Understanding how SARS-CoV-2 interacts with different mammalian angiotensin-converting enzyme II (ACE2) cell entry receptors will help elucidate determinants of intra- and cross-species virus transmission, facilitate development of effective new vaccines for both humans and livestock animals, and guide livestock farming and coronavirus screening procedures to ensure food supply security . In this work we applied laboratory directed evolution to several mammalian ACE2s with the goal of identifying conserved ACE2 mutations that increase spike binding affinity across multiple species . We found the Gln42Leu mutation increased ACE2-spike binding for human as well as four of four other mammalian ACE2s, while the Leu79Ile mutation had a similar effect for human and three of three mammalian ACE2 orthologs . These results are especially notable given the residues' high levels of representation, i.e , 83% for Gln42 and 56% for Leu79, among annotated mammalian ACE2s . We also found that substitutions at ACE2 position 34, which is relatively variable across mammalian ACE2s, increased binding for multiple ACE2 orthologs . Taken together, these results speak strongly to the plausibility of SARS-CoV-2 strains with increased ability to cross species transmission barriers . Our results can guide further computational and experimental studies to develop biomedical technologies and animal husbandry practices that help protect both humans and livestock from existing and future SARS-CoV-2 variants.