Displaying publications 41 - 60 of 141 in total

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  1. Fulltext Significant reduction of brain cysts caused by Toxoplasma gondii after treatment with spiramycin coadministered with metronidazole in a mouse model of chronic toxoplasmosis
    Chew WK, Segarra I, Ambu S, Mak JW
    Antimicrob Agents Chemother, 2012 Apr;56(4):1762-8.
    PMID: 22271863 DOI: 10.1128/AAC.05183-11
    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.
    Matched MeSH terms: Toxoplasmosis/drug therapy*; Toxoplasmosis/parasitology
  2. Fulltext Development of triplex real-time PCR and detection of Toxoplasma gondii DNA in infected mice tissues and spiked human samples
    Rahumatullah A, Khoo BY, Noordin R
    Trop Biomed, 2015 Jun;32(2):376-85.
    PMID: 26691266 MyJurnal
    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.
    Matched MeSH terms: Toxoplasmosis/diagnosis*; Toxoplasmosis/parasitology
  3. 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: Toxoplasmosis/diagnosis*; Toxoplasmosis/immunology
  4. Fulltext Recent advances in Toxoplasma gondii immunotherapeutics
    Lim SS, Othman RY
    Korean J Parasitol, 2014 Dec;52(6):581-93.
    PMID: 25548409 DOI: 10.3347/kjp.2014.52.6.581
    Toxoplasmosis is an opportunistic infection caused by the protozoan parasite Toxoplasma gondii. T. gondii is widespread globally and causes severe diseases in individuals with impaired immune defences as well as congenitally infected infants. The high prevalence rate in some parts of the world such as South America and Africa, coupled with the current drug treatments that trigger hypersensitivity reactions, makes the development of immunotherapeutics intervention a highly important research priority. Immunotherapeutics strategies could either be a vaccine which would confer a pre-emptive immunity to infection, or passive immunization in cases of disease recrudescence or recurrent clinical diseases. As the severity of clinical manifestations is often greater in developing nations, the development of well-tolerated and safe immunotherapeutics becomes not only a scientific pursuit, but a humanitarian enterprise. In the last few years, much progress has been made in vaccine research with new antigens, novel adjuvants, and innovative vaccine delivery such as nanoparticles and antigen encapsulations. A literature search over the past 5 years showed that most experimental studies were focused on DNA vaccination at 52%, followed by protein vaccination which formed 36% of the studies, live attenuated vaccinations at 9%, and heterologous vaccination at 3%; while there were few on passive immunization. Recent progress in studies on vaccination, passive immunization, as well as insights gained from these immunotherapeutics is highlighted in this review.
    Matched MeSH terms: Toxoplasmosis/therapy*
  5. Fulltext Recombinant dense granular protein (GRA5) for detection of human toxoplasmosis by Western blot
    Ching XT, Lau YL, Fong MY, Nissapatorn V, Andiappan H
    Biomed Res Int, 2014;2014:690529.
    PMID: 24987700 DOI: 10.1155/2014/690529
    Toxoplasma gondii infects all warm-blooded animals, including humans, causing serious public health problems and great economic loss for the food industry. Commonly used serological tests require costly and hazardous preparation of whole Toxoplasma lysate antigens from tachyzoites. Here, we have evaluated an alternative method for antigen production, which involved a prokaryotic expression system. Specifically, we expressed T. gondii dense granular protein-5 (GRA5) in Escherichia coli and isolated it by affinity purification. The serodiagnostic potential of the purified recombinant GRA5 (rGRA5) was tested through Western blot analysis against 212 human patient serum samples. We found that rGRA5 protein was 100% specific for analysis of toxoplasmosis-negative human sera. Also, rGRA5 was able to detect acute and chronic T. gondii infections (sensitivities of 46.8% and 61.2%, resp.).
