Acoustic non-destructive testing techniques have been developed to measure properties, such as stiffness, of standing trees and logs. Standing tree stiffness measurements are commonly calculated by inserting two probes into the “same face” of the tree stem, hitting one with a hammer, and measuring the propagation time of the resulting acoustic signal between two probes. Studies have suggested that these time of flight measurements are biased to measure the outerwood stiffness rather than that of the tree stem as a whole and may vary between different hammer hit strengths and hence between users. In this paper, we explore a multipath acoustic technique for measuring components of the anisotropic stiffness matrix of tree stems, which describes the mechanical properties of wood. This technique uses anisotropic acoustic wave propagation velocity measurements made along the surface and through a standing tree. This technique was used to calculate an average stiffness through the tree’s cross-section. Different acoustic/ultrasonic excitation techniques are investigated which obtain velocity measurements that are independent of the user.