Detection of Toxoplasma gondii Oocysts in Water Sample Concentrates by Real-Time PCR

Abstract

PCR techniques in combination with conventional parasite concentration procedures have potential for the sensitive and specific detection of Toxoplasma gondii oocysts in water. Three real-time PCR assays based on the B1 gene and a 529-bp repetitive element were analyzed for the detection of T. gondii tachyzoites and oocysts. Lower sensitivity and specificity were obtained with the B1 gene-based PCR than with the 529-bp repeat-based PCR. New procedures for the real-time PCR detection of T. gondii oocysts in concentrates of surface water were developed and tested in conjunction with a method for the direct extraction of inhibitor-free DNA from water. This technique detected as few as one oocyst seeded to 0.5 ml of packed pellets from water samples concentrated by Envirocheck filters. Thus, this real-time PCR may provide a detection method alternative to the traditional mouse assay and microscopy.

A Streamlined Strategy for Biohydrogen Production with Halanaerobium hydrogeniformans, an Alkaliphilic Bacterium

“…The rates of biohydrogen production were 0.37  and 0.88 μmol H 2 /h/mL for straw and switchgrass, respectively, as compared to a range of 0.59 μmol H 2 /h/mL with xylose or arabinose, and up to 2.35 μmol H 2 /h/mL with glucose. The rate of hydrogen production with switchgrass was highly similar to that found recently using A. thermophilum DSM 6725 at 70°C (Yang et al, 2009). The values of 0.37 and 0.88 μmol H 2 /h/mL for straw and switchgrass, respectively, are ∼12% of those generated by a fully grown microbial consortium producing H 2 from steam explosion-treated corn stover (Datar et al, 2007) or generated in various neutrophilic bioreactor configurations with glucose as the feedstock (Gavala et al, 2006).…”

Sensitive and Cost-Effective Detection ofToxoplasma Gondiiin Water Supplies of the Black Sea in Turkey by Loop-Mediated Isothermal Amplification (LAMP)

“…Although PCR-based assays have been used for detection of T. gondii in clinical specimens, there are few reports about real-time PcR detection of oocysts in water samples (15). the low numbers of T. gondii oocysts and the presence of PcR inhibitors in environmental samples pose a disadvantage to use of PcR for routine detection (8,12,16).…”

“…there are a number of different methods for diagnosis of toxoplasmosis, such as indirect fluorescent antibodytest (IFAT) and enzyme-linked immunosorbent assays (eliSA) (7), immunohistochemical staining and polymerase chain reaction (PCR) (2) and real-time PCR (15). However, unresolved issues regarding the effectiveness of these methods are still a major obstacle for water detection.…”

Large Scale Pre-Diagnosis ofToxoplasma GondiiDNA Genotyping by Real-Time PCR on Amniotic Fluid

“…The specificity rate of real-time PCR for the diagnosis of toxoplasmosis has been reported to be between 94% and 100% (1). Various publications reported methods for obtaining T. gondii DNA from amniotic fluid and protocols for RT PCR (10,17,18). In our study, real-time PCR was positive in 4 out of 300 fetuses.…”