A novel alfalfa variety designated AFX164040 and seed, plants and plant parts thereof are provided. Methods for producing an alfalfa plant comprise crossing alfalfa variety AFX164040 with another alfalfa plant. Methods for producing an alfalfa plant containing in its genetic material one or more traits transgenes or locus conversions introgressed into AFX164040 through backcross conversion and/or transformation are provided and the alfalfa seed, plant and plant part produced thereby. Alfalfa seed, plants or plant parts produced by crossing alfalfa variety AFX164040 or a locus or trait conversion of AFX164040 with another alfalfa plant or population are disclosed. Alfalfa populations derived from alfalfa variety AFX164040, methods for producing other alfalfa populations derived from alfalfa variety AFX164040 and the alfalfa populations and their parts derived by the use of those methods.