Improving Urine Sample Efficacy as a Convenient Alternative for Invasive Samples in Molecular Diagnosis of Toxoplasmosis.

Background: Diagnosis of some diseases is difficult due to invasive sampling. Urine has been candidate as a non-invasive and convenient alternative. It has many advantages and easy accessibility but some technical ills should be removed. Finding a suitable extraction method for improving urine DNA quantity and quality in altering invasive specimens for molecular diagnosis of some infectious diseases, was the main object of present research. Method: Toxoplasmosis was selected as an experimental model, regarding the congenital and ocular forms, its abundance and requirement to invasive sample for diagnosis. Samples prepared by adding some defined Toxoplasma gondii (RH strain) tachyzoites to normal urine. Several urine DNA extraction and purification methods comparatively were tested for finding the best one. The amount of extracted DNA assessed using Nanodrope spectrophotometer and a multiplex semi-nested PCR were designed for evaluating the results. Results: Urine samples with known number of tachyzoites were purified comparatively by five better methods. The results reviled that Cinnagen kit performed with more efficacies. It works well up to 1-5tachyzoites μl-1 of urine. The amount and quality of extracted DNA of more than 100 urine samples with defined tachyzoites were analyzed using a nested PCR method. Finally methods were enough sensitive to detect one tachyzoite DNA in final PCR reaction. Conclusion: This method was enough eligible and sensitive to perform molecular tests for different purposes of instance detecting toxoplasmosis by urine sample as a convenience and non invasive method; although it is better to perform some more experiments using patients samples comparing gold methods. [ABSTRACT FROM AUTHOR]