Development of High Resolution Melting Analysis as a Diag-nostic Tool for Molecular Detection of Toxoplasma Infection in Pregnant Women and HIV Positive Cases

Abstract

Background: Toxoplasma gondii is an obligate intracellular protozoan with worldwide distribution. Diagnosis of toxoplasmosis is a very critical issue, especially in pregnant women and immunocompromised patients. The aim of this study was rapid detection of T. gondii DNA in peripheral blood samples (PBS) employing HRM technique and using RE gene.
 Methods: Totally, 242 samples from pregnant women and human immunodeficiency virus (HIV) patients were collected from different hospitals and medical centers of Tehran during Oct 2017 to Dec 2018. High resolution melting analysis (HRM) using partial sequences of repetitive element (RE) gene was done and compared with ELISA test.
 Results: Overall, 51 were positive for acute toxoplasmosis that among them, 12 and 20 reported as positive in pregnant women and HIV+ patients, respectively using HRM technique. Among 70 patients in chronic phase of disease, 10 and 3 samples were reported as positive for pregnant women and HIV+ patients respectively. From 121 negative control, 3 (4.62%) samples associated with HIV+ patients, showed positive real-time PCR and HRM analysis results.
 Conclusion: For the first time, HRM technique via employing RE gene was used for detection of T. gondii infection in PBS. This method is suitable, helpful and in parallel with serological methods for early diagnosis of acute as well as active form of toxoplasmosis in pregnant women and HIV+ patients. The use of techniques based on melt curve and through employing next-generation dyes for diagnosis of T. gondii would be accessible for patients in developing countries.

Advances and Challenges in Diagnostics of Toxoplasmosis in HIV-Infected Patients

“…Nowadays, the use of the PCR method has been helpful in the improvement of diagnostic procedures, especially acute toxoplasmosis diagnosis [ 39 ]. Real-time PCR methods can detect fewer copies of the parasite genome compared to conventional PCR methods [ 85 ]. Currently, in Poland, according to the recommendations of the Polish AIDS Society, all symptomatic patients should be tested with PCR for the presence of parasite DNA in CSF [ 86 ].…”

The Therapeutic Efficacy of Zinc Oxide Nanoparticles on Acute Toxoplasmosis in BALB/c Mice

“…Immunocompetent patients with T. gondii infection are often asymptomatic ( 5 ), but immunocompromised patients may experience severe symptoms, including life-threatening conditions ( 5 , 6 ). Pregnant women with toxoplasmosis are at risk of complications such as miscarriage, blindness, and mental disorders in their fetuses ( 7 ).…”