Comparison of an in house and a commercial real-time polymerase chain reaction targeting Toxoplasma gondii RE gene using various samples collected from patients in Turkey

Abstract

BackgroundToxoplasma gondii is an opportunistic protozoan parasite that can infect all warm-blooded animals including humans and cause serious clinical manifestations. Toxoplasmosis can be diagnosed using histological, serological, and molecular methods. In this study, we aimed to detect T. gondii RE gene in various human samples by in house and commercial real time polymerase chain reactions.MethodsA total of 38 suspected cases of toxoplasmosis [peripheral blood (n:12), amnion fluid (n:11), tissue (n:9), cerebrospinal fluid (n:5), and intraocular fluid (n:1)] were included to the study. An in house and a commercial RT-PCR were applied to investigate the T. gondii RE gene in these samples.ResultsThe compatibility rate of the two tests was 94.7% (37/38). When the commercial RT-PCR kit was taken as reference, the sensitivity and specificity of in house RT-PCR test was 87.5 and 100%. When the in house RT-PCR test was taken as reference, the commercial RT-PCR kit has 100% sensitivity and 96.8% specificity. Incompatibility was detected in only in a buffy coat sample with high protein content.ConclusionsBoth the commercial and in house RT-PCR tests can be used to investigate T. gondii RE gene in various clinical specimens with their high sensitivity and specificity. In house RT-PCR assay can be favorable due to cost savings compared to using the commercial test.

Investigation of the role of stray cats for transmission of toxoplasmosis to humans and animals living in İzmir, Turkey

“…After screening the samples by commercial Real Time PCR, another Real Time PCR method [23], using hybridization probe was performed to confirm the presence of T. gondii RE gene by quantification and melting curve analyses as previously described [17,18,23,24]. During the in house Real Time PCR, the primers targeting the 134 bp region of T. gondii RE gene were 5'-AGGCGAGGGTGAGGATGA-3' (18nt, TOX-SE forward primer; final concentration: 0.…”

Multicenter Comparative Assessment of the TIB MolBiol Toxoplasma gondii Detection Kit and Four Laboratory-Developed PCR Assays for Molecular Diagnosis of Toxoplasmosis

“…Efforts towards Toxoplasma-PCR standardization and quality management include the use of commercialized PCR assays. Their performances, however, are variable and require proper evaluation by expert laboratories [5][6][7][8] -the more so since Toxoplasma parasitic loads in clinical samples are often low, which requires that PCR assays show a high sensitivity. For example in CT, the median concentration in the amniotic fluid (AF) has been reported to be around 10 tachyzoites/mL 9 .…”

Molecular Diagnosis of Toxoplasmosis

“…In the absence of a syndrome-based approach, it remains important to continue the optimization of the preanalytical and analytical steps and to evaluate the performances of the laboratorydeveloped methods and commercialized kits of qPCR assays targeting specifically T. gondii. 8,[10][11][12][13][14][15]18,20,21,28,30 6,7 or two other commercial kits of qPCR which have been previously evaluated. 10,12,13,15 All centers involved in the present study participate to the national external quality assessment on a regular basis with consistent results.…”

“…7 Since several years the growing number of commercially available methods has been facilitating the muchneeded standardization, but not all techniques are equal, and it is essential to correctly evaluate them before their implementation in routine diagnosis. 8,[10][11][12][13][14][15] To this aim, available commercialized PCR assays are evaluated by the "Molecular biology" group of the French National Reference Centre for Toxoplasmosis (http://cnrtoxoplasmose.chu-reims.fr, last access December 28, 2021). 8,10,11,15 Here, analytical performances, clinical sensitivity and specificity of the Toxoplasma RealCycler  Universal assay (Progenie Molecular, Valencia, Spain) were assessed for the diagnosis of toxoplasmosis using 168 characterized human samples in 8 referent centers.…”

“…40-0217-32). 12,15 1). the commercialized BioEvolution® test (BioEvolution®, Bussy-Saint-Martin, France) (Centers A, B).…”