    Matched MeSH terms: Toxoplasmosis/blood*
  6. Fulltext Genotyping of Toxoplasma gondii isolates from wild boars in Peninsular Malaysia
    Puvanesuaran VR, Noordin R, Balakrishnan V
    PLoS One, 2013;8(4):e61730.
    PMID: 23613920 DOI: 10.1371/journal.pone.0061730
    Toxoplasma gondii is a parasitic protozoan that infects nearly one-third of the world population. The present study was done to isolate and genotype T. gondii from wild boar from forests of Pahang, Malaysia. A total of 30 wild boars' blood, heads and hearts were obtained for this study and 30 (100.0%) were found to be seropositive when assayed with modified agglutination test (MAT ≥ 6). The positive samples were inoculated into mice and T. gondii was only isolated from samples that had strong seropositivity (MAT ≥ 1:24).The isolates were subjected to PCR-RFLP analysis and all the Peninsular Malaysia isolates of T. gondii are of clonal type I.
    Matched MeSH terms: Toxoplasmosis, Animal/parasitology*
  7. Cloning and expression of Toxoplasma gondii dense granule antigen 2 (GRA2) gene by Pichia pastoris
    Lau YL, Fong MY, Idris MM, Ching XT
    Southeast Asian J. Trop. Med. Public Health, 2012 Jan;43(1):10-6.
    PMID: 23082548
    Detection of Toxoplasma gondii infection is essential in pregnant women and immunosuppressed patients. Numerous studies have shown that the recombinant production of several Toxoplasma antigens, including dense granule antigens (GRAs) has high potential as diagnostic reagents. In the present study, we produced GRA2 using Pichia pastoris system. RNA of T. gondii RH strain tachyzoite was used as a template to produce cDNA clones of full-length GRA2 via reverse transcriptase PCR. Amplicons were inserted into pPICZalpha A and the recombinant plasmid transformed into P. pastoris, X-33 strain. The expressed recombinant protein was identified by SDS-PAGE and Western blotting. A recombinant protein of -28 kDa was produced, which could be detected by toxoplasmosis positive human sera indicating that the recombinant protein retained its antigenicity. The present study indicates that P. pastoris-expressed GRA2 should be useful for detection of Toxoplasma infection.
    Matched MeSH terms: Toxoplasmosis/diagnosis
  8. Isolation of viable Toxoplasma gondii cysts from brain samples for oral infection
    Puvanesuaran VR, Ibrahim N, Noordin R, Balakrishnan V
    Eur Rev Med Pharmacol Sci, 2012 Sep;16(9):1179-83.
    PMID: 23047500
    AIM: A method was developed to separate contaminant-free viable Toxoplasma gondii cysts from brain samples of infected mice for molecular biology studies and reinfection.
    MATERIALS AND METHODS: The mice brains were homogenized and washed with phosphate buffered saline (PBS) Tween 80 prior to fractionation using 19-22% dextran solution. Finally, the supernatant was purified by two-step membrane filtration (100-160 microm and < 10 microm) to obtain pure T. gondii cyst. The isolates were analyzed through microscopic observation, qPCR and by reinfection of new batch of mice.
    RESULTS: T. gondii cysts were best isolated with 21% dextran solution and two step filtration.
    CONCLUSIONS: The method was observed not to disrupt the integrity of the cysts containing bradyzoites. In addition, the isolated cysts in the filtrate were found to be contaminant-free, viable and able to infect healthy mice when introduced orally; which, mimics the natural infectivity pathway.
    Matched MeSH terms: Toxoplasmosis, Animal/parasitology*
  9. Optimization for high-level expression in Pichia pastoris and purification of truncated and full length recombinant SAG2 of Toxoplasma gondii for diagnostic use
    Ling LY, Ithoi I, Yik FM
    Southeast Asian J. Trop. Med. Public Health, 2010 May;41(3):507-13.
