Which soil layer contains the most humus?

The O horizon soil layer contains the most humus. This layer is the topmost layer of most types of soil. The O horizon layer is primarily composed of decomposed plant litter and humus.

Humus is a semi-soluble organic matter that is created from decomposed fungi and bacteria. Humus gives the upper layer of soil a dark brown to black color. Organisms add humus and nutrients to the soil through plant litter and the process of decomposition. This process influences the soil’s structure and fertility. Soil organisms, such as earthworms, beetles and termites, vertically redistribute humus within the mineral matter found beneath the soil's surface.