An Illumina MiSeq-Based Amplicon Sequencing Method for the Detection of Mixed Parasite Infections Using the Blastocystis SSU rRNA Gene as an Example.

Parasite mixed infections remain a relatively unexplored field in part due to the difficulties of unraveling complex mixtures of parasite DNA using classical methods of sequencing. Next-generation amplicon sequencing (NGS) is a powerful tool for exploring mixed infections of multiple genetic variants of the same parasite in clinical, environmental (water or soil), or food samples. Here, we provide a method for NGS-based detection of mixed parasite infections which uses the Blastocystis SSU rRNA gene as an example and includes steps for parasite concentration, DNA extraction, sequencing library preparation, and bioinformatic analysis.