Validation of dried blood spots in monitoring toxoplasmosis.

Due to the growing importance of toxoplasmosis worldwide, simple methods of diagnosis are important for epidemiologic studies. Dried blood spot (DBS) is a useful tool that surpasses venipuncture. DBS-Toxoplasma testing via a finger-stick could also be used in setting where phlebotomies might not be feasible, such as worldwide prenatal and newborn screening for congenital toxoplasmosis. This study included 101 study subjects were occupationally at-risk to Toxoplasma gondii infection and 33 as controls. Serum was collected from both groups for the detection of anti-Toxoplasma IgG antibodies by ELISA as a reference gold standard test. For the occupational at-risk group, capillary finger stick derived blood was blotted onto five sets of Whatman protein saver cards. Discs were stored as four sets; three sets at 4°C and eluted 1, 2 and 3 months of storage and one set at -20°C for 3 months then eluted. Additionally, one set was eluted immediately. Anti-Toxoplasma IgG antibodies were evaluated by ELISA from DBS eluted samples and compared to matched sera. DBS elutes from discs that were freshly prepared for anti-Toxoplasma IgG showed 100% sensitivity, specificity and diagnostic accuracy. Serologic testing using DBS showed very good diagnostic accuracy under all mentioned conditions of storage. Higher stability was obtained when the blood discs stored at 4°C for 1 month and up to 3 months at -20°C, with 98.18% sensitivity, 100% specificity and 99% diagnostic accuracy. DBS-Toxoplasma testing is characterized by simplicity in performance, cost-effectiveness and the ease of handling, to store and to transport, with high diagnostic accuracy.

Loop mediated isothermal amplification (LAMP) of Toxoplasma DNA from dried blood spots.

INTRODUCTION: The cosmopolitan protozoan Toxoplasma gondii is a major parasite of warm-blooded animals including man. Early and accurate diagnosis is a must for proper treatment that prevents life threatening sequels. Loop-mediated isothermal amplification (LAMP) is a novel technique that can amplify DNA with high sensitivity and specificity under isothermal conditions. AIM OF THE STUDY: To validate a LAMP-specific protocol for detection of Toxoplasma DNA using dried blood spots (DBS) from mice experimentally infected with the cystogenic Toxoplasma ME-49 strain. METHODS: In this study, the target DNA fragment was the Toxoplasma 529-bp repeat element that exists in 200-300 copies per T. gondii genome. The sensitivity of both LAMP and conventional PCR techniques was estimated in DBS samples from experimental mice at 1-week and 8-weeks post-infection. RESULTS: Out of 20 blood samples gathered on Whatman filter paper from mice at 1-week post-infection, 18 and 16 were positive by LAMP and conventional PCR, respectively. Neither techniques detected parasite DNA in blood at 8th week of infection. CONCLUSION: Dried blood spots are easy source of material for molecular studies. LAMP assay proved higher sensitivity than the conventional PCR in detecting parasitemia in early infection with the cystogenic Toxoplasma strain.

Sex dichotomy in the course of experimental latent toxoplasmosis.

Toxoplasma gondii is an opportunistic zoonotic protozoan that exceeds neurological and congenital impact sequence to reactivating latent toxoplasmosis especially under immunosuppression. Sex-associated hormones influence the severity of Toxoplasma infection. Thus, our study aimed to compare toxoplasmosis associated morbidity in both male and female mice and to monitor the response to anti-Toxoplasma therapeutics fortified with sex hormones in comparison to presently used drugs. Twenty male and 20 female mice were infected with ME49 Toxoplasma strain. The morbidity was assessed in the chronic stage in both sexes by estimating brain cyst burden, brain histopathological examination and monitoring serum anti-Toxoplasma IL-12 using ELISA method. Another 40 male and 40 female mice were infected with ME49 Toxoplasma strain then after 6 weeks received different treatment regimens including Atovaquone, Spiramycin, Metronidazole, Estradiol benzoate and Testoserone propionate either as a monotherapy or as a combination. Treatment response was monitored by scoring mice activity and brain cyst burden. This study showed that female mice demonstrated higher cyst burden and manifested more pathological reactions than male mice. While, the IL-12 serum level was significantly higher in male than female mice. Also, it is proved that the Toxoplasma cyst number was reduced significantly when used testosterone/atovaquone, or testosterone/spiramycin/metronidazole combined regimen in female mice groups. While for male mice, the combined therapy of spiramycin/metronidazole was the superior one. Accordingly, combined therapy with sex hormones is a promising strategy for discovering new therapeutic regimens for treating latent toxoplasmosis especially in female.

