A Triplex Nested PCR for Diagnosis of Plasmodium, Babesia and Toxoplasma in Blood to Promote Safety of Blood Transfusion Samples

BACKGROUND: The risk of transmission of some infectious agents has always been an important life-threatening side effect of blood transfusion. The aim of this study was to develop a triplex nested Polymerase Chain Reaction (tnPCR) to assess the presence of protozoan parasites Plasmodium, Toxoplasma and Babesia in blood samples concurrently. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of developed tnPCR method compared to gold standard methods were determined for all three parasite genera. METHODS: After selecting the genus level specific primers and setting up of tnPCR, blood samples were collected partially from Isfahan Blood Transfusion Organization (IBTO). Some different samples from human and animal were tested by this method in comparison with the gold standard methods (microscopic method for Plasmodium, Babesia and ELISA for Toxoplasma) in 2021. RESULTS: This tnPCR works well and the sensitivity, specificity, PPV, NPV and accuracy, of this molecular method were all 100% for Plasmodium spp., 93.33%, 99.16%, 93.33%, 99.16% and 99.25% for Babesia spp. and 100%,98.5%, 85.72%, 98.5% and 100% for Toxoplasma gondii respectively compared to standard methods. In average there were 100%, 99.22%, 95.24%, 99.5% and 99.75% contingency for all three parasites (α<0.05). The designed and provided method can detect one, two, or all three potentially dangerous pathogens simultaneously in one tube and one-step, in biological specimens as well as blood. CONCLUSION: The developed tnPCR worked well. It could be recommended for facilitating test, saving time, reducing the expense and cross contamination, subsequently the promotion of blood transfusion safety.