A multi-omics approach was utilized to identify altered biological responses and functions, and to prioritize contaminants to assess the risks of chemical mixtures in the Maumee Area of Concern (AOC), Maumee River, OH, USA. The Maumee AOC is designated by the United States Environmental Protection Agency as having significant beneficial use impairments, including degradation of fish and wildlife populations, bird or animal deformities or reproduction problems, and loss of fish and wildlife habitat. Tree swallow (Tachycineta bicolor) nestlings were collected at five sites along the Maumee River, which included wastewater treatment plants (WWTPs) and industrial land-use sites. Polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), polycyclic aromatic hydrocarbons (PAHs), polychlorinated dibenzo p dioxins and furans (PCDD/Fs), and chlorinated pesticide concentrations were elevated in Maumee tree swallows, relative to a remote reference site, Star Lake, WI, USA. Liver tissue was utilized for non-targeted transcriptome and targeted metabolome evaluation. A significantly differentially expressed gene cluster related to a downregulation in cell growth and cell cycle regulation was identified when comparing all Maumee River sites with the reference site. There was an upregulation of lipogenesis genes, such as PPAR signaling (HMGCS2, SLC22A5), biosynthesis of unsaturated fatty acids (FASN, SCD, ELOVL2, and FADS2), and higher lipogenesis related metabolites, such as docosapentaenoic acid (DPA), docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and arachidonic acid (AA) at two industrial land-use sites, Ironhead and Maumee, relative to WWTP sites (Perrysburg and SideCut), and the reference site. Toledo Water, in the vicinity of the other two industrial sites and also adjacent to a WWTP, showed a mix of signals between industrial land-use and WWTP land-use. PAHs, oxychlordane, and PBDEs were determined to be the most likely causes of the differentiation in biological responses, including de novo lipogenesis and biosynthesis of unsaturated fatty acids.