Molecular species identification of cryptic apple and snowberry maggots (Diptera: Tephritidae) in western and central Washington.

In Washington state, identification of the quarantine apple pest Rhagoletis pomonella (Walsh) is complicated by the presence of the cryptic species Rhagoletis zephyria Snow (Diptera: Tephritidae). Distinguishing the two flies is important because there is a zero tolerance policy for R. pomonella in apple production for export. Here, we attempt to distinguish the two species by scoring R. pomonella and R. zephyria populations from western and south-central Washington for a set of 11 nuclear markers, including four single nucleotide polymorphisms (SNPs) developed for rapid and inexpensive genotyping using Taqman real-time quantitative-polymerase chain reaction. We show that the four SNPs may be adequate in most cases for distinguishing whether a fly originated from apple or black hawthorn (the two major host plants for R. pomonella representing an economic risk) versus snowberry (the major host for R. zephyria, and not a commercial threat). However, directional introgression of R. zephyria alleles into R. pomonella can complicate the identification of flies of mixed ancestry based only on the four SNPs. Moreover, this problem is more acute in the sensitive apple-growing regions of central Washington where our results imply hybridization is common. Consequently, application of the four SNP quantitative-polymerase chain reaction assay can immediately assist ongoing apple maggot monitoring, while the development of additional genetic markers through next-generation sequencing would be valuable for increasing confidence in species identification and for assessing the threat posed by hybridization as R. pomonella further spreads into the more arid apple-growing regions of central Washington.