Comparison of real-time PCR detection methods for B1 and P30 genes of Toxoplasma gondii

Buchbinder, Susanne; Blatz, R; Rodloff, Arne C.

doi:10.1016/s0732-8893(02)00549-7

Cited by 40 publications

(32 citation statements)

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“…We also evaluated the biological sensitivity in artificial sample and showed that one parasite in one ml of AF could be detected if at least three reactions were performed. Only four different PCR protocols using the B1 gene as a target have been described in the literature and can detect the DNA equivalent from 0.05 to 2.5 tachyzoite per PCR reaction (analytical sensitivity) (Lin et al, 2000;Costa et al, 2000, Romand et al, 2004Buchbinder et al, 2003;Reishl et al, 2003;Edvinsson et al, 2006). These PCR protocols are now used by several teams to quantify T. gondii DNA in different types of samples (Costa et al, 2001;Cassaing et al, 2006;Martino et al, 2005).…”

Section: Discussionmentioning

confidence: 99%

Adapting a conventional pcr assay forToxoplasma gondiidetection to real-time quantitative pcr including a competitive internal control

et al. 2007

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Summary :We have developed a quantitative PCR assay (LightCycler ® ) using the pair of primers JW58 and JW59 for the detection of the 35-fold repeated B1 gene of Toxoplasma gondii. This real-time PCR, using fluorescence resonance energy transfert (FRET) hybridization probes, allows the quantification of T. gondii with several technical requirements not previously described: i) an internal amplification control (co-amplified in a single tube with the same primers), ii) Uracil-N-Glycosylase and iii) a standard curve corresponding to a serial dilution from a calibrated suspension of T. gondii ranging from 40 to 4.10 6 parasites in one ml of amniotic fluid (1 to 10 5 T. gondii/PCR). In artificial samples, one parasite could be detected if at least three reactions were performed. Résumé

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Section: Discussionmentioning

confidence: 99%

Adapting a conventional pcr assay forToxoplasma gondiidetection to real-time quantitative pcr including a competitive internal control

et al. 2007

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“…This method has been used for genome numbers estimation and diagnostic detection of parasites, such as Plasmodium spp. (Perandin et al 2004) and T. gondii (Buchbinder et al 2003).…”

Section: Introductionmentioning

confidence: 99%

Diagnostic and prognostic value of cell free circulating Schistosoma mansoni DNA: an experimental study

Eraky

Aly

2014

J Parasit Dis

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Searching for a more sensitive and accurate marker for schistosomiasis diagnosis and treatment follow up is a potential necessity. Hereby, we evaluated usefulness of circulating free DNA as a marker for schistosomiasis diagnosis, assessing drug efficacy and monitoring the control interventions impact using SYBR green real-time PCR. A batch of mice were infected by 90 ± 10 Schistosoma mansoni cercariae. Starting from the 2nd day post infection (p.i.), groups of 2 or 3 mice were sacrificed every 3 days until 30 days p.i. The remaining animals were treated by a single dose of 400 mg/kg mefloquine and sacrificed in group at 5, 10, 21 days post treatment (35, 40, 51 days p.i.). Using SYBR green real time qPCR, pooled sera DNA were extracted and amplified. The results showed that, circulating free S. mansoni DNA was detected from the 2nd day post infection (p.i.) onwards with gradual decrease in the cycle threshold value C t which indicates the gradual elevation of the DNA level (Log quantity was 2.6-3.1 IU/ml), As the infection progressed, DNA quantity was increased(Log quantity was 6.29 IU/ml). Initial increase of circulating free DNA was observed 10 days post treatment (40 days p.i.) (Log quantity was 7.38 IU/ ml). That was followed by a progressive decrease in DNA level by the end of 21st day, post treatment (51 p.i.) (Log quantity 4.35 IU/ml). In conclusion, circulating free S. mansoni DNA is a reliable marker in the diagnosis of schistosomiasis and for assessing drug efficacy and monitoring the impact of control interventions.

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“…Thus, for Toxoplasma gondii, another protozoan parasite, qrtPCR assays using exactly the same (B1 gene) target and different primers have been reported with analytical sensitivities varying by a 200-fold factor, i.e., from 0.05 to 10 genome equivalents per reaction tube (6,7,11,13,14,19,25,33,36). Still, such variations are due not only to the choice of the primers but also, at least as importantly, to the optimization of the method: indeed, identical primers may yield widely different performances in different laboratories (P. Bastien and the Molecular Biology Group of the French National Reference Center for Toxoplasmosis, unpublished data).…”

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confidence: 99%

Quantitative Real-Time PCR Is Not More Sensitive than “Conventional” PCR

2008

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Comparison of real-time PCR detection methods for B1 and P30 genes of Toxoplasma gondii

Cited by 40 publications

References 6 publications

Adapting a conventional pcr assay forToxoplasma gondiidetection to real-time quantitative pcr including a competitive internal control

Adapting a conventional pcr assay forToxoplasma gondiidetection to real-time quantitative pcr including a competitive internal control

Diagnostic and prognostic value of cell free circulating Schistosoma mansoni DNA: an experimental study

Quantitative Real-Time PCR Is Not More Sensitive than “Conventional” PCR

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