively by employing an indirect hemagglutination test (IHA). The results discovered that the prevalence of T. gondii in people was 2.44% and 6.1%, respectively based on antibodies of IgG and IgM, respectively. The prevalence was ranged from 0.3% to 5.4% during 2015-2018 based on IgM antibodies. For genders, the prevalence was 0.7% and 2.6% in males and females, respectively based on IgM antibodies. In different years, the prevalence was ranged from 4.9% to 14.0% based on IgG antibodies. The prevalence of T. gondii was 4.9% and 6.6% in males and femalesy based on IgG antibodies. The current results may be helpful for the implementation of preventive measures against Toxoplasma infection among people living in this region.

Matched MeSH terms: Toxoplasma
Screening and identification of host proteins interacting with Toxoplasma gondii SAG2 by yeast two-hybrid assay

Lai MY, Lau YL

Parasit Vectors, 2017 Oct 02;10(1):456.
PMID: 28969712 DOI: 10.1186/s13071-017-2387-y

BACKGROUND: The identification of receptors or binding partners of Toxoplasma gondii from humans is an essential activity. Many proteins involved in T. gondii invasion have been characterized, and their contribution for parasite entry has been proposed. However, their molecular interactions remain unclear.
RESULTS: Yeast two-hybrid (Y2H) experiment was used to identify the binding partners of surface antigens of T. gondii by using SAG2 as bait. Colony PCR was performed and positive clones were sent for sequencing to confirm their identity. The yeast plasmids for true positive clones were rescued by transformation into E. coli TOP 10F' cells. The interplay between bait and prey was confirmed by β-galactosidase assay and co-immunoprecipitation experiment. We detected 20 clones interacting with SAG2 based on a series of the selection procedures. Following the autoactivation and toxicity tests, SAG2 was proven to be a suitable candidate as a bait. Thirteen clones were further examined by small scale Y2H experiment. The results indicated that a strong interaction existed between Homo sapiens zinc finger protein and SAG2, which could activate the expressions of the reporter genes in diploid yeast. Co-immunoprecipitation experiment result indicated the binding between this prey and SAG2 protein was significant (Mann-Whitney U-test: Z = -1.964, P = 0.05).
CONCLUSIONS: Homo sapiens zinc finger protein was found to interact with SAG2. To improve the understanding of this prey protein's function, advanced investigations need to be carried out.

Matched MeSH terms: Toxoplasma
IFN-γ and IL-18 in conditioned media of parasite-infected host and IL-21-silenced colorectal cancer cells

Ge P, Ong CY, Abdalkareem AE, Khoo BY, Yuan B

Exp Ther Med, 2021 Feb;21(2):103.
PMID: 33335566 DOI: 10.3892/etm.2020.9535

The presence of certain soluble factors may provide a possible selective advantage for a parasite to gradually modify cell proliferation in neighbouring cells, which may result in chronic diseases. These soluble factors present in the conditioned medium also allow the parasite to invade rapidly into more host cells. The present study aimed to determine the levels of a group of type 1 T helper (Th1) cytokines in the conditioned media of host cells infected with parasites and in IL-21-silenced colorectal cancer cells. The conditioned media of human foreskin fibroblasts (HFFs) parasitized with the RH and ME49 strains of Toxoplasma gondii for 10 days were prepared, and subsequently the levels of the Th1 cytokines in the conditioned media were determined by ELISA. HFFs were incubated with the growth media containing selected soluble factors, and cell proliferation markers were subsequently analysed by reverse transcription-quantitative PCR. The mRNA expression level of cell proliferation markers was also examined in IL-21-silenced HCT116 cells, where the levels of soluble factors in the conditioned media were also determined as aforementioned. The results of the present study demonstrated that HFFs parasitized with ME49 released elevated levels of IFN-γ and lower levels of IL-18 into the conditioned medium compared with the controls. These phenomena were not observed in the conditioned medium of HFFs parasitized with RH. Similar levels of these soluble factors were also detected in the conditioned medium of IL-21-silenced HCT116 cells. The results of the present study also revealed that Ki67 and proliferating cell nuclear antigen mRNA expression was altered in host cells incubated with various levels of IFN-γ and IL-18, as well as in IL-21-silenced HCT116 cells compared with the respective controls. In conclusion, the current study provided preliminary evidence on the fundamental molecular mechanisms of host-parasite interactions that result in chronic diseases, which may aid in the treatment of these diseases in the relevant endemic regions.

Matched MeSH terms: Toxoplasma
Fulltext Susceptibility of Toxoplasma gondii to Ethanolic Extract of Tinospora crispa in Vero Cells

Sharif AA, Unyah NZ, Nordin N, Basir R, Wana MN, Alapid Ahmad A, et al.

Evid Based Complement Alternat Med, 2019;2019:2916547.
PMID: 31827548 DOI: 10.1155/2019/2916547

Background: Toxoplasmosis remains widely distributed globally and is one of the major neglected parasitic zoonotic infections. The infection is still endemic in most parts of the world due to poor control as well as challenges of the currently used medications which can be overcome by using natural products. This study evaluated the effect of ethanolic extract from the stem of Tinospora crispa (EETC) on host cell invasion and intracellular replication of Toxoplasma gondii.
Method: The stem powder of T. crispa was soaked in absolute ethanol for 72 hours. The resulting ethanolic extract was screened for the presence of phytochemicals. Vero cells monolayer in 96-well plate was infected with RH strain of T. gondii and treated with concentrations of the EETC, Veratrine alkaloid, and clindamycin ranging from 1.56 to 200 μg/mL. MTT assay was conducted after 24 hours to evaluate the cytotoxicity and antiparasitic activities of the EETC. Four and 24 hours treatment models were adapted to assess the infection index and intracellular proliferation of T.
Results: The study revealed that the EETC had no cytotoxic effects on Vero cells with IC50 = 179 μg/mL, as compared to clindamycin (IC50 = 116.5 μg/mL) and Veratrine alkaloid (IC50 = 60.4 μg/mL). The EETC had good anti-toxoplasma activities with IC50 = 6.31 μg/mL in comparison with clindamycin (IC50 = 8.33 μg/mL) and Veratrine alkaloid (IC50 = 14.25 μg/mL). The EETC caused more than 70% and 80% reduction in infection index and intracellular proliferation in both treatment models, respectively.
Conclusion: This in vitro study showed that the EETC contains promising phytochemicals effective against T. gondii and safe to the host cells.

Matched MeSH terms: Toxoplasma

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