    PMID: 20578535
    SAG2 is one of the major surface antigens of the intracellular protozoan parasite Toxoplasma gondii. In the present study, truncated recombinant SAG2(S) and full length recombinant SAG2(T) of T. gondii were optimally produced (approximately 15 mg/liter) in Pichia pastoris expression system using BMMY medium at pH 3, 25 degrees C in 0.5-1% methanol and a time-course of 1-2 days. The recombinant proteins were purified using a commercial gel filtration purification system obtaining approximately 33% recovery. The purified SAG2(S) and SAG2(T) showed molecular masses of 45 and 36 kDa by SDS-PAGE, respectively. The recombinant proteins were evaluated by Western blotting with patients' sera and demonstrated 90% sensitivity and 100% specificity for detection of toxoplasmosis. This study provided a means for large-scale expression and purification of SAG2, which should be useful for diagnosis of toxoplasmosis.
    Matched MeSH terms: Toxoplasmosis/diagnosis*
  10. Comparison of three forms of antigens in the demonstration of cell-mediated immune response in murine toxoplasmosis
    Rahmah N, Khairul Anuar A
    Biochem Biophys Res Commun, 1992 Dec 15;189(2):640-4.
    PMID: 1472034
    Mice were chronically infected with cysts of ME49 strain of Toxoplasma gondii. At different periods post-infection, their spleens were removed and single cell suspensions were made. Lymphocyte transformation experiments were performed on the lymphocyte suspensions using three different kinds of antigens of ME49 strain of T. gondii, namely soluble, excretory/secretory and cystic forms. The results showed that the pattern of lymphocyte responsiveness was dependent on the kind of antigen employed for induction of the blastogenesis. Using soluble and cystic forms of the antigen, different periods of lymphocyte suppression and lymphocyte proliferation were demonstrated. However, with the use of excretory/secretory antigen, no significant suppression of lymphocyte stimulation was noted throughout the course of infection. Thus excretory/secretory antigen may be the best form of antigen for stimulation of the cell-mediated immune response and hence it appears to be a good candidate for vaccine in toxoplasmosis.
    Matched MeSH terms: Toxoplasmosis, Animal/immunology*
  11. Demonstration of antigenic similarities and variations in excretory/secretory antigens of Toxoplasma gondii
    Rahmah N, Anuar AK
    Biochem Biophys Res Commun, 1992 Aug 31;187(1):294-8.
    PMID: 1520310
    C57BL/6 mice were orally infected with different doses of cysts of ME49 strain of Toxoplasma gondii to produce groups of acutely and chronically infected mice. Sera were obtained at different periods post-infection. SDS-PAGE was ran with excretory/secretory antigens of ME49 and RH strains of T. gondii, followed by Western blot analyses using the above sera and anti- IgA, IgM, IgG as conjugates. The SDS-PAGE profiles of the two antigens were similar. However the antigenic bands showed variations in all blots, most evidently in IgA blots of chronic sera. IgG blots showed greatest similarities in reactive bands. In IgM blots, more common bands were shown in chronic sera than in acute sera. Variations and similarities in prominence of some bands and time of their appearance were also noted, especially in IgM and IgG blots of chronic sera. Thus antigenic variations and similarities are present in excretory/secretory products of different strains of T. gondii.
    Matched MeSH terms: Toxoplasmosis, Animal/immunology*
  12. Prevalence of Toxoplasma gondii antibodies in goats in Sri Lanka
    Dorny P, Van Aken D
    Ann Trop Med Parasitol, 1992 Feb;86(1):83-5.
    PMID: 1616398
    Matched MeSH terms: Toxoplasmosis, Animal/epidemiology*
  13. Prevalence of antibodies including IgM to Toxoplasma gondii in Malaysians
    Thomas V, Sinniah B, Yap PL
    Southeast Asian J. Trop. Med. Public Health, 1980 Mar;11(1):119-25.
