Results of metabarcoding experiment aiming to rule out contamination by other animals in the bushmeat market or ecosystem

To rule out contamination of duiker samples by other animals in the bushmeat market or ecosystem, we applied a mammal-generic metabarcoding assay on two extracts from the animals that were MPXV-positive and TATV-like positive. For the MPXV-positive duiker sample from the DRC, no tissue remained after the first extraction and no extract remained after the capture experiments, so no experiment to rule out contamination was performed. Metabarcoding of duiker extracts was performed using the TagSteady library-preparation workflow to minimize tag-jumping and cross-sample contamination. Tagged mammal-generic 16S mam1/mam2 primers were used. PCRs were run in quadruplicate, and amplicons were run with negative controls. After end-repair, libraries were ligated to dual Illumina Y-adapters (P5–P7) and sequenced on an Illumina iSeq 100. Negative controls were sequenced alongside samples to monitor potential contamination. Here, we upload the paired fastq files for the pool along with details about the tag combinations are included in the respective csv files, that allow assignment of individual sequences to their respective samples. See manuscript for results of the taxonomic assignment of reads; no signs of non-duiker wild animal DNA was found in the extracts positive for orthopoxviruses.