AimThis study analyzed microvesicles and exosomes, called as extracellular vesicles (EVs) excreted in serum and cerebrospinal fluid (CSF) from patients with cerebral or gestational toxoplasmosis.
Symptomatic forms of toxoplasmosis are a serious public health problem and occur in around 10-20% of the infected people. Aiming to improve the molecular diagnosis of symptomatic toxoplasmosis in Brazilian patients, this study evaluated the performance of real time PCR testing two primer sets (B1 and REP-529) in detecting Toxoplasma gondii DNA. The methodology was assayed in 807 clinical samples with known clinical diagnosis, ELISA, and conventional PCR results in a 9-year period. All samples were from patients with clinical suspicion of several features of toxoplasmosis. According to the minimum detection limit curve (in C), REP-529 had greater sensitivity to detect T. gondii DNA than B1. Both primer sets were retrospectively evaluated using 515 DNA from different clinical samples. The 122 patients without toxoplasmosis provided high specificity (REP-529, 99.2% and B1, 100%). From the 393 samples with positive ELISA, 146 had clinical diagnosis of toxoplasmosis and positive conventional PCR. REP-529 and B1 sensitivities were 95.9% and 83.6%, respectively. Comparison of REP-529 and B1 performances was further analyzed prospectively in 292 samples. Thus, from a total of 807 DNA analyzed, 217 (26.89%) had positive PCR with, at least one primer set and symptomatic toxoplasmosis confirmed by clinical diagnosis. REP-529 was positive in 97.23%, whereas B1 amplified only 78.80%. After comparing several samples in a Brazilian referral laboratory, this study concluded that REP-529 primer set had better performance than B1 one. These observations were based after using cases with defined clinical diagnosis, ELISA, and conventional PCR.
This study investigated the potential of five miRNA candidates for cerebral toxoplasmosis/HIV co‐infection (CT/HIV) biomarkers. miR‐155‐5p, miR‐146a‐5p, miR‐21‐5p, miR‐125b‐5p and miR‐29c‐3p were tested in 79 plasma divided into groups: 32 CT/HIV patients; 27 individuals with asymptomatic toxoplasmosis (AT); and 20 individuals seronegative for toxoplasmosis (NC). From each was collected peripheral blood/EDTA for laboratory diagnosis. Blood cells for DNA extractions (molecular diagnosis), plasma for RNA extractions (gene expression) and ELISA (serological diagnosis). miRNA expression was performed by qPCR, and values were expressed in Relative Quantification (RQ). Among the five miRNAs, miR‐21‐5p and miR‐146a‐5p were up‐expressed in CT/HIV group when compared with AT and NC groups. RQ means for miR‐21‐5p and miR‐146a‐5p in CT/HIV group were 3.829 and 2.500, while in AT group, were 1.815 and 1.661, respectively. Differences between 3 groups were statistically significant (Kruskal‐Wallis ANOVA test), as well as CT/HIV and AT groups (Mann‐Whitney test). Plasma of CT/HIV and AT groups expressed similar levels of miR‐29c‐3p, miR‐155‐5p and miR‐125b‐5p. As NC group was different of CT/HIV and AT groups, differences between three groups were statistically significant (Kruskal‐Wallis ANOVA test). No difference was shown between CT/HIV and AT groups (Mann‐Whitney test). These results suggest the host miRNAs modulation by Toxoplasma gondii.
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