    PMID: 6996108
    A total of 736 sera collected from cord blood babies, children and adults of both sexes and of different age groups in Malaysia were tested using indirect fluorescent antibody technique for Toxoplasma antibodies. The RH strain of Toxoplasma gondii zoites were used as antigen. One hundred and twenty five sera which were reactive at 1:64 or high titres were tested with IgM specific conjugate. Results of the present studies showed that the prevalence of Toxoplasma antibody was highest among the Malays and lowest among children than among adults. The significance of Toxoplasma specific IgM was discussed.
    Matched MeSH terms: Toxoplasmosis/epidemiology
  14. Long term maintenance of Toxoplasma gondii (Rh strain) in Vero cell line and use of harvested antigens for immunodiagnosis
    Suresh K, Mak JW, Yong HS
    Southeast Asian J. Trop. Med. Public Health, 1991 Dec;22 Suppl:124-8.
    PMID: 1822869
    Thirty in vitro serial passages of Toxoplasman gondii cultures in Vero cell line performed once in every five days had a mean increase in parasite count of 74.4 +/- 14.8 times from that of initial counts. Long term cultures in Vero cell line did not alter the virulence of the parasite. The good correlation (r = 0.99) between the IFA titer and ELISA OD values using the parasite antigens from in vitro sources indicates that long term maintenance of T. gondii in culture does not affect significantly the ability to recognize antibodies to surface and soluble antigens. The results also show that soluble antigens containing host cells can be directly used for immunodiagnostic purposes without purification. The in vitro maintenance of T. gondii is safer and cheaper when compared to the in vivo method.
    Matched MeSH terms: Toxoplasmosis, Animal/diagnosis*
  15. Current public health status of some food-borne parasitic zoonoses in Malaysia
    Singh KI
    Southeast Asian J. Trop. Med. Public Health, 1991 Dec;22 Suppl:11-5.
    PMID: 1822865
    Matched MeSH terms: Toxoplasmosis/epidemiology
  16. Antibodies to Toxoplasma gondii from domestic animals in Malaysia
    Rajamanickam C, Cheah TS, Paramasvaran S
    Trop Anim Health Prod, 1990 Feb;22(1):61-2.
    PMID: 2321262
    Matched MeSH terms: Toxoplasmosis, Animal/epidemiology*
  17. Fulltext Experiences in the diagnosis and prevention of mental retardation of environmental origin in Singapore
    Paul FM
    Med J Malaysia, 1982 Dec;37(4):357-61.
    PMID: 6300621
    Consideration. is gzven to the recognition and prevention of carious types of mental retardation due to hazards of environmental origin. Observations are presented on congenital syphilis, congenital toxoplasmosis, congenital rubella, Singapore kernicterus, Japanese B encephalitis, and tuberculous meningitis. Appropriate preventiue measures have resulted in a significant reduction in Singapore of these conditions, and hence in a decreased frequency of environmentally determined mental retardation. and related disabilities.
    Matched MeSH terms: Toxoplasmosis, Congenital/complications
  18. IgG avidity Western blot using Toxoplasma gondii rGRA-7 cloned from nucleotides 39-711 for serodiagnosis of acute toxoplasmosis
    Deshpande PS, Kotresha D, Noordin R, Yunus MH, Saadatnia G, Golkar M, et al.
    Rev Inst Med Trop Sao Paulo, 2013 4 9;55(2):79-83.
    PMID: 23563759
    Toxoplasmosis is an important cause of congenital infection. The present study was performed to evaluate the usefulness of recombinant (r) GRA-7 cloned from nucleotides (n) 39-711 in discriminating between acute and chronic toxoplasmosis. First, commercial IgM, IgG and IgG avidity ELISAs were used to determine the serological profile of the sera. Serum samples were from 20 symptomatic patients with acute infection (low IgG avidity, IgM positive), 10 with chronic infection (high IgG avidity, IgM negative) and 10 with indeterminate IgG avidity (IgM positive) which were tested for IgG avidity status with an in-house developed IgG avidity Western blot using the rGRA-7 recombinant antigen. All 20 sera from cases of probable acute infection showed bands which either faded out completely or reduced significantly in intensity after treatment with 8 M urea, whereas the band intensities of the 10 serum samples from chronic cases remained the same. Of the 10 sera with indeterminate IgG avidity status, after treatment with 8 M urea the band intensities with six sera remained the same, two sera had completely faded bands and another two sera had significantly reduced band intensities. Discrimination between acute and chronic toxoplasmosis was successfully performed by the in-house IgG avidity Western blot.
