Validation of droplet digital Polymerase Chain Reaction for the detection and absolute quantification of Taenia solium eggs in spiked soil samples.

ABSTRACT

To enable the detection of taeniid eggs in environmental samples, a sensitive technology is required. In this study, we validated the effectiveness of a digital droplet Polymerase Chain Reaction (ddPCR) assay for detection, identification and absolute quantification of taeniid DNA from artificially contaminated soils with varying numbers of taeniid eggs using a set of universal primers, JB3 & JB4.5. The results showed that the number of cox1 copies detected increased gradually for both species with the number of taeniid eggs added to the different soil types. The highest cox1 DNA copies recovery for Taenia solium and T. lynciscapreoli was from the sand soil with lowest recovery being observed in clay soils. Therefore, ddPCR is a promising technology for screening of taeniid eggs from soil samples collected in the environment irrespective of the soil type and the number of eggs. The potential of the ddPCR protocol to detect taeniid egg DNA in spiked soil samples has great practical application for taeniid egg screening in soils from endemic areas. However, when universal primers are used in screening environmental samples, the identity of ddPCR positive samples must be confirmed by sequencing. In addition, more validation studies using species-specific primers and field soil samples is recommended.