Isothermal PCR for Feasible Molecular Diagnosis of Primary Toxoplasmosis in Women Recently Experienced Spontaneous Abortion.

AIM: The current study aimed to assess the practicability of a simple loop-mediated isothermal amplification (LAMP) about real-time quantitative PCR to diagnose primary toxoplasmosis among high-risk pregnant women. METHODS: Cloned Toxoplasma samples were used to calculate the analytical sensitivity while specificity was assessed using pooled DNA samples extracted from other parasitic stages. RESULTS: Both techniques showed 100% sensitivity and specificity and then applied to detect recent Toxoplasma infection in peripheral blood of 77 IgG negative women out of a total 139 women lately experienced spontaneous abortion. The 2 techniques obtained positive results in 8 samples confirming primary toxoplasmosis. CONCLUSION: Generally, LAMP assay is a simple, cost-effective molecular technique can be completed in less than half an hour to diagnose primary Toxoplasma infection. The technique can be applied in a minimally equipped laboratory by ordinary workers to screen the vulnerable groups. Further analysis using larger samples with the quantitative approach is recommended to confirm the sensitivity of this emergent molecular technique.

GENETIC POLYMORPHISM OF TOXOPLASMA GONDII FROM RECENTLY INFECTED ABORTED EGYPTIAN WOMEN.

Toxoplasmosis caused by Toxoplasma gondii is one of the most prevalent parasitic diseases in human beings. Human toxoplasmosis can be associated with serious clinical manifestations, particularly in developing fetus. The aim of the current study was to identify the possible lineage type of Toxoplasma gondii, molecularly detected in placental samples of women whose pregnancies were spontaneously terminated in the first trimester. Preliminary detection of Toxoplasma genomic materials was done by a SYBR green qPCR technology. Subsequent identification of Toxoplasma strain was done for the positive samples using PCR-restriction fragment length polymorphism (RFLP) at the SAG2 loci of T. gondii using restriction enzymes HhaI and Sau3AI. Out of 72 tested samples, Toxoplasma B1 gene was detected in 9 cases. Toxoplasma genotypes I and II in addition to unknown type were identified in 4, 3 and 2 cases respectively, while type III was not detected in our samples, hence excluded as a leading cause of abortion in humans in our preliminary study. Nevertheless, it remains uncertain to what extent the genotype of the parasite directly contributes to the clinical severity of human toxoplasmosis. Certainly, advanced molecular techniques targeting different Toxoplasma strains are crucial for better understanding of human toxoplasmosis. For more elucidation, additional studies are recommended intended for genetic characterization of such serious parasitic infection using larger number of samples.

Endothelial nitric oxide synthase (eNOS) (Glu298Asp) and urotensin II (UTS2 S89N) gene polymorphisms in preeclampsia: prediction and correlation with severity in Egyptian females.

BACKGROUND: Preeclampsia is a leading cause of maternal and fetal/neonatal morbidity and mortality. Early prediction of preeclampsia can minimize maternal and fetal complications. Gene polymorphisms are promising markers for early prediction of preeclampsia. AIM OF WORK: To assess the value of endothelial nitric oxide synthase (eNOS) (Glu298Asp) and urotensin II (UTS2 S89N) gene polymorphisms for early prediction of preeclampsia. METHODS: The preeclamptic group consisted of 53 pregnant who developed preeclampsia (35 mild and 18 severe), while the control group consisted of 65 age-matched pregnant females who completed uncomplicated pregnancies. eNOS and urotensin II gene polymorphisms were tested using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). RESULTS: Concerning the eNOS gene polymorphism, there were highly significant differences between the two groups regarding the GG genotype as well as the G and T allele frequency (p < 0.001) and a statistically significant differences regarding the GT, TT genotypes (p = 0.002, 0.0276, respectively). Concerning the urotensin II gene polymorphisms, there were highly significant differences regarding the SS, SN genotypes as well as the S and N allele frequency (p < 0.001), statistically significant differences regarding the NN genotype (p = 0.063). CONCLUSION: Women having mutation in any of the two studied genes are at risk to develop mild preeclampsia, and those having mutations in both genes are at risk to develop severe preeclampsia, while the females with normal pregnancy are protected by the higher percentage of expression of the normal (wild allelles) of both genes.

Placental protein 13 as an early predictor in Egyptian patients with preeclampsia, correlation to risk, and association with outcome.

INTRODUCTION: Placental protein 13 (PP13) is a protein expressed only in the placenta. It is involved in gluing the placenta to the uterus and remodeling the maternal arteries to expand them. Women who subsequently develop preterm preeclampsia have low first trimester maternal serum. AIM OF WORK: The aim of this work was to assess the value of PP13 as an early marker for screening of preeclampsia and to correlate it with the PP13 messenger RNA (mRNA). PATIENTS AND METHODS: As a part of the Antenatal Screening Project, 100 women in the first trimester of pregnancy were selected and subdivided into 2 groups: 50 women who developed preeclampsia in their third trimester (patient group) and 50 women who completed normal uncomplicated pregnancy until full term (control group). Placental protein 13 level was measured using the commercially available enzyme-linked immunosorbent assay kit and PP13 mRNA was tested using reverse transcription polymerase chain reaction. RESULTS: The maternal serum PP13 level in the preeclamptic group was (157.9 ± 45.5 pg/mL), which is significantly lower than that of the control group (225.3 ± 67.3 pg/mL), with highly statistically significant difference (P < 0.0001). The frequency of maternal PP13 mRNA expression was lower in the preeclamptic group (28%) compared to that in the control group (76%), with highly statistically significant difference (P < 0.0001). CONCLUSION: Combined serum PP13 level assay and PP13 mRNA expression are reliable markers for early detection of preeclampsia, and we recommend doing it as a routine investigation during the first trimester.