Genetic analysis of the rapid expansion of the Banded Killifish (Fundulus diaphanus) in Illinois

Recent investigations into the rapid expansion of Banded Killifish (Fundulus diaphanus) in Illinois has brought to light significant conservation implications. Previous data suggested that Illinois should be inhabited by the Western subspecies of Banded Killifish (F. d. menona), though morphological data from recently encountered populations reveals some populations might be comprised of the Eastern subspecies (F. d. diaphanus) and/or subspecies hybrids. Until now, the Illinois Endangered Species Protection Board has not had to recognize subspecies, and it was possible the recent expansion of Banded Killifish populations throughout Illinois could have led to the removal of its threatened status, regardless of the subspecies responsible for the expansion. The goal of our project was to investigate the genetic composition of expanding populations of Banded Killifish across Illinois, with an emphasis on determining whether this recovery of a rare taxa is of the native Western subspecies, an invasion of the non-native Eastern subspecies, or whether hybridization between subspecies is occurring. We compared the molecular characteristics of Banded Killifish populations in Illinois and surrounding areas to those from within the native ranges of the Western Banded Killifish and Eastern Banded Killifish. We collected Banded Killifish between 2017 and 2020 from nearly 100 sites around the Midwest – kettle lakes, southern Lake Michigan basin, the Chicago Area Waterway System (CAWS), upper Illinois River, lower Rock River, and Mississippi River – as well as areas representing known Western (e.g., Iowa, Minnesota) and Eastern (e.g., Maryland, Canada) subspecies. Genomic material was extracted from muscle tissues via common extraction kits and protocols and included the mitochondrial D-loop and nuclear X-src intron. Our results show that there are three conservation management units within Illinois – 1) the kettle lake region of McHenry and Lake counties, which is the only area not influenced by the non-native Eastern Banded Killifish; 2) the Lake Michigan – CAWS – Illinois River (including Wolf Lake and Powderhorn Lake in Cook County) population that is predominately made up of the Eastern subspecies and hybrids; and 3) the Mississippi River (including lower Rock River) population, which contains both subspecies as well as subspecies hybrids. Our data should provide the resource managers of Illinois a complete understanding of the expansion of Banded Killifish in Illinois and provide a crucial step in conserving the native Western Banded Killifish, while recognizing the existence of a non-native taxa.