Tracking Global Invasion Pathways of the Spongy Moth (Lepidoptera: Erebidae) to the U.S. using Stable Isotopes as Endogenous Biomarkers

The spread of invasive insect species causes enormous ecological damage and economic losses worldwide. A reliable method that tracks back an invaded insect's origin would be of great use to entomologists, phytopathologists and pest managers. The spongy moth (Lymantria dispar (L.d.), Linnaeus 1758) is a persistent invasive pest in the north-eastern United States (U.S.) and periodically causes major defoliations in temperate forests. We analysed field-captured (Europe, Asia, U.S.) and lab-reared L.d. specimens for their natal isotopic hydrogen and nitrogen signatures imprinted in their biological tissues (δ2H and δ15N) and compared these values to the long-term mean δ2H of regional precipitation (Global Network of Isotopes in Precipitation) and δ15N of regional plants at the capture site. We established the percentage of hydrogen-deuterium exchange for L.d. tissue (Pex=8.2%) using the comparative equilibration method and two-source-mixing-models, which allowed the extraction of the moth's natal δ2H value. We confirmed that the natal δ2H and δ15N values of our specimens are related to the environmental signatures at their geographic origins. With our regression models, we were able to isolate potentially invasive individuals and give estimations of their geographic origin. To enable the application of these methods on eggs, we established an egg-to-adult fraction factor for L.d. (Δegg-adult = 16.3‰ ± 4.3‰). Our models suggested that around 25% of the field-captured spongy moths worldwide were not native in the investigated capture sites. East Asia was the most frequently identified location of probable origin. Furthermore, our data suggested that eggs found on cargo ships in U.S. harbors in Alaska, California and Louisiana most probably originated from Asian L.d. in East Russia. These findings show that stable isotope biomarkers give a unique insight into invasive insect species pathways and thus, can be an effective tool to monitor the spread of insect pest epidemics.Microsoft Excel, 64-bitFunding provided by: Universität für Bodenkultur WienCrossref Funder Registry ID: http://dx.doi.org/10.13039/501100006380Award Number: Funding provided by: U.S. Department of AgricultureCrossref Funder Registry ID: http://dx.doi.org/10.13039/100000199Award Number: Funding provided by: OeAD-GmbHCrossref Funder Registry ID: http://dx.doi.org/10.13039/501100005203Award Number:MEASUREMENTSThe data was obtained at the University of Natural Resources and Life Sciences isotope laboratory (Tulln, Austria) between 2017 and 2019 by analysing tissue of spongy moths (Lymantira dispar, Lepidoptera: Erebidae, formally known as "gypsy moth") for their hydrogen stable isotope ratio (2H/1H = δ2H) using a TC/EA system (Thermo Fisher), a ConFlo III open split interface (Thermo Fisher) and a DeltaPLUS XP isotope-ratio mass spectrometer (IRMS, Thermo Fisher) nitrogen stable isotope ratio (15N/14N = δ15N) using a EA (Flash 2000) and IRMS Delta V system (Thermo Fisher) For δ2Hanalysis, quadruplet samples of 0.2±0.025 mg each of the moths' tarsus (lowest part of the leg, dominantly consisting of chitin) were weighed into silver capsules for solids (3.3x5 mm, IVA Analysentechnik). Eggs were sampled as a whole. A full complement of in-house water reference materials (measured against VSMOW and VSLAP) and the international standards IAEA CH7 PEF1 (δ2H = -100.3‰), USGS 43 Indian hair (δ2H = -44.4‰), IVA Casein 139443 (δ2H = -113.0‰) and IVA NBS22 (δ2H = -116.9‰) were used. The H3+ contribution ("H3-factor") was determined before every measurement. For the δ15N analysis, 2±0.3 mg of moth tissue were sampled into tin capsules for solids (5x8 mm, IVA Analysentechnik). Due to a lack of samples, multiple specimens were combined into one sample for the δ15N analysis. This introduced additional variance to the δ15N results. We used IVA Urea (δ15N = -0.36‰), IAEA 600 Caffeine (δ15N = 0.9‰) and IVA Casein 165389 (δ15N = 5.9‰) as reference materials