    Matched MeSH terms: Toxoplasmosis/diagnosis*
  19. Fulltext A Toxoplasma gondii 10 kDa in vitro excretory secretory antigen reactive with human IgM and IgA antibodies
    Saadatnia G, Ghaffarifar F, Khalilpour A, Amerizadeh A, Rahmah N
    Trop Biomed, 2011 Dec;28(3):606-14.
    PMID: 22433890 MyJurnal
    Toxoplasmosis can cause serious disease in immunocompromised patients and to congenitally infected foetuses. Appropriate laboratory investigations in potential cases of acute Toxoplasma infection are important. Excretory secretory antigen (ESA) is immunogenic during both human and experimental infections, therefore is considered as a good candidate for investigation into new infection markers. In this study, ESA was prepared from in vitro cultures of Toxoplasma gondii to identify T. gondii ESA antigenic component(s) that is/are most reactive with serum samples from probable acute cases of toxoplasmosis. Serum samples were obtained from several categories of individuals with the following Toxoplasma serology: Group I: IgM+ IgG+ (low IgG avidity) or IgM+ IgG- from sera of patients who had clinical query of toxoplasmosis (n=35). Group II: IgM- IgG+ (high IgG avidity) from chronically infected individuals (n=30). Group III: normal/healthy individuals with anti-Toxoplasma IgMIgG- (n=20). Group IV: individuals with other infections who had anti-Toxoplasma IgM- IgG- (n=10). The ESA was subjected to SDS-PAGE, followed by Western blot analysis using the above sera and probed with peroxidase conjugated anti-human IgM and IgA antibodies. The blots were then developed using chemiluminescence substrate. The selected antigenic band was excised from the gel after two dimensional electrophoresis and sent for mass spectrometry analysis using MALDI TOF-TOF. The most promising antigenic band was a 10 kDa protein which showed sensitivity of 80% in both IgM and IgA blots, and specificity of 96.7% with sera from other infections and healthy controls. The two best identifications for the 10 kDa band were ubiquitin (ribosomal protein CEP52 fusion protein) and polyubiquitin.
    Matched MeSH terms: Toxoplasmosis/diagnosis*
  20. Fulltext Characterisation of a truncated Toxoplasma gondii surface antigen 2 (SAG2) secreted by the methylotrophic yeast Pichia pastoris
    Lau YL, Shamilah H, Fong MY
    Trop Biomed, 2006 Dec;23(2):186-93.
    PMID: 17322821 MyJurnal
    A truncated form of surface antigen 2 (SAG2) of the protozoan parasite Toxoplasma gondii was cloned and expressed in the methylotrophic yeast Pichia pastoris. This recombinant antigen, designated as recSAG2-N, contained only the N-terminal half of the native SAG2. The recSAG2-N was secreted by the Pichia pastoris into the culture supernatant, and it was harvested by using the trichloroacetic acid precipitation method. Specificity of recSAG2-N was evaluated in western blot assays. Fifty human serum samples, including 32 from confirmed cases of toxoplasmosis, were tested. Results from the assays showed that recSAG2-N reacted with sera from the toxoplasmosis cases only. In vivo experiments showed that serum from mice which received recSAG2-N reacted with the native SAG2 of T. gondii.
    Matched MeSH terms: Toxoplasmosis/diagnosis
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