Alfalfa, Medicago sativa, is a link connecting crop production and animal husbandry and plays a dominant role in the development of the livestock sector. The productivity of alfalfa is adversely affected by aphids' feeding damage and their capacity to transmit viral plant pathogens. To increase alfalfa forage yield, it is imperative to control pest insects and use resistant varieties. The aim of this study was to identify the mechanism of M. sativa resistance to aphids by examining changes in the physiology, feeding behavior, and life history of the pests. The leaves of Gannong No. 5 (HA-3, aphid-resistant cultivar) had denser, longer trichome and thicker cortical parenchyma cell, and greater xylem thicknesses than those of Hunter River (Hu, aphid-susceptible cultivar). Nonprobing behaviors suggested that the spotted alfalfa aphid, Therioaphis trifolii, became more active in searching for suitable feeding sites on HA-3 than on Hu plants. Additionally, T. trifolii showed shorter durations for salivating into sieve elements and ingesting phloem sap on HA-3 plants. Life-table analysis showed that T. trifolii on HA-3 had longer developmental duration, higher mortality rate, and lower fecundity, net reproductive rate, intrinsic rate of increase, finite rate of increase and gross reproduction rate values than that on Hu plants. Moreover, relative fitness was significantly reduced in T. trifolii on HA-3 plants. The results of this study provided a basis for developing better control strategies for T. trifolii and studying the mechanisms of alfalfa resistance to aphids.