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. 2018 Jul-Sep;13(3):373-381.

Detection of Toxoplasma gondii in Acute and Chronic Phases of Infection in Immunocompromised Patients and Pregnant Women with Real-time PCR Assay Using TaqMan Fluorescent Probe

Parisa Mousavi  1 , Hossein Mirhendi  2 , Mehdi Mohebali  1   3 , Saeedeh Shojaee  1 , Hossein Keshavarz Valian  1   3 , Shirzad Fallahi  4 , Setareh Mamishi  5
Affiliations

Detection of Toxoplasma gondii in Acute and Chronic Phases of Infection in Immunocompromised Patients and Pregnant Women with Real-time PCR Assay Using TaqMan Fluorescent Probe

Parisa Mousavi et al. Iran J Parasitol. 2018 Jul-Sep.

Abstract

Background: Toxoplasma gondii, cause severe medical complications in infants and immune-compromised individuals. As using early, sensitive and rapid technique has major in diagnosis of toxoplasmosis, the present study was aimed to detect parasite by using from repetitive element (RE) and B1genes, in blood samples of seropositive immuno-compromised patients and pregnant women.

Methods: A total of 110 peripheral blood samples were collected from seropositive cases with anti-T. gondii antibodies, including immunocompromised patients and pregnant women. DNA was extracted by a commercial kit and subjected to TaqMan probe-based real-time PCR assay by using primers and probes specific for RE and B1 genes, separately. The data were analyzed by Kappa test and SPSS-22 software.

Results: In the pregnant women, 17 (68%) and 14 (56%) samples from 25 IgM+/ IgG+ cases and, 7 (25%) and 6 (21.4%) samples from 28 IgG+/IgM- cases were positive by RE and B1 real time PCR, respectively. Likewise, in immunocompromised group, 20 (66.6%) and 17 (56.6%) samples from 30 IgM+/ IgG+ cases and 2 (7.4%) and 2 (7.4%) samples from 27 IgG+/ IgM- cases were positive by RE and B1 real time PCR, respectively.

Conclusion: Probe-based real time PCR assay is a quantitative approach for early diagnosis of T. gondii infection in clinical samples. Moreover, this method can be more appropriate in diagnosis of acute and reactivated toxoplasmosis. In addition our results indicated that RE gene is more sensitive than B1 gene.

Keywords: B1 gene; Immunocompromised patients; Pregnant women; RE gene; Real time PCR; TaqMan fluorescent probe.

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Conflict of interest statement

Conflict of Interests The authors declare that there is no conflict of interest in this study.

Figures

Fig. 1:
Fig. 1:
A: Real-time amplification plot with 5 serial dilutions ranged from 5000 to 1 tachyzoites as the initial DNA template based on RE target. B: Standard curve for 5 serial dilutions of T. gondii tachyzoites in human EDTA blood. CT values were plotted against amount of tachyzoites based on RE repeated element. C: Real-time amplification plot with 5 serial dilution ranges of 5000 to 1 tachyzoites as the initial DNA template based on B1 target. D: standard curve for 5 serial dilutions of T. gondii tachyzoites in human EDTA blood. CT values were plotted against amount of tachyzoites based on B1 gene. (Rn, fluorescent signal.)
Fig. 2:
Fig. 2:
Real time PCR amplification plot for toxoplasmosis patients, positive and non-templet control based on RE (Image A) and B1 (Image B) genes
See this image and copyright information in PMC

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