Performance of rapid immunochromatographic assay in the diagnosis of Trichomoniasis vaginalis.

Trichomonas vaginalis accounts for nearly half of all curable sexually transmitted diseases worldwide with serious health consequences. Effort to increase the sensitivity of its diagnosis is critical to both control measures and epidemiologic studies. This study was conducted to evaluate the OSOM® Trichomonas Rapid Test (Sekisui Diagnostics, Framingham, MA, USA), a qualitative antigen-detection immunochromatographic (IC) assay in the diagnosis of vaginal trichomoniasis comparable to the conventional methods. The study enrolled 258 females aged 18-50 years classified into symptomatic (185) and asymptomatic (73) groups. Vaginal swab specimens were obtained for wet mount, stained preparation (Giemsa, acridine-orange), culture (InPouch TV™, modified Diamond's), and for rapid OSOM testing. Trichomonas vaginalis was detected in 67, 66, 71, 99, 96, and 97 using wet mount, acridine-orange stain, Giemsa stain, modified Diamond's, InPouch media, and OSOM test, respectively. In comparison to a composite reference standard (CRS) of wet mount microscopy and culture, OSOM test reported 97.98%, 99.37%, 98.98%, 98.75%, and 98.84% for sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy, respectively. The OSOM test proved to be a simple and objective test. This rapid point-of-care-test will contribute significantly in the diagnosis of vaginal trichomoniasis and will increase the understanding of its still vague epidemiology.

Echinococcosis granulosus in stray dogs and Echino-IHAT in the hunters in Cairo, Egypt.

A total of fifty stray dogs of both sex and of different ages were examined for natural infection with Echinococcus granulosus after post-mortem. Three main intestinal helminthes were recovered. These were in descending order of abundance Dipylidium caninum, Toxocara canis and E. granulosus. The latter worm was demonstrated in eight of them (16%). The sera of three out of 21 dog-hunters (14.3%) showed positive Echino-IHAT. Two of the hunters suffered Enterobius vermicularis and one of them had Taenia saginata as well. But, the abdominal X-ray of the two hunters was more or less normal and did not show any hydatid cyst.

Diagnostic potentials of copro-antigen detection based ELISA, compared to microscopy in intestinal amoebiasis.

Ninety three patients clinically presumed to have intestinal amoebiasis were chosen and stool samples were collected from all of them. Stool samples were subjected to microscopic examination and Entamoeba copro-antigens detection using Entamoeba and Entamoeba II tests. Out of 93 clinically positive samples, 51 (54.8%) were found positive by microscopy, while 49 (52.7%) were detected by Entamoeba test as having antigens specific for E. histolytica/E. dispar (88.24% sensitivity). Among 49 specimens, 4 were demonstrated as microscopy negative (90.48% specificity). Entamoeba II test demonstrated 16 specimens having antigens specific for the pathogenic E. histolytica among 49 positive by Entamoeba test, while 33 were detected as positive for nonpathogenic E. dispar copro-antigens. Copro-antigen assay using ELISA has shown to be more sensitive and specific than microscopy in different-tiation between pathogenic and nonpathogenic Entamoeba species. Extensive use of this technique allowed for revising the epidemiology of the true pathogenic E. histolytica and obviate the need for unnecessary chemotherapy with its costs and risk of side effects.