Fiber orientation is essential for the physical properties of composite materials. The theoretical parameters of a given reinforcement are usually known and widely used to predict the behavior of the material. However, manufacturing operations such as weaving or needling can produce deviations or fluctuations of fibers around the expected principal directions. These deviations can cause unexpected behavior of the material and should be taken into account in the quality assessment of the material. In this work, we propose an image processing approach to estimate true principal directions and fiber orientation distribution through image analysis of a single section of the material. The method applies to anisotropic materials with several main fiber directions and with cylindrical fibers bundled in threads. A thread-based labeling algorithm has been developed. It allows reliable estimation of the orientation of fibers and threads and provides directional fiber volume ratios. Our method has been successfully applied to the characterization of carbon reinforcement of composite materials.