The tomato pest Phthorimaea absoluta Meyrick is highly invasive but has not yet invaded North America. However, several morphologically similar species are already present, making detection of P. absoluta presence and invasion challenging. We designed a quantitative PCR molecular diagnostic to differentiate P. absoluta, P. operculella (Zeller), or Keiferia lycopersicella (Walsingham) (Lepidoptera: Gelechiidae) DNA. Additionally, we developed an RPA-Cas12a molecular diagnostic that allows for the isothermal detection of P. absoluta DNA, eliminating the need for a thermocycler. These results can be visualized simply using a UV light source and cell phone camera. We expect these diagnostics to improve quarantine and prevention measures against this serious agricultural threat.

File S1 contains the Python script used to identify potential Cas12a target sites in Phthorimaea absoluta. File S2 is a fasta file containing alignments of all three species used to design the quantitative PCR diagnostics for two genomic loci. File S3 is a fasta file containing alignments of all three species used to design the RPA-Cas12a daignostics for two genomic loci and one mitochondrial locus. Genome alignment files between the three species were obtained from Tabuloc et al. 2019 (https://doi.org/10.1007/s10340-019-01116-6). 

Python 3 with the "biopython" package is needed to run the python script in File S1.