Conservation genetics and habitat occupancy modeling for the frosted elfin butterfly, Callophrys irus

Butterflies and moths are declining globally from a series of conservation threats including climate change, habitat loss and fragmentation, and pesticide use. The frosted elfin butterfly, Callophrys irus, is a rare Lycaenid butterfly with isolated populations, many of which were presumed extirpated due to anthropogenic pressures. Historically, the range of C. irus extended from Ontario to Florida, and west to Texas and Wisconsin, with three subspecies described based on morphological differences. Host plant use for oviposition or as a larval food source varies across the range, with populations using either indigo (Baptisia spp.) or lupine (Lupinus spp.). Previous research had been at local scales, with no multi-state assessments of the population structure, subspecies differentiation, or core habitat components performed. The reported C. irus arsace subspecies, historically restricted to coastal regions of the South (Carolinas and Virginia), had not been observed in over two decades or evaluated genetically or systematically. Working with state and federal agencies, field surveys and non-lethal genetic sampling of frosted elfin was done for both indigo and lupine populations across 11 states on the East Coast. Field surveys in 2019 and 2021 recorded the presence of C. irus across multiple visits as well as habitat data (shrub cover, host plant count, etc.). Using this survey data, a habitat occupancy model was constructed to identify habitat factors that contributed to frosted elfin presence. The most significant predictor of frosted elfin occupancy was the total habitat patch area. The habitat area of the sites ranged from 0.0015 – 10 hectares, the median habitat area among all occupied patches was 1.31 hectares, and C. irus occupancy was predicted to increase by 7% for each additional hectare of habitat area available. Abundance of host plants was strongly correlated with habitat area, suggesting both factors should be considered in management plans. Non-lethal modern tissue samples collected in 2021 and 2022, and samples from preserved specimens in museum and private collections were obtained for genetic analysis. No extant populations within the historical range of C. irus arsace were found in the field survey effort, so this reported subspecies was represented only by preserved specimens. DNA was isolated from samples, with mitochondrial gene cytochrome c oxidase subunit I (COI) and the nuclear gene elongation factor one alpha (EF1α) amplified using polymerase chain reaction (PCR). Amplicons were Sanger sequenced and data were combined with available sequences from GenBank. Genetic differentiation was assessed by conducting haplotype analyses, constructing phylogenetic relationships, and conducting an analysis of molecular variation. There was no genetic evidence that the reported C. irus arsace subspecies was genetically distinct from C. irus irus. Haplotype and phylogenetic analyses identified subtle geographic clustering, particularly in mitochondrial DNA, with unique haplotypes in the far northern, southern, and western parts of the range. No host plant associated genetic differentiation was observed to distinguish indigo-specialized from lupine-specialized populations. These results represent the first range-wide genetic analysis of frosted elfin population structure and evaluation of key habitat covariates. The implications for management from our habitat occupancy model are straightforward and highly applicable. To improve the probability of frosted elfin occupancy, more managed pine barren habitat with more host plants is needed. The overall lack of genetic structure indicates that, if efforts remain as regionally local as possible, translocation or reintroduction of frosted elfin could result in established populations with regionally representative genetic diversity. Further, by combining this knowledge with the results of adjacent habitat occupancy model work, better habitat resources can be provided to frosted elfin butterflies before and as they establish in new locations. Thus, this research can contribute to future conservation efforts for the frosted elfin.

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